Objective　The microvasculature of the anterior lobe, posterior lobe and cerebellar vermis was observed under the optical microscope in 6 rats. Methods　The tannic acid-ferric chloride method (TAFM) was used to stain the vessels in cerebellum. Results　The arteries within the cortex and medulla of the cerebellum originated from pia mater arteries and central branches of the cerebellar arteries, the branches of pia mater arteries mostly penetrated into the cerebellar cortex vertically and formed capillary net in the molecular layer, purkinje layer and granular layer, the arteriole in the medulla of cerebellar posterior lobe branched and formed clawed-like figurations. These vessels beneath lobule cortex connected mutually by vessel anatomosis across the medulla. Conclusion　The arteriole in the cerebellum can be manifested distinctly in a strong three-dimension by TAFM and the vessel anatomosis in deep layer of cerebellum can be easily observed.

Objective To study problems related to drug-induced liver lesion caused by antituberculotic therapy (DLL).Methods Totally, 172 cases of DLL occurred in 1 464 patients with pulmonary tuberculosis after antituberculotic therapy, hospitalized during January 1995 to December 2002, were reviewed and analyzed retrospectively.Results Patients aged 20 to 60 years with DLL by antituberculolis therapy accounted for 70.3% of the total. Symptoms of digestive tract and change in liver function usually occurred within 8 weeks of intensive treatment (73.8%), and discontinuation of autituberculotic drugs was not needed for the mild cases, but needed for the severe cases with liver protective therapy. In total, 157 of the 172 cases (91.3%) recovered completely and 13 case improved (7.6%), two cases deteriorated and discharged, and doses of autituberculotic drugs should be reduced or stopped in 41 cases (23.8%) affecting their treatment efficacy.Conclusions DLL was liable to occur in patients of pulmonary tuberculosis with antituberculotic therapy, especially in the elderly men with body weight less than 50 kilograms, those with previous liver damage or infected with hepatotropic virus, alcohol drinking, complicated with extrapulmonary tuberculosis, with combination of isoniazid rifampicin and pyrazinamide.

BACKGROUND: Radix astragali has the effect of protecting cells from damage in ischemic reperfusion, whether pre-treatment with radix astragali can protect myocardial eells from apoptosis in ischemic reperfusion ? OBJECTIVE: To investigate the effect of pre-treatment with radix astragali on apoptosis and its relative genes in rats with ischemic myocardial reperfusion DESIGN: A randomized and controlled trial taking Wistar rats as experimental subjects.SETTING: The Basic Medical Department of Chengde Medical College and the Geriatric Department of the Affiliated Hospital.MATERIALS: The experiment was completed in the Imunnohistochemical Laboratory of Basic Medical Institute in Chengde Medical College from February to December in 2004. A total of 30 healthy male Wistar rats were selected, and at random classified as groups of radix astragali pre-treated (radix astragali), ischemic reperfusion and psuedo-operated (control), 10 rats for each group.METHODS: Radix astragali injection was given peritonealy for rats in radix astragali pre-treated group before operation, and the equivalent normai saline was given for those in ischemic reperfusion and psuedo-operated groups. One week later, the model of ischemic reperfusion was set up. After operation the myocardia in marginal zone of ischemic reperfusion were sampled, and the myocardia of the corresponding zone were taken for control group. The method of terminal (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) was used for assay of myocardial apoptosis rate, and the ABC immunohistochemical method was used for assay of myocardial bcl-2 (inhibiting apoptosis gene) and bax (promoting apoptosis gene).MAIN OUTCOME MEASURES: Apoptosis rates, and expression of bcl2 and bax genes of myocardia RESULTS: ① Apoptosis rate of myocardial cells: The rate in radix astragali group was decreased compared with that in ischemic reperfusion group [ (14.06 ±9.97) ％, (19.34±12.30) ％, t = 1.863, P ＜ 0.05].② Expression of bcl-2: There was no significant difference between radix astragali and ischemic reperfusion groups[(9.14±4.46) ％, (8.99±4.54) ％, P ＜ 0.05].③ Expression of bax: The expression in radix astragali group was decreased compared with that in ischemic reperfusion group [(12.65 ±7.23)％,(18.12±7.92) ％, t = 2.096, P ＜ 0.05]CONCLUSION: Pre-treatment with radix astragali can down-regulate the expression of promoting apoptosis gene so as to reduce the rate of myocardial cell apoptosis, hence it can protect the myocardial cells in ischemic reperfusion.

