ObjectiveTo investigate the efficacy and underlying mechanisms of Gynostemma pentaphyllum alcohol extract in improving glucose and lipid metabolism disorders in db/db mice through transcriptomics and gut microbiota analysis. MethodsEighteen db/db mice were randomly assigned to the model（DM） group， metformin（MET） group， and G. pentaphyllum alcohol extract（GP） group， with six mice in each group， based on stratification of fasting blood glucose and body weight. An additional six db/m mice were selected as the normal control（NC） group. Mice in the NC and DM groups were administered deionized water （10 mL·kg^-1） daily. The MET group received metformin （0.195 g·kg^-1） by gavage. The GP group was treated with G. pentaphyllum alcohol extract （3.9 g·kg^-1） by gavage for six weeks. Fasting blood glucose was measured every two weeks. After six weeks of intervention， serum levels of total cholesterol （TC）， triglycerides （TG）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， creatinine （CREA）， and blood urea nitrogen （BUN） were assessed. Enzyme-linked immunosorbent assay （ELISA） was used to measure insulin （FINS）， adiponectin （ADP）， and tumor necrosis factor-α （TNF-α）. Hematoxylin-eosin （HE） staining was used to observe liver histomorphology， periodic acid-Schiff （PAS） staining was employed to assess hepatic glycogen synthesis， and Oil Red O staining was used to detect hepatic lipid deposition. Liver transcriptomic data were used to identify differentially expressed genes in the liver and conduct enrichment analysis. Real-time PCR was employed to verify the expression levels of adiponectin gene （Adipoq）， peroxisome proliferator-activated receptor gamma coactivator-1α （PGC-1α）， AMP-activated protein kinase （AMPK）， peroxisome proliferator-activated receptor α （PPARα）， glucokinase （GCK）， forkhead box （Fox）O1， FoxO3， phosphoenolpyruvate carboxykinase （PEPCK）， and glucose-6-phosphatase （G6PC）. Metagenomic sequencing was conducted to analyze changes in gut microbiota composition. ResultsCompared with the NC group， the DM group exhibited significantly elevated fasting blood glucose （P<0.01）， serum AST， ALT， TC， TG， LDL-C， and HDL-C （P<0.01）. FINS， homeostatic model assessment for insulin resistance （HOMA-IR）， and the inflammatory cytokine TNF-α were significantly increased （P<0.01）， while ADP was significantly decreased （P<0.05）. Histological analysis confirmed severe hepatic steatosis and excessive lipid accumulation in the DM group， along with markedly reduced glycogen synthesis. Compared with the DM group， the GP group showed significantly decreased fasting blood glucose （P<0.01）， reduced serum TC， LDL-C， and HDL-C levels （P<0.05）， significantly decreased serum TG and AST levels （P<0.01）， significantly reduced FINS， HOMA-IR， and TNF-α levels （P<0.01）， and significantly increased ADP （P<0.01）. Hepatic steatosis and lipid deposition were significantly alleviated， while glycogen synthesis was markedly enhanced. Transcriptomic differential and enrichment analyses suggested that the mechanisms by which G. pentaphyllum alcohol extract improved hepatic glucose and lipid metabolism in db/db mice may involve regulation of the AMPK and FoxO signaling pathways. Real-time PCR results confirmed that expression of PGC-1α， PEPCK， G6PC， FoxO1， and FoxO3 was significantly downregulated following treatment with G. pentaphyllum alcohol extract （P<0.05， P<0.01）， whereas mRNA expression of Adipoq， PPARα， GCK， and AMPK was significantly upregulated （P<0.05， P<0.01）. Metagenomic analysis showed that the relative abundance of Lactobacillus， Alistipes， and Akkermansia species was higher in the GP group than in the DM group. ConclusionG. pentaphyllum alcohol extract may improve glucose and lipid metabolism disorders in db/db mice by regulating the hepatic AMPK/PPARα pathway to suppress lipid deposition and alleviate hepatic steatosis， by inhibiting gluconeogenesis through the AMPK/PGC-1α and FoxO pathways to lower fasting blood glucose， and by increasing the abundance of beneficial gut bacteria such as Lactobacillus， Alistipes， and Akkermansia to restore gut microbiota balance.

