Inhibition of IGF2BP1 attenuates the progression of endometriosis through PTBP1.

INTRODUCTION: Endometriosis (EMs), manifested by pain and infertility, is a chronic inflammatory disease. The precise pathophysiology of this disease remains uncertain. Insulin-like growth factor-2 mRNA-binding protein 1 (IGF2BP1) and polypyrimidine tract-binding protein 1 (PTBP1) have both been found to regulate proliferation, apoptosis, and invasion. This study aimed to investigate the effects of IGF2BP1/PTBP1 in treating EMs. MATERIALS AND METHODS: qRT-PCR and western blotting were employed to quantify IGF2BP1 and PTBP1 expression in six patients with EMs (mean age 33.83 years). The correlation analysis, STRING database prediction, and RNA immunoprecipitation were utilized to identify the relationship between IGF2BP1 and PTBP1. Ectopic endometrial volume, weight, HE staining, and IGF2BP1 silencing were utilized to estimate the effects of IGF2BP1 in EMs model rats. qRT-PCR, CCK-8, 5-ethynyl-2'-deoxyuridine (EDU) labeling, Transwell assay, and flow cytometry were utilized to assess the effects of IGF2BP1/PTBP1 on the proliferation, migration, invasion, and apoptosis of ectopic endometrial stromal cells (eESCs). Furthermore, western blotting was employed to evaluate expressions of PCNA, VEGF, and E-cadherin in EMs rats and eESCs. RESULTS: The mRNA and protein levels of IGF2BP1 and PTBP1 in the ectopic and eutopic endometrium of EMs patients were significantly increased. RNA immunoprecipitation revealed a close interaction of IGF2BP1 with PTBP1. Additionally, the endometrial volume, weight, and histopathologic scores in rats were significantly reduced after IGF2BP1 silencing. IGF2BP1 silencing also decreased the expression of PCNA and VEGF, and increased E-cadherin expression in endometrial tissues of EMs rats. Moreover, IGF2BP1 silencing inhibited proliferation, migration, and invasion and promoted apoptosis through PTBP1 in eESCs. CONCLUSIONS: IGF2BP1 exhibits potential beneficial properties in the management of EMs by interacting with PTBP1, thereby highlighting IGF2BP1 as a promising therapeutic target for EMs.

Neurotensin modulates ovarian vascular permeability via adherens junctions.

Neurotensin (NTS) is a 13-amino acid peptide which is highly expressed in the mammalian ovary in response to the luteinizing hormone surge. Antibody neutralization of NTS in the ovulatory follicle of the cynomolgus macaque impairs ovulation and induces follicular vascular dysregulation, with excessive pooling of red blood cells in the follicle antrum. We hypothesize that NTS is an essential intrafollicular regulator of vascular permeability. In the present study, follicle injection of the NTS receptor antagonist SR142948 also resulted in vascular dysregulation. To measure vascular permeability changes in vitro, primary macaque ovarian microvascular endothelial cells (mOMECs) were enriched from follicle aspirates and studied in vitro. When treated with NTS, permeability of mOMECs decreased. RNA sequencing (RNA-Seq) of mOMECs revealed high mRNA expression of the permeability-regulating adherens junction proteins N-cadherin (CDH2) and K-cadherin (CDH6). Immunofluorescent detection of CDH2 and CDH6 confirmed expression and localized these cadherins to the cell-cell boundaries, consistent with function as components of adherens junctions. mOMECs did not express detectable levels of the typical vascular endothelial cadherin, VE-cadherin (CDH5) as determined by RNA-Seq, qPCR, western blot, and immunofluorescence. Knockdown of CDH2 or CDH6 via siRNA abrogated the NTS effect on mOMEC permeability. Collectively, these data suggest that NTS plays an ovulation-critical role in vascular permeability maintenance, and that CDH2 and CDH6 are involved in the permeability modulating effect of NTS on the ovarian microvasculature. NTS can be added to a growing number of angiogenic regulators which are critical for successful ovulation.

Afadin-nectin forces its way to the front.

Force transmission at cell-cell junctions critically regulates embryogenesis, tissue homeostasis, and diseases including cancer. The cadherin-catenin linkage has been considered the keystone of junctional force transmission, but new findings challenge this paradigm, arguing instead that the nectin-afadin linkage plays the more important role in mature junctions in the intestinal epithelium.

