Antidementia medication acetylcholinesterase inhibitors have therapeutic benefits on osteoporotic bone by attenuating osteoclastogenesis and bone resorption.

This study was designed to determine whether the use of acetylcholinesterase inhibitors (AChEIs), a group of drugs that stimulate acetylcholine receptors and are used to treat Alzheimer's disease (AD), is associated with osteoporosis protection and inhibition of osteoclast differentiation and function. Firstly, we examined the effects of AChEIs on RANKL-induced osteoclast differentiation and function with osteoclastogenesis and bone resorption assays. Next, we investigated the impacts of AChEIs on RANKL-induced nuclear factor κB and NFATc1 activation and expression of osteoclast marker proteins CA-2, CTSK and NFATc1, and dissected the MAPK signaling in osteoclasts in vitro by using luciferase assay and Western blot. Finally, we assessed the in vivo efficacy of AChEIs using an ovariectomy-induced osteoporosis mouse model, which was analyzed using microcomputed tomography, in vivo osteoclast and osteoblast parameters were assessed using histomorphometry. We found that Donepezil and Rivastigmine inhibited RANKL-induced osteoclastogenesis and impaired osteoclastic bone resorption. Moreover, AChEIs reduced the RANKL-induced transcription of Nfatc1, and expression of osteoclast marker genes to varying degrees (mainly Donepezil and Rivastigmine but not Galantamine). Furthermore, AChEIs variably inhibited RANKL-induced MAPK signaling accompanied by downregulation of AChE transcription. Finally, AChEIs protected against OVX-induced bone loss mainly by inhibiting osteoclast activity. Taken together, AChEIs (mainly Donepezil and Rivastigmine) exerted a positive effect on bone protection by inhibiting osteoclast function through MAPK and NFATc1 signaling pathways through downregulating AChE. Our findings have important clinical implications that elderly patients with dementia who are at risk of developing osteoporosis may potentially benefit from therapy with the AChEI drugs. Our study may influence drug choice in those patients with both AD and osteoporosis.

Single-cell analyses reveal distinct expression patterns and roles of long non-coding RNAs during hESC differentiation into pancreatic progenitors.

BACKGROUND: Deep understanding the differentiation process of human embryonic stem cells (hESCs) is essential for developing cell-based therapeutic strategy. Substantial efforts have been made to investigate protein-coding genes, yet it remains lacking comprehensive characterization of long non-coding RNAs (lncRNAs) during this process. METHODS: hESCs were passaged every 5-6 days and had maintained stable karyotype even until the 50th generation. Pancreatic progenitor specification of in vitro differentiation from hESCs was performed and modified. The nuclei were stained with 4,6-Diamidino-2-phenylindole (DAPI). Droplet-based platform (10X Genomics) was applied to generate the single-cell RNA sequencing (scRNA-seq) data. The quality of the filtered read pairs was evaluated by using FastQC. Batch effects were removed using the size factor method. Dimension reduction and unsupervised clustering analyses were performed using Seurat R package. The Monocle 2 and MetaCell algorithms were used to order single cells on a pseudotime course and partition the scRNA-seq data into metacells, respectively. Co-expression network was constructed using WGCNA. Module- and hub-based methods were adopted to predict the functions of lncRNAs. RESULTS: A total of 77,382 cells during the differentiation process of hESCs toward pancreatic progenitors were sequenced. According to the single-cell map, the cells from different time points were authenticated to constitute a relatively homogeneous population, in which a total of 7382 lncRNAs could be detected. Through further analyzing the time course data, conserved and specific expression features of lncRNAs during hESC differentiation were revealed. Based upon pseudotime analysis, 52 pseudotime-associated lncRNAs that grouped into three distinct expression patterns were identified. We also implemented MetaCell algorithm and network-based methods to explore the functional mechanisms of these lncRNAs. Totally, 464 lncRNAs, including 49 pseudotime-associated lncRNAs were functionally annotated by either module-based or hub-based methods. Most importantly, we demonstrated that the lncRNA HOTAIRM1, which co-localized and co-expressed with several HOX genes, may play crucial role in the generation of pancreatic progenitors through regulation of exocytosis and retinoic acid receptor signaling pathway. CONCLUSIONS: Our single-cell analyses provide valuable data resources for biological researchers and novel insights into hESC differentiation processes, which will guide future endeavors to further elucidate the roles of lncRNAs.

