Exploring Mycobacterium tuberculosis infection-induced alterations in gene expression in macrophage by microarray hybridization

Tuberculosis remains a major global threat to human health, with 8 million cases of clinical tuberculosis and 3 million deaths annually[1] (www.stoptb.org/tuberculosis/#facts.html). Mycobacterium tuberculosis (Mtb) is the causative agent of tuberculosis. The emergence of multi-drug resistant strains of Mtb and co-infection with the human immunodeficiency virus (HIV) have further emphasized the need for effective prevention and treatment of tuberculosis. Mtb is a facultative intracellular pat…     

内蒙古东部地区狼的食性初步研究

本文采用粪便分析法和食痕分析法对内蒙古东部狼的食性进行初步研究。结果表明, 该地区狼的主要食物是大、中型有蹄类（出现频率53.8%, 相对生物量85.17%）, 粪便中家畜出现频率为40.3%, 相对生物量为75.67%。该地区狼的食性同黑龙江狼的食性相比, 存在地区差异。通过饲喂实验, 提出狼粪便中所含猎物的干重与鲜重间的线性回归方程为Y=2.76+0.02x（r=0.99）, 应用此方程可估计出狼消耗各种猎物的相对生物量和数量。在该地区狼对家畜危害比较严重, 要适当猎取同时也应保持其适当数量。     

Selection of Reference Genes for Gene Expression Studies Related to Intramuscular Fat Deposition in Capra hircus Skeletal Muscle

The identification of suitable reference genes is critical for obtaining reliable results from gene expression studies using quantitative real-time PCR (qPCR) because the expression of reference genes may vary considerably under different experimental conditions. In most cases, however, commonly used reference genes are employed in data normalization without proper validation, which may lead to incorrect data interpretation. Here, we aim to select a set of optimal reference genes for the accurate normalization of gene expression associated with intramuscular fat (IMF) deposition during development. In the present study, eight reference genes (PPIB, HMBS, RPLP0, B2M, YWHAZ, 18S, GAPDH and ACTB) were evaluated by three different algorithms (geNorm, NormFinder and BestKeeper) in two types of muscle tissues (longissimus dorsi muscle and biceps femoris muscle) across different developmental stages. All three algorithms gave similar results. PPIB and HMBS were identified as the most stable reference genes, while the commonly used reference genes 18S and GAPDH were the most variably expressed, with expression varying dramatically across different developmental stages. Furthermore, to reveal the crucial role of appropriate reference genes in obtaining a reliable result, analysis of PPARG expression was performed by normalization to the most and the least stable reference genes. The relative expression levels of PPARG normalized to the most stable reference genes greatly differed from those normalized to the least stable one. Therefore, evaluation of reference genes must be performed for a given experimental condition before the reference genes are used. PPIB and HMBS are the optimal reference genes for analysis of gene expression associated with IMF deposition in skeletal muscle during development.     

Structural basis for targeting the ribosomal protein S1 of Mycobacterium tuberculosis by pyrazinamide

Pyrazinamide (PZA) is a first‐line drug for tuberculosis (TB) treatment and is responsible for shortening the duration of TB therapy. The mode of action of PZA remains elusive. RpsA, the ribosomal protein S1 of Mycobacterium tuberculosis (Mtb), was recently identified as a target of PZA based on its binding activity to pyrazinoic acid (POA), the active form of PZA. POA binding to RpsA led to the inhibition of trans‐translation. However, the nature of the RpsA–POA interaction remains unknown. Key questions include why POA exhibits an exquisite specificity to RpsA of Mtb and how RpsA mutations confer PZA resistance. Here, we report the crystal structures of the C‐terminal domain of RpsA of Mtb and its complex with POA, as well as the corresponding domains of two RpsA variants that are associated with PZA resistance. Structural analysis reveals that POA binds to RpsA through hydrogen bonds and hydrophobic interactions, mediated mainly by residues (Lys303, Phe307, Phe310 and Arg357) that are essential for tmRNA binding. Conformational changes induced by mutation or sequence variation at the C‐terminus of RpsA abolish the POA binding activity. Our findings provide insights into the mode of action of PZA and molecular basis of PZA resistance associated with RpsA mutations.     

The complete mitochondrial genome and phylogenetic analysis of Nyctereutes procyonoides

The complete mitochondrial genome sequence of the raccoon dog (Nyctereutes procyonoides) was determined by using the long and accurate polymerase chain reaction. The entire mitochondrial genome sequence is 16,713 bp in length contains two ribosomal RNA genes, 13 protein-coding genes, 22 transfer RNA genes and 1 control region. Most mitochondrial genes are encoded on the H strand, except for the ND6 gene and 8 tRNA genes. The base compositions of mitochondrial genomes present clearly A–T skew. All the transfer RNA genes can be folded into the typical cloverleaf-shaped structure except tRNA-Ser (AGY), which lacks the dihydrouridine arm. Protein-coding genes mainly initiate with ATG and terminate with TAA. Some reading frame intervals and overlaps are found in the mitochondrial genome. The control region can be divided into three domains: the extended termination associated sequences (ETASs) domain, the central conserved domain and the conserved sequence blocks (CSBs) domain. Three conserved sequence blocks (CSBs) and one extended termination associated sequences (ETAS-1) is found in the control region. The phylogenetic analysis based on the concatenated data set of 14 genes in the mitochondrial genome of Canidae shows that the raccoon dog has close phylogenetic position with the red fox (Vulpes vulpes) and they constitute a clade which has an equil evolutionary position with the clade formed by the genera Canis and Cuon.     

