Identification of Hub Genes and Potential Pathogenesis in Gastric Cancer Based on Integrated Gene Expression Profile Analysis.

OBJECTIVE: Gastric cancer (GC) is one of the most common malignancies and ranks third in terms of cancer-related mortality. This study aims to identify the hub genes and potential mechanisms in GC using a bioinformatics approach. METHODS: Microarray data GSE54129, GSE79973, GSE55696 were extracted from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) was identified using Benjamini-Hochberg method in the limma package. GO and KEGG pathway enrichment analyses of the DEGs were conducted. Furthermore, protein-protein interaction network was constructed the STRING platform, and the hub genes were discovered using Maximal Clique Centrality method via cytoHubba. The predictive significance of hub genes was evaluated through GSE15459 dataset. RESULTS: A total of 73 genes was identified as DEGs in GC. Volcano plots and heatmaps of DEGs were visualized. Functional enrichment analysis revealed that the genes were mostly enriched in response to xenobiotic stimulus, digestion, cellular hormone metabolic process, extracellular matrix structural constituent, calcium-dependent cysteine-type endopeptidase activity, aromatase activity, apical part of cell, basal part of cell, and apical plasma membrane. Regarding KEGG pathway-enrichment, the genes were mainly involved in Drug metabolism-cytochrome P450, Retinol metabolism, Chemical carcinogenesis-DNA adducts, Gastric acid secretion, and Metabolism of xenobiotics by cytochrome P450. By combining the results of Cytohubba, the top five intersecting genes identified were SPP1, INHBA, MMP7, THBS2 and FAP. Kapplan-Meier analysis results showed that these 5 hub genes were highly related to the overall survival of patients. CONCLUSION: SPP1, INHBA, MMP7, THBS2, and FAP were identified as prospective biomarkers and therapeutic targets for GC that might be utilized for prognostic evaluation and scheme selection.

Everolimus exerts anticancer effects through inhibiting the interaction of matrix metalloproteinase-7 with syndecan-2 in colon cancer cells.

Previous work showed that matrix metalloproteinase-7 (MMP-7) regulates colon cancer activities through an interaction with syndecan-2 (SDC-2) and SDC-2-derived peptide that disrupts this interaction and exhibits anticancer activity in colon cancer. Here, to identify potential anticancer agents, a library of 1,379 Food and Drug Administration (FDA)-approved drugs that interact with the MMP-7 prodomain were virtually screened by protein-ligand docking score analysis using the GalaxyDock3 program. Among five candidates selected based on their structures and total energy values for interacting with the MMP-7 prodomain, the known mechanistic target of rapamycin kinase (mTOR) inhibitor, everolimus, showed the highest binding affinity and the strongest ability to disrupt the interaction of the MMP-7 prodomain with the SDC-2 extracellular domain in vitro. Everolimus treatment of the HCT116 human colon cancer cell line did not affect the mRNA expression levels of MMP-7 and SDC-2 but reduced the adhesion of cells to MMP-7 prodomain-coated plates and the cell-surface localization of MMP-7. Thus, everolimus appears to inhibit the interaction between MMP-7 and SDC-2. Everolimus treatment of HCT116 cells also reduced their gelatin-degradation activity and anticancer activities, including colony formation. Interestingly, cells treated with sirolimus, another mTOR inhibitor, triggered less gelatin-degradation activity, suggesting that this inhibitory effect of everolimus was not due to inhibition of the mTOR pathway. Consistently, everolimus inhibited the colony-forming ability of mTOR-resistant HT29 cells. Together, these data suggest that, in addition to inhibiting mTOR signaling, everolimus exerts anticancer activity by interfering with the interaction of MMP-7 and SDC-2, and could be a useful therapeutic anticancer drug for colon cancer.NEW & NOTEWORTHY The utility of cancer therapeutics targeting the proteolytic activities of MMPs is limited because MMPs are widely distributed throughout the body and involved in many different aspects of cell functions. This work specifically targets the activation of MMP-7 through its interaction with syndecan-2. Notably, everolimus, a known mTOR inhibitor, blocked this interaction, demonstrating a novel role for everolimus in inhibiting mTOR signaling and impairing the interaction of MMP-7 with syndecan-2 in colon cancer.

The dependence between glycodelin and selected metalloproteinases concentrations in bovine placenta during early gestation and parturition with and without retained foetal membranes.

