ObjectiveTo investigate the effects of modified Ditan tang on genes related to the transcription-translation feedback loop （TTFL） of biorhythm in the rat model of non-alcoholic fatty liver disease （NAFLD） and its mechanism for prevention and treatment of NAFLD. MethodsSixty-five healthy SPF male SD rats were randomly assigned into blank （n=20）， model （n=15）， and low-， medium-， and high-dose （2.68， 5.36， and 10.72 g·kg^-1·d^-1， respectively） modified Ditan tang （n=10） groups. Other groups except the blank group were fed a high-fat diet for 12 weeks. The modified Ditan tang groups were treated with the decoction at corresponding doses by gavage， and the blank and model groups were treated with an equal volume of normal saline from the 9th week for 4 weeks. The levels of triglyceride （TG）， total cholesterol （TC）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） in the serum were measured by an automatic biochemical analyzer. TG and non-esterified fatty acid （NEFA） assay kits were used to measure the levels of TG and NEFA in the liver. The pathological changes in the hypothalamus and liver were observed by hematoxylin-eosin staining， and the lipid deposition in the liver was observed by oil red O staining. The levels of brain-muscle ARNT-like protein 1 （BMAL1/ARNTL） in the hypothalamus and liver were determined by immunohistochemical staining. The mRNA and protein levels of BMAL1， circadian locomotor output cycles kaput （CLOCK）， period circadian clock 2 （PER2）， and cryptochrome1 （Cry1） in the hypothalamus and liver were determined by Real-time PCR and Western blot， respectively. ResultsCompared with the blank group， the model group showed elevated levels of TG， TC， LDL-C， AST， and ALT （P<0.01） and a lowered level of HDL-C （P<0.05） in the serum， elevated levels of TG and NEFA in the liver （P<0.01）， pyknosis and deep staining of hypothalamic neuron cells， and a large number of vacuoles in the brain area. In addition， the model group showed lipid deposition in the liver， up-regulated mRNA and protein levels of CLOCK and BMAL1 （P<0.01）， and down-regulated mRNA and protein levels of Cry1 and PER2 （P<0.01） in the hypothalamus and liver. Compared with the model group， all the three modified Ditan tang groups showed lowered levels of TG， TC， LDL-C， ALT， and AST （P<0.05， P<0.01） and an elevated level of HDL-C （P<0.05） in the serum， and lowered levels of TG and NEFA （P<0.05， P<0.01） in the liver. Furthermore， the three groups showed alleviated pyknosis and deep staining of hypothalamic neuron cells， reduced lipid deposition in the liver， down-regulated mRNA and protein levels of CLOCK and BMAL1 （P<0.05， P<0.01）， and up-regulated mRNA and protein levels of Cry1 and PER2 （P<0.05， P<0.01） in the hypothalamus and liver. ConclusionModified Ditan tang can reduce lipid deposition in the liver and regulate the expression of CLOCK， BMAL1， Cry1， and PER2 in the TTFL of NAFLD rats.

Kupffer cells (KCs), as residents and sentinels of the liver, are involved in the formation of hepatic fibrosis (HF). However, the biological functions of circular RNAs (circRNAs) in KCs to HF have not been determined. In this study, the expression levels of circRNAs, microRNAs, and messenger RNAs (mRNAs) in KCs from a mouse model of HF mice were investigated using microarray and circRNA-Seq analyses. circDcbld2 was identified as a candidate circRNA in HF, as evidenced by its up-regulation in KCs. Silver staining and mass spectrometry showed that Wtap and Igf2bp2 bind to cirDcbld2. The suppression of circDcbld2 expression decreased the KC inflammatory response and oxidative stress and inhibited hepatic stellate cell (HSCs) activation, attenuating mouse liver fibrogenesis. Mechanistically, Wtap mediated the N ⁶-methyladenosine (m6A) methylation of circDcbld2, and Igf2bp2 recognized m6A-modified circDcbld2 and increased its stability. circDcbld2 contributes to the occurrence of HF by binding miR-144-3p/Et-1 to regulate the inflammatory response and oxidative stress. These findings indicate that circDcbld2 functions via the m6A/circDcbld2/miR-144-3p/Et-1 axis and may act as a potential biomarker for HF treatment.

