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ABSTRACT: Eph kinases and their ephrin ligands (EFN) are all cell surface molecules, capable of transmitting signals in both directions (1, 2). Such bidirectional signaling is called forward (from EFNs to Ephs) and reverse (from Ephs to EFNs) signaling. Eph family kinases have 15 members, divided into A and B subfamilies. Ephrin ligands have 9 members, also classified into A and B families. Ephs and ephrins interact promiscuously, but EphAs mainly interact with EFNAs, and EphBs with EFNBs. EphB family kinases and their ephrin ligands (EFN) are expressed in the T cell compartment.
In this study, using mice with T cell-specific EFNB2 gene knockout (EFNB2 KO mice), we investigated T cell development and function after EFNB2 deletion. EFNB2 KO mice presented normal thymus weight and cellularity. Their thymocyte subpopulations, such as CD4CD8 double positive cells and CD4 and CD8 single positive cells, were normally distributed, but there was a significant relative increase of CD4CD8 double negative cells. Flow cytometry analysis revealed that there was a moderate increase in the DN3 subpopulation. This augmented percentage of DN cells was further confirmed in competitive repopulation chimeras, suggesting that EFNB2 is involved in thymocyte development. The EFNB2 KO mice had normal T cell numbers and percentages in the spleen, and the T cells were able to be activated and to proliferate normally upon TCR ligation. Further, EFNB2 KO naïve CD4 cells were capable of differentiating into Th1, Th2, Th17 and Treg cells similar to WT naïve CD4 cells.
Our results suggest the involvement of EFNB2 in thymocyte development. However, heavy redundancy among Eph/EFN family members prevents the occurrence of detrimental phenotypes in the T cell compartment caused by T cell-specific EFNB2 gene null mutation.
Molecular Immunology 06/2012; 52(3-4):141-7. · 2.90 Impact Factor
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ABSTRACT: Eph kinases constitute the largest receptor tyrosine kinase family, and their ligands, ephrins (Efns), are also cell surface molecules. Although they are ligands, Efns can transduce signals reversely into cells. We have no prior knowledge of the role played by any members of this family of kinases or their ligands in blood pressure (BP) regulation. In the present studies, we investigated the role of Efnb1 in vascular smooth muscle cell (VSMC) contractility and BP regulation. We revealed that reverse signaling through Efnb1 led to a reduction of RhoA activation and VSMC contractility in vitro. Consistent with this finding, ex vivo, there was an increase of RhoA activity accompanied by augmented myosin light chain phosphorylation in mesenteric arteries from mice with smooth muscle-specific conditional Efnb1 gene knock-out (KO). Small interfering RNA knockdown of Grip1, a molecule associated with the Efnb1 intracellular tail, partially eliminated the effect of Efnb1 on VSMC contractility and myosin light chain phosphorylation. In support of these in vitro and ex vivo results, Efnb1 KO mice on a high salt diet showed a statistically significant heightened increment of BP at multiple time points during stress compared with wild type littermates. Our results demonstrate that Efnb1 is a previously unknown negative regulator of VSMC contractility and BP and that it exerts such effects via reverse signaling through Grip1.
Journal of Biological Chemistry 03/2012; 287(19):15557-69. · 4.77 Impact Factor
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Hongyu Luo,
Zenghui Wu,
Johanne Tremblay,
Eric Thorin,
Junzheng Peng,
Julie L Lavoie,
Bing Hu,
Ekatherina Stoyanova,
Guy Cloutier,
Shijie Qi,
Tao Wu,
Mark Cameron,
Jiangping Wu
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ABSTRACT: Eph kinases constitute the largest receptor tyrosine kinase family, and their ligands, ephrins (Efns), are also cell surface molecules. Our study is the first to assess the role of Ephb6 in blood pressure (BP) regulation. We observed that EphB6 and all three of its Efnb ligands were expressed on vascular smooth muscle cells (VSMC) in mice. We discovered that small arteries from castrated Ephb6 gene KO males showed increased contractility, RhoA activation, and constitutive myosin light chain phosphorylation ex vivo compared with their WT counterparts. Consistent with this finding, castrated Ephb6 KO mice presented heightened BP compared with castrated WT controls. In vitro experiments in VSMC revealed that cross-linking Efnbs but not Ephb6 resulted in reduced VSMC contractions, suggesting that reverse signaling through Efnbs was responsible for the observed BP phenotype. The reverse signaling was mediated by an adaptor protein Grip1. Additional experiments demonstrated decreased 24-h urine catecholamines in male Ephb6 KO mice, probably as a compensatory feedback mechanism to keep their BP in the normal range. After castration, however, such compensation was abolished in Ephb6 KO mice and was likely the reason why BP increased overtly in these animals. It suggests that Ephb6 has a target in the nervous/endocrine system in addition to VSMC, regulating a testosterone-dependent catecholamine compensatory mechanism. Our study discloses that Ephs and Efns, in concert with testosterone, play a critical role in regulating small artery contractility and BP.