Elevated Insulin-Like Growth Factor-1-Induced Female Rats Perpetuate the Polycystic Ovary Syndrome Phenotype: Pathological Mechanism of Insulin-Like Growth Factor-1 in Polycystic Ovary Syndrome.

OBJECTIVES: This study was designated to establish a polycystic ovary syndrome (PCOS) rat model with recombinant human insulin-like growth factor-1 (RH-IGF-1). We made assessment on the characteristics of hyperinsulinemia and hyperandrogenism in the rat model. DESIGN: This study performed the characteristics of PCOS upon RH-IGF-1 injection and evaluated the disease process of PCOS syndrome caused by the insulin-resistant pathological condition of IGF-1 based on the comparative study of in vivo test. SETTING: The experiment was conducted in the experimental research center of Yinzhou NO.2 hospital, Ningbo, Zhejiang Province, China. MATERIALS AND METHODS: Thirty-four female Sprague Dawley immature rats aged 21 days were randomly divided into two groups. Those treated with RH-IGF-1 2 mg/100 g daily were in RH-IGF-1 group (n = 20), and those with 0.9% sodium chloride 0.2 mL/100 g daily were in the saline group (n = 14). The experiment was carried out in two stages. In stage I, rats were anesthetized upon the first estrous cycle in the saline group with tissue and blood samples collected (n = 7), and rats in the RH-IGF-1-treated group were anesthetized on the 5th day after vaginal opening (VO) (n = 10). In stage II, rats in the saline group were anesthetized after three complete cycles (n = 7), meanwhile, while on the 15th day after VO (n = 10) for those in the RH-IGF-1 group. RESULTS: We have found that compared with the control group, rats injected with RH-IGF-1 expressed an early VO, disordered estrous cycle, polycystic ovaries, and significantly increased ovarian weight/body weight ratio. And from the perspective of hormone secretion, their androgen increased significantly and the insulin resistance index also elevated distinctly, possessing main characteristics similar to PCOS. LIMITATIONS: In this study, we were limited by the inability to examine IGF-1 in hypothalamus. IGF-1 in hypothalamus and in vitro experiments would be taken into consideration for further study in the future. CONCLUSIONS: These findings suggest that IGF-1 may be a key factor in the pathogenesis of PCOS, and the increase of androgen may be the pathological result, not the cause of PCOS.

LncRNA HCG11 represses ovarian cancer cell growth via AKT signaling pathway.

AIM: Ovarian cancer is a main contributor of cancer-relevant deaths among women worldwide due to high incidence and mortality. Mounting evidence has unveiled that lncRNAs play critical roles in malignancies, including ovarian cancer. Although the tumor suppressor function of HCG11 in prostate cancer and glioma has been proved, investigations on HCG11 role in ovarian cancer are still scarce. METHODS: Gene or protein expression was quantified by RT-qPCR or western blot. HCG11 effects on ovarian cancer were assessed by functional assays. Bioinformatics analysis and mechanism experiments were implemented to identify the association among HCG11, miR-1270, and PTEN. RESULTS: HCG11 was weakly expressed in ovarian cancer and functioned as a tumor suppressor in ovarian cancer by retarding cell proliferation, migration, and EMT. Besides, HCG11 could bind to miR-1270 and PTEN was a target gene of miR-1270. Mechanically, HCG11 competitively bound with miR-1270 to upregulate PTEN. From rescue experiments, HCG11 impeded AKT/mTOR pathway to retard ovarian cancer cell growth by miR-1270/PTEN. CONCLUSIONS: HCG11 was a tumor suppressor in ovarian cancer cells and additionally, HCG11 regulated AKT/mTOR pathway to hinder ovarian cancer cell growth via modulating miR-1270/PTEN, indicating that HCG11 may represent a promising target for effective treatment of ovarian cancer patients.

