Macrophages enhance cisplatin resistance in gastric cancer through the transfer of circTEX2.

Cisplatin-based chemotherapy is often used in advanced gastric cancer (GC) treatment, yet resistance to cisplatin may lead to treatment failure. Mechanisms underlying cisplatin resistance remain unclear. Recent evidence highlighted the role of macrophages in cancer chemoresistance. Macrophage-derived exosomes were shown to facilitate intercellular communication. Here, we investigated the cisplatin resistance mechanism based on macrophage-derived exosomes in gastric cancer. Cell growth and apoptosis detection experiments revealed that M2-polarized macrophages increased the resistance of GC cells to cisplatin. qRT-PCR, RNAase R assay, actinomycin D assay and cell nucleo-cytoplasmic separation experiments confirmed the existence of circTEX2 in macrophage cytoplasm, with a higher expression level in M2 macrophages than that in M1 macrophages. Further experiments showed that circTEX2 acted as microRNA sponges for miR-145 and regulated the expression of ATP Binding Cassette Subfamily C Member 1 (ABCC1). Inhibition of the circTEX2/miR-145/ABCC1 axis blocked the cisplatin resistance of gastric cancer induced by M2 macrophages, as evidenced by in vitro and in vivo experiments. In conclusion, our research suggests that the exosomal transfer of M2 macrophage-derived circTEX2 enhances cisplatin resistance in gastric cancer through miR-145/ABCC1. Additionally, communication between macrophages and cancer cells via exosomes may be a promising therapeutic target for the treatment of cisplatin-resistant gastric cancer.

Nanocomposites based on nanoceria regulate the immune microenvironment for the treatment of polycystic ovary syndrome.

The immune system is closely associated with the pathogenesis of polycystic ovary syndrome (PCOS). Macrophages are one of the important immune cell types in the ovarian proinflammatory microenvironment, and ameliorate the inflammatory status mainly through M2 phenotype polarization during PCOS. Current therapeutic approaches lack efficacy and immunomodulatory capacity, and a new therapeutic method is needed to prevent inflammation and alleviate PCOS. Here, octahedral nanoceria nanoparticles with powerful antioxidative ability were bonded to the anti-inflammatory drug resveratrol (CeO2@RSV), which demonstrates a crucial strategy that involves anti-inflammatory and antioxidative efficacy, thereby facilitating the proliferation of granulosa cells during PCOS. Notably, our nanoparticles were demonstrated to possess potent therapeutic efficacy via anti-inflammatory activities and effectively alleviated endocrine dysfunction, inflammation and ovarian injury in a dehydroepiandrosterone (DHEA)-induced PCOS mouse model. Collectively, this study revealed the tremendous potential of the newly developed nanoparticles in ameliorating the proinflammatory microenvironment and promoting the function of granulosa cells, representing the first attempt to treat PCOS by using CeO2@RSV nanoparticles and providing new insights in combating clinical PCOS.

CircSOD2 polarizes macrophages towards the M1 phenotype to alleviate cisplatin resistance in gastric cancer cells by targeting the miR-1296/STAT1 axis.

Cisplatin is the first-line drug for gastric cancer (GC). Cisplatin resistance is the most important cause of poor prognosis for GC. Increasing evidence has identified the important role of macrophage polarization in chemoresistance. CircRNAs are newly discovered non-coding RNAs, characterized by covalently closed loops with high stability. Previous studies have reported a significant difference between circRNA profiles expressed in classically activated M1 macrophages, and those expressed in alternatively activated M2 macrophages. However, the underlying mechanism behind the regulation of GC cisplatin resistance by macrophages remains unclear. In our study, we observed the aberrant high expression of circSOD2 in M1 macrophages derived from THP-1. These expression patterns were confirmed in macrophages from patients with GC. Detection of the M1 and M2 markers confirmed that overexpression of circSOD2 enhances M1 polarization. The viability of cisplatin-treated GC cells was significantly reduced in the presence of macrophages overexpressing circSOD2, and cisplatin-induced apoptosis increased dramatically. In vivo experiments showed that macrophages expressing circSOD2 enhanced the effect of cisplatin. Moreover, we demonstrated that circSOD2 acts as a microRNA sponge for miR-1296 and regulates the expression of its target gene STAT1 (signal transducer and activator of transcription 1). CircSOD2 exerts its function through the miR-1296/STAT1 axis. Inhibition of circSOD2/miR-1296/STAT1 may therefore reduce M1 polarization. Overexpression of circSOD2 promotes the polarization of M1 macrophages and enhances the effect of cisplatin in GC. CircSOD2 is a novel positive regulator of M1 macrophages and may serve as a potential target for GC chemotherapy.

