International Journal of Molecular and Cellular Medicine最新文献

Polycystic ovary syndrome (PCOS) is the most prevalent endocrine disorder of women in reproductive age with significant effects on reproductive and metabolic functions. Many molecular players may be involved in PCOS pathology; however, miRNAs possess great ability in gene expression control in normal ovarian function and folliculogenesis. We appraised the relative expression of miR-146a, miR-222, miR-9, and miR-224 in serum and follicular fluid (FF) of PCOS patients compared to control subjects. PCOS (n = 35) and control (n = 30) subjects were recruited in the study during their enrolment in IVF cycles. Serum and FF of human subjects were collected and stored. Total RNA was isolated from samples and cDNA was synthesized using miRNA-specific stem-loop RT primers. Quantitative real-time PCR was used to evaluate the expression of miRNAs relative to U6 expression. The predictive value of miRNAs' expression for discrimination of PCOS patients from control subjects was evaluated by receiver-operating characteristic (ROC) curve analysis. miR-224 was not detected in serum and FF samples. Significantly, higher levels of miR-146a and miR-9 in serum of PCOS group were detected. In contrast, relative expression of miR-146a and miR-9 significantly decreased in FF. In PCOS group, relative expression of all detected miRNAs was elevated in serum in comparison to FF, whereas in control group no change was noticed. Combination of FF miRNAs showed improved predictive value with area under the ROC curve (AUC) of 0.84, 93.8% sensitivity, and 83.3% specificity. Contradicting alternations of miRNAs in serum and FF are indicative of different sources of miRNAs in body fluids. Presumptive target genes of studied miRNAs in signalling pathways may show the potential role of these miRNA in folliculogenesis.

Normal drugs exhibit activities against both normal and cancer cells. Furthermore, cancer cells may develop resistance to these drugs that alternative treatment must be explored. The main objective of this study was to examine the anticancer activity of Schiff base against Tongue Squamous Cell Carcinoma Fibroblasts (TSCCF) and normal human gingival fibroblasts (NHGF) and to propose its mechanism. A Novel Schiff base ligand was synthesized from the reaction of 5-C-2-4-NABA (5-chloro-2-((4-nitrobenzylidene) amino) benzoic acid). These Schiff bases possessed azomethine group (-HC=N-) and aromatic group (CH) as analyzed by Fourier transforms infrared (FTIR) spectroscopy and UV-Vis spectra. The in vitro cytotoxicity screening assay suggested promising activity against TSCCF with IC₅₀ of 446.68 µg/mL, but insignificant activity against NHGF cells (IC₅₀ of 977.24 µg/mL) after 72 h. The evidence of apoptotic induction was supported by DAPI staining of apoptotic nuclei with reduced cell numbers, suggesting that Schiff base could induce apoptotic bodies in cancer cells being observed. Based on the Schiff base structure, the anti-cancer mechanism may be attributed to the -HC=N- azomethine group. For the first time, our findings highlighted the anticancer activities of the new Schiff base against oral cancer cell lines.

Severe Acute Respiratory Syndrome Corona Virus 2 (SARS-CoV-2) infecting mechanism depends on hosting angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) as essential components and androgens as regulators for inducing the expression of these components. Therefore, hyperandrogenism-related disease such as polycystic ovary syndrome (PCOS) in insulin resistant women in reproductive-age is a high-risk factor for SARS-CoV-2 infection. Here, we describe the signaling pathways that might increase the susceptibility and severity of this new pandemic in PCOS women with insulin resistance (IR). Luteinizing hormone and insulin increase the risk of SARS-CoV-2 infection in these patients via the induction of steroidogenic enzymes expression through cAMP-response element binding protein and Forkhead box protein O1 (FOXO1), respectively. TMPRSS2 expression is activated through phosphorylation of FOXO1 in ovaries. In other words, SARS-CoV-2 infection is associated with temporary IR by affecting ACE2 and disturbing β-pancreatic function. Therefore, PCOS, IR, and SARS-CoV-2 infection are three corners of the triangle that have complicated relations, and their association might increase the risk of SARS-CoV-2 infection and severity.

