Metformin enhances the survival ability of ovarian granulosa cells in polycystic ovary syndrome by promoting LINC00548 to suppress androgen receptor/klotho pathway.

BACKGROUND: Metformin (Met) has been reported to play the key role in the pathogenesis of polycystic ovary syndrome (PCOS). However, the precise mechanisms underlying the actions of Met in PCOS remain incompletely understood. This study aimed to confirm the mechanism of Met interacting with a long non-coding RNA LINC00548 in PCOS. METHODS: Ovarian granulosa cells (OGCs) were incubated 500 nM dihydrotestosterone (DHT) to construct PCOS in vitro model and then were treated 20 µM Met. A series of cell experiments including Cell Counting Kit-8, Terminal uridine nucleotide end labeling, and flow cytometry were performed to confirm the changes of OGC survival ability. Quantitative real-time polymerase chain reaction was conducted to determine the levels of LINC00548, whereas Western blotting was applied to determine the levels of androgen receptor (AR) and klotho. RESULTS: Met improved the cell viability and suppressed cell apoptosis in DHT-treated OGCs. LINC00548 downregulated in DHT-treated OGCs was upregulated by Met, and its overexpression further enhanced the positive effects of Met on the survival ability of DHT-treated OGCs. In addition, Met could induce the upregulation of LINC00548 to suppress the activation of AR/klotho pathway in DHT-treated OGCs. CONCLUSION: Overall, this study discovers that Met enhances the survival ability of OGCs in PCOS by elevating LINC00548 expression to suppress AR/klotho pathway.

WTAP/IGF2BP3-mediated GBE1 expression accelerates the proliferation and enhances stemness in pancreatic cancer cells via upregulating c-Myc.

BACKGROUND: Pancreatic cancer (PC) is one of the most malignant cancers with highly aggressiveness and poor prognosis. N6-methyladenosine (m6A) have been indicated to be involved in PC development. Glucan Branching Enzyme 1 (GBE1) is mainly involved in cell glycogen metabolism. However, the function of GBE1 and Whether GBE1 occurs m6A modification in PC progression remains to be illustrated. METHODS: The clinical prognosis of GBE1 was analyzed through online platform. The expression of GBE1 was obtained from online platform and then verified in normal and PC cell lines. Lentivirus was used to generated GBE1 stable-overexpression or knockdown PC cells. Cell Counting Kit (CCK-8), colony formation assay, sphere formation assay and flow cytometry assay were conducted to analyze cell proliferation and stemness ability in vitro. Subcutaneous and orthotopic mouse models were used to verify the function of GBE1 in vivo. RNA immunoprecipitation (RIP) assay, RNA stability experiment and western blots were conducted to explore the molecular regulation of GBE1 in PC. RESULTS: GBE1 was significantly upregulated in PC and associated with poor prognosis of PC patients. Functionally, GBE1 overexpression facilitated PC cell proliferation and stemness-like properties, while knockdown of GBE1 attenuated the malignancy of PC cells. Importantly, we found the m6A modification of GBE1 RNA, and WTAP and IGF2BP3 was revealed as the m6A regulators to increase GBE1 mRNA stability and expression. Furthermore, c-Myc was discovered as a downstream gene of GBE1 and functional rescue experiments showed that overexpression of c-Myc could rescue GBE1 knockdown-induced PC cell growth inhibition. CONCLUSIONS: Our study uncovered the oncogenic role of GBE1/c-Myc axis in PC progression and revealed WTAP/IGF2BP3-mediated m6A modification of GBE1, which highlight the potential application of GBE1 in the targeted therapy of PC.

Hepatitis B time series in Xinjiang, China (2006-2021): change point detection based on the Mann-Kendall-Sneyers test.

Hepatitis B is a major global challenge, but there is a lack of epidemiological research on hepatitis B incidence from a change point perspective. This study aimed to fill this gap by identifying significant change points and trends in hepatitis time series in Xinjiang, China. The datasets were obtained from the Xinjiang Information System for Disease Control and Prevention. The Mann-Kendall-Sneyers (MKS) test was used to detect change points and trend changes on the hepatitis B time series of 14 regions in Xinjiang, and the effectiveness of this method was validated by comparing it with the binary segmentation (BS) and segment regression (SR) methods. Based on the results of change point analysis, the prevention and control policies and measures of hepatitis in Xinjiang were discussed. The results showed that 8 regions (57.1%) with at least one change fell within the 95% confidence interval (CI) in all 14 regions by the MKS test, where five regions (Turpan (TP), Hami (HM), Bayingolin (BG), Kyzylsu Kirgiz (KK), Altai (AT)) were identified at one change point, two change points existed for two regions (Aksu (AK), Hotan (HT)) and three change points was detected in 1 region (Bortala (BT)). Most of the change points occurred at both ends of the sequence. More change points indicated an upward trend in the front half of the sequence, while in the latter half, many change points indicated a downward trend prominently. Finally, in comparing the results of the three change point tests, the MKS test showed a 61.5% agreement (8/13) with the BS and SR.