[Banxia Xiexin Decoction inhibiting colitis-associated colorectal cancer infected with Fusobacterium nucleatum by regulating Wnt/ß-catenin pathway].

This paper investigates the intervention effect and mechanism of Banxia Xiexin Decoction(BXD) on colitis-associated colorectal cancer(CAC) infected with Fusobacterium nucleatum(Fn). C57BL/6 mice were randomly divided into a control group, Fn group, CAC group [azoxymethane(AOM)/dextran sulfate sodium salt(DSS)](AOM/DSS), model group, and BXD group. Except for the control and AOM/DSS groups, the mice in the other groups were orally administered with Fn suspension twice a week. The AOM/DSS group, model group, and BXD group were also injected with a single dose of 10 mg·kg~(-1) AOM combined with three cycles of 2.5% DSS taken intragastrically. The BXD group received oral administration of BXD starting from the second cycle until the end of the experiment. The general condition and weight changes of the mice were monitored during the experiment, and the disease activity index(DAI) was calculated. At the end of the experiment, the colon length and weight of the mice in each group were compared. Hematoxylin-eosin(HE) staining was used to observe the pathological changes in the colon tissue. Enzyme-linked immunosorbent assay(ELISA) was used to detect the levels of interleukin(IL)-2, IL-4, and IL-6 inflammatory factors in the serum. Immunohistochemistry(IHC) was used to detect the expression of Ki67, E-cadherin, and ß-catenin in the colon tissue. Western blot was used to detect the protein content of Wnt3a, ß-catenin, E-cadherin, annexin A1, cyclin D1, and glycogen synthase kinase-3ß(GSK-3ß) in the colon tissue. The results showed that compared with the control group, the Fn group had no significant lesions. The mice in the AOM/DSS group and model group had decreased body weight, increased DAI scores, significantly increased colon weight, and significantly shortened colon length, with more significant lesions in the model group. At the same time, the colon histology of the model group showed more severe adenomas, inflammatory infiltration, and cellular dysplasia. The levels of IL-4 and IL-6 in the serum were significantly increased, while the IL-2 content was significantly decreased. The IHC results showed low expression of E-cadherin and high expression of Ki67 and ß-catenin in the model group, with a decreased protein content of E-cadherin and GSK-3ß and an increased protein content of Wnt3a, ß-catenin, annexin A1, and cyclin D1. After intervention with BXD, the body weight of the mice increased; the DAI score decreased; the colon length increased, and the tumor decreased. The histopathology showed reduced tumor proliferation and reduced inflammatory infiltration. The levels of IL-6 and IL-4 in the serum were significantly decreased, while the IL-2 content was increased. Meanwhile, the expression of E-cadherin was upregulated, and that of Ki67 and ß-catenin was downregulated. The protein content of E-cadherin and GSK-3ß increased, while that of Wnt3a, ß-catenin, annexin A1, and cyclin D1 decreased. In conclusion, BXD can inhibit CAC infected with Fn, and its potential mechanism may be related to the inhibition of Fn binding to E-cadherin, the decrease in annexin A1 protein level, and the regulation of the Wnt/ß-catenin pathway.

[Downregulation of SETDB1 suppresses epithelial mesenchymal transition and invasion and metastasis in oral cancer via SOX7 methylation].