Pregnancy course depends on the appropriate connection between the mother and the developing foetus. Pregnancy is completed when the placenta is timely expelled. Placental retention is one of the possible pregnancy complications. Extracellular matrix, including adhesive proteins and enzymes that can break down collagens, seems to be responsible for it. The aim of the present study was to examine the impact of one of the adhesive proteins - glycodelin (Gd) - on selected metalloproteinases degrading collagens (MMP2, MMP3, MMP7). Placental tissues from healthy pregnant cows collected during early-mid pregnancy (2nd month n = 7, 3rd month n = 8, 4th month n = 6) and in cows that properly released placenta (NR; n = 6) and cows with retained foetal membranes (R; n = 6) were experimental material. The concentrations of glycodelin and protein content of selected metalloproteinases were measured by ELISA in the maternal and foetal placental homogenates as well as in the culture of epithelial cells derived from the maternal part of the placenta. The presence of these protein molecules was confirmed by Western Blotting. In the bovine placenta, the concentrations of examined proteins exhibit significant changes during placental formation. Gd, MMP3 and MMP7 concentrations decrease with pregnancy progress (between the 2nd and 4th month), while MMP2 concentrations were on the same level in this period. During parturition, concentrations of Gd and MMP3 were significantly higher in the R group compared to the NR group. In parallel, MMP2 concentrations did not show significant differences between the groups (NR vs R), and MMP7 concentrations decreased significantly in the maternal part of the placenta in cows with retained foetal membranes (R). Obtained results show correlations between the gestational age and proteins' (Gd, MMP3, MMP7) concentration, both in the maternal and foetal part of the placenta.

The effect and mechanism of low-molecular-weight heparin on the decidualization of stromal cells in early pregnancy.

OBJECTIVE: This study aimed to analyze the effect of low-molecular-weight heparin (LMWH) on the decidualization of stromal cells in early pregnancy and explore the effect of LMWH on pregnancy outcomes. METHODS: Recurrent spontaneous abortion (RSA) mouse model (CBA/J × DBA/2) and normal pregnant mouse model (CBA/J × BALB/c) were established. The female mice were checked for a mucus plug twice daily to identify a potential pregnancy. When a mucus plug was found, conception was considered to have occurred 12 h previously. The pregnant mice were divided randomly into a normal pregnancy control group, an RSA model group, and an RSA + LMWH experimental group (n = 10 mice in each group). Halfway through the 12th day of pregnancy, the embryonic loss of the mice was observed; a real-time quantitative polymerase chain reaction was used to detect the messenger ribonucleic acid (mRNA) expressions of prolactin (PRL) and insulin-like growth factor-binding protein 1 (IGFBP1) in the decidua of the mice. Additionally, the decidual tissues of patients with RSA and those of normal women in early pregnancy who required artificial abortion were collected and divided into an RSA group and a control group. Decidual stromal cells were isolated and cultured to compare cell proliferation between the two groups, and cellular migration and invasion were detected by membrane stromal cells. Western blotting was used to detect the protein expressions of proliferating cell nuclear antigen (PCNA), cyclin D1, matrix metalloproteinase- (MMP) 2, and MMP-7 in stromal cells treated with LMWH. RESULTS: Compared with the RSA group, LMWH significantly reduced the pregnancy loss rate in the RSA mice (p < 0.05). Compared with the RSA group, the LMWH + RSA group had significantly higher expression levels of PRL and IGFBP1 mRNA (p < 0.01). LMWH promoted the proliferation, migration, and invasion of human decidual stromal cells; compared with the control group, the expression levels of MMP-2, MMP-7, cyclin D1, and PCNA proteins in the decidual stromal cells of the LMWH group increased (p < 0.05). CONCLUSIONS: The use of LMWH can improve pregnancy outcomes by enhancing the proliferation and migration of stromal cells in early pregnancy and the decidualization of stromal cells.

Matrix metalloproteinase-7 and claudin-7 as novel identified therapeutic targets for restoration of intestinal epithelial barrier in inflammatory bowel diseases.

Intestinal tight junction disruption and mucosal immune dysregulation contribute to pathogenesis and progression of inflammatory bowel diseases (IBD). A proteolytic enzyme matrix metalloproteinase 7 (MMP-7), which is highly expressed in intestinal tissue, is implicated to IBD and other immune overactivation-associated diseases. In the issue of the Frontiers in Immunology, Ying Xiao and colleagues demonstrate that MMP-7-mediated claudin-7 degradation promotes IBD pathogenesis and disease progression. Therefore, inhibition of MMP-7 enzymatic activity can be a therapeutic strategy for the treatment of IBD.

[Aqueous extract of Epimedium sagittatum mitigates pulmonary fibrosis in mice].

