Adrenal Hyperplasia, Congenital ; diagnosis ; Female ; Humans ; Infant, Newborn ; Male ; Neonatology ; Pregnancy ; Prenatal Diagnosis

Objective To investigate the relatonahip between TNF-α and ventricular arrhythmias after acute myocardial infarction(AMI)and its mechanism.Method Both the clinical and animal experiments were done.(1)Clinical experiment:Eighty patients with AMI were included in Union Hospital,Tongji Medical College,Huazhong University of Science and Techology,from May 2005 to November 2006 according to the WHO diagnostic criteria.Co-infection of diseases such as severe upper respiratory infection,lung infection,high fever,cancer,et al were excluded.The relationship between the levels of TNF-α and arrhythmias were observed at different times after AMI.A straight line correlation,analysis Was done.(2)Animal experiment:Different concentrations of TNF-αwere added to isohted rat hearts for observing the arrhythrnia effects.The effect of TNF-α on intracellular Ca2+ concentration was detected by laser confocal technique.All data were analyzed by SNK-q test using SPSS 13.0 sofeware prograrn.Results(1)The plasma levels of TNF-α were significantly associated with the Lown class of PVC after AMI and they were higher in AMI of anterior wall[(46.41±10.34)pg/ml]than other positions [(28.25±6.35)pg/ml,P<0.05].2)The frequency of ventricular arrhythmias was interrelated with the concentralions of TNF-α.Using etanercept beforehand,TNF-α induced a slight increase of intracellular Ca2+ intensity (P<0.05).Conclusions There was a relationship between TNF-αlevels and ventricular arrhythmisa in patients with AMI.Animal experiments confirmed the isolated heart perfusion with TNF-α induced ventricular arrhytrnias.Expression of TNF-α after AMI was related with the occurrence of ventricular arrhythrnias.The effect might be associated with the increased inuaeellular Ca2+ intensity caused by TNF-α.

BackgroundRetinal pigment epithelial(RPE) cells can secrete platelet-derived growth factor (PDGF) and PDGF receptor(PDGFR).Studies have shown that PDGF plays a key role in the formation of proliferative vitreous retinopathy(PVR). ObjectiveThis study was to investigate the proliferation and apoptosis changes of RPE after blockage of the PDGFR-α expression by antisense oligonucleotide ( ASODN ) in vitro. Methods Human RPE cells strain was cultured in low glucose DMEM with 10％ fetal bovine serum.Logarithmic phase cells were collected and incubated in 96-well plate at the density of 5 × 105 cells/hole.PDGFR-α ASODN was transfected into RPE cells at different concentrations for 48 hours.The cells of the blank control group were regularly cultured without any transfection.The changes of PDGFR-α expression were detected by reverse transcription-polymerase chain reaction(RT-PCR),and the proliferation of RPE was detected by MTT as the A490 value.Hoechst 33258 fluorescence staining was used to determine the apoptosis of RPE.Flow cytometry method (FCM) was applied to detect the change of cell cycle and apoptosis rate of RPE cells. ResultsThe A490 values of RPE cells were 1.45±0.12,1.07±0.06,0.65±0.05 in blank control group,1.0 μmol/L Lipo-ASODN group and 2.0 μmol/L Lipo-ASODN group with the significant difference(P=0.00 ),and that of 1.0 μmol/L Lipo-ASODN group and 2.0 μ mol/L Lipo-ASODN group were significantly lower than the blank control group ( P =0.00,0.00).Hoechst 33258 staining showed that the apoptosis cells were obviously more in Lipo-ASODN group compared with blank control group.PDGFR-α ASODN transfection induced an increase of percentage of RPE cells in G0/G1 phase( F =206.70,P =0.00),and the apoptosis rates in 1.0 μmol/L Lipo-ASODN group and 2.0 μmol/L Lipo-ASODN group were significantly enhanced in comparison with blank control group ( 37.8 ± 1.3 vs 10.5 ± 0.1,61.2 ± 1.9 vs 10.5 ± 0.1 ) ( F =1808.90,P =0.00 ).Expression intensity of PDGFR-α mRNA in RPE cells in Lipo-ASODN groups was lower. ConclusionsBlocking the PDGFR-α expression with ASODN technology can suppress proliferation and induce apoptosis of RPE cells.Intensity of PDGFR-α mRNA expression in RPE cells is ASODN dose-dependent.ASODN targeted to PDGFR-α offers an experimental basis of the gene therapy for PVR.

