乳酸菌素治疗HBV感染与肠源性内毒素血症的临床观察

目的 研究慢性肝病的内毒素血症发生率以及乳酸菌素治疗的效果。方法 用鲎试验（ＬＬＴ）检测。结果 检测１１７例临床各类型慢性肝病的肠源性内毒素血症阳性率,结果为４０％～８０％,以重症肝炎最高。内毒素与肿瘤坏死因子互为因果对肝病发生发展产生重要影响。检测结果显示慢性肝病ＬＬＴ阳性病人ＴＮＦ－α较ＬＬＴ阴性及正常人明显提高,差别有显著性（Ｐ〈０．０５）,应用乳酸菌素治疗肝病肠源性内毒血症３０例,鲎试验转阴率为７０．６％,显著高于对照组的６．６％,Ｐ〈０．０１。结论 乳酸菌素可作为肝病的辅助治疗的有效药物之一。     

肝病肠道菌群失调与肠源性内毒素血症

当今肝病肠道菌群失调与肠源性内毒素血症(intestinal endotoxemia,IETM)的关系日益受到重视.在肝病时发生肠道菌群微生态失调,部分革兰阴性菌大量增殖,后者导致肠源性内毒素血症发生率升高,而内毒素血症在肝病的维持和发展中起到重要的作用,加重肝病.因此,肝病-肠道菌群失调-肠源性内毒素血症形成一个恶性循环,其中关键的一个环节就是肠道菌群失调所致肠源性内毒素血症.本文将就肝病时肠道菌群失调所致肠源性内毒素血症以及微生态疗法阻断此环节的研究加以综述.     

内毒素和细胞因子在酒精性肝病中的作用

酒精性肝病发病机制比较复杂,其中内毒素、细胞因子导致的肝细胞凋亡发挥了关键作用.酒精致肠道粘膜通透性增加,引起高内毒素血症,内毒素作用于kupffer细胞产生过量细胞因子如肿瘤坏死因子a(TNF-a)等,TNF-a与肝细胞表面的相应受体结合,诱导肝细胞凋亡,引起肝细胞损害.     

失血性休克后IL—1活性变化及其与肠源性内毒素血症的关系

目的：观察失血性休克后血浆ＩＬ－１活性的变化及其与肠源性内毒素血症的关系。方法：复制容量控制失血性休克及肠源性内毒素血症模型,改良ＬＡＦ法及显色基质偶氮法检测ＩＬ－１及ＥＴ。结果：普通ＳＤ大鼠３０％失血后２～３ｈ及６～１０ｈ血浆ＩＬ－１活性升高,３ｈ后ＥＴ水平显著升高;失血前胃内灌注双岐杆菌或失血后静注ｒＢＰＩ,大鼠血浆ＥＴ水平无明显升高,ＩＬ－１活性第１峰仍然存在,但第２峰消失。无菌ＳＤ大鼠１０％失血、灌注ＬＰＳ后１ｈ血浆ＥＴ水平升高,２ｈ后ＩＬ－１活性升高。结论：失血性休克后ＩＬ－１活性呈双峰型升高,第１峰与内毒素无关,第２峰为内毒素刺激峰,内毒素主要来源于肠道     

TAA 致慢性肝病内毒素血症大鼠清道夫受体、CD14 表达变化*

目的:观察清道夫受体(SR)和脂多糖受体CD14在TAA介导的慢性肝病内毒素血症大鼠肝组织中的表达。方法:通过大鼠持续灌胃给小剂量(12mg/kg.d)TAA建立大鼠肝损伤内毒素血症模型,HE染色光镜观察肝脏病理变化;改良赖氏法检测大鼠血清ALT、AST;改良过氯酸法测定血清内毒素含量;酶联免疫法检测大鼠血清CD4+和CD8+;免疫组化染色方法观察大鼠肝组织清道夫受体和CD14的表达。结果:TAA诱导后,大鼠肝脏出现片状坏死并可见灶性炎症;血浆ALT、AST及内毒素水平显著升高(P<0.05或P<0.01);血清CD4+、CD8+T细胞明显降低(P<0.01);肝组织CD14表达上调,清道夫受体表达下调,和正常大鼠相比,差异显著(P<0.05)。结论:肝组织SR表达下降和CD14表达增强可能是TAA介导慢性肝病内毒素血症的重要机制。     