This study aims to investigate the intervention effect of the aqueous extract of Epimedium sagittatum Maxim on the mouse model of bleomycin(BLM)-induced pulmonary fibrosis, so as to provide data support for the clinical treatment of pulmonary fibrosis. Ninety male C57BL/6N mice were randomized into normal(n=10), model(BLM, n=20), pirfenidone(PFD, 270 mg·kg~(-1), n=15), and low-, medium-, and high-dose E. sagittatum extract(1.67 g·kg~(-1), n=15; 3.33 g·kg~(-1), n=15; 6.67 g·kg~(-1), n=15) groups. The model of pulmonary fibrosis was established by intratracheal instillation of BLM(5 mg·kg~(-1)) in the other five groups except the normal group, which was treated with an equal amount of normal saline. On the day following the modeling, each group was treated with the corresponding drug by gavage for 21 days. During this period, the survival rate of the mice was counted. After gavage, the lung index was calculated, and the morphology and collagen deposition of the lung tissue were observed by hematoxylin-eosin(HE) and Masson staining, respectively. The levels of reactive oxygen species(ROS) in lung cell suspensions were measured by flow cytometry. The levels of glutathione peroxidase(GSH-Px), total superoxide dismutase(T-SOD), and malondialdehyde(MDA) the in lung tissue were measured. Terminal-deoxynucleoitidyl transferase-mediated nick-end labeling(TUNEL) was employed to examine the apoptosis of lung tissue cells. The content of interleukin-6(IL-6), chemokine C-C motif ligand 2(CCL-2), matrix metalloproteinase-8(MMP-8), transforming growth factor-beta 1(TGF-ß1), alpha-smooth muscle actin(α-SMA), E-cadherin, collagen Ⅰ, and fibronectin in the lung tissue was measured by enzyme-linked immunosorbent assay(ELISA). The expression levels of F4/80, Ly-6G, TGF-ß1, and collagen Ⅰ in the lung tissue were determined by immunohistochemistry. The mRNA levels of CCL-2, IL-6, and MMP-7 in the lung tissue were determined by qRT-PCR. The content of hydroxyproline(HYP) in the lung tissue was determined by alkaline hydrolysation. The expression of α-SMA and E-cadherin was detected by immunofluorescence, and the protein levels of α-SMA, vimentin, E-cadherin in the lung tissue were determined by Western blot. The results showed the aqueous extract of E. sagittatum increased the survival rate, decreased the lung index, alleviated the pathological injury, collagen deposition, and oxidative stress in the lung tissue, and reduced the apoptotic cells. Furthermore, the aqueous extract of E. sagittatum down-regulated the protein levels of F4/80 and Ly-6G and the mRNA levels of CCL-2, IL-6, and MMP-7 in the lung tissue, reduced the content of IL-6, CCL-2, and MMP-8 in the alveolar lavage fluid. In addition, it lowered the levels of HYP, TGF-ß1, α-SMA, collagen Ⅰ, fibronectin, and vimentin, and elevated the levels of E-cadherin in the lung tissue. The aqueous extract of E. sagittatum can inhibit collagen deposition, alleviate oxidative stress, and reduce inflammatory response by regulating the expression of the molecules associated with epithelial-mesenchymal transition, thus alleviating the symptoms of bleomycin-induced pulmonary fibrosis in mice.

Renogrit attenuates Vancomycin-induced nephrotoxicity in human renal spheroids and in Sprague-Dawley rats by regulating kidney injury biomarkers and creatinine/urea clearance.

Vancomycin, is widely used against methicillin-resistant bacterial infections. However, Vancomycin accumulation causes nephrotoxicity which leads to an impairment in the filtration mechanisms of kidney. Traditional herbal medicines hold potential for treatment of drug-induced nephrotoxicity. Herein, we investigated protective properties of plant-based medicine Renogrit against Vancomycin-induced kidney injury. Phytometabolite analysis of Renogrit was performed by UHPLC. Spheroids formed from human proximal tubular cell (HK-2) were used for in vitro evaluation of Vancomycin-induced alterations in cell viability, P-gp functionality, NAG, KIM-1 levels, and mRNA expression of NGAL and MMP-7. The in vivo efficacy of Renogrit against Vancomycin-induced nephrotoxicity was further evaluated in Sprague-Dawley (SD) rats by measurement of BUN, serum creatinine, and their respective clearances. Moreover, eGFR, kidney-to-body weight ratio, GSH/GSSG ratio, KIM-1, NAG levels and mRNA expression of KIM-1 and osteopontin were also analyzed. Changes in histopathology of kidney and hematological parameters were also observed. Renogrit treatment led to an increase in cell viability, normalization of P-gp functionality, decrease in levels of NAG, KIM-1, and reduction in mRNA expression of NGAL and MMP-7. In Vancomycin-challenged SD rats, Renogrit treatment normalized altered kidney functions, histological, and hematological parameters. Our findings revealed that Renogrit holds a clinico-therapeutic potential for alleviating Vancomycin-associated nephrotoxicity.

Circulating biomarkers of extracellular matrix dysregulation are associated with adverse post-stage 2 outcomes in infants with single ventricle heart disease.

