[Expression of DARS2 in colorectal cancer and its clinical significance].

Objective: To investigate the expression of DARS2 and its clinical significance in colorectal cancer. Methods: In this study, bioinformatics tools, especially gene expression profile interactive analysis 2 (GEPIA2), were used to conduct an in-depth analysis of DARS2 expression in colorectal cancer tissues. Immunohistochemical staining was carried out in 108 colorectal cancer specimens and 30 normal colorectal tissues obtained from the First Affiliated Hospital of Nanchang University, Nanchang, China. Colorectal cancer cell lines (HCT116 and SW480) were transfected with small interfering RNA (siRNA) and DARS2 overexpression plasmid to examine the effects of DARS2 knockdown and overexpression on cell function. To assess the effects on cell function, CCK8 and transwell migration assays were used to assess proliferation and cell motility, respectively. Additionally, protein immunoblotting was employed to scrutinize the expression of proteins associated with the epithelial-mesenchymal transition of colorectal cancer cells. Results: DARS2 exhibited a pronounced upregulation in expression within colorectal cancer tissues compared to their normal epithelial counterparts. Furthermore, DARS2 expression was higher in colorectal cancer of stage Ⅲ-Ⅳ than those of stage Ⅰ-Ⅱ, exhibiting a significant correlation with N staging, M staging, and pathological staging (P<0.05). Kaplan-Meier analyses showed a decreased overall survival rate in colorectal cancer with DARS2 expression compared to those without DARS2 expression (P<0.05). In the siRNA transfection group, there was a significant reduction in cell proliferation and migration (P<0.01 and P<0.05, respectively). Conversely, the transfection of DARS2 overexpression plasmids substantially increased both cell proliferation and migration (P<0.05). Additionally, immunoblotting revealed that DARS2 knockdown led to an upregulation of E-cadherin expression and a downregulation of N-cadherin and vimentin expression. In contrast, DARS2 overexpression resulted in increased N-cadherin and vimentin expression, coupled with reduction in E-cadherin expression. Conclusions: There is a strong association between DARS2 expression and colorectal cancer progression. Silencing DARS2 inhibits cell proliferation and migration, exerting a discernible influence on the epithelial-mesenchymal transition process.

[Epidemic situation of malaria in Lishui City from 2013 to 2018].

OBJECTIVE: To analyze the epidemic situation and epidemiological characteristics of malaria in Lishui City from 2013 to 2018, so as to provide the evidence for formulating the malaria control strategy. METHODS: The data pertaining malaria cases in Lishui City from 2013 to 2018 were captured from National Notifiable Communicable Disease Reporting System and the Information System for Parasitic Diseases Control and Prevention, and the epidemiological features of malaria cases were analyzed. RESULTS: A total of 119 malaria cases were reported in Lishui City from 2013 to 2018, including 101 cases with falciparum malaria (84.87%), 6 cases with vivax malaria (5.04%), 8 cases with ovale malaria (6.72%), and 4 cases with mixed infection (3.36%). Among the 119 cases, there were one local case with blood transfusion-induced malaria and 118 cases with over- seas imported malaria. There were 98.32% of the imported malaria cases acquiring infection in African countries, and most cases were reported in Qingtian County (60.50%) and Liandu District (22.69%). In addition, 86.55% of the malaria cases were detected in individuals at ages of 20 to 50 years, and most cases were found in oversea workers (52.94%) and businessmen (38.65%). CONCLUSIONS: Most of the malaria cases in Lishui City are imported from Africa, and the monitoring and health education pertaining to malaria control knowledge requires to be intensified among high-risk populations.

MiR-199a modulates autophagy and inflammation in rats with cerebral infarction via regulating mTOR expression.

