Granulocyte-Macrophage Colony-Stimulating Factor and Tumor Necrosis Factor Alpha in Patients with Human Immunodeficiency Virus (HIV) Type 1 Infection

Variations in cytokine production in patients with human immunodeficiency virus (HIV) infection could be involved in the physiopathology and in the progression of the disease. Therefore we studied the level of granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor α (TNFα) produced in patients with HIV infection at stage II (asymptomatic seropositives) and stage IV (AIDS) of the CDC classification, by using an enzyme amplified sensitivity immunoassay. We measured the level of GM-CSF and TNFα in supernatant of phytohemagglutinin-activated peripheral blood mononuclear cells from patients and healthy individuals. In one out of 10 stage II patients and 4 out of 14 stage IV patients, we obtained higher levels of GM-CSF than the mean + 2 S.D. of controls, but in 3 stage IV patients with very low CD4+ T lymphocyte counts (< 50/mm^–3) compared to other patients, the GM-CSF values were very low. High levels of TNFα were detected in 3 out of 10 stage II and 6 out of 11 stage IV patients. The high values of TNFα were associated with high values of GM-CSF in stage II and in most of AIDS patients except those with very low CD4⁺ T cell counts, who produced low levels of GM-CSF. Plasma levels of cytokines were evaluated in 10 stage II, 22 stage IV patients and 20 controls. Increased levels of GM-CSF (more than 9 pg/ml) were observed in the plasma from 8 out of 10 stage II patients and 17 out of 22 stage IV patients. The tendency that increased levels of GM-CSF were associated with increased levels of TNFα was observed in plasma from stage IV patients. We report a disarray of GM-CSF production in patients with HIV infection that could be involved in clinical manifestations and progression of the disease.     

Characterization of Recombinant Integrase of Human Immunodeficiency Virus Type 1 (Isolate Bru)

Integration of the human immunodeficiency virus type 1 (HIV-1) DNA into the human genome requires the virusencoded integrase protein. The recombinant integrase protein of HIV-1 (isolate Bru) was prepared by constructing a plasmid based on pET-15b encoding the integrase gene. Integrase of HIV-1 was purified using a bacterial expression system (Escherichia coli). The main kinetic parameters of HIV-1 integrase (K _m = (3.7 ± 0.2)·10^–10 M, k _cat = (1.2 ± 0.3)·10^–7 sec^–1) were determined using an oligonucleotide duplex constructed on the basis of the U5-terminal sequence of proviral HIV-1 DNA as the substrate. Inhibition of integrase by aurintricarbonic acid ([I]₅₀ = 6.3 ± 0.4 M) and dependence of integrase activity on Mg²⁺ and Mn²⁺ concentration were studied.     

硒与艾滋病关系研究的新进展

硒是人体必需微量元素,硒与癌症、动脉硬化、心脏病的关系已为人们所熟悉.近年来硒在艾滋病发生发展中的作用越来越引起人们的关注.在HIV感染者及AIDS病人具有渐进性硒衰竭的症状,而病毒及T细胞有可能具有编码硒蛋白的功能,其中一种病毒编码的硒蛋白可能对HIV的表达起抑制作用.在体外,硒对HIV-1的复制具有调节作用.这些为硒的临床应用提供了潜在的前景.     

Microtubule-associated Proteins 1 (MAP1) Promote Human Immunodeficiency Virus Type I (HIV-1) Intracytoplasmic Routing to the Nucleus