Children with single ventricle heart disease (SVHD) experience morbidity due to inadequate pulmonary blood flow. Using proteomic screening, our group previously identified members of the matrix metalloproteinase (MMP), tissue inhibitor of metalloproteinase (TIMP), and fibroblast growth factor (FGF) families as potentially dysregulated in SVHD. No prior study has taken a targeted approach to mapping circulating levels of these protein families or their relationship to pulmonary vascular outcomes in SVHD. We performed a prospective cohort study of 70 SVHD infants pre-Stage 2 palliation and 24 healthy controls. We report targeted serum quantification of 39 proteins in the MMP, TIMP, and FGF families using the SomaScan platform. Clinical variables were extracted from the medical record. Twenty of 39 tested proteins (7/14 MMPs, 2/4 TIMPs, and 11/21 FGFs) differed between cases and controls. On single variable testing, 6 proteins and no clinical covariates were associated with both post-Stage 2 hypoxemia and length of stay. Multiple-protein modeling identified increased circulating MMP 7 and MMP 17, and decreased circulating MMP 8 and FGFR2 as most associated with post-Stage 2 hypoxemia; increased MMP 7 and TIMP 4 and decreased circulating MMP 1 and MMP 8 were most associated with post-operation length of stay. The MMP, TIMP, and FGF families are altered in SVHD. Pre-Stage 2 imbalance of extracellular matrix (ECM) proteins-increased MMP 7 and decreased MMP 8-was associated with multiple adverse post-operation outcomes. Maintenance of the ECM may be an important pathophysiologic driver of Stage 2 readiness in SVHD.

Plasma Exosomal microRNA Profile Reveals miRNA 148a-3p Downregulation in the Mucosal-Dominant Variant of Pemphigus Vulgaris.

The mucosal-dominant variant of pemphigus vulgaris (MPV) is an autoimmune disease characterized by oral mucosal blistering and circulating pathogenic IgG antibodies against desmoglein 3 (Dsg3), resulting in life-threatening bullae and erosion formation. Recently, microRNAs (miRNAs) have emerged as promising players in the diagnosis and prognosis of several pathological states. For the first time, we have identified a different expression profile of miRNAs isolated from plasma-derived exosomes (P-EVs) of MPV patients positive for antibodies against Dsg3 (Dsg3-positive) compared to healthy controls. Moreover, a dysregulated miRNA profile was confirmed in MPV tissue biopsies. In particular, a strong downregulation of the miR-148a-3p expression level in P-EVs of MPV patients compared to healthy controls was demonstrated. Bioinformatics prediction analysis identifies metalloproteinase-7 (MMP7) as a potential miR-148a-3p target. An in vitro acantholysis model revealed that the miR-148a-3p expression level was dramatically downregulated after treatment with Dsg3 autoantibodies, with a concomitant increase in MMP7 expression. The increased expression of MMP7 leads to the disruption of intercellular and/or extracellular matrix adhesion in an in vitro cellular model of MPV, with subsequent cell dissociation. Overexpression of miR-148a-3p prevented cell dissociation and regressed MMP7 upregulation. Our findings suggest a pivotal role of P-EV cargo in regulating molecular mechanisms involved in MPV pathogenesis and indicate them as potential MPV therapeutic targets.

Interplay between EZH2/ß-catenin in stemness of cisplatin-resistant HNSCC and their role as therapeutic targets.

The Wnt/ß-catenin signaling pathway is associated with the regulation of cancer stem cells, and it can be driven by epigenetic modifications. Here, we aim to identify epigenetic modifications involved in the control of the Wnt/ß-catenin signaling and investigate the role of this pathway in the accumulation of cancer stem cells (CSC) and chemoresistance of Head and Neck Squamous Cell Carcinoma (HNSCC). Quantitative-PCR, western blot, shRNA assay, viability assay, flow cytometry assay, spheres formation, xenograft model, and chromatin immunoprecipitation were employed to evaluate the Wnt/ß-catenin pathway and EZH2 in wild-type and chemoresistant oral carcinoma cell lines, and in the populations of CSC and non-stem cells. We demonstrated that ß-catenin and EZH2 were accumulated in cisplatin-resistant and CSC population. The upstream genes of the Wnt/ß-catenin signaling (APC and GSK3ß) were decreased, and the downstream gene MMP7 was increased in the chemoresistant cell lines. The inhibition of ß-catenin and EZH2 combined effectively decreased the CSC population in vitro and reduced the tumor volume and CSC population in vivo. EZH2 inhibition increased APC and GSK3ß, and the Wnt/ß-catenin inhibition reduced MMP7 levels. In contrast, EZH2 overexpression decreased APC and GSK3ß and increased MMP7. EZH2 and ß-catenin inhibitors sensitized chemoresistant cells to cisplatin. EZH2 and H3K27me3 bounded the promoter of APC, leading to its repression. These results suggest that EZH2 regulates ß-catenin by inhibiting the upstream gene APC contributing to the accumulation of cancer stem cells and chemoresistance. Moreover, the pharmacological inhibition of the Wnt/ß-catenin combined with EZH2 can be an effective strategy for treating HNSCC.