A in vitro platelet aggregation bioassay was developed for the quality control of XST capsules. The in vitro anti-platelet aggregation effect in rats was observed to detect the bioactivity of XST capsules. Panax notoginseng saponins and Xuesaitong lyophilizedpowder for injection were taken as standard control substances to determine the potency. According to the results, XST capsules showeda significant inhibitory effect on thrombin-induced platelet aggregation in a dose-dependent manner. The in vitro anti-platelet activity oflyophilized powder for injection was stabler than that of Panax notoginseng saponins, and so suitable to serve as a standard control substance. The biological potency of XST capsules compared with standard control substance was detected by using parallel line assay. According to the results, the established bioassay method had a good repeatability (RSD 2.92%). The sample test results could pass thereliability test(linear deviation P > 0.05, parallel deviation P > 0.05). This bioassay method could be used as one of the complementary quality control methods for XST capsules.
Animals ; Capsules ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Male ; Panax notoginseng ; chemistry ; Platelet Aggregation ; drug effects ; Platelet Aggregation Inhibitors ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Saponins ; pharmacology

OBJECTIVE: To investigate the effect of estrogen replacement therapy (ERT) in the early phase on the atherosclerosis and the level of plasminogen activator inhibitor-1(PAI-1). METHODS: Twenty-eight rabbits were randomly assigned to 4 groups: Group A, sham operation (n=7); Group B, ovariectomized without estradiol (n=7); Group C, ovariectomized with low-dose estradiol (n=7); and Group D, ovariectomized with high-dose estradiol (n=7). All rabbits were given 1% cholesterol diet for 12 weeks. Levels of blood lipid, estradiol, and PAI-1 were measured before the operation and at the end of the 4th and 12th weeks. Twelve weeks later, we took the aortas for pathological analysis and calculated the areas of atherosclerotic plaque. RESULTS: After 12 weeks, the estradiol level of Group B was significantly lower than that of Group A, and that of Group D was obviously higher than Group A. There was no significant difference between Group C and A. The concentrations of total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) in Group B significantly increased compared with Group A (P<0.01). The levels of TC and LDL-C of Group C and D were significantly lower than those of Group A. Whereas the concentrations of triglyceride (TG) and high density lipoprotein cholesterol (HDL-C) in Group B were lower than those of Groups A, C and D (P<0.01). In contrast to Groups A, C and D, the level of PAI-1 was significantly higher in the Group B (P<0.01), without significant differences among Groups A, C and D. The area of atherosclerotic lesion of aorta in Group B was significantly bigger than that of Group A, C and D. The areas of aortic atherosclerotic plaque in Group C and D were obviously smaller than those of Group A (P<0.01). CONCLUSION Transdermal estrogen replacement therapy in the early phase can improve the metabolism of the serum lipids, reduce the level of PAI-1, and probably provide the protective effect on the atheroma formation.
Administration, Cutaneous ; Animals ; Atherosclerosis ; blood ; drug therapy ; pathology ; Cholesterol ; blood ; Estradiol ; administration & dosage ; Estrogen Replacement Therapy ; Female ; Ovariectomy ; Plasminogen Activator Inhibitor 1 ; blood ; Rabbits ; Triglycerides ; blood

OBJECTIVEThe major cause of congenital adrenal hyperplasia (CAH) is 21-hydroxylase deficiency, which accounts for 90% - 95% of all cases in most populations. This study was conducted to characterize the molecular basis of the 21-hydroxylase deficiency and to obtain the spectrum of the CYP21 gene mutations in a group of Chinese patients, and analyze the relationship of genotype and phenotype.

METHODSTo detect the distribution of gene mutations in Chinese population samples from 52 patients with 21-hydroxylase deficiency from 51 families were collected, including two parents samples in 30 patients and one parents sample in 10 patients. Blood samples were obtained for extraction of peripheral blood lymphocytes. Polymerase chain reaction (PCR) followed by nesed PCR were used to study the 21-hydroxylase gene (CYP21) mutations. The primary PCR amplified two overlapping CYP21-specific DNA fragments, The product of the nested PCR which used products from the primary PCR was analysed by restriction fragment length polymorphism (RFLP) or amplification-created restriction site (ACRS). All patients were studied by 6 mutations, including P30L, I2g (intron 2 nt 656 c/a-->g splice mutation), E3Delta8nt (exon 3 codon111-codon113 8 bp deletion), I172N, V281L and Q318X.