Research Progress on Biological Evidence Identification in Fire Scenes.

Biological evidence is relatively common evidence in criminal cases, and it has strong probative power because it carries DNA information for individual identification. At the scene of fire-related cases, the complex thermal environment, the escape of trapped people, the firefighting and rescue operations, and the deliberate destruction of criminal suspects will all affect the biological evidence in the fire scene. Scholars at home and abroad have explored and studied the effectiveness of biological evidence identification in fire scenes, and found that the blood stains, semen stains, bones, etc. are the main biological evidence which can be easily recovered with DNA in fire scenes. In order to analyze the research status and development trend of biological evidence in fire scenes, this paper systematically sorts out the relevant research, mainly including the soot removal technology, appearance method of typical biological evidence, and possibility of identifying other biological evidence. This paper also prospects the next step of research direction, in order to provide reference for the identification of biological evidence and improve the value of biological evidence in fire scenes.

Multiple change point analysis of hepatitis B reports in Xinjiang, China from 2006 to 2021.

Objective: Hepatitis B (HB) is a major global challenge, but there has been a lack of epidemiological studies on HB incidence in Xinjiang from a change-point perspective. This study aims to bridge this gap by identifying significant change points and trends. Method: The datasets were obtained from the Xinjiang Information System for Disease Control and Prevention. Change points were identified using binary segmentation for full datasets and a segmented regression model for five age groups. Results: The results showed four change points for the quarterly HB time series, with the period between the first change point (March 2007) and the second change point (March 2010) having the highest mean number of HB reports. In the subsequent segments, there was a clear downward trend in reported cases. The segmented regression model showed different numbers of change points for each age group, with the 30-50, 51-80, and 15-29 age groups having higher growth rates. Conclusion: Change point analysis has valuable applications in epidemiology. These findings provide important information for future epidemiological studies and early warning systems for HB.

Clinical Efficacy of Bolus of Six Drugs Including Rehmannia as an Adjunct to Metformin in the Treatment of Senile Type-2 Diabetes Mellitus and its Influence on Insulin Resistance, Inflammatory Factors and Blood Glucose-related Indicators.

Objective: To investigate the clinical efficacy of Bolus of Six Drugs Including Rehmannia (Liuwei Dihuang pill) as an adjunct to metformin in the treatment of senile Type-2 diabetes mellitus and its influence on insulin resistance, inflammatory factors and blood glucose-related indexes. Methods: This is a Retrospective study. Eighty senile Type-2 diabetes mellitus admitted to Baoding Hospital of Traditional Chinese Medicine from January 2019 to December 2021 were enrolled and divided into two groups using the random number method. Patients in the control group were given oral metformin, while those in the observation group were treated with Bolus of six Drugs Including Rehmannia as an adjunct to metformin. The clinical efficacy, blood glucose-related indicators, insulin-related indicators, inflammatory factors-related indicators and adverse drug reactions were compared between the two groups. Results: The overall response in the observation group was higher than that in the control group(P<0.05). After treatment, the levels of FPG, 2hPG and HbA1 being more significantly lower in the observation group than that in the control group(P<0.05). Moreover, the levels of FINS, HOMA-IR and HOMA-IS were all significantly improved in the observation group than that the control group (P<0.05). HOMA-ß levels in the observation group were significantly higher than those in the control group (P<0.05). The levels of TNF-α, IL-6 and IL-8 in the observation group were significantly lower than those in the control group (P<0.05). Conclusion: Bolus of six Drugs Including Rehmannia as an adjunct to metformin is a regimen with satisfactory safety profile for the treatment of senile Type-2 diabetes mellitus.

Correlation between SMADs and Colorectal Cancer Expression, Prognosis, and Immune Infiltrates.

