Objective To investigate the effects of simvastatin on the production of interleukin (IL)-17and B-cell activating transcription factor (BATF) in the peripheral blood mononuclear cells (PBMCs) of rheumatoid arthritis (RA) patients and healthy individuals.Methods PBMCs were isolated from heparinized blood of healthy donors or RA patients using Ficoll-Hypaque density gradient centrifugation.The cells were stimulated by PMA and ionomycin in the absence or presence of simvastatin or MVA at 37 ℃ 5％CO2.The mRAN level of IL-17,BATF and GAPDH was detected by RT-PCR; the protein level of IL-17 in supernatants was assayed by ELISA kit; and the protein level of BATF was detected by Western Blotting.The comparison between the two groups was carried out by paired-t test and Chi-square test was used for muhi-group comparison.Results PBMCs of healthy donors [(69.2±12.2) vs (8.1±2.2) pg/ml,P＜0.05; (76.6±14.7) vs (10.2±7.2) pg/ml,P＜0.05] and RA patients [(79.6±12.7) vs (15.8±5.8) pg/ml,P＜0.05; (90.3±9.7) vs (12.9±7.9) pg/ml,P＜0.05] were stimulated with PMA and ionomycin to produce high levels of IL-17.After treatment with simvastatin,the expression and secretion level of IL-17 in healthy controls and RA PBMCs were markedly decreased.The inhibition of simvastatin on the production of IL-17 was reversed by mevalonic acid (MVA),but no significant changes of BATF after treating with simvastatin.Conclusion Simvastatin inhibits the production of IL-17 in the PBMCs at gene and protein levels,which is not targeted at suppressing the expression of IL-17 transcription factor BATF.

Objective:To investigate the relationship between herpesviridae and malignant or benign laryngeal diseases.Method:128 paraffin-embedded laryngeal squamous cell carcinoma and laryngeal epithelium hyperplastic lesions were detected by polymerase chain reaction (PCR) and PCR-ISH for herpesviridae. Result:HSV-1 was detected in 10 cases by PCR,among them 3 were laryngeal squamous cell carcinoma (LSCC),1 was carcinoma in situ(CIS),4 were laryngeal polyps and 2 were laryngeal keratosis. Except 1 LSCC and 1 CIS, 8 of 10 cases were positive while detected by PCR-ISH. In benign diseases, signals were shown from basal layer to superficial cell; in malignant lesions, the signals were scattered in the diseases.Conclusion:Most of laryngeal diseases were not related to herpesviridae, but HSV-1 may acts as initiator in the development of a few cases.

Objective To characterize and quantify the CD4 +CD25 + regulatory T (Treg) cell population in peripheral blood of patients with ankylosing spondylitis (AS) and to determine the influence of treatment with tumor necrosis factor (TNF)-a inhibitors on them.Methods Peripheral blood mononuclear cells (PBMC) were isolated from 25 patients with active AS,in which 10 patients were treated with 12 weeks of etanercept,and 21 healthy subjects.CD4+CD25high T cells were analyzed using flow cytometry,and mRNA expression of FOXP3 was determined by real-time polymerase chain reaction (PCR).Proliferation of T cells to PHA was measured by WST-1 assay using depleted CD25+ cells by immunomagnetic sorting.Results There was no significant difference in the percentage of CD4+CD25high cells in peripheral blood between patients with active AS and controls (P＞0.05).However,PBMC from patients with active AS expressed reduced levels of FOXP3 mRNA (P＜0.01) which were inversely correlated with C-reactive protein (CRP)(P＜0.01).CD4+CD25+ cells in peripheral blood of both active AS patients and controls exhibited suppressive capacity on the proliferation of effector T cells in vitro (both P＜0.01).Treatment with etanereept increased significantly CD4+CD25high cells and FOXP3 mRNA expression (both P＜0.01),with negative correlations between these increases and decrease in CRP levels (P＜0.05 and P＜0.01,respectively).Conclusion In AS patients,peripheral FOXP3-expressing CD4 +CD25 + Treg cells are abnormal,and are up-regulated by etanercept treatment.This suggests a possible pathogenesis of AS and a potential mechanism for clinical efficacy of TNF-α inhibitors.