Journal of Biological Chemistry 02/2012; 287(9):6819-29. · 4.77 Impact Factor
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Hongyu Luo,
Zenghui Wu,
Johanne Tremblay,
Eric Thorin,
Junzheng Peng,
Julie L. Lavoie,
Bing Hu,
Ekatherina Stoyanova,
Guy Cloutier,
Shijie Qi,
Tao Wu,
Mark Cameron,
Jiangping Wu
[show abstract]
[hide abstract]
ABSTRACT: Eph kinases constitute the largest receptor tyrosine kinase family, and their ligands, ephrins (Efns), are also cell surface
molecules. Our study is the first to assess the role of Ephb6 in blood pressure (BP) regulation. We observed that EphB6 and
all three of its Efnb ligands were expressed on vascular smooth muscle cells (VSMC) in mice. We discovered that small arteries
from castrated Ephb6 gene KO males showed increased contractility, RhoA activation, and constitutive myosin light chain phosphorylation
ex vivo compared with their WT counterparts. Consistent with this finding, castrated Ephb6 KO mice presented heightened BP compared
with castrated WT controls. In vitro experiments in VSMC revealed that cross-linking Efnbs but not Ephb6 resulted in reduced VSMC contractions, suggesting that
reverse signaling through Efnbs was responsible for the observed BP phenotype. The reverse signaling was mediated by an adaptor
protein Grip1. Additional experiments demonstrated decreased 24-h urine catecholamines in male Ephb6 KO mice, probably as
a compensatory feedback mechanism to keep their BP in the normal range. After castration, however, such compensation was abolished
in Ephb6 KO mice and was likely the reason why BP increased overtly in these animals. It suggests that Ephb6 has a target
in the nervous/endocrine system in addition to VSMC, regulating a testosterone-dependent catecholamine compensatory mechanism.
Our study discloses that Ephs and Efns, in concert with testosterone, play a critical role in regulating small artery contractility
and BP.
Journal of Biological Chemistry 02/2012; 287(9):6819-6829. · 4.77 Impact Factor
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ABSTRACT: Pno1 is a protein that plays a role in proteasome and ribosome neogenesis in yeast. So far, its functions in mammalian cells have not been investigated. To understand its function in mammals, we performed in situ hybridization analysis of Pno1 expression in different development stages and generated Pno1 gene knockout (KO) and transgenic (Tg) mice lineages. The results showed early lethality of homozygous Pno1 KO lineage caused, as demonstrated in parallel by ex vivo experiments, by arrest of embryo development before compaction stage. Though, heterozygous (HET) mice with 50% of normal Pno1 mRNA concentration were fertile and showed no obvious anomalies. The lymphoid organs of HET mice were normal in size, weight and cellularity, with normal T and B cell subpopulations. TCR-triggered activation and proliferation of HET T cells were normal. Proteasome activities in HET organs were uncompromised. Tg mice with actin promoter-driven Pno1 expression were also fertile, with no apparent anomalies, although they expressed 2-5-fold higher Pno1 mRNA levels. The lymphoid organs of Tg mice were of normal size, weight and cellularity with normal T and B cell sub-populations. TCR-triggered activation and proliferation of Tg T cells were normal. Tg organs and tissues presented normal proteasome activity as did their wild type counterparts. Tagged Pno1 over-expression in L cells and density gradient fractionation established that Pno1 existed in large complexes with sedimentation rates between 20S and 26S, bigger than mature 26S proteasomes. Pno1 in fractions did not coincide with 40S or 60S ribosome subunits. Our study indicates that Pno1 is essential for cellular functions, but only a small percentage of its normal level is sufficient, and excessive amounts are neither harmful nor useful. The nature of the large complexes it associates with remains to be identified, but it is certain that they are not mature proteasomes or ribosomes.