健胃益气汤对大鼠胃粘膜急性损伤的细胞保护作用

目的建立无水乙醇致大鼠胃粘膜损伤试验模型,研究预先给予健胃益气汤(GWYQT)对急性胃粘膜损伤的细胞保护作用。为临床推广应用该方治疗胃粘膜损伤提供科学理论和实验依据。方法①动物分组、建立动物模型。②取大鼠胃腺区组织制作切片并作HE及PAS染色,光学显微镜观察胃粘膜组织的病理变化。③用免疫组织化学ABC法对大鼠胃粘膜组织细胞凋亡相关蛋白Bcl-2、Bax等进行检测并通过图像分析对以上结果做定性、定位、定量分析及统计学处理。结果健胃益气汤预防用药能维护胃粘膜粘液含量及厚度,降低胃粘膜组织细胞Bax蛋白表达,增加Bcl-2蛋白表达,实验对照组与中药组比较,有显著性差异(P<0.01)。结论给予中药健胃益气汤可有效保护胃粘膜,减轻胃粘膜细胞损伤,抑制细胞凋亡。     

Trehalose-6-phosphate Phosphatase from Mycobacterium tuberculosis induces humoral and cellular immune responses

Trehalose-6-phosphate phosphatase is an enzyme strictly essential for the growth of mycobacteria. Subcellular fractionation of Mycobacterium tuberculosis and M. bovis bacillus Calmette-Guérin (BCG) located the trehalose-6-phosphate phosphatase in the cell wall and membrane fractions. Trehalose-6-phosphate phosphatase induced an increased Th1-type immune response in mice, characterized by an elevated level of interferon-gamma in antigen-stimulated splenocyte culture and a strong IgG2a antibody response. The trehalose-6-phosphate phosphatase was recognized by the sera of tuberculosis patients and BCG-vaccinated donors. The mycobacterial trehalose-6-phosphate phosphatase is an immunodominant antigen, and it may be a candidate for vaccine development for the control of tuberculosis.     

Recombinant BCG coexpressing Ag85B, ESAT-6 and mouse-IFN-gamma confers effective protection against Mycobacterium tuberculosis in C57BL/6 mice

In this study, the protective efficacy of a novel recombinant bacille Calmette Géurin (BCG) strain (rBCG-AEI) expressing fusion protein the antigen 85B (Ag85B)- the 6-kDa early secreted antigen target (ESAT-6)-IFN-gamma against Mycobacterium tuberculosis H37Rv in mice was evaluated. The immunogenicity study showed that rBCG-AEI could induce higher specific antibody titers and significantly increase cellular immune response than BCG, or rBCG-A strain (expressing Ag85B), or rBCG-AE strain (expressing fusion protein Ag85B-ESAT-6). The protective experiment demonstrated that rBCG-AEI could confer similar or even better protective efficacy against M. tuberculosis infection compared with others in organ bacterial loads, lung histopathology and net weight gain or loss. The results suggested that rBCG-AEI is a potential candidate for further study.     

Chemical properties and immunostimulatory activity of a water-soluble polysaccharide from the clam of Hyriopsis cumingii Lea

A water-soluble polysaccharide HCLPS-1 was isolated from clam of Hyriopsis cumingii Lea by hot water, DEAE-cellulose and Sephadex-G200, gel-permeation chromatography and tested for its immunostimulatory activities. Its structural characteristics were investigated by UV, IR, HPLC, NMR spectroscopy, methylation analysis and GLC–MS. HCLPS-1 consisted of glucose and xylose with the molar ratio of 35:1. Based on the data obtained, HCLPS-1 was determined to has a main chain of (1 → 4)-linked β-d-glucopyranosyl residues, with a weight-average molecular weight of about 1.56 × 10⁵ Da. HCLPS-1 not only could significantly promote concanavalin A (Con A), lipopolysaccharide (LPS)-stimulated splenocytes proliferation in concentration-dependent manner in vitro, but also increase Con A- and LPS-induced splenocytes proliferation in mice immunized with the sheep red blood cell (SRBC). Meanwhile, HCLPS-1 remarkably promoted 2, 4-dinitrofluorobenzene (DNFB)-induced delayed-type hypersensitivity (DTH) reactions. These results suggested that HCLPS-1 could improve both specific and non-specific cellular immune response and might be explored as a potential natural immunomodulator.