Background: In recent years, the incidence and mortality of colorectal cancer (CRC) are increasing, and the 5-year survival rate of advanced metastatic CRC is poor. Small mothers against decapentaplegic (SMAD) superfamily are intracellular signal transduction proteins associated with the development and prognosis of a variety of tumors. At present, no study has systematically analysed the relationship between SMADs and CRC. Methods: Here, R3.6.3 was used to analyse the expression of SMADs in pan-cancer and CRC. Protein expression of SMADs were analysed by Human Protein Atlas (HPA). Gene expression profiling interactive analysis (GEPIA) was used to evaluate the correlation between SMADs and tumor stage in CRC. The effect of R language and GEPIA on prognosis was analysed. Mutation rates of SMADs in CRC were determined by cBioPortal, and potentially related genes were predicted using GeneMANIA. R analysis was used to correlate immune cell infiltration in CRC. Results: Both SMAD1 and SMAD2 were found to be weakly expressed in CRC and correlated with the immune invasion level. SMAD1 was correlated with patient prognosis, and SMAD2 was correlated with tumor stage. SMAD3, SMAD4, and SMAD7 were all expressed at low levels in CRC and associated with a variety of immune cells. SMAD3 and SMAD4 proteins were also expressed at low levels, and SMAD4 had the highest mutation rate. SMAD5 and SMAD6 were overexpressed in CRC, and SMAD6 was also associated with patient overall survival (OS) and CD8+ T cells, macrophages, and neutrophils. Conclusions: Our results reveal innovative and strong evidence that SMADs can be used as biomarkers for the treatment and prognosis of CRC.

Genkwanin Alleviates Mitochondrial Dysfunction and Oxidative Stress in a Murine Model of Experimental Colitis: The Participation of Sirt1.

OBJECTIVE: Genkwanin is a biologically active O-methylated flavone extracted from Daphne genkwa. An increasing number of studies have described the modulatory effects of genkwanin on human diseases, including antitumor, anti-inflammatory, and antioxidant activities. However, little is known about whether genkwanin might be a therapeutic agent for inflammatory bowel disease or its possible underlying mechanisms. MATERIALS AND METHODS: Forty C57BL/6 male mice were orally administered dextran sulfate sodium (DSS) to generate the colitis model, and genkwanin was orally administered at the indicated concentrations. Body weight, disease activity index, colon length, and H&E staining were used to evaluate colitis. Oxidative stress and antioxidant levels were measured by detecting ROS generation and malondialdehyde, superoxide dismutase and glutathione levels. The levels of proinflammatory cytokines (TNF-α, IL-1ß, IFNγ and IL-6) were measured using ELISAs. Cell viability was determined using the CCK-8 assay. Mitochondrial function was evaluated by measuring the oxygen consumption rate, mtDNA content, and activities of electron transfer chain (ETC) complexes I, II, and IV. The expression of SIRT1, Nrf2 and its target genes was determined using qRT-PCR and western blotting. SIRT1 was depleted by lentivirus-mediated knockdown. RESULTS: In this study, oral administration of genkwanin alleviated colitis induced by oral administration of DSS in mice, as evidenced by reduced weight loss, colon length shortening and histopathology scores. Furthermore, genkwanin relieved oxidative stress and reduced the production of proinflammatory cytokines. In vitro assays revealed that genkwanin administration inhibited reactive oxygen species (ROS) production and improved mitochondrial function in human intestinal epithelial cells. Genkwanin also upregulated the expression of SIRT1, and lentivirus-mediated SIRT1 knockdown partially abrogated the protective effect of genkwanin on oxidative stress and mitochondrial dysfunction. CONCLUSIONS: Findings from our murine model and cell culture experiments provide a promising basis for genkwanin to be studied as a treatment for IBD in clinical trials.

Synthesis, crystal structure and thermal properties of an unsymmetrical 1,2,4,5-tetrazine energetic derivative.

A new unsymmetrical s-tetrazine derivative, namely 4-({2-[6-(3,5-dimethyl-1H-pyrazol-1-yl)-1,2,4,5-tetrazin-3-yl]hydrazin-1-ylidene}methyl)phenol (DPHM), C14H14N8O, was synthesized based on 3-(3,5-dimethylpyrazol-1-yl)-6-hydrazinyl-s-tetrazine (DPHT). The structure was characterized by elemental analysis and single-crystal X-ray diffraction. Crystal structure determination shows that DPHM crystallizes in the monoclinic P21/c space group with high coplanarity and a zigzag layered structure. In addition, its thermal behaviour was investigated by DSC and TG-DTG methods. The thermal safety of DPHM was evaluated by self-accelerating decomposition temperature (TSADT), critical temperature of thermal explosion (Tb), entropy of activation (ΔS=), enthalpy of activation (ΔH=) and free energy of activation (ΔG=). Meanwhile, the kinetic parameters and specific heat capacity of DPHM were also determined. The results show that DPHM has better stability and detonation properties than 3-(2-benzylidenehydrazin-1-yl)-6-(3,5-dimethylpyrazol-1-yl)-s-tetrazine (DAHBTz), due to the introduction of a hydroxy group, which increases the number of hydrogen-bond interactions and improves the stability and density of DPHM. This study demonstrates that the performance of an explosive can be optimized through structural modification.

The correlation between exposure to BPA and the decrease of the ovarian reserve.