PURPOSE: To explore the mechanism of SETDB1 inhibiting epithelial mesenchymal transition (EMT),migration and invasion in oral cancer via SOX 7 methylation. METHODS: SETDB1 and SOX7 mRNA and protein expression levels in KB cells of oral cancer and oral mucosal epithelial ATCC cells were determined by qRT-PCR and Western blot (WB). SETDB1 si-RNA was structured, then transfect into KB cells of oral cancer by liposome-mediated method. siRNA-SETDB1 was the experimental group (si-S), siRNA empty vector was the negative control group (si-N), and untransfected KB cells were the blank control group(NC). SETDB1 mRNA and protein expression levels were detected by qRT-PCR and Western blot(WB), to verify the transfection effect. The methylation levels of SOX7 were determined by pyrosequencing. The expression of N-cadherin, Vimentin, ß-catenin, and Slug proteins was detected by WB. Cell viability was measured by MTT assay, migration ability was tested by scratch healing assay, and invasion ability was tested by Transwell chamber assay. Statistical analysis was performed with SPSS 21.0 software package. RESULTS: The results of Rt-qPCR and WB showed that the SETDB1 mRNA and protein expression decreased significantly in si-S group(P＜0.05). Pyrosequencing test results showed that the regulation of SETDB1 could significantly reduce the SOX7 methylation rate and increased the SOX7 protein expression. WB results showed that knockdown of SETDB1 significantly inhibited the expression of EMT-related proteins N-cadherin, Vimentin, ß-catenin and Slug in oral cancer KB cells (P＜0.05). The results of cell functology experiments showed that knockdown of SETDB1 could significantly inhibit survival, migration and invasion of KB cells. CONCLUSIONS: Downregulation of SETDB1 could suppress EMT, migration and invasion of oral cancer cells by regulating SOX7 methylation level, providing new ideas and targets for the diagnosis and treatment of oral cancer.

The secreted protein Amuc_1409 from Akkermansia muciniphila improves gut health through intestinal stem cell regulation.

Akkermansia muciniphila has received great attention because of its beneficial roles in gut health by regulating gut immunity, promoting intestinal epithelial development, and improving barrier integrity. However, A. muciniphila-derived functional molecules regulating gut health are not well understood. Microbiome-secreted proteins act as key arbitrators of host-microbiome crosstalk through interactions with host cells in the gut and are important for understanding host-microbiome relationships. Herein, we report the biological function of Amuc_1409, a previously uncharacterised A. muciniphila-secreted protein. Amuc_1409 increased intestinal stem cell (ISC) proliferation and regeneration in ex vivo intestinal organoids and in vivo models of radiation- or chemotherapeutic drug-induced intestinal injury and natural aging with male mice. Mechanistically, Amuc_1409 promoted E-cadherin/ß-catenin complex dissociation via interaction with E-cadherin, resulting in the activation of Wnt/ß-catenin signaling. Our results demonstrate that Amuc_1409 plays a crucial role in intestinal homeostasis by regulating ISC activity in an E-cadherin-dependent manner and is a promising biomolecule for improving and maintaining gut health.

Expression of HIF-1α, Ki67, SMA and E-cadherin in endometriosis, endometrial and ovarian carcinoma.

INTRODUCTION: Endometriosis is a benign gynecological condition that shares many characteristics with cancer cells, including immune evasion, survival, adhesion, invasion, and angiogenesis. The simultaneous investigation of tissue hypoxia, EMT, and proliferative index in endometriosis, endometrial, and ovarian carcinomas may provide new insight into the evolution and progression of gynecological neoplasms.

Sulforaphane Inhibits Adhesion and Migration of Cisplatin- and Gemcitabine-Resistant Bladder Cancer Cells In Vitro.

Only 20% of patients with muscle-invasive bladder carcinoma respond to cisplatin-based chemotherapy. Since the natural phytochemical sulforaphane (SFN) exhibits antitumor properties, its influence on the adhesive and migratory properties of cisplatin- and gemcitabine-sensitive and cisplatin- and gemcitabine-resistant RT4, RT112, T24, and TCCSUP bladder cancer cells was evaluated. Mechanisms behind the SFN influence were explored by assessing levels of the integrin adhesion receptors ß1 (total and activated) and ß4 and their functional relevance. To evaluate cell differentiation processes, E- and N-cadherin, vimentin and cytokeratin (CK) 8/18 expression were examined. SFN down-regulated bladder cancer cell adhesion with cell line and resistance-specific differences. Different responses to SFN were reflected in integrin expression that depended on the cell line and presence of resistance. Chemotactic movement of RT112, T24, and TCCSUP (RT4 did not migrate) was markedly blocked by SFN in both chemo-sensitive and chemo-resistant cells. Integrin-blocking studies indicated ß1 and ß4 as chemotaxis regulators. N-cadherin was diminished by SFN, particularly in sensitive and resistant T24 and RT112 cells, whereas E-cadherin was increased in RT112 cells (not detectable in RT4 and TCCSup cells). Alterations in vimentin and CK8/18 were also apparent, though not the same in all cell lines. SFN exposure resulted in translocation of E-cadherin (RT112), N-cadherin (RT112, T24), and vimentin (T24). SFN down-regulated adhesion and migration in chemo-sensitive and chemo-resistant bladder cancer cells by acting on integrin ß1 and ß4 expression and inducing the mesenchymal-epithelial translocation of cadherins and vimentin. SFN does, therefore, possess potential to improve bladder cancer therapy.

