Characterization of small RNAs derived from Citrus exocortis viroid (CEVd) in infected tomato plants

In plants, RNA silencing provides an adaptive immune system that inactivates pathogenic nucleic acids, guided by 21-24-mer RNAs of pathogen origin. The characterization of pathogen-related small RNAs (sRNAs) is relevant to uncovering the strategies used by pathogens to evade host defense responses. Several groups have reported the detection of viroid-derived sRNAs during infections, although the origin of these sRNAs and their chemical characteristics were poorly understood. Here, we describe the in vivo cleavage of Citrus exocortis viroid (CEVd) RNA into sRNAs of 21-22 nt, that are phosphorylated at their 5'-end and methylated at 3'. Our studies suggested that the CEVd genomic RNA might be the predominant in vivo substrate for cleavage by Dicer-like enzyme(s), which preferentially targeted residues mainly located within the right-end domain of the viroid. Further analysis on the accumulation levels of specific miRNAs controlling major regulators of leaf development and the miRNA pathway and the levels of their target mRNAs provided evidence that the endogenous tomato miRNA pathway was not affected by CEVd infection.     

中国汉族两个马凡综合征家系FBN1基因的一个新突变

目的 对两个常染色体显性遗传的马凡综合征家系进行基因诊断,并探讨其临床特点。方法 完成家系调查和系谱分析,通过聚合酶链式反应和直接测序的方法对收集到的两个家系中的成员进行原纤维蛋白1(fibrillin 1,FBN1)基因的突变检测。结果 两个家系均呈常染色体显性遗传模式。对两个家系成员进行FBN1基因突变检测发现,在两个家系的患者中发现一个相同的突变位点,即FBN1第27号外显子3463位碱基由G变为A( 3463G＞A),导致原纤维蛋白1第1 155位氨基酸由天冬氨酸变为天冬酰胺(Asp 1155Asn),而两个家系的正常成员及选取的100名健康对照中均未发现该突变位点。结论 先证者均符合Ghent标准诊断为马凡综合征,基因诊断发现两家系中相同的突变位点3463G＞A为中国汉族马凡综合征患者中首次报道。     

IL-7对中国HIV/AIDS患者病程的影响

目的 了解IL-7对中国HIV／AIDS患者病程的影响。方法应用超敏感酶免法对66例中国HIV／AIDS感染者及8例健康对照者血浆IL-7水平进行定量检测，分析其与CD^＋T细胞绝对值、血浆病毒载量及HIV表型的相关性；并且在体外研究rhIL-7对人PBMC中T淋巴细胞增殖及CXCR4表达的影响。结果中国HIV／AIDS患者血浆IL-7水平高于健康对照（P〈0．05），与CD4^＋T细胞绝对值负相关（P〈0．01），与血浆病毒载量正相关（P〈0．05）。rhIL-7可在体外促进T淋巴细胞增殖反应及CXCR4表达。结论中国HIV／AIDS患者血浆IL-7水平升高，且与疾病进展密切相关，可作为疾病进展的相关标志之一。     

Allele dependent silencing of COL1A2 using small interfering RNAs

Osteogenesis imperfecta (OI) is generally caused by a dominant mutation in Collagen I, encoded by the genes COL1A1 and COL1A2. To date there is no satisfactory therapy for OI, but inactivation of the mutant allele through small interfering RNAs (siRNA) is a promising approach, as siRNAs targeting each allele of a polymorphism could be used for allele-specific silencing irrespective of the location of the actual mutations. In this study we examined the allele dependent effects of several tiled siRNAs targeting a region surrounding an exonic COL1A2 T/C polymorphism (rs1800222) in heterozygous primary human bone cells. Relative abundances of COL1A2 alleles were determined by cDNA sequencing and overall COL1A2 abundance was analyzed by quantitative PCR. One of the siRNAs decreased overall COL1A2 abundance by 71% of which 75% was due to silencing of the targeted T-allele. In conclusion, allele-preferential silencing of Collagen type I genes may be a future therapeutic approach for OI.     