Epstein-Barr virus (EBV) represents one of the most important viral carcinogens. EBV nuclear antigen-1 (EBNA1) can induce the expression of different cellular and viral genes. In this study, we evaluated the EBNA1 effects on the expression patterns of human papillomavirus type 18 (HPV-18) E6 and E7 oncogenes and three cellular genes, including BIRC5, c-MYC, and STMN1, in a cervical adenocarcinoma cell line. HeLa cells were divided into three groups: one transfected with a plasmid containing the EBNA1 gene, one transfected with a control plasmid, and one without transfection. In all three groups, the expression levels of E6, E7, BIRC5, c-MYC, and STMN1 genes were checked using real-time PCR. Pathological staining was used to examine changes in cell morphology. Real-time PCR results showed that the expression level of HPV-18 E6 (P=0.02) and E7 (P=0.02) oncogenes significantly increased in HeLa cells transfected with the EBNA1 plasmid compared to cells transfected with control plasmid. Also, the presence of EBNA1 induced the expression of BIRC5 and c-MYC, which increased tenfold (P=0.03) and threefold (P=0.02), respectively. Regarding the STMN1 cellular gene, although the expression level in HeLa cells transfected with EBNA1 plasmid showed a twofold increase, this change was insignificant (P=0.11). Also, EBNA1 expression caused the creation of large HeLa cells with abundant cytoplasm and numerous nuclei. The EBV-EBNA1 could increase the expression levels of HPV-18 E6 and E7 viral oncogenes as well as c-MYC and BIRC5 cellular genes in the HeLa cell line. These findings indicate that the simultaneous infection of cervical cells with HPV-18 and EBV might accelerate the progression of cervical cancer.

MicroRNAs (miRNAs) have emerged as essential gene expression regulators associated with human diseases such as colorectal cancer (CRC). The purpose of this study was to evaluate the expression of miR-330-3p and its target gene BMI1 in tissue samples of patients with CRC, polyp, and healthy adjacent tissue samples and their association with clinicopathological and demographic factors such as age, tumor stage, grade, and lymph node invasion of the tumor. Following the extraction of total RNA from approximately 50 mg of colon and rectum tissue of 82 patients with CRC, 13 polypoid lesions, and 26 marginal healthy tissues using RiboEx reagent, cDNA synthesis was performed, and then quantitative real-time PCR was used to detect the expression levels of miR-330-3p and BMI1. Alterations in the gene expression were assessed using the 2^{(-∆∆ CT)} method. The expression of miR-330-3p in all of the CRC samples was significantly lower than in adjacent healthy tissues and polyp (P<0.001). BMI1 was up-regulated in 97.9% of CRC tissue compared to healthy adjacent tissues and polyps (P<0.001). A negative reverse correlation between the miR-330-3p and BMI1 gene was observed in the CRC samples (r= -0.882, P<0.001). Down-regulation of miR-330-3p and BMI1 overexpression strongly correlates with higher tumor stage and lymph node invasion. The AUC for miR-330-3p and BMI1expression was 0.982 (sensitivity, 98.5%; specificity, 78.8%), and 0.971 (sensitivity, 97.6%; specificity, 84.6%) (P<0.001), respectively. Our results indicated that miR-330-3p and BMI1 expression probably could be considered potential diagnostic or prognostic biomarkers for CRC patient.

Current cancer therapies include chemotherapy, radiation therapy, immunotherapy, and surgery. Despite these treatment methods, a major point in cancer treatment is early detection. RNAs (mRNA, miRNAs, and LncRNA) can be used as markers to improve cancer diagnosis and treatment. This research examined how radiotherapy affected CCL5, miR-214, and MALAT-1 gene expression in the immune pathway in peripheral blood samples from radiation therapy-treated breast cancer patients. Before and after radiotherapy, peripheral blood was collected from 15 patients in four steps. Blood samples were collected in an outpatient facility from 20 healthy female volunteers with no history of malignant or inflammatory conditions. RNA was extracted from the blood samples and cDNA was synthesized. CCL5, miR-214, and MALAT-1 gene expression were determined by real-time polymerase chain reaction (RT-PCR). CCL5 protein levels in the serum were determined in 80 samples (60 BC and 20 healthy controls) using Quantikine Enzyme-Linked Immunosorbent Assay (ELISA) kits (R&D Systems). The data were then statistically evaluated. There was a significant difference between CCL5 levels in tumoral and adjacent normal blood samples (p < 0.05). The results also show that the level of gene expression and serum concentration of CCL5 protein in different phases of radiotherapy is significantly different. On the other hand, the expression level of the miR-214 gene was significantly decreased in patients compared to the control group, but this decrease was not significant for the MALAT-1 gene (p< 0.05). Also, after each stage of radiotherapy, the expression level of these two genes showed a decrease, but in the fourth week after radiotherapy, this decrease was significant (p< 0.05). Radiotherapy is associated with a decrease in the expression of miR-214 and MALAT-1, as a result, an increase in the expression of CCL5. An increase in the concentration of CCL5 protein is accompanied by an increase in the level of monocytes, which ultimately causes the infiltration of macrophages and can ultimately cause cancer recurrence. It is suggested that these genes can probably be used as diagnostic and therapeutic radiotherapy markers in breast cancer.