Overexpression of miR-144-3p alleviates polycystic ovaries syndrome through targeting expression of HSP-70.

Increasing microRNAs are shown to be participate in polycystic ovarian syndrome (PCOS) pathogenesis. Nevertheless, the biological effects of miR-144-3p and its detailed mechanisms in PCOS are to be investigated. The purpose of our work was to study the function of miR-144-3p in PCOS. Currently, Expression of miR-144-3p was greatly reduced in PCOS patients and PCOS rat models. In addition, HSP-70 expression was greatly elevated PCOS. Cell proliferation assays and flow cytometry assay were carried out following the overexpression of miR-144-3p in ovarian granulosa cells from PCOS rat models. We observed that miR-144-3p overexpression induced the proliferation and repressed cell apoptosis while loss of miR-144-3p demonstrated an opposite process. Then, PCOS rat models were classified to four groups: LV-NC group, LV-miR-144-3p group, Anti-control group, and Anti-miR-144-3p group. In response to loss of miR-144-3p, we found E2, T, and LH serum levels were elevated and FSH serum level was inhibited. Upregulation of miR-144-3p exhibited an opposite process. Moreover, HSP-70 was a direct target of miR-144-3p. Furthermore, increased expression of HSP-70 rescued the effects of miR-144-3p on ovarian granulosa cell growth and apoptosis. In addition, knockdown of HSP-70 alleviated endocrine disorders and abnormal ovarian weight in vivo. To sum up, miR-144-3p might function as a novel target for PCOS treatment via targeting HSP-70.

DNA Methylation and Recurrent Pregnancy Loss: A Mysterious Compass?

Recurrent pregnancy loss (RPL) is a common and severe pathological pregnancy, whose pathogenesis is not fully understood. With the development of epigenetics, the study of DNA methylation, provides a new perspective on the pathogenesis and therapy of RPL. The abnormal DNA methylation of imprinted genes, placenta-specific genes, immune-related genes and sperm DNA may, directly or indirectly, affect embryo implantation, growth and development, leading to the occurrence of RPL. In addition, the unique immune tolerogenic microenvironment formed at the maternal-fetal interface has an irreplaceable effect on the maintenance of pregnancy. In view of these, changes in the cellular components of the maternal-fetal immune microenvironment and the regulation of DNA methylation have attracted a lot of research interest. This review summarizes the research progress of DNA methylation involved in the occurrence of RPL and the regulation of the maternal-fetal immune microenvironment. The review provides insights into the personalized diagnosis and treatment of RPL.

Identification of two methylated fragments of an SDC2 CpG island using a sliding window technique for early detection of colorectal cancer.

Colorectal cancer (CRC) is one of the most common cancer types globally with a 5-year survival rate of < 50% in China. Aberrant DNA methylation is one of the hallmarks of tumor initiation, progression, and metastasis. Here, we investigated the clinical performance of two differentially methylated regions (DMRs) in SDC2 CpG islands for the detection of CRC. A sliding window technique was used to identify the DMRs, and methylation-specific PCR assay was used to assess the DMRs in 198 CRC samples and 54 normal controls. Two DMRs (DMR2 and DMR5) were identified using The Cancer Genome Atlas (TCGA) data, and the hypermethylation of DMR2 and DMR5 was detected in 90.91% (180/198) and 89.90% (178/198) of CRC samples, respectively. When combining DMR2 and DMR5, the sensitivity for CRC detection was 94.4% higher than that of DMR2 or DMR5 alone. Based on the above results, we propose using DMR2 and DMR5 as a sensitive biomarker to detect CRC.

