Comparative analysis of surgical modalities for the gynecomastia treatment: efficiency and aesthetic outcomes.

This retrospective study rigorously compares the clinical efficacy of three surgical methodologies for treating gynecomastia while providing guidance for future surgical modality selection. We analyzed records of 77 gynecomastia patients treated between January 2015 and October 2022. Patients were categorized into three groups: Group A (subcutaneous gland resection via areola incision), Group B (liposuction combined with single-hole endoscopic gland resection), and Group C (liposuction combined with three-hole endoscopic gland resection). Parameters assessed included patient demographics, intraoperative bleeding, surgical duration, hospitalization duration, costs, postoperative drainage, complications, and patient satisfaction. Group A had significantly shorter operation time and lower cost than Groups B and C (P < 0.05). There were no significant differences in postoperative drainage (P > 0.05). Group A had a higher incidence of subcutaneous fluid complications. All groups achieved 100% overall postoperative efficiency. Group B demonstrated superior outcomes for scarring and patient satisfaction. All three surgical modalities effectively treat gynecomastia. Circumareolar incision subcutaneous gland resection is optimal for mild to moderate cases due to reduced operation time and cost. Liposuction with single-hole endoscopic gland resection and three-hole endoscopic gland resection offers fewer complications and discreet incisions. Notably, the liposuction and single-hole endoscopic approach yielded superior postoperative patient satisfaction, aligning with minimally invasive principles and warranting broad clinical application.

LncRNA SEMA3B-AS1 suppresses the tumor-initiating characteristics of triple negative breast cancer via engaging in MLL4-mediated H3K4 trimethylation.

Long noncoding RNAs (lncRNAs) are crucial regulators of tumor-initiating cells (TICs) and hold particular importance in triple negative breast cancer (TNBC). Yet, the precise mechanisms by which TIC-associated lncRNAs influence TNBC remain unclear. Our research utilized The Cancer Genome Atlas Breast Cancer (BC) data set to identify prognostic lncRNAs. We then conducted extensive assays to explore their impact on the tumor-initiating phenotype of TNBC cells and the underlying mechanisms. Notably, we found that low expression of lncRNA SEMA3B-AS1 correlated with unfavorable survival in BC patients. SEMA3B-AS1 was also downregulated in TNBC and linked to advanced tumor stage. Functional experiments confirmed its role as a TIC-suppressing lncRNA, curtailing mammosphere formation, ALDH + TIC cell proportion, and impairing clonogenicity, migration, and invasion. Mechanistic insights unveiled SEMA3B-AS1's nuclear localization and interaction with MLL4 (mixed-lineage leukemia 4), triggering H3K4 methylation-associated transcript activation and thus elevating the expression of SEMA3B, a recognized tumor suppressor gene. Our findings emphasize SEMA3B-AS1's significance as a TNBC-suppressing lncRNA that modulates TIC behavior. This study advances our comprehension of lncRNA's role in TNBC progression, advocating for their potential as therapeutic targets in this aggressive BC subtype.

High blood galectin-3 level associated with risk of frailty in aging.

Background: Frailty is one of the most problematic expressions of population aging, but its underlying mechanism has not been fully elucidated. Circulating galectin-3 (Gal-3) is involved in the pathogenesis of many age-related diseases. This study aims to explore the influence of circulating Gal-3 on the regulation of frailty and aging and to identify the potential mechanism further. Methods: In this cross-sectional analysis, the Fried frailty phenotype (FP) was assessed among 149 community elderly residents in Shanghai. Peripheral blood mononuclear cells (PBMCs) were isolated by the Ficoll-Paque density gradient method, and differentially expressed genes (DEGs) encoding transcription factors in frailty were detected by Illumina and bioinformatics analyzed with R software. Gene Ontology (GO) enrichment analyses and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to explore the functional roles of these DEGs and the target genes related to frailty phenotypes. The serum Gal-3 concentration was tested by enzyme-linked immunosorbent assay (ELISA). Mouse frailty phenotype was used to construct an in vivo model of frailty, after which the serum levels of circulating Gal-3 and its gene expression levels in mouse tissues were determined. Results: Participants' mean age was 72.04 ± 7.05 years. In total, 21.48% were frail and 36.91% were pre-frail. The mean serum Gal-3 concentration was 46.34 ± 17.99 ng/mL in frail participants, 32.30 ± 8.14 ng/mL in pre-frail participants, and 26.00 ± 5.87 ng/mL in non-frail individuals (p < 0.001). Significant positive correlations between serum Gal-3 level and FP score, SARC-F score, C-reactive protein (CRP), interleukin-6, etc., were observed. In addition, the KEGG pathway and GO enrichment analyses showed that 265 DEGs in PBMCs of frail participants were mainly related to inflammatory response, translation, RNA binding, protein binding, ribosome, and primary immunodeficiency. LGALS3 was identified as the overlapping gene between frailty-related DEGs and aging-related DEGs. The elevated serum Gal-3 concentration in the in vivo model of frailty was consistent with the results in participants. Conclusion: In both community-dwelling older adults and aged mice, serum Gal-3 concentration was positively correlated with frailty. This circulating mediator may be a promising indicator of frailty. Clinical trial registration: Chinese Clinical Trial Registry identifier, ChiCTR2000036399.