基质金属蛋白酶MMP-7及其抑制因子TIMP-1在子宫内膜异位症中的表达及意义

目的 观察基质金属蛋白酶-7(matrix metalloproteinase-7,MMP-7)和基质金属蛋白酶抑制因子-1(tissue inhibitors of matrix metalloproteinase-1,TIMP-1)在子宫内膜异位症(Endmetriosis,EMs)中的表达,探讨二者在EMs发病机制中的作用.方法 采用免疫组化链霉菌抗生物素蛋白-过氧化物酶连接(SP)法检测35例EMs患者的在位内膜,异位内膜及20例时照组为因肌壁间或浆膜下子宫肌瘤在我科行子宫切除术,且月经周期正常的子宫内膜中MMP-7和TIMP-1表达情况,检测结果采用SPSS软件进行统计学分析.结果 MMP-7在EMs在位内膜中的阳性表达率为57.1%,异位内膜中的阳性表达率45.7%,与对照组子宫内膜组织的表达(阳性率为40.0%)相比无显著性差异(P＞0.05).研究组异位内膜中的表达强度明显高于对照组,两者比较,差异有统计学意义(P＜0.05).TIMP-1在两组所有标本中均有阳性表达.TIMP-1在研究组异位内膜中的表达强度低于在位内膜,两者比较,差异有统计学意义(P＜0.05).研究组在位内膜中的表达强度稍低于对照组,两组比较,差异无统计学意义(P＞0.05).结论 MMP-7在EMs的发生过程中起了重要作用.MMP-7在位内膜中表达增强,异位内膜TIMP-1表达减弱,是EMs患者在位内膜、异位内膜细胞具有侵袭能力的原因之一.     

A novel cultured tissue model of rat aorta: VSMC proliferation mechanism in relationship to atherosclerosis

Development of a cultured tissue experimental model of rat aorta was explored in order to study mechanism of vascular smooth muscle (VSMC) proliferation. This particular model has potential with regard to amelioration of atherosclerosis and other vascular diseases in comparison to whole animal and cell culture models. The aorta segments of rats were divided into 4 experimental groups: the injured endothelium, injured endothelium plus BQ123, without injured endothelium and without injured endothelium plus BQ123. Each of group was subdivided into a further 2 subgroups and cultured with 20% serum and with serum-free DMEM. Each group cultured in vitro for 5, 8 and 13 days respectively. The control group was not cultured in vitro. Bromodeoxyuridine (BrDU 8x10(-4) mol/l) was added into the cultured medium of all groups, 24 h prior to harvesting. These segments were fixed in 4% paraformaldehyde for paraffin slice used to HE and immunocytochemical staining and other aorta segments were used to detect the expressions of hypertension-related gene-1 (HRG-1) and smooth muscle 22 alpha (SM22alpha) by RT-PCR. ET-1 content in the supernatant was detected with radioimmunology. Proliferous VSMC can be observed on artery segments cultured in vitro, and conspicuous plaques were developed on model vascular wall cultured for 13 days. Labeled cells increased with an increase in culture time but were not seen in the control group. A greater number of labeled cells were observed in injured endothelium group cultured in 20% serum DMEM. Hyperplasia was inhibited after BQ123 was added into the medium, suggesting that serum and ET-1 are important factors that lead to VSMC proliferation. Expressions of HRG-1 and SM22alpha were decreased while the aorta segments were cultured in vitro, minimum or even absent mRNA expressions of HRG-1 and SM22alpha were detected in injured endothelium cultured in 20% serum DMEM and increased in injured endothelium plus BQ123 group cultured. ET-1 content in the supernatant increased in injured endothelium cultured in 20% serum DMEM. These results show that the phenotypic transform and VSMC proliferation on cultured artery segments were related not only to serum culture, but also to ET-1 secreting. ET-1 and serum may be the main factors of contributing to the proliferation and phenotypic transform. This model provides a favorable experimental platform for research into the mechanism of vascular proliferous diseases as well as its prevention and treatment.     

Eighth case of Li-Campeau syndrome in a Turkish patient caused by a novel pathogenic variant in UBR7 and expanding the phenotype

Li-Campeau syndrome (LICAS) is an autosomal recessive disorder characterized by developmental delay, intellectual disability, genital anomalies, congenital heart defects, and dysmorphic features. LICAS is caused by biallelic pathogenic variants in the UBR7 gene, acting as an E3 ubiquitin-protein ligase. Using exome sequencing (ES), we identified a homozygous novel pathogenic splice site variation c.1185+1G>C in UBR7 in a 32-month-old male from a nonconsanguineous Turkish family with clinical features of LICAS. Sanger sequencing revealed the heterozygous state of parents for this variant and confirmed the co-segregation study. The variant may lead to the loss of function of UBR7 and is in a highly conserved residue. Bioinformatic prediction analysis using in silico algorithms supports the pathogenic effect of the splice site variant in the UBR7.     