Toxicological effects of deltamethrin on quail cerebrum: Weakened antioxidant defense and enhanced apoptosis.

Deltamethrin is the most common type II synthetic pyrethroid insecticide, and has posed widespread residues to environment. However, whether deltamethrin has potential toxic effects on quail cerebrum remains greatly obscure. Accordingly, we investigated the impact of chronic exposure to deltamethrin on oxidative stress and apoptosis in quail cerebrum. Quails upon 12-week exposure of deltamethrin (0, 15, 30, or 45 mg/kg body weight intragastric administration) were used as a cerebrum injury model. The results showed that deltamethrin treatment led to cerebral injury dose-dependently through the weakened antioxidant defense by downregulating nuclear factor erythroid-2-related factor 2 (Nrf2) and its downstream proteins levels and mRNA expression. Furthermore, deltamethrin treatment induced apoptosis in cerebrum by decreasing B-cell lymphoma gene 2 (Bcl-2) level, as well as increasing Jun N-terminal kinase3, caspase-3, and Bcl-2-associated X protein levels. Simultaneously, toll-like receptor 4 (TLR4) downstream inflammation-related genes or proteins were significantly up-regulated by deltamethrin dose-dependently. Altogether, our study demonstrated that chronic exposure to deltamethrin induces inflammation and apoptosis in quail cerebrums by promoting oxidative stress linked to inhibition of the Nrf2/TLR4 signaling pathway. These results provide a novel knowledge on the chronic toxic effect of deltamethrin, and establish a theoretical foundation for the evaluation of pesticide-induced health risk.

CircPSMC3 alleviates the symptoms of PCOS by sponging miR-296-3p and regulating PTEN expression.

Polycystic ovary syndrome (PCOS), the most common female endocrine disease that causes anovulatory infertility, still lacks promising strategy for the accurate diagnosis and effective therapeutics of PCOS attributed to its unclear aetiology. In this study, we determined the abnormal reduction in circPSMC3 expression by comparing the ovarian tissue samples of PCOS patients and normal individuals. The symptom relief caused by up-regulation of circPSMC3 in PCOS model mice suggested the potential for further study. In vitro functional experiments confirmed that circPSMC3 can inhibit cell proliferation and promote apoptosis by blocking the cell cycle in human-like granular tumour cell lines. Mechanism study revealed that circPSMC3 may play its role through sponging miR-296-3p to regulate PTEN expression. Collectively, we preliminarily characterized the role and possible insights of circPSMC3/miR-296-3p/PTEN axis in the proliferation and apoptosis of KGN cells. We hope that this work provides some original and valuable information for the research of circRNAs in PCOS, not only to better understand the pathogenesis but also to help provide new clues for seeking for the future therapeutic target of PCOS.

Deltamethrin induces liver fibrosis in quails via activation of the TGF-ß1/Smad signaling pathway.

Deltamethrin (DLM) is an important member of the pyrethroid pesticide family, and its widespread use has led to serious environmental and health problems. Exposure to DLM causes pathological changes in the liver of animals and humans and can lead to liver fibrosis. However, the mechanism of DLM-induced liver fibrosis remains unclear. Therefore, to address its potential molecular mechanisms, we used both in vivo and in vitro methods. Quails were treated in vivo by intragastric administration of different concentrations of DLM (0, 15, 30, or 45 mg kg-1), and the chicken liver cancer cell line LMH was treated in vitro with various doses of DLM (0, 50, 200, or 800 µg mL-1). We found that DLM treatment in vivo induced liver fibrosis in a dose-dependent manner through the promotion of oxidative stress, activation of transforming growth factor-ß1 (TGF-ß1) and phosphorylation of Smad2/3. Treatment of LMH cells with different concentrations of DLM similarly induced oxidative stress and also decreased cell viability. Collectively, our study demonstrates that DLM-induced liver fibrosis in quails occurs via activation of the TGF-ß1/Smad signaling pathway.