Combined therapy with pioglitazone and FGF21 mRNA synergistically ameliorates metabolic disorders in NAFLD rats.

Background: Current study aims to investigate the ameliorative effect of pioglitazone (PIO) combined with mRNA encoding FGF21 (termed mFGF21) on the metabolic disorders in rats with nonalcoholic fatty liver disease (NAFLD) and its potential mechanism. Methods: In vitro functional activity of FGF21 protein expressed by mFGF21 was evaluated in human adipose-derived stem cells (hASCs). The pharmacokinetic profiles of FGF21 protein expressed by mFGF21 were investigated in normal SD rats and NAFLD rats, respectively. Results: As the results, it showed that the PIO could enhanced in vitro functional activity of FGF21 protein expressed from mFGF21 in hASCs. Not only that, mFGF21 turns the body into a processing plant for endogenous protein expression, which enhanced the pharmacokinetic profiles of FGF21 proteins. Combined treatment with PIO and mFGF21 significantly reduced body weight, fasting blood glucose levels, insulin levels and lipid metabolism in NAFLD rats compared with control or both two monotherapy groups. The results of H&E staining and Western blot revealed that combined treatment with PIO and mFGF21 significantly decreased hepatic fat accumulation in NAFLD rats by activating the SHP1/AMPK signaling pathway. Conclusions: Our finding collectively demonstrated that PIO and mFGF21 combination therapy could synergistically ameliorate metabolic disorders in NAFLD rats.

STK24 Promotes Progression of LUAD and Modulates the Immune Microenvironment.

Objective: Recent studies have shown that serine/threonine-protein kinase 24 (STK24) plays an important role in cancer development. However, the significance of STK24 in lung adenocarcinoma (LUAD) remains to be determined. This study is aimed at investigating the significance of STK24 in LUAD. Methods: STK24 was silenced and overexpressed by siRNAs and lentivirus, respectively. Cellular function was assessed by CCK8, colony formation, transwell, apoptosis, and cell cycle. mRNA and protein abundance was checked by qRT-PCR and WB assay, respectively. Luciferase reporter activity was evaluated to examine the regulation of KLF5 on STK24. Various public databases and tools were applied to investigate the immune function and clinical significance of STK24 in LUAD. Results: We found that STK24 was overexpressed in lung adenocarcinoma (LUAD) tissues. High expression of STK24 predicted poor survival of LUAD patients. In vitro, STK24 enhanced the proliferation and colony growth ability of A549 and H1299 cells. STK24 knockdown induced apoptosis and cell cycle arrest at G0/G1 phase. Furthermore, Krüppel-like factor 5 (KLF5) activated STK24 in lung cancer cells and tissues. Enhanced lung cancer cell growth and migration triggered by KLF5 could be reversed by silencing of STK24. Finally, the bioinformatics results showed that STK24 may be involved in the regulation of the immunoregulatory process of LUAD. Conclusion: KLF5 upregulation of STK24 contributes to cell proliferation and migration in LUAD. Moreover, STK24 may participate in the immunomodulatory process of LUAD. Targeting KLF5/STK24 axis may be a potential therapeutic strategy for LUAD.

Hepatic-Targeted Nano-enzyme with Resveratrol Loading for Precise Relief of Nonalcoholic Steatohepatitis.