Potentially important markers in thyroid neoplasia: Claudin-1 and MMP-7.

Background: Thyroid carcinomas are the most common malignant endocrine tumors, and various immunohistochemical markers are tested in routine practice to reduce diagnostic differences, as well as to elucidate carcinogenesis and detect malignancy. Disruption of basement membranes and the extracellular matrix is an important step in tumor carcinogenesis and progression. The claudin and matrix metalloproteinase families are also thought to be effective in this process. Aim: In this retrospective study, the comparative expression of claudin-1 and MMP-7 immunomarkers in normal tissues and thyroid neoplasia were investigated. Materials and Methods: Immunohistochemical staining was performed for claudin-1 and matrix metalloproteinase 7 (MMP-7) in 112 sections, including 24 follicular adenomas, 22 follicular carcinomas, 24 medullary carcinomas, 24 papillary carcinomas, and 18 single dominant nodules from thyroid lesions. Results: A significant staining difference for claudin-1 was observed in follicular carcinoma and medullary carcinoma, papillary carcinoma, and single dominant nodules compared to normal thyroid tissue. A statistically significant staining difference was observed for MMP-7 in follicular adenoma, medullary carcinoma, and papillary carcinoma compared to normal thyroid tissue. Conclusions: These results indicate that claudin-1 and MMP-7 are important in the diagnosis, differential diagnosis, and carcinogenesis of follicular adenoma, follicular carcinoma, medullary carcinoma, papillary carcinoma, and single dominant nodules.

Unbiased Human Kidney Tissue Proteomics Identifies Matrix Metalloproteinase 7 as a Kidney Disease Biomarker.

SIGNIFICANCE STATEMENT: Although gene expression changes have been characterized in human diabetic kidney disease (DKD), unbiased tissue proteomics information for this condition is lacking. The authors conducted an unbiased aptamer-based proteomic analysis of samples from patients with DKD and healthy controls, identifying proteins with levels that associate with kidney function (eGFR) or fibrosis, after adjusting for key covariates. Overall, tissue gene expression only modestly correlated with tissue protein levels. Kidney protein and RNA levels of matrix metalloproteinase 7 (MMP7) strongly correlated with fibrosis and with eGFR. Single-cell RNA sequencing indicated that kidney tubule cells are an important source of MMP7. Furthermore, plasma MMP7 levels predicted future kidney function decline. These findings identify kidney tissue MMP7 as a biomarker of fibrosis and blood MMP7 as a biomarker for future kidney function decline. BACKGROUND: Diabetic kidney disease (DKD) is responsible for close to half of all ESKD cases. Although unbiased gene expression changes have been extensively characterized in human kidney tissue samples, unbiased protein-level information is not available. METHODS: We collected human kidney samples from 23 individuals with DKD and ten healthy controls, gathered associated clinical and demographics information, and implemented histologic analysis. We performed unbiased proteomics using the SomaScan platform and quantified the level of 1305 proteins and analyzed gene expression levels by bulk RNA and single-cell RNA sequencing (scRNA-seq). We validated protein levels in a separate cohort of kidney tissue samples as well as in 11,030 blood samples. RESULTS: Globally, human kidney transcript and protein levels showed only modest correlation. Our analysis identified 14 proteins with kidney tissue levels that correlated with eGFR and found that the levels of 152 proteins correlated with interstitial fibrosis. Of the identified proteins, matrix metalloprotease 7 (MMP7) showed the strongest association with both fibrosis and eGFR. The correlation between tissue MMP7 protein expression and kidney function was validated in external datasets. The levels of MMP7 RNA correlated with fibrosis in the primary and validation datasets. Findings from scRNA-seq pointed to proximal tubules, connecting tubules, and principal cells as likely cellular sources of increased tissue MMP7 expression. Furthermore, plasma MMP7 levels correlated not only with kidney function but also associated with prospective kidney function decline. CONCLUSIONS: Our findings, which underscore the value of human kidney tissue proteomics analysis, identify kidney tissue MMP7 as a diagnostic marker of kidney fibrosis and blood MMP7 as a biomarker for future kidney function decline.

Melatonin inhibits chondrosarcoma cell proliferation and metastasis by enhancing miR-520f-3p production and suppressing MMP7 expression.

