Moderate hypothermia alters interleukin-6 and interleukin-1alpha reactions in ischemic brain in mice

Female C57BL/6 mice were decapitated and their brains were removed and cultured at either 37 or 33 degrees C for 48 h to investigate whether or not moderate hypothermia alters the cytokine reactions in the ischemic brain. The interleukin-6 and interleukin-1alpha levels in the culture media were significantly elevated in a time-dependent manner. The interleukin-6 levels after the incubation at 33 degrees C were significantly lower than those at 37 degrees C. The interleukin-1alpha levels at 33 degrees C were significantly higher than those at 37 degrees C. The interleukin-1alpha levels incubated with interleukin-6 antibody were significantly higher than those without IL-6 antibody. At 37 degrees C, the mRNA expression of interleukin-6 was observed from 2 to 48 h after incubation, but the same expression of interleukin-1alpha was only detected until 12 h. Accordingly, the ischemic brain incubated at 33 degrees C showed a decreased interleukin-6 production in comparison with that at 37 degrees C and the level of interleukin-6 showed negative feedback for the production of interleukin-1alpha. The temperature should, therefore, be carefully considered when evaluating the cytokine reaction for cerebral ischemia.     

A comparison of oral, rectal, and tympanic membrane-derived temperature changes after ingestion of liquids and smoking

Ambulatory patients frequently ingest liquids or smoke just before temperature measurement. The change in body temperature measurements over time following ingestion of ice water, hot water, and smoking were investigated. Twenty-two healthy, afebrile study subjects sequentially ingested temperature-controlled water and smoked a cigarette. Simultaneous oral and auditory canal temperatures were measured over 15 minutes following ingestion. Auditory canal temperatures were obtained with an infrared detection probe; we designated this process a tympanic membrane-derived (TMD) temperature. To determine the correlation between rectal and TMD temperatures, 100 patients had simultaneous measurements at both sites. Mean initial temperatures were rectal, 37.1 +/- 0.5 degrees C (mean +/- S.D.); oral, 36.4 +/- 0.4 degrees C; and TMD, 37.4 +/- 0.4 degrees C. Maximal mean oral temperature change was greatest at 1.5 minutes after hot, +0.9 +/- 0.1 degrees C, and cold, -1.2 +/- 0.2 degrees C, water. This change was statistically significant for seven minutes at the 95% confidence level (analysis of variance test with Dunnett's multiple range test for significance). There was no significant change in the TMD temperature with any ingestion. The Pearson correlation coefficient for 107 pairs of rectal and TMD temperatures, r = 0.90 (P less than .001), was excellent. It was concluded that hot and cold liquids significantly influence oral temperature measurement for seven to nine minutes following ingestion. TMD temperature is unaffected by liquid ingestion and may allow accurate measurement of body temperature. Further studies are needed to determine the accuracy of TMD temperature over a wide range of body temperature in diverse clinical settings.     

Effects of long-term whole-body cold exposures on plasma concentrations of ACTH, beta-endorphin, cortisol, catecholamines and cytokines in healthy females

OBJECTIVE: Cold therapy is used to relieve pain and inflammatory symptoms. The present study was designed to determine the influence of long-term regular exposure to acute cold temperature. Two types of exposure were studied: winter swimming in ice-cold water and whole-body cryotherapy. The outcome was investigated on humoral factors that may account for pain alleviation related to the exposures. MATERIAL AND METHODS: During the course of 12 weeks, 3 times a week, a group of healthy females (n = 10) was exposed to winter swimming (water 0-2 degrees C) for 20 s and another group (n = 10) to whole-body cryotherapy (air -110 degrees C) for 2 min in a special chamber. Blood specimens were drawn in weeks 1, 2, 4, 8 and 12, on a day when no cold exposure occurred (control specimens) and on a day of cold exposures (cold specimens) before the exposures (0 min), and thereafter at 5 and 35 min. RESULTS: Plasma ACTH and cortisol in weeks 4-12 on time-points 35 min were significantly lower than in week 1, probably due to habituation, suggesting that neither winter swimming nor whole-body cryotherapy stimulated the pituitary-adrenal cortex axis. Plasma epinephrine was unchanged during both experiments, but norepinephrine showed significant 2-fold to 3-fold increases each time for 12 weeks after both cold exposures. Plasma IL-1-beta, IL-6 or TNF alpha did not show any changes after cold exposure. CONCLUSIONS: The main finding was the sustained cold-induced stimulation of norepinephrine, which was remarkably similar between exposures. The frequent increase in norepinephrine might have a role in pain alleviation in whole-body cryotherapy and winter swimming.     