OBJECTIVE: This study aimed to evaluate whether exposure to bisphenol A (BPA) affects the ovarian reserve. METHODS: Follicular fluid (FF) was collected from diminished ovarian reserve (DOR) and non-DOR patients who underwent in vitro fertilization or intracytoplasmic sperm injection. ELISA was used to detect the BPA and hormones levels in 54 cases of DOR and 67 cases of non-DOR. A total of 64, five-week-old SPF C57BL/6 mice were randomly divided into four groups, of which three were exposed to 5, 50, and 500 µg/kg/day of BPA solution, and one was exposed to con oil only as the control. The weight and estrus of each mouse were recorded daily, and the E2 hormone and anti-Müllerian hormone (AMH) in the serum were detected by ELISA. The expression levels of AMH mRNA and protein were also detected. RESULTS: The BPA levels in the FF of DOR patients were significantly higher than those of non-DOR patients (234.048±81.736 ng/L vs. 193.300±67.225 ng/L, P<0.01); The AMH and E2 levels in the FF of DOR patients were lower than those of non-DOR patients ([555.689+74.224] pg/ml vs. [587.178+77.731] pg/ml, P<0.05, [209.720+31.556] pg/ml vs. [221.845+32.632] pg/ml, P<0.05). The BPA concentration was correlated with the AMH and E2 levels in the FF (rBPA & AMH=-0.312, P<0.05; rBPA & E2=-0.290, P<0.05); in the animal experiment, the levels of serum AMH and E2 as well as the expression levels of the AMH gene and protein in the BPA treatment group displayed downward trends. The concentrations of the 5 and 500 µg/kg/day groups decreased significantly (P<0.05). CONCLUSION: The increased BPA in the FF may promote the pathogenesis of DOR. BPA did not present a single-dose effect on the mouse ovary. Sub-chronic exposure to a low dose of BPA can reduce the ovarian reserve in female mice.

Investigation of changes in endocannabinoids and N-acylethanolamides in biofluids, and their correlations with female infertility.

Female infertility is a worldwide medical problem, and the scarcity of infertility biomarkers has hindered the ability to launch preventive and therapeutic measures in a timely manner. Intriguingly, alterations in endocannabinoids (eCBs) and N-acylethanolamides (NAEs) have been observed in the biofluids of infertile females. Therefore, a hypothesis of using eCB and NAEs in biofluids as infertility biomarkers was proposed by several researchers; however, little evidence exists to verify the hypothesis. To investigate their correlations with female infertility, we developed a magnetic liquid microextraction-chemical derivatization (MLME-CD) method coupled with liquid chromatography-tandem mass spectrometry for the quantification of eCBs and NAEs in biofluids. The target compounds were first purified with magnetic toluene as sorbents, and then labeled with 4-(N,N-dimethyamino)benzoyl chloride (4-DMABC). The MLME-CD method offered several advantages, including reliable quantification results by preventing the isomerization of eCB, high throughput by requiring 20min for sample preparation, and good sensitivity with limits of detection at 3.0-54.3 fmol. The intra-day and inter-day relative standard deviations were below 14.5%, and the recoveries were 87.4%-117.9%. Concentrations of eCBs and NAEs in the serum of 49 infertile women and 53 fertile women (controls), and in the ovarian follicular fluid of 21 infertile women and 20 controls were then quantified. Using unpaired t test analysis indicated significant differences in AEA and PEA in serum, and OEA in follicular fluid between infertile women and healthy controls, and the areas under the curve were in the range of 0.605-0.707.

Massively Parallel Sequencing (MPS) of Cell-Free Fetal DNA (cffDNA) for Trisomies 21, 18, and 13 in Twin Pregnancies.

Massively parallel sequencing (MPS) technology has become increasingly available and has been widely used to screen for trisomies 21, 18, and 13 in singleton pregnancies. This study assessed the performance of MPS testing of cell-free fetal DNA (cffDNA) from maternal plasma for trisomies 21, 18, and 13 in twin pregnancies. Ninety-two women with twin pregnancies were recruited. The results were identified through karyotypes of amniocentesis or clinical examination and follow-up of the neonates. Fluorescent in-situ hybridization was used to examine the placentas postnatally in cases of false-positive results. The fetuses with autosomal trisomy 21 (n = 2) and trisomy 15 (n = 1) were successfully detected via MPS testing of cffDNA. There was one false-positive for trisomy 13 (n = 1), and fluorescence in-situ hybridization (FISH) identified confined placental mosaicism in this case. For twin pregnancies undergoing second-trimester screening for trisomy, MPS testing of cffDNA is feasible and can enhance the diagnostic spectrum of non-invasive prenatal testing, which could effectively reduce invasive prenatal diagnostic methods. In addition to screening for trisomy 21, 18, and 13 by cffDNA, MPS can detect fetal additional autosomal trisomy. False-positive results cannot completely exclude confined placental mosaicism.