Chondrosarcoma has a high propensity to metastasize and responds poorly to chemotherapy and radiation treatment. The enzymatic activity of matrix metalloproteinases (MMPs) is very important in chondrosarcoma metastasis. Melatonin exhibits anticarcinogenic activity in many types of cancers by suppressing the expression of certain MMP family members, but this has not yet been clearly determined in chondrosarcoma. Our study demonstrates that MMP7 plays an essential role in chondrosarcoma cell proliferation, migration, and anoikis resistance. We also found that MMP7 is highly expressed in chondrosarcomas. Our in vitro and in vivo investigations show that melatonin strongly inhibits chondrosarcoma cell proliferation, migration, and anoikis resistance by directly suppressing MMP7 expression. Melatonin reduced MMP7 synthesis by promoting levels of miR-520f-3p expression, which were downregulated in human chondrosarcoma tissue samples. Pharmacological inhibition of miR-520f-3p markedly reversed the effects of melatonin upon chondrosarcoma proliferation and metastasis. Thus, our study suggests that melatonin has therapeutic potential for reducing the tumorigenesis and metastatic potential of chondrosarcoma via the miR-520f-3p/MMP7 axis.

Effect of CUX1 on the Proliferation of Hu Sheep Dermal Papilla Cells and on the Wnt/ß-Catenin Signaling Pathway.

CUT-like homeobox 1 protein (CUX1), also called CUX, CUTL1, and CDP, is a member of the DNA-binding protein homology family. Studies have shown that CUX1 is a transcription factor that plays an important role in the growth and development of hair follicles. The aim of this study was to investigate the effect of CUX1 on the proliferation of Hu sheep dermal papilla cells (DPCs) to reveal the role of CUX1 in hair follicle growth and development. First, the coding sequence (CDS) of CUX1 was amplified by PCR, and then CUX1 was overexpressed and knocked down in DPCs. A Cell Counting Kit-8 (CCK8), 5-ethynyl-2-deoxyuridine (EdU), and cell cycle assays were used to detect the changes in the proliferation and cell cycle of DPCs. Finally, the effects of overexpression and knockdown of CUX1 in DPCs on the expression of WNT10, MMP7, C-JUN, and other key genes in the Wnt/ß-catenin signaling pathway were detected by RT-qPCR. The results showed that the 2034-bp CDS of CUX1 was successfully amplified. Overexpression of CUX1 enhanced the proliferative state of DPCs, significantly increased the number of S-phase cells, and decreased the number of G0/G1-phase cells (p < 0.05). CUX1 knockdown had the opposite effects. It was found that the expression of MMP7, CCND1 (both p < 0.05), PPARD, and FOSL1 (both p < 0.01) increased significantly after overexpression of CUX1 in DPCs, while the expression of CTNNB1 (p < 0.05), C-JUN, PPARD, CCND1, and FOSL1 (all p < 0.01) decreased significantly. In conclusion, CUX1 promotes proliferation of DPCs and affects the expression of key genes of the Wnt/ß-catenin signaling pathway. The present study provides a theoretical basis to elucidate the mechanism underlying hair follicle development and lambskin curl pattern formation in Hu sheep.

Synergistic Protective Effect of Interactions of Quercetin with Lycopene Against Ochratoxin A-Induced Ulcerative Colitis.

Ulcerative colitis is a type of inflammatory bowel disease responsible for the inflammation of the innermost lining of the colon and rectum. The present study's objective is to determine the potential synergistic impact of quercetin (QR) and lycopene (LP) in ulcerative colitis (UC) induced in rats by ochratoxin A (OTA) by biochemical and morphological alterations. QR and LP were administered alone and in combination with the OTA for 7 days. OTA administration caused UC generation, resulting in significant changes in body weight percentage, disease activity index (DAI), macroscopic evaluation, colon weight/length ratio, and histological score. In addition to the above parameters, it also leads to elevated oxidative stress, i.e. increased malondialdehyde (MDA), nitric oxide (NO), myeloperoxidase (MPO), and hydroxyproline levels and decreased superoxide dismutase (SOD) and reduced glutathione (GSH) levels. Histological changes in the colon architecture were also observed suggestive of extensive mucosal damage. In addition, a high level of matrix metalloproteinase 7 (MMP7) was observed in immunohistochemistry, and a high level of gene expression of osteopontin (OPN), runt-related transcription factor 2 (RUNX2), MMP-7, and interleukin-6 (IL-6) was observed in OTA administered animals. The combination of QR and LP significantly restored the per cent body weight loss and DAI score and improved macroscopic and histological changes, colon weight/length ratio, and macroscopic damages. It also improved the biochemical parameters to near-normal levels, i.e. reduced MDA, NO, MPO, and hydroxyproline levels and increased SOD and GSH levels. In addition, OPN, Runx2, MMP-7, and IL-6 gene expression decreased compared to the OTA-induced UC group. Outcomes of the present study indicate the potential of QR + LP as anti-inflammatory and immunomodulatory agents against OTA-induced UC in rats.