In vitro antimetastatic potential of pseudolaric acid B in HSC-3 human tongue squamous carcinoma cell line.

OBJECTIVE: Pseudolaric acid B (PAB) is a novel diterpenoid derived from the traditional Chinese medicinal herb Cortex pseudolaricis that exerts anticancer, anti-inflammatory, and immunomodulatory properties. While the anticancer potential of PAB has been studied, its effects on metastasis have not been well-studied. This study aims to determine the inhibitory effects of PAB on HSC-3 human tongue squamous cell carcinoma (TSCC) cell line. DESIGN: Cell viability and soft agar colony formation assays were conducted to assess cellular proliferation and in vitro tumorigenic capacity of TSCC cells, respectively. Additionally, wound healing, transwell migration, and invasion assays were conducted to monitor the aggressive behavior of TSCC cells. Furthermore, Western blotting analysis was conducted to reveal the signaling pathways involved in the modulation of epithelial-mesenchymal transition (EMT). RESULTS: The migratory and invasive capacities of HSC-3 cells were suppressed by PAB irrespective of their proliferation states. PAB's effects on EMT involved upregulation of E-cadherin expression and downregulation of Twist; these were concomitantly accompanied by downregulated phosphorylation of epidermal growth factor receptor (EGFR). CONCLUSIONS: PAB suppresses human TSCC in vitro by regulating Twist/E-cadherin through the EGFR signaling pathway. PAB may have potential as a candidate antimetastatic drug for TSCC treatment.

Comparative Characterization of Human Meibomian Glands, Free Sebaceous Glands, and Hair-Associated Sebaceous Glands Based on Biomarkers, Analysis of Secretion Composition, and Gland Morphology.

Meibomian gland dysfunction (MGD) is one of the main causes of dry eye disease. To better understand the physiological functions of human meibomian glands (MGs), the present study compared MGs with free sebaceous glands (SGs) and hair-associated SGs of humans using morphological, immunohistochemical, and liquid chromatography-mass spectrometry (LCMS)-based lipidomic approaches. Eyelids with MGs, nostrils, lips, and external auditory canals with free SGs, and scalp with hair-associated SGs of body donors were probed with antibodies against cytokeratins (CK) 1, 8, 10, and 14, stem cell markers keratin 15 and N-cadherin, cell-cell contact markers desmoglein 1 (Dsg1), desmocollin 3 (Dsc3), desmoplakin (Dp), plakoglobin (Pg), and E-cadherin, and the tight junction protein claudin 5. In addition, Oil Red O staining (ORO) was performed in cryosections. Secretions of MGs as well as of SGs of nostrils, external auditory canals, and scalps were collected from healthy volunteers, analyzed by LCMS, and the data were processed using various multivariate statistical analysis approaches. Serial sections of MGs, free SGs, and hair-associated SGs were 3D reconstructed and compared. CK1 was expressed differently in hair-associated SGs than in MGs and other free SGs. The expression levels of CK8, CK10, and CK14 in MGs were different from those in hair-associated SGs and other free SGs. KRT15 was expressed differently in hair-associated SGs, whereas N-cadherin was expressed equally in all types of glands. The cell-cell contact markers Dsg1, Dp, Dsc3, Pg, and E-cadherin revealed no differences. ORO staining showed that lipids in MGs were more highly dispersed and had larger lipid droplets than lipids in other free SGs. Hair-associated SGs had a smaller number of lipid droplets. LCMS revealed that the lipid composition of meibum was distinctively different from that of the sebum of the nostrils, external auditory canals, and scalp. The 3D reconstructions of the different glands revealed different morphologies of the SGs compared with MGs which are by far the largest type of glands. In humans, MGs differ in their morphology and secretory composition and show major differences from free and hair-associated SGs. The composition of meibum differs significantly from that of sebum from free SGs and from hair-associated SGs. Therefore, the MG can be considered as a highly specialized type of holocrine gland that exhibits all the histological characteristics of SGs, but is significantly different from them in terms of morphology and lipid composition.

