Researchers commonly use cyclization recombination enzyme/locus of X-over P1 (Cre/loxP) technology-based conditional gene knockouts of model mice to investigate the functional roles of genes of interest in Sertoli and Leydig cells within the testis. However, the shortcomings of these genetic tools include high costs, lengthy experimental periods, and limited accessibility for researchers. Therefore, exploring alternative gene silencing techniques is of great practical value. In this study, we employed adeno-associated virus (AAV) as a vector for gene silencing in Sertoli and Leydig cells. Our findings demonstrated that AAV serotypes 1, 8, and 9 exhibited high infection efficiency in both types of testis cells. Importantly, we discovered that all three AAV serotypes exhibited exquisite specificity in targeting Sertoli cells via tubular injection while demonstrating remarkable selectivity in targeting Leydig cells via interstitial injection. We achieved cell-specific knockouts of the steroidogenic acute regulatory ( Star ) and luteinizing hormone/human chorionic gonadotropin receptor (Lhcgr) genes in Leydig cells, but not in Sertoli cells, using AAV9-single guide RNA (sgRNA)-mediated gene editing in Rosa26-LSL-Cas9 mice. Knockdown of androgen receptor ( Ar ) gene expression in Sertoli cells of wild-type mice was achieved via tubular injection of AAV9-short hairpin RNA (shRNA)-mediated targeting. Our findings offer technical approaches for investigating gene function in Sertoli and Leydig cells through AAV9-mediated gene silencing.
Animals ; Male ; Leydig Cells/metabolism* ; Mice ; Dependovirus/genetics* ; Sertoli Cells/metabolism* ; Gene Silencing ; Genetic Vectors ; Testis/cytology*

OBJECTIVE: To evaluate the immediate effect of Kuanxiong Aerosol (KXA) on perioperative coronary microcirculation in patients with unstable angina (UA) suffering from elective percutaneous coronary intervention (PCI). METHODS: From February 2021 to July 2023, UA inpatients who underwent PCI alone in the left anterior descending (LAD) branch were included. Random numbers were generated to divide patients into the trial group and the control group at a ratio of 1:1. The index of coronary microcirculation resistance (IMR) was measured before PCI, and the trial group was given two sprays of KXA, while the control group was not given. IMR was measured again after PCI, cardiac troponin I (cTnI) and creatine kinase isoenzyme-MB (CK-MB) were detected before and 24 h after surgery, and major cardiovascular adverse events (MACEs) were recorded for 30 days. The data statistics and analysis personnel were blinded. RESULTS: Totally 859 patients were screened, and 62 of them were involved into this study. Finally, 1 patient in the trial group failed to complete the post-PCI IMR and was excluded, 30 patients were included for data analysis, while 31 patients in the control group were enrolled in data analysis. There was no significant difference in baseline data (age, gender, risk factors, previous history, biochemical index, and drug therapy, etc.) between the two groups. In addition, differences in IMR, cTnI and CK-MB were not statistically significant between the two groups before surgery. After PCI, the IMR level of the trial group was significantly lower than that of the control group (19.56 ± 14.37 vs. 27.15 ± 15.03, P=0.048). Besides, the incidence of perioperative myocardial injury (PMI) was lower in the trial group, but the difference was not statistically significant (6.67% vs. 16.13%, P=0.425). No MACEs were reported in either group. CONCLUSIONS KXA has the potential of improving coronary microvascular dysfunction. This study provides reference for the application of KXA in UA patients undergoing elective PCI. (Registration No. ChiCTR2300069831).
Humans ; Percutaneous Coronary Intervention ; Male ; Microcirculation/drug effects* ; Female ; Angina, Unstable/physiopathology* ; Pilot Projects ; Middle Aged ; Aged ; Drugs, Chinese Herbal/pharmacology* ; Aerosols ; Troponin I/blood* ; Coronary Circulation/drug effects* ; Elective Surgical Procedures