Objective To investigate the effects of 3-N-butylphthalide ( NBP ) pretreatment on the score of neurological deficit , oxidative stress and pathomorphology in rats with cerebral ischemia reperfusion injury ( CIRI ) . Methods Ninety male SD rats were randomly divided into sham operation group ( Sham group ) , model group ( IR group), NBP pretreatment low dose group (NBPⅠgroup), NBP pretreatment middle dose group(NBPⅡgroup) and NBP pretreatment high dose group(NBPⅢgroup), 18 rats per group.Pretreatment was given once a day within 1 week before establishing the model of cerebral ischemia reperfusion injury .The model of middle cerebral artery occlusion ( MCAO) was subjected by suture method .The score of neurological deficit was executed after ischemia for 2h and reperfusion for 24h in all the rats.The cerebral infarction was observed by TTC staining .The pathologic change of brain was observed by HE staining under the microscope .Hydroxylamine method was used to detect activity of SOD , chemical colorimetry method was used to measure activity of GSH-PX, and TBA method was used to detect content of MDA .Results (1) In Sham group, the score of neurological deficit and the percentage of infarction volume were zero , the morphology of nerve cell was regular , and activity of SOD, GSH-PX and content of MDA of brain tissue were normal .(2) Compared with IR group , the score of neurological deficit was significantly reduced in NBP pretreatment groups (all P<0.01); the score of neurological deficit was decreased progressively in turn in NBP Ⅰ,Ⅱ,Ⅲgroup (all P<0.05).(3) Compared with IR group, the percentage of infarction volume was cut down progressively in turn in NBPⅠ,Ⅱ,Ⅲgroup (all P<0.05), and neuron injury was also induced obviously in NBP pretreatment groups .(4) Activity of SOD, GSH-PX was largely increased , and content of MDA was greatly decreased in NBP pretreatment groups ( P<0.01 ) .Activity of SOD , GSH-PX went up progressively in turn , and contents of MDA were cut down progressively in turn in NBP Ⅰ,Ⅱ,Ⅲgroup ( all P<0.05 ) .Conclusion 3-N-butylphthalide can significantly up-regulate the activity of SOD and GSH-PX, decrease the content of MDA , reduce the percentage of infarction volume , and relieve the damage of nerve cell to preventively protect the rats with cerebral ischemia reperfusion injury .

Objective To observe microvascular architecture and free radical metabolism in hippocampus after focal cerebral ischemia/reperfusion and to explore the effect of NBP (3-n-butylphthalide). Methods Fifty-four SD rats were ran?domly divided into NBP pretreatment group, ischemia/reperfusion group and sham operation group (n=18 in each group). The model of middle cerebral artery occlusion(MCAO)was established by suture method. The neurological scores were counted and the volume of infarction was measured;TA-Fe method was applied to observe the microvascular architecture of hippo?campus, Mivnt image analysis system was used to analyze the microvessel density(MVD)and the microvessel area density (MVA)of hippocampus quantitatively;The activity of SOD and content of MDA were measured by colorimetric method. Re?sults Compared to the ischemia reperfusion(IR)group, the neurological scores and the volume of infarction were decreased sharply in NBP group. What′s more, the activity of SOD, MVD and MVA were all enhanced but the content of MDA and the count of closed microvessels were both reduced(P < 0.01). Conclusion NBP can improve microvascular architecture of hippocampus and reduce the free radical injury. There is a protective effect on hippocampus of rats who suffered focal cere?bral ischemia reperfusion.

OBJECTIVE: To detect potential mutation in a pedigree affected with autosomal recessive non-syndromic deafness. METHODS: Mutation analysis was carried out by next generation sequencing, and suspected mutations were verified by Sanger sequencing. RESULTS: A heterozygous c.235delC mutation of the GJB2 gene, together with compound heterozygous mutations of the OTOF gene [c.1194T>A (p.D398E) and c.2180A>G (p.N727S)] were detected in the proband. The sister of the proband (also had hearing loss) has carried a heterozygous c.235delC mutation in the GJB2 gene, in addition with a heterozygous c.2180A>G(p.N727S) mutation of the OTOF gene. By Sanger sequencing, a heterozygous IVS1+2T>A mutation was further detected in the non-coding region of the GJB2 gene in both sisters. CONCLUSION The compound heterozygous c.235delC and IVS1+2T>A mutations of the GJB2 gene probably account for the hearing loss in the two sisters, among which IVS1+2T>A is considered as a novel pathogenic mutation of the GJB2 gene.
Connexins ; genetics ; DNA Mutational Analysis ; Female ; Hearing Loss, Sensorineural ; genetics ; High-Throughput Nucleotide Sequencing ; Humans ; Membrane Proteins ; genetics ; Mutation ; Pedigree

