人预存天然抗体与猪胰腺细胞的异种反应性研究

目的 分析人预存天然抗体与猪各种胰腺细胞的异种反应性及其同种型。方法 将患者和正常人血清预先加热灭活并按1：2、1：4、1：8、1：16、1：32倍比稀释,分别同猪胰腺冰冻切片于37℃下孵育30min,冲洗后组织进一步用免疫荧光标记的羊抗人IgG和IgG抗体于37℃下孵育30min。采用间接免疫荧光染色法检测人预存天然抗体与猪胰腺细胞的反应性。结果 55．6％的患者和55．0％的正常人体内均存在着抗猪内分泌细胞的天然抗体,但患者血清中强阳性的百分率更高。100％的患者和95．0％的对照血清都可同一种或多种胰腺细胞反应。预存天然抗体的同种型包括IgG和IgM两种。结论 鉴于预存抗体同猪胰腺细胞的强阳性反应,移植前行配型试验和移植物预处理对移植的胰岛素细胞的存活是必要的。     

猪后肾帽状间充质细胞发育为肾小球足细胞的过程

目的:探讨中国实验用小型猪肾小球足细胞的发育过程。方法:应用过碘酸-希夫氏染色观察中国实验用小型猪胚胎不同时间点(胚胎28d至出生后21d,以周为单位,共17个时间点)肾小球发育过程中的形态学变化。应用免疫荧光技术检测猪胚肾不同阶段(帽状间充质、肾小囊体、逗号形体、"S"形体、毛细血管袢期肾小球、成熟肾小球)足细胞的发育过程。结果:猪在胚胎第28天(E28d)后肾已开始发育,可见典型的帽状间充质、肾小囊体、逗号形体和"S"形体;E35d可见肾小球形成,包括近皮质的不成熟肾小球以及近髓质的成熟肾小球。胚肾组织免疫荧光染色显示:胚肾早期足细胞标志物WT1表达于Six2阳性的后肾帽状间充质细胞,相继表达于肾小囊体、整个逗号形体、逗号形体尾部以及"S"形体下端,最终局限于肾小球足细胞。结论:中国实验用小型猪的足细胞来源于Six2阳性的后肾帽状间充质细胞,经过肾小囊体、逗号形体、"S"形体、毛细血管袢期肾小球阶段,发育成为成熟肾小球的足细胞。     

猪抗人淋巴细胞免疫球蛋白联合环孢菌素A治疗重型再障的临床疗效

目的探讨使用国产猪抗人淋巴细胞免疫球蛋白（p-ALG）联合环孢菌素A（CsA）治疗重型再生障碍性贫血（SAA）的疗效及观察不良反应。方法应用p-ALG（30mg/kg/d,静脉滴注）联合CsA（起始量为5mg/kg/d）治疗SAA 8例,辅以促造血、成分血输注、造血生长因子等支持治疗。检测治疗前后外周血象及骨髓象变化,以流式细胞仪测定治疗前后外周血T淋巴细胞亚群的改变情况,观察不良反应及其处理效果。结果随访8例中,基本治愈2例,缓解3例,明显进步1例,早期死亡2例,总有效率75%。治疗显效时患者的外周血CD3＋CD4＋（Th）细胞比例升高,CD3＋CD8＋（Ts）细胞比例降低,Th/Ts比值升高,与治疗前比较,差异有统计学意义（P〈0.01）。结论使用国产p-ALG联合CsA作为初诊SAA的首次治疗方案效果良好,治疗费用较马或兔抗胸腺细胞免疫球蛋白（ATG）联合CsA方案更经济。     

Ex vivo decrease in uranium diffusion through intact and excoriated pig ear skin by a calixarene nanoemulsion

Cutaneous contamination by radionuclides is a major concern in the nuclear industry. In case of skin exposure to uranium, no efficient emergency treatment is available to remove the actinide from the skin. For this purpose, we developed a nanoemulsion containing calixarene molecules displaying good chelating properties towards uranium. In this paper, we describe the ability of this formulation to trap uranium and limit its transfer from the cutaneous contaminated site into the blood. Uranium percutaneous diffusion kinetics was assessed with Franz cells over 24 h through intact and excoriated pig ear skin biopsies, after or without application of the nanoemulsion. Uranium distribution in the skin layers was analysed by SIMS microscopy. The results showed that prompt application of the calixarene nanoemulsion allows a 94% and 98% reduction of the amount of uranium diffused respectively through intact and excoriated skin. The formulation is still efficient in case of delayed application up to 30 minutes since the 24 h-uranium transfer through excoriated skin is reduced by 71%. Besides, no accumulation of uranium or uranium-calixarene chelate was observed in the different skin layers. In conclusion, this study demonstrated the efficiency of the calixarene nanoemulsion, which can be regarded as a promising treatment for uranium cutaneous contamination.     