贝飞达对慢性肝病患者纤维化指标及TNF-α的影响

目的探讨贝飞达在治疗慢性肝病患者时,对血清HA、NL、IV-C及TNF-α的影响。方法血清HA、LN、IV-及TNF-α检测采用放免法。结果60例肝功能减退者,经3个月贝飞达治疗与对照组比较。HA、LN、IV-C及TNF-α值下降水平平均低于对照组,经统计学处理差异有显著性。结论应用微生态调节剂贝飞达对慢性肝病患者进行抗纤维化治疗。通过调整肠道菌群失调。控制内毒素血症,可以降低血清HA、LN、IV-C及TNF-α水平,从而达到预防和控制纤维化的目的。     

家兔血浆内毒素的鲎试验定量测定法及其应用

本文介绍采用过氯酸法处理血浆,应用国产鲎试剂和基质定量检测动物血中微量内毒素的鲎试验合成基质偶氮显色法。该法变异系数为3.8—4.6％,回收率为84—112％,对革兰氏阴性细菌具有特异性。应用该法对实验性大肠杆菌败血症家兔及实验性肺炎球菌感染家兔的血浆内毒素定量测定结果表明,均有不同程度的内毒素血症出现。     

大鼠内毒素血症不同时期血浆CGRP和背根神经节CGRP mRNA水平的变化

本文采用逆转录聚合酶链反应（RT－PCR）方法测定大鼠内毒素血症不同时期胸腰段背根神经节降钙素基因相关肽（CGRP）mRNA水平的改变,结合血浆CGRP水平的改变,以期全面了解大鼠内毒素血症不同时期CGRP释放与合成的变化。结果显示：注射内毒素（5mg／kg）后30min时,大鼠血浆CGRP开始增高,而背根神经节CGRPmRNA水平无明显变化;注射内毒素后3h时,血浆CGRP及背根神经节CGRPmRNA均明显增高．分别为142％和32％,8h时则进一步增高,分别为216％和85％。提示内毒素不仅刺激外周组织释放CGRP,而且还能通过某些机制激活背根神经节CGRPmRNA的转录,使CGRP合成增加,以作为CGRP大量释放的重要补充来源。     

内毒素耐受的机制研究

内毒素是革兰阴性菌细胞壁的成分,能够激发机体的免疫反应。当细菌释放大量的内毒素到血液,即可引起内毒素血症,内毒素血症可以伴随多种疾病出现,引起致死性感染性休克,循环功能衰竭,其病死率极高。内毒素耐受是指机体接受小剂量内毒素刺激后对后续内毒素刺激的反应性降低,表现为促炎因子释放减少而抗炎因子释放增加,机体发热,缺氧,低血压,休克的症状减轻。内毒素耐受的发生机制极其复杂,受机体内多种因素的调节,但目前尚无明确的结论。近年来,有关其机制的研究有许多报道,其中,对内毒素耐受的信号机制的研究最为广泛,大量的研究表明内毒素的主要受体,细胞内的信号蛋白,负调控因子以及转录因子可能在内毒素耐受的发生过程中起重要作用。也有报道表明免疫细胞的凋亡,染色体修饰和基因重排以及小RNA的参与可能诱导内毒素耐受的发生。本文从细胞、分子水平对内毒素耐受的发生机制进行综述,拟对炎症性疾病如内毒素血症的预防和治疗提供理论依据。     

复合益生菌活菌制剂对慢性肝病患者血内毒素的影响

探讨复合益生菌活菌制剂对慢性肝病患者血内毒素(ET)的影响,明确此类药物在慢性肝病治疗中的应用价值,我们选择了60例住院慢性肝病患者随机分组作对照观察,现报告如下.     