Detection of HPV E6/E7 mRNA in the diagnosis of cervical cancer and precancerous lesions after kidney transplantation.

OBJECTIVE: To explore the value of human papillomavirus (HPV) E6/E7 mRNA detection in the diagnosis of cervical cancer and its precancerous lesions after kidney transplantation. METHODS: One hundred and sixty-six women who underwent cervical cancer screening after kidney transplantation were selected and received thinprep cytology test (TCT), HPV DNA and HPV E6/E7 mRNA tests. A biopsy under colposcopy was performed for those with suspicious test results. The positive rates of TCT, HPV DNA and HPV E6/E7 mRNA expressions in patients with different biopsy pathological grades, the positive rates of HPV E6/E7 mRNA in TCT and HPV DNA positive patients were compared. Besides, the relationship between the results of the three detections and the pathological results of cervical biopsy as well as the diagnostic efficacy of cervical cancer and its precancerous lesions were compared. RESULTS: Among the 166 women undergoing cervical cancer screening, 87 cases received histopathological biopsy, of which, the positive expression rates of HPV E6/E7 mRNA in the negative, cervical intraepithelial neoplasia (CIN) I, CIN II, CIN III and invasive carcinoma (ICC) patients were 51.43%, 54.55%, 66.67%, 81.82% and 100.00%, respectively. The positive expression rates of HPV E6/E7 mRNA in TCT and HPV DNA-positive patients were 47.50% and 51.96%, respectively; those rates for diagnosis of ≥CIN II were significantly greater than that of ≤CIN I (both P<0.01). Receiver operating characteristic curve revealed that the areas under the concentration-time curve of TCT, HPV DNA and HPV E6/E7 mRNA detection for cervical cancer and precancerous lesions were 0.723, 0.833, 0.929, respectively. Their sensibilities were 76.89%, 83.30% and 92.38%, and their specificities were 77.04%, 88.47% and 94.47%, respectively. CONCLUSION: HPV E6/E7 mRNA detection effectively improves the diagnostic sensitivity and specificity of cervical cancer and precancerous lesions, thereby avoiding over-examination and over-treatment.

IFI16 promotes human embryonic stem cell trilineage specification through interaction with p53.

Transcriptional regulation plays an essential role in the self-renewal and differentiation of human embryonic stem cells (hESCs). However, how external signals disrupt the self-renewal regulatory network and further drive hESC differentiation remains largely unknown. Here, we found the immune regulative protein, gamma-interferon-inducible protein 16 (IFI16) was involved in the regulation of both self-renewal and differentiation gene expression during hESC trilineage specification through interaction with p53. IFI16 expression levels were upregulated through JNK activation. IFI16 knockdown delayed the downregulation of self-renewal gene expression and suppressed the upregulation of differentiation gene expression, while IFI16 overexpression accelerated trilineage specification. Furthermore, IFI16 stabilized p53-binding in the genome through IFI16-p53 interaction and differentially regulated self-renewal and differentiation gene expression. Together, our results suggest a particular role of IFI16 in differential gene expression regulation during trilineage specification of hESCs in a manner that is dependent on the genome-wide profile of p53-binding directed by IFI16-p53 interaction.

Multi-Omics Integration Reveals a Competitive Endogenous RNAs Network for the Identification of Progression Biomarkers and the Stratification of Patients Diagnosed With Nephroblastoma.

Specific types of nephroblastoma (Wilms' tumor, WT) are known to associate with poor overall survival. Emerging experimental evidence has demonstrated that competitive endogenous RNA (ceRNA) networks have important roles in regulating cancer occurrence, but the roles of ceRNAs in regulating the WT progression and the patient outcomes remain unclear. Using the multi-omics data of 132 WT patients collected from TARGET database, an integration analysis pipeline was performed to construct a highly reliable ceRNA network. As results, a total of 147 nodes (116 mRNAs, 15 miRNAs, and 16 lncRNAs) were identified and used to explore the underlying mechanism for WT progression. WGCNA analysis further identified several prognostic molecules, including hsa-mir-93, LINC00087 and RP5-1086K13, that significantly associated with the overall survival rate. And, enrichment analysis verified the participation of these molecules in tumor-related pathways, such as those controlling autophagy and cadherin-mediated adhesion. Importantly, the WT patients were classified into three categories according to the ceRNA network, which significantly correlated with the overall survival. In conclusion, the ceRNA network could be a promising tool to further validate the prognostic biomarkers and categories of patients diagnosed with WT.

