[Show abstract][Hide abstract] ABSTRACT: Yes-associated protein 1 (YAP1) is a transcriptional coactivator in the Hippo signaling pathway. Increased YAP1 activity promotes the growth of tumors, including that of colorectal cancer (CRC). Verteporfin, a drug that enhances phototherapy to treat neovascular macular degeneration, is an inhibitor of YAP1. We found that verteporfin inhibited tumor growth independently of its effects on YAP1 or the related protein TAZ in genetically or chemically induced mouse models of CRC, in patient-derived xenografts, and in enteroid models of CRC. Instead, verteporfin exhibited in vivo selectivity for killing tumor cells in part by impairing the global clearance of high-molecular weight oligomerized proteins, particularly p62 (a sequestrome involved in autophagy) and STAT3 (signal transducer and activator of transcription 3; a transcription factor). Verteporfin inhibited cytokine-induced STAT3 activity and cell proliferation and reduced the viability of cultured CRC cells. Although verteporfin accumulated to a greater extent in normal cells than in tumor cells in vivo, experiments with cultured cells indicated that the normal cells efficiently cleared verteporfin-induced protein oligomers through autophagic and proteasomal pathways. Culturing CRC cells under hypoxic or nutrient-deprived conditions (modeling a typical CRC microenvironment) impaired the clearance of protein oligomers and resulted in cell death, whereas culturing cells under normoxic or glucose-replete conditions protected cell viability and proliferation in the presence of verteporfin. Furthermore, verteporfin suppressed the proliferation of other cancer cell lines even in the absence of YAP1, suggesting that verteporfin may be effective against multiple types of solid cancers.
[Show abstract][Hide abstract] ABSTRACT: MDI 301 is a novel 9-cis retinoic acid derivative in which the terminal carboxylic acid group has been replaced by a picolinate ester. MDI 301, a retinoic acid receptor-α - agonist, suppressed the growth of several human myeloid leukemia cell lines (HL60, NB4, OCI-M2, and K562) in vitro and induced cell-substrate adhesion in conjunction with upregulation of CD11b. Tumor growth in HL60-injected athymic nude mice was reduced. In vitro, MDI 301 was comparable to all-trans retinoic acid (ATRA) whereas in vivo, MDI 301 was slightly more efficacious than ATRA. Most importantly, unlike what was found with ATRA treatment, MDI 301 did not induce a cytokine response in the treated animals and the severe inflammatory changes and systemic toxicity seen with ATRA did not occur. A retinoid with these characteristics might be valuable in the treatment of promyelocytic leukemia, or, perhaps, other forms of myeloid leukemia.
[Show abstract][Hide abstract] ABSTRACT: The oncofetal antigen - immature laminin receptor protein (OFA/iLRP) has been linked to metastatic tumor spread for several years. The present study, in which two highly-specific, high-affinity OFA/iLRP-reactive mouse monoclonal antibodies were examined for ability to suppress tumor cell growth and metastatic spread in the A20 B-cell leukemia model and the B16 melanoma model, provides the first direct evidence that targeting OFA/iLRP with exogenous antibodies can have therapeutic benefit. While the antibodies were modestly effective at preventing tumor growth at the primary injection site, both antibodies strongly suppressed end-organ tumor formation following intravenous tumor cell injection. Capacity of anti-OFA/iLRP antibodies to suppress tumor spread through the blood in the leukemia model suggests their use as a therapy for individuals with leukemic disease (either for patients in remission or even as part of an induction therapy). The results also suggest use against metastatic spread with solid tumors.
[Show abstract][Hide abstract] ABSTRACT: Facial erythema and telangiectasia are commonly associated with the erythematotelangiectatic subtype of rosacea (ETR). It is important for clinicians to recognize that these findings can also be associated with a subtype of photoaging, which we term telangiectatic photoaging (TP).
To demonstrate that ETR and TP are distinct dermatologic disorders.
A case-control observational study comparing clinical, histologic, and gene expression features of 26 participants with ETR, 20 with TP, and 11 age- and sex-matched controls in the Program for Clinical Research in Dermatology at University of Michigan.
