‘Ischemic tolerance’ phenomenon detected in various brain regions

We investigated the effects of mild and non-lethal ischemic insult on neuronal death following subsequent lethal ischemic stress in various brain regions, using a gerbil model of bilateral cerebral ischemia. Single 10-min ischemia consistently caused neuronal damage in the hippocampal CA1, CA2, CA3 and CA4, layer III/IV of the cerebral cortex, dorsolateral part of the caudoputamen and ventrolateral part of the thalamus. On the other hand, in double ischemia groups, 2-min ischemic insult 2 days before 10-min ischemia exhibited significant protection in the CA1 and CA3 of the hippocampus, the cerebral cortex, the caudoputamen and the thalamus. Five-min ischemic insult 2 days before 10-min ischemia also showed protective effect in the same areas as those of 2-min ischemia except for the CA1 region of the hippocampus, while 1-min ischemic insult exhibited no protective effect in any brain regions. In the immunoblot analysis, both 2- and 5-min ischemia caused increased synthesis of heat shock protein 72 (HSP 72) in the hippocampus, but 1-min ischemia did not. The present study demonstrated that the ‘ischemic tolerance’ phenomenon was widely found in the brain and also suggested that ischemic treatment severe enough to cause HSP 72 synthesis might be needed for induction of ‘ischemic tolerance’.     

微嵌合体表达与移植肾免疫耐受的实验研究

目的探讨肾移植术后受者微嵌合体表达与免疫耐受的相关性。方法采集接受男性供体肾脏移植术后女性受者不同生存期外周血及尿沉渣标本,采用Y染色体三对引物SRY1、DYZ11st和DYZ12nd,应用PCR和RT-PCR方法检测标本中微嵌合体特异性代表Y染色体DNA和mRNA的表达,同时测定移植肾功能。结果在130例供者为男性的女性肾移植受者检测到嵌合阳性者98例,占75.39%(n=130),嵌合阴性者32例,占24.61%。嵌合阳性与阴性两组比较,移植肾平均存活时间分别为(8.7±3.5)年和(5.4±3.3)年,P<0.05;嵌合阳性者发生过排斥反应的有11例,占11.22%,而嵌合阴性者为9例,占28.13%,P<0.05;血清肌酐在嵌合阳性者(74.3±32.5)mol/L,而嵌合阴性者为(113.6±37.8)mol/L,P<0.05。结论受者体内嵌合阳性与阴性者相比具有更好的移植肾功能和低排斥率,肾移植术后受者生存期愈长嵌合阳性率愈高,提示肾移植术后受者体内的嵌合状态与免疫耐受具有相关性。     

Short-term suppression of plasma free fatty acids fails to improve insulin sensitivity when intramyocellular lipid is elevated.

AIMS: Metabolic responses to manipulation of the plasma free fatty acid (FFA) concentration were assessed in six healthy men via cross-over design to determine whether FFAs independently influence insulin sensitivity. METHODS: Intramyocellular lipid (IMCL) was measured by proton magnetic resonance spectroscopy and insulin sensitivity via frequently sampled intravenous glucose tolerance test (IVGTT) after 67 h of two identical low carbohydrate/high fat (LC) diets which were used to elevate IMCL and plasma FFAs. To uncouple the influence of FFAs and IMCL on insulin sensitivity, FFAs were suppressed 30 min prior to and during IVGTT in one treatment [LC + nicotinic acid (NA)] by NA ingestion. RESULTS: Vastus lateralis IMCL was significantly elevated in LC (13.3 +/- 1.1 x 10(-3)) and LC + NA (13.5 +/- 1.1 x 10(-3)) (P < 0.01 for both), but was not different between conditions (P > 0.05). Plasma FFAs were raised in LC (0.79 +/- 0.08 mmol/l) and LC + NA (0.80 +/- 0.11 mmol/l) (P < 0.01 for both) and were significantly reduced by NA ingestion prior to (0.36 +/- 0.05 mmol/l, P < 0.01) and during IVGTT (P < 0.05) in LC + NA. Despite marked differences in plasma FFA availability, insulin sensitivity and glucose tolerance were not different between LC and LC + NA (P > 0.05 for both). CONCLUSIONS: Plasma FFAs appear to exert no immediate effect on insulin sensitivity/glucose tolerance independent of their action on intracellular lipid moieties. Further research is required to elucidate the duration of FFA suppression required to restore insulin sensitivity following lipid-induced insulin resistance.     

The short insulin tolerance test for determination of insulin sensitivity: a comparison with the euglycaemic clamp.