Nonalcoholic steatohepatitis (NASH) is characterized by massive lipid deposition in hepatocytes and is often associated with hepatic inflammation and other severe metabolic syndromes. The intervention of NASH can prevent its further progression into hepatocarcinoma. In this study we have successfully constructed liver-targeted Ce-based hollow mesoporous nanocarriers loaded with bioactive drugs. This may provide an effective approach for eliminating NASH. Liver-section-specific targeting was realized by covalently linked galactose (Gal), which can be specifically recognized by receptors in the membranes of hepatocytes. Meanwhile, resveratrol (Res), a drug used to treat NASH, was efficiently loaded into the pores and cavity of CeO2 (Res@H-CeO2 -Gal). In steatotic HepG2 cells (free fatty acid induction), this nanosystem was found to enhance cellular Res internalization for improved anti-lipogenesis activity. In mice with NASH, Res@H-CeO2 -Gal increased Res delivery to liver sections for a reduction in lipid accumulation and enhanced anti-inflammatory activity from the antioxidant capacity of Ce-based nanocarriers. This effectively recovered NASH mice to the normal state. These findings show that the hepatic targeting and Res delivery nanoplatform could act as a safe and promising strategy for the elimination of NASH and other liver diseases.

Effects of nutritional supplement and resistance training for sarcopenia in patients with inflammatory bowel disease: A randomized controlled trial.

BACKGROUND: Nutritional supplementation and resistance training are broadly recommended as part of the treatment of sarcopenia, but studies that have evaluated interventions in inflammatory bowel disease patients with sarcopenia are lacking. The aim of this study was to evaluate the effects of nutritional supplementation and resistance training for improving height-adjusted appendicular skeletal muscle mass (ASM/H2) and medical indices in patients with inflammatory bowel disease. METHODS: This randomized, double-blind, placebo-controlled trial of forty-five participants was performed at Huadong Hospital Affiliated to Fudan University in Shanghai from September 2020 to June 2021. Eligible participants were randomly assigned to receive whey protein (10 g/d) or placebo (10 g/d) for 8 weeks while completing a resistance training program (3 times a week). Data such as ASM/H2 and other medical indices were collected at baseline and at 4 and 8 weeks of intervention. RESULTS: Fifteen participants were allocated to the resistance training and whey protein (RT+WP) group, and thirteen participants were allocated to the resistance training and placebo (RT+placebo) group. The ASM/H2 significantly increased in the RT+WP group after 4 and 8 weeks of intervention, and the ASM/H2 of the RT+WP group was significantly higher than that of the RT+placebo group after 4 and 8 weeks of intervention (F = 1.092, P = .035). Both interventions significantly increased albumin (F = 7.214, P = .003). Hemoglobin and creatinine significantly increased in the RT+WP group (F = 3.592, P = .035; F = 3.922, P = .033, respectively). In addition, a significant group × time interaction was not observed for body mass index, 5-time chair stand test time, 3-metre walk speed, grip strength, waist circumference, hip circumference, or waist-to-hip ratio (P > .05). CONCLUSIONS: Nutritional supplementation may be effective in improving sarcopenia, as well as many other physiological indicators during resistance training.

Effects of Diets With Different Protein Levels on Lipid Metabolism and Gut Microbes in the Host of Different Genders.

The purpose of this experiment was to investigate the effects of different protein levels on lipid metabolism and gut microbes in mice of different genders. A total of 60 mice (30 female and 30 male) were randomly assigned to six groups and fed female mice with low protein diet (FLP), basal protein diet (FBD), and high protein diet (FHP). Similarly, the male mice fed with low protein diet (MLP), basal protein diet (MBD), and high protein diet (MHP). The low protein diet contained 14% CP, the basal diet contained 20% CP, and the high protein diet contained 26% CP. The results of the study showed that both basal and high protein diets significantly reduced the perirenal adipose tissues (PEAT) index in male mice compared to low protein diet (p < 0.05). For the gut, the FHP significantly increased the relative gut weight compared to the FBD and FLP (p < 0.05). At the same time, the FHP also significantly increased the relative gut length compared with the FBD and FLP (p < 0.05). The MHP significantly increased TC concentration compared with the MLP (p < 0.05), and the MBD tended to increase TC concentration compared with the MLP in serum (p = 0.084). The histomorphology result of the jejunum and ileum showed that a low protein diet was beneficial to the digestion and absorption of nutrients in the small intestine of mice. While different protein levels had no effect on the total number of fecal microbial species in mice, different protein levels had a significant effect on certain fecal microbes in mice, the absolute abundance of Verrucomicrobia in the feces of male mice was significantly higher in both high and basal protein diets than in the low protein diet (p < 0.05). The high protein diet significantly reduced the absolute abundance of Patescibacteria in the feces of female mice compared to both the basal and low protein diets (p < 0.05). The absolute abundance of Patescibacteria in male feces was not affected by dietary protein levels (p > 0.05). Taken together, our results suggest that a low protein diet can alter fat deposition and lipid metabolism in mice, and that it benefited small intestinal epithelial structure and microbes.

