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 calculate both the epidemic and intensity thresholds for different levels in Beijing and to establish a tiered alert system in the 2018-2019 influenza season as well as to evaluate the performance of calculated thresholds.Method Weekly count of influenza-like illness and percentage of influenza-like illness (ILI％) of the last five influenza seasons were modeled by 'moving epidemic method'(MEM) to calculate the influenza epidemic and intensity thresholds at different levels.A cross-validation procedure was used to evaluate the performance.Indicators of Matthew correlation coefficient,Youden's index,sensitivity and specificity were calculated.Results For weekly count of influenza-like illness,data showed that the epidemic threshold for 2018-2019influenza season was 12 984 and the medium,high and very high intensity thresholds were 22 503,37 589,47 157,respectively.Matthew correlation coefficient of the epidemic threshold was 62％ and youden's index as 60％,sensitivity as 69％,specificity as 91％.Data on weekly ILI％,the epidemic threshold for 2018-2019 influenza season was 1.66％,with medium,high and very high intensity thresholds as 2.46％,3.84％ and 4.66％,respectively.The overall Matthew correlation coefficient of the epidemic threshold was 59％,with 54％ for the Youden's index,sensitivity as 60％ and specificity as 94％.Conclusions MEM produced a good specific signal for detecting the influenza epidemics and the accuracy of the method was acceptable.The early warning performance regarding the application of weekly count on influenza-like illness was slightly better than ILI％.This method could be applied in the practical influenza epidemic alert "work in Beijing".

OBJECTIVE: To explore the value of galactose-deficient IgA1 (Gd-IgA1) in the early diagnosis of Henoch-Schönlein purpura nephritis (HSPN) in children. METHODS: A total of 67 hospitalized children who were definitely diagnosed with HSPN between January and April 2018 and 58 hospitalized children with Henoch-Schönlein purpura (HSP) were enrolled in the study. Twenty children undergoing routine physical examinations served as controls. The levels of serum and urine Gd-IgA1 were determined using ELISA. The receiver operating characteristic curve was used to analyze the value of serum Gd-IgA1 and urine Gd-IgA1/urine creatinine ratio in the diagnosis of HSPN. RESULTS: The level of serum Gd-IgA1 and urine Gd-IgA1/urine creatinine ratio in children with HSP or HSPN were significantly higher than those in healthy control group (P<0.01), with a significantly greater increase observed in children with HSPN (P<0.01). Serum Gd-IgA1 ≥1 485.57 U/mL and/or urine Gd-IgA1/urine creatinine ratio ≥105.74 were of favorable value in the diagnosis of HSPN. During the six-month follow-up of the 49 children with HSP, the incidence of HSPN was 47% (23/49), which included a 100% incidence in children with serum Gd-IgA1 ≥1 485.57 U/mL and a 73% incidence in children with urine Gd-IgA1/urine creatinine ratio ≥105.74. CONCLUSIONS Serum and urine Gd-IgA1 is of favorable clinical value in the early diagnosis of HSPN.
Child ; Early Diagnosis ; Galactose ; Glomerulonephritis, IGA ; Humans ; Immunoglobulin A ; Purpura, Schoenlein-Henoch

The present study aims to investigate the lignan constituents from Sambucus williamsii and their proliferation effects on osteoblast-like UMR106 cells. Seven compounds were isolated and purified by macroporous resin D101, silica gel, Sephadex LH-20, Toyopearl HW-40, ODS column chromatographies and Preparative HPLC(C-18). Their structures were elucidated by spectroscopic methods as threo-guaiacylglycerol-beta-0-4'-conifery ether (1), lirioresinol A (2), 1-hydroxypinoresinol (3), 5-methoxybalanophonin (4), balanophonin (5), 5-methoxy-trans-dihydrodehydrodiconiferyl alcohol (6), and p-hydroxybenzaldehyde (7). Compounds 3-7 were obtained from this genus for the first time. The proliferation effects of all isolated compounds on osteoblast-like UMR106 cells were determined. Compounds 1-7 (1 x 10(-12)-1 x 10(-7) mol x L(-1)) increased UMR106 cell proliferation to some extent.
Cell Line ; Cell Proliferation ; drug effects ; Lignans ; isolation & purification ; pharmacology ; Osteoblasts ; cytology ; drug effects ; Plant Stems ; chemistry ; Sambucus ; chemistry