Objective:To preliminarily investigate the mechanism of MMP-9 blocking CD73 detachment from RPE cells surface and preventing and treating experimental autoimmune pigment membranitis(EAU).Methods:RPE cells isolated from wild-type C57BL/6 and CD73 gene knockout (CD73 -/-) mice were cultured in vitro, and treated with lipopolysaccharide and TNF-α to induce CD73 detachment from RPE surface. According to whether MMP-9 inhibitor CTK8G1150 was added at the same time (the final concentration was 5.0 mol/L) or not, RPE cells cultured in the two types of mice were respectively set as MMP-9 inhibitor intervention group and non-intervention control group. The cells in each group were treated with the intervention of a solvent, 1 μmol/Ladenosine monophosphate (AMP), 1 μmol/L AMP, and 3 μmol/L 5' -α,β-methylene adenosine diphosphate (APCP) (AMP+APCP). The stimulating effect of RPE cells in different groups on CD4 + T cell proliferation was detected by tritiated thymidine incorporation. Adoptive immune induced EAU in wild-type B6 mice and CD73 -/- mice, respectively. The receptor mice were randomly divided into the MMP-9 inhibitor intervention group and the non-intervention control group, and CTK8G1150 or the solvent were injected into the subretinal cavity 4, 7 and 10 days after adoptive immunity. CD73 mRNA and protein expression in RPE cells of recipient mice were detected by real-time quantitative PCR (RT-PCR) and Western blot. One-way ANOVA was used to analyze all experimental data. Results:When the stimulation mode was AMP, the proliferation of CD4 + T cells in the C57BL/6 MMP-9 inhibitor intervention group decreased significantly compared with the nonintervention group ( F=13.28, P<0.01). When the stimulation mode was solvent and AMP+APCP, there was no statistically significant difference in the proliferation capacity of CD4 + T cells between the two groups ( F=7.78, 6.58; P>0.05). There was no statistically significant difference in the proliferation capacity of CD4 + T cells between the CD73 -/- MMP-9 inhibitor intervention group and the non-intervention group ( F=5.24, 6.12, 7.04; P>0.05). RT-PCR results showed that there was no statistically significant difference in the relative expression of CD73 mRNA in RPE cells between the MMP-9 inhibitor group and the non-intervention control group( F=6.54, P>0.05). Western blot results showed that the expression of CD73 protein in RPE cells in the MMP-9 inhibitor group of B6 receptor mice was significantly increased compared with the control group ( F=15.24, P<0.01). Conclusion:MMP-9 inhibitor blocks CD73 detachment from RPE cells surface and has a protective effect on EAU.

OBJECTIVETo detect potential mutations of COL1A1 and COL1A2 genes in four Chinese pedigrees affected with osteogenesis imperfecta (OI) and provide prenatal diagnosis for a fetus at 18th gestational week.

METHODSAll coding regions and exon/intron boundaries of the COL1A1 and COL1A2 genes were analyzed with targeted next-generation sequencing (NGS). Suspected mutations were confirmed with Sanger sequencing in the probands, unaffected relatives and 200 unrelated healthy individuals. Prenatal diagnosis for a high-risk fetus was carried out through Sanger sequencing.

RESULTSThe probands of families 1 and 2 have respectively carried a c.760G>A (p.Gly254Arg) and a c.608G>T (p.Gly203Val) mutation of the COL1A1 gene. For family 3, the proband and his daughter have carried a novel c.299-1G>C splicing mutation of the COL1A1 gene. The same mutation was not found in the fetus of this family. For family 4, the proband has carried a novel c.1990G>C (p.Gly664Arg) mutation of the COL1A2 gene. The four mutations were not found in the unaffected relatives and 200 unrelated healthy individuals.

CONCLUSIONThe mutations of the COL1A1 and COL1A2 genes probably underlie the disease in the four families. NGS combined with Sanger sequencing can provide an effective and accurate method for their genetic and prenatal diagnosis.

Adult ; Child, Preschool ; Collagen Type I ; genetics ; DNA Mutational Analysis ; Female ; High-Throughput Nucleotide Sequencing ; Humans ; Infant, Newborn ; Male ; Mutation ; Osteogenesis Imperfecta ; genetics ; Prenatal Diagnosis

Objective: To carry out genetic testing for a family with two pregnancies affected with hydrops fetalis and dilated cardiomyopathy (DCM) of the fetus. Methods: DNA was extracted from fetal tissue as well as peripheral blood samples from the couple. Single nucleotide polymorphism array (SNP array) and next-generation sequencing (NGS) were carried out to screen potential mutation. Suspected mutation was validated with PCR and Sanger sequencing. Results: The manifestation of fetal echocardiography was consistent with DCM. No obvious abnormality was found by SNP array analysis. A hemizygous c. 481G>A (p.G161R) mutation of the TAZ gene was detected in the male fetus by NGS and confirmed by Sanger sequencing. The mutation was inherited from his mother. Conclusion Barth syndrome due to the c. 481G>A mutation of the TAZ gene probably underlies the recurrent hydrops fetalis and fetal DCM in this family.

Objective: To analyze variants of RUNX2 gene in two pedigrees affected with cleidocranial dysplasia and provide prenatal diagnosis for them. Methods: For the two probands, the coding sequences of the RUNX2 gene were analyzed with PCR and bidirectional Sanger sequencing. To verify the results, peripheral blood samples were collected from their parents and 100 healthy controls. For family 1, umbilical cord blood was also collected for prenatal genetic diagnosis. Results: In family 1, the proband and the fetus both carried a heterozygous c. 578G>C (p.Arg193Pro) mutation. For family 2, the proband was found to carry a heterozygous c. 909C>A (p.Tyr303X) mutation. The same mutations were not found among their parents and 100 healthy controls. Neither mutation was reported previously. Conclusion Variants of the RUNX2 gene probably underlie the cleidocranial dysplasia in both pedigrees. The results enabled prenatal diagnosis for the affected family.