Figure 6 - uploaded by Nina Steele
Content may be subject to copyright.
Expression of human CD163+ macrophages in huNRGS mice post-injury. (A) Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue. Heat/intensity corresponds to cell abundance in a plot region for the population indicated above the plot. Among the intact cells (red gate), human leukocytes were defined as CD45+ events (purple gates). Expression of CD45+/CD163+ (green gate) cells in uninjured and injured stomach tissues. SPADE tree analysis of (B) uninjured and (C) injured stomach tissues collected from huNRGS mice 3 days post-injury. (D) Quantification of CD45+/CD163+ cells in uninjured and injured huNRGS mouse stomachs. *p < 0.05 compared with uninjured tissue, n = 4 mice per group. Immunofluorescence staining of CD44v9 (red) and human-specific CD163 (green) in gastric tissue collected from (E) NRGS and (F, G) huNRGS mice 3 days post-injury. (H) Quantification of CD163+ cells per high-power field in NRGS and huNRGS mice. ND = not detected; n = 4 mice per group. Scale bar tile scan = 200 μm, scale bar higher magnification = 50 μm. 

Expression of human CD163+ macrophages in huNRGS mice post-injury. (A) Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue. Heat/intensity corresponds to cell abundance in a plot region for the population indicated above the plot. Among the intact cells (red gate), human leukocytes were defined as CD45+ events (purple gates). Expression of CD45+/CD163+ (green gate) cells in uninjured and injured stomach tissues. SPADE tree analysis of (B) uninjured and (C) injured stomach tissues collected from huNRGS mice 3 days post-injury. (D) Quantification of CD45+/CD163+ cells in uninjured and injured huNRGS mouse stomachs. *p < 0.05 compared with uninjured tissue, n = 4 mice per group. Immunofluorescence staining of CD44v9 (red) and human-specific CD163 (green) in gastric tissue collected from (E) NRGS and (F, G) huNRGS mice 3 days post-injury. (H) Quantification of CD163+ cells per high-power field in NRGS and huNRGS mice. ND = not detected; n = 4 mice per group. Scale bar tile scan = 200 μm, scale bar higher magnification = 50 μm. 

Source publication
Article
Full-text available
Cluster-of-differentiation gene 44, in particular CD44 variant isoform 9 (CD44v9), emerges during regeneration of the gastric epithelium in response to injury. In particular, CD44v9 is expressed within Spasmolytic Polypeptide/TFF2-Expressing Metaplasia (SPEM) glands during gastric repair, but the function is unknown. Here we tested the hypothesis t...

Contexts in source publication

Context 1
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 2
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 3
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 4
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 5
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 6
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 7
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 8
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 9
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 10
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 11
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 12
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 13
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 14
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 15
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 16
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 17
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 18
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 19
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 20
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 21
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 22
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 23
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 24
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 25
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 26
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 27
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 28
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 29
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 30
... are recruited to the ulcer site and are known to secrete growth factors and cytokines necessary for wound repair [2]. This is of significance given that it has been documented that M2 macrophages promote the advancement of SPEM in the presence of inflammation [14]. Indeed, we observed an increase in the myeloid cell number at the ulcer site of young mice 3 days after ulcer induction (supplementary material, Figure S6A). By marked contrast to the young group, aged mice did not show recruitment of macrophages 3 days after ulcer induction (supplementary material, Figure S6B). Tissue was collected from the ulcerated region, and the expression of T H 2 cytokines was analysed by qRT-PCR (supplementary material, Figure S6C, D). Compared with the aged mice, within the injured tissue collected from young mouse stomachs 3 days post-injury, there was a significant increase in IL-13, IL-4, and IL-33 (supplementary material, Figure S6C, D) that is consistent with the expression of IL-33 and IL-13 as promoters of SPEM [20]. However, within the injured tissue of stomachs collected from aged mice, these increases were not observed (supple- mentary material, Figure S6D). Stomachs from both the uninjured and the injured groups of mice were collected, enzymatically dissociated, and F4/80+ macrophages were isolated by FACS (supplementary material, Figure S6E). Gene expression indicative of M1 and M2 phe- notypes was examined from FACS-sorted macrophages (supplementary material, Figure S6F, G). We observed that while the expression of M1 markers (supplemen- tary material, Figure S6F) was low or undetected in macrophages isolated from injured tissue, these cells expressed significant amounts of M2 markers (supple- mentary material, Figure S6G). These data demonstrate that there is a skewed cytokine microenvironment observed in the young versus aged animals, which correlates with an increase in M2 macrophage markers in the young injured ...