Semen-derived enhancer of viral infection(SEVI) is a peptide fragment (PAP248-286) from prostatic acid phosphatase(PAP), which can enhance human immunodeficiency virus infection. The mechanisms of SEVI include: (1) SEVI with several cationic amino acid residues reduced electrostatic repulsion between HIV virus and the target cells; (2) The disorder state of SEVI in the human body fluids was helpful to the interaction between virus and the target cell membranes; (3) SEVI could capture HIV particles directly and speed the velocity of virus on the surface of the target cells and improve adsorption and fusion. Currently, the substances of inhibiting SEVI activity include: EGCG from green tea, small molecule compound of aminoquinoline Surfen, ThT analogs BTA-EG6. Those compounds might block the combination of HIV and SEVI or prevent the formation of amyloid fibers, and then reduce the enhancement of SEVI. The studies on the biological characteristics and mechanisms of SEVI have a big benefit for the prevention and treatment of HIV infection.
HIV Infections ; etiology ; transmission ; Humans ; Male ; Semen ; physiology ; Sexually Transmitted Diseases, Viral ; etiology ; Static Electricity

Objective To study the natural history of HIV-1 infection among intravenous drug users (IDUs) detected in late 1989 in the study area and the factors related to survival of these IDUs infected with HIV. Methods 196 injecting drug users first detected during August and December, 1989 were observed in Ruili county, Yunnan province. Data gathered from the 20-year follow-up program was collected and analyzed retrospectively. Results After 20 years' follow-up period, 90.3% of the 196 IDUs with HIV infection died, 5.1% of them were still alive, and 4.6% were lost. The crude pre-AIDS mortality rate was 98.1/1000 person-years, and the AIDS mortality rate was 54.9/1000 person-years. Malaria, septicemia were the main causes of death among the natural diseases whereas overdose and accidental causes were the principal causes related to those non-disease deaths.The median survival time from sero-conversion to death was 8.6 years (95%CI: 7.6-9.7). The median survival time from sero-conversion to death due to AIDS was 11.3 years (95%CI: 10.3-12.8) with the incubation time as around 10.3 years. People older than 30 years at seroconversion and length of drug usage were associated with shorter survival time, with hazards ratios as 1.9 and 0.7, respectively.Conclusion A high pre-AIDS mortality was observed among IDUs. Both the median survival time from sero-conversion to death and the HIV incubation period were shorter than that observed in the developed countries. Age of HIV infection seemed to have a strong effect on survival.

Beijing has been one of the epicenters attacked most severely by the SARS-CoV (severe acute respiratory syndrome-associated coronavirus) since the first patient was diagnosed in one of the city's hospitals. We now report complete genome sequences of the BJ Group, including four isolates (Isolates BJ01, BJ02, BJ03, and BJ04) of the SARS-CoV. It is remarkable that all members of the BJ Group share a common haplotype, consisting of seven loci that differentiate the group from other isolates published to date. Among 42 substitutions uniquely identified from the BJ group, 32 are non-synonymous changes at the amino acid level. Rooted phylogenetic trees, proposed on the basis of haplotypes and other sequence variations of SARS-CoV isolates from Canada, USA, Singapore, and China, gave rise to different paradigms but positioned the BJ Group, together with the newly discovered GD01 (GD-Ins29) in the same clade, followed by the H-U Group (from Hong Kong to USA) and the H-T Group (from Hong Kong to Toronto), leaving the SP Group (Singapore) more distant. This result appears to suggest a possible transmission path from Guangdong to Beijing/Hong Kong, then to other countries and regions.
Genome, Viral ; Haplotypes ; Humans ; Mutation ; Open Reading Frames ; Phylogeny ; SARS Virus ; genetics

OBJECTIVETo demonstrate the effectiveness of a retrovirus-based plasmid vector coupled with nanometer material-polyamidoamine (PAMAM) dendrimer in stable gene expression of FVIII in vitro and to study the cytotoxicity of PAMAM.

