Gastric cancer (GC) is a major cause of global cancer mortality. Genetic variations in DNA repair genes can modulate DNA repair capability and, consequently, have been associated with risk of developing cancer. We have previously identified a T to C point mutation at nucleotide 889 (T889C) in DNA polymerase beta (POLB) gene, a key enzyme involved in base excision repair in primary GCs. The purpose of this study was to evaluate the mutation and expression of POLB in a larger cohort and to identify possible prognostic roles of the POLB alterations in GC. Primary GC specimens and their matched normal adjacent tissues were collected at the time of surgery. DNA, RNA and protein samples were isolated from GC specimens and cell lines. Mutations were detected by PCR-RFLP/DHPLC and sequencing analysis. POLB gene expression was examined by RT-PCR, tissue microarray, Western blotting and immunofluorescence assays. The function of the mutation was evaluated by chemosensitivity, MTT, Transwell matrigel invasion and host cell reactivation assays. The T889C mutation was detected in 18 (10.17%) of 177 GC patients. And the T889C mutation was associated with POLB overexpression, lymph nodes metastases and poor tumor differentiation. In addition, patients with- the mutation had significantly shorter survival time than those without-, following postoperative chemotherapy. Furthermore, cell lines with T889C mutation in POLB gene were more resistant to the treatment of 5-fluorouracil, cisplatin and epirubicin than those with wild type POLB. Forced expression of POLB gene with T889C mutation resulted in enhanced cell proliferation, invasion and resistance to anticancer drugs, along with increased DNA repair capability. These results suggest that POLB gene with T889C mutation in surgically resected primary gastric tissues may be clinically useful for predicting responsiveness to chemotherapy in patients with GC. The POLB gene alteration may serve as a prognostic biomarker for GC.
Esophageal cancer (EC) is a lethal cancer with an extremely aggressive nature and poor survival rate. However, the molecular mechanisms driving the occurrence and progression of EC are not well understood. MicroRNAs (miRNAs) are small RNA molecules that regulate the expression of protein-coding genes. miRNA-mediated gene regulation plays an important role in EC. By cross-referencing studies from NCBI, we found that microRNA-375 (miR-375) is one of the most frequently downregulated miRNAs in EC. We assessed expression of miR-375 in EC cell lines and primary EC tissues and their matched normal tissues. We found significant downregulation of miR-375 in both cell lines and EC tissues. Forced expression of miR-375 attenuated EC cell proliferation and invasion. Human epidermal growth factor receptor 2 (HER2, ERBB2), a known proto-oncogene, was identified here as one of the potential target genes of miR-375. Ectopic expression of miR-375 significantly suppressed the expression of ERBB2 and subsequently downregulated one of its target genes, vascular endothelial growth factor A (VEGFA), which is related to cancer invasion and metastasis. These findings suggest that miR-375 acts as a tumor suppressor by blocking the ERBB2/VEGFA pathway with the potential to modulate the occurrence and/ or progression of EC.
Increased expression of progesterone receptor (PR) has been reported in gastric cancer (GC). We have previously identified a functional T889C point mutation in DNA polymerase beta (POLB), a DNA repair gene in GC. To provide a detailed analysis of molecular changes associated with the mutation, human cDNA microarrays focusing on 18 signal transduction pathways were used to analyze differential gene expression profiles between GC tissues with T889C mutant in POLB gene and those with wild type. Among the differentially expressed genes, notably, PR was one of the significantly up-regulated genes in T889C mutant POLB tissues, which were subsequently confirmed in POLB gene transfected AGS cell line. Interestingly, patients with T889C mutation and PR positivity were associated with higher incidence of intraperitoneal metastasis (IM). In vitro studies indicate that PR expression was upregulated in AGS cell line when transfected with T889C mutant expression vector. Cotransfection of T889C mutant allele and PR gene induced cell migration in the cell line. These data demonstrated that T889C mutation-associated PR overexpression results in increased IM. Therefore, T889C mutation-associated PR overexpression may serve as a biomarker for an adverse prognosis for human GC.
Lichen sclerosus (LS) is a chronic inflammatory skin disorder with unknown pathogenesis. The aberrant expression of microRNAs (miRNAs) is considered to exert a crucial role in LS. We used the next-generation sequencing technology (RNASeq) for miRNA profiling and Ingenuity Pathway Analysis (IPA) for molecular network analysis. We performed qRT-PCR, miRNA transfection and Matrigel assays for functional studies. We identified a total of 170 differentially expressed miRNAs between female LS and matched adjacent normal tissue using RNASeq, with 119 upregulated and 51 downregulated. Bioinformatics analysis revealed molecular networks that may shed light on the pathogenesis of LS. We verified the expression of a set of miRNAs that are related to autoimmunity, such as upregulated miR-326, miR-142-5p, miR-155 and downregulated miR-664a-3p and miR-181a-3p in LS tissue compared to the matched adjacent normal tissue. The differentially expressed miRNAs were also verified in blood samples from LS patients compared to healthy female volunteers. Functional studies demonstrated that a forced expression of miR-142-5p in human dermal fibroblast PCS-201-010 cells resulted in decreased cell proliferation and migration. These findings suggest that differentially expressed miRNAs may play an important role in LS pathogenesis; therefore, they could serve as biomarkers for LS management.
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