ObjectiveTo investigate the efficacy and underlying mechanisms of Gynostemma pentaphyllum alcohol extract in improving glucose and lipid metabolism disorders in db/db mice through transcriptomics and gut microbiota analysis. MethodsEighteen db/db mice were randomly assigned to the model（DM） group， metformin（MET） group， and G. pentaphyllum alcohol extract（GP） group， with six mice in each group， based on stratification of fasting blood glucose and body weight. An additional six db/m mice were selected as the normal control（NC） group. Mice in the NC and DM groups were administered deionized water （10 mL·kg^-1） daily. The MET group received metformin （0.195 g·kg^-1） by gavage. The GP group was treated with G. pentaphyllum alcohol extract （3.9 g·kg^-1） by gavage for six weeks. Fasting blood glucose was measured every two weeks. After six weeks of intervention， serum levels of total cholesterol （TC）， triglycerides （TG）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， creatinine （CREA）， and blood urea nitrogen （BUN） were assessed. Enzyme-linked immunosorbent assay （ELISA） was used to measure insulin （FINS）， adiponectin （ADP）， and tumor necrosis factor-α （TNF-α）. Hematoxylin-eosin （HE） staining was used to observe liver histomorphology， periodic acid-Schiff （PAS） staining was employed to assess hepatic glycogen synthesis， and Oil Red O staining was used to detect hepatic lipid deposition. Liver transcriptomic data were used to identify differentially expressed genes in the liver and conduct enrichment analysis. Real-time PCR was employed to verify the expression levels of adiponectin gene （Adipoq）， peroxisome proliferator-activated receptor gamma coactivator-1α （PGC-1α）， AMP-activated protein kinase （AMPK）， peroxisome proliferator-activated receptor α （PPARα）， glucokinase （GCK）， forkhead box （Fox）O1， FoxO3， phosphoenolpyruvate carboxykinase （PEPCK）， and glucose-6-phosphatase （G6PC）. Metagenomic sequencing was conducted to analyze changes in gut microbiota composition. ResultsCompared with the NC group， the DM group exhibited significantly elevated fasting blood glucose （P<0.01）， serum AST， ALT， TC， TG， LDL-C， and HDL-C （P<0.01）. FINS， homeostatic model assessment for insulin resistance （HOMA-IR）， and the inflammatory cytokine TNF-α were significantly increased （P<0.01）， while ADP was significantly decreased （P<0.05）. Histological analysis confirmed severe hepatic steatosis and excessive lipid accumulation in the DM group， along with markedly reduced glycogen synthesis. Compared with the DM group， the GP group showed significantly decreased fasting blood glucose （P<0.01）， reduced serum TC， LDL-C， and HDL-C levels （P<0.05）， significantly decreased serum TG and AST levels （P<0.01）， significantly reduced FINS， HOMA-IR， and TNF-α levels （P<0.01）， and significantly increased ADP （P<0.01）. Hepatic steatosis and lipid deposition were significantly alleviated， while glycogen synthesis was markedly enhanced. Transcriptomic differential and enrichment analyses suggested that the mechanisms by which G. pentaphyllum alcohol extract improved hepatic glucose and lipid metabolism in db/db mice may involve regulation of the AMPK and FoxO signaling pathways. Real-time PCR results confirmed that expression of PGC-1α， PEPCK， G6PC， FoxO1， and FoxO3 was significantly downregulated following treatment with G. pentaphyllum alcohol extract （P<0.05， P<0.01）， whereas mRNA expression of Adipoq， PPARα， GCK， and AMPK was significantly upregulated （P<0.05， P<0.01）. Metagenomic analysis showed that the relative abundance of Lactobacillus， Alistipes， and Akkermansia species was higher in the GP group than in the DM group. ConclusionG. pentaphyllum alcohol extract may improve glucose and lipid metabolism disorders in db/db mice by regulating the hepatic AMPK/PPARα pathway to suppress lipid deposition and alleviate hepatic steatosis， by inhibiting gluconeogenesis through the AMPK/PGC-1α and FoxO pathways to lower fasting blood glucose， and by increasing the abundance of beneficial gut bacteria such as Lactobacillus， Alistipes， and Akkermansia to restore gut microbiota balance.