Caffeine intake and the risk of incident kidney stones: a systematic review and meta-analysis.

BACKGROUND: Kidney stone disease is increasingly common in the general population, with a high recurrence rate after stone removal. It has been proven that caffeine consumption can reduce the risk of diseases, such as stroke and dementia. However, the effect of caffeine intake on the incidence of kidney stones has not been determined. This systematic review and meta-analysis were performed to evaluate the association of caffeine intake with the risk of incident kidney stones. METHODS: PubMed, Web of Science, Scopus, Cochrane and Google Scholar were searched using terms related to coffee, caffeine and kidney stones to find eligible articles up to December 2021. Articles with clear diagnostic criteria for kidney stone disease and the exact intake dose of caffeine were included. The incidence of kidney stone disease was the main outcome. Summarized risk estimates and 95% CIs for the highest and lowest categories of caffeine intake were calculated using a random effects model. RESULTS: Seven studies were included in the final meta-analysis, with 9707 cases of kidney stones and a total of 772,290 cohort members. Compared with the lowest category of caffeine intake, the pooled relative risk (RR) was 0.68 ([95% CI 0.61-0.75], I2 = 57%) for the highest category of caffeine intake. Subgroup analyses showed that caffeine intake had an inverse relationship with the incidence of kidney stones in all subgroups. CONCLUSION: This study suggests that a higher caffeine intake may be associated with a lower risk of incident kidney stones.

Spread and Molecular Characteristics of Enterobacteriaceae Carrying fosA-Like Genes from Farms in China.

In this study, we aimed to investigate the occurrence and molecular characteristics of fosfomycin-resistant Enterobacteriaceae isolates from pig, chicken and pigeon farms in Guangxi Province of China. A total of 200 fosfomycin-resistant strains were obtained from food animals and their surrounding environments, with the fosA, fosA3, and fosA7.5 genes being detected in 26% (52/200), 10% (20/200), and 5% (10/200), respectively. Surprisingly, three fosA7.5-producing E. coli isolates were found to be concomitant with fosA3. Most of the fosA-like-gene-positive isolates were multidrug-resistant strains and consistently possessed blaCTX-M-1/CTX-M-9, floR, and blaTEM genes. Only fosA3 was successfully transferred to the recipient strains, and the 29 fosA3-carrying transconjugants exhibited high-level resistance to fosfomycin (MIC ≥ 512 µg/mL). Multilocus sequence typing (MLST) combined with enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) analyses indicated that fosA3 or fosA7.5 genes were spread by horizontal transfer as well as via clonal transmission between E. coli. We used the PCR mapping method to explore the genetic contexts of fosA-like genes, and two representative strains (fEc.1 and fEcg99-1) were fully sequenced. Six different genetic structures surrounding fosA3 were detected and one infrequent context was discovered among the conjugable fosA3-positive E. coli isolates. The five genetic environments of fosA were identified and found to be highly similar to the partial sequence of transposon Tn2921. Furthermore, whole-genome sequencing (WGS) results showed that fosA7.5 was colocalized with mcr-3, blaCMY-63, sul3, tet(A), dfrA, and a number of virulence-related factors on the same chromosomes of strains, and various insertion sequences (IS3/ISL3) were detected upstream or downstream of fosA7.5. The phylogenetic analysis revealed that both fosA7.5- and fosA3-carrying E. coli ST602 and fosA7.5-carrying E. coli ST2599 were closely related to E. coli isolates from humans, which may indicate that they pose a threat to human health. IMPORTANCE Here, we report the widespread and complex genetic environments of fosA-like genes in animal-derived strains in China. The fosA7.5 gene was identified in this study and was found to confer resistance to fosfomycin. The high prevalence of fosA-like genes in farms indicates that food animals serve as a potential reservoir for the resistance genes. This study also discovered that fosfomycin resistance genes were always associated with mobile elements, which would accelerate the transmission of fosA-like genes in strains. Importantly, E. coli ST602 and ST2599 carrying fosA3 or fosA7.5 from food animals had high similarity to E. coli isolates from humans, suggesting that fosA-like genes can be transmitted to humans through the food chain, thus posing a serious threat to public health. Therefore, the prevalence of fosA-like genes isolated from animals should be further monitored.