The mechanism of the contribution of ICAM-1 to epithelial-mesenchymal transition (EMT) in bladder cancer.

Bladder cancer is one of the most prevalent cancers worldwide. Moreover, if not optimally treated, bladder cancer is a significant burden on healthcare systems due to multiple recurrences which often require more aggressive therapies. Therefore, targeted anti-cancer therapies, developed based on an in-depth understanding of specific proteins and molecular mechanisms, are promising in cancer treatment. Here, for the first time, we presented the new approaches indicating that intracellular adhesion molecule-1 (ICAM-1) may play a potential role in enhancing therapeutic effectiveness for bladder cancer. In the present study, we presented that ICAM-1 expression as well as its regulation in bladder cancer is strongly correlated with the high expression of N-cadherin. Importantly, the presence of N-cadherin and its regulator-TWIST-1 was abolished when ICAM-1 was silenced. We identified also that ICAM-1 is capable of regulating cellular migration, proliferation, and EMT progression in bladder cancer cells via the N-cadherin/SRC/AKT/GSK-3ß/ß-catenin signaling axis. Therefore, we propose ICAM-1 as a novel metastatic marker for EMT progression, which may also be used as a therapeutic target in bladder cancer.

Inhibition of PD-L1 expression in non-small cell lung cancer may reduce vasculogenic mimicry formation by inhibiting the epithelial mesenchymal transformation process.

Non-small cell lung cancer (NSCLC) is a kind of highly malignant tumor. Studies have shown that Vasculogenic mimicry (VM) may be responsible for dismal prognosis in NSCLC. Immunotherapy with programmed death-1 (PD-1) or programmed death ligand-1 (PD-L1) has significantly altered the treatment of assorted cancers, including NSCLC, but its role and mechanism in the formation of Vasculogenic mimicry (VM) in NSCLC remains unclear. This study aimed to investigate the role of the anti-PD-L1 antibody in the formation of VM in NSCLC and its possible mechanisms. The results showed that anti-PD-L1 antibody therapy could inhibit the growth of NSCLC-transplanted tumors and reduce the formation of VMs. In addition, this study found that anti-PD-L1 antibodies could increase the expression of the epithelial-mesenchymal transition (EMT) related factor E-cadherin. zinc finger E-box binding homeobox 1 (ZEB1) is an important transcription factor regulating EMT. Knocking down ZEB1 could significantly inhibit tumor growth, as well as the expression of VE-cadherin and mmp2, while remarkably increase the expression of E-cadherin. During this process, the formation of VM was inhibited by knowing down ZEB1 in both in vitro and in vivo experiments of the constructed ZEB1 knockdown stable transfected cell strains. Therefore, in this study, we found that anti-PD-L1 antibodies may reduce the formation of VMs by inhibiting the EMT process.

ID3 is a novel target gene of p53 and modulates lung cancer cell metastasis.

The tumor suppressor p53 prevents cancer development by regulating dozens of target genes with diverse biological functions. Although numerous p53 target genes have been identified to date, the dynamics and function of the regulatory network centered on p53 have not yet been fully elucidated. We herein identified inhibitor of DNA-binding/differentiation-3 (ID3) as a direct p53 target gene. p53 bound the distal promoter of ID3 and positively regulated its transcription. ID3 expression was significantly decreased in clinical lung cancer tissues, and was closely associated with overall survival outcomes in these patients. Functionally, ID3 deficiency promoted the metastatic ability of lung cancer cells through its effects on the transcriptional regulation of CDH1. Furthermore, the ectopic expression of ID3 in p53-knockdown cells restored E-cadherin expression. Collectively, the present results demonstrate that ID3 plays a tumor-suppressive role as a downstream effector of p53 and impedes lung cancer cell metastasis by regulating E-cadherin expression.

SERPINB5 promotes colorectal cancer invasion and migration by promoting EMT and angiogenesis via the TNF-α/NF-κB pathway.