A novel presenilin 1 mutation, I202F occurring at a previously predicted pathogenic site causing autosomal dominant Alzheimer's disease

We report a novel presenilin-1 (PSEN1) mutation, I202F occurring in a Welsh kindred with familial Alzheimer's disease. The average age at onset was 53 years. The I202F mutation occurs in alignment with previously reported PSEN1 mutations in the fourth transmembrane domain and confirms that PSEN1 mutations line up along transmembrane alpha-helices.     

Evaluation of direct immunofluorescence test with PCR for detection of novel influenza A (H1N1) virus during 2009 pandemic

During the 2009 novel influenza (H1N1) pandemic, the sensitivity of direct immunofluorescence assay (DFA) for H1N1 infection was 62% (266/429) of that of the polymerase chain reaction (PCR) test. The sensitivity of the DFA differed significantly with the age of patients: the sensitivity was the highest (71.8%) for patients aged <10 years and the lowest for patients aged ≥30 years. The sensitivity of DFA in patients aged ≥30 years was 40.7%. Furthermore, the sensitivity (67.3%, 171/254) of DFA was higher for patients who had a high temperature at admission. An increase in the incidence of H1N1 infection did not influence the sensitivity of DFA (62.1% vs. 62%; p=0.984) test, but resulted in a decrease in the negative predictive value, from 92.4% (700/757) to 69.6% (247/355). PCR may be useful as the initial test for diagnosing H1N1 infection in patients aged ≥30 years with a normal temperature at presentation.     

A novel type X collagen gene mutation (G595R) associated with Schmid-type metaphyseal chondrodysplasia

Metaphyseal chondrodysplasia of the Schmid type (MCDS) is a skeletal dysplasia affecting the long bone metaphyses; it is characterized by short stature, bowlegs, and coxa vara. The spine is generally not involved. Here we report a novel missense mutation of the type X collagen gene in a sporadic case of MCDS. The mutation was a heterozygous single base-pair transition of G-to-A at nucleotide 1783, which predicted a substitution of glycine by arginine at codon 595 (G595R) in the carboxyl-terminal noncollagenous domain. Interestingly, another mutation of the same codon, in which glycine is substituted by glutamic acid (G595E), was previously reported to cause spondylometaphyseal dysplasia (SMD), another group of skeletal dysplasias with involvement of the spine in addition to the long tubular bones. The novel G595R mutation identified in the present study supports the concept of type X collagenopathy. Received: September 10, 1999 / Accepted: October 25, 1999     

Human B7-1 is more efficient than B7-2 in providing co-stimulation for alloantigen-specific T cells

Besides a signal via the T cell receptor/CD3 complex, an additional co-stimulatory signal is required for optimal T cell activation. This signal can be delivered by interaction of either B7-1 or B7-2 expressed by antigen-presenting cells with CD28 on the T cells. Comparison of the function of B7-1 and B7-2 in different experimental animal systems generated conflicting data on the roles for the co-stimulatory molecules. We therefore investigated whether there are differences between B7-1 and B7-2-mediated co-stimulation in an alloantigen-specific primary T cell response induced by B7-transfected human cell lines of epithelial origin. Both transfected keratinocyte cell lines efficiently induce T cell proliferation and the ratios of stimulator versus responder cells are similar. The kinetics of proliferation and interleukin (IL)-2, IL-4 and interferon-γ production are also comparable between both transfectant lines. However, despite equal B7 expression levels, it is consistently found that the magnitude of the B7-1-induced T cell proliferation was higher than that of B7-2. Comparison of precursor frequencies of helper T lymphocytes responsive with either B7-1 or B7-2 revealed that the frequency of B7-1-responsive T cells was higher than that of B7-2, and that the frequency of cells activated by a combination of B7-1 and B7-2 did not differ significantly from that of B7-1 alone. We therefore conclude that the B7-2-responsive T cells are part of the B7-1-responsive population, and that B7-1 on keratinocytes is more efficient in providing co-stimulation for alloantigen-specific T cells.