The current study aimed to clarify the roles of apolipoprotein A5 (ApoA5) and milk fat globule-epidermal growth factor 8 (Mfge8) in regulating myocardial lipid deposition and the regulatory relationship between them. The serum levels of ApoA5 and Mfge8 in obese and healthy people were compared, and the obesity mouse model induced by the high-fat diet (HFD) was established. In addition, primary cardiomyocytes were purified and identified from the hearts of suckling mice. The 0.8 mmol/L sodium palmitate treatment was used to establish the lipid deposition cardiomyocyte model in vitro. ApoA5-overexpressing adenovirus was used to observe its effects on cardiac function and lipids. The expressions of the fatty acid uptake-related molecules and Mfge8 on transcription or translation levels were detected. Co-immunoprecipitation was used to verify the interaction between ApoA5 and Mfge8 proteins. Immunofluorescence was used to observe the co-localization of Mfge8 protein with ApoA5 or lysosome-associated membrane protein 2 (LAMP2). Recombinant rMfge8 was added to cardiomyocytes to investigate the regulatory mechanism of ApoA5 on Mfge8. The results showed that participants in the simple obesity group had a significant decrease in serum ApoA5 levels (P < 0.05) and a significant increase in Mfge8 levels (P < 0.05) in comparison with the healthy control group. The adenovirus treatment successfully overexpressed ApoA5 in HFD-fed obese mice and palmitic acid-induced lipid deposition cardiomyocytes, respectively. ApoA5 reduced the weight of HFD-fed obese mice (P < 0.05), shortened left ventricular isovolumic relaxation time (IVRT), increased left ventricular ejection fraction (LVEF), and significantly reduced plasma levels of triglycerides (TG) and cholesterol (CHOL) (P < 0.05). In myocardial tissue and cardiomyocytes, the overexpression of ApoA5 significantly reduced the deposition of TG (P < 0.05), transcription of fatty acid translocase (FAT/CD36) (P < 0.05), fatty acid-binding protein (FABP) (P < 0.05), and fatty acid transport protein (FATP) (P < 0.05), and protein expression of Mfge8 (P < 0.05), while the transcription levels of Mfge8 were not significantly altered (P > 0.05). In vitro, the Mfge8 protein was captured using ApoA5 as bait protein, indicating a direct interaction between them. Overexpression of ApoA5 led to an increase in co-localization of Mfge8 with ApoA5 or LAMP2 in cardiomyocytes under lipid deposition status. On this basis, exogenous added recombinant rMfge8 counteracted the improvement of lipid deposition in cardiomyocytes by ApoA5. The above results indicate that the overexpression of ApoA5 can reduce fatty acid uptake in myocardial cells under lipid deposition status by regulating the content and cellular localization of Mfge8 protein, thereby significantly reducing myocardial lipid deposition and improving cardiac diastolic and systolic function.