The transition of cancer cells from epithelial state to mesenchymal state awarded hepatocellular carcinoma (HCC) stem cell properties and induced tumorigenicity, drug resistance, and high recurrence rate. Reversing the mesenchymal state to epithelial state by inducing mesenchymal-epithelial remodeling could inhibit the progression of HCC. Using high-throughput screening, chrysin was selected from natural products to reverse epithelial-mesenchymal transition (EMT) by selectively increasing CDH1 expression. The target identification suggested chrysin exerted its anti-HCC effect through covalently and specifically binding threonine 205 (Thr205) of alpha-enolase (ENO1). For the first time, we revealed that ENO1 bound β-catenin mRNA, and recruited YTHDF2 to identify the m6A modified β-catenin in the 3'-UTR region to degrade β-catenin mRNA. Eventually, the CDH1 gene expression was improved through the regulation of β-catenin mRNA. ENO1/β-catenin mRNA interaction might be a promising target for cellular plasticity reprogramming. Moreover, chrysin could mediate mesenchymal‒epithelial remodeling through increasing degradation of β-catenin mRNA by promoting the binding of ENO1 and β-catenin mRNA. To the best of our knowledge, chrysin is the first reported small molecule inducing β-catenin mRNA degradation through binding to ENO1. The water-soluble derivative of chrysin may be a natural product-derived lead compound for circumventing metastasis, recurrence, and drug resistance of HCC by mediating mesenchymal‒epithelial remodeling.

In recent years,3D bioprinting technology has developed rapidly,becoming an essential tool in the fields of cancer research,tissue engineering,disease modeling and mechanistic studies.This paper reviewed the fundamental principles of bioprinting technology and its current applications in cancer research and tissue engineering.Bioprinting is an additive manufacturing technology that constructs complex three-dimensional tissue structures by digitally controlling the layer-by-layer deposition of biomaterials and living cells.The core steps of bioprinting include designing a 3D model,selecting appropriate bioprinting techniques and materials,printing layer by layer,followed by post-processing involving cell culture and functionalization.In cancer research,3D bioprinting can create complex tumor models that simulate the tumor microenvironment,revealing new mechanisms of tumor initiation and progression.Traditional in vitro models,such as 2D cell cultures or animal models,often fail to accurately replicate the complexity of human tumors.However,3D bioprinted tumor models,which mimic the dynamic interactions between tumor cells and their environment such as immune cells,stroma and blood vessels,offer a more biomimetic platform for studying tumor growth,invasion and metastasis.These models provide a research platform that closely mirrors actual tumor behavior.Additionally,Bioprinted models and scaffolds can be leveraged in personalized precision therapies by efficiently constructing patient-specific 3D models from their own cells.These models enable the prediction of patient's sensitivity to drugs and radiotherapy.Additionally,localized scaffolds can be developed to meet individual patient needs,allowing for the formulation of appropriate drug types and dosages.Furthermore,3D-printed scaffolds can support drug delivery by targeting specific areas,reducing drug-related side effects.They can also be used to facilitate local immunotherapy,cytokine therapy,cancer vaccines,and chimeric antigen receptor cell therapy,enhancing therapeutic outcomes.In tissue engineering,traditional tissue repair methods often struggle to address the complex requirements of constructing intricate tissue structures.3D bioprinting offers a novel solution by enabling the creation of complex tissue architectures and promoting tissue regeneration.Basic tissues,such as bone,cartilage and skin,which have higher regenerative capacities,are gradually being incorporated into clinical practice.Significant progress has also been made in the repair and reconstruction of more complex organs like the liver and heart,though considerable challenges remain before these advancements can be fully translated into clinical applications.Finally,this paper discussed the current challenges and future directions of 3D bioprinting in these fields,aiming to provide reference for researchers.