热休克及化疗刺激对人白血病细胞K562中HSP70 mRNA及MDR1 mRNA表达的影响

为了研究热休克、Adriamycin（ADM）化疗应激刺激下热休克蛋白70（HSP70）、多药耐药基因MDRl的表达情况，对K562细胞予以43℃高温热休克和（或）加以ADM化疗，然后采用免疫组织化学方法检测HSP70的表达，MTT法检测K562细胞的生长抑制率，RT—PCR检测HSP70 mRNA和MDRl mRNA的表达，流式细胞术检测P-糖蛋白（P—gP）的表达。结果显示：①HSP70蛋白的合成在43℃热作用后恢复37℃孵育2小时达到高峰并聚集在胞核和核仁内，呈“核内迁移”现象；3小时后HSP70表达开始降低，5小时后HSP70的合成逐渐降低至正常，并恢复以胞浆表达为主。②K562细胞在43℃高温热休克刺激下HSP70mRNA表达增加，随着37℃孵育时间的延长表达量有增多趋势并持续一段时间，MDRlmRNA的表达与之相似，二者在37℃孵育2小时分别增加4倍和5．8倍。③ADM化疗刺激后细胞内P—gP表达增加；ADM化疗、43℃热休克刺激后再加ADM化疗，K562细胞中HSP70 mRNA、MDRl mRNA表达均上调，且43℃热休克刺激处理后再加入ADM作用二者的表达要高于单加ADM化疗组。结论：热休克和ADM化疗刺激的K562细胞中HSP70和MDRl表达增加，有利于维护肿瘤细胞的稳定性，HSP70可能与耐药有关。     

Platelet cold agglutinins: a flow cytometric analysis.

Spontaneous EDTA-independent cold platelet agglutination is a rare phenomenon that produces pseudothrombocytopenia when blood samples are analyzed in automated cell counters. We report a case of platelet cold agglutinins and an analysis by flow cytometry. A 49 year old woman presented with abnormal vaginal bleed secondary to uterine fibroids. Platelet clumping was observed in blood samples taken in EDTA-, heparin- and citrate-containing tubes. In flow cytometric tests, patient serum agglutinated 16% of normal platelets at 22 degrees C, and 7% of platelets after incubation at 37 degrees C; in contrast, 3% and < 1% of platelets were agglutinated at 22 and 37 degrees C, respectively, after incubation with normal serum. Minimal agglutination (< 10%) was observed with patient serum at a titre of 1:5 or at temperatures > 30 degrees C. After incubation at 4 degrees C, IgM antibody and C3 were increased on the patient's platelets; no significant amount of IgM or C3 was detected on normal platelets. The specificity of the platelet cold agglutinin was determined by competitive inhibition by monoclonal anti-CD41(GPIIbIIIa). Before the addition of monoclonal antibody, patient's serum agglutinated 16% of normal platelets at 22 degrees C; after addition of anti-CD41 only 2% of the platelets were agglutinated. This blocking effect was not observed with anti-CD42. The patient's platelets functioned normally as determined by CD62 and CD63 expression in response to thrombin, normal platelet aggregation in response to collagen, ADP, and ristocetin, and a normal template bleeding time. In summary, platelet agglutination by a platelet cold agglutinin was quantitated by flow cytometry, the responsible antibody was characterized as a low titre IgM with minimal activity > 30 degrees C, and competitive binding studies supported the GPIIbIIIa complex as the binding site for the antibody. Since the antibody did not affect platelet function, we believe that these patients will not suffer complications from their platelet cold agglutinin, but it could pose a problem under circumstances such as cardiac surgery with hypothermia.     

The effect of local temperature on the response of an extremity to indirect heating in man

Venous occlusion plethysmography was used to measure blood flow through the extremities of five normal subjects before and during exposure to a standard heat load with one extremity maintained at a cold (15 degrees C) and the other at a neutral (35 degrees C) temperature. In the foot, local cold (15 degrees C) delayed the onset and reduced by about 92% the vasodilatation during release of sympathetic vasoconstrictor tone caused by body heating. Similar results were obtained in the hand exposed to cold (15 degrees C) but the suppression of the reflex vasodilatation in response to body heating was about 66%. This difference between the hand and foot did not appear to be related to the greater length of the foot since the distal half of the foot reacted like the total foot: local cooling suppressed the reflex vasodilatation by 88%. The results suggest that sympathetic release tests should not be carried out in a laboratory with an environmental temperature as low as 15 degrees C, especially when the circulation to the foot is under investigation.     

An intron enhancer activates the immunoglobulin-related Hemolin gene in Hyalophora cecropia

Hemolin is the only insect member of the immunoglobulin (Ig) superfamily reported to be up-regulated during an immune response. In diapausing pupae of Hyalophora cecropia the gene is expressed in fat body cells and in haemocytes. Like the mammalian Ig kappa light chain gene, the Hemolin gene harbours an enhancer including a kappaB motif in one of its introns. This motif binds the H. cecropia Rel factor Cif (Cecropia immunoresponsive factor). The Hemolin third intron also mediates transient reporter gene expression in immunoresponsive Drosophila mbn-2 cells. Co-transfections of Drosophila SL2 cells showed that the Drosophila Rel factor Dif (Dorsal-related immunity factor), transactivates reporter gene constructs through the intron. Moreover, a 4.8-fold synergistic activation was obtained when Dif is combined with the rat C/EBP (CCAAT/enhancer element-binding protein) and human HMGI (high mobility group protein I). This is the first report of an insect immune-related gene that is up-regulated by an enhancer activity conferred through an intron.     