Capsaicin is a natural product which is extracted from pepper and has the potential to be used in cancer treatment because of its anti- proliferative effects. The aim of the study was to determine the effect of capsaicin on the hepatocellular carcinoma cell proliferation and the expressions of related genetic markers as Ki-67, PI3K/AKT/mTOR and epigenetic markers as miR-126 and piR-Hep-1. The inhibitory concentration of capsaicin in HepG2 cells was determined. piR-Hep-1 and miR-126 expressions and Ki-67, PI3K, AKT and mTOR gene expressions were examined by RT-PCR. The inhibitory concentration of capsaicin for HepG2 cells was 200 nM and the decreased proliferation was observed at 24^th hour. As epigenetic markers, an up regulation of miR-126 and down regulation of piR-Hep-1 expression were determined after treatment. Moreover, Ki-67, PI3K and mTOR gene expressions decreased while AKT gene expression increased after the treatment (p<0.001). According to the obtained data, capsaicin has an impact on proliferation both genetically and epigenetically. Furthermore, treatment of capsaicin effects miR-126 and piR-Hep-1 expressions which effect carcinogenesis in different way. Moreover, there are some clues which indicate that these two small non-coding RNA might affect each other and share the same target molecules post-transcriptionally.

The NF-kB signaling pathway was introduced as a key pathway in carcinogenesis that is induced by inflammation in gastrointestinal malignancies. The RelA transcription factor is an important component of this signaling pathway. Furthermore, CD44 is implicated in the tumorigenesis and metastasis of gastric cancer. The aim of this study was to assay the effect of RELA knockout on CD44 expression in MKN45 cells. CRISPR/Cas9 was used to knock out RELA in MKN-45. The median fluorescence intensity (MFI) of CD44 before and after RELA knockout is analyzed in MKN45. The CRISPR/Cas9 vector pSpCas9 (BB)-2A-Puro (PX459) was used for gRNA cloning (two guides). The MKN-45 cell line was co-transfected. The purified co-transfected cells with puromycin were cultured and used for the RELA gene expression assay by real-time PCR. Flow cytometry was used for the analysis of the MFI of CD44+ in MKN45. The results showed that 180 nucleotide sequences between exon 2 and exon 3 of RELA were deleted in MKN45. RELA expression significantly (P<0.001) decreased after CRISPR/Cas9 knockout. Compared to the control group, the MFI of CD44 in transfected cells significantly decreased (P <0.001). Knockout of RELA significantly decreased CD44 expression in MKN45 cells. It can be concluded that the NF-kB signaling pathway via RELA is related to CD44 expression and consequently the tumorigenesis of gastric cancer. More studies about this relationship are recommended.

Sulforaphane (SFN) is an organosulfur product of found isothiocyanates in vegetables. The chemopreventive effects of SFN have revealed that there is a link between excessive consumption of SFN-rich vegetables and cancer formation without possible toxicological consequences. We aimed to evaluate the cellular outcome of SFN from a toxicological perspective, particularly for renal cells including clear cell adenocarcinoma (769-P) and human embryonic renal epithelial (293T) cells. The viability/cytotoxicity experiments were performed with methyl thiazole diphenyl tetrazolium (MTT) and lactate dehydrogenase (LDH) assays. IC₅₀-dependent, non-cytotoxic concentrations were used for the determination of cell cycle status and apoptosis by using flow cytometry and western blot. A certain concentration of SFN effectively altered apoptotic/necrotic behavior in 769-P compared to the control group 293T. Cell cycle status remained stable while showing a decreased proliferation profile for 769-P cells. The percentage of the S phase from the cell cycle in 293T cells significantly reduced without affecting proliferation status. The use of SFN as an alternative to traditional treatments might be considered for the battle against renal cell carcinoma but the current findings showed that caution should be applied particularly for renal cells. Our study will provide a basis for future in vivo studies to support traditional cancer therapies.

The aim of the present study was to examine the hypothesis that miR-33-5p attenuates morphine state-dependent (StD) memory via the µ opioid receptor by regulating cyclic AMP response element-binding protein (CREB). The effects of post-training morphine and morphine StD memory and their interaction with pre-test naloxone were evaluated using a single-trial inhibitory avoidance paradigm. Then, the hippocampal miR-33-5p gene and pCREB/CREB protein expression profiles were evaluated using quantitative real-time PCR and western blotting, respectively. We found that while post-training morphine and morphine StD memory respectively up- and down-regulate the miR-33-5p expression profile in the hippocampus, the reverse results are true for the expression of pCREB/CREB. Pre-test naloxone antagonized the response. Overall, our findings suggest that the expression levels of miR-33-5p in the hippocampus set the basis for morphine StD memory with low miR-33-5p enabling state dependency. The mechanism is mediated via miR33-5p and CREB signaling with the interaction of the µ opioid receptor. This finding may be used as a potential strategy for ameliorating morphine-induced memory-related disorders.