The polymerase 1 and transcript release factor (PTRF)-cytoplasmic phospholipase A2 (cPLA2) phospholipid remodeling pathway facilitates tumor proliferation in glioma. Nevertheless, blockade of this pathway leads to the excessive activation of oncogenic receptors on the plasma membrane and subsequent drug resistance. Here, CD26/dipeptidyl peptidase 4 (DPP4) was identified through screening of CRISPR/Cas9 libraries. Suppressing PTRF-cPLA2 signaling resulted in the activation of the epidermal growth factor receptor (EGFR) pathway through phosphatidylcholine and lysophosphatidylcholine remodeling, which ultimately increased DPP4 transcription. In turn, DPP4 interacted with EGFR and prevented its ubiquitination. Linagliptin, a DPP4 inhibitor, facilitated the degradation of EGFR by blocking its interaction with DPP4. When combined with the cPLA2 inhibitor AACOCF3, it exhibited synergistic effects and led to a decrease in energy metabolism in glioblastoma cells. Subsequent in vivo investigations provided further evidence of a synergistic impact of linagliptin by augmenting the sensitivity of AACOCF3 and strengthening the efficacy of temozolomide. DPP4 serves as a novel target and establishes a constructive feedback loop with EGFR. Linagliptin is a potent inhibitor that promotes EGFR degradation by blocking the DPP4-EGFR interaction. This study presents innovative approaches for treating glioma by combining linagliptin with AACOCF3 and temozolomide.

Aim To investigate the therapeutic effect of the MW-9 on ulcerative colitis(UC)and reveal the underlying mechanism, so as to provide a scientific guidance for the MW-9 treatment of UC. Methods The model of lipopolysaccharide(LPS)-stimulated RAW264.7 macrophage cells was established. The effect of MW-9 on RAW264.7 cells viability was detected by MTT assay. The levels of nitric oxide(NO)in RAW264.7 macrophages were measured by Griess assay. Cell supernatants and serum levels of inflammatory cytokines containing IL-6, TNF-α and IL-1β were determined by ELISA kits. Dextran sulfate sodium(DSS)-induced UC model in mice was established and body weight of mice in each group was measured. The histopathological damage degree of colonic tissue was assessed by HE staining. The protein expression of p-p38, p-ERK1/2 and p-JNK was detected by Western blot. Results MW-9 intervention significantly inhibited NO release in RAW264.7 macrophages with IC50 of 20.47 mg·L-1 and decreased the overproduction of inflammatory factors IL-6, IL-1β and TNF-α(P<0.05). MW-9 had no cytotoxicity at the concentrations below 6 mg·L-1. After MW-9 treatment, mouse body weight was gradually reduced, and the serum IL-6, IL-1β and TNF-α levels were significantly down-regulated. Compared with the model group, MW-9 significantly decreased the expression of p-p38 and p-ERK1/2 protein. Conclusions MW-9 has significant anti-inflammatory activities both in vitro and in vivo, and its underlying mechanism for the treatment of UC may be associated with the inhibition of MAPK signaling pathway.