PLoS ONE 01/2012; 7(9):e46093. · 4.09 Impact Factor
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ABSTRACT: Eph kinases are the largest family of cell surface receptor tyrosine kinases. The ligands of Ephs, ephrins (EFNs), are also cell surface molecules. Ephs interact with EFNs transmitting signals in both directions, i.e., from Ephs to EFNs and from EFNs to Ephs. EFNB1 is known to be able to co-stimulate T cells in vitro and to modulate thymocyte development in a model of foetal thymus organ culture. To further understand the role of EFNB1 in T cell immunity, we generated T-cell-specific EFNB1 gene knockout mice to assess T cell development and function in these mice.
The mice were of normal size and cellularity in the thymus and spleen and had normal T cell subpopulations in these organs. The bone marrow progenitors from KO mice and WT control mice repopulated host spleen T cell pool to similar extents. The activation and proliferation of KO T cells was comparable to that of control mice. Naïve KO CD4 cells showed an ability to differentiate into Th1, Th2, Th17 and Treg cells similar to control CD4 cells.
Our results suggest that the function of EFNB1 in the T cell compartment could be compensated by other members of the EFN family, and that such redundancy safeguards the pivotal roles of EFNB1 in T cell development and function.
BMC Immunology 12/2011; 12:68. · 2.53 Impact Factor
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ABSTRACT: Erythropoietin-producing hepatocellular kinases (Eph kinases) constitute the largest family of cell membrane receptor tyrosine kinases, and their ligand ephrins are also cell surface molecules. Because of promiscuous interaction between Ephs and ephrins, there is considerable redundancy in this system, reflecting the essential roles of these molecules in the biological system through evolution. In this study, both Efnb1 and Efnb2 were null-mutated in the T cell compartment of mice through loxP-mediated gene deletion. Mice with this double conditional mutation (double KO mice) showed reduced thymus and spleen size and cellularity. There was a significant decrease in the DN4, double positive, and single positive thymocyte subpopulations and mature CD4 and CD8 cells in the periphery. dKO thymocytes and peripheral T cells failed to compete with their WT counterparts in irradiated recipients, and the T cells showed compromised ability of homeostatic expansion. dKO naive T cells were inferior in differentiating into Th1 and Th17 effectors in vitro. The dKO mice showed diminished immune response against LCMV infection. Mechanistic studies revealed that IL-6 signaling in dKO T cells was compromised, in terms of abated induction of STAT3 phosphorylation upon IL-6 stimulation. This defect likely contributed to the observed in vitro and in vivo phenotype in dKO mice. This study revealed novel roles of Efnb1 and Efnb2 in T cell development and function.
Journal of Biological Chemistry 12/2011; 286(48):41135-52. · 4.77 Impact Factor
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ABSTRACT: IL-7 plays vital roles in thymocyte development, T cell homeostasis, and the survival of these cells. IL-7 receptor α (IL-7Rα) on thymocytes and T cells is rapidly internalized upon IL-7 ligation. Ephrins (Efns) are cell surface molecules and ligands of the largest receptor kinase family, Eph kinases. We discovered that T cell-specific double gene knock-out (dKO) of Efnb1 and Efnb2 in mice led to reduced IL-7Rα expression in thymocytes and T cells, and that IL-7Rα down-regulation was accelerated in dKO CD4 cells upon IL-7 treatment. On the other hand, Efnb1 and Efnb2 overexpression on T cell lymphoma EL4 cells retarded IL-7Rα down-regulation. dKO T cells manifested compromised STAT5 activation and homeostatic proliferation, an IL-7-dependent process. Fluorescence resonance energy transfer and immunoprecipitation demonstrated that Efnb1 and Efnb2 interacted physically with IL-7Rα. Such interaction likely retarded IL-7Rα internalization, as Efnb1 and Efnb2 were not internalized. Therefore, we revealed a novel function of Efnb1 and Efnb2 in stabilizing IL-7Rα expression at the post-translational level, and a previously unknown modus operandi of Efnbs in the regulation of expression of other vital cell surface receptors.
