两种人化SCID小鼠的建立及其免疫应答性

将人外周血淋巴细胞（ＰＢＬ）和腹腔淋巴结淋巴细胞（ＰＬＮＬ）移植入严重联合免疫缺陷疾病（ＳｅｖｅｒｅＣｏｍｂｉｎｅｄＩｍｍｕｎｏｄｅｆｉｃｉｅｎｔＤｉｓｅａｓｅ，ＳＣＩＤ）小鼠腹腔，建立人化ＳＣＩＤ小鼠（即ＳＣＩＤ－ｈＰＢＬ，ＳＣＩＤ－ｈＰＬＮＬ）。两个月后，两种人化小鼠体内淋巴器官都可以检测到人Ｔ、Ｂ淋巴细胞分布；小鼠血清人源性ＩｇＧ和抗ＥＢ病毒壳抗原ＩｇＧ抗体水平（ＩｇＧ／ＶＣＡ）比较显示，用Ｂ９５８死细胞作为ＶＣＡ来源所诱导的实验组１０只小鼠，ＩｇＧ／ＶＣＡ阳性率为７０％（７／１０），对照组为１７％（２／１２）。１４只ＳＣＩＤ－ｈＰＬＮＬ小鼠血清内人ＩｇＧ／ＶＣＡ的几何平均滴度（ＧＭＴ）和血清ＩｇＧ浓度在１∶１０８和９６．２±５６．４μｇ／Ｌ；在另８只ＳＣＩＤ－ｈＰＢＬ中为１∶７．９和１３．８４±６．０μｇ／Ｌ。实验提示，ＳＣＩＤ－ｈＰＬＮＬ小鼠较ＳＣＩＤ－ｈＰＢＬ小鼠更适合于人源性特异ＩｇＧ的诱导。     

HLA多态性与HIV感染及AIDS发病相关性的研究

为研究人类白细胞抗原（ＨＬＡ）和人免疫缺陷病毒（ＨＩＶ）易感性的关系，本文分析比较了以下三组人群的ＨＬＡ表型频率和单倍型频率：①１７２例正常人；②１７例血清ＨＩＶ阴性高危人群；③１８０例血清ＨＩＶ阳性患者，其中２１例发展为艾滋病（ＡＩＤＳ），３７例６个月内ＣＤ４＋淋巴细胞降低至少２０％（ＣＤ４ｄｅｃｌｉｎｅ）。在１７２例对照和１８０例血清ＨＩＶ阳性受试者的比较中发现，其ＨＬＡ表型频率和单倍型频率没有显著差别，提示ＨＩＶ感染与ＨＬＡ无关联。然而，ＨＬＡ－Ｂ２１、ＨＬＡ－Ｂ８、ＨＬＡ－Ｂ３５表型及ＨＬＡ－Ａ１－Ｂ８、ＨＬＡ－Ｂ８－ＤＲ３、ＨＬＡ－Ａ１－Ｂ８－ＤＲ３单倍型与ＣＤ４阳性淋巴细胞下降，或与血清ＨＩＶ感染发展成ＡＩＤＳ病显著相关。１７例ＨＩＶ血清阴性高危人群中，无一携带单倍型ＨＬＡ－Ａ１－Ｂ８－ＤＲ３，提示该单倍型可能与ＨＩＶ感染的抗性相关。     

Epstein-Barr病毒壳抗原的纯化及其在检查血清抗体中的应用

为了建立纯化Ｅｐｓｔｅｉｎ－Ｂａｒｒ（ＥＢ）病毒壳抗原（ＥＢＶ－ＶＣＡ）的方法，用于酶联免疫吸附试验（ＥＬＩＳＡ）检测人血清中的相关抗体，我们用重组昆虫病毒在感染的ｓｆ９细胞中表达ＥＢＶ－ＶＣＡ。感染的细胞经裂解后，层析纯化表达产物。用纯化的ＶＣＡ作为抗原包被ＥＬＩＳＡ板或硝基纤维膜，检查血清中ＶＣＡ／ＩｇＡ和ＶＣＡ／ＩｇＧ抗体，为ＥＢＶ感染的检测和ＮＰＣ的诊断发展了一个敏感、特异和简便的方法。     