After cell entry, HIV undergoes rapid transport toward the nucleus using microtubules and microfilaments. Neither the cellular cytoplasmic components nor the viral proteins that interact to mediate transport have yet been identified. Using a yeast two-hybrid screen, we identified four cytoskeletal components as putative interaction partners for HIV-1 p24 capsid protein: MAP1A, MAP1S, CKAP1, and WIRE. Depletion of MAP1A/MAP1S in indicator cell lines and primary human macrophages led to a profound reduction in HIV-1 infectivity as a result of impaired retrograde trafficking, demonstrated by a characteristic accumulation of capsids away from the nuclear membrane, and an overall defect in nuclear import. MAP1A/MAP1S did not impact microtubule network integrity or cell morphology but contributed to microtubule stabilization, which was shown previously to facilitate infection. In addition, we found that MAP1 proteins interact with HIV-1 cores both in vitro and in infected cells and that interaction involves MAP1 light chain LC2. Depletion of MAP1 proteins reduced the association of HIV-1 capsids with both dynamic and stable microtubules, suggesting that MAP1 proteins help tether incoming viral capsids to the microtubular network, thus promoting cytoplasmic trafficking. This work shows for the first time that following entry into target cells, HIV-1 interacts with the cytoskeleton via its p24 capsid protein. Moreover, our results support a role for MAP1 proteins in promoting efficient retrograde trafficking of HIV-1 by stimulating the formation of stable microtubules and mediating the association of HIV-1 cores with microtubules.     

Extracellular HIV-1 Nef increases migration of monocytes

Infiltration of human immunodeficiency virus type 1 (HIV-1)-infected and uninfected monocytes/macrophages in organs and tissues is a general phenomenon observed in progression of acquired immunodeficiency syndrome (AIDS). HIV-1 protein Nef is considered as a progression factor in AIDS, and is released from HIV-1-infected cells. Here, we show that extracellular Nef increases migration of monocytes. This effect is (i) concentration-dependent, (ii) reaches the order of magnitude of that induced by formyl-methyonyl-leucyl-proline (fMLP) or CC chemokine ligand 2 (CCL2)/monocyte chemotactic protein (MCP)-1, (iii) inhibited by anti-Nef monoclonal antibodies as well as by heating, and (iv) depends on a concentration gradient of Nef. Further, Nef does not elicit monocytic THP-1 cells to express chemokines such as CCL2, macrophage inhibitory protein-1alpha (CCL3) and macrophage inhibitory protein-1beta (CCL4). These data suggest that extracellular Nef may contribute to disease progression as well as HIV-1 spreading through affecting migration of monocytes.     

17种植物中蛋白质提取物的抗HIV—1活性

以化合物对ＨＩＶ－１诱导Ｃ８１６６细胞形成合胞体的抑制实验和化合物对ＨＩＶ－１感染细胞的保护实验作为初筛方法,筛选了来源７科１７种植物的４７个样品,其中８个样品经测定是核糖体失活蛋白,其余为粗提蛋白。     

Increased Neopterin Levels in Brains of Patients with Human Immunodeficiency Virus Type 1 Infection

Postmortem levels of native neopterin (D-erythro-neopterin) were measured in cerebral cortical samples from 44 human immunodeficiency virus type 1-infected and eight uninfected, nonneurological control patients. Cerebral cortical gray and white matter neopterin levels for the controls ranged from 0.5 to 7.2 pmol/mg of protein in contrast to neopterin levels in brains of the virus-infected patients, which frequently were more than threefold and occasionally more than 30-fold higher than mean control levels. Cortical neopterin levels did not correlate with severity of the acquired immunodeficiency syndrome dementia complex, but subcortical levels correlated with the presence of active human immunodeficiency virus type 1 infection, as reflected by pathological evidence of multinucleated giant cell encephalitis. Evidence of opportunistic cytomegalovirus infections in approximately 25% of the human immunodeficiency virus type 1-infected patients was associated with enhanced levels of neopterin in frontal cortex.     