SET nuclear proto-oncogene gene expression is associated with microsatellite instability in human colorectal cancer identified by co-expression analysis.

BACKGROUNDS AND AIMS: Microsatellite instability (MSI) is one of the promising biomarkers in human colorectal cancers (CRCs), and it is influenced by an intricate gene interaction network. Hence, we aimed to identify and validate hub genes associated with MSI CRC and to illustrate its underlying mechanisms. METHODS: Weighted gene co-expression network analysis (WGCNA) was used to investigate potential regulatory targets and relationships between key modules and hub genes associated with MSI CRC. RESULTS: In the red module (r = 0.83), SET nuclear proto-oncogene (SET) was selected due to its high intra-modular connectivity and module membership. In the test sets, SET expression was downregulated in MSI CRCs compared to that in microsatellite stability (MSS) CRCs. SET expression level had a good performance in stratifying patients into MSI or MSS CRCs (area under the curve = 0.953). Moreover, the BRAF V600E mutation was highly associated with SET expression, and MSI/HLA- samples showed lower levels of SET mRNA expression than MSS/HLA- samples. Finally, gene set enrichment analysis (GSEA) indicated that patients in the SET low expression group were enriched in base excision repair. CONCLUSION: SET was identified and validated as a novel potential biomarker in MSI CRCs, and SET probably acts through regulating the base excision repair pathway.

Prognostic significance of autophagy-related proteins expression in resected human gastric adenocarcinoma.

Gastric adenocarcinoma (GC) is one of the most common malignancies in the world and one of the most frequent causes of cancer-related death. Autophagy is a highly regulated catabolic pathway responsible for the degradation of long-lived proteins and damaged intracellular organelles. However, the mechanism and guiding significance of autophagy in the development and progression of GC have remained to be elucidated. This study aimed to explore the clinicopathological significances and prognostic values of autophagy-related proteins AMBRA1 and Beclin-1 in GC. Quantum dots based immunofluorescence histochemistry (QDs-IHC) was performed to observe the expression of AMBRA1 and Beclin-1 proteins in the tissue microarrays including 163 specimens of GC and 20 noncancerous gastric tissues. Simultaneously, AMBRA1 and Beclin-1 proteins were detected by Western blotting in the 10 fresh GC and corresponding normal gastric tissues. The results showed that the expression levels of both AMBRA1 and Beclin-1 proteins were higher in GC tissues than in noncancerous gastric tissues by QDs-IHC and Western blotting (P<0.05). High AMBRA1 expression was detected in 90 of 163 (55.2%) GCs and high Beclin-1 expression was detected in 83 of 163 (50.9%) GCs. High AMBRA1 expression was closely related to depth of invasion, and lymph nodes metastasis (P<0.05). High expression of Beclin-1 protein was correlated with tumor grade (P<0.05). Positive correlation was observed between AMBRA1 and Beclin-1. Survival analysis indicated the high expression of AMBRA1 and Beclin- 1 was an independent factor in predicting poor overall survival (OS) of GC patients. These findings suggest the high expression of AMBRA1 and Beclin-1 proteins is significantly correlated with GC progression. High AMBRA1 and Beclin-1 expression heralds worse outcome of GC patients, suggesting a novel candidate prognostic marker and a therapeutic target for GC.

Transfection of colorectal cancer cells with chemokine MCP-3 (monocyte chemotactic protein-3) gene retards tumor growth and inhibits tumor metastasis.