Epstein—Barr病毒壳抗原的纯化及其在检查血清抗体…

为了建立纯化Ｅｐｓｔｅｉｎ－Ｂａｒｒ（ＥＢ）病毒壳抗原（ＥＢＶ－ＶＣＡ）的方法，用于酶联免疫吸附试验（ＥＬＩＳＡ）检测人因清中的相关抗体，我们用重组昆虫病毒在感染的ｓｆ９细胞中表达ＥＢＶ－ＶＣＡ。感染的细胞经裂解后，层析纯化表达产物。用经的ＶＣＡ作为抗原包虫被ＥＬＩＳＡ板或硝基纤维膜，检查血清中ＶＣＡ／ＩｇＡ和ＶＣＡ／ｇＧ抗体，为ＥＢＶ感染的的检测和ＮＰＣ的诊断发展了一个敏感、特异和简便的方法。     

腺病毒、合胞病毒对外周血单个核细胞的IL-2受体表达及TNFα产生的影响

近年来国内外均注意到细胞因子及受体与病毒感染的关系。作者用１００ＴＣＩＤ５０的７型腺病毒（ＡＤＶ）及呼吸道合胞病毒（ＲＳＶ）Ｌｏｎｇ株刺激正常人体外培养的外周血淋巴细胞（ＰＢＭＣ），ＡＰＡＡＰ法检测淋巴细胞ＩＬ－２受体阳性率，酶联免疫法检测淋巴细胞上清液的ＴＮＦａ。初步观察了ＡＤＶ、ＲＳＶ对人ＰＢＭＣ的ＩＬ－２受体表达、ＴＮＦα产生的影响。经２００Ｕｇ／ｍｌ的ＰＨＡ活化的ＰＢＭＣ加入ＡＤＶ、ＲＳＶ后对照组ＩＬ－２＋细胞百分率为３４．３％，ＡＤＶ组为１７．３％，ＲＳＶ组为１７％，较对照组均显著降低…     

抗病毒蛋白MxA的诱导和检测方法的实验研究

目的 研究抗病毒蛋白ＭｘＡ的表达及其活性。方法用ＩＦＮα２ｂ或３型腺病毒（Ａ＿３）分别对ＷＩ－３８细胞或人外周血单个核细胞（ＰＢＭＣｓ）作用 １２或２４ ｈ，并用Ｗｅｓｔｅｍ ｂｌｏｔ法或ＦＡＣＳ法，分别对Ｍ蛋白的表达进行定性和定量检测。采用微量细胞病变抑制法，对重组的ＭｘＡ和ＩＦＮα２ｂ进行抗病毒效应实验。结果 （１）低浓度的 ＩＦＮα２ｂ（＞ １×1０～＃ＩＵ／Ｌ）和Ａｄ＿３（＞２００ ＴＣＩＤ＿５０），均可诱导相应细胞表达 ＭｘＡ；（２）１０μｇ／Ｌ ＭｘＡ可抵抗２０个 ＴＣＩＤ＿（５０）的 ＨＳＶ－Ｉ、Ｐｏｌｉｏ． Ｖ感染 Ｖｅｒｏ细胞、Ａｄ＿３感染ＨｅＬａ细胞，抵抗２００个ＴＣＩＤ＿（５０）的ＶＳＶ感染Ｗｉｓｈ细胞。结论ＭｘＡ只能由干扰素（ＩＮＦα2ｂ）或病毒诱导产生，其具有广谱的抗病毒活性。     

从器官移植受者PBMC中分离鉴定HHV┐

从器官移植受者ＰＢＭＣ中分离鉴定ＨＨＶ┐６罗敏华施凯肖扬名人类疱疹病毒６型（ＨＨＶ－６）是１９８６年由Ｓａ－ｉａｈｕｄｄｉｎ首先从淋巴增生及ＡＩＤＳ病人外周血单个核细胞（ＰＢＭＣ）中分离到的新型疱疹病毒。该病毒在正常人群中感染率很高，在免疫功能受抑制...     