Comparative Fitness of Multi-Dideoxynucleoside-Resistant Human Immunodeficiency Virus Type 1 (HIV-1) in an In Vitro Competitive HIV-1 Replication Assay

We examined whether human immunodeficiency virus type 1 (HIV-1) fitness was altered upon the acquisition of a set or subset of five mutations (A62V, V75I, F77L, F116Y, and Q151M) in the pol gene, which confers resistance to multiple dideoxynucleosides (MDR), as well as the zidovudine resistance-associated mutation T215Y, using a competitive HIV-1 replication assay in a setting of an HXB2D genetic background. Target H9 cells were exposed to a 50:50 mixture of paired infectious molecular clones, and HIV-1 in the culture supernatant was transmitted to new cultures every 7 to 10 days. The polymerase-encoding region of the virus was sequenced at various time points, and the relative proportion of the two viral populations was determined. In the absence of drugs, the comparative order for replicative fitness was HIV-162/75/77/116/151 > HIV-177/116/151 > HIV-1151 > wild-type HIV-1 (HIV-1wt) > HIV-175/77/116/151 > HIV-1151/215 > HIV-1215. In the presence of zidovudine or didanosine, the order was HIV-162/75/77/116/151 > HIV-177/116/151 > HIV-175/77/116/151 > HIV-1151 > HIV-1215. HIV-1215S(TCC), a putative intermediate infectious clone for HIV-1215, replicated comparably to HIV-1wt, while two putative intermediates for HIV-1151 [HIV-1151L(CTG) and HIV-1151K(AAG)] replicated much less efficiently than HIV-1wt and HIV-1151, suggesting that for HIV-1151 to develop, two base substitutions are likely to occur concurrently or within a short interval. These data may illustrate the molecular basis by which HIV-1151 emerges much less frequently than HIV-1215. The present data also demonstrate that several MDR HIV-1 variants are more fit than HIV-1wt in the absence of drugs and that resistance-associated mutations and drug pressure are critical variates for HIV-1 fitness.     

人免疫缺陷病毒1型TAT蛋白点突变对其反式激活作用的影响

不同免疫缺陷病毒（ＨＩＶ－１,ＨＩＶ－２和ＳＩＶ）的Ｔａｔ均有３个高度保守的结构域：Ｃｙｓ富集域、核心域和碱性氨基酸富集域,用ＰＣＲ定点突变法在ＨＩＶ－１Ｔａｔ蛋白的这些区域引入单氨基酸或多氨基酸突变;构建了以ＨＩＶ－１ＬＴＲ－１５８到＋８Ｏ区域为启动子,含有不同突变点的突变Ｔａｔ基因表达质粒;以荧光酶基因为报告基因,瞬时共转染Ｊｕｒｋａｔ细胞：分析不同氨基酸突变对Ｔａｔ的反式激活作用的影响。结果发现,突变后的Ｔａｔ蛋白的活性均极大地降低或丧失,表明这些序列内的氨基酸的确定性对Ｔａｔ的活性至关重要。     

The T-cell receptor Vβ repertoire in naive and human immunodeficiency virus-1 (HIV-1) infected chimpanzees

Using a panel of human T-cell receptor (TCR) variable region β chain (Vβ) polymerase chain reaction (PCR) primers, we performed cross-sectional and longitudinal analyses of the TCR Vβ repertoire in naive and HIV-1 infected chimpanzees. We demonstrate that our TCR PCR primer panel will support amplification of chimpanzee cDNA from most of the TCR Vβ families. However, no differences in TCR Vβ expression were found between the naive and HIV-1 infected chimpanzees, unlike the TCR Vβ repertoire perturbation found in HIV-1 infected human subjects. This finding suggests that a complete TCR repertoire in HIV-1 infected chimpanzees is associated with the maintenance CD4 + T-cell numbers and lack of progression to AIDS.     

Influence of Regulatory Genes of Type 1 Human Immunodeficiency Virus on Proliferation and Differentiation of Murine Embryonic Stem Cells

Spontaneous formation of embryoid bodies and subsequent differentiation of some cells into cardiomyocytes were demonstrated on murine embryonic stem cells of R1 line. The lines of embryonic stem cells were obtained that had been transfected with genetic constructs carrying expressing regulatory genes of the human immunodeficiency virus tat and nef and green protein gene (GFP). The transfection of embryonic stem cells with the gene tat stimulated their proliferative activity, while this activity decreased in the cells transfected with the gene nef. The time necessary for the formation of embryoid bodies by all lines of transfected cells was similar to that in the control cells. In the cultures of cells transfected with nef and tat, the number of embryoid bodies and the percentage of embryoid bodies with contracting cardiomyocytes were higher and lower than in the control, respectively. Thus, an inverse correlation was observed between the effects of regulatory genes of the human immunodeficiency virus on proliferation and differentiation embryonic stem cells.     