Background There is a lack of research evidence on the association between sugar-sweetened beverage (SSB) consumption and gestational diabetes mellitus (GDM) in China. Objective To explore the association between frequency of SSB consumption before pregnancy and risk of GDM in pregnant women in Shaanxi Province, and to provide a scientific basis for targeted interventions to control maternal blood glucose. Methods The recruitment to the China Birth Cohort study started in October 2020. Pregnant women at 6-16 weeks who had their first prenatal examination at five hospitals in Shaanxi Province were recruited. A maternal health questionnaire was used to collect basic information about pregnant women. A semi-quantitative food frequency questionnaire was used to collect the consumption of carbonated beverages, fruit and vegetable juice beverages, coffee beverages, and milk tea beverages in one year before pregnancy, which were summed to obtain the SSB consumption. Pregnant women were divided into three groups according to SSB consumption, namely <1 serving·week⁻¹, 1-4 servings·week⁻¹, and ≥5 servings·week⁻¹. GDM was confirmed by oral glucose tolerance test (OGTT) between 24-28 weeks of gestation. A binary logistic regression model was applied to explore the association between SSB consumption and risk of GDM. Multiple linear regression was applied to investigate the associations between SSB consumption (per 1-serving·d⁻¹ increase) and OGTT fasting plasma glucose, 1-hour glucose, and 2-hour glucose. Results A total of 3811 pregnant women were finally enrolled in this study, of which 752 developed GDM, with an incidence rate of 19.7%. The incidence rates of GDM in pregnant women with SSB consumption frequency of <1 serving·week⁻¹, 1-4 servings·week⁻¹, and ≥5 servings·week⁻¹ were 18.0%, 21.1%, and 26.8%, respectively. After adjusting for maternal age, pre-pregnancy body mass index (BMI), education, number of children born, family history of diabetes, smoking, alcohol consumption, physical activity level, and total energy intake, the risk of GDM increased by 26% (OR=1.26, 95%CI: 1.05, 1.50) in the 1-4 servings·week⁻¹ group and by 76% (OR=1.76, 95%CI: 1.31, 2.38) in the ≥5 servings·week⁻¹ group compared to the <1 serving·week⁻¹ SSB consumption group, respectively. Further stratified analysis revealed no interaction effect (P_interaction>0.05) between SSB consumption and maternal age, pre-pregnancy BMI, or first labor or not. For each additional SSB consumption per day, the risk of GDM increased by 94% (OR=1.94, 95%CI: 1.37, 2.75); and the maternal OGTT 1-hour glucose and 2-hour glucose increased by 0.33 mmol·L⁻¹ and 0.18 mmol·L⁻¹, respectively (P<0.05), and no significant increase in fasting plasma glucose was found (P>0.05). Conclusion Higher SSB consumption before pregnancy increases the risk of GDM in pregnant women.

Objective:To investigate the epidemiological characteristics of lower extremity deep vein thrombosis (DVT) in patients with femoral fracture.Methods:Retrospectively analyzed were the data of 2,571 patients with femoral fracture who had been treated at the Third Hospital of Hebei Medical University from January 2019 to December 2019. There were 1,079 males and 1,492 females, aged from 14 to 96 years (average, 67.1 years). There were 1,158 femoral neck fractures, 951 femoral intertrochanteric fractures, 309 femoral shaft fractures, and 153 femoral condylar fractures. 2,414 patients were treated surgically while 157 patients non-surgically. Color Doppler ultrasonography of both lower extremities was performed to determine the occurrence of DVT before operation and every week after operation for patients undergoing surgical treatment, and within 48 hours after admission and every week during hospitalization for those undergoing non-surgical treatment. The incidence and location of DVT were recorded for different femoral fractures.Results:The incidence of DVT in this cohort was 35.5%(913/2,517), that of proximal DVT 5.3%(135/2,571), and that of distal DVT 30.3% (778/2,571). In patients with femoral neck fracture, femoral intertrochanteric fracture, femoral shaft fracture and femoral condylar fracture, the incidence of DVT was respectively 28.8% (334/1,158), 44.7% (425/951), 30.7% (95/309) and 38.6% (59/153), the incidence of proximal DVT was respectively 2.7% (31/1,158), 5.6%(53/951), 9.7% (30/309) and 13.7% (21/153), and the incidence of distal DVT was respectively 26.2% (303/1,158), 39.1% (372/951), 21.0% (65/309) and 24.8%(38/153). The incidence of DVT in the femoral vein and above, popliteal vein, tibiofibular vein and intermuscular vein in this cohort was respectively 2.3%(60/2,571), 2.9%(75/2,571), 6.4%(165/2,571) and 23.8%(613/2,571).Conclusions:The incidence of DVT may be high in patients with femoral fracture, and the proximal DVT with a high risk of pulmonary embolism may occur more in patients with femoral condylar fracture.