This study aimed to investigate the role of SERPINB5 in colorectal cancer (CRC). We established knockdown and overexpression models of SERPINB5 in CRC cells and conducted bioinformatics analysis to assess the clinicopathological significance of SERPINB5 expression in CRC patients. Human CRC cells were transfected with LV-SERPINB5 and sh-SERPINB5 lentivirus for subsequent functional and mechanistic studies. Results showed that high SERPINB5 expression correlated positively with CEA levels, N stage and lymphatic infiltration, while displaying a negative correlation with progression-free survival. Overexpression of SERPINB5 in CRC cells upregulated the expression of TNF-α, p-NF-κB/p65, N-cadherin, MMP2 and MMP9, accompanied by decreased E-cadherin expression. In addition, SERPINB5 overexpression enhanced the migration, invasion, and proliferation of CRC cells. Furthermore, overexpression of SERPINB5 in CRC cells increased VEGFA expression, and the conditioned medium from SERPINB5-overexpressing CRC cells promoted tube formation of HUVECs. Conversely, overexpression of SERPINB5 in HUVECs decreased VEGFA expression and inhibited tube formation. Notably, these changes in CRC cells were reversed by QNZ, a specific inhibitor of the TNF-α/NF-κB pathway. In summary, our findings revealed that high SERPINB5 expression correlated with poor progression-free survival in CRC patients. Moreover, SERPINB5 could induce EMT and angiogenesis by activating the TNF-α/NF-κB pathway, thereby promoting the invasion and migration of CRC cells.

Cadherin-17 (CDH17) expression in human cancer: A tissue microarray study on 18,131 tumors.

Cadherin-17 (CDH17) is a membranous cell adhesion protein predominantly expressed in intestinal epithelial cells. CDH17 is therefore considered a possible diagnostic and therapeutic target. This study was to comprehensively determine the expression of CDH17 in cancer and to further assess the diagnostic utility of CDH17 immunohistochemistry (IHC). A tissue microarray containing 14,948 interpretable samples from 150 different tumor types and subtypes as well as 76 different normal tissue types was analyzed by IHC. In normal tissues, a membranous CDH17 staining was predominantly seen in the epithelium of the intestine and pancreatic excretory ducts. In tumors, 53 of 150 analyzed categories showed CDH17 positivity including 26 categories with at least one strongly positive case. CDH17 positivity was most common in epithelial and neuroendocrine colorectal neoplasms (50.0%-100%), other gastrointestinal adenocarcinomas (42.7%-61.6%), mucinous ovarian cancer (61.1%), pancreatic acinar cell carcinoma (28.6%), cervical adenocarcinoma (52.6%), bilio-pancreatic adenocarcinomas (40.5-69.8%), and other neuroendocrine neoplasms (5.6%-100%). OnIy 9.9% of 182 pulmonary adenocarcinomas were CDH17 positive. In colorectal adenocarcinomas, reduced CDH17 staining was linked to high pT (p = 0.0147), nodal metastasis (p = 0.0041), V1 (p = 0.0025), L1 (p = 0.0054), location in the right colon (p = 0.0033), and microsatellite instability (p < 0.0001). The CDH17 expression level was unrelated to tumor phenotype in gastric and pancreatic cancer. In summary, our comprehensive overview on CDH17 expression in human tumors identified various tumor entities that might often benefit from anti-CDH17 therapies and suggest utility of CDH17 IHC for the distinction of metastatic gastrointestinal or bilio-pancreatic adenocarcinomas (often positive) from primary pulmonary adenocarcinomas (mostly negative).

Cholangiocyte organoids to study drug-induced injury.

BACKGROUND: Drug induced bile duct injury is a frequently observed clinical problem leading to a wide range of pathological features. During the past decades, several agents have been identified with various postulated mechanisms of bile duct damage, however, mostly still poorly understood. METHODS: Here, we investigated the mechanisms of chlorpromazine (CPZ) induced bile duct injury using advanced in vitro cholangiocyte cultures. Intrahepatic cholangiocyte organoids (ICOs) were driven into mature cholangiocyte like cells (CLCs), which were exposed to CPZ under cholestatic or non-cholestatic conditions through the addition of a bile acid cocktail. RESULTS: CPZ caused loss of monolayer integrity by reducing expression levels of tight junction protein 1 (TJP1), E-cadherin 1 (CDH1) and lysyl oxidase homolog 2 (LOXL2). Loss of zonula occuludens-1 (ZO-1) and E-cadherin was confirmed by immunostaining after exposure to CPZ and rhodamine-123 leakage further confirmed disruption of the cholangiocyte barrier function. Furthermore, oxidative stress seemed to play a major role in the early damage response by CPZ. The drug also decreased expression of three main basolateral bile acid transporters, ABCC3 (ATP binding cassette subfamily C member 3), SLC51A/B (solute carrier family 51 subunit alpha/beta) and multidrug resistance transporter ABCB1 (ATP binding cassette subfamily B member 1), thereby contributing to bile acid accumulation. CPZ did not induce an inflammatory response by itself, but addition of TNFα revealed a synergistic effect. CONCLUSION: These results show that ICOs present a model to identify toxic drugs affecting the bile ducts while providing mechanistic insights into hepatotoxicity.