The Saccharomyces cerevisiae BEM1 homologue in Neurospora crassa promotes co‐ordinated cell behaviour resulting in cell fusion

Directed growth or movement is a common feature of microbial development and propagation. In polar growing filamentous fungi, directed growth requires the interaction of signal sensing machineries with factors controlling polarity and cell tip extension. In Neurospora crassa an unusual mode of cell–cell signalling mediates mutual attraction of germinating spores, which subsequently fuse. During directed growth of the two fusion partners, the cells co‐ordinately alternate between two physiological stages, probably associated with signal sending and receiving. Here, we show that the Saccharomyces cerevisiae BEM1 homologue in N. crassa is essential for the robust and efficient functioning of this MAP kinase‐based signalling system. BEM1 localizes to growing hyphal tips suggesting a conserved function as a polarity component. In the absence of BEM1, activation of MAK‐2, a MAP kinase essential for germling fusion, is strongly reduced and delayed. Germling interactions become highly instable and successful fusion is greatly reduced. In addition, BEM1 is actively recruited around the forming fusion pore, suggesting potential functions after cell–cell contact has been established. By genetically dissecting the contribution of BEM1 to additional various polarization events, we also obtained first hints that BEM1 might function in different protein complexes controlling polarity and growth direction.     

The RHO1-GAPs SAC7, BEM2 and BAG7 control distinct RHO1 functions in Saccharomyces cerevisiae

In Saccharomyces cerevisiae, the small GTPase RHO1 plays an essential role in the control of cell wall synthesis and organization of the actin cytoskeleton. Several regulators for RHO1 are known, including the GTPase-activating proteins (GAPs) SAC7 and BEM2. Here we show that BAG7, identified as the closest homologue of SAC7, also acts as a GAP for RHO1 in vitro and in vivo. Furthermore, we find that BAG7, SAC7, and BEM2 are functionally different in vivo. Overexpression of BAG7 or SAC7,but not BEM2, suppresses the cold sensitivity of a sac7 mutation and the lethality of RHO1 hyperactivation in response to cell wall damage. In contrast, overexpression of BEM2 or SAC7, but not BAG7, downregulates the RHO1-controlled PKC1-MPK1 pathway, and disruption of BEM2 or SAC7, but not BAG7, results in increased MPK1 activation. We conclude that BEM2 and SAC7, but not BAG7, are involved in the control of the RHO1-mediated activation of MPK1, whereas BAG7 and SAC7, but not BEM2, are involved in the regulation of other RHO1 functions. This suggests that different RHO1GAPs control different RHO1 effector pathways, thus ensuring their individual regulation at the appropriate place and time.     

Boundary element method analysis for the bioheat transfer equation.

In this paper, the boundary element method (BEM) approach is applied to solve the Pennes (1948) bioheat equation. The objective is to develop the BEM formulation and demonstrate its feasibility. The basic BEM formulations for the transient and steady-state cases are first presented. To demonstrate the usefulness of the BEM approach, numerical solutions for 2-D steady-state problems are obtained and compared to analytical solutions. Further, the BEM formulation is applied to model a conjugate problem for an artery imbedded in a perfused heated tissue. Analytical solution is possible when the conduction in the x-direction is negligible. The BEM and analytical results have very good agreement.     

Control of cellular morphogenesis by the Ip12/Bem2 GTPase-activating protein: possible role of protein phosphorylation

The IPL2 gene is known to be required for normal polarized cell growth in the budding yeast Saccharomyces cerevisiae. We now show that IPL2 is identical to the previously identified BEM2 gene. bem2 mutants are defective in bud site selection at 26 degrees C and localized cell surface growth and organization of the actin cytoskeleton at 37 degrees C. BEM2 encodes a protein with a COOH-terminal domain homologous to sequences found in several GTPase-activating proteins, including human Bcr. The GTPase-activating protein-domain from the Bem2 protein (Bem2p) or human Bcr can functionally substitute for Bem2p. The Rho1 and Rho2 GTPases are the likely in vivo targets of Bem2p because bem2 mutant phenotypes can be partially suppressed by increasing the gene dosage of RHO1 or RHO2. CDC55 encodes the putative regulatory B subunit of protein phosphatase 2A, and mutations in BEM2 have previously been identified as suppressors of the cdc55-1 mutation. We show here that mutations in the previously identified GRR1 gene can suppress bem2 mutations. grr1 and cdc55 mutants are both elongated in shape and cold- sensitive for growth, and cells lacking both GRR1 and CDC55 exhibit a synthetic lethal phenotype. bem2 mutant phenotypes also can be suppressed by the SSD1-vl (also known as SRK1) mutation, which was shown previously to suppress mutations in the protein phosphatase- encoding SIT4 gene. Cells lacking both BEM2 and SIT4 exhibit a synthetic lethal phenotype even in the presence of the SSD1-v1 suppressor. These genetic interactions together suggest that protein phosphorylation and dephosphorylation play an important role in the BEM2-mediated process of polarized cell growth.     