Preliminary Study on Risk Factors for Morbidity of Nonalcoholic Fatty Liver Disease in High-Income Male Population.

OBJECTIVES: Believed to be a result of metabolic syndrome and unhealthy lifestyle, the incidence of nonalcoholic fatty liver disease (NAFLD) has become a serious public health problem. Among the high-income male population, metabolic syndrome and unhealthy lifestyle are particularly prominent. Therefore, we conducted a survey on 375 high-income male subjects, expecting to understand the risk factors and related factors for morbidity of NAFLD among the high-income male population being physically examined in Shanghai. METHODS: A cross-sectional study was applied to 375 high-income male subjects (including 190 patients with NAFLD and 185 non-NAFLD subjects) who were examined in the special needs clinic at Huadong Hospital affiliated to Fudan University. In combination with medical history, physical examination, and laboratory test results and by use of a self-made NAFLD health questionnaire, the basic data of the research objects were collected and the obtained data were subject to a correlation analysis. RESULTS: This study investigated 375 high-income males, and the morbidity rate of NAFLD was 50.67%. The NAFLD group was higher than the non-NAFLD group in terms of body weight, BMI, systolic blood pressure, and diastolic blood pressure (P < 0.05). Hypertension (OR = 2.944), diabetes (OR = 7.278), and hyperuricemia (OR = 1.922) are the risk factors for NAFLD; compared with no metabolic diseases, one (OR = 1.848), two (OR = 2.417), and three metabolic diseases (OR = 14.788) are risk factors for the development of NAFLD. Compared with the non-NAFLD group, the NAFLD group had a higher level of WBC, RBC, Hb, PLT, FPG, HbA1c, ALT, AST, GGT, ALP, TP, and UA (P < 0.05). There was a statistically significant difference in the intake of supper and staple foods between the NAFLD group and the non-NAFLD group, and the highly greasy diet was a risk factor for NAFLD (OR = 2.173) as opposed to the nongreasy diet. CONCLUSION: High-income male population is a high-risk group of NAFLD. Most of the patients with NAFLD have abnormal biochemical indicators as opposed to the healthy population and are more likely to be complicated with other chronic diseases or abnormal health status. And the occurrence of hypertension, diabetes, and hyperuricemia is the risk factor for the development of NAFLD. At the same time, the number of metabolic diseases complicated is also a risk factor for NAFLD as compared with the absence of complications with such metabolic diseases. Compared with a diet that is not greasy, the fact that high-income male NAFLD patients have a very greasy diet increases the risk of NAFLD.

Long-term effects and benefits of Helicobacter pylori eradication on the gastric mucosa in older individuals.

BACKGROUND: The current international consensus report indicated that all Helicobacter pylori (H. pylori)-positive patients should be treated. This study aimed to evaluate the long-term effects and benefits of H. pylori eradication on the gastric mucosa in the elderly population. METHODS: We performed a retrospective cohort study with 311 individuals aged ≥60 years, including 83 with persistent H. pylori infection (persistent group), 128 with successful H. pylori eradication (eradicated group), and 100 without H. pylori infection (control group). The results of endoscopy and mucosal histology were investigated at baseline and followed up for 5 and 10 years. RESULTS: In the 5 to 10-year follow-up, there was a significant difference in the atrophy score among the three groups (P < 0.001); however, no significant difference was observed in the intestinal metaplasia (IM) score (P > 0.05). There was no significant difference in the cumulative incidence of gastric neoplastic lesion (GNL) between the eradicated and persistent groups during the 5 to 10-year follow-up period (P > 0.05). The baseline IM score of patients with GNL was significantly higher than that of those without GNL in the eradicated and control groups (P < 0.05). In all patients with GNL, the mean interval time between baseline and diagnosis of GLN was more than 6 years. The severity of baseline mucosal IM (odds ratio: OR 3.092, 95% confidence interval [CI]: 1.690-5.655, P < 0.001) and H. pylori infection (OR: 2.413, 95%CI: 1.019-5.712, P = 0.045) significantly increased the risk for GNL. CONCLUSIONS: Older patients with a life expectancy of less than 5 to 10 years, especially those with moderate to severe gastric mucosal IM, may not benefit from the eradication of H. pylori to prevent gastric cancer.