Findings of clinical history and examination, light and electron microscopy, immunohistochemical analyses, and real-time quantitative reverse-transcriptase polymerase chain reaction gene expression.
Transient erythema was greater in the ETR group (38% graded moderate to severe) than in the TP (0%; P < .001) and control groups (0%; P = .002). Nontransient erythema was also greater in the ETR group (50% graded moderate to severe) than in the TP (25%; P = .03) and control groups (0%; P < .001). Participants with ETR tended to have erythema and telangiectasia primarily on the central face (79%), whereas those with TP tended to have more lateral involvement (57%; P < .001). Those with ETR had significantly less clinical evidence of photodamage (0% graded 6-8 on a photonumeric scale) than those with TP (40% graded 6-8; P = .01). Histologically, there was less evidence of photodamage in ETR than in TP, which had wispy collagen and solar elastosis surrounding blood vessels. Immunohistologic analysis demonstrated greater geometric mean immunostained area by mast cell tryptase staining in ETR samples (0.018%) than in TP (0.004%; P = .01) or control samples (0.001%; P < .001) but no increase in mast cell number, indicative of greater mast cell degranulation. Gene expression of matrix metalloproteinase-3 was 4-fold greater in ETR samples than in TP samples (P = .004) and 5-fold higher than in control samples (P = .004). Gene expression of the neuropeptides calcitonin gene-related peptide (CGRP-α) and substance P was significantly increased in ETR compared with TP (9-fold [P < .001] and 5-fold [P = .002], respectively) and control samples (10-fold [P < .001] and 28-fold [P < .001], respectively).
Telangiectatic photoaging is characterized by less transient and nontransient erythema, a more lateral distribution of erythema and telangiectasia, less neurogenic mast cell activation, and less MMP-mediated matrix remodeling than ETR. These data demonstrate that TP is a distinct clinical entity from ETR that can be distinguished on the basis of clinical, histologic, and gene expression findings.
[Show abstract][Hide abstract] ABSTRACT: The move away from animal models for skin safety testing is inevitable. It is a question of when, not if. As skin safety studies move away from traditional animal-based approaches, a number of replacement technologies are becoming available. Human skin in organ culture is one such technology. Organ-cultured skin has several features that distinguish it from other technologies. First and foremost, organ-cultured skin is real skin. Almost by definition, therefore, it approximates the intact skin better than other alternative models. Organ culture is an easy-to-use and relatively inexpensive approach to preclinical safety assessment. Although organ culture is not likely to replace high-throughput enzyme assays or monolayer culture/skin equivalent cultures for initial compound assessment, organ culture should find use when the list of compounds to be evaluated is small and when simpler models have narrowed the dose range. Organ-cultured skin also provides a platform for mechanistic studies.
Expert Review of Dermatology 01/2014; 7(3). DOI:10.1586/edm.12.24
[Show abstract][Hide abstract] ABSTRACT: In order to advance a culture model of human colonic neoplasia, we developed methods for the isolation and in vitro maintenance of intact colonic crypts from normal human colon tissue and adenomas. Crypts were maintained in three-dimensional Matrigel culture with a simple, serum-free, low Ca 2+ (0.15 mM) medium. Intact colonic crypts from normal human mucosa were viably maintained for 3-5 days with preservation of the in situ crypt-like architecture, presenting a distinct base and apex. Abnormal structures from adenoma tissue could be maintained through multiple passages (up to months), with expanding buds/tubules. Immunohistochemical markers for intestinal stem cells (Lgr5), growth (Ki67), differentiation (E-cadherin, cytokeratin 20 (CK20) and mucin 2 (MUC2)) and epithelial turnover (Bax, cleaved Caspase-3), paralleled the changes in function. The epithelial cells in normal crypts followed the physiological sequence of progression from proliferation to differentiation to dissolution in a spatially and temporally appropriate manner. Lgr5 expression was seen in a few basal cells of freshly isolated crypts, but was not detected after 1-3 days in culture. After 24 h in culture, crypts from normal colonic tissue continued to show strong Ki67 and MUC2 expression at the crypt base, with a gradual decrease over time such that by days 3-4 Ki67 was not expressed. The differentiation marker CK20 increased over the same period, eventually becoming intense throughout the whole crypt. In adenoma-derived structures, expression of markers for all stages of progression persisted for the entire time in culture. Lgr5 showed expression in a few select cells after months in culture. Ki67 and MUC2 were largely associated with the proliferative budding regions while CK20 was localized to the parent structure. This ex vivo culture model of normal and adenomatous crypts provides a readily accessible tool to help understand the growth and differentiation process in human colonic epithelium.