Animals ; Humans ; Mice ; Myocytes, Cardiac/metabolism* ; Obesity/physiopathology* ; Male ; Apolipoprotein A-V/blood* ; Lipid Metabolism/physiology* ; Milk Proteins/blood* ; Myocardium/metabolism* ; Diet, High-Fat ; Antigens, Surface/physiology* ; Mice, Inbred C57BL ; Cells, Cultured ; Female

Twelve compounds were isolated from the rice fermentation extracts of Penicillium expansum GY618 by silica column chromatography, Sephadex gel column chromatography, ODS column chromatography and semi preparative HPLC methods. They were determined as 11-hydroxyl-penicitrinone F (1), penicitrinone F (2), betulin (3), erythrodiol (4), ergosterol (5), ergost-5α,8α-epidioxy-6,22-dien-3β-ol (6), (5α,8α-epidioxy-(22E,24R)-ergosta-6,9(11),22-trien-3β-ol) (7), 5α,8α-epidioxy-(22E,24R)-23-methylergosta-6,22-dien-3β-ol (8), 5α,8α-epidioxy- 23,24(R)-dimethylcholesta-6,9(11),22-trien-3β-ol (9), dankasterone A (10), (17R)-4-hydroxy-17-methylincisterol (11) and ergosta-4,6,8(14),22-tetraen-3-one (12), through mass spectrometer, nuclear magnetic resonance (NMR) and comparison with the literature. Compound 1 was a new compound and compounds 2-4, 6-12 were isolated from Penicillium expansum fungus for the first time. The tyrosinase inhibitory activity experiment showed that compounds 1, 3 and 12 showed certain inhibitory activity against tyrosinase with IC₅₀ values of (75 ± 9), (69 ± 8) and (64 ± 2) μmol·L^-1, respectively. The IC₅₀ of other compounds were all greater than 100 μmol·L^-1, while IC₅₀ of the positive control kojic acid was (46 ± 4) μmol·L^-1.

AIM To study the protective effect of Gynura divaricate polysaccharide on gouty nephropathy(GN)induced by dry yeast combined with adenine in rats.METHODS Sixty male SD rats were randomly divided into the normal group,the model group,the allopurinol group(42 mg/kg),and the low-dose,medium-dose and high-dose G.divaricate polysaccharide groups(140,280,560 mg/kg).All the rats except those of the normal group were induced into GN models by intragastrical dosing of yeast(5 g/kg)and adenine(100 mg/kg)and intervened with corresponding drug administration simultaneously.After 35 days,the rats had their levels of creatinine(Cr)and uric acid(UA)in serum and urine detected and their fraction excretion of uric acid(FEUA)value determined;their kidney mass and volume measured and their levels of kidney index and density calculated;their renal pathological changes checked by HE staining;their renal GLUT9,URAT1,ABCG2 and OAT1 mRNA expressions dectected by RT-qPCR;and their renal GLUT9,URAT1,ABCG2 and OAT1 protein expressions dectected by Western blot.RESULTS Compared with the model group,each dose of G.divaricate polysaccharide group displayed decreased levels of kidney mass,kidney volume and kidney index(P＜0.01);increased levels of kidney density(P＜0.05,P＜0.01);decreased serum levels of UA and Cr(P＜0.01);increased urine levels of UA and Cr(P＜0.01);increased FEUA value(P＜0.01);decreased GLUT9,URAT1 mRNA and protein expressions(P＜0.05,P＜0.01);and increased ABCG2,OAT1 mRNA and protein expressions(P＜0.05,P＜0.01);and more alleviated renal histological aberrations.CONCLUSION G.divaricate polysaccharide exerts good protective effects against yeast/adenine-induced GN in rats probably through down-regulating protein expression of GLUT9,URAT1 and up-regulating ABCG2 and OAT1.

Objective To investigate the distribution and antimicrobial resistance profiles of clinically isolated Haemophilus influenzae and Moraxella catarrhalis in hospitals across China from 2015 to 2021,and provide evidence for rational use of antimicrobial agents.Methods Data of H.influenzae and M.catarrhalis strains isolated from 2015 to 2021 in CHINET program were collected for analysis,and antimicrobial susceptibility testing was performed by disc diffusion method or automated systems according to the uniform protocol of CHINET.The results were interpreted according to the CLSI breakpoints in 2022.Beta-lactamases was detected by using nitrocefin disk.Results From 2015 to 2021,a total of 43 642 strains of Haemophilus species were isolated,accounting for 2.91％of the total clinical isolates and 4.07％of Gram-negative bacteria in CHINET program.Among the 40 437 strains of H.influenzae,66.89％were isolated from children and 33.11％were isolated from adults.More than 90％of the H.influenzae strains were isolated from respiratory tract specimens.The prevalence of β-lactamase was 53.79％in H.influenzae strains.The H.influenzae strains isolated from children showed higher resistance rate than the strains isolated from adults.Overall,779 strains of H.influenzae did not produce β-lactamase but were resistant to ampicillin(BLNAR).Beta-lactamase-producing strains showed significantly higher resistance rates to these antimicrobial agents than the β-lactamase-nonproducing strains.Of the 16 191 M.catarrhalis strains,80.06％were isolated from children and 19.94％isolated from adults.M.catarrhalis strains were mostly susceptible to both amoxicillin-clavulanic acid and cefuroxime,evidenced by resistance rate lower than 2.0％.Conclusions The emergence of antibiotic-resistant H.influenzae due to β-lactamase production poses a challenge for clinical anti-infective treatment.Therefore,it is very important to implement antibiotic resistance surveillance for H.influenzae and guide rational antibiotic use.All local clinical microbiology laboratories should actively improve antibiotic susceptibility testing and strengthen antibiotic resistance surveillance for H.influenzae.

