青蒿琥酯皮肤擦剂在小鼠和兔体内的药代动力学研究

将青蒿琥酯溶于苯二甲酸二甲酯,加适量氨酮制成皮肤擦剂。给兔脱毛后,皮肤涂抹此擦剂25mg/kg后,血药浓度达峰时间平均为2 h,峰浓度平均为1.80μg/ml。药物在兔体内平均驻留时间为3.54 h,清除半衰期约为2.46 h。给小鼠脱毛皮肤涂抹擦剂6.7,31.3和71.4 mg/kg,血药浓度在给药后0.5～4 h达高峰,峰浓度分别为0.82,2.05和7.11μg/ml,体内药物平均驻留时间为3.39,2.79及3.54 h,清除半衰期为2.35,1.93及2.45 h。可见,给兔及小鼠皮肤擦剂后,青蒿琥酯吸收良好,血药浓度维持时间较长。     

Chromosome 11p15.5 regional imprinting: comparative analysis of KIP2 and H19 in human tissues and Wilms' tumors

The imprinted H19 gene is frequently inactivated in Wilms' tumors (WTs) either by chromosome 11p15.5 loss of heterozygosity (LOH) or by hypermethylation of the maternal allele and it is possible that there might be coordinate disruption of imprinting of multiple 11p15.5 genes in these tumors. To test this we have characterized total and allele- specific mRNA expression levels and DNA methylation of the 11p15.5 KIP2 gene in normal human tissues, WTs and embryonal rhabdomyosarcoma (RMS). Both KIP2 alleles are expressed but there is a bias with the maternal allele contributing 70-90% of mRNA. Tumors with LOH show moderate to marked reductions in KIP2 mRNA relative to control tissues and residual mRNA expression is from the imprinted paternal allele. Among WTs without LOH most cases with H19 inactivation also have reduced KIP2 expression and most cases with persistent H19 expression have high levels of KIP2 mRNA. In contrast to the extensive hypermethylation of the imprinted H19 allele, both KIP2 alleles are hypomethylated and WTs with biallelic H19 hypermethylation lack comparable hypermethylation of KIP2 DNA. 5-aza-2'-deoxycytidine (aza-C) increases H19 expression in RD RMS cells but does not activate KIP2 expression. These data indicate coordinately reduced expression of two linked paternally imprinted genes in most WTs and also suggest mechanistic differences in the maintenance of imprinting at these two loci.      

A novel mis-sense mutation (G1381A) in the G6PD gene identified in a Chinese man

Objective　To detect new mutations among 29 glucose-6-phosphate dehydrogenase (G6PD) deficient individuals from Yunnan province. Methods　The nitroblue tetrazolium (NBT) method was used to screen G6PD deficient individuals. Mutation was identified by single strand conformation polymorphism (SSCP), amplification created restriction site (ACRS), amplification refractory mutation system (ARMS) and DNA sequencing. Results　Among 29 cases, 18 cases of G1388A, 1 case of C1004A, and 1 case of G1381A were identified. Nine cases remained to be defined. The G1381A mutation is a novel mis-sense mutation, with a substitution of threonine for alanine (A461T). The resultant G6PD had reduced enzymatic activity. In addition, G1381A caused a restriction site of Stu I to disappear, providing a rapid method for the detection of this mutation. Conclusion　A novel mis-sense mutation G1381A was found. This mutation results in a substitution of threonine for alanine, producing enzyme with reduced activity. The loss of the Stu I restriction site offers a rapid method for the detection of this mutation.     

Associations between both genetic and environmental biomarkers and lung cancer: evidence of a greater risk of lung cancer in women smokers

This molecular epidemiologic case-control study of lung cancer incorporated three complementary biomarkers: the glutathione S- transferase M1 (GSTM1) null genotype, a potential marker of susceptibility, and polycyclic aromatic hydrocarbon-DNA adducts (PAH- DNA) and sister chromatid exchanges (SCE), both indicators of environmentally induced genetic damage. Associations between biomarkers and lung cancer were investigated, as were possible gene-environment interactions between the GSTM1 null genotype and tobacco smoke exposure. Subjects included 136 primary non-small cell lung cancer surgical patients and 115 controls at the Columbia Presbyterian Medical Center. Questionnaire and Tumor Registry data, pre-treatment blood samples and biomarker measurements on blood were obtained. Overall, GSTM1 null genotype was significantly associated with lung cancer [odds ratio (OR) = 2.04, 95% confidence interval (CI) = 1.13-3.68]. ORs for GSTM1 and lung cancer were significant in females (2.50, 1.09-5.72) and smokers (2.25, 1.11-4.54) and not significant in males (1.4, 0.58-3.38) and non-smokers (0.88, 0.18-4.33). However, ORs for males versus females and smokers versus non-smokers did not differ significantly. The OR for GSTM1 and lung cancer in female smokers was 3.03 (1.09- 8.40), compared with 1.42 (0.53-4.06) in male smokers. In contrast to PAH-DNA adducts in leukocytes, SCE did not differ between cases and controls. Neither biomarker differed significantly between the two GSTM1 genotypes. The combined effect of elevated PAH-DNA adducts and GSTM1 genotype on case-control status (16.19, 1.2-115) appeared multiplicative. Results suggest that the effect of the GSTM1 null genotype is greatest in female smokers, which is consistent with other evidence that indicates that women are at higher risk of lung cancer than males, given equal smoking. Persons with both the GSTM1 deletion and elevated PAH-DNA adducts may represent a sensitive subpopulation with respect to carcinogens in tobacco smoke and other environmental media.      