Context 31
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 32
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 33
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 34
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 35
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 36
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 37
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 38
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 39
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 40
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 41
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...
Context 42
... determine the immune phenotype following ulcer injury in huNRGS mice, mass cytometry by time-of-flight (CyTOF) analysis was performed on uninjured and injured stomachs 3 days after ulcer induction. Bivariate plots compare human-derived immune cells measured in mouse uninjured and injured stomach tissue ( Figure 6A). Heat/intensity corresponds to cell abundance in a plot region for the popula- tion indicated above the plot ( Figure 6A). Intact cells (31.44%, red gate) were defined using event length and an iridium-based cell marker (Ir-191 intercalator) ( Figure 6A). Among the intact cells, human leukocytes were defined as CD45+ events ( Figure 6A). CyTOF analysis showed that compared with the uninjured huN- RGS mouse stomachs, hCD45+ cells within the injured huNRGS mouse stomachs highly expressed CD163 ( Figure 6A), a scavenger receptor expressed exclusively on cells of the monocyte/macrophage lineage, in partic- ular M2 macrophages [22]. In support of this, CD163+ cells were negative for T-cell markers CD4 and CD3 (supplementary material, Figure S7A, B). We identified cell subpopulations using spanning-tree progression analysis of density-normalized events (SPADE) [23] ( Figure 6B, C). SPADE analysis is an unsupervised hierarchical clustering technique in which the size of each node is the relative cell frequency and the colour is the median intensity of the expression of each marker ( Figure 6B, C). Compared with the uninjured stomachs of huNRGS mice ( Figure 6B, D), there was a signifi- cant increase in human-specific CD45+/CD163+ cells infiltrating the injured tissue ( Figure 6C, D). Infiltration of CD163+ macrophages into the injured epithelium was confirmed by immunofluorescence ( Figure 6F, H), which was not observed in the non-engrafted NRGS mice ( Figure 6E, H). Importantly, macrophage infil- tration correlated with elevated CD44v9 expression ( Figure 6F, G). These data demonstrate a central role for the human immune response in ulcer repair and identified the infiltration of CD163+ macrophages into the ulcer site. Moreover, macrophage infiltration correlated with increased expression of CD44v9 within the regenerating gastric ...

Citations

... Notably, CD44v6 delineates a marker indicative of invasive intramucosal carcinoma and precancerous manifestations. Concurrently, CD44v9 surfaces during gastric epithelial repair post-injury, co-expressing alongside other SPEM-associated markers (55,56). Cumulative evidence underscores the affirmative correlation between CD44 and its variant isoforms with GC onset and progression, pivotal for diagnostic, therapeutic, and prognostic stratifications (57). ...
Article
Full-text available
Gastric cancer (GC) ranks among the top five most diagnosed cancers globally, with particularly high incidence and mortality rates observed in Asian regions. Despite certain advancements achieved through early screening and treatment strategies in many countries, GC continues to pose a significant public health challenge. Approximately 20% of patients infected with Helicobacter pylori develop precancerous lesions, among which metaplasia is the most critical. Except for intestinal metaplasia (IM), which is characterized by goblet cells appearing in the stomach glands, one type of mucous cell metaplasia, spasmolytic polypeptide-expressing metaplasia (SPEM), has attracted much attention. SPEM represents a specific epithelial cell alteration within the gastric mucosa, characterized by the expressing trefoil factor 2 (TFF2) in basal glands, resembling the basal metaplasia of deep antral gland cells. It primarily arises from the transdifferentiation of mature chief cells, mucous neck cells (MNCs), or isthmus stem cells. SPEM is commonly regarded as a precursor lesion in the development of gastric inflammation and subsequent carcinogenesis. The formation of SPEM is intricately associated with chronic gastric inflammation, Helicobacter pylori infection, and various other environmental and genetic factors. Recently, with the profound exploration of the biological and molecular mechanisms underlying SPEM, a deeper understanding of its role in GC initiation and progression has emerged. This review summarizes the role, molecular mechanisms, and clinical significance of SPEM in the onset and progression of GC.
