Lesser prevalence of polyps/WNT-dysregulation and concomitant upregulation of gamma-catenin/MYC point to alternate pathways in colorectal cancer in India.

Colorectal cancer (CRC) is known to follow adenoma carcinoma sequence (ACS) in majority of the tumors and the driver variants and associated pathways are well delineated. However, most of the published data are from the west and information in other ethnicities is sparse. We therefore comprehensively evaluated the CRC tumors from Indian ethnicity for the prevalence of ACS. In this cohort study, clinical data of 100,497 patients who attended hospital between 2013 and 2018 were accessed. Tumors from patients (n = 130) with CRC who were treated primarily by surgery were included. DNA and RNA were isolated to assess variants (direct sequencing) and WNT-pathway dysregulation in genes related to ACS. Global gene expression was generated and analyzed on microarrays (Affymetrix; N = 10) and next generation sequencing platforms (Illumina; N = 25). Gene expression at mRNA (qRT-PCR) and protein level (IHC) of JUP/CTNNB1/MYC were assessed. Correlation between expression of JUP and MYC was evaluated by Karl Pearson's correlation coefficient. The prevalence of polyps was 16.75%, while 18.26% variants in APC/CTNNB1, 20.00% in KRAS, and 18.33% WNT dysregulation were noted. Interestingly, 29/60 (48.33%) tumors showed only MYC upregulation with normal APC/CTNNB1 expression. Global gene expression and validation in an independent tumor cohort confirmed concomitant upregulation of JUP (gamma-catenin) & MYC (r = 0.71; p = 0.001) at mRNA and protein in sizeable number of tumors (45/96; 46.88%). Our study provides evidence for limited prevalence of ACS in the Indian ethnicity. Preventive colonoscopies for early identification and management of CRC may not be an effective strategy in this ethnicity.

ß-catenin and γ-catenin are dispensable for T lymphocytes and AML leukemic stem cells.

The ß-catenin transcriptional coregulator is involved in various biological and pathological processes; however, its requirements in hematopoietic cells remain controversial. We re-targeted the Ctnnb1 gene locus to generate a true ß-catenin-null mutant mouse strain. Ablation of ß-catenin alone, or in combination with its homologue γ-catenin, did not affect thymocyte maturation, survival or proliferation. Deficiency in ß/γ-catenin did not detectably affect differentiation of CD4+T follicular helper cells or that of effector and memory CD8+ cytotoxic cells in response to acute viral infection. In an MLL-AF9 AML mouse model, genetic deletion of ß-catenin, or even all four Tcf/Lef family transcription factors that interact with ß-catenin, did not affect AML onset in primary recipients, or the ability of leukemic stem cells (LSCs) in propagating AML in secondary recipients. Our data thus clarify on a long-standing controversy and indicate that ß-catenin is dispensable for T cells and AML LSCs.

Effects of quercetin, a natural phenolic compound, in the differentiation of human mesenchymal stem cells (MSC) into adipocytes and osteoblasts.

Natural phenols may have beneficial properties against oxidative stress, which is associated with aging and major chronic aging-related diseases, such as loss of bone mineral mass (osteoporosis) and diabetes. The main aim of this study was to analyze the effect of quercetin, a major nutraceutical compound present in the "Mediterranean diet", on mesenchymal stem-cell (MSC) differentiation. Such cells were induced to differentiate into osteoblasts or adipocytes in the presence of two quercetin concentrations (0.1 and 10µM). Several physiological parameters and the expression of osteoblastogenesis and adipogenesis marker genes were monitored. Quercetin (10µM) inhibited cell proliferation, alkaline phosphatase (ALPL) activity and mineralization, down-regulating the expression of ALPL, collagen type I alpha 1 (COL1A1) and osteocalcin [bone gamma-carboxyglutamate protein (BGLAP)] osteoblastogenesis-related genes in MSC differentiating into osteoblasts. Moreover, in these cultures, CCAAT/enhancer-binding protein alpha (CEBPA) and peroxisome proliferator-activated receptor gamma 2 (PPARG2) adipogenic genes were induced, and cells differentiated into adipocytes were observed. Quercetin did not affect proliferation, but increased adipogenesis, mainly at 10-µM concentration in MSC induced to differentiate to adipocytes. ß- and γ-catenin (plakoglobin) nuclear levels were reduced and increased, respectively, in quercetin-treated cultures. This suggests that the effect of high concentration of quercetin on MSC osteoblastic and adipogenic differentiation is mediated via Wnt/ß-catenin inhibition. In conclusion, quercetin supplementation inhibited osteoblastic differentiation and promoted adipogenesis at the highest tested concentration. Such possible adverse effects of high quercetin concentrations should be taken into account in nutraceutical or pharmaceutical strategies using such flavonol.

Mechanisms underlying the reversal of acantholysis in pemphigus by a cholinergic receptor agonist / 中华皮肤科杂志

