明天的医学向何处去——平衡医学方法论略谈

《明天的医学向何处去》一文是王佑三同志应中国中医研究院、中西医结合研究会1990～2010年中医药研究的重大进展与突破预测研究课题组和中国人体科学学会之约所撰。本报征得作者同意稍加删节后发表,期望引起对“明天的医学”的关注。     

自发免疫耐受大鼠移植肝内调节性T细胞的生物学特性

目的研究大鼠肝移植后自发免疫耐受的形成与移植肝内CD4~ CD25~ 调节性T细胞(Tr细胞)的关系。方法按供、受者不同将实验分为3组。急性排斥组:DA大鼠为供者,LEW大鼠为受者;自发耐受组:LEW大鼠为供者,DA大鼠为受者;同基因组:供、受者均为DA大鼠。各组均建立大鼠原位肝移植模型。分别在肝移植术后4、7、14、30和90 d时采用密度梯度离心法分离移植肝内淋巴细胞,免疫磁珠分离(MACS)法分选出CD4~ CD25~ Tr细胞;用流式细胞术(FCM)检测细胞纯度,同时分析CD4~ CD25~ Tr细胞比例的变化;体外细胞增殖试验研究CD4~ CD25~ Tr细胞对CD4~ CD25~-T细胞的免疫抑制作用。结果肝移植早期,急性排斥组和自发耐受组移植肝内CD4~ CD25~ Tr细胞比例均明显增加,其中急性排斥组增加更为明显;移植后4 d左右,两组CD4~ CD25~ Tr细胞比例开始下降,急性排斥组的下降幅度较大;移植后30 d,自发耐受组受者的移植肝内CD4~ CD25~ Tr细胞比例达到第2次高峰,约在移植后90 d时下降至正常生理水平。移植后7 d左右,急性排斥组受者均因发生排斥反应而死亡,而自发耐受组受者均存活。此外,CD4~ CD25~ Tr细胞能有效抑制CD4~ CD25~-T细胞的增殖。结论CD4~ CD25~ Tr细胞是一种具有特异免疫调节功能的T细胞亚群,其主动的免疫抑制功能可能是诱导大鼠肝移植自发免疫耐受的机制之一。     

EFFECTS OF ANGIOTENSINS II AND III ON GLOMERULOTUBULAR BALANCE IN RATS

1. The role of angiotensin as a modulator of proximal glomerulotubular (GT) balance was investigated in anaesthetized rats by examining the relationship between glomerular filtration rate (GFR) and absolute proximal reabsorption (APR) during removal of endogenous angiotensin II (AII) and III (AIII) with enalaprilat (CEI) and then during their subsequent replacement by intravenous infusions. 2. Enalaprilat lowered mean arterial blood pressure (MABP) and increased renal blood flow (RBF), GFR, urine flow rate and sodium excretion. Filtration fraction (FF) was not altered. Absolute proximal reabsorption, derived from fractional lithium clearance, increased by only 48% of the change expected for 'perfect' GT balance. 3. Angiotensin II replacement corrected MABP, GFR and plasma renin level, but reduced RBF and increased FF; APR was decreased and GT balance was restored. Urine flow and sodium excretion remained above control values with AII. 4. Replacement with AIII did not correct the hypotension but completely reversed the renal and renin responses to enalaprilat and restored GT balance without affecting FF. 5. It was concluded that the relation between proximal reabsorption and GFR is considerably modified by the intrarenal angiotensin concentration. The findings are best explained by a direct stimulation of proximal tubular sodium transport by angiotensin at the concentrations existing in anaesthetized rats.     

Methionine-induced acidosis in the weanling rat

Whole body balances of non-metabolizable base (NB) were studied in weanling rats fed a cereal-based diet with or without added l -methionine. In response to l -methionine loading (2.5 mmol day^-1) the rats exhibited a significant reduction in rates of food intake (109 vs. 160 g per 8 days) and body growth (3 vs. 52 g per 8 days); fractional oxidation of absorbed dietary amino acid sulphur increased from 0.28 to 0.64; and mean urinary sulphate excretion increased from 2.3 to 14.8 mmol per 8 days. Mean rates of renal excretion of NB and filtered titratable organic acid decreased from 20 to ?11 mmol per 8 days and from 22 to 8 mmol per 8 days. Balances of calcium and NB remained at reference values despite a decrease in whole blood ‘base excess’ from ?1.0 to ?6.4 mmol 1–¹. The concentration of NB in plasma rose but slightly. It is concluded that L-methionine loading in the weanling rat leads to extracellular non-carbonic acidosis subject to renal compensation. This acidosis is due not to retention of H⁺ released by ionization of endogenous sulphuric acid but possibly to accumulation of (acid) organic metabolites of methionine which are efficiently conserved by the kidneys. The rise in sulphuric acid production leads to an adaptive decrease in fractional reabsorption of filtered sulphate. Even during inhibited growth, absorbed dietary NB is retained and deposited in the skeleton, probably as calcium carbonate.     