... Moreover, the present study demonstrated that CD44v9 + epithelial cells could be scattered or clustered in normal tissues. Similar studies reported that the CD44v9 expression could occur in the regenerated gastric epithelium cells or gastric epithelium cells infected with H. pylori (36)(37)(38), as well as the expansion of stem cells of gastric tissues (37). ...
Article
Full-text available
The specificity and sensitivity of the current diagnostic and prognostic biomarkers for gastric cancer (GC) are limited. The present study aimed to evaluate the diagnostic and prognostic significance of cluster-of-differentiation gene 44 variant isoform 9 (CD44v9) and T cell immunoglobulin and mucin domain-containing protein 3 (TIM3) expression levels alone or combined in the tumor tissues of patients with GC and reveal the roles of CD44v9 and TIM3 in the cytokeratin (CK)⁺ and CK⁻ regions. Multiplex immunofluorescence staining was performed for CD44v9, TIM3 and CK using a tissue microarray. The tissues were divided into three regions based on CK expression: Total, CK⁺, and CK⁻ regions. The diagnostic and prognostic value was evaluated using receiver operating characteristic curves, Kaplan-Meier and Cox regression analyses. The results demonstrated that the density of cells expressing CD44v9, TIM3 and co-expressing CD44v9 and TIM3 (CD44v9/TIM3) in both the CK⁺ and CK⁻ regions of tumor tissues was significantly higher than those in normal tissues (P<0.001). Moreover, the expression of CD44v9 in the CK⁻ region was significantly positively correlated with age and tumor grade (P<0.05), and the expression of CD44v9/TIM3 in the CK⁻ region of tumor tissues was significantly positively correlated with age, tumor grade and metastasis (P<0.05). Furthermore, the area under the curve for TIM3 expression in the CK⁺ region was 0.709, with a sensitivity of 45.83% and a specificity of 85.54% (P<0.001). High expression of CD44v9 in the CK⁻ region was also significantly associated with poor survival and independently predicted a poor prognosis in patients with GC (hazard ratio, 2.387; 95% confidence interval, 1.384–4.118; P<0.01). In conclusion, dividing tissue regions based on CK expression is important for the diagnosis of GC. The expression of TIM3 in the CK⁺ region demonstrated diagnostic potential for GC, and high expression of CD44v9 in the CK⁻ region was an independent prognostic risk factor for patients with GC.
... mesenchyme(Lee et al, 2023), basement membrane(Bertaux-Skeirik et al, 2017; Khurana et al, 2013), and the immune system(Bockerstett et al, 2020;De Salvo et al, 2021; Jeong et al, 2021;Meyer et al, 2020;Petersen et al, 2018;Petersen et al, 2014). Unfortunately, we are currently limited in our study of the dynamic trafficking changes in cathartocytosis in living cells because we must work in fixed tissue. ...
Preprint
Injury can cause differentiated cells to undergo massive reprogramming to become proliferative to repair tissue via a cellular program called paligenosis. Gastric digestive-enzyme-secreting chief cells use paligenosis to reprogram into progenitor-like Spasmolytic-Polypeptide Expressing Metaplasia (SPEM) cells. Stage 1 of paligenosis is to downscale mature cell architecture via a process involving lysosomes. Here, we noticed that sulfated glycoproteins (which are metaplasia and cancer markers in mice and humans) were not digested during paligenosis but excreted into the gland lumen. Various genetic and pharmacological approaches showed that endoplasmic reticulum membranes and secretory granule cargo were also excreted and that the process proceeded in parallel with, but was independent lysosomal activity. 3-dimensional light and electron-microscopy demonstrated that excretion occurred via unique, complex, multi-chambered invaginations of the apical plasma membrane. As this lysosome-independent cell cleansing process does not seem to have been priorly described, we termed it "cathartocytosis". Cathartocytosis allows a cell to rapidly eject excess material (likely in times of extreme stress such as are induced by paligenosis) without waiting for autophagic and lysosomal digestion. We speculate the ejection of sulfated glycoproteins (likely mucins) would aid in downscaling and might also help bind and flush pathogens (like H pylori which causes SPEM) away from tissue.