AIM: To evaluate the possibility of the induction of anti-tumor immune response by transfecting the colorectal cancer cells with chemokine MCP-3 gene. METHODS: Mouse MCP-3 gene was transduced into mouse colorectal cancer cells CMT93 by using of Liposome. G418-resistant clones were selected and the MCP-3 mRNA expression was detected by RT-PCR. The chemotactic activity of MCP-3 in the cell culture supernatant was detected by Chemotaxis assay. The tumorigenicity of wild type CMT93 and CMT93 gene transfectants were detected by in vivo experiments. The immune cell infiltrations in tumor tissue and tumor metastasis were detected histopathologically. RESULTS: MCP-3 mRNA expression was detected by RT-PCR in gene-transfected cells (CMT93/MCP-3), but not in control groups. And MCP-3 secreted in the cell culture supernatant possessed chemotatic activity. The results from in vivo experiments showed that the tumorigenicity of CMT93/MCP-3 had not decreased, but the tumors derived from CMT93/MCP-3 cells grew more slowly than those from CMT93 cells (1.021+/-0.253) cm(2) vs (1.769+/-0.371) cm(2), P<0.05) or CMT93/mock cells (1.021+/-0.253) cm(2) vs (1.680 +/-0.643)cm(2), P<0.05). Histophathological results showed few immune cells infiltrating in the tumor tissue derived from the controls. In the tumor tissue derived from CMT93/MCP-3, infiltrating immune cells increased. In addition, no tumor metastasis was found in all mice inoculated with CMT93/MCP-3 tumor cells. But all mice had tumor metastasis in CMT93 controls and 4 in 5 mice had tumor metastasis in CMT93/mock controls. CONCLUSION: The results suggested that the transfection of chemokine MCP-3 gene could promote the induction of anti-colorectal cancer immunity, but the tumor growth could not be inhibited completely by merely MCP-3 gene transfection.

[Active immunity for anti-colorectal cancer induced by chemokine MCP-3 gene transfection].

BACKGROUND & OBJECTIVE: Chemokines play an important role in the infiltration of immune cells to tumor tissues. Anti-tumor immune response had been elicited in many tumor models by the chemokine gene transfection. The aim of this study was to evaluate the possibility of inducing anti-colorectal cancer active immune response by transfection of mouse colorectal cancer CMT93 cells with chemokine MCP-3 gene. METHODS: Mouse MCP-3 gene was transduced into mouse colorectal cancer cells CMT93 by using of liposome. G418-resistant clones were selected and the MCP-3 mRNA expression was detected by RT-PCR. The chemotactic activity of MCP-3 in the cell culture supernatant was detected by chemotaxis assay. In vivo experiments were performed to observe the tumorigenicity of wild type CMT93 and MCP-3 gene modified tumor cells. The immune cell infiltration in tumor tissues and tumor metastasis were detected histopathologically. RESULTS: RT-PCR detection showed MCP-3 was expressed in MCP-3 gene-transfected G418-resistant clones(CMT93/MCP-3), but not in wild type CMT93. In chemotaxis assay, the results showed that the cell culture supernatant of CMT93/MCP-3 possess obviously chemotactic activity. The chemotactic index of the CMT93/MCP-3 supernatant was 5.57(P < 0.05). The supernatants from the control groups did not possessed the chemotactic activity. In vivo experiments showed that the tumorigenicity of CMT93/MCP-3 had not decreased significantly compared to wild type CMT93, but the tumors grew more slowly from CMT93/MCP-3 than from the controls (P < 0.05). In the tumor tissue from CMT93/MCP-3, obvious infiltrated immune cells were found, and few immune cells infiltrated in the tumor tissue from the controls. In the mice inoculated with CMT93/MCP3 tumor cells, tumor metastasis was inhibited significantly, its metastasis rate was 0(0/7), lower than that of CMT93 (100%, 4/4) and CMT93/mock (80%, 4/5) (P < 0.05). CONCLUSION: Transfection with chemokine MCP-3 gene can induce anti-colorectal cancer active immune response, but the tumor growth cannot be inhibited completely by merely MCP-3 gene transfection.

Expression of B7 costimulation molecules by colorectal cancer cells reducestumorigenicity and induces anti-tumor immunity.