A Narrative Review of Gut-Muscle Axis and Sarcopenia: The Potential Role of Gut Microbiota.

Sarcopenia is a multifactorial disease related to aging, chronic inflammation, insufficient nutrition, and physical inactivity. Previous studies have suggested that there is a relationship between sarcopenia and gut microbiota,namely, the gut-muscle axis. The present review highlights that the gut microbiota can affect muscle mass and muscle function from inflammation and immunity,substance and energy metabolism, endocrine and insulin sensitivity, etc., directly or indirectly establishing a connection with sarcopenia, thereby realizing the "gut-muscle axis".

Impact of a low-carbohydrate and high-fiber diet on nonalcoholic fatty liver disease.

BACKGROUND AND OBJECTIVES: To study the effects of a low-carbohydrate and high-fiber diet and education on patients with nonalcoholic fatty liver disease. METHODS AND STUDY DESIGN: We randomly divided 44 patients with nonalcoholic fatty liver disease into two groups: low-carbohydrate and high-fiber diet and education (intervention group), and education alone (control group). Liver and kidney function, fasting plasma glucose, insulin resistance index, body composition, and controlled attenuation parameter were detected before and after the intervention. RESULTS: After 2 months, the body fat, body weight, abdominal circumference, and visceral fat area, fasting plasma glucose, insulin resistance index, and levels of serum alanine aminotransferase, aspartate transaminase, uric acid, and insulin of the intervention group were significantly lower than before (p<0.05). In the female intervention group, the insulin resistance index and levels of serum alanine aminotransferase, uric acid, triglyceride, fasting plasma glucose, and C-peptide were lower and the level of serum high-density lipoprotein cholesterol was higher than in the female control group (p<0.05). In the male intervention group, the levels of serum alanine aminotransferase, triglyceride, and fasting plasma glucose were lower and the level of serum high-density lipoprotein cholesterol was higher compared with the male control group (p<0.05). CONCLUSIONS: A low-carbohydrate and high-fiber diet and education can effectively reduce the body weight and body fat of patients with nonalcoholic fatty liver disease and improve metabolic indicators such as liver enzymes, blood glucose, blood lipid, and uric acid. Our female patients showed significantly better improvement in the indicators than our male patients.

Circadian Clock Genes in the Metabolism of Non-alcoholic Fatty Liver Disease.

Non-alcoholic fatty liver disease (NAFLD) is a common disease, which is characterized by the accumulation of triglycerides in the hepatocytes without excess alcohol intake. Circadian rhythms can participate in lipid, glucose, and cholesterol metabolism and are closely related to metabolism seen in this disease. Circadian clock genes can modulate liver lipid metabolism. Desynchrony of circadian rhythms and the influences imparted by external environmental stimuli can increase morbidity. By contrast, synchronizing circadian rhythms can help to alleviate the metabolic disturbance seen in NAFLD. In this review, we have discussed the current research connections that exist between the circadian clock and the metabolism of NAFLD, and we have specifically focused on the key circadian clock genes, Bmal1, Clock, Rev-Erbs, Rors, Pers, Crys, Nocturnin, and DECs.

The histone demethylase KDM4D promotes hepatic fibrogenesis by modulating Toll-like receptor 4 signaling pathway.