... SPEM lineages have been further characterized as diastase-periodic acid Schiff-positive lineages expressing TFF2 and MUC6 and promoting GSII-agglutinin bonds, similar to mucus-secreting cells of the deep antral glands (12). Furthermore, this lineage expresses several other anal genealogical markers, including CD44 variant 9 (CD44v9) and clusterin, where CD44v9 marks the SPEM in the corpus and is involved in regeneration after gastric epithelial cell injury (7,13,14). These studies have shown a high degree of similarity between the spectra of SPEM and the deep antral glands. ...
Article
Full-text available
Gastric cancer ranks as one of the most prevalent cancers worldwide. While the incidence of gastric cancer in Western countries has notably diminished over the past century, it continues to be a leading cause of cancer-related mortality on a global scale. The majority of gastric cancers in humans are attributed to chronic Helicobacter pylori infection and the progression of gastric cancer is often preceded by gastritis, atrophy, metaplasia and dysplasia. However, the precise mechanisms underlying the development of gastric cancer remain ambiguous, including the formation of gastric polyps and precancerous lesions. In humans, two types of precancerous metaplasia have been identified in relation to gastric malignancies: Intestinal metaplasia and spasmolytic polypeptide-expressing metaplasia (SPEM). The role of SPEM in the induction of gastric cancer has gained recent attention and its link with early-stage human gastric cancer is increasingly evident. To gain insight into SPEM, the present study reviewed the role and research progress of SPEM in gastric cancer.
... The CD44v9positive group has a higher recurrence rate than the CD44v9-negative group [35]. Although CD44v9 emerges in response to injury and contributes to the gastric epithelium [36], there are no reports of CD44v9 directly causing GMA. Further functional studies of rs58618380 in CAPZA1 and CD44v9 are required. ...
Article
Full-text available
Background Helicobacter pylori secretes cytotoxin-associated gene A (CagA) into the gastric epithelium, causing gastric mucosal atrophy (GMA) and gastric cancer. In contrast, host cells degrade CagA via autophagy. However, the association between polymorphisms in autophagy-related genes and GMA must be fully elucidated. Results We evaluated the association between single nucleotide polymorphisms (SNPs) in autophagy-related genes (low-density lipoprotein receptor-related protein 1, LRP1 ; capping actin protein of muscle Z-line alpha subunit 1, CAPAZ1 ; and lysosomal-associated membrane protein 1, LAMP1 ) and GMA in 200 H. pylori -positive individuals. The frequency of the T/T genotype at rs1800137 in LRP1 was significantly lower in the GMA group than in the non-GMA group ( p = 0.018, odds ratio [OR] = 0.188). The frequencies of the G/A or A/A genotype at rs4423118 and T/A or A/A genotype at rs58618380 of CAPAZ1 in the GMA group were significantly higher than those in the non-GMA group ( p = 0.029 and p = 0.027, respectively). Multivariate analysis revealed that C/C or C/T genotype at rs1800137, T/A or A/A genotype at rs58618380, and age were independent risk factors for GMA ( p = 0.038, p = 0.023, and p = 0.006, respectively). Furthermore, individuals with the rs1800137 C/C or C/T genotype of LRP1 had a 5.3-fold higher susceptibility to GMA. These genetic tests may provide future directions for precision medicine for individuals more likely to develop GMA. Conclusion LRP1 and CAPZA1 polymorphisms may be associated with the development of GMA.
