Objective To establish TLC identification method,HPLC fingerprint,chemical pattern recognition,and multi-components quantitative analysis methods of Spatholobus suberectus,and identify the quality of 16 batches of Spatholobus su-berectus from different origins.Methods The TLC method was used to identify Spatholobus suberectus qualitatively.HPLC fin-gerprint of Spatholobus suberectus was established.The quality of Spatholobus suberectus was evaluated by chemical pattern recogni-tion technologies,and the contents of Gallocatechin,protocatechuic acid,catechin,epigallocatechin and fermononetin were deter-mined by HPLC.Results TLC method of Spatholobus suberectus was established.Ten common peaks were identified in 16 bat-ches of Spatholobus suberectus,and 6 components were identified by reference substances.The similarity of fingerprint was 0.769-0.990,indicating good similarity.The samples were divided into 3 groups by cluster analysis.Results of the principal component a-nalysis showed that the top 3 samples in the list of comprehensive scores were S16、S1 1、S14.Marker compounds that cause the quality difference of Spatholobus suberectus were screened out through the orthogonal partial least squares discriminant analysis,which were fermononetin,peaks 7 and epigallocatechin.Gallocatechin,protocatechuic acid,catechin,epigallocatechin,and ferm-ononetin had a good linearity in the concentration range(r＞0.999).The content determination analysis showed that there were sig-nificant differences in the contents of the 5 index components among Spatholobus suberectus from different origins.Conclusion The established TLC and HPLC fingerprints of Spatholobus suberectus were stable and reliable,and the HPLC method for multiple active components method provides scientific support for improving the quality control method of Spatholobus suberectus.

Objective To establish TLC identification method,HPLC fingerprint,chemical pattern recognition,and multi-components quantitative analysis methods of Spatholobus suberectus,and identify the quality of 16 batches of Spatholobus su-berectus from different origins.Methods The TLC method was used to identify Spatholobus suberectus qualitatively.HPLC fin-gerprint of Spatholobus suberectus was established.The quality of Spatholobus suberectus was evaluated by chemical pattern recogni-tion technologies,and the contents of Gallocatechin,protocatechuic acid,catechin,epigallocatechin and fermononetin were deter-mined by HPLC.Results TLC method of Spatholobus suberectus was established.Ten common peaks were identified in 16 bat-ches of Spatholobus suberectus,and 6 components were identified by reference substances.The similarity of fingerprint was 0.769-0.990,indicating good similarity.The samples were divided into 3 groups by cluster analysis.Results of the principal component a-nalysis showed that the top 3 samples in the list of comprehensive scores were S16、S1 1、S14.Marker compounds that cause the quality difference of Spatholobus suberectus were screened out through the orthogonal partial least squares discriminant analysis,which were fermononetin,peaks 7 and epigallocatechin.Gallocatechin,protocatechuic acid,catechin,epigallocatechin,and ferm-ononetin had a good linearity in the concentration range(r＞0.999).The content determination analysis showed that there were sig-nificant differences in the contents of the 5 index components among Spatholobus suberectus from different origins.Conclusion The established TLC and HPLC fingerprints of Spatholobus suberectus were stable and reliable,and the HPLC method for multiple active components method provides scientific support for improving the quality control method of Spatholobus suberectus.

Moutan Cortex and Paeonia Radix are also traditional Chinese herbal medicines. The market is in great demand. However, the market sales of Moutan Cortex and Paeonia Radix have problems such as pesticide residues, excessive heavy metals and degraded quality, which affect their safety and effectiveness. The establishment of a pollution-free and planting system can effectively reduce the pesticide residue and heavy metal content and improve the quality of the medicinal materials while ensuring the production of Moutan Cortex and Paeonia Radix. The article uses field data which based on P.suffruticosa and P.lactiflora cultivation, combined with growth habits and distribution of origin, this research has developed a pollution-free and planting technology system for P.suffruticosa and P.lactiflora. The system includes planting base regionalization, field management, fertilization, pest control and so on, which provides reference for P.suffruticosa and P.lactiflora.

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.

Objective To investigate the effect ofulinastatin on severe heat-stroke with acute lung injury induced by severe heat stroke.Methods Thirty severe heat stroke patients were divided into conventional group (n=15) and ulinastatin group (n=15) randomly,with another 80 healthy adults serving as controls.The baseline data such as age,gender,onset period and APACHE Ⅱ scores were recorded and compared between the two groups on admission.Peripheral leucocyte counts,oxygenation index and Murray scores were determined on the 1st,3rd and 5th day.The concentration of inflammatory mediators in bronchoalveolar lavage fluid (BALF) and alveolar macrophage supernatant were detected by enzyme-linked immunosorbent assay (ELISA).Western blotting and real-time PCR were used to measure expression of triggering receptor-1 on myeloid cells (TREM-1) on alveolar macrophages.Furthermore,comparison was made in terms of the ventilation period,ICU stay time and mortality in 28 days between the two groups.Results No differences were found in age,gender,onset period and APACHE Ⅱ scores between the two groups (P＞0.05).Compared with the conventional group,peripheral leucocyte counts and Murray scores in the ulinastatin group significantly decreased on the 3rd and 5th day (P＜0.05,P＜0.01).But oxygenation index was higher in the ulinastatin group than in the conventional group (P＜0.05).The concentration of TNF-α and IL-6 in BALF was lower in the ulinastatin group than in the conventional group (on the 3rd day:P＜0.05,P＜0.01;on the 5th day:P＜0.01,P＜0.01).The concentration of TNF-α and IL-6 in alveolar macrophage supernatant was lower in the ulinastatin group than in the conventional group (on the 3rd day:-P＜0.05,P＜0.01;on the 5th day:P＜0.01,P＜0.05).The expression of TREM-1 protein on alveolar macrophages were lower in the ulinastatin group than in the conventional group (on the 3rd day P＜0.01;on the 5rd day P＜0.05).TREM-1 mRNA was lower in the ulinastatin group than in the conventional group (on the 3rd day:P＜0.05;on the 5th day:P＜0.05).Eventually,the treatment of ulinastatin shorten ventilation period and ICU stay time (P＜0.01,P＜0.05).Nonetheless,it failed to show any improvement in terms of the mortality during 28 days (P＞0.05).Conclusion Our study exhibited that ulinastatin had good effect on the heat stroke patients with acute lung injury and it helped reduce the inflammatory reaction of pulmonary tissues.The underlying mechanism of these effects might lie in its ability to reduce heat stroke-induced inflammatory secretion and expression of TREM-1 on alveolar macrophage.