Journal of Biological Chemistry 11/2011; 286(52):44976-87. · 4.77 Impact Factor
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ABSTRACT: TGF-βi is a secreted protein and is capable of binding to both extracellular matrix (ECM) and cells. It thus acts as a bifunctional molecule enhancing ECM and cell interactions, a lack of which results in dysfunction of many cell types. In this study, we investigated the role of TGF-βi in the function and survival of islets. Based on DNA microarray followed by quantitative PCR confirmation, TGFβi gene showed drastic increase in expression in islets after culture. We demonstrated that recombinant TGF-βi could preserve the integrity and enhance the function of cultured islets. Such a beneficial effect was mediated via signaling through FAK. Exogenous TGF-βi was capable of sustaining high-level FAK phosphorylation in isolated islets, and FAK knockdown by small interfering RNA in islets resulted in compromised islet function. TGF-βi transgenic (Tg) islets showed better integrity and insulin release after in vitro culture. In vivo, β-cell proliferation was detectable in Tg but not wild-type pancreata. At age above 12 mo, Tg pancreata contained giant islets. Tg mice displayed better glucose tolerance than that of the controls. Tg islets were more potent in lowering blood glucose when transplanted into syngeneic mice with streptozotocin-induced diabetes, and these transplanted islets also underwent regeneration. Our results indicate that TGF-βi is a vital trophic factor promoting islet survival, function, and regeneration. At least some of its beneficial effect was mediated by signaling through FAK.
The Journal of Immunology 04/2011; 186(10):5833-44. · 5.79 Impact Factor
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ABSTRACT: Ran (Ras-related nuclear protein), a Ras family GTPase, is involved in multiple cellular functions, including the regulation of DNA replication, cell cycle progression, nuclear structure formation, RNA processing-exportation, and nuclear protein importation. Ran(+/-) embryonic stem (ES) cells were produced in an attempt to generate Ran null mutant mice. Even after an extremely large number of blastocyst injections, no Ran(+/-) chimeric mice could be generated. Ran(+/-) ES cell-derived fibroblasts showed reduced Ran protein expression, and manifested augmented nuclear abundance of AP-1 factors (c-Jun and c-Fos) upon cytokine stimulation. Our experiments demonstrated that intracellular Ran protein levels controlled the nuclear presence of certain transcription factors, such as c-Fos and c-Jun.
FEBS letters 10/2010; 584(22):4623-6. · 3.54 Impact Factor
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ABSTRACT: Ras-related nuclear protein (Ran) is a Ras family GTPase, and its documented functions are the regulation of DNA replication, cell cycle progression, nuclear structure formation, RNA processing and exportation, and nuclear protein importation. In this study, we performed detailed mapping of Ran expression during mouse ontogeny using in situ hybridization. High Ran expression was found in various organs and tissues including the thymus cortex and spleen white pulp. Ran was induced in T cells 24 h after their activation. The function of Ran in the immune system was investigated using Ran transgenic (Tg) mice. In Ran Tg T cells, there was compromised activation marker expression, lymphokine secretion, and proliferation upon T cell receptor activation in vitro when compared with wild type T cells. Tg mice also manifested defective delayed type hypersensitivity in vivo. Upon PMA and ionomycin stimulation, Tg T cells were defective in nuclear accumulation of AP-1 factors (c-Jun and c-Fos) but not NF-kappaB family members. Our experiments showed that Ran had important regulatory function in T cell activation. One of the possible mechanisms is that intracellular Ran protein levels control the nuclear retention for selective transcription factors such as c-Jun and c-Fos of AP-1, which is known to be critical in T cell activation and proliferation and lymphokine secretion.
Journal of Biological Chemistry 12/2009; 285(8):5488-96. · 4.77 Impact Factor
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ABSTRACT: TNF-like ligand 1A (TL1A), a member of the TNF superfamily, is the ligand of DR3 and DcR3. Several types of cells, such as endothelial cells, monocytes/macrophages, dendritic cells, and CD4 and CD8 T cells, are capable of producing this cytokine. In present study, we demonstrated that TL1A aggravated collagen-induced arthritis in mice. It increased collagen-induced arthritis penetrance and clinical scores as well as the severity of the pathological findings. TL1A administration led to the occurrence of multiple enlarged germinal centers in the spleen, and it boosted serum anti-collagen Ab titers in vivo. In vitro, TL1A augmented TNF-alpha production by T cells upon TCR ligation, and it greatly enhanced Th17 differentiation and IL-17 production. We further showed that human rheumatoid arthritis (RA) synovial fluids had elevated TL1A titers, and human chrondrocytes and synovial fibroblasts were capable of secreting TL1A upon TNF-alpha or IL-1beta stimulation. Taken together, these data suggest that TL1A secretion in lymphoid organs might contribute to RA initiation by promoting autoantibody production, and TL1A secretion stimulated by inflammatory cytokines in RA joints might be a part of a vicious circle that aggravates RA pathogenesis.