感染性休克患者的负液体平衡与预后的关系

目的探究感染性休克治疗的前3天患者的液体平衡状态与预后的关系。方法采用回顾性对照研究。查看1999年1月至2003年12月收入首都医科大学附属复兴医院ICU的感染性休克患者病例记录,入选病例必须严格符合感染性休克的诊断标准,且既往无肾功能不全病史。采集病例相关数据以及诊断后第1、2、3天的液体平衡值。比较不同组别患者的急性生理和慢性健康评分(APACHEⅡ)、继发器官衰竭评分(SOFA)、液体平衡和病死率等数据。对影响患者预后的独立危险因素进行Logistic回归分析,确定和描述感染性休克患者的预后与在前3天的液体复苏治疗中出现的负平衡((0mL)相关因素的关系。结果负液体平衡患者与未出现负液体平衡患者2组的病死率差异有统计学意义(52.4%vs87.5%,χ2=5.303,P=0.021)。通过对入组时患者年龄、APACHEⅡ评分、第1天和第3天SOFA评分和正负平衡等影响患者预后的独立危险因素Logistic回归分析,表明前3天的治疗中,若有1d出现负液体平衡即可成为影响患者预后的独立危险因素(P=0.035)。结论在感染性休克前3天的治疗中,若有1d出现液体平衡负值即可成为影响感染性休克患者预后的独立因素,对感染性休克的28d生存预后有较强的预测性。在前3天的治疗中出现液体平衡为负值((0mL)的感染性休克患者的生存率比液体平衡为正值的患者的28d生存率高。     

Long-term neuroretinal full-thickness transplants in a large animal model of severe retinitis pigmentosa

Background The purpose of this study was to explore neuroretinal transplantation in a large animal model of severe retinitis pigmentosa and to establish graft development, long-term survival, graft-host integration, and effects on the host retina. Methods Rhodopsin transgenic pigs, aged 6 months, received in one eye a fetal full-thickness neuroretinal sheet in the subretinal space by means of vitrectomy and retinotomy. Six months postoperatively, eyes were studied in the light microscope and with immunohistochemical markers. Full-field electroretinography (ERG) was performed at 4 and 6 months. Results Laminated grafts with well-organized photoreceptors, rod bipolar cells, and Müller cells were found in five of six eyes. Neuronal connections between graft and host retina were not seen. In the five eyes containing a graft, the number of surviving rods in the host retina was significantly higher compared with unoperated eyes. The ERG did not reveal any significant difference in b-wave amplitude between operated and control eyes, but the cone-derived response in operated eyes increased significantly from 4 to 6 months while the rod response in control eyes decreased significantly. Conclusions Fetal full-thickness neuroretina can be transplanted safely to an eye with severe retinal degeneration. In their major part, the transplants develop a normal laminated morphology and survive for at least 6 months. Graft and host retinal neurons do not form connections. Retinal function in the host is reduced initially by the surgical trauma, but the presence of a well-laminated graft counteracts this effect and rescues rods from degeneration. Supported by The Foundation Fighting Blindness (grant# C-NC02-798-0078), The Faculty of Medicine, University of Lund, The Swedish Research Council, The Princess Margaretas Foundation for Blind Children, The 2nd ONCE International Award for New Technologies for the Blind.     