Use of a screen for synthetic lethal and multicopy suppressee mutants to identify two new genes involved in morphogenesis in Saccharomyces cerevisiae.

Genes CDC24 and CDC42 are required for the establishment of cell polarity and for bud formation in Saccharomyces cerevisiae. Temperature-sensitive (Ts-) mutations in either of these genes cause arrest as large, unbudded cells in which the nuclear cycle continues. MSB1 was identified previously as a multicopy suppressor of Ts- cdc24 and cdc42 mutations. We have now sequenced MSB1 and constructed a deletion of this gene. The predicted amino acid sequence does not closely resemble any other in the available data bases, and the deletion does not produce any readily detectable phenotype. However, we have used a colony-sectoring assay to identify additional genes that appear to interact with MSB1 and play a role in bud emergence. Starting with a strain deleted for the chromosomal copy of MSB1 but containing MSB1 on a high-copy-number plasmid, mutants were identified in which MSB1 had become essential for viability. The new mutations defined two genes, BEM1 and BEM2; both the bem1 and bem2 mutations are temperature sensitive and are only partially suppressed by MSB1. In bem1 cells, a single copy of MSB1 is necessary and sufficient for viability at 23 or 30 degrees C, but even multiple copies of MSB1 do not fully suppress the growth defect at 37 degrees C. In bem2 cells, a single copy of MSB1 is necessary and sufficient for viability at 23 degrees C, multiple copies are necessary for viability at 30 degrees C, and even multiple copies of MSB1 do not suppress the growth defect at 37 degrees C. In a wild-type background (i.e., a single chromosomal copy of MSB1), both bem1 and bem2 mutations cause cells to become large and multinucleate even during growth at 23 degrees C, suggesting that these genes are involved in bud emergence. This suggestion is supported for BEM1 by other evidence obtained in a parallel study (J. Chant, K. Corrado, J. Pringle, and I. Herskowitz, submitted for publication). BEM1 maps centromere distal to TYR1 on chromosome II, and BEM2 maps between SPT15 and STP2 on chromosome V.     

PiggyBac-based screening identified BEM4 as a suppressor to rescue growth defects in och1-disrupted yeast cells

Glycoengineered yeast cells, which express human-compatible glycan structures, are particularly attractive host cells to produce therapeutic glycoproteins. Disruption of OCH1 gene, which encodes an α-1,6-mannosyltransferase required for mannan-type N-glycan formation, is essential for the elimination of yeast-specific N-glycan structures. However, the gene disruption causes cell wall defects leading to growth defects. Here, we tried to identify factors to rescue the growth defects of och1Δ cells by in vivo mutagenesis using piggyBac (PB)-based transposon. We isolated a mutant strain, named 121, which could grow faster than parental och1Δ cells. The PB element was introduced into the promoter region of BEM4 gene and upregulated the BEM4 expression. Overexpression of BEM4 suppressed growth defects in och1Δ cells. The slow grow phenotypes were partially rescued by expression of Rho1p, whose function is regulated by Bem4p. Our results indicate that BEM4 would be useful to produce therapeutic proteins in glycoengineered yeast without the growth defects.     

Application of Different Variants of the BEM in Numerical Modeling of Bioheat Transfer Problems

Heat transfer processes proceeding in the living organisms are described by the different mathematical models. In particular, the typical continuous model of bioheat transfer bases on the most popular Pennes equation, but the Cattaneo-Vernotte equation and the dual phase lag equation are also used. It should be pointed out that in parallel are also examined the vascular models, and then for the large blood vessels and tissue domain the energy equations are formulated separately. In the paper the different variants of the boundary element method as a tool of numerical solution of bioheat transfer problems are discussed. For the steady state problems and the vascular models the classical BEM algorithm and also the multiple reciprocity BEM are presented. For the transient problems connected with the heating of tissue, the various tissue models are considered for which the 1st scheme of the BEM, the BEM using discretization in time and the general BEM are applied. Examples of computations illustrate the possibilities of practical applications of boundary element method in the scope of bioheat transfer problems.     