RESULTSThrough analysis of 52 patients with 21-hydroxylase deficiency, in 5 patients no mutations were detected, in 17 patients only one mutated allele could be characterized, two different mutations were identified in 21 patients, three mutations were detected in 2 patients. Totally, in 73% of alleles the genotypes could be detected. The most common mutation was I2g, which present on 31% affected alleles, then followed by I172N, Q318X, V281L, P30L, E3Delta8nt, accounting for 23%, 14%, 9%, 3%, 2% of all identified mutations respectively, which included multiple mutations accounting for 6%. The most frequent molecular defects of the salt-wasting form were the I2g (45.7%), Q318X (26%). Of the simple virilizing form, the dominant mutations were I172N (40.7%) and I2g (18.5%).

CONCLUSIONSix different mutations were examined in this study, and the detected mutations accounted for 73% affected alleles, in which I2g and I172N were the most common mutations (accounting for 54%). Correlation between genotypes and phenotypes was compatible with the reported data. Two rounds of PCR followed by RFLP or ACRS analysis may provide important information for genetic counseling and for prenatal diagnosis.

Adrenal Hyperplasia, Congenital ; diagnosis ; enzymology ; genetics ; Child ; Child, Preschool ; China ; Family Health ; Female ; Gene Frequency ; Genotype ; Humans ; Infant ; Male ; Phenotype ; Point Mutation ; genetics ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Steroid 21-Hydroxylase ; genetics ; metabolism

OBJECTIVETo establish a rapid method of detecting CYP21 gene mutations.

METHODSFifty Chinese patients with 21-hydroxylase deficiency and some of their families were investigated. Blood samples were obtained for extraction of peripheral blood lymphocytes. A search for restriction sites discriminating between the morbid and the normal in CYP21 gene was made by the computer program DNAssist. PCR-based amplication-created restriction site(PCR-ACRS) was performed at I172N and R356W which are not natural recognition sequence. In addition, I172N and R356W were analysed in five families which conform to the applicability of PCR-ACRS.

RESULTSIn 50 identified 21-hydroxylase deficient Chinese patients, 21 were found to have I172 N (3 were homozygote, 18 were heterozygote); 8 were found to have R356W, all of them were heterozygote. By analysing the families, the findings were consistent with the characteristics of autosomal recessive genetic deficiency.

CONCLUSIONAnalysis of CYP21 gene point mutations using PCR-ACRS is relatively simple, accurate and feasible.

Adrenal Hyperplasia, Congenital ; enzymology ; genetics ; China ; DNA ; genetics ; metabolism ; DNA Mutational Analysis ; methods ; Deoxyribonucleases, Type II Site-Specific ; metabolism ; Female ; Humans ; Male ; Mutation ; Polymerase Chain Reaction ; methods ; Reproducibility of Results ; Sensitivity and Specificity ; Steroid 21-Hydroxylase ; genetics ; metabolism

Objective:To observe the effect of electroacupuncture (EA) on the expressions of acetylcholine (ACh) and mucin 5AC (MUC5AC) in the lungs of rats with chronic obstructive pulmonary disease (COPD),and explore the mechanism of EA in treating COPD.Methods:Thirty Sprague-Dawley (SD) rats were randomly divided into a control group,a COPD group,and an EA group,with 10 rats in each group.The control group was a group of normal rats.The COPD rat model was induced by cigarette smoke combined with lipopolysaccharide (LPS).The COPD rats were treated with EA at bilateral Feishu (BL 13) and Zusanli (ST 36) in the EA group,30 min each time,once a day,successively for 14 d.The lung function was tested.The contents of ACh and MUC5AC in lungs and bronchoalveolar lavage fluid (BALF) were detected by enzyme-linked immunosorbent assay (ELISA).Pearson method was used to analyze the correlation between pulmonary function and the content of MUC5AC in lungs.The mRNA and protein expressions of MUC5AC in lung tissues were detected by real-time polymerase chain reaction (RT-PCR) and Western blot (WB),respectively.The immune response of MUC5AC was observed by immunohistochemistry.Results:Eight rats were left in each group,and the other two died.Compared with the control group,the total airway resistance (Raw) increased significantly and dynamic compliance (Cdyn) decreased significantly in the COPD group (P＜0.01);compared with the COPD group,the Raw level declined significantly and Cdyn increased significantly in the EA group (P＜0.01).The contents of ACh and MUC5AC in the lungs and BALF were remarkably higher in the COPD group compared with those in the control group (P＜0.01,P＜0.001);compared with the COPD group,the contents of ACh and MUC5AC were significantly lower in the EA group (P＜0.05,P＜0.001).There was a negative correlation between MUC5AC content and lung function (P＜0.001).The mRNA and protein expressions of MUC5AC in the lungs were significantly higher in the COPD group than in the control group (P＜0.001);compared with the COPD group,the expressions were significantly lower in the EA group (P＜0.01).Compared with the control group,the immune response of MUC5AC in the airway epithelium significantly increased in the COPD group (P＜0.001);the immune response of MUC5AC was significantly lower in the EA group compared with that in the COPD group (P＜0.001).Conclusion:EA treatment can improve the lung function of COPD rats,which may be related to its effect in the down-regulation of ACh and MUC5AC contents in the lungs as well as the inhibition of mucus hypersecretion.