Objective To understand the changing composition and antibiotic resistance of bacterial species in the clinical isolates from outpatient and emergency department(hereinafter referred to as outpatients)and inpatient children over time in various hospitals,and to provide laboratory evidence for rational antibiotic use.Methods The data on clinically isolated pathogenic bacteria and antimicrobial susceptibility of isolates from outpatients and inpatient children in the CHINET program from 2015 to 2021 were collected and analyzed.Results A total of 278 471 isolates were isolated from pediatric patients in the CHINET program from 2015 to 2021.About 17.1％of the strains were isolated from outpatients,primarily group A β-hemolytic Streptococcus,Escherichia coli,and Staphylococcus aureus.Most of the strains(82.9％)were isolated from inpatients,mainly SS.aureus,E.coli,and H.influenzae.The prevalence of methicillin-resistant S.aureus(MRSA)in outpatients(24.5％)was lower than that in inpatient children(31.5％).The MRSA isolates from outpatients showed lower resistance rates to the antibiotics tested than the strains isolated from inpatient children.The prevalence of vancomycin-resistant Enterococcus faecalis or E.faecium and penicillin-resistant S.pneumoniae was low in either outpatients or inpatient children.S.pneumoniae,β-hemolytic Streptococcus and S.viridans showed high resistance rates to erythromycin.The prevalence of erythromycin-resistant group A β-hemolytic Streptococcus was higher in outpatients than that in inpatient children.The prevalence of β-lactamase-producing H.influenzae showed an overall upward trend in children,but lower in outpatients(45.1％)than in inpatient children(59.4％).The prevalence of carbapenem-resistant Klebsiella pneumoniae(CRKpn),carbapenem-resistant Pseudomonas aeruginosa(CRPae)and carbapenem-resistant Acinetobacter baumannii(CRAba)was 14％,11.7％,47.8％in outpatients,but 24.2％,20.6％,and 52.8％in inpatient children,respectively.The prevalence of multidrug-resistant E.coli,K.pneumoniae,Proteus mirabilis,P.aeruginosa and A.baumannii strains was lower in outpatients than in inpatient children.The prevalence of fluoroquinolone-resistant E.coli,ESBLs-producing K.pneumoniae,ESBLs-producing P.mirabilis,carbapenem-resistant E.coli(CREco),CRKpn,and CRPae was lower in children in outpatients than in inpatient children,but the prevalence of CRAba in 2021 was higher than in inpatient children.Conclusions The distribution of clinical isolates from children is different between outpatients and inpatients.The prevalence of MRSA,ESBL,and CRO was higher in inpatient children than in outpatients.Antibiotics should be used rationally in clinical practice based on etiological diagnosis and antimicrobial susceptibility test results.Ongoing antimicrobial resistance surveillance and prevention and control of hospital infections are crucial to curbing bacterial resistance.

