成人骨髓间充质干细胞的体外培养及生物学特性

目的:观察成人骨髓间充质干细胞体外分离培养方法及其生物学特性,为骨髓间充质干细胞的临床应用奠定技术基础。方法:实验于2005-12在吉林省四平市中心医院中心实验室进行。实验材料:人骨髓间充质干细胞,Ficoll淋巴细胞分离液,胎牛血清,牛血清白蛋白,α-MEM培养基,胰蛋白酶,FITC标记的CD44和CD90。实验方法:①人骨髓间充质干细胞的分离与培养:采集成人骨髓,密度梯度法分离单个核细胞,条件培养基培养和扩增,倒置光学显微镜下观察细胞形态及生长情况。②人骨髓间充质干细胞生长曲线的测定:取生长良好的P1,P5,P10细胞,用胰酶消化,接种于24孔板培养,每天各取3孔消化计数细胞,取平均值,连续培养7d,绘制生长曲线。③人骨髓间充质干细胞表面标志的测定:取生长良好P5细胞进行免疫荧光染色鉴定(CD44,CD90并设同型对照)。④人骨髓间充质干细胞的冻存及复苏:收集生长良好细胞,计数细胞密度及冻前存活率。将细胞加入到细胞冻存液中液氮冷冻保存。30d后复苏细胞,每日倒置光学显微镜下观察细胞生长状况。结果:①倒置显微镜下观察细胞形态:倒置显微镜下观察细胞呈纤维形,漩涡状生长。②人骨髓间充质干细胞的生长曲线分析:传代的细胞生长较原代要快,从第10代开始细胞生长开始出现缓慢征象。③细胞表面标记检测结果:免疫荧光染色后细胞膜着色明显,CD44,CD90阳性率均大于90%。④细胞冻存及复苏生长特性分析:椎虫蓝计数细胞存活率90%以上,增殖能力强,和未冻存过的传代细胞具有同样的生长特性。结论:应用密度梯度法分离法能获得较高纯度的骨髓间充质干细胞,能在体外长期培养,在传代培养5代内生长旺盛且生物学特性稳定。     

Extruded erythroblast nuclei are bound and phagocytosed by a novel macrophage receptor

Resident bone marrow macrophages (RBMM) play an important role in clearance of nuclei extruded from late-stage erythroblasts (Eb). To investigate the nature of macrophage receptors involved in this process, extruded erythroblast nuclei (EEN) were purified by cultivation of erythroblasts with erythropoietin, followed by density gradient centrifugation. By electron microscopy, the majority of free nuclei had an intact plasma membrane. EEN bound avidly to RBMM in a divalent cation-independent manner at both 4 degrees C and 37 degrees C. The use of specific monoclonal antibodies (MoAbs) and inhibitors showed that this adhesive interaction was not mediated by previously characterized macrophage receptors involved in recognition of either developing hematopoietic cells or apoptotic cells. The EEN receptor was expressed on resident macrophages isolated from murine bone marrow, spleen, lymph node, and peritoneal cavity, but was completely absent from alveolar macrophages. Despite high levels of EEN binding to RBMM, few were phagocytosed even after prolonged culture. Phorbol myristate acetate (PMA) was found to stimulate phagocytosis, suggesting that this is a regulated process. These findings indicate that EEN are recognized by a novel macrophage receptor and that recognition may be triggered during the membrane remodeling that accompanies enucleation.     

Neuroeconomics for the Study of Social Cognition in Adolescent Depression

Traditional social‐cognitive approaches for investigating interpersonal problems in adolescent depression are limited. An important functional domain studied in adolescent depression is reward, but experimental paradigms have largely been nonsocial. In this article, we propose the methods and concepts of neuroeconomics may address this gap. We begin by discussing a well‐established social reward model for vulnerability to adolescent depression. We then show how neuroeconomics may extend this model by offering the tools to examine the mechanics of social exchanges, in behavioral and neural terms, that maintain (or pose vulnerability to) depression. In doing so, we propose a neuroeconomic model of adolescent depression in which depression is defined as a perturbation of interpersonal motivational/reward exchange. This model serves to guide future research.     

Parkinson's disease in relation to pesticide exposure and nuclear encoded mitochondrial complex I gene variants

Parkinson's disease (PD) is a common age-related neurodegenerative disorder thought to result from the integrated effects of genetic background and exposure to neuronal toxins. Certain individual nuclear-encoded mitochondrial complex I gene polymorphisms were found to be associated with approximately 2-fold risk variation in an Australian case-control sample. We further characterized this sample of 306 cases and 321 controls to determine the mutual information contained in the 22 SNPs and, additionally, level of pesticide exposure: five distinct risk sets were identified using grade-of-membership analysis. Of these, one was robust to pesticide exposure (I), three were vulnerable (II, III, IV), and another (V) denoted low risk for unexposed persons. Risk for individual subjects varied > 16-fold according to level of membership in the vulnerable groups. We conclude that inherited variation in mitochondrial complex I genes and pesticide exposure together modulate risk for PD.     