[Objective]To investigate the etiology,clinical manifestation,treatment and prognosis of optic disc vas-culitis.[Method]Twenty-four eyes of 21 patients were enrolled in this retrospective study.Eye examinations,treatment, and effect were recorded.[Result]Six were male and 15 were female.The age was between 19 and 43 years old(average:28.7±1.6).85.7% of the patients referred to the clinic with mild to moderate decreased vision.Edema of the optic disc can be seen in both types while tortuous veins can also be found in type 2.Similar characteristics were noticed in OCT,FFA, and etc.With a follow-up of 4.52±0.98 months after treatment(prednisone:initial dose 1.0-1.2 mg/kg),the BCVA of the affected eyes improved significantly.[Conclusion]Optic disc vasculitis is affected by autoimmune disorder,infection,hy-perlipidemia,and etc.Edema of the optic disc with/without tortuous veins and retinal hemorrhage can be noticed.Similar diseases should be excluded in avoidance of misdiagnosis. Systemic examination and complete solution should be per-formed.Glucocorticoid helps to improve the visual function.The application of anti-VEGF is effective in secondary macu-lar edema.However,the long-term efficacy is awaiting being confirmed.

OBJECTIVETo explore the therapeutic efficiency of human telomerase reverse transcriptase promoter (hTERTp) mediated horseradish peroxidase (HRP) catalyzed effects of indole-3-acetic (IAA) on laryngeal squamous cell carcinoma with different radiosensitivity in vivo.

METHODSHuman laryngeal squamous cell carcinoma Hep-2 and Hep-2R cells were transplanted into nude mice. After growing to about 30 approximately 50 mm3, the tumor-bearing mice were randomly divided into eight groups: Hep-2 line: combined group (A), gene group (B), radiation group (C) and blank group (D); Hep-2R line: combined group (AR), gene group (BR), radiation group (CR) and blank group (DR). The phTERTp-HRP was delivered by intratumoral injection and the IAA by intraperitoneal injection, combined with 2 Gy daily radiation to a total dose of 30 Gy. The tumor volume was recorded. The cell apoptosis was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. The expression of HRP protein was detected by AP immunohistochemisty.

RESULTSThe tumor growth of combined groups was attenuated significantly and the tumor volume of Hep-2R blank group was the largest. The inhibition rate of each group was: A: 54.8%, B: 10.0%, C: 31.9%; AR: 52.7%, BR: 24.8%, CR: 17.0%. In the combined groups, necrosis and apoptosis of tumor cells were observed under the light microscope and the apoptotic index [A (16.6 +/- 1.3)% vs. AR (17.6 +/- 1.3)%] of tumor cells was highest (P < 0.05). The HRP protein expression of BR (33.3 +/- 8.9)% was higher than that of B (21.9 +/- 5.7)%, which was directly up-regulated in the tumors (45.0% vs. 54.8%, P < 0.05) after radiation.

CONCLUSIONIn the Hep-2- and Hep-2R-transplantation tumors in nude mice, hTERTp can be induced by radiation and enhance the expression of horseradish peroxidase (HRP) gene according to telomerase activity. hTERTp-HRP/IAA system, which has synergistic effects with radiation and inhibits the tumor growth by induction of apoptosis and necrosis, may be a new gene-radiation strategy for the treatment of laryngeal carcinoma.

Animals ; Apoptosis ; Cell Line, Tumor ; Combined Modality Therapy ; Female ; Genetic Therapy ; methods ; Horseradish Peroxidase ; genetics ; metabolism ; Humans ; Indoleacetic Acids ; metabolism ; pharmacology ; Laryngeal Neoplasms ; metabolism ; pathology ; therapy ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Promoter Regions, Genetic ; Radiation Tolerance ; Radiotherapy ; methods ; Random Allocation ; Telomerase ; genetics ; Tumor Burden