Knockdown of CENPU inhibits cervical cancer cell migration and stemness through the FOXM1/Wnt/ß-catenin pathway.

Currently, the clinical outcome of cervical cancer (CC) is still undesirable, and it is urgent to explore more treatment strategies for CC. In this study, the effects of CENPU on migration and stemness of CC was studied. The CENPU expression were retrieved from The Cancer Genome Atlas (TCGA). The effects of CENPU on the viability and proliferation of cells were evaluated by CCK-8 assay and colony formation assay. Wound healing assay and invasion assay were chosen to assess migration and invasion of cells. Tumorsphere-forming assay was applied for testing the stemness. Western blot analysis was applied for assessing the level of CENPU, Nanog, Oct4, FOXM1, ß-catenin, c-myc and MMP-7. The tumor sizes and volumes were also measured. The TCGA data and WB assay suggested that CENPU was upregulated in CC. CENPU knockdown would inhibit the viability of CC cells and prohibit the migration and invasion of cells. Tumorsphere-forming assay and WB results suggested that CENPU silencing decreased the sphere formation rate and the expression of Nanog and Oct4. Moreover, CENPU knockdown suppressed the expression of FOXM1, ß-catenin, c-myc and MMP-7 by WB. In vivo study demonstrated that CENPU knockdown inhibited the growth of CC, indicated by reduced sizes and volumes of CC. In summary, our results suggested that knockdown of CENPU inhibited CC migration and stemness through the FOXM1/Wnt/ß-catenin pathway.

Alantolactone Inhibits Melanoma Cell Culture Viability and Migration and Promotes Apoptosis by Inhibiting Wnt/ß-Catenin Signaling.

BACKGROUND: Melanoma is a highly invasive and metastatic malignant tumor originating from melanocytes and is associated with a poor prognosis. Surgical resection and chemotherapy are currently the main therapeutic options for malignant melanoma; however, their efficacy is poor, highlighting the need for the development of new, safe, and effective drugs for the treatment of this cancer. OBJECTIVE: To investigate the effects of alantolactone (ALT) on the proliferative, migratory, invasive, and apoptotic ability of malignant melanoma cells and explore its potential anticancer mechanism. METHODS: Melanoma cells (A375 and B16) were treated with different concentrations (4, 6, 8, and 10 µmol/L) of ALT, with DMSO and no treatment serving as controls. The effects of the different concentrations of the drug on cell proliferation were assessed by crystal violet staining and CCK-8 assay. The effects on cell migration and invasion were detected by wound healing and Transwell assays, respectively. Flow cytometry was used to evaluate the effects of the drug on apoptosis and the cell cycle. ALT target genes in melanoma were screened using network pharmacology. Western blotting was used to measure the expression levels of the proliferation-related protein PCNA; the apoptosisrelated proteins Bax, Bcl-2, and caspase-3; the invasion and metastasis-related proteins MMP-2, MMP-7, MMP-9, vimentin, E-cadherin, and N-cadherin; and the canonical Wnt signaling pathway-related proteins ß-catenin, c-Myc, and p-GSK3ß. In addition, an l model of melanoma was established by the subcutaneous injection of A375 melanoma cells into nude mice, following which the effects of ALT treatment on malignant melanoma were determined in vivo. RESULTS: Compared with the controls, the proliferative, migratory, and invasive capacity of ALT-treated melanoma cells was significantly inhibited, whereas apoptosis was enhanced (P<0.01), showing effects that were exerted in a dose-dependent manner. The expression levels of the pro-apoptotic proteins Bax and caspase-3, as well as those of the interstitial marker E-cadherin, were upregulated in melanoma cells irrespective of the ALT concentration (P<0.05). In contrast, the expression levels of the anti-apoptotic protein Bcl-2, the proliferation-related protein PCNA, and the invasion and metastasis-related proteins MMP-2, MMP-7, MMP-9, N-cadherin, and vimentin were downregulated (P<0.05). The network pharmacology results indicated that GSK3ß may be a key ALT target in melanoma. Meanwhile, western blotting assays showed that ALT treatment markedly suppressed the expression of ß-catenin as well as that of its downstream effector c-Myc, and could also inhibit GSK3ß phosphorylation. CONCLUSION: ALT can effectively inhibit the culture viability, migration, and invasion of A375 and B16 melanoma cells while also promoting their apoptosis. ALT may exert its anti-melanoma effects by inhibiting the Wnt/ß-catenin signaling pathway. Combined, our data indicate that ALT has the potential as an effective and safe therapeutic drug for the treatment of melanoma.

Expression of MMP-2, MMP-7, MMP-9, and TIMP-1 by Inflamed Mucosa in the Initial Diagnosis of Ulcerative Colitis as a Response Marker for Conventional Medical Treatment.