Objective:To explore the repair effect of resveratrol combined with Schwann cell-like cells(SCLCs)dif-ferentiated from adipose-derived stem cells(ADSCs)on sciatic nerve injury in rats.Methods:ADSCs were primarily cultured and induced to differentiate into SCLCs.Cell morphology was observed by scanning electron microscopy.West-ern Blot method was used to detect the expressions of S100 calcium-binding protein β(S100β),p75 neurotrophin receptor(p75NTR),and glial fibrillary acidic protein(GFAP).Rats were randomly divided into Control group(Con-trol),Schwann cell-like cell group(SCLCs),resveratrol group(Res),and resveratrol+Schwann cell-like cell group(Res+SCLCs).Eight weeks after the successful establishment of the sciatic nerve injury model,the sciatic nerve func-tion index(SFI)of each group was detected by footprint experiment;the mechanical withdrawal threshold(MWT)was measured by von Frey filament stimulation needle;The wet weight ratio(WR)of the tibialis anterior muscle was deter-mined by weighing method;Western Blot and RT-qPCR methods were used to detect the expressions of neurotrophin-3(NT-3),nerve growth factor(NGF),insulin-like growth factor-1(IGF-1),and brain-derived neurotrophic factor(BDNF)at the injury site.Results:After 8 days of induction of ADSCs,the cells had elongated poles and increased extracellular components;S100β,p75NTR,and GFAP proteins were highly expressed.After treatment with SCLCs,Res,and Res+SCLCs,the SFI and WR of the treatment groups were significantly better than those of the Control group(P＜0.05);the MWT of rats in the Res+SCLCs group and SCLCs group was reduced(P＜0.05).Western Blot re-sults showed that the expressions of NT-3,IGF-1,NGF,and BDNF proteins in rats in the Res+SCLCs group were higher than those in other groups(P＜0.05);The expressions of NT-3,NGF,and BDNF proteins in rats in the SCLCs group were higher than those in the Control group(P＜0.05);the expressions of NT-3 and NGF proteins in rats in the Res group were higher than those in the Control group(P＜0.05).RT-qPCR results showed that the expressions of NT-3,IGF-1,NGF,and BDNF mRNA in rats in the Res+SCLCs group were highly expressed;the expressions of NT-3,IGF-1,NGF,and BDNF mRNA in rats in the SCLCs group were higher than those in the Control group(P＜0.05);The expressions of IGF-1 and NGF mRNA in rats in the Res group were higher than those in the Control group(P＜0.05).Conclusion:Res combined with SCLCs differentiated from ADSCs has a good repair effect on sciatic nerve inju-ry in rats.

The prevalence rate of nonalcoholic fatty liver disease(NAFLD)in postmenopausal women is significantly higher than that in premenopausal women and even exceeds that in men of the same age group,and exercise intervention remains an effective method for the prevention and treatment of NAFLD in postmenopausal women.In addition,postmenopausal women with NAFLD often have comorbidities such as sarcopenia,osteoporosis,cardiovascular diseases,and diabetes,which requires targeted exercise prescriptions and proactive intervention for potential comorbidities.Through a literature review,this article provides targeted recommendations for exercise intervention prescriptions in postmenopausal women with NAFLD and related comorbidities.