The glucose clamp technique is currently regarded as the standard test for measuring insulin sensitivity against which other methods are compared but is unsuitable for routine screening of patients outside a hospital base. There is thus a need for a simpler test to measure insulin sensitivity. We have therefore compared the glucose disappearance rate KITT in the first 15 min of the insulin tolerance test (ITT) with the M and M/I values derived from the standard euglycaemic clamp in nine normal subjects and eight subjects with Type 2 (non-insulin dependent) diabetes mellitus and coexisting obesity. All subjects underwent the ITT and euglycaemic clamp in random order. Nine subjects later had a repeat ITT to determine the reproducibility of the test. In the ITT, 0.1 U kg-1 body weight, human Actrapid insulin was given as an IV bolus and simultaneous arterialized and venous blood samples were obtained every minute for 15 min. The first order rate constant for the disappearance of glucose KITT over the period 3-15 min was taken as a measure of insulin sensitivity. The euglycaemic clamp was performed with an insulin infusion of 50 mU kg-1 h-1 for 120 min and a variable rate glucose infusion to maintain blood glucose concentration at 0.5 mmol l-1 below fasting level to minimize the effect of endogenous insulin secretion. The ratio of the mean rate of glucose infused (M, mumol kg-1 min-1) to the plasma insulin over the last 30 min of the clamp was taken as a measure of tissue sensitivity to insulin (M/I) assuming endogenous glucose output was suppressed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differences in the hypothermic response to ethanol in rats selectively bred for oral ethanol preference and nonpreference

The duration of retention of tolerance to ethanol was tested in the alcohol-preferring (P) and alcohol-nonpreferring (NP) rats lines, using ethanol-induced hypothermia as a measure of tolerance. Rats received two injections of ethanol (3.5 g/kg) body wt, IP) and the time between the injections was 1, 2, or 3 days. When one day separated the two injections, tolerance to the hypothermic effect of a second “test” injection was found in both lines. When 2 or 3 days separated the two injections, the P line showed a loss of tolerance and the NP line showed sensitization to ethanol. Sensitization in the NP line grew stronger when the interval between injections was increased from 2 to 3 days. The duration of retention of tolerance to ethanol-induced hypothermia in the P line was shorter than has previously been reported for motor impairment in this line. It appears that the duration of tolerance retention in the P line depends on the test used to measure tolerance. Sensitization to ethanol in the NP line may be associated with low oral ethanol intake. This research was supported, in part, by grants AA08312, AA03243, and AA07611 from the PHS     

Characterization of T cell epitopes in αs1-casein in cow''s milk allergic, atopic and non-atopic children

BACKGROUND: One to two percent of infants suffer from IgE-mediated allergic reactions against cow's milk proteins. Most children develop clinical tolerance, but approximately 15% are still allergic by the age of 10 years. Little is known about the T cell epitopes in individual cow's milk protein in relation to allergy and tolerance. OBJECTIVE: To identify T cell epitopes in alphas1-casein, the most abundant milk protein, and to investigate T cell responses toward these epitopes in allergic, atopic and non-atopic children. METHODS: Allergen-specific T cell lines (TCLs) were derived from peripheral blood mononuclear cells of 11 cow's milk allergic, nine atopic and nine non-atopic children. T cell responses were measured to alphas1-casein and to overlapping peptides (18-mers), spanning the alphas1-casein molecule. Proliferation was determined by incorporation of (3)H-thymidine, and cytokine production (IL-10, IL-13 and IFN-gamma) was measured by ELISA. RESULTS: Four main regions (amino acid (AA) residues 43-66, 73-96, 91-114 and 127-180) in the alphas1-casein molecule were immunogenic to T cells, among which the AA residues 133-156 spanned the immunodominant part. Only subtle differences were found in peptide recognition between the subject groups. Some of the peptides induced slightly Th1- or Th2-skewed cytokine responses. The increased levels of IL-10 in response to alphas1-casein observed in TCLs from atopic children appeared not to be linked to recognition of specific IL-10-inducing epitopes. CONCLUSIONS: The immunodominant sequence in alphas1-casein is spanned by AA residues 133-156. Tolerance towards alphas1-casein in atopic children may be mediated by an overall induction of IL-10 and not by recognition of certain T cell epitopes. The identified T cell epitopes in children with cow's milk allergy may be useful targets in developing peptide immunotherapy.     