Objective To explore whether a single bout of exhaustive exercise can activate autophagy in mice skeletal muscles and to investigate the effect and possible mechanisms of exercise-induced autophagy on antioxidant defense function in vivo.Methods Thirty healthy 4-week-old male C57BL/6 mice were randomly divided into a control group(n=6) and an exercise group(n=24).The latter was subjected to a single bout of exhaustive treadmill exercise.Gastrocnemius muscles of 6 mice on both sides were then isolated right after,as well as 6h,12h and 24h after the running.The manganese super oxide dismutase(MnSOD),total superoxide dismutase(T-SOD) and Copper-Zinc superoxide dismutase (CuznSOD) activities were determined using the xanthine oxidase method and the content of the total anti-oxygen capability(T-AOC) using the colorimetric method.The expression of Beclin1,P62 and Bcl2 mRNAs,as well as their proteins were determined using the fluorescent quantitative PCR and Western blotting.Results The expression of Beclin1 mRNA in gastrocnemius muscles of the exercise group increased more significantly than that of the control group right after the exercise,as well as 6 hours later(P＜0.01).P62 mRNA expression also increased significantly(P＜0.01) 24h after the exercise.In contrast,the expression of P62 protein at 0h,12h and 24h after the exercise decreased more significantly(P＜0.05 or P＜0.01) than that of the control group.Similarly,the expression of Bcl2 protein at 0h,6h and 12h increased significantly(P＜0.05 or P＜0.01).The MnSOD and CuznSOD activity increased significantly right after the exercise(P＜0.05).The T-SOD activity decreased significantly(P＜ 0.01) 6h after the exercise.T-AOC content increased significantly 6h,12h and 24h after the exercise (P＜0.05 or P＜0.01).The P62 protein expression was negatively correlated with T-AOC content.Conclusion The single bout of exhaustive treadmill exercise can activate the expression of such autophagyrelated factors as Beclin1 mRNA and P62 protein and increase the autophagy to a certain extent to maintain antioxidant defensive function of the skeletal muscle.

OBJECTIVE:To establish the method for simultaneous determination of 6 components in Lonicera japonica,and to study the content changes of them before and after before and after carbonized. METHODS:UPLC method was adopted. The deter-mination was performed on Agilent Eclipse Plus C18 RRHD column with mobile phase consisting of 0.1% phosphoric acid solu-tion-acetonitrile(gradient elution)at the flow rate of 0.2 mL /min. The detection wavelength was set at 350 nm,and column tem-perature was 25 ℃. The sample size was 1 μL. RESULTS:The linear ranges of chlorogenic acid,rutin,galuteolin,isochlorogenic acid A,isochlorogenic acid B and isochlorogenic acid C were 21.2-424 μg(r=0.9993),1.17-23 μg(r=0.9995),2.18-43 μg(r=0.9998),5.10-102 μg(r=0.9993),2.60-52 μg(r=0.9991),4.95-99 μg(r=0.9998),respectively. RSDs of precision,stability and repeatability tests were all lower than 2.0%. Recoveries were 97.11%-99.76%(RSD=1.20%,n=6),95.20%-99.90%(RSD=2.20%,n=6),95.71%-100.30%(RSD=2.20%,n=6),95.00%-96.98%(RSD=0.88%,n=6),96.47%-103.00%(RSD=2.40%, n=6),95.78%-103.80%(RSD=3.20%,n=6). Compared with before processing,the contents of rutin,isochlorogenic acid B and isochlorogenic acid C in L. japonica were increased along with processing,the contents of chlorogenic acid and isochlorogenic acid A were decreased significantly,while the content of galuteolin had no significant change. CONCLUSIONS:The method is sim-ple,precise,stable and repeatable,and can be used for simultaneous determination of 6 components in L. japonica. Those chemi-cal components have certain changes before and after carbonized.