On the role of long incubation periods in the dynamics of acquired immunodeficiency syndrome (AIDS)

In this study, we investigate systematically the role played by the reproductive number (the number of secondary infections generated by an infectious individual in a population of susceptibles) on single group populations models of the spread of HIV/AIDS. Our results for a single group model show that if R 1, the disease will die out, and strongly suggest that if R > 1 the disease will persist regardless of initial conditions. Our extensive (but incomplete) mathematical analysis and the numerical simulations of various research groups support the conclusion that the reproductive number R is a global bifurcation parameter. The bifurcation that takes place as R is varied is a transcritical bifurcation; in other words, when R crosses 1 there is a global transfer of stability from the infection-free state to the endemic equilibrium, and vice versa. These results do not depend on the distribution of times spent in the infectious categories (the survivorship functions). Furthermore, by keeping all the key statistics fixed, we can compare two extremes: exponential survivorship versus piecewise constant survivorship (individuals remain infectious for a fixed length of time). By choosing some realistic parameters we can see (at least in these cases) that the reproductive numbers corresponding to these two extreme cases do not differ significantly whenever the two distributions have the same mean. At any rate a formula is provided that allows us to estimate the role played by the survivorship function (and hence the incubation period) in the global dynamics of HIV. These results support the conclusion that single population models of this type are robust and hence are good building blocks for the construction of multiple group models. Our understanding of the dynamics of HIV in the context of mathematical models for multiple groups is critical to our understanding of the dynamics of HIV in a highly heterogeneous population.     

APOBEC3G与Vif在HIV-1感染中的作用

载脂蛋白B mRNA编辑催化多肽样（apolipoprotein B mRNA-editing catalytic polypeptide-like,APOBEC）蛋白是一组胞嘧啶脱氨基酶,具有天然的抗病毒活性,对多种病毒具有抑制作用,特别是逆转录病毒. APOBEC3蛋白能够抑制人类免疫缺陷病毒(HIV-1)的感染,其中APOBEC3G和APOBEC3F的作用最强. APOBEC3G能够通过胞嘧啶脱氨基作用和非胞嘧啶脱氨基作用抑制病毒感染. HIV-1病毒感染因子(Vif) 蛋白主要经泛素-蛋白酶体途径介导APOBEC3G降解,从而拮抗其抗病毒作用. APOBEC3G和Vif之间相互作用的研究对于寻求新的抗HIV治疗靶点具有重要意义.     

Optimized Infectivity of the Cell-Free Single-Cycle Human Immunodeficiency Viruses Type 1 (HIV-1) and Its Restriction by Host Cells

The infectivity of retroviruses such as HIV-1 in plasma or cultured media is less than 0.1% in general, the mechanisms of which are not yet fully understood. One possible explanation among others is the potential presence of large numbers of defective virions in a virus pool, which limits the apparent infectivity of HIV virions. To test this hypothesis, we have varied the culture conditions used to generate single-cycle HIV-1 virions. Among these culture variables, virion harvest time, media change after transfection, and envelope plasmid input can all improve HIV-1 infectivity by reducing the number of defective virions. A harvest time of 18–24 hours post transfection as opposed to 48 hours, and a media change six hours post transfection both improve viral infectivity. An optimal quantity of envelope plasmid input during transfection was also found. Collectively, these conditions increased the infectivity of HIV-1 virions by sevenfold compared to normally reported values in TZM-bl indicator cell lines. These conditions also increased the infectivity of HIV-1 in CD4⁺ T cells, suggesting that these conditions work by increasing the intrinsic infectivity of a virus pool. Nevertheless, these improvements on virion infectivity were marginal compared to the impact of host cells on HIV infection, which can decrease the apparent infectivity by 19-fold even for the most optimized viruses. These results suggest that the infectivity of HIV-1 virions can be optimized by reducing the number of defective virions; however, viral-cell interactions may pose a major barrier for HIV-1 infectivity.     