OBJECTIVE: The aim of this study was to investigate the roles of micro ribonucleic acid (miR)-199a in rats with cerebral infarction by regulating mammalian target of rapamycin (mTOR). MATERIALS AND METHODS: A total of 36 Sprague-Dawley rats were randomly assigned into three groups, including: sham group (n=12), model group (n=12) and miR-199a mimics group (n=12). In sham group internal and external carotid arteries were exposed. The ischemia-reperfusion model was successfully established using suture embolization in the other two groups. After modeling, rats in sham group and model group were intraperitoneally injected with normal saline. However, rats in miR-199a mimics group were injected with miR-199a mimics. Following intervention for 3 d, sampling was conducted. Neurological deficit was evaluated in rats based on the Zea-Longa scoring system. Hematoxylin-eosin (HE) staining was performed to observe neuronal morphology. The expression of mTOR was detected using immunohistochemistry, and the relative expression level of tau protein was determined via Western blotting (WB). Besides, the messenger RNA (mRNA) expressions of mTOR and tau were detected by quantitative Polymerase Chain Reaction (qPCR). Finally, inflammatory factor content was measured through enzyme-linked immunosorbent assay (ELISA). RESULTS: Model group and miR-199a mimics group exhibited a substantially higher Zea-Longa score than sham group (p<0.05). Compared with model group, the Zea-Longa score rose prominently in miR-199a mimics group (p<0.05). According to the results of HE staining, the structure of neurons in sham group was clear and intact, while the structure of neurons in model group was disordered. Meanwhile, neuronal morphology in miR-199a mimics group was significantly worse than that in model group (p<0.05). Immunohistochemistry results demonstrated that the positive expression level of mTOR was considerably upregulated in both model group and miR-199a mimics group in comparison with sham group (p<0.05). Moreover, its positive expression level in miR-199a mimics group was markedly higher that in model group (p<0.05). Based on the results of WB, model and miR-199a mimics groups exhibited a remarkably higher relative expression level of tau protein than sham group (p<0.05). However, the relative expression level of tau protein in miR-199a mimics group was prominently higher than that in model group (p<0.05). QPCR results manifested that the relative mRNA expression levels of mTOR and tau in model group and miR-199a mimics group were dramatically higher than those in sham group (p<0.05). Compared with those in model group, the relative mRNA expression levels of mTOR and tau increased significantly in miR-199a mimics group (p<0.05). ELISA results revealed that model group and miR-199a mimics group had prominently higher content of inflammatory factors than sham group (p<0.05). In addition, content of inflammatory factors in miR-199a mimics group was considerably higher than that in model group (p<0.05). CONCLUSIONS: MiR-199a modulates mTOR expression to exert important regulatory effects on the autophagy and inflammation in rats with cerebral infarction.

[Expression and mechanism of Twist2 in glioma].

Objective: To investigate the significance of Twist2 in glioma and whether it is involved in the malignant transformation of glioma by epithelial-mesenchymal transition (EMT). Methods: Using immunohistochemical method detected the expression level of Twist2 in 60 cases of gliomas (including WHO grades Ⅱ, Ⅲ and Ⅳ, each for 20 cases) and 20 cases of non-tumor brain tissues. Real-time fluorescence quantitative PCR and Western blot were used to detect the expression level of Twist2 mRNA and protein in 61 cases of fresh glioma tissue (WHO grade Ⅱ 16 cases, Ⅲ 21 cases, Ⅳ 24 cases) and 12 cases of adjacent tissues, and the expression levels of E-cadherin, N-cadherin and vimentin were also investigated in fresh glioma tissue. Results: Immunohistochemistry results showed that the percentages of Twist2 expression in glioma was 90%(54/60) compared with 30%(6/20) in non-tumor brain tissues(P<0.01). The percentages of Twist2 expression were 75% (15/20), 95% (19/20), and 100% (20/20) in the WHO gradesⅡ, Ⅲ and Ⅳ gliomas, respectively. WHO grades Ⅳ and Ⅲ were significantly higher than that of WHO grade Ⅱ (P<0.01). There was no significant difference between WHO grade Ⅳand WHO Ⅲ glioma (P>0.05). Real-time fluorescence quantitative PCR and Western blot showed that the expression level of Twist 2 in gliomas was significantly higher than that in para-cancerous tissues (P<0.01), and those in WHO grades Ⅳ and Ⅲ gliomas were significantly higher than that in WHO grade Ⅱ glioma (P<0.01). There was no significant difference between WHO grade Ⅳand grade Ⅲ glioma (P>0.05). Detection of key protein expression in EMT by Western blot displayed that the expression of E-cadherin was negatively associated with Twist2 in glioma (r=-0.972, P<0.01). The expression of N-cadherin and vimentin was positively associated with Twist2 in glioma(r=0.971, P<0.01; r=0.968, P<0.01). Conclusions: The expression of Twist2 in human glioma is positively correlated with the malignant grade of glioma, which may be involved in the malignant progression of glioma by EMT.