[Show abstract][Hide abstract] ABSTRACT: Progressive bone mineral loss and increasing bone fragility are hallmarks of osteoporosis. A combination of minerals isolated from the red marine algae, Lithothamnion sp. was examined for ability to inhibit bone mineral loss in female mice maintained on either a standard rodent chow (control) diet or a high-fat western diet (HFWD) for 5, 12, and 18 months. At each time point, femora were subjected to μ-CT analysis and biomechanical testing. A subset of caudal vertebrae was also analyzed. Following this, individual elements were assessed in bones. Serum levels of the 5b isoform of tartrate-resistant acid phosphatase (TRAP) and procollagen type I propeptide (P1NP) were also measured. Trabecular bone loss occurred in both diets (evident as early as 5 months). Cortical bone increased through month 5 and then declined. Cortical bone loss was primarily in mice on the HFWD. Inclusion of the minerals in the diet reduced bone mineral loss in both diets and improved bone strength. Bone mineral density was also enhanced by these minerals. Of several cationic minerals known to be important to bone health, only strontium was significantly increased in bone tissue from animals fed the mineral diets, but the increase was large (5-10 fold). Serum levels of TRAP were consistently higher in mice receiving the minerals, but levels of P1NP were not. These data suggest that trace minerals derived from marine red algae may be used to prevent progressive bone mineral loss in conjunction with calcium. Mineral supplementation could find use as part of an osteoporosis-prevention strategy.
Biological trace element research 10/2013; 156(1). DOI:10.1007/s12011-013-9820-7 · 1.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The extracellular Ca(2+)-sensing receptor (CaSR) is a robust promoter of differentiation in colonic epithelial cells and functions as a tumor suppressor in colon cancer. CaSR mediates its biologic effects through diverse mechanisms. Loss of CaSR expression activates a myriad of stem cell-like molecular features that drive and sustain the malignant and drug-resistant phenotypes of colon cancer. This CaSR-null phenotype, however, is not irreversible and induction of CaSR expression in CaSR-null cells promotes cell death mechanisms and restores drug sensitivity. The CaSR also functions as a tumor suppressor in breast cancer and promotes cellular sensitivity to cytotoxic drugs. BRCA1 and CaSR functions intersect in breast cancer cells, and CaSR activation can rescue breast cancer cells from the deleterious effect of BRCA1 mutations.
[Show abstract][Hide abstract] ABSTRACT: The goal of this study was to determine if a multimineral natural product derived from red marine algae could reduce colon polyp formation in mice on a high-fat diet. C57BL/6 mice were maintained for up to 18 mo either on a high-fat "Western-style" diet or on a low-fat diet (AIN 76A), with or without the multimineral-supplement. To summarize, colon polyps were detected in 22 of 70 mice (31%) on the high-fat diet but in only 2 of 70 mice (3%) receiving the mineral-supplemented high-fat diet (P < 0.0001). Colon polyps were detected in 16 of 70 mice (23%) in the low-fat group; not significantly different from high-fat group but significantly higher than the high-fat-supplemented group (P = 0.0006). This was in spite of the fact that the calcium level in the low-fat diet was comparable to the level of calcium in the high-fat diet containing the multimineral-product. Supplementation of the low-fat diet reduced the incidence to 8 of 70 mice (11% incidence). Taken together, these findings demonstrate that a multimineral natural product can protect mice on a high-fat diet against adenomatous polyp formation in the colon. These data suggest that increased calcium alone is insufficient to explain the lower incidence of colon polyps.