... As a follow-up to the above studies, Kuo et al. examined the association between serum TFF2 levels and the expression of miR-21, 155 and 223 in gastric mucosa for SPEM and reported that the above molecules might have diagnostic values Kuo et al., 2019). Subsequent studies successively showed that GSII (Shimizu et al., 2016), CD44v9 (Bertaux-Skeirik et al., 2017;Zavros, 2017), Clusterin (Vange et al., 2017), SRY-related high mobility group box gene 9 (SOX9) (Serizawa et al., 2016), human epididymis protein 4 (HE4) (Nozaki et al., 2008;Jeong et al., 2021) and myelin and lymphocyte protein 2 (MAL2) (Weis et al., 2014) were associated with the expression and proliferation of SPEM cells. In recent studies, aquaporin 5 (AQP5), Trop2 and DDIT4 were shown to reflect parietal cell loss and the severity of SPEM development and could also predict a higher risk of GC (Riera et al., 2020;Lee et al., 2021;Miao et al., 2021). ...
Article
Full-text available
Background Spasmolytic polypeptide expression metaplasia (SPEM) occurs in the corpus of the stomach and is closely related to inflammations caused by H. pylori infection. Recently, SPEM was suggested as one of the dubious precancerous lesions of gastric cancer (GC). Thus, further research on SPEM cell transdifferentiation and its underlying mechanisms could facilitate the development of new molecular targets improving the therapeutics of GC. Using bibliometrics, we analyzed publications, summarized the research hotspots and provided references for scientific researchers engaged in related research fields. Methods We searched the Web of Science Core Collection (WoSCC) for publications related to SPEM-GC from 2002 to 2022. The VOSviewer, SCImago, CiteSpace and R software were used to visualize and analyze the data. Gene targets identified in the keyword list were analyzed for functional enrichment using the KEGG and GO databases. Results Of the 292 articles identified in the initial search, we observed a stable trend in SPEM-GC research but rapid growth in the number of citations. The United States was the leader in terms of quality publications and international cooperation among them. The total number of articles published by Chinese scholars was second to the United States. Additionally, despite its low centrality and average citation frequency, China has become one of the world’s most dynamic countries in academics. In terms of productivity, Vanderbilt University was identified as the most productive institution. Further, we also observed that Gastroenterology was the highest co-cited journal, and Goldenring Jr. was the most prolific author with the largest centrality. Conclusion SPEM could serve as an initial step in diagnosing gastric precancerous lesions. Current hotspots and frontiers of research include SPEM cell lineage differentiation, interaction with H. pylori, disturbances of the mucosal microenvironment, biomarkers, clinical diagnosis and outcomes of SPEM, as well as the development of proliferative SPEM animal models. However, further research and collaboration are still required. The findings presented in this study can be used as reference for the research status of SPEM-GC and determine new directions for future studies.
... These data further support that macrophages are recruited to the site of gastric ulcer and that Hh signaling is needed for the recruitment of macrophages to the site of gastric injury. SmoKO mice exhibit loss of CD44v9 and IL-13 at the site of injury The induction of metaplasia has been shown to correlate with the infiltration of M2 macrophages and a cytokine signaling network of IL-33 and IL-13 15,16 . In particular, our group has demonstrated that the infiltration of macrophages to the gastric epithelium during bacterial infection and regeneration is dependent on Hh signaling 7,14 . ...
... In particular, our group has demonstrated that the infiltration of macrophages to the gastric epithelium during bacterial infection and regeneration is dependent on Hh signaling 7,14 . In addition, CD44v9 marks a reparative cell lineage at the ulcer margin 16 . A loss of CD44v9 at the ulcer margin in SmoKO mice (Fig. 3B) compared to controls (Fig. 3A). ...
... Loss of Shh signaling in the myeloid cell lineage observed in the LysMcre/+;Smof/f (SmoKO) mice correlated with not only a loss of macrophage recruitment to the injured stomach but also disruption of epithelial cell regeneration. Induction of spasmolytic polypeptide/TFF2-expressing metaplasia (SPEM) is critical to driving regeneration of the gastric epithelium 16,30 . Cluster-ofdifferentiation gene 44 variant isoform 9 (CD44v9) emerges during regeneration of the gastric epithelium in response to injury and is known to drive SPEM glands. ...