Expression levels of a gene signature in hiPSC associated with lung adenocarcinoma stem cells and its capability in eliciting specific antitumor immune-response in a humanized mice model.

BACKGROUND: Previous studies have reported that cancer stem cells (CSCs) play a key role in tumorigenesis, metastasis, and recurrence. CSC-based vaccination confers better protection in tumor cells. However, isolation and cultivation of CSCs are difficult. This study aimed to explore the similarities between CSCs and induced pluripotent stem cells (iPSCs). METHODS: ALDH1+ cancer stem cells were isolated from lung adenocarcinoma patients and their gene expression patterns compared with human induced pluripotent stem cells (hiPSCs). In addition, a tumor vaccine was developed using hiPSC and unmethylated cytosine-guanine (CpG). Finally, the antitumor properties of the vaccine were evaluated in a humanized mouse model. RESULTS: Preimmunization of iPSC+CpG elicited stronger antigen presentation and cytotoxic T cell response which suppressed the growth of tumors. Adoptive transfer of spleen T cells from the vaccine preimmunized mice inhibited tumor growth in unvaccinated recipients without any side effects. CONCLUSIONS: This study suggests a universal strategy for tumor therapy which simplifies future clinical procedures. Therefore, the application of hiPSC elicits tumor protective responses.

MiR-182 inhibits kidney fibrosis by regulating transforming growth factor ß1/Smad3 pathway in autosomal dominant polycystic kidney disease.

The aim of the present study was to investigate the molecular mechanism of miR-182 in kidney fibrosis in polycystic kidney disease (PKD). We measured the expression of miR-182 in kidney tissue of autosomal dominant PKD. Additionally, we investigated the relationship between miR-182 and fibrotic protein by transfecting miR-182 mimics and miR-182 inhibitor into polycystic kidney cyst-lined epithelial cells, respectively. Furthermore, we observed the interaction between transforming growth factor ß1 (TGF-ß1) and miR-182 and fibrinogen factors of cyst-lined epithelial cells after TGF-ß1 intervention, and measured the expression of Smad2 and Smad3 protein. Results are presented as follows: (a) MiR-182 was positively correlated with fibrosis of cyst-lined epithelial cells; (b) TGF-ß1 could induce fibrosis of cyst-lined epithelial cells; (c) the expression of miR-182 had a remarkably impact on the fibrosis induced by TGF-ß1, but had little effect on the expression of TGF-ß1; (d) the expression of Smad3 protein in TGF-ß1 induce-cyst-lined epithelial cells was increased. TGF-ß1 and miR-182 promoting the fibrosis of polycystic kidney cyst-lined epithelial cells may be mediated by the TGF-ß1/Smad3 signaling pathway, of which Smad3 was an important regulator.

The Effects of Chloroquine and Hydroxychloroquine on ACE2-Related Coronavirus Pathology and the Cardiovascular System: An Evidence-Based Review.

The ongoing pandemic of coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a serious threat to global public health and there is currently no effective antiviral therapy. It has been suggested that chloroquine (CQ) and hydroxychloroquine (HCQ), which were primarily employed as prophylaxis and treatment for malaria, could be used to treat COVID-19. CQ and HCQ may be potential inhibitors of SARS-CoV-2 entry into host cells, which are mediated via the angiotensin-converting enzyme 2 (ACE2), and may also inhibit subsequent intracellular processes which lead to COVID-19, including damage to the cardiovascular (CV) system. However, paradoxically, CQ and HCQ have also been reported to cause damage to the CV system. In this review, we provide a critical examination of the published evidence. CQ and HCQ could potentially be useful drugs in the treatment of COVID-19 and other ACE2 involved virus infections, but the antiviral effects of CQ and HCQ need to be tested in more well-designed clinical randomized studies and their actions on the CV system need to be further elucidated. However, even if it were to turn out that CQ and HCQ are not useful drugs in practice, further studies of their mechanism of action could be helpful in improving our understanding of COVID-19 pathology.