Nutrition and Cancer 10/2012; 64(7):1020-8. DOI:10.1080/01635581.2012.713160 · 2.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Gadolinium, a member of the lanthanoid family of transition metals, interacts with calcium-binding sites on proteins and other biological molecules. The overall goal of the present investigation was to determine if gadolinium could enhance calcium-induced epithelial cell growth inhibition in the colon. Gadolinium at concentrations as low as 1-5 μM combined with calcium inhibits proliferation of human colonic epithelial cells more effectively than calcium alone. Gadolinium had no detectable effect on calcium-induced differentiation in the same cells based on change in cell morphology, induction of E-cadherin synthesis, and translocation of E-cadherin from the cytosol to the cell surface. When the colon epithelial cells were treated with gadolinium and then exposed to increased calcium concentrations, movement of extracellular calcium into the cell was suppressed. In contrast, gadolinium treatment had no effect on ionomycin-induced release of stored intracellular calcium into the cytoplasm. Whether these in vitro observations can be translated into an approach for reducing abnormal proliferation in the colonic mucosa (including polyp formation) is not known. These results do, however, provide an explanation for our recent findings that a multi-mineral supplement containing all of the naturally occurring lanthanoid metals including gadolinium are more effective than calcium alone in preventing colon polyp formation in mice on a high-fat diet.
Biological trace element research 09/2012; 150(1-3). DOI:10.1007/s12011-012-9503-9 · 1.75 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Ulcerative dermatitis (UD) is a common, spontaneous condition in mice with a C57BL/6 background. Although initial lesions may be mild, UD is a progressive disease that often results in ulcerations or debilitating fibrotic contractures. In addition, lesions typically are unresponsive to treatment. Euthanasia is often warranted in severe cases, thereby affecting study outcomes through the loss of research subjects. Because the clinical assessment of UD can be subjective, a quantitative scoring method and documentation of the likely time-frame of progression may be helpful in predicting when animals that develop dermatitis should be removed from a study. Such a system may also be helpful in quantitatively assessing success of various treatment strategies and be valuable to clinical laboratory animal veterinarians. In this 1.5-y, prospective cohort study, we followed 200 mice to monitor the development and course of UD. Mice were examined every 2 wk. A clinical sign (alopecia, pruritus, or peripheral lymphadenopathy) was not identified that predicted development of UD lesions in the subsequent 2-wk period. Once UD developed, pruritus, the character of the lesion (single or multiple crust, coalescing crust, erosion, or ulceration), and the size of the lesion were the only parameters that changed (increased) over the course of the disease. Pruritus was a factor in the rapid progression of UD lesions. We used these findings to develop a quantitative scoring system for the severity of UD. This enhanced understanding of the progression of UD and the quantitative scoring system will enhance the monitoring of UD.
Journal of the American Association for Laboratory Animal Science: JAALAS 09/2012; 51(5):586-93. · 1.12 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We recently show that CCN3 is a counter-regulatory molecule for the pro-fibrotic protein CCN2, and a potentially novel fibrosis therapy. The goal of this study was to assess the role of CCN3 in fibroproliferative/fibrotic responses in human dermal fibroblasts exposed to Omniscan, one of the gadolinium-based contrast agents associated with development of nephrogenic systemic fibrosis (NSF) a rare but life-threatening disease thought to be complication of NMR diagnostics in renal impaired patients. Human dermal fibroblasts were exposed to Omniscan; or to platelet-derived growth factor (PDGF) and transforming growth factor-β (TGF-β) as controls. Proliferation was assessed along with matrix metalloproteinase-1, tissue inhibitor of metalloproteinases-1 and type 1 procollagen in the absence and presence of CCN3. In parallel, CCN3 production was assessed in control and Omniscan-treated cells. The results showed that PDGF stimulated fibroblast proliferation, production of Timp-1 and MMP-1 whereas exogenous CCN3 inhibited, in a dose response manner, cell proliferation (approx. 50 % max.) and production of MMP-1 (approx 35 % max.) but had little effect on TIMP-1. TGF-β stimulated type 1 procollagen production but not proliferation, Timp-1 or MMP-1 compared to non-TGF-ß treated control cells, and CCN3 treatment blocked (approx. 80 % max.) this up-regulation. Interestingly, untreated, control fibroblasts produced high constitutive levels of CCN3 and concentrations of Omniscan that induced fibroproliferative/fibrogenic changes in dermal fibroblasts correspondingly suppressed CCN3 production. The use of PDGF and TGF-β as positive controls, and the study of differential responses, including that to Omniscan itself, provide the first evidence for a role of fibroblast-derived CCN3 as an endogenous regulator of pro-fibrotic changes, elucidating possible mechanism(s). In conclusion, these data support our hypothesis of a role for fibroblast-derived CCN3 as an endogenous regulator of pro-fibrotic changes in these cells, and suggest that CCN3 may be an important regulatory molecule in NSF and a target for treatment in this and other fibrotic diseases.