Article
Full-text available
Sonic Hedgehog (Shh), secreted from gastric parietal cells, contributes to the regeneration of the epithelium. The recruitment of macrophages plays a central role in the regenerative process. The mechanism that regulates macrophage recruitment in response to gastric injury is largely unknown. Here we tested the hypothesis that Shh stimulates macrophage chemotaxis to the injured epithelium and contributes to gastric regeneration. A mouse model expressing a myeloid cell-specific deletion of Smoothened (LysMcre/+;Smof/f) was generated using transgenic mice bearing loxP sites flanking the Smo gene (Smo loxP) and mice expressing a Cre recombinase transgene from the Lysozyme M locus (LysMCre). Acetic acid injury was induced in the stomachs of both control and LysMcre/+;Smof/f (SmoKO) mice and gastric epithelial regeneration and macrophage recruitment analyzed over a period of 7 days post-injury. Bone marrow-derived macrophages (BM-Mø) were collected from control and SmoKO mice. Human-derived gastric organoid/macrophage co-cultures were established, and macrophage chemotaxis measured. Compared to control mice, SmoKO animals exhibited inhibition of ulcer repair and normal epithelial regeneration, which correlated with decreased macrophage infiltration at the site of injury. Bone marrow chimera experiments using SmoKO donor cells showed that control chimera mice transplanted with SmoKO bone marrow donor cells exhibited a loss of ulcer repair, and transplantation of control bone marrow donor cells to SmoKO mice rescued epithelial cell regeneration. Histamine-stimulated Shh secretion in human organoid/macrophage co-cultures resulted in macrophage migration toward the gastric epithelium, a response that was blocked with Smo inhibitor Vismodegib. Shh-induced macrophage migration was mediated by AKT signaling. In conclusion, Shh signaling acts as a macrophage chemoattractant via a Smo-dependent mechanism during gastric epithelial regeneration in response to injury.
... Indeed, certain genes upregulated in metaplastic corpus epithelium are highly expressed in antral glands at homeostasis, including DMBT1 (Sousa et al., 2012;Garay et al., 2017) and GKN3 (Menheniott et al., 2010;Bockerstett et al., 2020), but by the same token, various genes appear to be unique to metaplasia in the corpus, neither expressed in corpus nor antral glands at homeostasis. These include genes such as HE4 (Jeong et al., 2021;Nozaki et al., 2008;O'Neal et al., 2013), CD44v9 (Fan et al., 1996;Bertaux-Skeirik et al., 2015;Bertaux-Skeirik et al., 2017;Tsugawa et al., 2019), and CLU (Weis et al., 2013;Shimizu et al., 2016), among others (Weis et al., 2014). A more appropriate term to describe these glandular changes (and which will be used in this review to refer to these metaplastic changes) is spasmolytic polypeptide-expressing metaplasia (or SPEM), which is characterized by the expression of the antral spasmolytic polypeptide in the bases of corpus glands (Schmidt et al., 1999) and accurately describes the characteristic features of this metaplastic entity. ...
Article
Full-text available
The human stomach functions as both a digestive and innate immune organ. Its main product, acid, rapidly breaks down ingested products and equally serves as a highly effective microbial filter. The gastric epithelium has evolved mechanisms to appropriately handle the myriad of injurious substances, both exogenous and endogenous, to maintain the epithelial barrier and restore homeostasis. The most significant chronic insult that the stomach must face is Helicobacter pylori (Hp), a stomach-adapted bacterium that can colonize the stomach and induce chronic inflammatory and pre-neoplastic changes. The progression from chronic inflammation to dysplasia relies on the decades-long interplay between this oncobacterium and its gastric host. This review summarizes the functional and molecular regionalization of the stomach at homeostasis and details how chronic inflammation can lead to characteristic alterations in these developmental demarcations, both at the topographic and glandular levels. More importantly, this review illustrates our current understanding of the epithelial mechanisms that underlie the pre-malignant gastric landscape, how Hp adapts to and exploits these changes, and the clinical implications of identifying these changes in order to stratify patients at risk of developing gastric cancer, a leading cause of cancer-related deaths worldwide.
... SPEM can be identified by the expression of a number of proteins not expressed in healthy epithelium (CD44v9, GKN3). 20,21,45 As previously reported, SPEM staining was present throughout the corpus of TxA23 mice. 20 In contrast, there was little to no SPEM observed in age-matched TxA23xIl4ra -/mice ( Figure 5B). ...