The Journal of Immunology 10/2009; 183(8):5350-7. · 5.79 Impact Factor
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ABSTRACT: Drak2 is a member of the death-associated protein family and a serine threonine kinase. In this study, we investigated its role in beta cell survival and diabetes. Drak2 mRNA and protein were rapidly induced in islet beta cells after stimulation by inflammatory lymphokines known to be present in type 1 diabetes. Drak2 up-regulation was accompanied by increased beta cell apoptosis. beta cell apoptosis caused by the said stimuli was inhibited by Drak2 knockdown using small interfering RNA. Conversely, transgenic Drak2 overexpression led to aggravated beta cell apoptosis triggered by the stimuli. Further in vivo experiments demonstrated that Drak2 transgenic islets were more vulnerable to streptozocin insult. We established that inducible NO synthase was upstream and caspase-9 was downstream of Drak2 in its signaling pathway. Purified Drak2 could phosphorylate ribosomal protein S6 (p70S6) kinase in an in vitro kinase assay. Drak2 overexpression in NIT-1 cells led to enhanced p70S6 kinase phosphorylation, whereas Drak2 knockdown in these cells reduced it. These mechanistic studies proved that p70S6 kinase was a bona fide Drak2 substrate.
The Journal of Immunology 05/2009; 182(8):4762-70. · 5.79 Impact Factor
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ABSTRACT: Drak2 is a serine threonine kinase in the death-associated protein family. In this study, we investigated its role in free fatty acid (FFA)-induced islet apoptosis. Drak2 mRNA and protein were rapidly induced in islet beta-cells after FFA stimulation. Such Drak2 upregulation was accompanied by increased beta-cell apoptosis, which was inhibited by Drak2 knockdown using siRNA. Conversely, transgenic (Tg) Drak2 overexpression led to aggravated beta-cell apoptosis triggered by FFA. Drak2 overexpression in islets compromised the increase of anti-apoptotic factors, such as Bcl-2, Bcl-xL and Flip, upon FFA assault. Further in vivo experiments demonstrated that Drak2 Tg mice presented compromised glucose tolerance in a diet-induced obesity model. Our data show that Drak2 is detrimental to islet survival in the presence of excessive lipid.
Journal of Cellular Biochemistry 10/2008; 105(4):1073-80. · 2.87 Impact Factor
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ABSTRACT: In this study, we investigated the diagnostic value of serum death decoy receptor 3 (DcR3) for systemic lupus erythematosus (SLE). The possible pathogenic role of DcR3 in SLE was also assessed. Serum DcR3 levels of 90 SLE patients, 11 patients with rheumatic conditions and 123 healthy controls were determined by ELISA. In all, 43% of the SLE patients, 9% of patients with rheumatic conditions and 2.4% of the normal healthy individuals presented elevated serum DcR3 levels. A higher percentage of DcR3-positive SLE patients, compared with DcR3-negative SLE patients, showed abnormally high serum IgE levels, a surrogate marker of T(h)2-type immune responses. To determine the cause and effect relationship of DcR3 expression and a T(h)2-prone status, we studied young DcR3 transgenic (Tg) mice, whose transgene was driven by an actin promoter. These mice had IL-4 overproduction and augmented serum IgE levels, signs of dominant T(h)2 immune responses. To determine possible SLE pathogenic roles of DcR3, the T-cell-depleted bone marrow of DcR3 Tg mice was transplanted into lethally irradiated syngeneic C57BL/6 female mice. The recipients developed an SLE-like syndrome. They presented anti-dsDNA and anti-nuclear antibodies, along with renal and liver pathology compatible with that of SLE. In total, 90% of Tg bone marrow-transplanted mice, compared with 20% of wild-type bone marrow-transplanted mice, perished within 12 months after the transplantation. Our results showed that serum DcR3 could serve as an additional parameter for SLE diagnosis and that DcR3 secreted from cells of hematopoietic origin was SLE pathogenic in mice.