Characterization of bud emergence 46 (BEM46) protein: Sequence,structural, phylogenetic and subcellular localization analyses

The bud emergence 46 (BEM46) protein from Neurospora crassa belongs to the α/β-hydrolase superfamily. Recently, we have reported that the BEM46 protein is localized in the perinuclear ER and also forms spots close by the plasma membrane. The protein appears to be required for cell type-specific polarity formation in N. crassa. Furthermore, initial studies suggested that the BEM46 amino acid sequence is conserved in eukaryotes and is considered to be one of the widespread conserved “known unknown” eukaryotic genes. This warrants for a comprehensive phylogenetic analysis of this superfamily to unravel origin and molecular evolution of these genes in different eukaryotes. Herein, we observe that all eukaryotes have at least a single copy of a bem46 ortholog. Upon scanning of these proteins in various genomes, we find that there are expansions leading into several paralogs in vertebrates. Usingcomparative genomic analyses, we identified insertion/deletions (indels) in the conserved domain of BEM46 protein, which allow to differentiate fungal classes such as ascomycetes from basidiomycetes. We also find that exonic indels are able to differentiate BEM46 homologs of different eukaryotic lineage. Furthermore, we unravel that BEM46 protein from N. crassa possess a novel endoplasmic-retention signal (PEKK) using GFP-fusion tagging experiments. We propose that three residues namely a serine 188S, a histidine 292H and an aspartic acid 262D are most critical residues, forming a catalytic triad in BEM46 protein from N. crassa. We carried out a comprehensive study on bem46 genes from a molecular evolution perspective with combination of functional analyses. The evolutionary history of BEM46 proteins is characterized by exonic indels in lineage specific manner.     

The solution structure of an N-terminally truncated version of the yeast CDC24p PB1 domain shows a different beta-sheet topology

Phox and Bem1 (PB1) domains mediate protein-protein interactions via the formation of homo- or hetero-dimers. The C-terminal PB1 domain of yeast cell division cycle 24 (CDC24p), a guanine-nucleotide exchange factor involved in cell polarity establishment, is known to interact with the PB1 domain occurring in bud emergence MSB1 interacting 1 (BEM1p) during the regulation of the yeast budding process via its OPR/PC/AID (OPCA) motif. Here, we present the structure of an N-terminally truncated version of the Sc CDC24p PB1 domain. It shows a different topology of the beta-sheet than the long form. However, the C-terminal part of the structure shows the conserved PB1 domain features including the OPCA motif with a slight rearrangement of helix alpha1. Residues which are important for the heterodimerization with BEM1p are structurally preserved.     

Characterization of Two Novel Propachlor Degradation Pathways in Two Species of Soil Bacteria

Propachlor (2-chloro-N-isopropylacetanilide) is an acetamide herbicide used in preemergence. In this study, we isolated and characterized a soil bacterium, Acinetobacter strain BEM2, that was able to utilize this herbicide as the sole and limiting carbon source. Identification of the intermediates of propachlor degradation by this strain and characterization of new metabolites in the degradation of propachlor by a previously reported strain of Pseudomonas (PEM1) support two different propachlor degradation pathways. Washed-cell suspensions of strain PEM1 with propachlor accumulated N-isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain PEM1 grew on propachlor with a generation time of 3.4 h and a K_s of 0.17 ± 0.04 mM. Acinetobacter strain BEM2 grew on propachlor with a generation time of 3.1 h and a K_s of 0.3 ± 0.07 mM. Incubations with strain BEM2 resulted in accumulation of N-isopropylacetanilide, N-isopropylaniline, isopropylamine, and catechol. Both degradative pathways were inducible, and the principal product of the carbon atoms in the propachlor ring was carbon dioxide. These results and biodegradation experiments with the identified metabolites indicate that metabolism of propachlor by Pseudomonas sp. strain PEM1 proceeds through a different pathway from metabolism by Acinetobacter sp. strain BEM2.