Article
Full-text available
Background aims: It is well established that chronic inflammation promotes gastric cancer-associated metaplasia, but little is known regarding the mechanisms by which immune cells and cytokines regulate metaplastic cellular changes. The goals of this study were to identify interleukin 13 (IL-13)-producing immune cells, determine the gastric epithelial cell response(s) to IL-13, and establish the role(s) of IL-13 in metaplasia development. Methods: Experiments utilized an established mouse model of autoimmune gastritis (TxA23), TxA23xIl4ra-/- mice, which develop gastritis but do not express the IL-4/IL-13 receptor subunit IL-4Rα, and TxA23xIL-13-YFP mice, which express yellow fluorescent protein in IL-13-producing cells. Flow cytometry was used to measure IL-13 secretion and identify IL-13-producing immune cells. Mouse and human gastric organoids were cultured with IL-13 to determine epithelial cell response(s) to IL-13. Single-cell RNA sequencing was performed on gastric epithelial cells from healthy and inflamed mouse stomachs. Mice with gastritis were administered IL-13-neutralizing antibodies and stomachs were analyzed by histopathology and immunofluorescence. Results: We identified six unique subsets of IL-13-producing immune cells in the inflamed stomach. Organoid cultures showed that IL-13 acts directly on gastric epithelium to induce a metaplastic phenotype. IL-4Rα-deficient mice did not progress to metaplasia. Single-cell RNA sequencing determined that gastric epithelial cells from IL-4Rα-deficient mice upregulated inflammatory genes but failed to upregulate metaplasia-associated transcripts. Neutralization of IL-13 significantly reduced and reversed metaplasia development in mice with gastritis. Conclusions: IL-13 is made by a variety of immune cell subsets during chronic gastritis and promotes gastric cancer-associated metaplastic epithelial cell changes. Neutralization of IL-13 reduces metaplasia severity during chronic gastritis.
... In contrast to unaffected gastric glands showing no expressions of GSII, CD44v9, or AQP5 at the base, regenerating glands adjacent to ulcerated mucosa prominently showed CD44v9 expression at the base as described previously ( Figure 4A). 27 Most of the CD44v9expressing basal gland cells strongly expressed AQP5 and GSII ( Figure 4A). These findings indicate that AQP5 is expressed in SPEM lineages as part of the regenerative mucosa surrounding acute ulcers. ...
Article
Full-text available
BACKGROUND & AIMS Metaplasia in the stomach is highly associated with development of intestinal-type gastric cancer. Two types of metaplasias, spasmolytic polypeptide-expressing metaplasia (SPEM) and intestinal metaplasia (IM) are considered precancerous lesions. However, it remains unclear how SPEM and IM are related. Here, we investigated a new lineage-specific marker for SPEM cells, aquaporin 5 (AQP5), to assist in the identification of these two metaplasias. METHODS Drug- or Helicobacter felis (H. felis) infection-induced mouse models were used to identify the expression pattern of AQP5 in acute or chronic SPEM. Gene-manipulated mice treated with or without drug were employed to investigate how AQP5 expression is regulated in metaplastic lesions. Metaplastic samples from transgenic mice and human gastric cancer patients were evaluated for AQP5 expression. Immunostaining with lineage-specific markers was used to differentiate metaplastic gland characteristics. RESULTS Our results revealed that AQP5 is a novel lineage-specific marker for SPEM cells which are localized at the base of metaplastic glands initially and expand to dominate glands after chronic H. felis infection. Additionally, AQP5 expression was upregulated early in chief cell reprogramming and was promoted by interleukin-13 (IL-13). In humans, metaplastic corpus showed highly branched structures with AQP5-positive SPEM. Human SPEM cells strongly expressing AQP5 were present at the bases of incomplete IM glands marked by TROP2, but were absent from complete IM glands. CONCLUSIONS AQP5-expressing SPEM cells are present in pyloric metaplasia and TROP2-positive incomplete IM and may be an important component of metaplasia which can predict a higher risk for gastric cancer development.