Objective:Integrating genes and GEO database related cuproptosis chips,to analyze connection between cupropto-sis genes,immune infiltration and myocardial infarction(MI),construct risk prediction model,predict Western medicine and Chi-nese medicine,analyze miRNA-mRNA regulatory network,and to provides a new research direction for the future study of MI-related cuproptosis mechanism and immune infiltration.Methods:By GEO database retrieval of MI related chips,standardized processing and MI-related cuproptosis genes screening were performed,and immunoinfiltration analysis and quantification were performed based on the treated gene expression matrix,correlation between immune infiltrating cells and function was analyzed,as well as their differ-ences in MI group and the control group.Cuproptosis genes that most related to MI in immune infiltration were screened out,and the risk model was constructed to analyze the risk probability of cuproptosis genes in MI.The Enrichr website and Coremine Medical data-base were used to predict cuproptosis-related genes in MI in Western medicine and traditional Chinese medicine.Finally,the up-stream mirnas of FDX1 and SLC31A1 were predicted by miRTarBase,Starbase and Targetscan databases,and miRNA-mRNA regula-tory networks were constructed.Results:Correlation of immune infiltration showed that Tfh cells and B cells had the strongest positive correlation(r=0.68),while regulatory T cells and iDC had the strongest negative correlation(r=-0.63);the difference analysis of im-mune infiltration showed that the differences among mast cells,NK cells and Th1 cells in the MI group at the cellular level were the most significant(P<0.001);and the differences in APC co-inhibition and MHCⅠ at the functional level were the most significant(P<0.001).Six genes with the highest correlation between immune cells and immune function were screened out:ATP7A,DLD,FDX1,LIAS,LIPT1 and SLC31A1.Results of the risk model showed that the high levels of FDX1 and SLC31A1 were negatively correlated with the risk prediction of MI.A total of 21 Western medicines and 30 traditional medicines were predicted by database comparison.miRTarBase,Starbase and Targetscan databases predicted 9 upstream miRNAs of cuproptosis-related genes in MI,including has-miR-122-5p.Conclusion:Tfh cells,B cells,para-inflammatory,typeⅠinterferon response and other related immune cells and functions may play important roles in pathogenesis and prognosis of MI.FDX1 and SLC31A1 as the key genes of cuproptosis process,are negatively correlated with MI.A total of 30 kinds of traditional Chinese medicines including Spirulina,sheep liver,Wen ezhu,Pian jianghuang and Yujin may have potential value in treatment of MI.Finally,9 miRNAs including has-miR-122-5p may play an important role in the regulation of cuproptosis in myocardial infarction.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Objective To observe the clinical effect of Tongyuan Acupuncture combined with Modified Linggui Zhugan Decoction in the treatment of polycystic ovary syndrome(PCOS)with spleen deficiency and phlegm-damp syndrome.Methods A total of 86 patients with PCOS of spleen deficiency and phlegm-damp type were randomly divided into observation group and control group,43 cases in each group.The control group was treated with Metformin Hydrochloride Tablets combined with Ethinylestradiol and Cyproterone Acetate Tablets.The observation group was treated with Tongyuan Acupuncture combined with Modified Linggui Zhugan Decoction on the basis of the treatment of the control group.Both groups were treated for three consecutive months.After three months of treatment,the clinical efficacy of the two groups was evaluated.The changes of sex hormone indexes estradiol(E2),testosterone(T),luteinizing hormone(LH),follicle stimulating hormone(FSH)and prolactin(PRL)expression levels were observed before and after treatment in the two groups.The changes of glucose metabolism levels including insulin resistance index(HOMA-IR),fasting insulin(FINS)and fasting blood glucose(FPG)expression levels were compared before and after treatment between the two groups.Results(1)The total effective rate was 90.70%(39/43)in the observation group and 74.42%(32/43)in the control group.The curative effect of the observation group was superior to that of the control group,and the difference was statistically significant(P＜0.05).(2)After treatment,HOMA-IR,FINS and FPG in the two groups were significantly improved(P＜0.05),and the improvement of HOMA-IR,FINS and FPG in the observation group was superior to that in the control group,the difference being significant(P＜0.05).(3)After treatment,the levels of E2,T,LH,FSH and PRL in the two groups were significantly improved(P＜0.05),and the levels of E2,T,LH,FSH and PRL in the observation group were significantly superior to those in the control group,the differences were statistically significant(P＜0.05).Conclusion Tongyuan Acupuncture combined with Modified Linggui Zhugan Decoction in the treatment of spleen deficiency and phlegm-damp type of PCOS can effectively improve the sex hormone level of patients and regulate the glucose metabolism of patients.The clinical effect is significant.

The analysis of nano-micro nuclear particles has attracted significant attention due to the crucial role of their elemental and isotopic characteristics in tracing the origins of particulate matter and assessing its potential risks to human health and the environment.However,challenges persist in obtaining accurate and consistent element profiles and ratios for small-sized nanoparticles due to their low level and the transient nature.In this study,a high-speed digital oscilloscope was integrated with multiple collector inductively coupled plasma mass spectrometry(MC-ICP-MS)to develop a high time-resolution"Event-triggered signal capture"(ETSC)system for single particle analysis.This innovative approach enabled the analysis of element/isotope within rare earth nanoparticles at ag-fg level.The ETSC accurately recorded the complete profile of single particle,event captured by the electron multiplier with nanosecond time resolution,allowing for high-sensitivity element analysis and high-precision isotope analysis of single particles.The results demonstrated that the ETSC system could achieve quantitative analysis of ag levels of ytterbium(Yb)in 50-nm rare earth-doped particles,with a detection limit as low as 38 ag for Yb.Moreover,the isotopic precision of single particle analysis for 173/171Yb could reach 0.047(standard deviation),and the standard error for isotopic analysis of multiple particles could achieve a level of 2‰-3‰(permil)for 173/171Yb.Finally,the capability of ETSC system to analyze environmental samples was demonstrated through the analysis of doped ytterbium oxide nanoparticles.All these findings demonstrated that the ETSC provided a unique method for elemental and isotopic analysis of single nuclear particles.