Objective To evaluate a new enzyme-linked immunospot assay (TSPOT-TB) for the diagnosis of latent tuberculosis infection in patients with rheumatic diseases.Methods The rapid TSPOT-TB assay was applied to detect ESAT-6 and CFP-10 specific T cells in blood samples from 126 rheumatic disease patients.A PPD skin test was performed on all patients simultaneously.Results The positive rate of TSPOT assay was 23.8% and that of PPD skin test was 34.9%.The overall agreement between the 2 tests was 71.4%.Among PPD (-) patients (n=82),11 were TSPOT (+) ( 13.4% ).Among PPD (+) patients (n=44),25 were TSPOT(-) ( 56.8% ).The patients who got BCG vaccination showed a significantly higher rate of positive results of PPD skin test than those who did not(41% vs 19%,P＜0.05).While in TSPOT assay,the BCG vaccination did not show any influence on TSPOT results (22% vs 27%,P＞0.05).Conclusion BCG vaccina-tion affects the results of PPD test in patients with rheumatic diseases,but has no influence on TSPOT results.The infection rate of latent tuberculosis in patients with rheumatic diseases in our study is 23.8% detected by TSPOT.

Objective To compare the efficacy of the conventional PPD skin test and a new enzymelinked immunospot assay(TSPOT-TB)for diagnosing latent tuberculosis infection(LTBI)in patients with rheumatic diseases.Methods Two hundred and sixty rheumatic patients were enrolled,and all were screened for LTBI based on clinical history,chest X-ray,PPD skin test or TSPOT.Results The positive rate of TSPOT assay was 24.1%and that of PPD skin test was 39.4%.The overall concordance rate between the 2tests was 61.0%.Among PPD negative patients (n=149).29 were TSPOT(+)(19.5%).Among PPD(+)patients(n=98),69 were TSPOT(-)(70.0%).The patients who got BCG vaccination or had history of tuberculosis infection showed a significantly higher rate of positive result of PPD skin test than those who did not (P<0.05 or P<0.01).While in TSPOT assay,the BCG vaccination or history of tuberculosis infection did not show influence on TSPOT results(P>0.05).Of the 127 patients who received biological agents after screening for LTBI,9 patients were pretreated with isoniazide.Twenty-seven patients stopped biological agent treatment because of the positive results of PPD or TSPOT.Twenty three patients who had positive PPD but negative TSPOT results received biological agent treatment without isoniazide,and none of them developed active tubereulosis after 6 to 18 months of follow-up.Conclusion BCG vaccination affects the result of PPD test in rheumatic patients,but has no influence on TSPOT results.The infection rate of latent tuberculosis of rheumatic patients in our research is 23.8%detected by TSPOT.

Abstract:Objective To better understand the cholinergic receptors in vestibular hair cells (VHC) and their subtypes, and to investigate the effects of cholinergic agonists on intracellular calcium concentration ([Ca2+]i ) in guinea pig VHCs.Methods VHCs were isolated from guinea pig crista ampullaris by enzymatic and mechanical methods. The effect of cholinergic agonists on [Ca2+]i was examined using laser scanning confocal microscopy and the Ca2+ sensitive dye Fluo-3.Results The results showed that the addition of acetylcholine (ACh) and carbachol (CCh), muscarnic and nicotinic agonists, induced [Ca2+]i increases in all the VHCs, whereas acetylcholine bromide (ACh-Br), a nicotinic agonist, induced the [Ca2+]i increase in only a small percentage of VHCs. The ACh or CCh-induced Ca2+ response could be partially suppressed by atropine. In the presence of 0.1?mmol/L atropine, the amplitudes of ACh or CCh-induced [Ca2+]i responses became significantly smaller than those in atropine free medium (P<0.01). Conclusions The results suggest the existence of cholinergic receptors in guinea pig VHCs. It is the muscarnic agonists rather than nicontic receptors that dominate [Ca2+]i variation. Atropine can suppress muscarnic agonist-induced Ca2+ responses.

OBJECTIVETo assess the value of combined chromosome karyotype analysis and multiplex ligation probe amplification (MLPA) assay for the prenatal diagnosis of fetuses with abnormalities detected by ultrasonography.

METHODSWith informed consent obtained, 72 pregnant women with ultrasound detected fetal structural abnormalities underwent percutaneous umbilical cord blood sampling. Routine karyotype analysis and MLPA assay were used to detect potential chromosomal deletions and duplications.

RESULTSFive cases were found with an abnormal karyotype. In addition, the MLPA has detected 2 chromosomal microdeletions and 1 microduplication. Together the two methods have yielded a detection rate of 11.11%.

CONCLUSIONFor fetal abnormalities revealed by ultrasonography, combined karyotype analysis and MLPA assay can provide a better option for its efficiency and simplicity.