Histone Deacetylase Inhibitors Reduce Cysts by Activating Autophagy in Polycystic Kidney Disease.

BACKGROUND: Histone deacetylase inhibitors (HDACi) have therapeutic effects on various models of renal diseases including autosomal dominant polycystic kidney disease (ADPKD), but the molecular mechanism is unclear. OBJECTIVES: Here, we studied the role of trichostatin A (TSA), a specific HDACi, in regulating cyst growth to test the possibility that HDACi might help manage ADPKD by enhancing autophagy. RESULTS: Autophagy protein expression was higher in cultured Pkd1 knockout (Pkd1-/-) cells, an in vitro model of cystogenesis, compared with control cells. TSA prevented cyst formation in Pkd1-/- cells. We further tested whether TSA could not reduce the size of an already established cyst after inhibition of autophagy by chloroquine in Pkd1-/- cells. In vivo, treatment with TSA significantly slowed cyst growth in Pkd1-/- mice. Moreover, TSA treatment stimulated AMPK and inactivated mTOR during cyst growth in Pkd1-/- cells and kidneys in mice. CONCLUSIONS: Our results suggest that HDACi may prevent cyst formation by activation of the AMPK pathway and autophagy. They also imply that HDACi could have therapeutic potential for ADPKD treatment.

PYNOD reduces microglial inflammation and consequent neurotoxicity upon lipopolysaccharides stimulation.

PYNOD, a nod-like receptors (NLR)-like protein, was indicated to inhibit NF-κB activation, caspase-1-mediated interleukin (IL)-1ß release and cell apoptosis in a dose-dependent manner. Exogenous addition of recombinant PYNOD to mixed glial cultures may suppress caspase-1 activation and IL-1ß secretion induced by Aß. However, to the best of our knowledge, there no study has focused on the immunoregulatory effects of PYNOD specifically in microglia. The present study aimed to explore the roles of PYNOD involved in the lipopolysaccharides (LPS)-induced microglial inflammation and consequent neurotoxicity. Murine microglial BV-2 cells were transfected with pEGFP-C2-PYNOD (0-5.0 µg/ml) for 24 h and incubated with or without LPS (1 µg/ml) for a further 24 h. Cell viability was determined using MTT assay and the secretion of nitric oxide (NO), IL-1ß and caspase-1 was measured using the Griess method or ELISA. Protein expression levels of NF-κB p65 and inducible nitric oxide synthase (iNOS) were detected by immunofluorescent staining and/or western blot analysis. Co-culture of BV-2 cells with human neuroblastoma cell line SK-N-SH was performed in Transwell plates and the cell viability and apoptosis (using flow cytometry) of SK-N-SH cells were determined. Results indicated that PYNOD overexpression inhibited NO secretion and iNOS protein expression induced by LPS in BV-2 cells, with no detectable cytotoxicity. PYNOD overexpression also reduced the secretion of IL-1ß and caspase-1 from BV-2 cells upon LPS stimulation. These effects were dose-dependent. Additionally, PYNOD overexpression prevented LPS-induced nuclear translocation of NF-κB p65 in BV-2 cells. The growth-inhibitory and apoptosis-promoting effects of BV-2 cells towards SK-N-SH cells were alleviated as a result of PYNOD overexpression. In conclusion, PYNOD may mitigate microglial inflammation and consequent neurotoxicity.

Spatial-Resolution Cell Type Proteome Profiling of Cancer Tissue by Fully Integrated Proteomics Technology.