Characterization of the immunological memory state generated in mice susceptible to Leishmania major following exposure to low doses of L. major and resulting in resistance to a normally pathogenic challenge

BALB/c mice are susceptible to a high-dose infection of the protozoan Leishmania major, which induces a parasite-specific antibody, Th2-like response, exclusive of a significant and protective cell-mediated Th1 component. We have shown, in contrast, that infection with a low number of parasites induces cell-mediated immunity exclusive of antibody production, and results in resistance to substantial subsequent high-dose infection. Low-dose exposure thus constitutes effective vaccination. In the present study, we analyze lymphokine production by parasite-specific T cells from these low-dose exposed, resistant mice and from normal, susceptible mice following high-dose infection. Two findings stand out. First, the parasite-specific T cells in mice rendered resistant appear not to be in an activated, effector state at the time of parasite challenge, as assessed by lack of lymphokine production on short-term stimulation with parasite antigens, but to be rather in a memory state. Second, the ratio of parasite antigen-dependent production of interferon-γ to that of interleukin-4 by spleen cells of low-dose exposed and normal mice upon high-dose challenge takes a dramatically different course. This ratio is similar in both groups of mice shortly after challenge, but increases dramatically in the resistant and declines dramatically in the control mice over a period of weeks, such that these ratios differ by about 60-fold 12 weeks after the high-dose challenge. In addition, we show that a similar state of resistance occurs following low-dose infection with a more virulent strain of L. major. In toto, our observations suggest that resistance may be generally achievable by low-dose exposure and may be associated with a memory state which, when activated by parasite challenge, results in the evolution of the response over weeks such that the protective, Th1 component becomes ever more dominant over the Th2 component.     

冠状动脉粥样硬化性心脏病患者心外膜脂肪组织的生物信息学研究

背景 心血管疾病（CVD）是常见病和多发病,患病率和死亡率呈快速上升趋势。动脉粥样硬化（AS是缺血性CVD的病理基础,研究表明心外膜脂肪组织（EAT）通过分泌外泌体（EXO）和生物活性物质促进AS进展,但其作用机制仍需进一步研究。目的 通过生物信息学方法挖掘冠状动脉粥样硬化性心脏病（CAD）患者EAT中的关键基因,探讨免疫细胞浸润情况,联合CAD患者EXO间差异表达基因（DEGs）推测EAT来源EXO间DEGs并进行验证,从细胞及分子水平上探讨EAT在CAD疾病过程中的作用机制。方法 从基因表达综合数据库（GEO）中下载关于EAT的数据集GSE64554、GSE120774,根据临床信息将EAT的测序数据分为CAD组和健康对照组,使用R语言及相关软件包进行生物信息学分析。首先使用R语言筛选CAD组与健康对照组EAT间DEGs,并进行GO富集分析和KEGG通路富集分析,构建蛋白质-蛋白质相互作用（PPI）网络,评估所选基因的生物学功能及可能参与其调控的转录因子。构建GSE64554数据集中EAT的加权基因共表达网络（WGCNA）,获取同CAD表型相关的基因模块,将所获EAT间DEGs与模...     

A macrophage inflammatory protein homolog encoded by guinea pig cytomegalovirus signals via CC chemokine receptor 1

Cytomegaloviruses encode homologs of cellular immune effector proteins, including chemokines (CKs) and CK receptor-like G protein-coupled receptors (GPCRs). Sequence of the guinea pig cytomegalovirus (GPCMV) genome identified an open reading frame (ORF) which predicted a 101 amino acid (aa) protein with homology to the macrophage inflammatory protein (MIP) subfamily of CC (β) CKs, designated GPCMV-MIP. To assess functionality of this CK, recombinant GPCMV-MIP was expressed in HEK293 cells and assayed for its ability to bind to and functionally interact with a variety of GPCRs. Specific signaling was observed with the hCCR1 receptor, which could be blocked with hMIP −1α in competition experiments. Migration assays revealed that GPCMV-MIP was able to induce chemotaxis in hCCR1-L1.2 cells. Antisera raised against a GST-MIP fusion protein immunoprecipitated species of ∼12 and 10 kDa from GPCMV-inoculated tissue culture lysates, and convalescent antiserum from GPCMV-infected animals was immunoreactive with GST-MIP by ELISA assay. These results represent the first substantive in vitro characterization of a functional CC CK encoded by a cytomegalovirus.     

Historical evolution,overview, and therapeutic manipulation of co-stimulatory molecules

Co-stimulatory molecules are key mediators in the regulation of immune responses and knowledge of its different families, structure, and functions has improved in recent decades. Understanding the role of co-stimulatory molecules in pathological processes has allowed the development of strategies to modulate cellular functions. Currently, modulation of co-stimulatory and co-inhibitory molecules has been applied in clinical applications as therapeutic targets in diseases and promising results have been achieved.