Journal of Cell Communication and Signaling 05/2012; 6(2):97-105. DOI:10.1007/s12079-012-0164-4
[Show abstract][Hide abstract] ABSTRACT: Goal: The goal of this study was to determine if a multi-mineral natural product derived from the skeletal remains of the red marine algae, Lithothamnion calcareum, could inhibit gastro-intestinal tumor formation. Several dietary features of the western style diet are known to contribute to liver and colon tumorigenesis, but a role for a deficiency in essential minerals has not been explored. Methods: To address this question, C57BL/6 mice were maintained for 5, 12 and 18 months on either a low fat diet or a high fat "Western-style" diet with or without a mineral-rich product, as a dietary supplement. At the end of incubation period, animals were sacrificed and colon polyp formation was assessed by stereomicroscopy and confirmed histologically. Liver masses were also identified grossly and characterized histologically. Results: Liver: While all the livers appeared normal at 5 months, numerous liver masses were visible in the high fat diet mice sacrificed after 12 and 18 months. The majority of masses were in male mice (20 out of 100 males versus 3 out of 100 females, p=0.0002) and in male animals on high fat diet than on low fat diet (15 out of 50 versus 5 out of 50, p=0.0228).The multi-mineral supplement reduced the number of liver masses in male mice (4 of 50 mice in the mineral-supplemented diet groups compared to 16 of 50 mice without the supplement, p=0.0050). Histological evaluation revealed a total of 17 neoplastic lesions (9 adenomas and 8 hepatocellular carcinomas). Out of 8 hepatocellular carcinomas, 7 were found in unsupplemented diet groups and only one in mice on supplemented high fat diet. Colon: High fat diet induced colon polyps in two female mice at 5 months. Several polyps were detected at 12 and 18 months, most of these lesions were found in female mice (34 out of 100 females versus 11 out of 100 males, p=0.0002). Supplement reduced the incidence of polyps in the presence of high-fat from 18 of 50 female mice to 2 of 50 (p<0.0001). Colon polyps were detected in 15 of 50 mice in the low-fat unsupplemented diet group; not significantly different from the non-supplemented high fat group or supplemented low fat group but significantly higher than the high fat supplemented group (p=0.0009). This was in spite of the fact that the calcium level in the low fat diet was comparable to the level of calcium in the high fat supplemented diet. Histologically, there were 8 tumors identified as adenocarcinomas, 7 as adenomas, and 7 as foci of hyperplasia. Out of 8 adenocarcinomas, 7 were found in unsupplemented diet groups, one found in mice on supplemented low fat diet and zero in mineral-rich high fat diet group. Conclusion: These findings demonstrate that a multi-mineral natural product can protect mice on a high fat diet against adenomatous polyp formation in the colon and liver injury. These data suggest that a multi-mineral approach to colon polyp chemoprevention may be more effective than supplementation with calcium alone and the protective role for dietary minerals in liver cancer.
AACR 103rd Annual Meeting 2012-- Mar 31-Apr 4, 2012; 03/2012