HIV感染者肠粘膜病毒特异性抗体产生和多克隆B细胞激活

ＨＩＶ感染者肠粘膜病毒特异性抗体产生和多克隆Ｂ细胞激活［英］／ＥｒｉｋｓｓｏｎＫ…／／ＡＩＤＳ．—１９９５，９（７），—６９５～７００为探讨ＨＩＶ感染患者的粘膜Ｂ细胞活化的情况，该作者检测了ＨＩＶ感染患者肠粘膜总的抗体分泌细胞（ＡＳＣ）和ＨＩＶ—特异...     

动物模型与HIV／CNS相关疾病

ＨＩＶ－１感染人体中的神经系统（ＣＮＳ）的神经病变的研究难度，导致急需ＨＩＶ－１感染ＣＮＳ的动物模型，已建立的模型可概述ＨＩＶ／ＣＮＳ疾病的某些层面。ＳＩＶ感染恒河猴导致神经病理改变及神经行为功能失调，ＦＩＶ感染猫引起感觉中枢电生理变化及睡眠紊乱，鼠白血病病毒混合感染小鼠，其脑和行为变化与人某些改变有比较价值，ＳＣＩＤ鼠接受ＨＩＶ感染的人体器官或细胞嫁接（ＨｕＳＣＩＤ）它们经历的神经病变与人类死于     

CVIDB细胞增殖和分化功能

ＣＶＩＤＢ细胞增殖和分化功能耿排力，金子英雄，近藤直实（青海医学院微生物免疫学教研室西宁８１０００１）近年对常见多变型免疫缺陷（ＣＶＩＤ）的发病机制进行了大量研究。我们用ｒＩＬ－２刺激经金黄色葡萄球菌ｃｏｗａｎＩ株（ＳＡＣ）活化的健康人及ＣＶＩＤ患儿...     

Frequency of Tk and Hprt lymphocyte mutants and bone marrow micronuclei in mice treated neonatally with zidovudine and didanosine

The nucleoside analog zidovudine (3'-azido-3'-deoxythymidine, AZT), by itself or in combination with other anti- retroviral drugs, is used perinatally to prevent mother to child transmission of human immunodeficiency virus type 1. AZT is mutagenic in vitro and mutagenic and carcinogenic when administered to neonatal mice. A previous study indicated that the anti-retroviral agent didanosine (2',3'-dideoxyinosine, ddI) potentiated the mutagenicity of AZT in the thymidine kinase (TK) gene of cultured human TK6 lymphoblastoid cells. We have evaluated whether or not ddI affects the in vivo genotoxicity of AZT by breeding C57Bl/6N/Tk+/- female mice with C3H/HeNMTV male mice and treating the offspring daily on postnatal days 1-8 with 200 mg/kg ddI alone or in combination with 200 mg/kg AZT. One day after the last dose, bone marrow polychromatic erythrocytes (PCEs) were obtained to assess the induction of micronuclei; 3 weeks following treatment, the induction of mutants was determined in the hypoxanthine-guanine phosphoribosyltransferase (Hprt) and Tk genes of splenic T lymphocytes from B6C3F1/Tk+/- mice. The mixture of AZT and ddI, but not ddI alone, caused a significant increase in micronucleated PCEs. When assessed 3 weeks after dosing, ddI did not induce mutations in the Hprt or Tk genes. The mixture of AZT and ddI also did not induce mutations in the Hprt gene, but did induce a significant increase in Tk mutants, similar to that observed previously with AZT alone. The induction of mutations in the Tk gene by the mixture of AZT and ddI was associated with loss of the wild-type Tk+ allele. These data indicate that, under the conditions of this experiment, ddI is not mutagenic in neonatal B6C3F1/Tk+/- mice and that it does not potentiate the mutagenicity of AZT.     