Increasing attention has been focused on cell type proteome profiling for understanding the heterogeneous multicellular microenvironment in tissue samples. However, current cell type proteome profiling methods need large amounts of starting materials which preclude their application to clinical tumor specimens with limited access. Here, by seamlessly combining laser capture microdissection and integrated proteomics sample preparation technology SISPROT, specific cell types in tumor samples could be precisely dissected with single cell resolution and processed for high-sensitivity proteome profiling. Sample loss and contamination due to the multiple transfer steps are significantly reduced by the full integration and noncontact design. H&E staining dyes which are necessary for cell type investigation could be selectively removed by the unique two-stage design of the spintip device. This easy-to-use proteome profiling technology achieved high sensitivity with the identification of more than 500 proteins from only 0.1 mm2 and 10 µm thickness colon cancer tissue section. The first cell type proteome profiling of four cell types from one colon tumor and surrounding normal tissue, including cancer cells, enterocytes, lymphocytes, and smooth muscle cells, was obtained. 5271, 4691, 4876, and 2140 protein groups were identified, respectively, from tissue section of only 5 mm2 and 10 µm thickness. Furthermore, spatially resolved proteome distribution profiles of enterocytes, lymphocytes, and smooth muscle cells on the same tissue slices and across four consecutive sections with micrometer distance were successfully achieved. This fully integrated proteomics technology, termed LCM-SISPROT, is therefore promising for spatial-resolution cell type proteome profiling of tumor microenvironment with a minute amount of clinical starting materials.

α2A-adrenergic blockade attenuates septic cardiomyopathy by increasing cardiac norepinephrine concentration and inhibiting cardiac endothelial activation.

Cardiomyopathy is a common complication associated with increased mortality in sepsis, but lacks specific therapy. Here, using genetic and pharmacological approaches, we explored the therapeutic effect of α2A-adrenergic receptor (AR) blockade on septic cardiomyopathy. CLP-induced septic rats were treated with BRL44408 (α2A-AR antagonist), prazosin (α1-AR antagonist) and/or reserpine. CLP-induced cardiomyopathy, indicated by reduced dP/dt and increased cardiac troponin I phosphorylation, was attenuated by BRL44408, this was associated with reduced cardiac TNF-α and endothelial VCAM-1 expression, cardiomyocyte apoptosis and related signal molecule phosphorylation. BRL44408 increased cardiac norepinephrine (NE) concentration in CLP rats. Pretreatment with reserpine that exhausts cardiac NE without affecting the circulating NE concentration or with prazosin partially abolished the cardioprotection of BRL44408 and reversed its inhibitory effects on myocardial TNF-α, apoptosis and related signal molecule phosphorylation, but not on VCAM-1 expression in septic rats. These effects of BRL44408 were confirmed by α2A-AR gene deletion in septic mice. Furthermore, α2-AR agonist not only enhanced LPS-induced TNF-α and VCAM-1 expression in cardiac endothelial cells that express α2A-AR, but also enhanced LPS-induced cardiac dysfunction in isolated rat hearts. Our data indicate that α2A-AR blockade attenuates septic cardiomyopathy by promoting cardiac NE release that activates myocardial α1-AR and suppressing cardiac endothelial activation.

TRAF3 delays cyst formation induced by NF-κB signaling.

This study aims to investigate the effects of TNF receptors associated factor 3 (TRAF3) on the signaling pathway and expression of downstream products of nuclear factor kappa B (NF-κB) in the epithelial cells of renal ducts in individuals with polycystic kidney disease (PKD). We observe the TRAF3 genic overexpression of the epithelial cells, which form a tubular branch structure, in polycystic kidneys and to explore the protective effect of TRAF3 on the cystogenesis and progression of PKD. Western blotting analysis was conducted to examine the signaling changes of NF-κB in PKD the epithelial cells and TRAF3 transgenic PKD epithelial cells. Changes in the downstream apoptosis factor and cell proliferation in PKD epithelial cells and TRAF3 transgenic PKD epithelial cells were detected. A three-dimensional matrigel culture experiment was performed to examine abnormal tubulomorphogenesis in vitro. The overexpression of TRAF3 significantly inhibited the signaling pathway of NF-κB in the PKD epithelial cells, downregulated the expression of downstream factors Bcl-2 and Bcl-xl, and significantly decreased cystic epithelial cell proliferation. Additional branch structures were observed in the PKD epithelial cells with a three-dimensional culture compared to wildtype cells. TRAF3 may likely induce apoptosis and resistance to proliferation and may be a new target to inhibit the cyst formation in PKD by regulating the NF-κB signaling pathway Bcl-2 and Bcl-xl activity. © 2017 IUBMB Life, 69(3):170-178, 2017.