International Immunology 07/2008; 20(8):1067-75. · 3.41 Impact Factor
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ABSTRACT: B-cell translocation gene 2 (BTG2) belongs to the anti-proliferative gene family. According to previous in vitro studies, BTG2 overexpression leads to delayed cell cycling. We investigated BTG2 expression during mouse ontogeny and its immune and circadian functions in this study. In situ hybridization showed that BTG2 was expressed at high levels in the central nervous system, liver, stomach, thymus, spleen, skin, adrenal gland, pituitary gland and salivary glands during embryonic days (E10-E17), postnatal days (P1 and P10) and adult stages. Expression was observed in organs and tissues from adult mice with and without a robust proliferation program. Thus, the gene might have important functions that are both related and unrelated to proliferation. BTG2 expression was induced after in vitro T-cell receptor stimulation in T cells using anti-CD3 antibodies. However, transgenic (Tg) mice with actin promoter-driven expression of BTG2 showed normal in vitro and in vivo T-cell responses, such as thymus development, T-cell activation marker expression, T-cell proliferation and migration, as well as in vivo delayed-type hypersensitivity reactions. Although BTG2 was expressed in the suprachiasmatic nucleus and pineal gland in the brain, BTG2 Tg mice had no abnormal circadian behavior. Our data on BTG2 expression during ontogeny provide useful clues for the further investigation of BTG2 function. Additional studies are warranted to examine its role in immune and other systems.
International Immunology 04/2008; 20(3):317-26. · 3.41 Impact Factor
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ABSTRACT: Decoy receptor 3 (DcR3), a tumor necrosis factor receptor family member, is a secreted protein that can enhance cell survival by interfering with multiple apoptosis pathways. This study was undertaken to investigate the role of DcR3 in the pathogenesis of autoimmune disease.
We generated transgenic mice with actin promoter-driven expression of human DcR3 and investigated the development of autoimmune disease in these mice.
T cell immune responses were compromised in young DcR3-transgenic mice. Beyond 5-6 months of age, transgenic mice developed a systemic lupus erythematosus (SLE)-like syndrome, with numerous features of the disease. They produced autoantibodies against double-stranded DNA. Their kidneys showed pathologic changes indicative of glomerular nephritis and IgG and C3 deposition, and proteinuria, leukocyturia, and hematuria, were evident. Aged transgenic mice also developed skin lesions and lymphocyte infiltration in the liver, and exhibited leukopenia, anemia, and thrombocytopenia. The SLE-like syndrome penetrance in DcR3-transgenic mice was sex associated, occurring in approximately 60% of females versus 20% of males. Exogenous recombinant DcR3 or endogenous DcR3 produced by transgenic T cells effectively protected T cells against activation-induced apoptosis in vitro. Probably as a consequence of this, CD4 cells with a phenotype of previous activation were increased in the peripheral blood of transgenic mice beyond 6 months of age.
These results show that DcR3 overexpression could lead to an SLE-like syndrome in mice.
Arthritis & Rheumatism 12/2007; 56(11):3748-58. · 7.87 Impact Factor
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ABSTRACT: Riken 2810430M08 is a gene with unknown functions. According to in situ hybridization (ISH), it presented a pattern of temporal expression, peaking in the mid-gestation (embryonic Days e9-e14) stage in most tissues. In the late-gestation stage and during the adulthood, Riken 2810430M08 was expressed in some tissues with and without a robust proliferation program. Thus, the gene might have important functions that are related and unrelated to proliferation. Its expression was induced after T-cell receptor stimulation in T cells. However, transgenic mice with actin promoter-driven expression of Riken 2810430M08 showed normal in vitro and in vivo T cells responses, such as fetal thymus development, adult T cells activation marker expression, lymphokine secretion, proliferation and migration, and delayed type hypersensitivity (DTH). The expression of Riken 2810430M08 during ontogeny has provided useful clues for further investigation. Additional studies are warranted to examine its role in immune and other systems.
