Influence of antioxidants on the contractile response of heat-stressed human umbilical artery smooth muscle cells / 解放军医学杂志

Objective To study the change of the contractile response of human umbilical artery smooth muscle cells (HUASMCs) during the heat stress, and explore the effect of the antioxidant on the changes. Methods HUASMCs were randomly divided into control group, heat stress group, antioxidant preprocessing group. Cells were stimulated by norepinephrine (NE) at a low concentration (0.05mg/L) and at a normal concentration (1.0mg/L) and cultured in the thermostatic water bath (41℃) for 0.5, 1, 1.5 or 2h, respectively. After stimulated by NE, proportion of the cell surface area contraction was measured to reflect the contractile response of each group. Results Compared with control group, regardless of the NE concentration: in heat stress group, contractile response at 1h increased significantly (P0.05), but in the antioxidant preprocessing group, the contractile response was more significant to the normal NE concentration than to the low NE concentration (P<0.05 or 0.01). Regardless of the NE concentration, the contractile response was lower in the antioxidant preprocessing group than in the heat stress group. Conclusions In the course of heat stress, the contractile response of HUASMCs presents as time-related change. The usage of antioxidant may correct the over-response of HUASMCs to NE in the early heat stress stage, but cannot correct the reduction of the contractile response in the middle and advanced stage.

Effects of gradient heat stress on phagocytosis of liver Kupffer cells in vitro / 解放军医学杂志

Objective To investigate the effect of gradient heat stress on phagocytosis of hepatic Kupffer cells (KCs) in vitro in rats. Methods Rat Kupffer cells were isolated in vitro and the temperature for gradient heat stress was set at 37, 39, 41 and 43℃. After thermal stimulation, cell injury was detected by PI and Hochest33342 staining. CCK-8 assay was used to investigate difference in cellular proliferation rate over 24h between the groups. Flow cytometry was used to investigate the influence of heat stress on the phagocytosis of KCs. Results Compared to the normal control group, cells in each heat stress group exhibited varying degrees of damage, especially cells in 43℃ group. The ratio of damage cells increased with the increase of heat stress severity (P<0.05). Proliferation assay indicated that the proliferation rate of cells in each heat stress group was significantly decreased in comparison with normal control group 6h after heat stress (P<0.05). After 12h recovery, decrease in proliferation rate was observed only in 43℃ group (P<0.001), and no difference in the rate of proliferation could be observed between the heat stress groups and normal control group after 24h recovery. Flow cytometry showed, that the phagocytosis of KCs decreased in heat stress groups compared with control group, especially in 43℃ group (P<0.05). This phenomenon disappeared after 24h recovery. Conclusion Heat stress can inhibit the phagocytosis of rat liver KCs through its cytotoxic effect on KCs, and subsequently inhibits its proliferative ability. Further investigation of the effect of heat stress on KCs may help understand the pathogenesis of heat stress.

Effects of MAPKs signaling on heat stress-induced apoptosis of pulmonary microvascular endothelial cells and its mechanism / 解放军医学杂志

Objective To investigate the effect of mitogen-activated protein kinases (MAPKs) activation on the heat stressinduced apoptosis of pulmonary microvascular endothelial cells (PMVECs).Methods A mouse model of severe heat stroke was made and TUNEL and immunohistochemistry were employed to detect lung tissue damage.MACS separation was used for isolation of neonatal PMVECs,and TUNEL was utilized to detect the apoptosis of PMVECs.Western blotting was used for determining the MAPKs activation during heat stress recovery (0,2,6h).The monolayer permeability of endothelial cells was detected in terms of transmembrane resistance (TEER) and horseradish peroxidase (HRP).Cells were pretreated with MAPKs activation inhibitors to examine the effect of heat stress on the monolayer cell permeability and apoptosis.Results In mice with severe heat stroke,extensive apoptosis of PMVECs was found in their pulmonary tissues.TUNEL revealed that the number of apoptotic cells increased over time during heat stress recovery period and heat stress could activate MAPKs in PMVECs.Compared with heat stress group,in the cells pretreated with p38 or ERK activation inhibitor PD98059 and SB203580,the monolayer permeability and apoptosis increased while in cells pretreated withJNK inhibitor SP600125,the cellular permeability and apoptosis decreased.Conclusion In mice with severe heat stoke,PMVECs might experience apoptosis and p38 and ERK could inhibit apoptosis while JNK could promote apoptosis.