Ginsenosides from stems and leaves of ginseng prevent ethanol-induced lipid accumulation in human L02 hepatocytes.

OBJECTIVE: To investigate the effect of ginsenosides from stems and leaves of ginseng on ethanol-induced lipid deposition in human L02 hepatocytes. METHODS: L02 cells were exposed to ethanol for 36 h and treated with or without ginsenosides. The viability of L02 cells was evaluated by methylthiazolyldiphenyl-tetrazolium bromide assay and the triglyceride (TG) content was detected. Lipid droplets were determined by oil red O staining. Intracellular reactive oxygen species (ROS) production and the mitochondrial membrane potential were tested by flow cytometry. The ATP level was measured by reverse phase high performance liquid chromatography. The expression of cytochrome p450 2E1 (CYP2E1) and peroxisome proliferator-activated receptor α (PPARα) was detected by reverse transcriptase-polymerase chain reaction and Western blotting, respectively. RESULTS: Ethanol exposure resulted in the increase of TG level, lipid accumulation and ROS generation, and the decrease of mitochondrial membrane potential and ATP production in the cells. However, ginsenosides significantly reduced TG content (9.69±0.22 µg/mg protein vs. 4.93±0.49 µg/mg protein, P<0.01), and ROS formation (7254.8±385.7 vs. 5825.2±375.9, P<0.01). Meanwhile, improvements in mitochondrial membrane potential (10655.33±331.34 vs. 11129.52±262.35, P<0.05) and ATP level (1.20±0.18 nmol/mg protein vs. 2.53±0.25 nmol/mg protein, P<0.01) were observed by treatment with ginsenosides. Furthermore, ginsenosides could down-regulate CYP2E1 expression (P<0.01) and upregulate PPARα expression (P<0.01) in ethanol-treated cells. CONCLUSIONS: Ginsenosides could prevent ethanol-induced hepatocyte steatosis in vitro related to the inhibition of oxidative stress and the improvement of mitochondrial function. In addition, the modulation of CYP2E1 and PPARα expression may also play an important role in the protective effect of ginsenosides against lipid accumulation.

Risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery / 中国内镜杂志

Objective To investigate the risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery. Methods 80 patients with cervical cancer were recruited in the study, who underwent radical surgery with laparoscopic surgery from March 2013 to December 2016. The patients were followed up after surgery, and the overall survival and disease free survival was analyzed with tumor recurrence and death as the terminal events. And the risk factors associated with clinical prognosis were identified by using Cox regression analysis. Results The patients were followed up from 12 months to 46 months, and the median period was 39 months. There were 16 recurrences and 6 deaths during the period of follow-up, yielding a disease-free survival of (41.85 ± 1.06) year and an overall survival of (44.86 ± 0.74) year. Cox regression analysis demonstrated that tumor size, clinical stage, lymph node metastasis and vascular invasion were independent risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery. Conclusion Tumor size, clinical stage, lymph node metastasis and vascular invasion were independent risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery.

Risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery / 中国内镜杂志

Objective To investigate the risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery. Methods 80 patients with cervical cancer were recruited in the study, who underwent radical surgery with laparoscopic surgery from March 2013 to December 2016. The patients were followed up after surgery, and the overall survival and disease free survival was analyzed with tumor recurrence and death as the terminal events. And the risk factors associated with clinical prognosis were identified by using Cox regression analysis. Results The patients were followed up from 12 months to 46 months, and the median period was 39 months. There were 16 recurrences and 6 deaths during the period of follow-up, yielding a disease-free survival of (41.85 ± 1.06) year and an overall survival of (44.86 ± 0.74) year. Cox regression analysis demonstrated that tumor size, clinical stage, lymph node metastasis and vascular invasion were independent risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery. Conclusion Tumor size, clinical stage, lymph node metastasis and vascular invasion were independent risk factors associated with clinical prognosis of cervical cancer treated with laparoscopic surgery.

p38 MAPK signal pathway involved in anti-inflammatory effect of Chaihu-Shugan-San and Shen-ling-bai-zhu-San on hepatocyte in non-alcoholic steatohepatitis rats.