Adult ; Chromosomes ; genetics ; Female ; Fetus ; abnormalities ; Humans ; Karyotyping ; methods ; Ligation ; methods ; Middle Aged ; Pregnancy ; Prenatal Diagnosis ; methods ; Young Adult

Objective　To investigate the relationship between tinnitus and glucose metabolism in auditory cortex and whether positron emission tomography (PET) can be an objective tool in measuring tinnitus. Methods　Eleven right-handed patients with severe tinnitus and ten right-handed control subjects participated in the 18　 F-FDG/PET study. Analysis with regions of interests was used to calculate asymmetry indices according to the formula: ［(L-R)×100/［(L+R)÷2］］. Results　Glucose metabolism in the auditory cortex of tinnitus patients was asymmetric between the left and right auditory cortices, with that of the left being much higher than that of the right. The asymmetry indices of tinnitus patients was significantly higher than that of the control group (unpaired t test, P<0.001). This revealed that the increased metabolic activity was present in the predominant left hemisphere with a significant focus on the superior and transverse temporal gyri (Brodmann areas 41 and 42, respectively corresponding to primary and secondary auditory cortex), and the results were independent of the subjective localization of the tinnitus sensation. Conclusion　It is suggested that the increased metabolism in the left auditory cortex is related to the tinnitus sensation. PET is capable of providing objective evidence for tinnitus and may be used as a potential tool in measuring tinnitus.

OBJECTIVETo investigate the genetic mechanism of maternal nonsyndromic inherited sensorineural hearing loss(SNHL), to identify the incidence of the 7445(G) mutation in such pedigrees and sporadic patients with SNHL, and to provide the theoretical evidence for the diagnosis of this disease.

METHODSBlood samples were obtained from 2 pedigrees and 14 sporadic patients with SNHL. DNA was extracted from the isolated leukocytes. The mitochondrial DNA (mtDNA) fragments were amplified by PCR. The 1555(G), 3243(G) and 7445(G) mutation was detected by Alw 26 I, Apa I and Xba I restriction endonuclease digestion respectively. The sequence of 12S rRNA, tRNA(Leu(UUR)) and tRNA(Ser(UCN)) was examined.

RESULTSRestriction endonuclease digestion analysis showed that 12 individuals from 2 pedigrees carried homoplasmic 7445(G) mutation, which was of maternal inheritance. Six individuals from 2 pedigrees and 14 sporadic patients did not have 7445(G) mutation. All individuals did not have 1555(G) and 3243(G) mutation. The sequence analysis further showed that none of them carried homoplasmic 1555(G) and 3243(G) mutation, 12 individuals had (nt)7445 A--> G substitution in tRNA(Ser(UCN)) gene.

CONCLUSIONThe incidence of 7445(G) mutation in such pedigrees is higher than that in sporadic patients. Screening for mtDNA 7445(G) mutation combined with 1555(G) examination is of much value to clinical use.

DNA Mutational Analysis ; methods ; DNA, Mitochondrial ; genetics ; Female ; Hearing Loss, Sensorineural ; genetics ; Humans ; Male ; Pedigree ; Point Mutation ; RNA, Transfer, Ser ; genetics

OBJECTIVESTo study the status of cochlear mitochondrial DNA (mtDNA) and to determine the location of mtDNA deletion in aged mice.

METHODSWe detected cochlear mtDNA in 2, 7 - 10 and 17 - 19 month old mice by nested polymerase chain reaction (PCR) and DNA sequencing.

RESULTSmtDNA3867bp deletions were found in the cochleae of aged mice. The deletion occurred within nt9103-nt12970 and were flanked by 15 base pair direct repeats. Comparing the incidence of mtDNA3867bp deletions, 17 - 19 month old mice (7/8) were significantly higher than 7 - 10 month old mice (4/16). The deletion was not observed in 2 month old mice (0/7). The ratio of deleted mtDNA/total mtDNA in 17 - 19 month old mice was higher than in 7 - 10 month old mice (P < 0.001).

CONCLUSIONCochlear mtDNA 3867bp deletion in aged mice may be related to presbycusis.

Aging ; genetics ; Animals ; Base Sequence ; Cochlea ; metabolism ; DNA, Mitochondrial ; analysis ; genetics ; Mice ; Molecular Sequence Data ; Oxidative Phosphorylation ; Presbycusis ; etiology ; Sequence Deletion