Melatonin serves as a novel treatment in bladder fibrosis through TGF-ß1/Smad and EMT.

BACKGROUND: Melatonin (MEL) is an indole amine molecule primarily produced in the pineal gland. Melatonin has been shown in numerous studies to have antifibrotic effects on the kidney, liver, and other organs. However, it is still unclear how melatonin works in bladder fibrosis. We explored how melatonin affects animals with bladder fibrosis and the underlying mechanisms. MATERIALS AND METHODS: MEL was used to treat human bladder smooth muscle cells (HBdSMCs) after they were stimulated with transforming growth factor-ß1 (TGF-ß1) in vitro. Proteomic analysis and bioinformatic analysis of the altered expression of these proteins were subsequently performed on HBdSMCs from the different processing methods. To construct an in vivo bladder fibrosis model, we injected protamine sulfate (PS) and lipopolysaccharide (LPS) twice a week into the rat bladder for six weeks. After two weeks of PS/LPS treatment, the mice in the treatment group were treated with MEL (20 mg/kg/d) for 4 weeks. Finally, we detected the expression of fibrosis markers from different perspectives. The TGF-ß1/Smad pathway and epithelial-mesenchymal transition (EMT) in cell and bladder tissues were also identified. Further proteomic analysis was also performed. RESULTS: In vitro, we found that TGF-ß1 treatment enhanced the expression of the fibrosis markers collagen III and α-SMA in HBdSMCs. E-cadherin expression decreased while the TGF-ß1/Smad pathway was activated. Vimentin and N-cadherin expression was also elevated at the same time. Similar findings were observed in the LPS group. After MEL treatment, the expression of collagen III and α-SMA decreased, the expression of E-cadherin increased, and the expression of vimentin and N-cadherin also decreased. According to our quantitative proteomics analysis, CCN1 and SQLE may be important proteins involved in the development of bladder fibrosis. MEL decreased the expression of these genes, leading to the relief of bladder fibrosis. Bioinformatics analysis revealed that the extracellular space structure related to metabolic pathways, actin filament binding, and stress fibers can serve as a pivotal focus in the management of fibrosis. CONCLUSION: Melatonin attenuates bladder fibrosis by blocking the TGF-ß1/Smad pathway and EMT. CCN1 appears to be a possible therapeutic target for bladder fibrosis.

D-beta-hydroxybutyrate up-regulates Claudin-1 and alleviates the intestinal hyperpermeability in lipopolysaccharide-treated mice.

The hyperpermeability of intestinal epithelium is a key contributor to the occurrence and development of systemic inflammation. Although D-beta-hydroxybutyrate (BHB) exhibits various protective effects, whether it affects the permeability of intestinal epithelium in systemic inflammation has not been clarified. In this study, we investigated the effects of BHB on the intestinal epithelial permeability, the epithelial marker E-cadherin and the tight junction protein Claudin-1 in colon in the lipopolysaccharide (LPS)-induced systemic inflammation mouse model. Intraperitoneal injection of LPS was used to induce systemic inflammation and BHB was given by oral administration. The permeability of intestinal epithelium, the morphological changes of colonic epithelium, the distribution and generation of colon E-cadherin, and the Claudin-1 generation and its epithelial distribution in colon were detected. The results confirmed the intestinal epithelial hyperpermeability and inflammatory changes in colonic epithelium, with disturbed E-cadherin distribution in LPS-treated mice. Besides, colon Claudin-1 generation was decreased and its epithelial distribution in colon was weakened in LPS-treated mice. However, BHB treatments alleviated the LPS-induced hyperpermeability of intestinal epithelium, attenuated the colonic epithelial morphological changes and promoted orderly distribution of E-cadherin in colon. Furthermore, BHB up-regulated colon Claudin-1 generation and promoted its colonic epithelial distribution and content in LPS-treated mice. In conclusion, BHB may alleviate the hyperpermeability of intestinal epithelium via up-regulation of Claudin-1 in colon in LPS-treated mice.