Objective To study the roles of micheliolide on ovarian cancer cells. Methods Firstly, human ovarian cancer cell lines HeyA8, SKOV3 and A2780/DDP were treated with different concentration of micheliolide (0.25, 0.5, 1, 2.5, 5, 10, 20, 50 μmol/L) for 72 hours, then methyl thiazolyl tetrazolium (MTT) assay was used wo detect the growth of the human ovarian cancer cell lines and the stongest inhibited cell line were selected for the following test. Secondly, after HeyA8 cell line was treated with different concentration (5, 10, 20μmol/L) of micheliolide for 24 hours, the HeyA8 cell apoptosis was measured byflow cytometry. Thirdly, the expression of RelA mRNA in HeyA8 cell was detected through real-time PCR, the expressions of nuclear factor κB(NF-κB)signal pathway related protein RelA and the activited cysteinyl aspartate specific proteinase (caspase-9) were detected by western blot analysis. Results (1) The growth of HeyA8, SKOV3 and A2780/DDP cells were all significantly inhibited after being treated with different concentration of micheliolide for 72 hours and the roles of inhibition were all concentration dependant (P<0.05). The half maximal inhibitory concentration (IC50) of HeyA8, SKOV3 and A2780/DDP were (9.8±2.2), (12.0±2.1) and (12.8±1.8)μmol/L, respectively. We chose HeyA8 cell to do the following expreriments because of its best inhibited effect. (2) After HeyA8 cell was treated with micheliolide of different concentrations, as the concentration increased (20 and 0μmol/L, for example), the apoptosis rate of HeyA8 cell raised from (7.2±1.0)%to (17.4±1.1)%, the percentage of survived cells reduce from (92.8 ± 1.3)% to (82.6 ± 1.4)%,and the relative mRNA level of RelA decreased from 1.00 ± 0.13 to 0.18 ± 0.00 (P<0.01); furthermore, the expression of RelA protein was weaken and the activited caspase-9 protein expression was increased gradually. Conclusions Micheliolide plays a significantly inhibited role in HeyA8, SKOV3 and A2780/DDP cells. The inhibited role of micheliolide inovarian cancer cells might through inhibiting nuclear factor-kappa B (NF-кB) signaling pathway, and inducing the expression of activited caspase-9 protein to promoting apoptosis of HeyA8 cell.

Objective To investigate the diagnostic value of blood homocysteine,ankle -brachial index and brachial -ankle pulse wave velocity in elderly patients with coronary heart disease (CHD).Methods 97 patients with routine coronary angiography were classified into CHD group (65 cases)and non -CHD group (32 cases) according to the results of coronary angiography.There were 24 cases with single -vessel disease in 65 CHD cases, 21 cases with double -vessel disease and 20 patients with multivessel disease of CHD.Basic clinical parameters,age, gender,TC,TG,LDL -C,HDL -C,etc and blood HCY,ABI,baPWV levels were compared among groups.Results The age of double -vessel disease group,multivessel disease group was significantly higher than that in single -vessel disease group(t =3.721,3.927,all P <0.05).HCY,ABI,baPWV in CHD group were (18.29 ±2.73)μmol/L, (0.97 ±0.16),(16.38 ±1.27)m/s,which had statistically significant differences compared with non -CHD group (HCY:t =5.701,P <0.01;ABI:t =6.138,P <0.01;baPWV:t =15.132,P <0.01 ).There were no significant differences between single -vessel disease group and non -CHD group on the ABI(all P >0.05),and the ABI of multi -vessel disease,double vessel disease group were significantly lower than that of non -CHD group (all P <0.01).HCY,baPWV of CHD group were significantly higher than non -CHD group(all P <0.01 ),double vessel disease,HCY multivessel disease group,ABI,baPWV average water with single -vessel disease group were signifi-cantly different(all P <0.01 ).With the increase of coronary lesions involved,the blood HCY,baPWV showed an increasing trend and ABI showed an decreasing trend.Conclusion Combined detection of HCY,baPWV and ABI has great value in early detection and early intervention of CHD in the elderly.