BACKGROUND: Traditional Chinese Medicine (TCM), has over thousands-of-years history of use. Chaihu-Shugan-San (CSS), and Shen-ling-bai-zhu-San (SLBZS), are famous traditional Chinese herbal medicine formulas, which have been used in China, for the treatment of many chronic diseases. MATERIALS AND METHODS: This study investigated the anti-inflammatory effects of CSS and SLBZS on signaling molecules involved in p38 mitogen-activated protein kinase (p38 MAPK), pathway on hepatocytes of non-alcoholic steatohepatitis (NASH), rats induced by high fat diet. SD male rats were randomly divided into 8 groups: negative control group, model control group, high (9.6g/kg/day)/low (3.2g/kg/day)-dose CSS group, high (30g/kg/day)/low (10g/kg/day)-dose SLBZS group, high (39.6g/kg/day)/low (13.2g/kg/day)-dose integrated group. The rats of NASH model were induced by feeding a high-fat diet. After 16, wks, Hepatocytes were isolated from 6, rats in each group by collagenase perfusion. The liver histopathological changes and serum inflammatory cytokines TNF-α, IL-6 were determined. The proteins of TLR4, phosphor-p38 MAPK and p38 MAPK involved in p38 MAPK signal pathway were assayed. RESULTS: The statistical data indicated the NASH model rats reproduced typical histopathological features of NASH in human. CSS and SLBZS ameliorated lipid metabolic disturbance, attenuated NASH progression, decreased the levels of TNF-α and IL-6 in serum, as well as inhibited TLR4 protein expression, p38 MAPK phosphorylation, and activation of p38 MAPK. In conclusion, CSS and SLBZS might work as a significant anti-inflammatory effect on hepatocyte of NASH by inhibiting the activation of TLR4, p-p38 MAPK and p38 MAPK involved in p38 MAPK signal pathway. CONCLUSION: To some extent, CSS and SLBZS may be a potential alternative and complementary medicine to protect against liver injury, alleviate the inflammation reaction, moderate NASH progression.

Berberine inhibits norepinephrine-induced apoptosis in neonatal rat cardiomyocytes via inhibiting ROS-TNF-α-caspase signaling pathway.

OBJECTIVE: To determine the effect of berberine (Ber) on norepinephrine (NE)-induced apoptosis in neonatal rat cardiomyocytes. METHODS: The cultured neonatal rat cardiomyocytes were treated with NE in the presence or absence of Ber. The activity of lactate dehydrogenase (LDH) in the culture medium was examined, and apoptosis of cardiomyocytes was assessed by Hoechst 33258, isothiocyanate (FITC)-conjugated annexin-V, and propidine iodide (PI) staining. In addition, the activities of caspases-2 and-3 were measured by a fluorescent assay kit. The level of secreted tumor necrosis factor α (TNF-α) and production of intracellular reactive oxygen species (ROS) were also determined. RESULTS: NE at a concentration of 50 µ mol/L induced an obvious increase in the activity of LDH in the culture medium (P<0.05), which was inhibited by coincubation with 0.5, 1.0, or 2.0 µ mol/L Ber (P<0.05). Ber also significantly attenuated NE-induced apoptosis in a dose-dependent manner (P<0.01). Moreover, Ber at a dose of 2 µ mol/L markedly decreased the ROS and TNF-α productions (P <0.05) and inhibited the activation of caspases-2 and -3 in cardiomyocytes exposed to NE (P<0.05)h. CONCLUSION: The present study suggested that Ber could reduce NE-induced apoptosis in neonatal rat cardiomyocytes through inhibiting the ROS-TNF-α-caspase signaling pathway.