Leaf Extract from European Olive (Olea europaea L.) Post-Transcriptionally Suppresses the Epithelial-Mesenchymal Transition and Sensitizes Gastric Cancer Cells to Chemotherapy.

The overall survival of patients with the advanced and recurrent gastric cancer (GC) remains unfavorable. In particular, this is due to cancer spreading and resistance to chemotherapy associated with the epithelial-mesenchymal transition (EMT) of tumor cells. EMT can be identified by the transcriptome profiling of GC for EMT markers. Indeed, analysis of the TCGA and GTEx databases (n = 408) and a cohort of GC patients (n = 43) revealed that expression of the CDH2 gene was significantly decreased in the tumors vs. non-tumor tissues and correlated with the overall survival of GC patients. Expression of the EMT-promoting transcription factors SNAIL and ZEB1 was significantly increased in GC. These data suggest that targeting the EMT might be an attractive therapeutic approach for patients with GC. Previously, we demonstrated a potent anti-cancer activity of the olive leaf extract (OLE). However, its effect on the EMT regulation in GC remained unknown. Here, we showed that OLE efficiently potentiated the inhibitory effect of the chemotherapeutic agents 5-fluorouracil (5-FU) and cisplatin (Cis) on the EMT and their pro-apoptotic activity, as was demonstrated by changes in the expression of the EMT markers (E- and N-cadherins, vimentin, claudin-1) in GC cells treated with the aforementioned chemotherapeutic agents in the presence of OLE. Thus, culturing GC cells with 5-FU + OLE or Cis + OLE attenuated the invasive properties of cancer cells. Importantly, upregulation of expression of the apoptotic markers (PARP cleaved form) and increase in the number of cells undergoing apoptosis (annexin V-positive) were observed for GC cells treated with a combination of OLE and 5-FU or Cis. Collectively, our data illustrate that OLE efficiently interferes with the EMT in GC cells and potentiates the pro-apoptotic activity of certain chemotherapeutic agents used for GC therapy.

Multifaceted impact of adipose conditioned media: Obesity-driven promotion of prostate cancer and cancer stem cell dynamics.

Obesity is an established risk factor for the development and progression of prostate cancer (PC). This study used adipose conditioned media (ACM) from differentiated adipocytes to assess its effect on PC development and aggressiveness. Due to limited research on ACM's impact on isolated PC stem cells (PCSCs), we also examined CD44+ PCSCs. ACM notably boosted interleukin-1ß (IL-1ß), IL-6, and IL-8 production in normal prostate epithelial cells and LNCaP cells. It also increased IL-6 and IL-8 production in PC3 and CD44+ LNCaP cells, and IL-1ß and IL-6 production in CD44+ PC3 cells. This indicates that ACM induces the production of inflammatory cytokines in both cancer and prostate epithelial cells. Furthermore, ACM promoted proliferation in androgen receptor (AR)-negative PC3 cells, CD44+ PC3 PCSCs, and nonmalignant RWPE cells, without affecting AR-positive LNCaP cells. In addition, ACM-enhanced invasion and migration potential in both PC3 and CD44+ PC3 cells. Western blot analysis indicated the involvement of NF-κB and AKT pathways in ACM-induced proliferation in PC3 cells and NF-κB in PCSCs. In ACM-treated PC3 cells, E-cadherin was downregulated, while N-cadherin, Snail, vimentin, fibronectin, and Twist were upregulated, suggesting ACM-induced invasion via classical epithelial-to-mesenchymal transition (EMT) pathways. In response to ACM, PCSCs exhibited increased expression of E-cadherin, Snail, and vimentin, which are partial EMT markers promoting stemness and resistance to apoptosis. In addition, increased expressions of Nanog, Oct3/4, survivin, and Bcl-2 were observed. Although the molecules we studied have diverse effects on cellular regulation, our data emphasize obesity's multifaceted role in promoting and aggressing PC, notably affecting PCSC populations.