正常人左心室功能指标的参考值及其预计公式的初步研究报告*

目的: 当前评估左心室容量和功能仍常用正常值范围,个体化分析也仅使用体表面积进行校正。尚缺少个体化因素相关的大样本参考值和预计公式。方法: 本研究纳入美国加州洛杉矶县南湾地区1200名健康志愿者,其中男807女393,年龄20岁-94岁,心脏CT造影（CTA）,经过高精度三维成像技术处理,计算左心室容积在收缩和舒张过程中的连续动态变化,测定左心室（LV）容量和功能指标：舒张末期容积（EDV)、收缩末期容积(ESV)、每搏输出量(SV)、射血分数(EF)和心输出量(CO)。将以上指标与一般特征指标进行多因素相关分析,以探索正常人预计值计算公式。结果: 男性除LVEF小于女性外（P<0.001）,其余各指标均大于女性（P<0.001）。多元线性回归分析提示, 性别、年龄、身高和体质量均为EDV、ESV、SV的独立影响因子(P<0.001); 而CO仅受年龄、性别、体质量显著影响(P<0.001),但与身高无关(P>0.05)。CO的预测公式CO (L·min^-1)= 6.963+0.446(Male) -0.037×年龄(yr)+0.013×体质量(kg)。结论: 性别、年龄、身高、体质量均影响左心室容量和功能,建立预测值计算公式,对心血管疾病的无创评估和个体化精准医疗具有重要参考价值。     

利用改进的酚-氯仿法从猪毛囊中提取基因组DNA

为提高从猪毛囊组织中提取基因组DNA的效率, 文章在借鉴从其他组织提取基因组DNA方法的基础上, 对经典的酚-氯仿法的反应体系和步骤进行了改进。利用改进的酚－氯仿抽提法, 从猪的毛囊组织中快速、高效地提取了高质量基因组DNA。利用该方法从1~6根猪毛囊中提取的基因组DNA可满足基于PCR技术的相关分子生物学实验需要。     

Isolation and osmotic relations of developing megagametophytes of Torenia fournieri

A new method is reported to isolate and handle living megagametophytes of Torenia fournieri at any developmental stage. The stages were determined using light microscopy and delimited by correlating floral morphological traits. When significant changes in the osmotic pressure were found during development, enzyme solutions contained different concentrations of osmoticum. Osmotic pressure is lowest in the megaspore, increases until the four-nucleate stage and then gradually decreases until complete embryo sac formation. In enzymatic solutions containing appropriate concentrations of osmoticum, protoplasts of megaspores, two-, four-, eight-nucleate embryo sacs, egg cells, synergids and central cells were successfully isolated. The living protoplasts were collected by micromanipulator, transferred into microdroplets and tested for viability. Received: 1 June 1998 / Revision accepted: 20 May 1999     

Bigendothelin-1 (1-21) fragment during early sepsis modulates tau, p38-MAPK phosphorylation and nitric oxide synthase activation

Earlier we have demonstrated that inhibition of endothelin biosynthesis ameliorates endotoxemia-induced inducible nitric oxide synthase (iNOS) activation and phosphorylation of p38-mitogen activated protein kinase (pp38-MAPK). Therefore, in the present study, we tested the hypothesis that activation of endothelin (ET)-1 biosynthesis using bigET-1 during early sepsis would upregulate iNOS and affect myocardial function in the rat. Male Sprague-Dawley rats (350–400 g) were anesthetised using Nembutal® (50 mg/kg, i.p.) and jugular vein, tail artery (Mean arterial pressure, MAP) and right carotid arteries (advanced to left ventricle, LV) were cannulated. The rats were randomly divided into saline-, bigET-1- and C-terminal fragment of bigET-1(bigET-1(22-38))-treated groups. Sepsis was induced using i.p. injection of cecal inoculum obtained from a donor rat (200 mg/kg in 5 ml 5% sterile dextrose water, D₅W). Sham animals received an i.p. injection of D₅W (5 ml/kg). MAP and LVP were recorded and cardiodynamic parameters were calculated at 0, 2, 6, 12 and 24 h post sham or sepsis-induction. A significant elevation in LV isovolumic relaxation rate constant (tau), LV end diastolic pressure (LVEDP) and rate pressure product (RPP) was observed in vehicle-treated septic group at 24 h. BigET-1 significantly increased concentration of LV ET-1 both in sham and septic groups. BigET-1 elevated tau and LVEDP both in sham and septic animals as early as 12 h which persisted through 24 h. However, bigET-1(22-38) elevated LVEDP in septic group at 24 h but not in sham group. BigET-1 accentuated the levels of plasma nitric oxide byproduct (NOx) levels in both sham and septic animals at 6, 12 and 24 h. Sepsis increased myocardial iNOS at 24 h. BigET-1 significantly upregulated expression of myocardial iNOS and pp38-MAPK. The data suggest that increased substrate availability for ET-1 at the time of sepsis-induction contributes in diastolic dysfunction, iNOS activation and p38-MAPK phosphorylation. (Mol Cell Biochem 271: 225–237, 2005)     

Hypercholesterolemia attenuates postischemic ventricular dysfunction in the isolated rabbit heart