Genetic damage detected in CD-1 mouse pups exposed perinatally to 3'-azido-3'-deoxythymidine or dideoxyinosine via maternal dosing, nursing, and direct gavage: II. Effects of the individual agents compared to combination treatment

We previously reported extraordinary increases in micronucleated erythrocytes in CD-1 mouse pups exposed to 3'-azido-3'-deoxythymidine (AZT) and dideoxyinosine (ddI; 50/250, 75/375, 150/750 mg/kg/day AZT/ddI) by gavage throughout gestation and lactation, followed by direct pup dosing beginning postnatal day (PND) 4 (Bishop et al. [2004]: Environ Mol Mutagen 43: 3-9). That study was conducted to explore the potential for genetic damage in newborns exposed perinatally to antiretrovirals in order to reduce maternal-infant transmission of HIV-1. Because dramatic increases in frequencies of micronucleated erythrocytes were seen in exposed pups, additional studies were conducted to clarify the relative contribution of each drug to the observed damage. Pregnant CD-1 mice were administered AZT (50, 75, 150 mg/kg/day) or ddI (250, 375, 750 mg/kg/day) by gavage twice daily in equal fractions beginning prior to mating and continuing throughout gestation and lactation. Direct pup dosing (same regimens) began on PND 4. Peripheral blood erythrocytes of male pups were screened for micronuclei on PNDs 1, 4, 8, and 21. Significant increases in micronucleated erythrocytes were observed in pups and dams exposed to AZT at all doses and sampling times. The highest micronucleus levels were observed in pups on PND 8 after the initiation of direct dosing. In contrast, effects seen in pups and dams treated with ddI were minimal. These results demonstrate that AZT, a component of many anti-HIV combination therapies, induces chromosomal damage in perinatally exposed neonatal mice. Comparison of micronucleated cell frequencies induced by AZT alone or in combination with ddI suggests that ddI potentiates AZT-induced chromosomal damage following direct exposure.     

Clinical and immunologic effects of combination therapy with intravenous immunoglobulins and AZT in HIV-infected patients

30 patients with HIV infection were enrolled to evaluate the clinical efficacy and toxicity of zidovudine (AZT), 0.5 g/day p.o. (Group A) vs. AZT 0.5 g/day p.o. plus intravenous immunoglobulins (IVIG), 0.4 g/kg of body weight for three consecutive days, followed by one treatment of 0.6 g/kg of body weight every fourth week (Group B), over a period of one year. The study was open and randomized. The treatment groups were compared using the following study variables: 1) type of infections, recurrences and severity; 2) change in CD4+ T and CD8+ T cell count; 3) change in platelet count; 4) change in TNF alpha serum levels; 5) the probability of not developing an opportunistic infection over a period of 12 months. Patients from Group B developed less pathological events in comparison to Group A. No significative differences were evident with regard to values of T cell subsets obtained before and after treatment in each group and between the two groups. On the contrary, in 12 out of 15 patients from Group B there was a significant increase in platelet count. In both groups there was a significant decrease of mean serum levels of TNF alpha when a comparison was made between time 12 vs. time 6. However, when data were expressed as single values, in three subjects from Group B TNF alpha was still detectable by time 12 vs. 9 individuals in Group A. The cumulative probabilities of developing an opportunistic infection over the 12 months of treatment in the Group A subjects were significantly higher than in the Group B subjects (p less than 0.01). Adverse effects--nausea and gastric pain--were reported for 3 individuals (20%) from Group A and 4 patients (26%) from Group B. In conclusion, patients treated with AZT are especially likely to benefit from IVIG prophylaxis.     