INTRODUCTION: Experimental and clinical data involve matrix metalloproteases (MMPs) and their tissue inhibitors (TIMPs) in the pathogenesis of inflammatory bowel diseases. However, the impact of MMPs/TIMPs expression by inflamed mucosa on medical response therapy has scarcely been investigated. METHODS: The expression of MMP-2, MMP-7, MMP-9, and TIMP-1 was determined by immunohistochemical analysis in inflamed mucosa samples at diagnosis in 82 patients with ulcerative colitis (UC; 22 never-treated with corticosteroids, 28 nonresponders, and 32 responders to corticosteroid therapy) and 15 patients with acute diverticulitis (AD). The global expression (score value) of each factor was analyzed by computer-generated image analysis. RESULTS: UC samples showed higher MMP-2 and MMP-9 expression but lower TIMP-1 expression than the AD samples (p < 0.0001, for all). High MMP-9 and TIMP-1 scores were significantly associated with no need for corticosteroid treatment (p < 0.001 and p = 0.017, respectively); whereas higher score in the MMP-7 expression was significantly associated with nonresponse to corticosteroid therapy (p = 0.037). In addition, in this latter UC subgroup, MMP-7 correlated positively with the younger age of the patients and with the extension of the disease (p = 0.030 and p = 0.010, respectively). CONCLUSION: Our results suggest the relevance of MMPs and TIMPs for predicting treatment response to both 5-aminosalicylates and corticosteroids in UC.

Matrix metalloproteinase 7 is associated with clinical and pathological characteristics of salivary adenoid cystic carcinomas.

OBJECTIVES: The aim of this study was to investigate the relationship between matrix metalloproteinase-7 (MMP-7) expression and the clinical and pathological characteristics of salivary adenoid cystic carcinomas (SACC) of the palatal minor salivary gland. METHODS: In this study, 58 samples of SACC and 10 samples of normal salivary gland tissue were examined. Immunohistochemistry was used to detect MMP-7 and vascular endothelial growth factor A (VEGF-A) expression in SACC and normal tissues. The clinical and pathological characteristics of the patients with SACC were collected. RESULTS: Of the 58 SACC samples, 44 were positive for MMP-7, and the expression rate was 75.9%. No expression was detected in the 10 normal salivary gland tissues. The level of MMP-7 expression in the SACC and normal samples was significantly different. The level of expression of MMP-7 in the SACC samples did not correlate with age, sex or pathological type but did correlate with pathological grade, nerve infiltration and clinical stage. There was a positive correlation between VEGF-A and MMP-7 expression. CONCLUSIONS: The SACC samples showed high expression of MMP-7, which was associated with tumour differentiation, invasiveness and clinical stage. The detection of MMP-7 positively correlated with the detection of VEGF-A in SACC.

NOVA1 promotes NSCLC proliferation and invasion by activating Wnt/ß-catenin signaling.

BACKGROUND: Neuro-oncological ventral antigen 1 (NOVA1) is a neuron-specific RNA-binding protein which regulates alternative splicing in the developing nervous system. Recent research has found that NOVA1 plays a significant role in carcinogenesis. In this paper, we examine the role of NOVA1 in non-small cell lung cancer (NSCLC) and its underlying molecular mechanisms. METHODS: The expression of NOVA1 in NSCLC was detected by immunohistochemistry and correlations between NOVA1 expression and clinicopathological factors were analyzed by chi-square tests. Kaplan-Meier survival analysis and the Cox regression model were used to evaluate the predictive effect of prognostic factors. Western blotting, Cell Counting Kit-8, colony formation, apoptosis, migration and invasion assays were used to detect the effects of silencing (si)NOVA1 RNA on Wnt/ß-catenin signaling and biological behavior in NSCLC cell lines. RESULTS: Our study showed that expression of NOVA1 was up-regulated and significantly correlated with poor differentiation (p = 0.020), advanced TNM stage (P = 0.001), T stage (P = 0.001) and lymph node metastasis (P = 0.000) as well as the expression of ß-catenin (P = 0.012) in NSCLC. The down-regulation of NSCLC by siRNA significantly inhibited proliferation, migration and invasion and promoted apoptosis in NSCLC cells. Expression of Wnt signaling molecules, including ß-catenin, activated ß-catenin, cyclin D1, matrix metalloproteinase (MMP)-2 and MMP-7, was also significantly reduced by siNOVA1. The inhibition of Wnt/ß-catenin signaling in A549 and H1299 cells by siNOVA1 was reversed after treatment with a ß-catenin expression plasmid. CONCLUSION: The present study suggests that NOVA1 may serve as a potential prognosis biomarker in NSCLC. High NOVA1 expression was associated with poor survival rate. Finally, in vitro experiments verified that NOVA1 promotes NSCLC cell proliferation and invasion by regulating Wnt/ß-catenin signaling.