Objective To evaluate the reversal effect of a cholinergic receptor agonist on acantholysis in pemphigus,and to investigate its mechanism.Methods Human HaCaT keratinocytes were co-cultured with pemphigus vulgaris immunoglobulin G (PV-IgG) to establish a cell model of pemphigus,then classified into two groups to be incubated with the cholinergic receptor agonist carbachol for 12 hours (test group) or remain untreated (control group).Cell dissociation assay was performed to quantitatively estimate the reversal effect of carbachol on acantholysis,and immunofluorescence assay to qualitatively assess the changes of desmosomal proteins.Radio-immunoprecipitation assay (RIPA) lysis buffer and Triton X-100 were used to lyse HaCaT cells to obtain total proteins and cytoplasmic proteins,and Western blot was conducted to determine the expression levels of adhesion-related proteins desmoglein 3 (Dsg3) and plakoglobin (PG) on the surface of HaCaT cells,as well as the phosphorylation levels of p38 mitogen activated protein kinase (p38 MAPK) and epidermal growth factor receptor (EGFR) at different time points.Quantitative polymerase chain reaction (qPCR) was performed to detect the mRNA expressions of the above surface proteins,and coimmunoprecipitation assay to qualitatively evaluate the interaction between Dsg3 and PG.Results The number of cell debris was significantly lower in the test group than in the control group (18.67 ± 2.52 vs.46.67 ± 2.03,t =11.22,P＜0.01).Immunofluorescence assay showed that carbachol could reverse the internalization of desmosomal molecules induced by PV-IgG.In the pemphigus cell model,the levels of total Dsg3 and PG as well as non-desmosomal Dsg3 were decreased,while the level of non-desmosomal PG increased,and the interaction between Dsg3 and PG was attenuated.When the pemphigus cell model was co-cultured with carbachol,these above changes were reversed.Carbachol also increased the mRNA levels (expressed as 2-△△Ct) of Dsg3 and PG from 1.428 ± 0.215 and 1.563 ± 0.247 in the control group to 4.974 ± 0.948 (t =3.65,P =0.01) and 13.420 ± 1.715 (t =6.85,P ＜ 0.01) in the test group respectively.In phosphorylation assay,carbachol inhibited the phosphorylation of EGFR,but had no significant effect on that of p38 MAPK.Conclusions The cholinergic receptor agonist carbachol can reverse acantholysis in pemphigus,likely by inhibiting the internalization of Dsg3 and PG,enhancing their expressions and interaction,and suppressing the phosphorylation of the key signaling molecule for acantholysis,EGFR.

Expression of E-cadherin and alpha - , beta - , gamma - catenin proteins in endometrial carcinoma / 대한산부인과학회잡지

OBJECTIVES: E-cadherin is a transmembrane protein that is one of the key players involved in cell to cell adhesion. Loss of E-cadherin expression is suggested to promote tumor invasion and distant metastasis in tumor development. Recently, it has been proposed E-cadherin function requires its linkage to the cytoskeleton through catenins. So defects in catenins may cause defective E-cadherin function and promote tumor invasion. We intend to evaluate the expression of E-cadherin and alpha-, beta-, gamma- catenin in tissues of human endometrial carcinoma to analyze the patterns of cell adhesion molecules' expression in endometrial carcinoma and to investigate the relationship between status of cell adhesion molecules and various clinicopathological factors. MATERIALS AND METHODS: The present study investigated the immunohistochemical expression of E-cadherin and alpha-, beta-, gamma- catenin in 33 paraffin embedded formalin fixed tissues of endometrial carcinomas. RESULTS: Aberrant E-cadherin, alpha-, beta-, gamma- catenin expression was observed in 33.3(11 of 33), 27.3(9 of 33), 18.2 (6 of 33), and 51.5(17 of 33) % of the specimens, respectively. Statistically significant correlation was found between aberrant expression of E-cadherin and lymph node metastasis and cell types other than endometrioid adenocarcinoma. Aberrant pattern of gamma- catenin expression also correlated with deep myometrial invasion. But alpha-, beta- catenin expression were not correlated with any clinicopathological parameters. Using Kaplan-Meier curves, abnormal expression of E-cadherin correlated closely with poor survival (p<0.05). CONCLUSION: We revealed aberrant expression of these cell adhesion molecules in part of patients with endometrial carcinoma. Aberrant expression of E-cadherin was correlated with lymph node metastasis and cell types other than endometrioid adenocarcinoma and aberrant expression of gamma-catenin was related with deep myometrial invasion.

The expression and tyrosine phosphorylation of E-cadherin, beta-, gamma-catenin and EGFR after treatment of EGF and TGF-alpha in Cervical Cancer Cell Lines / 대한부인종양콜포스코피학회잡지

OBJECTIVES: Cadherin/catenin adhesion complex is fundamentally involved in epithelial cancer invasion and metastasis. E-cadherin and EGFR colocalize on the basolateral membrane of epithelial cell and EGF down-regulate E-cadherin expression. In the invasion and metastasis of cancer, E-cadherin expression is decreased and growth factors receptor is overexpressed. The present study was aimed to find the role of E-cadherin, beta-and gamma-catenin, growth factors and its receptors in cervical cancer cell lines. METHODS: The cervical cancer cell cultures were treated with different time duration of EGF 30 ng/ml and TGF-a 10 ng/ml(0, 10 min, 20 min, 30 min, 1 hr, 2 hr, 4 hr, 8 hr, 24 hr). The change in cancer cell morphology and the changes in E-cadherin, beta- and gamma-catenin, EGFR and activated EGFR expression were studied with a western blot analysis and an immunoprecipitation. RESULTS: Through a western blot analysis, E-cadherin 120 kDa band and EGFR 170 kDa band were expressed in CaSki, HT-3 and ME-180 cell line, which showed epithelial contact growth. 1n these 3 cell lines, expression of E-cadherin did not decrease with time dependent manner. after the treatment of EGF and TGF- alpha. The expression of EGFR decreased and activated EGFR expression increased in 30 minutes to 1 hour but decreased subsequently. When the cells treated with EGF, there were no change in beta-and gamma-catenin expression with there dependent manner. The tyrosine phosphorylation of beta-and gamma-catenin increased in 30 minutes to 1 hour but decreased subsequently with activated EGFR. CONCLUSION: This study showed that an activated EGFR which has involved with tyrosine phosphorylation of beta- and gamma-catenin influenced by growth factors rather than expression of E-cadherin, has a role in the invasion and metastasis of the cervical cancer.