AIM:To study the tumorigenicity of colorectal cancer cells transfected with B7 gene and the anti-tumor immunity induced by B7 gene modified colorectal cancer cells.METHODS:B7 gene was transfected into mouse colon cancer cell line CMT93.The transfectants were selected in DMEM containing 800mg/L G418, and B7 molecules were detected by immunohistochemistry.Experiments in vivo include: (1)5X10(6) B7(+) CMT93 cells were inoculated into the back of C57BL/6 mice subcutanously to determine their tumorigenicity (n = 4). As control, wild type CMT93 cells were inoculated the same as the experimental group (n= 3). (2) The mice primed by B7(+) CMT93 cells whose tumors vanished were rechallenged with wild type CMT93 to observe the immune protection of these mice against the wild type CMT93 (n = 4). Non-primed 4 native mice inoculated with wild type CMT93 were used as control.With in vivo cytotoxicity assay, the mice were immunized with B7 (+) CMT93 or the wild type CMT93 by intraperitoneal injection (n = 4X2). The spleen cells and the abdominal cavity infiltrating lymphocytes were obtained and cultured for two days. Cytotoxicity of these cells against the B7 gene modified or wild type CMT93 was detected by MTT assay.RESULTS:B7 high expression clones were obtained after the transfection of the B7 gene into CMT93 cells by electroporation. Immunohistochemistry results showed mainly membrance staining and partly cytoplasm staining in B7 gene transfected CMT93 cells. in vivo experiments: (1)After the inoculation of the B7(+) CMT93 cells in the back of C57BL/6 mice, they lost their tumorigenicity greatly (P < 0.01). All the small tumors growing in the early period in the experimental group vanished in one month, and the tumors in control group grew progressively. (2) No tumors were found in all 4 mice primed by B7(+) CMT93 cells after they were rechallenged with wild type CMT93. In the control group all mice had grown tumors (P < 0.05). In vitro cytotoxicity assay, the CTLs induced by B7(+) CMT93 had a higher cytotoxity against the wild type CMT93 than that induced by wild type CMT93 (P < 0.05), and the cytotoxity of CTLs induced by B7(+) CMT93 against B7(+) CMT93 cells was higher than that against wild type CMT93 cells (P < 0.05).CONCLUSION:The results suggest that the expression of costimulation B7 molecules by colorectal cancer cells can decrease their tumorigenicity greatly, and the B7 molecule can augment the activation of the CTLs against colorectal cancer, and it plays an important role in CTL effector function as well.

Radioimmunoimaging of colorectal cancer using (99m)Tc labeled monoclonal antibody.

AIM:To determine whether Hb3 and its fragment F(ab')(2) have practical value in radioimmunoimaging of colorectal cancer.METHODS:Intact Hb3 was purified by hydroxylapatite chromatography.The fragment F(ab') (2) was prepared by cold digestion and purified as intact Hb3.Hb3 and its fragment F(ab') (2) were labeled with 99mTc by direct labeling method using SnCl(2) as reducing agent. The radioactive doses ranged from 15 to 40 mCi.The imaging was accomplished by single photon emission computered tomograph (SPECT) with imaging time ranging from 2.5 to 48 hours. In this study, 10 patients were selected. Among them, 7 were administered with intact Hb3, and 3 with F(ab') (2) fragment. All the patients were diagnosed as having colorectal adenocarcinoma.RESULTS:After purification, intact Hb3 and its fragment F(ab') (2) were fit for radioimmunoimaging. The percentage of labeling of (99m)Tc to Hb3 or F(ab') (2) was 80.6%-91.5%. Among the 10 patients, 3 of 7 patients administered with intact Hb3 had positive scans, the other 4 had negative scans, and 2 of 3 patients administered with F(ab') (2)had positive scans, the other 1 had negative scans.CONCLUSION:The results showed that both intact Hb3 and its F(ab') (2) have some practical value in radioimmunoimaging of colorectal cancer, and the effects of imaging with F(ab') (2) was better than that with intact Hb3.