The effects of the chronic administration of cholesterol on the stunned myocardium have not been studied. The objective was to determine the effect of a cholesterol enriched diet on postischemic ventricular dysfunction. In group 1 (G1, n = 7 isolated rabbit hearts underwent a follow up of ventricular function during 30 min in aerobic conditions. In group 2 (G2, n = 6) G1 was repeated but the animals were subjected to a 1% cholesterol enriched diet during 4 weeks (hypercholesterolemic animals). In group 3 (G3, n = 8) hearts underwent 15 min of global ischemia followed by 30 min of reperfusion. In Group 4 (G4, n = 11) G3 was repeated, but in hypercholesterolemic animals. Since cholesterol decreased the inotropism in basal situation, and this makes the comparison between groups difficult, we performed a Group 5 (G5, n = 7), in which G4 protocol was repeated but isoproterenol (8 g/kg/min) was administered 10 min before ischemia, in order to match the preischemic inotropic state with respect to the normocholesterolemic ones. G1 and G2 maintained a stable inotropism during the 30 min of perfusion. The preischemic left ventricular developed pressure (LVDP) in G3 and G4 was 91.4± 4.3 and 70.8± 3.4 mmHg (p< 0.05), respectively, and after 30 min of reperfusion differences were not observed between G3 and G4. Nevertheless, when LVDP is expressed as a percentage, we detected an attenuation of postischemic systolic alterations in hypercholesterolemic animals (67.3± 3.6 in G4 vs. 90.8± 3.1% in G3, p< 0.05). When LVDP in G5 was increased until matching the one of G3, there were no differences after 30 min of reperfusion. Left ventricular end diastolic pressure increased 285± 46%, 61± 25% (p< 0.05 vs. G3 and G5) and 216± 25% in G3, G4 and G5 at 30 min of reperfusion. There were no differences either in the values of tau or infarct size between groups. Thus, in hypercholesterolemic animals, a decrease of the preischemic inotropism exists and there is an attenuation of the stunned myocardium. When contractility of the normo and hypercholesterolemic animals is matched, the beneficial effect disappears.     

Emergence of hematopoietic stem cells in the human embryo

We have previously identified a novel site of hematopoietic cell production within the human embryo, which is localised in the ventral wall of the dorsal aorta and vitelline artery. Cells emerging in that territory between 27 and 40 days of gestation exhibit the expected phenotypic, molecular, and functional properties of hematopoietic stem cells and are the first multipotent, lympho-myeloid progenitors that appear in human ontogeny. We have next demonstrated that vascular endothelial cells sorted stringently, by flow cytometry, from the human yolk sac and embryonic aorta exhibit dramatic blood-forming potential in culture. These results suggest a filiation between vascular endothelium and hematopoietic cells in the course of early human ontogeny. More preliminary data indicate that a subpopulation of vascular endothelial cells in the bone marrow may retain this hematogenous potential until adult stages.     

Avian HSC emergence, migration, and commitment toward the T cell lineage

To date three sites of emergence of hemopoietin cells have been identified during early avian development: the yolk sac, the intraaortic clusters and recently the allantois. However, the contributions of the hematopoietic stem cell (HSC) populations generated by these different sites to definitive hematopoiesis and their migration routes are not fully unraveled. Experimental embryology as well as the establishment of the genetic cascades involved in HSC emergence help now to draw a better scheme of these processes.     

Choroideremia gene product affects trophoblast development and vascularization in mouse extra-embryonic tissues

Choroideremia (CHM) is a hereditary eye disease caused by mutations in the X-linked CHM gene. Disruption of the Chm gene in mice resulted in prenatal death of Chm-/Y males and Chm-/Chm+ females that had inherited the mutation from their mothers. Male chimeras and Chm+/Chm- females with paternal transmission of the mutation were viable and had photoreceptor degeneration reminiscent of human choroideremia. Here, we show that Chm-/Y males and Chm-/Chm+ females were retarded at e7.5 and died before e11.5 due to multiple defects of the extra-embryonic tissues. Mutant embryos exhibited deficiency of diploid trophoblasts associated with overabundance of giant cells. In yolk sac and placenta, severe defects in vasculogenesis were obvious. Chm-/Y males exhibited more pronounced phenotypes than Chm-/Chm+ females. The lethal genotypes could be rescued by tetraploid aggregation. Chm-/Chm+ females, but not Chm-/Y males, could also be rescued when their Chm+/Chm- mothers were mated with Mus spretus males. Backcross analysis suggested that the viability of interspecies hybrid Chm-/Chm+ females may be due to expression from the Chm allele on the M. spretus X-chromosome rather than a modifier effect. Our results demonstrate that Chm is essential for diploid trophoblast development and plays a role in the vascularization in placenta and yolk sac.     

Immunocytochemical analysis of cubilin-mediated endocytosis of high density lipoproteins (HDL) in epithelial cells of the rat visceral yolk sac

Cubilin was recently shown to function as an endocytic receptor for high density lipoprotein (HDL) holoparticles and apolipoprotein A–I (apo A–I), the main protein constituent of HDL. In the present study, we analyzed the distribution and intracellular trafficking of cubilin and HDL in rat visceral yolk sac epithelial cells. After epithelial cells were loaded with apolipoprotein E-free HDL for 30 min in vitro, double immunofluorescence showed that the apical cytoplasm of the cells was strongly stained with anti-cubilin antibodies and anti-apo A–I/HDL. Furthermore, double immunogold electron-microscopic observations revealed the distinct localization of cubilin and HDL in endocytic vacuoles. In early endosomes, both were colocalized on the membrane. Although, in late endosomes, cubilin was also localized on the membrane, HDL was mainly located in the matrix. Both were found in the matrix in lysosomes. In addition, cubilin was markedly localized in apical tubules (ATs), which are generally accepted as being receptor recycling compartments. Thus, HDL is internalized through cubilin-mediated endocytosis and is finally transported to lysosomes. By contrast, cubilin is mainly translocated to ATs for recycling, although some of the cubilin is degraded in lysosomes. Quantitative analysis further revealed that cubilin was not concentrated on the membranes of ATs, although it accumulated in the AT area. Some HDL were also observed in the AT area. These findings suggest that the translocation of cubilin and HDL to ATs from early endosomes occurs through a simple sorting mechanism based on the geometry of these compartments and the bulk membrane and volume flow.