CYP450, genetics and Parkinson's disease: gene x environment interactions hold the key

The ecogenetic theory contends that most cases of Parkinson's disease (PD) result from the actions of environmental factors in genetically susceptible individuals on a background of normal ageing. This notion is supported by epidemiologic data; family history of PD and exposures to environmental toxins such as pesticides increase risk, while cigarette smoking reduces risk. As a result, polymorphic genes that code for metabolic enzymes have been considered as candidates for conferring differential risk for PD. Given their prominence in xenobiotic metabolism, the cytochrome P450 (CYP) genes have come under great scrutiny. The activity of CYP2D6 is largely determined by genetic variability and common sequence variants exist in human populations that lead to poor metaboliser (PM) phenotypes. These have been extensively studied as genetic risk factors for PD with inconsistent results. However, these studies have disregarded interactive effects (e.g. gene x environment interactions) despite the assertions of the ecogenetic theory. Data from our group and others suggest that the CYP2D6 PM genotype interacts with certain environmental factors such as pesticide exposure and cigarette smoking to confer differential risk for PD. Previous failure to consider such interactions might, in part, explain the inconsistencies observed in the CYP2D6 genetic risk-factor literature. Our data illustrate, using CYP2D6 as an exemplar, that it is crucial to consider both genetic and environmental factors, and their interactions, in any examination of risk factors for PD.     

番荔枝种子化学成分研究

从番荔枝Annona squamosa L.种子的乙醇提取物分得三个polyketide类化合物:squamostatin-8(Ⅰ),annonin-vl(Ⅱ)和desacetyluvaricin(Ⅲ)以及胡萝卜甙(daucosterol,IV)。其中Ⅰ是一种新的polyketide,经光谱解析方法,测定了Ⅰ的化学结构式和部分碳原子的立体化学.     

An assessment of MMP and TIMP gene expression in cell lines and stroma - tumour differences in microdissected breast cancer biopsies.

To examine matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinases (TIMP) mRNA levels in archival breast cancer biopsies, we employed microdissection to separate tumour tissue from the surrounding breast tissue, or stroma and RT-PCR to determine gross qualitative and small quantitative differences in the patterns of expression. In this study, a significant correlation (p < 0.05, by Mann-Whitney U analysis) between TIMP-2 expression and lymph node involvement was identified, while MMP-11 and TIMP-1 expression patterning also significantly (p < 0.05) differed between those tumours showing calcification and those that did not. When compared by Spearmans' rho correlation analysis, a significant association (p < 0.05, rho = 0.404) was identified in the pattern of MMP-2 and MMP-9 gene expression. In this study, the use of microdissection and a systematic strategy of RT-PCR analysis have allowed us to investigate localized MMP and MMP inhibitor expression within breast tumours. We have identified patterns of gene expression that may further reveal aspects of breast carcinogenesis, and a robust method for examining changes in clinically important genes using archival biopsies and across stroma-tumour boundaries.     

The ACE deletion polymorphism is not associated with Parkinson's disease

The deletion allele (D allele) polymorphism in the angiotensin converting enzyme (ACE) gene is associated with increased levels of the neuropeptide substance P in the basal ganglia and substantia nigra. A reduction of substance P levels in the brain occurs in Parkinson's disease (PD) and has been implicated in the pathogenesis of the disease. We investigated the hypothesis that the D allele may be protective towards PD by examining the frequency of the ACE (I/D) polymorphism in 178 PD cases (male:female ratio = 1.4) and 192 controls (male:female ratio = 1.5). ACE (I/D) genotype was determined using polymerase chain reaction and 3% agarose gel electrophoresis. Unadjusted chi-square analysis revealed no significant difference between genotype frequencies (chi2 = 3.30, p > 0.10) or allele frequencies (chi2 = 2.52, p > 0.10) between patient and control groups, although PD patients were less likely to be homozygous (OR = 0.80, 95% CI = 0.49-1.29) or heterozygous (OR = 0.80, 95% CI = 0.59-1.06) for the D allele. A stepwise logistic regression analysis of the ACE deletion and risk factor data confirmed that there was no significant association between the ACE deletion (D allele) polymorphism and PD (OR = 0.62, 95% CI = 0.35-1. 10, p = 0.10). This study does not support the hypothesis that the D allele of the ACE gene confers a protective effect with respect to PD.     

Dermatomyositis panniculitis: A case report

Dermatomyositis‐related panniculitis is a rare cutaneous manifestation of dermatomyositis. There are few reported cases in the medical literature. We present the case of a 60‐year‐old woman with a 2‐year history of dermatomyositis and recent biopsy‐confirmed panniculitis treated with prednisone, cyclophosphamide and i.v. immunoglobulin.