[Role of ash2 (absent, small, or homeotic)-like and Jumonji domain-containing protein 3 on histone methylation of interferon-gamma gene and their associations with vascular damage of Kawasaki disease].

Objective: To investigate the impacts of ash2 (absent, small, or homeotic)-like (Ash2L) and Jumonji domain-containing protein 3 (Jmjd3) on histone methylation of interferon-gamma(IFN-γ) gene and association with vascular damage of Kawasaki disease (KD) in acute phase. Methods: This study was performed among 36 children with KD in acute phase (KD group) and 28 age-matched health children (control group), who were treated or underwent physical examination in our hospital between February 2015 and June 2016. Patients were further divided into KD groups with or without coronary artery lesions (KD-CAL(+) , 16 cases; KD-CAL(-), 20 cases). All KD patients were treated with intravenous immunoglobulin. The proportion of type 1 helper T(Th1) cells and protein levels of IFN-γ, T-box expressed in T cells(T-bet), phosphorylated signal transducer and activator of transcription 1(pSTAT1) and phosphorylated signal transducer and activator of transcription 4(pSTAT4) were analyzed by flow cytometry.Chromatin immunoprecipitation was performed to determine histone methylation (histone H3 tri-methyl K4(H3K4me3), histone H3 tri-methyl K27(H3K27me3)) and binding levels of Ash2L, Jmjd3 and Ezh2 associated with IFN-γ in CD4(+) T cells. Quantitative real-time PCR was used to determine mRNA levels of IFN-γ, interferon γ receptor 1(IFN-γR1), interferon γ receptor 2(IFN-γR2), interleukin 12 receptor subunit beta 1(IL-12Rß1), interleukin 12 receptor subunit beta 2(IL-12Rß2), interleukin 18 receptor subunit beta α(IL-18Rα), interleukin 18 receptor subunit beta ß(IL-18Rß), tumor necrosis factor receptor 1(TNFR1), toll-like receptor 4(TLR4), receptor interacting serine/threonine kinase 1(RIP-1) and myeloid differentiation primary response gene 88(MyD88) in CD4(+) T cells. Plasma concentrations of IFN-γ, interleukin 12(IL-12), interleukin 18(IL-18) and tumor necrosis factor α(TNF-α) were measured by enzyme-linked Immunosorbent assay. Results: (1)The proportion of Th1 and its protein level of IFN-γ were significantly higher in KD group than those in control group and higher in KD-CAL(+) group than in KD-CAL(-) group (all P<0.05), and lower after treatment than before treatment (all P<0.05). (2)Compared with control group, mRNA level of IFN-γ and IFN-γ-associating H3K4me3 was increased, while level of IFN-γ associating H3K27me3 in CD4(+) T cells was reduced in KD group (all P<0.05), which resulted in a higher rate of H3K4me3/H3K27me3 (P<0.05) in KD group, which was positively correlated with IFN-γ mRNA in KD group(r=0.55, P<0.05). Similar results were found between KD-CAL(+) group and KD-CAL(-) group (all P<0.05). Level of IFN-γ associating H3K27me3 was increased, and mRNA level of IFN-γ and IFN-γ associating H3K4me3 was decreased after treatment than before treatment (all P<0.05). (3)Expression of T-bet protein and binding levels of Ash2L and Jmjd3 with IFN-γ gene were significantly higher in KD group than those in control group(all P<0.05), higher in KD-CAL(+) group than those in KD-CAL(-) group (all P<0.05). These parameters were significantly lower after treatment than before treatment (all P<0.05). Binding level of Ezh2 with IFN-γ gene was similar among various groups (all P>0.05). (4)In comparison with control or after treatment, surface receptors(IFN-γR1/2, IL-12Rß1/2, IL-18Rα/ß, TNFR1 and TLR4) and its downstream molecules(pSTAT1, pSTAT4, RIP(1) and MyD88) in CD4(+) T cells, and plasma concentrations of inflammatory cytokines(IFN-γ, IL-12, IL-18 and TNF-α) were found to be higher in KD group(all P<0.05). These parameters were also higher in KD-CAL(+) group than in KD-CAL(-) group (all P<0.05). Conclusion: Aberrant histone methylation of IFN-γ associating H3K4me3 and H3K27me3 caused by over-binding of Ash2L and Jmjd3 might be involved in immune dysfunction and vascular damage in KD in the acute phase.