Journal of Cellular Biochemistry 06/2007; 101(1):89-98. · 2.87 Impact Factor
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Suzy D C Bianco,
Ji-Bin Peng,
Hitomi Takanaga,
Yoshiro Suzuki,
Alessandra Crescenzi,
Claudine H Kos,
Liyan Zhuang,
Michael R Freeman,
Cecilia H A Gouveia,
Jiangping Wu, Hongyu Luo,
Theodora Mauro,
Edward M Brown,
Matthias A Hediger
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ABSTRACT: We report the phenotype of mice with targeted disruption of the Trpv6 (Trpv6 KO) epithelial calcium channel. The mice exhibit disordered Ca(2+) homeostasis, including defective intestinal Ca(2+) absorption, increased urinary Ca(2+) excretion, decreased BMD, deficient weight gain, and reduced fertility. Although our Trpv6 KO affects the closely adjacent EphB6 gene, the phenotype reported here is not related to EphB6 dysfunction. INTRODUCTIOn: The mechanisms underlying intestinal Ca(2+) absorption are crucial for overall Ca(2+) homeostasis, because diet is the only source of all new Ca(2+) in the body. Trpv6 encodes a Ca(2+)-permeable cation channel responsible for vitamin D-dependent intestinal Ca(2+) absorption. Trpv6 is expressed in the intestine and also in the skin, placenta, kidney, and exocrine organs.
To determine the in vivo function of TRPV6, we generated mice with targeted disruption of the Trpv6 (Trpv6 KO) gene.
Trpv6 KO mice are viable but exhibit disordered Ca(2+) homeostasis, including a 60% decrease in intestinal Ca(2+) absorption, deficient weight gain, decreased BMD, and reduced fertility. When kept on a regular (1% Ca(2+)) diet, Trpv6 KO mice have deficient intestinal Ca(2+) absorption, despite elevated levels of serum PTH (3.8-fold) and 1,25-dihydroxyvitamin D (2.4-fold). They also have decreased urinary osmolality and increased Ca(2+) excretion. Their serum Ca(2+) is normal, but when challenged with a low (0.25%) Ca(2+) diet, Trpv6 KO mice fail to further increase serum PTH and vitamin D, ultimately developing hypocalcemia. Trpv6 KO mice have normal urinary deoxypyridinoline excretion, although exhibiting a 9.3% reduction in femoral mineral density at 2 months of age, which is not restored by treatment for 1 month with a high (2%) Ca(2+) "rescue" diet. In addition to their deranged Ca(2+) homeostasis, the skin of Trpv6 KO mice has fewer and thinner layers of stratum corneum, decreased total Ca(2+) content, and loss of the normal Ca(2+) gradient. Twenty percent of all Trpv6 KO animals develop alopecia and dermatitis.
Trpv6 KO mice exhibit an array of abnormalities in multiple tissues/organs. At least some of these are caused by tissue-specific mechanisms. In addition, the kidneys and bones of Trpv6 KO mice do not respond to their elevated levels of PTH and 1,25-dihydroxyvitamin D. These data indicate that the TRPV6 channel plays an important role in Ca(2+) homeostasis and in other tissues not directly involved in this process.
Journal of Bone and Mineral Research 03/2007; 22(2):274-85. · 6.37 Impact Factor
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ABSTRACT: Drak2 is a death-associated protein family serine-threonine kinase. Its expression and roles in the immune system were investigated in this study. According to in situ hybridization, Drak2 expression was ubiquitous at the mid-gestation stage in embryos, followed by more focal expression in various organs in the perinatal period and adulthood, notably in the thymus, spleen, lymph nodes, cerebellum, suprachiasmatic nuclei, pituitary, olfactory lobes, adrenal medulla, stomach, skin, and testes. Drak2 transgenic (Tg) mice were generated using the human beta-actin promoter. These Tg mice showed normal T cell versus B cell and CD4 versus CD8 populations in the spleen, but their spleen weight cellularity was lower in comparison with wild type mice. After TCR activation, the proliferation response in Drak2 Tg T cells was normal, although their interleukin (IL)-2 and IL-4 but not interferon-gamma production was augmented. Activated Drak2 Tg T cells demonstrated significantly enhanced apoptosis in the presence of exogenous IL-2. At the molecular level, Drak2 Tg T cells manifested a lower increase of anti-apoptotic factors during activation; such a change probably rendered the cells vulnerable to subsequent IL-2 insults. The compromised apoptosis in Drak2 Tg T cells was associated with reduced numbers of T cells with the memory cell phenotype (CD62L(lo)) and repressed secondary T cell responses in delayed type hypersensitivity. Our study demonstrates that Drak2 expresses in the T cell compartment but is not T cell-specific; it plays critical roles in T cell apoptosis and memory T cell development.
Journal of Biological Chemistry 06/2006; 281(18):12587-95. · 4.77 Impact Factor