Expression and characterization of Gag protein of HIV-1（CN） in Pichia pastoris

To express the core protein of HIV-1 of Chinese prevalent strain （HIV-1 （CN）） in Pichia pastoris, the fulllength gag gene was inserted into the secretory expression vector pHILS1. Linearized recombinant plasmid pHILGAG by Sail was electrotransformed into the yeast strain GS115, and the yeast transformants were identified by PCR. To induce the interest protein to be expressed, the PCR positive transformants were inoculated in the medium of BMGY and BMMY, mRNA of the strain was detected by RT-PCR, and the expressed protein was analyzed by SDS-PAGE, Western blotting and thin layer scanning. mRNA （1.3 kb） was amplified by RT-PCR. SDS-PAGE and Western blotting analysis showed that the molecular mass of the expressed protein was 55 kD, which was similar to the expected value, and the expressed protein could react with McAb to HIV-1 p24. Thin layer scanning analysis demonstrated that the whole amount of the expressed protein was approximately 13 % of the soluble protein in the supernatant. The recombinant yeast had good genetic stability. The optimal expression conditions of the engineering yeast were as follows： BMMY medium, 80-90% of dissolved oxygen, 1% methanol, and 3-day-cultivation course. Gag proteins were expressed under the optimal expression condition and purified via gel filtration chromatography. The purity of the interest protein was up to 85 %. After the purified proteins were inoculated into BALB/c mice, the anti-HIV-1 antibodies in the immunized mice could be detected by Western blotting.     

Factor V G1691A (Leiden) and prothrombin G20210A single-nucleotide polymorphisms in type 2 diabetes mellitus

The association of the single nucleotide polymorphisms (SNPs) G1691A in coagulation factor V (FV)-Leiden and G20210A in prothrombin (PRT) genes with type 2 diabetes mellitus (T2DM) were analyzed in 112 T2DM patients (58 males, 54 females; mean age 55.24 +/- 13.5 years) and 249 healthy control subjects (118 males, 131 females; mean age 53.03 +/- 13.8 years). No association was found for FV-Leiden with T2DM, as the frequency of the G/G (82.1% vs. 85.5%), G/A (17.0% vs. 14.1%), and A/A (0.9% vs. 0.4%) genotypes was not different between patients and controls, respectively (P = 0.644). Similarly, lack of association of PRT G20210A with T2DM was seen among the population studied, and the frequency of the G/G (92.9% vs. 97.2%), G/A (6.3% vs. 2.8%), and A/A (0.9% vs. 0.0%) genotypes was similar among patients and controls, respectively (P = 0.094). Neither FV-Leiden nor PRT G20210A was associated with, and no evidence for interactions between these mutations was seen in, T2DM.     

Structure and strength of the distal radius in female patients with rheumatoid arthritis: A case‐control study

The purpose of this work was to investigate the volumetric bone mineral density (vBMD), bone microstructure, and mechanical indices of the distal radius in female patients with rheumatoid arthritis (RA). We report a cross‐sectional study of 66 middle‐aged female RA patients and 66 age‐matched healthy females. Areal BMD (aBMD) of the hip, lumbar spine, and distal radius was measured by dual‐energy X‐ray absorptiometry (DXA). High‐resolution peripheral quantitative computed tomography (HR‐pQCT) was performed at the distal radius, yielding vBMD, bone microstructure, and mechanical indices. Cortical and trabecular vBMD were 3.5% and 10.7% lower, respectively, in RA patients than controls, despite comparable aBMD. Trabecular microstructural indices were –5.7% to –23.1% inferior, respectively, in RA patients compared to controls, with significant differences in trabecular bone volume fraction, separation, inhomogeneity, and structural model index. Cortical porosity volume and percentage were 128% and 93% higher, respectively, in RA patients, with stress being distributed more unevenly. Fourteen RA patients had exaggerated periosteal bone apposition primarily affecting the ulnovolar aspect of the distal radius. These particular patients were more likely to have chronic and severe disease and coexisting wrist deformity. The majority of the differences in density and microstructure between RA patients and controls did not depend on menstrual status. Recent exposure to glucocorticoids did not significantly affect bone density and microstructure. HR‐pQCT provides new insight into inflammation‐associated bone fragility in RA. It detects differences in vBMD, bone microstructure, and mechanical indices that are not captured by DXA. At the distal radius, deterioration in density and microstructure in RA patients involved both cortical and trabecular compartments. Excessive bone resorption appears to affect cortical more than trabecular bone at distal radius, particularly manifested as increased cortical porosity. Ulnovolar periosteal apposition of the distal radius is a feature of chronic, severe RA with wrist deformity. © 2013 American Society for Bone and Mineral Research.