Objective To investigate the changing antibiotic resistance profiles of E.coli isolated from patients in the 52 hospitals participating in the CHINET program from 2015 to 2021.Methods Antimicrobial susceptibility was tested for clinical isolates of E.coli according to the unified protocol of CHINET program.WHONET 5.6 and SPSS 20.0 software were used for data analysis.Results Atotal of 289 760 nonduplicate clinical strains ofE.coli were isolated from 2015 to 2021,mainly from urine samples(44.7±3.2)％.The proportion of E.coli strains isolated from urine samples was higher in females than in males(59.0％vs 29.5％).The proportion of E.coli strains isolated from respiratory tract and cerebrospinal fluid samples was significantly higher in children than in adults(16.7％vs 7.8％,0.8％vs 0.1％,both P＜0.05).The isolates from internal medicine department accounted for the largest proportion(28.9±2.8)％with an increasing trend over years.Overall,the prevalence of ESBLs-producing E.coli and carbapenem resistant E.coli(CREco)was 55.9％and 1.8％,respectively during the 7-year period.The prevalence of ESBLs-producing E.coli was the highest in tertiary hospitals each year from 2015 to 2021 compared to secondary hospitals.The prevalence of CREco was higher in children's hospitals compared to secondary and tertiary hospitals each year from 2015 to 2021.The prevalence of ESBLs-producing E.coli in tertiary hospitals and children's hospitals and the prevalence of CREco in children's hospitals showed a decreasing trend over the 7-year period.The prevalence of CREco in secondary and tertiary hospitals increased slowly.Antibiotic resistance rates changed slowly from 2015 to 2021.Carbapenem drugs(imipenem,meropenem)were the most active drugs amongβ-lactams against E.coli(resistance rate≤2.1％).The resistance rates of E.coli to β-lactam/β-lactam inhibitor combinations(piperacillin-tazobactam,cefoperazone-sulbactam),aminoglycosides(amikacin),nitrofurantoin and fosfomycin(for urinary isolates only)were all less than 10％.The resistance rate of E.coli strains to antibiotics varied with the level of hospitals and the departments where the strains were isolated,especially for cefazolin and ciprofloxacin,to which the resistance rate of E.coli strains from children in non-ICU departments was significantly lower than that of the strains isolated from other departments(P＜0.05).The E.coli isolates from ICU showed higher resistance rate to most antimicrobial agents tested(excluding tigecycline)than the strains isolated from other departments.The E.coli strains isolated from tertiary hospitals showed higher resistance rates to the antimicrobial agents tested(excluding tigecycline,polymyxin B,cefepime and carbapenems)than the strains from secondary hospitals and children's hospitals.Conclusions E.coli is an important pathogen causing clinical infection.More than half of the clinical isolates produced ESBL.The prevalence of CREco is increasing in secondary and tertiary hospitals over the 7-year period even though the overall prevalence is still low.This is an issue of concern.

Objective:To formulate a community teacher standard for the training of general practitioner program (direction in hospice care).Methods:This was a qualitative study. Twenty-nine experts in general practice and hospice care from various teaching hospitals and community health service centers were invited to participate in two rounds of Delphi consultation from April to August 2022. Based on the results of the consultation, a standard for community faculty training in comprehensive care and end-of-life care for general practitioners was formulated, and the weight coefficients of the indicators were calculated.Results:Of the 29 experts, 11 (37.9%) were male, their age was (52.9±8.9) years, and their working life was (17.0±7.2) years. In the two rounds of expert consultation, the expert positive coefficient was 100.0% (29/29) and 96.6% (28/29), the expert authority coefficient was 0.789 and 0.851, and the expert opinion coordination coefficient W of the importance of all indicators was 0.124 and 0.131, respectively ( χ2=123.01, 131.71, P<0.05). Finally, a set of community-based teacher standard for general practitioner specialty training (direction in hospice care) was established, which consisted of 4 first-level indicators and 30 second-level indicators. The four first-level indicators were basic literacy, clinical qualification and ability, teaching qualification and ability, and scientific research ability, all with the weight coefficients of 0.254, 0.254, 0.252 and 0.240, respectively. Conclusion:The standard of teachers in the community base of general practitioner specialty training (direction in hospice care) established in this study is more scientific, which is conducive to the comprehensive evaluation of teachers in the community training base.