HIV(+)/AIDS并发肺结核与单纯肺结核患者临床对照分析

提高对HIV(+)/AIDS并发肺结核临床表现的认识。方法:选取1998年以来我院收治的HIV(+)/AIDS并发肺结核的病例共47例为观察组(A组),及同期住院的HIV(-)单纯肺结核病例50例为对照组(B组)进行回顾性对照分析。结果:(1)A组发烧和体重下降较B组更常见,而咳嗽和咯血较B组少见;(2)A组痰抗酸杆菌阳性率显著低于B组;(3)A组结核杆菌培养阳性率显著低于B组,而耐多药结核(MDR-TB)所占比率显著高于B组;(4)肺结核的X线表现为弥漫性浸润或粟粒性阴影的,A组多于B组,而A组影像学空洞率显著低于B组;(5)A组合并肺外结核较B组多见,A组淋巴系统较B组常发生结核病变,A组全身血液播散性结核病的发病率明显高于B组;(6)PPD结核菌素反应阳性率A组显著低于B组,A组PPD阳性率与外周CD4+T淋巴细胞数相关。结论:HIV(+)/AIDS患者并发肺结核临床表现不典型。     

多聚TAR-CoreRNA诱饵对人免疫缺陷病毒1型Tat蛋白活性的抑制作用

为了抑制Ｔａｔ蛋白的反式激活作用,在细胞内大量表达外源ＴＡＲＲＮＡ使其与Ｔａｔ蛋白结合,从而竞争性抑制其与ＨＩＶ－１ＬＴＲ的ＴＡＲＲＮＡ元件结合．构建了以ＨＩＶ－１ＬＴＲＹＬ１５８（－１５８～＋１８０）为启动子,分别含有４,８和１５个拷贝的ＴＡＲ－ＣｏｒｅＲＮＡ诱饵（ｄｅｃｏｙ）表达质粒;以荧光酶基因为报告基因,检测了瞬时共转染体系中含不同拷贝数的ＴＡＲ－ＣｏｒｅＤＮＡ转录产物对Ｔａｔ蛋白反式激活作用的影响．结果证明,ＴＡＲ－ＣｏｒｅＲＮＡ诱饵对Ｔａｔ蛋白活性具有很强的抑制作用,其抑制程度与ＴＡＲ－ＣｏｒｅＤＮＡ串联体的拷贝数有关．     

The role of HIV replicative fitness in perinatal transmission of HIV

Perinatal transmission of Human immunodeficiency virus(HIV),also called mother-to-child transmission(MTCT),accounts for 90% of infections in infants worldwide and occurs in 30%-45% of children born to untreated HIV-1 infected mothers.Among HIV-1 infected mothers,some viruses are transmitted from mothers to their infants while others are not.The relationship between virologic properties and the pathogenesis caused by HIV-1 remains unclear.Previous studies have demonstrated that one obvious source of selective pressure in the perinatal transmission of HIV-1 is maternal neutralizing antibodies.Recent studies have shown that viruses which are successfully transmitted to the child have growth advantages over those not transmitted,when those two viruses are grown together.Furthermore,the higher fitness is determined by the gp120 protein of the virus envelope.This suggests that the selective transmission of viruses with higher fitness occurred exclusively,regardless of transmission routes.There are many factors contributing to the selective transmission and HIV replicative fitness is an important one that should not be neglected.This review summarizes current knowledge of the role of HIV replicative fitness in HIV MTCT transmission and the determinants of viral fitness upon MTCT.