Luminal Salmonella Endotoxin Affects Epithelial and Mast Cell Function in the Proximal Colon of Pigs

Background: The effects of proximal small-bowel resection on absorption and synthesis of cholesterol are unclear. Methods: To study cholesterol absorption and synthesis after proximal gut resections of variable length, plasma plant sterols, cholestanol, and cholesterol precursors were measured 1 and 2 months after 50% and 75% proximal small-bowel resection or transection. To examine the effect of increased crypt cell proliferation and brush border development on cholesterol absorption, the results were related to the mucosal morphology, crypt cell proliferation, and disaccharidase activities of the remaining small bowel. Results: Campesterol levels in proportion to cholesterol decreased markedly more, and those of cholestanol markedly less, than would be expected simply due to the amount of proximal small intestine removed, whereas sitosterol proportions decreased in proportion to the length of gut resection. Campesterol proportions markedly (P = 0.06) increased between 1 and 2 months after 50% resection but remained unchanged after 75% resection. Crypt cell proliferation was only increased in the 75% resection group (P < 0.05). The longer the proximal gut resection, the lower was the mucosal enzyme activity. Both resection groups showed increased plasma cholesterol precursor proportions and crypt depth (P < 0.05), whereas villus height remained unchanged. After massive proximal resection campesterol and sitosterol proportions were inversely related to crypt cell proliferation (r = -0.86-0.83, P < 0.01). Conclusions: Increased crypt cell proliferation activated by massive proximal gut resection may act as a previously unrecognized factor in aggravating cholesterol malabsorption and retarding its recovery during the early postoperative period. These findings warrant further investigation.     

媒介动物(蚊、猪)体内黄病毒分子流行病学调查研究

〔目的〕通过对珠海注册养猪场及珠海口岸地区的蚊虫体内黄病毒的带毒率、宿主动物(猪群)及其人群进行血清黄病毒类(日本脑炎、登革热等)抗体水平的调查研究,为控制黄病毒在人群中的流行提供科学依据。〔方法〕采集养猪场猪血清、养猪场和口岸蚊标本、养猪场从业人员、入境的东南亚船员和城区部分发热病人血清,采用酶联免疫吸附试验(ELISA)法分别检测猪群和人群血清中的日本脑炎、登革热抗体;使用紫外灯诱蚊法在养猪场和口岸捕捉蚊类,经鉴定种类后,按20只蚊一组制成蚊悬液后,采用细胞培养、逆转录-聚合酶链反应(RT-PCR)以及荧光定量PCR技术检测蚊体内日本脑炎病毒、登革和西尼罗病毒等黄病毒的携带情况。〔结果〕1.猪血清日本脑炎病毒IgG抗体:结果已发表于《中国国境卫生检疫杂志》2007年第3期《珠海市部分注册猪场日本脑炎调查研究》。2.蚊媒带毒情况检测:2005年1月～2006年12月共捕获的2763只成蚊,经鉴定隶属4属8种,其中白纹伊蚊所占比例最高(32.75%),其次为致倦库蚊(26.06%)、三带喙库蚊(25.30%)、海滨库蚊(7.75%)、中华按蚊(3.84%)、骚扰阿蚊(3.18%),其他(常型曼蚊和巨型阿蚊等)(1.12%);在132组蚊研磨液的病毒细胞培养中,有7份标本出现CPE病变,使用黄病毒通用引物,RT-PCR反应能扩增出相应的阳性条带,其中白纹伊蚊组阳性3份,致倦库蚊组阳性3份,三带喙库蚊组阳性1份,但用日本脑炎病毒、登革病毒Ⅰ～Ⅳ型和西尼罗病毒等特异性引物,未能扩增出相应的阳性条带,说明可能存在着其它黄病毒类的病毒感染;采用荧光定量PCR法,对蚊悬液和细胞培养液日本脑炎、登革和西尼罗病毒进行检测,从1组海滨库蚊标本中检出日本脑炎病毒核酸阳性,但强度较弱,这与珠海地区是乙脑低发地区相一致。3.人群登革热和日本脑炎血清抗体检测:从东南亚的入境船员、本地发热病人和养猪场从业人员等血清标本511份中,登革病毒抗体IgG总阳性率为7.24%,其中以东南亚的入境船员的阳性率最高,为12.76%,没有检出登革病毒抗体IgM和日本脑炎IgM抗体。〔结论〕珠海地区存在日本脑炎的主要传播媒介,并且在蚊媒中携带有日本脑炎病毒;蚊标本细胞培养出现细胞病变和RT-PCR扩增出黄病毒基因的特异性片段,提示在捕获蚊标本中可能存在着其他病毒感染的可能;虽然养猪场猪群日本脑炎感染率不高,但存在着日本脑炎病毒的隐性感染或曾经感染过,提示不能够放松对乙型脑炎的预防控制工作,应采取积极的预防措施,防止人群和猪场日本脑炎的发生与流行。此外,研究结果表明东南亚船员有较高的登革热感染率,因此,在登革热流行期间,我国口岸应加强对来自东南亚国家交通工具员工和旅客的登革热的监测,以及有可能藏带、孳生蚊虫的废旧物品、集装箱等的检验检疫工作,以防止登革热的传入与流行。     