BACKGROUND: Accumulating evidence has revealed the pivotal role of epigenetic regulation in the pathogenesis of liver disease. However, the epigenetic mechanism that accounts for hepatic stellate cells (HSCs) activation in liver fibrosis remains largely unknown. METHODS: Primary HSCs were used to screen the differentially expressed histone H3 lysine methyltransferases and demethylases during HSC activation. Loss-of-function experiments were applied to determine the cellular functions of KDM4D in HSCs. Transcriptome analysis was applied to explore the downstream targets of KDM4D. Real-time qPCR, western blotting, immunohistochemical staining, and chromatin immunoprecipitation were performed to uncover the underlying mechanism concerning KDM4D during liver fibrogenesis. FINDINGS: KDM4D was identified as a remarkable up-regulated histone H3 demethylase during HSC activation. The overexpression profile of KDM4D was confirmed in three fibrosis animal models and human fibrotic liver tissues. In vitro Kdm4d knockdown impaired the collagen gel contraction and migration capacity of primary HSCs. In established CCl4-induced mice model, Kdm4d knockdown inhibited fibrosis progression, and promoted fibrosis reversal, with enhanced thinning and splitting of fibrotic septa, as well as a dramatic decrease in collagen area. Whole gene transcriptome analysis showed the regulatory role of KDM4D in Toll-Like Receptor (TLR) signaling pathway. Mechanistically, KDM4D catalyzed histone 3 on lysine 9 (H3K9) di-, and tri-demethylation, which promoted TLR4 expression, and subsequently prompted liver fibrogenesis by activating NF-κB signaling pathways. INTERPRETATION: KDM4D facilitates TLR4 transcription through demethylation of H3K9, thus activating TLR4/NF-κB signaling pathways in HSCs, contributing to HSC activation and collagen crosslinking, further, hepatic fibrosis progression. FUND: Shanghai New Hundred Talents Program, Shanghai Municipal Commission of Health and Family Planning, Key Developing Disciplines Program, Shanghai Key disciplines program of Health and Family Planning and Shanghai Sailing Program.

Huperzine A attenuates nonalcoholic fatty liver disease by regulating hepatocyte senescence and apoptosis: an in vitro study.

OBJECTIVE: This study was undertaken to detect if free fatty acids (FFA) induce hepatocyte senescence in L-02 cells and if huperzine A has an anti-aging effect in fatty liver cells. METHODS: L-02 cells were treated with a FFA mixture (oleate/palmitate, at 3:0, 2:1, 1:1, 1:2 and 0:3 ratios) at different concentrations. Cell viability and fat accumulation rate were assessed by a Cell Counting Kit 8 and Nile Red staining, respectively. The mixture with the highest cell viability and fat accumulation rate was selected to continue with the following experiment. The L-02 cells were divided into five groups, including the control group, FFA group, FFA + 0.1 µmol/L huperzine A (LH) group, FFA + 1.0 µmol/L huperzine A (MH) group and FFA + 10 µmol/L huperzine A (HH) group, and were cultured for 24 h. The expression of senescence-associated ß-galactosidase (SA-ß-gal) was detected by an SA-ß-gal staining kit. The expression levels of aging genes were measured by qRT-PCR. The expression levels of apoptosis proteins were detected by a Western blot. ELISA kits were used to detect inflammatory factors and oxidative stress products. The expression of nuclear factor (NF-κB) and IκBα were detected by immunofluorescence. RESULTS: The FFA mixture (oleate/palmitate, at a 2:1 ratio) of 0.5 mmol/L had the highest cell viability and fat accumulation rate, which was preferable for establishing an in vitro fatty liver model. The expression of inflammatory factors (TNF-α and IL-6) and oxidants Malonaldehyde (MDA), 4-hydroxynonenal (HNE) and reactive oxygen species (ROS) also increased in the L-02 fatty liver cells. The expression levels of aging markers and aging genes, such as SA-ß-gal, p16, p21, p53 and pRb, increased more in the L-02 fatty liver cells than in the L-02 cells. The total levels of the apoptosis-associated proteins Bcl2, Bax, Bax/Bcl-2, CyCt and cleaved caspase 9 were also upregulated in the L-02 fatty liver cells. All of the above genes and proteins were downregulated in the huperzine A and FFA co-treatment group. In the L-02 fatty liver cells, the expression of IκBα decreased, while the expression of NF-κB increased. After the huperzine A and FFA co-treatment, the expression of IκBα increased, while the expression of NF-κB decreased. CONCLUSION: Fatty liver cells showed an obvious senescence and apoptosis phenomenon. Huperzine A suppressed hepatocyte senescence, and it might exert its anti-aging effect via the NF-κB pathway.