Mechanism of continuous venovenous hemofiltration combined with ulinastatin for the treatment of septic shock / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the molecular mechanisms of continuous venovenous hemofiltration (CVVH) combined with ulinastatin (ULI) (CVVH-ULI) for the treatment of septic shock.</p><p><b>METHODS</b>Human umbilical endothelial cells (HUVECs) were incubated with serums isolated from normal healthy people (control), septic shock patients treated with conventional therapy (CT) or treated with CVVH combined with ULI (CVVH-ULI). Endothelial permeability was evaluated by the leakage of FITC-labeled albumin. The morphological changes of F-actin was evaluated by Rhodamine-phalloidin. The phosphorylated levels of p38 were determined by Western blot. Cells were then treated with p38inhibitor (SB203580), or DMSO, followed by incubation with serum from septic shock patients treated with conventional therapy. Endothelial permeability and F-actin rearrangements were also evaluated as noted above.</p><p><b>RESULTS</b>Serum from CT group increased endothelial permeability, F-actin rearrangements, and phosphorylated levels of p38, which were inhibited by CVVH-ULI treatment. Moreover, in CT group, the serum-induced endothelial hyperpermeability and F-actin rearrangements were inhibited by SB203580, the inhibitor of p38.</p><p><b>CONCLUSION</b>CVVH combined with ulinastatin decreases endothelial hyperpermeability induced by septic shock through inhibiting p38 MAPK pathways.</p>

Protective effects of ulinastatin against acute lung injury induced by heatstroke in mice / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the protective effect of ulinastatin (UTI) against acute lung injury induced by heatstroke in mice.</p><p><b>METHODS</b>Sixty C57/BL6 mice were randomly divided into 6 groups, with 10 mice in each: control group, heatstroke group, UTI pretreatment group, saline pretreatment group, UTI post-treatment group, saline post-treatment group. The control mice were housed at a controlled room temperature of (22∓1) degrees; celsius, and the other groups were placed inside a temperature and humidity controlled chamber pre-set at 37 degrees; celsius and 60%. The two UTI groups were intraperitoneally injected with UTI at 5×10(4) U/kg 10 min before or after heat stress, and the two saline groups were given then equal amounts of saline in the same manner. The core body temperature of mice was monitored by a mercury thermometer every 30 min in the first 1.5 h during heating. The core temperature was measured, then every 15 min until it reached 42.7 degrees; celsius, which was taken as the onset of heatstroke. The animals were allowed to recover passively at ambient temperature for 6 h. The lung histopathological changes, protein concentration in BALF, lung wet/dry weight ratios, lung water content, and pulmonary microvascular permeability were assayed after 6 h of recovery at 37 degrees;celsius.</p><p><b>RESULTS</b>Compared with the control group, the heatstroke model group and two saline groups displayed more severe lung damage and pathological morphology changes, and the lung wet/dry weight ratio, protein concentration in BALF, lung water content and pulmonary microvascular permeability were also significantly increased. These effects were significantly alleviated in UTI treated group. Pretreat ment with UTI significantly prolonged the time to Tc≥42.7 degrees; celsius but had no effect on lung injury induced by heatstroke.</p><p><b>CONCLUSION</b>UTI can reduce the pulmonary edema and inflammatory exudation in acute lung injury caused by heatstroke.</p>

Research progress of long non-coding RNA in regulating cell apoptosis and autophagy / 中国病理生理杂志

Longnon-codingRNAs(lncRNAs)areagroupofRNAslongerthan200bpwithoutprotein-coding capacity and play an important role in epigenetics , transcription regulation and post-transcription regulation .Recently, studies have shown that dysregulation of lncRNAs caused cell cycle disorders , and abnormal activities of cell apoptosis and autophagy , contributing to a variety of diseases .Therefore , the role of lncRNAs in regulating cell death is expected to lead to the discovery of potential novel diagnostic markers and therapeutic targets .