Effect of combination interleukin-3 (IL-3) and granulocytemacrophage colony stimulating factor (GM-CSF) on hematopoiesis administered to retrovirus-infected immunodeficient mice receiving dose-escalation zidovudine (AZT)

We have previously demonstrated that continuous administration of dose-escalation zidovudine (AZT) in either normal or LP-BM5 MuLV immunodeficient virus-infected mice (MAIDS) was associated with the development of anemia, neutropenia, and thrombocytopenia. Hematopoietic growth factors/cytokines are being evaluated to determine their efficacy in ameliorating the hematopoietic toxicity associated with AZT. In normal mice receiving AZT, an increase in only plasma erythropoietin and not GM-CSF, Meg-CSF or TNF-α has been reported. This article describes studies that investigated the effect of combination interleukin-3 (IL-3) and granulocyte-macrophage colony stimulating factor (GM-CSF) administered in normal non-viral, viral-infected, and viral-infected C57BL6 mice receiving dose-escalation AZT, i.e. 0.1 mg/ml, 1.0 mg/ml, and 2.5 mg/ml placed in the drinking water. Non-viral control mice responded to IL-3/GM-CSF by increasing erythropoiesis, myelopoiesis and platelet production measured by increased bone marrow and spleen derived erythroid, myeloid and platelet precursor stem cells cultured in semi-solid media. Virus-infected control mice not receiving IL-3/GM-CSF developed pancytopenia. Administration of IL-3/GM-CSF to virus-infected mice receiving dose-escalation AZT did not ameliorate the peripheral pancytopenia associated with immunodeficiency disease and AZT treatment, even though erythroid, myeloid and platelet precursor progenitor cells were increased at certain times when compared to either normal or viral-infected mice receiving IL-3/GM-CSF. These results indicate that the combination use of IL-3 and GM-CSF in vivo is only a partially effective growth factor/cytokine treatment to ameliorate the hematopoietic toxicity associated with the use of the anti-viral drug zidovudine.     

清胰汤对L-精氨酸诱发的重症急性胰腺炎小鼠胰腺p-STAT3表达的影响

目的: 观察L-精氨酸(L-Arg)诱发的重症急性胰腺炎小鼠胰腺组织p-STAT3表达的变化,以及清胰汤对p-STAT3表达的影响,从而探讨STAT3在急性胰腺炎中的作用和清胰汤治疗急性胰腺炎的机制。方法: 健康雄性成年昆明种小鼠30只,随机分为3组(n=10):对照组、模型组和清胰汤组。除对照组外,其余各组给予腹腔注射20 % L-精氨酸(3 g/kg,间隔1 h再注射1次);清胰汤组在第2 次腹腔注射20 %L-Arg 30 min 后给予清胰汤浓缩液灌胃(10 mL/kg),之后每天灌胃2次。在造模后72 h麻醉处死动物检测血清淀粉酶活性;取胰腺组织计算胰腺湿重比,HE染色观察胰腺病理学改变;取肺组织匀浆检测髓过氧化物酶(MPO)的活性,HE染色观察肺病理学改变; Western blotting及real-time PCR分别检测胰腺组织p-STAT3蛋白及单核细胞趋化蛋白-1(MCP－1)mRNA的表达变化。结果: L-Arg诱发急性胰腺炎72 h后,血清淀粉酶活性明显升高、胰腺湿重比增加、肺组织MPO显著增加,与对照组比较差异显著(P<0.05);而清胰汤组血清淀粉酶的活性、胰腺湿重比、MPO水平明显降低,与模型组相比差异显著(P<0.01);模型组72 h胰腺及肺可见明显病理损伤,胰腺组织p-STAT3蛋白及MCP-1 mRNA的表达明显增强;清胰汤治疗组胰腺及肺病理损伤减轻,胰腺组织p-STAT3蛋白及MCP-1 mRNA的表达减少。结论: L-Arg诱发的重症急性胰腺炎小鼠胰腺组织STAT3蛋白表达明显增加,STAT3活化可能参与了L-Arg诱发的急性胰腺炎进展;抑制胰腺STAT3活化是清胰汤治疗急性胰腺炎的作用机制之一。     