口服抗原对巨噬细胞共刺激分子CD80/CD86表达的影响

目的探讨口服抗原对巨噬细胞(M)共刺激分子CD80/CD86表达的影响及其在诱导妊娠免疫耐受中的作用。方法自然流产小鼠(CBA/J×DBA/2)分为免疫组和未免疫组,免疫组分别口服滋养细胞膜抗原(TMA2)和卵清蛋白(OVA),以正常妊娠小鼠(CBA/J×BALB/c)作为对照组。采用双标记流式细胞分析技术,分别检测各组小鼠脾脏及肠系膜淋巴结(MLN)CD80 M和CD86 M的表达。结果MLN内,未免疫组CD80 M表达明显高于对照组(P<0.05),而CD86 M含量明显低于对照组(P<0.001);TMA2免疫组CD80 M含量明显低于未免疫组(P<0.05);OVA免疫组CD86 M含量明显高于未免疫组(P<0.001)。脾脏内,CD80 M表达在各组间比较无显著差异;CD86M表达,未免疫组明显低于对照组(P<0.05),TMA2免疫组显著低于未免疫组(P<0.05),OVA免疫组则显著高于未免疫组(P<0.001)。结论流产的发生与M表面共刺激信号CD80/CD86异常有关;口服适当抗原可改变CD80/CD86 M表达模式,诱导妊娠免疫耐受的形成。     

Rat Cytomegalovirus Infection Interferes with Anti-CD4 mAb-(RIB 5/2) Mediated Tolerance and Induces Chronic Allograft Damage

In order to assess the role of heterologous immunity on tolerance induction (TI) by signal 1 modification, the influence of rat cytomegalovirus infection (RCMVI) on TI by a non-depleting monoclonal anti-CD4 mAb (monoclonal antibody) (RIB 5/2) in a rat kidney transplant (KTx) model was investigated. Orthotopic rat KTx (Dark Agouty (DA)-->Lewis (LEW)) was performed after TI with RIB 5/2 [10 mg/kg body weight (BW); day -1, 0, 1, 2, 3; i.p. (intraperitoneal route)]. RCMVI (5x10E5 Plaque forming units [PFU] i.p.) was simultaneously conducted to KTx, 50 days after KTx, and 14 days before and after KTx. RIB 5/2 induced robust allograft tolerance even across the high-responder strain barrier. RCMVI broke RIB 5/2-induced tolerance regardless of the time of RCMVI but did not induce acute graft failure during the 120 days follow-up. RCMVI induced a significant chronic deterioration of allograft function (p<0.01) and enhanced morphological signs of chronic allograft damage (p<0.05). Cellular infiltrates and major histo-compatibility complex (MHC)-expression were more pronounced (p<0.05) in the infected groups. RCMVI induced not only RCMV-specific T-cell response but also enhanced the frequency of alloreactive T cells. RCMV interferes with anti-CD4 mAb-induced tolerance and leads to chronic allograft damage. The data we presented suggest a potentially important role of viral infections and their prophylaxis in clinical TI protocols.     

Intranasal treatment with ovalbumin but not the major T cell epitope ovalbumin 323–339 generates interleukin-10 secreting T cells and results in the induction of allergen systemic tolerance

BACKGROUND: Nasal administration of major peptide T cell epitopes gives contradictory data on the induction of peripheral tolerance. OBJECTIVE: To compare the prophylactic effect of intranasal treatment (INT) on the development of an allergic response, using either ovalbumin (OVA) or its major T cell epitope OVA 323-339 (OVAp). METHODS: BALB/c mice were treated intranasally with OVA or OVAp and subsequently immunized s.c. with OVA. Anti-OVA-specific antibody, T cell proliferation and cytokine responses were analysed. In an adoptive transfer model using OVAp specific TCR transgenic (Tg) T cells from D011.10 mice, in vivo tracking and characterization of transferred T cells in the cervical, inguinal and bronchial lymph nodes (BLN) and in the spleen were determined by FACS analysis. RESULTS: Prophylactic INT with OVA induced T cell tolerance towards subsequent OVA s.c. immunizations, inhibiting OVA specific T cell proliferation, IgE and IgG1 production, in contrast to INT with OVAp, which was unable to induce tolerance. In vivo analysis of transferred OVA-specific TCR Tg T cells showed that INT with OVA induced a preferential activation of T cells in BLN, as opposed to a broad, systemic activation with OVAp. In vivo, OVAp INT led to faster and more sustained cell division cycles than OVA INT. Ex vivo, tolerance to OVA was associated with the generation of IL-10 secreting CD4(+) T cells in BLN of OVA-treated mice only. CONCLUSION: INT with OVA but not with OVAp led to regional (as opposed to systemic) T cell activation and the induction of IL-10 secreting CD4(+) T cells in BLN, potentially critical steps in the induction of T cell-specific tolerance via the nasal route.