Study on the pathological changes of the lung and brain in mice during heat stress / 中华急诊医学杂志

Objective To prepare mouse model with heat stress and determine its pathological changes of the lung and brain during heat stress. Methods BALB/c mouse were randomly (random number) divided into two groups, control group and heat stress group. The animals in the control group were sham- heated at a temperature of ( 25 ± 0.5) ℃ and humidity of (35 ± 5 ) %. The animals of heat stress group were placed in a prewarmed incubator maintained at (35.5 ± 0.5) ℃ and relative humidity of (60 ± 5) %. Rectal temperature (Tc) was monitored, and when Tc respectively reached 39 ℃, 40 ℃ , 41 ℃ and 42 ℃, those study animals were killed. The other animals were removed from the incubator and allowed to cool at an ambient temperature of (25 ±0. 5)℃ and humidity of (35 ±5)% , respectirvely for 12 and 24 hrs when Tc reached 41 ℃ , and for 6 hrs when Tc reached 42 ℃. The lung and brain of all the animals were isolated. Hematoxylin and eosin stain and light microscope were used to detect their pathological changes. Results All the animals displayed uniform response to the heat stress. Low degree of heat stress could induced obviously pathological changes of the lung, progressively greater damage to lung with further congestion of lung matrix, asystematic hemorrhage of alveolar space, abscission of alveolar epithelial cell and disappear of pulmonary alveolus tissue structure were detected with the rise of Tc to 42 ℃. However, absorption of congestion and hemorrhage and recovery of pulmonary alveolus tissue structure could also be seen with cooling at ambient temperature. With low degree of heat stress, the brain only showed moderate edema. Neuronal denaturation and necrosis were detected when Tc reached to 42 ℃. Interestingly, the lesions of brain further aggravated even through cooling treatment after Tc reached to 42 ℃ , but recovery could been observed after cooling treatment followed with Tc of 41 ℃. Conclusions The pathological changes of the lung and brain showed distinctive lesions to heat stress and cooling treatment, and these changes were correlated with the timing and time of cooling treatment, which provide the experimental basis to further study the mechanisms between the heatstroke and multiple organ dysfunction syndrome (MODS).

A UNIVERSAL PRIMER U2 LABELING METHOD FOR MICROARRAY ANALYSIS / 解剖学报

Objective To report a new method of fluorescent labeling technique in microarray studies: universal primer U2 labeling( UPL). The efficiency was compared of the UPL with that of random primer, restriction display labeling method and the reverse transcription coupled random primer spiking labeling method(RT-PSL). Methods Influenza viral RNA was labeled with both UPL and the conventional random primer labeling method as well as two other more laborious labeling methods( RD-direct and RD-incorporate), and hybridized with influenza virus oligonucleotide microarrays. The signals extracted from the microarrays were analyzed using SPSS 10.0 software. Results The fluorescent intensity, signal-to-noise ration(SNR), true positive ratio(TPR) of probes and labeling reproducibility of UPL were demonstrated to be higher than those of the Random primer approaches.Conclusion These results established that UPL is a valid new labeling protocol, which may have wide applications in the research and development of the microarray technology.

DETECTION OF SPERMATOZOAL TOTAL RNAS BY LABORATORY ON CHIP GEL ELECTROPHORESIS / 解剖学报

Objective Detecting of spermatozoal total RNAs by laboratory on chip gel electrophoresis so that it could provide better total RNAs for the sequent experiments, and spur the development of spermatozoal molecular biology. Methods Sperms of healthy adults were collected and then total RNAs were extracted by RNeasy mini kit(QIAGEN), detection and quality control were performed by loboratory on chip gel electrophoresis system. Meantime, the control RNAs were extracted from lymphocytes. Results It was found that there were a plenty of genes expressed in healthy sperms. Electrophoretic graphs showed that the total RNAs of spermatozoal had 2 bands which went ahead a little comparing to the normal somatic cells. The former peak appeared keenness, and the latter was broad and showed like a reversed U. The ratio of them was largely more than 2, no extra peaks were found in electrophoretic graph. Conclusion A simple,intuitionistic method to detect and control the quality of the healthy adults' spermatozoal total RNAs had been successfully constructed by using laboratory on chip gel electrophorosis.