METHODSThe retrovirus-based plasmid vector pLNC-FVIII BD was generated by cloning a B-domain-deleted (760aa - 1639aa) FVIII cDNA (FVIIIBD cDNA) into retroviral vector pLNCX. The complex that contained PAMAM and pLNC-FVIII BD transfer FVIII BD cDNA into NIH3T3 cell line. In day 2, 5, 10, 15, 30 after transferring, the antigen and procoagulant activity of human FVIII in the cell culture medium were measured by ELISA assay and one-stage method, respectively. RT-PCR was performed for the detection of FVIII BD mRNA. Inhibitory percentage of cell vitality was used for cytotoxicity of PAMAM.

RESULTSHuman FVIII was expressed for 30 days by transfected cells. The mean procoagulant activity of secreted FVIII in these 30 days was 0.929 U/ml, and the FVIII antigen was 0.188 micro g/ml by 10(6) cells in 24 hours, respectively. The level of FVIII didn't significantly decreased during these days. Inhibitory percent of cell vitality was only 5.32%.

CONCLUSIONPAMAM could effectively transfer pLNC-FVIII BD into NIH3T3 cells and FVIII could be stably and effectively expressed by the transfected cells. Cytotoxicity of PAMAM was low.

Animals ; Dendrimers ; Factor VIII ; genetics ; Genetic Vectors ; genetics ; Mice ; NIH 3T3 Cells ; Plasmids ; Polyamines ; pharmacology ; Retroviridae ; genetics

OBJECTIVETo explore the gene mutation type of an inherited coagulation factor VII deficiency pedigree.

METHODSFVII:Ag, FVII:C, FVIIa were detected to classify deficiency type. FVII gene mutations were analysed in the proband and her family members by DNA directly sequencing. Biostructural pathology of the identified mutation was analysed by molecular modeling.

RESULTSHomozygosity of C-->T transition at position 11514 in exon 8 resulting in Thr359Met was identified in the proband, and heterozygosity for Thr359Met was confirmed in her parents, her son and some other family members. Thr359Met induces CRM-deficiency. It is found by computer simulated molecular model that the replacement of Thr by Met which has a larger and longer side chain might cause steric hindrance, and change the number of H-bonds.

CONCLUSIONSHomozygous missense mutation Thr359Met was found in a pedigree of hereditary FVII deficiency. This mutation might change the configuration of protein molecule and result in severe FVII deficiency.

Adolescent ; Adult ; Aged ; DNA Mutational Analysis ; Factor VII ; genetics ; Factor VII Deficiency ; genetics ; Female ; Homozygote ; Humans ; Male ; Middle Aged ; Mutation, Missense ; Pedigree ; Polymerase Chain Reaction

Objective：This study was designed to establish a salivary EBV-EA IgA and DNase IgA test technique, and seek a fast and specific diagnostic technique for nasopharyngeal carcinoma (NPC). Methods：Polypeptides of EBV-DNase(ED) and EA-D was synthesized as catching antigens. With ELISA technique, IgA/ED and IgA/EA-D were evaluated respectively in saliva and serum from NPC patients and healthy volunteers. Results：After statistic analysis of the optical density(OD) values of samples, the diagnostic criteria of NPC in the examination of either IgA/ED or IgA/EA-D was defined as following：OD≥ 0.3 for serum and OD≥ 0.45 for saliva. Significantly statistical difference existed between the values of either IgA/ED or IgA/EA-D titer in patients with NPC and the values in healthy volunteers,P<0.0001. The coincidence rates between the diagnosis of above IgA/ED or IgA/EA-D titer testes and corresponding histological diagnosis were 72.6％ － 77.3％ . Conclusion： The Elisa test to detect salivary IgA/ED and IgA/EA-D with synthesized polypeptides is a simple, repeatable, and cheap technique with stability and sensitivity. However its coincidence rate with histology should be improved.?