Transplacental drug transfer and frequency of Tk and Hprt lymphocyte mutants and peripheral blood micronuclei in mice treated transplacentally with zidovudine and lamivudine

In previous studies, we have shown that zidovudine (3'-azido-3'-deoxythymidine; AZT), but not lamivudine [(-)2',3'-dideoxy-3'-thiacytidine; 3TC], is genotoxic when administered to neonatal mice, and that 3TC when coadministered with AZT does not alter the responses observed with AZT alone (Von Tungeln et al. [2002] Carcinogenesis 23:1427-1432). We now have investigated the transplacental transfer of these drugs and the induction of mutants and micronuclei in the neonatal offspring. From gestational day 12 until parturition, female C57BL/6N and C57BL/6N/Tk(+/-) mice, which had been mated to male C3H/HeNMTV mice, were treated daily by gavage with AZT, 3TC, or a combination of AZT and 3TC. In both dams and fetuses, AZT was found at much higher levels than its metabolites, AZT 5'-glucuronide and 3'-azido-3'-deoxythymidine. In the neonates, AZT and the mixture of AZT and 3TC caused a decrease in the percentage of reticulocytes (RETs) and an increase in the percentage of micronucleated RETs and micronucleated normochromatic erythrocytes. When assessed 3 weeks after birth, AZT and the combination of AZT and 3TC increased the thymidine kinase (Tk) mutant frequency in male mice; at 5 weeks, 3TC increased the Tk mutant frequency in female mice. The increase in Tk mutants in mice treated with AZT and the mixture of AZT and 3TC was associated with loss of the wild-type (Tk(+)) allele (loss of heterozygosity; LOH) and a pattern of discontinuous LOH. These data indicate that AZT, 3TC, and the combination of AZT and 3TC are transplacental mutagens and that the increase in mutants resulting from AZT is due mainly to large-scale genetic alterations.     

Mutagenicity of zidovudine, lamivudine, and abacavir following in vitro exposure of human lymphoblastoid cells or in utero exposure of CD-1 mice to single agents or drug combinations

Experiments were performed to investigate the impact of zidovudine (AZT), lamivudine (3TC), and abacavir (ABC) on cell survival and mutagenicity in two reporter genes, hypoxanthine-guanine phosphoribosyltransferase (HPRT) and thymidine kinase (TK), using cell cloning assays for assessing the effects of individual drugs/drug combinations in (1) TK6 human lymphoblastoid cells exposed in vitro and (2) splenic lymphocytes from male CD-1 mice exposed transplacentally on days 12-18 of gestation. In TK6 cells, dose-related increases in HPRT and TK mutant frequencies were found following 3 days of exposure to AZT or 3TC alone (33, 100, or 300 microM), or to equimolar amounts of AZT-3TC. Compared with single drug exposures, AZT-3TC coexposures generally yielded enhanced elevations in HPRT and TK mutant frequencies. Mutagenicity experiments with ABC alone, or in combination with AZT-3TC, were complicated by the extreme cytotoxicity of ABC. Exposure of cells either to relatively high levels of AZT-3TC short-term (100 microM, 3 days), or to peak plasma-equivalent levels of AZT-3TC for an extended period (10 microM, 30 days), resulted in similar drug-induced mutagenic responses. Among sets of mice necropsied on days 13, 15, or 21 postpartum, Hprt mutant frequencies in T-cells were significantly elevated in the AZT-only (200 mg/kg bw/day) and AZT-3TC (200 mg AZT + 100 mg 3TC/kg bw/day) groups at 13 days of age. These results suggest that the mutagenicity by these nucleoside analogs is driven by cumulative dose, and raises the question of whether AZT-3TC has greater mutagenic effects than AZT alone in perinatally exposed children.