Regeneration of autotransplanted splenic tissue is not proportional to total weight and size of transplanted fragments

Young minipigs were used as a model for splenic autotransplantation in children. After splenectomy about 7% of the spleen was implanted in the greater omentum either as a single slice or as three small slices. Six months later there was no difference in the amount of regenerated splenic tissue or lymphocyte content between these two groups. The regenerated tissue weighed 3.98±1.9 g, which is not less than after whole spleen transplantation and is only about 6% of the splenic weight of control minipigs. The regenerated splenic tissue contained only about 3% of normal splenic lymphocyte numbers. Based on data from the literature on splenic regeneration in man and on the present experiments in animals, about 10% of the traumatized spleen seems to be sufficient for splenic regeneration after intraomental implantation. Offprint requests to: R. Pabst     

烫伤、内毒素对豚鼠胃肠动力影响的实验研究

目的:探讨烫伤后胃肠动力障碍发生机制.方法:豚鼠30只,随机分为烫伤、内毒素及对照组,在烫伤及内毒素腹腔注射1小时后测定豚鼠胃肠推进距离、肠道组织降钙素基因相关肽(CGRP)、Na -K 、Mg2 、Ca2 、及Ca2 -Mg2 -ATP酶含量.结果:①烫伤及内毒素组肠道推进功能受到抑制,烫伤组抑制更明显.②烫伤及内毒素组CGRP含量较对照组增多,烫伤组增高幅度更明显.③烫伤及内毒素组豚鼠肠道组织中各ATP酶的含量较对照组明显减少,两组ATP酶减少程度无明显差异.结论:①烫伤及内毒素抑制豚鼠的肠道推进功能,烫伤比内毒素组抑制更明显.②烫伤及内毒素组CGRP含量较对照组明显增多,烫伤组CGRP增高幅度更明显.③烫伤及内毒素组各ATP酶的含量较对照组减少,烫伤组、内毒素组各ATP酶减少程度无明显差异.     

Purification and characterization of human endopeptidase 3.4.24.16. Comparison with the porcine counterpart indicates a unique cleavage site on neurotensin

We have purified and characterized human brain endopeptidase 3.4.24.16. The enzyme behaved as a 72 kDa protein and belonged to the metalloprotease family. Human endopeptidase 3.4.24.16 cleaved neurotensin at a unique site at the Pro¹⁰-Tyr¹¹ bond, leading to the formation of neurotensin(1–10) and neurotensin(11–13). The kinetic parameters displayed by human endopeptidase 3.4.24.16 towards a series of natural neuropeptides indicated that bradykinin was the most efficiently proteolysed. Angiotensin I, dynorphins 1–8 and 1–9 and substance P also behaved as good substrates while neuromedin N, angiotensin II, leucine and methionine enkephalin and neurokinin A resisted degradation by human endopeptidase 3.4.24.16. We have purified the porcine counterpart of endopeptidase 3.4.24.16 and compared its ability to cleave neurotensin with that of the enzyme from human origin. It appeared that, besides a major production of neurotensin(1–10), an additional formation of neurotensin(1–8) was observed with the pig enzyme, suggesting a cleavage of neurotensin not only at the Pro¹⁰-Tyr¹¹ bond but also at the Arg⁸-Arg⁹ peptidyl bond. The latter cleavage appeared reminiscent of endopeptidase 3.4.24.15 since this peptidase was reported to cleave neurotensin at the Arg⁸-Arg⁹ bond. Our study indicated that neurotensin(1–10) formation by porcine endopeptidase 3.4.24.16 could be potently blocked with the selective endopeptidase 3.4.24.16 dipeptide inhibitor Pro-Ile without interfering with neurotensin(1–8) formation. By contrast, the formation of the latter product was highly potentiated by dithiothreitol and inhibited by the endopeptidase 3.4.24.15 inhibitor Cpp-Ala-Ala-Tyr-pAB, two effects that were not observed for neurotensin(1–10) production. Altogether, our results indicate that porcine endopeptidase 3.4.24.16 cleaves neurotensin at a unique site, leading to the formation of rteurotensin(1–10) and that the production of neurotensin(1–8) is due to contaminating endopeptidase 3.4.24.15.     

An analysis of methylphenidate induced gnawing in guinea pigs

20 compounds acting on the central nervous system and the autonomic nervous system were tested in guinea pigs for their ability to induce gnawing. Only methylphenidate induced vigorous gnawing similar to that produced by apomorphine and amphetamine. Methylphenidate differs in its mode of action from both apomorphine and amphetamine. In the guinea pig, phenylethyl configuration with OH groups at para and meta positions of the phenyl ring does not seem to be an essential criterion for inducing gnawing as suggested for the rat (Ernst, 1965). Catecholamines do not appear to play any significant role in the mediation of methylphenidate gnawing, or even in the gnawing response itself in guinea pigs, since increase in the level of dopamine and other catecholamines does not induce gnawing.Communication No. 1394 of Central Drug Research Institute, Lucknow, India.