Gene transcription during exposure to, and recovery from, cold and desiccation stress in Drosophila melanogaster

We exposed adult male Drosophila melanogaster to cold, desiccation or starvation, and examined expression of several genes during exposure and recovery. Frost was expressed during recovery from cold, and was up-regulated during desiccation. Desiccation and starvation (but not cold) elicited increased expression of the senescence-related gene smp-30. Desat2 decreased during recovery from desiccation, but not in response to starvation or cold. Hsp70 expression increased after 1 h of recovery from cold exposure, but was unchanged in response to desiccation or starvation stress, and Hsp23 levels did not respond to any of the stressors. We conclude that D. melanogaster's responses to cold and desiccation are quite different and that care must be taken to separate exposure and recovery when studying responses to environmental stress.     

Immunochemical studies on an IgG lambda cryoglobulin in cold-induced urticaria

A type I cryoglobulinaemia associated with cold-induced urticaria was demonstrated in a 64-year-old woman without primary disease. The cryoglobulin contained only IgG lambda as disclosed by immunofixation technique. Different physicochemical studies indicated that the IgG lambda component was monomeric at temperatures above 35 degrees C, but became polymerized below 35 degrees C. In addition crossed immunoelectrophoresis of plasma fibronectin from the patient showed a heterogeneous precipitate at low temperatures but a homogeneous precipitate at 25 degrees C indicating a complex formation at low temperature between IgG lambda and fibronectin. Fibronectin, however, was not essential for the cold precipitation of the cryoglobulin. The precipitation phenomenon at low temperatures was found to be a result of the physicochemical properties of the cryoglobulin itself unrelated to the antibody specificities tested. The importance of performing the immunochemical and physicochemical techniques at low temperature (7 degrees C) and at high temperature (35 degrees C) to gain knowledge of the nature of the protein, is emphasized. We conclude that only results obtained by relevant laboratory procedures might lead to correct classification and understanding of cryoglobulinaemia.     

Insect cold hardiness: the role of mitogen‐activated protein kinase and Akt signalling in freeze avoiding larvae of the goldenrod gall moth,Epiblema scudderiana

Larvae of the goldenrod gall moth, Epiblema scudderiana, use the freeze avoidance strategy of cold hardiness to survive the winter. Here we report that protein kinase‐dependent signal transduction featuring mitogen‐activated protein kinase (MAPK) signalling cascades (extracellular signal regulated kinase, c‐jun N‐terminal kinase and p38 MAPK pathways) and the Akt (also known as protein kinase B, or PKB) pathway could be integral parts of the development of cold hardiness by E. scudderiana. We used Luminex technology to assess the protein levels and phosphorylation status of key components and downstream targets of those pathways in larvae in response to low temperature acclimation. The data showed that MAPK pathways (both total protein and phosphorylated MAPK targets) were inhibited after 5°C acclimation, but not ?15°C exposure, as compared with the 15°C control group. However, total heat shock protein 27 (HSP27) levels increased dramatically by ～12‐fold in the ?15°C acclimated insects. Elevated HSP27 may facilitate anti‐apoptotic mechanisms in an Akt‐dependent fashion. By contrast, both 5 and ?15°C acclimation produced signs of Akt pathway activation. In particular, the inhibitor phosphorylated Glycogen Synthase Kinase 3a (p‐GSK3) levels remained high in cold‐exposed larvae. Additionally, activation of the Akt pathway might also facilitate inhibition of apoptosis independently of GSK3. Overall, the current study indicates that both MAPK and Akt signal transduction may play essential roles in freeze avoidance by E. scudderiana.     

Proteomics of cryoprotective dehydration in Megaphorura arctica Tullberg 1876 (Onychiuridae: Collembola)

The Arctic springtail, Megaphorura arctica Tullberg 1876 (Onychiuridae: Collembola), is one of the few organisms known to survive the extreme stresses of its environment by using cryoprotective dehydration. We have undertaken a proteomics study comparing M. arctica, acclimated at -2°C, the temperature known to induce the production of the anhydroprotectant trehalose in this species, and -6°C, the temperature at which trehalose expression plateaus, against control animals acclimated at +5°C. Using difference gel electrophoresis, and liquid chromatography tandem mass spectrometry, we identified three categories of differentially expressed proteins with specific functions, up-regulated in both the -2°C and -6°C animals, that were involved in metabolism, membrane transport and protein folding. Proteins involved in cytoskeleton organisation were only up-regulated in the -6°C animals.