Objective To investigate the protective effect of heat shock protein (HSP) 70 on the acute lung injury (ALI) of rats with heat stroke.Methods Sixty four rats were randomly (by employing a random number table) assigned into a sham-heated group (Sham group),heat stress group (HS group),and HS plus gluttamine treatment group (HS+GLN group) and HS plus quercet in treatment group (HS+QU group),16 each.All rats were housed in a artificial climate chamber,with the rats in the sham groups exposed to a temperature of 23 ℃ and humidity of 55％ ± 5％,while the rats of HS,HS+GLN and HS+QU groups to an ambient temperature of 39 ℃ and humidity of 65％.During heat stress or sham heating,rectal temperature (Tr),systolic blood pressure (SBP) and pulse rate (PR) were monitored to observe the difference in heat stress response among the groups.The time point at which the SBP started to drop from the peak level was taken as the point of HS onset.At the onset of HS,heat exposure was terminated,then the rats were immediately removed from the chamber,and returned to room temperature.The rats were scarified 0h and 6h after HS onset respectively.After bronchoalveolar lavage fluid (BALF) was collected,the lungs of all animals were harvested for pathological examination of lung injury.The concentrations of IL-1β,TNF-α and IL-6 in BALF and HSP70 in lung homogenate were measured by using an enzyme linked immunosorbent assay kit.Results Compared with HS and HS+QU groups,the rats in HS+GLN group required significantly greater heat load to induce HS (P＜0.001),and had longer survival time span after HS onset.Compared with Sham group,the concentration of HSP70 in lung homogenate in HS group increased in a time-dependent manner (P＜0.001).In comparison with HS group,the concentration of HSP70 in lung homogenate from HS+GLN group was significantly elevated at each time point (P＜0.001),while the treatment with QU significantly inhibited the expression of HSP70 (P＜0.001).The concentration of IL-1β,TNF-α and IL-6 in BALF significantly decreased in HS+GLN group compared with those in HS group and HS+QU group (P＜0.001).The pathological results showed that the lung injury was milder in HS+GLN group,while the opposite in HS+QU group.Conclusion HSP70 could protect HS rats against ALI by enhancing their thermo-tolerance and inhibiting inflammatory response.

Objective To investigate the role of oxidative stress in acute liver injury in a heat stroke model of conscious rats,and to explore its underlying mechanism.Methods Thirty-two rats were randomly (by using a random number table) assigned into a sham-heated control group (Sham group,n=8),a sham-heated group treated with NAC (Sham-NAC group,n=8),a heat stroke group (HS group,n=8) and a heat stoke group treated with NAC (HS-NAC,n=8).Rats were prepared with pre-warm chamber to initiate heat stoke.The change of rectum temperature (Tr),heart rate (HR) and systolic blood pressure (SBP) were monitored,and the time point of HS onset was recorded.Rats were sacrificed 12h after HS onset.ALT,serum TBIL,IL-6,IL-1β,TNF-α,MDA,T-SOD and GSH in the liver homogenates were measured.Liver tissues were harvested for determining the concentration of reactive oxygen species (ROS),neutrophil infiltration and the histological changes.Results During HS onset,no significant differences were observed in Tr,HR,SBP and heat exposure time between HS group and HS-NAC group (P＞0.05).However,the survival time was significantly longer in HS-NAC group than in HS group (P=0.039).12 hours after HS onset,the concentrations of ROS and MDA in the liver homogenates were significantly higher in HS group than in the other groups (P=0.000),while the concentrations of T-SOD and GSH were much lower than in the other groups (P=0.000).The serum concentrations of ALT and TBIL were significantly higher in HS group than in the other groups (P=0.000).Compare with HS group,the pathological injury was alleviated in HS-NAC group (P=0.000).The neutrophil infiltration level and the concentrations of IL-6,IL-1 β and TNF-α in liver tissue were significantly higher in HS group than in HS-NAC group (P=0.000).Conclusion Oxidative stress may play an important role in the pathogenesis of HS liver injury through its cytotoxic effect and by inducing inflammatory responses.

There exist a series of problems in heat stroke treatment,such as,pathogenesis is still unclear,clinical classification is too simple and has no intrinsic relation with pathophysiological process and prognosis,missing of indexes for hierarchical diagnosis and prognosis prediction,and lack of targeted therapeutic norms.All of these factors could lead to high mortality and disability by heat stroke.Our research team started an epidemiological investigation of heat stroke since 2002.On the basis of discovering organ injury rule,system info and treatment technology on critical medicine were applied to heat stroke treatment.Research on organ injury mechanism for heatstroke was carried out based on translational medicine idea,and periodic research results were also achieved.A series of key technologies for heat stroke treatment were obtained.These technologies were popularized in 30 hospitals across the country,thus improving ability of heat stroke treatment.

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.

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 (P<0.05), while at 2h, it was reduced significantly (P<0.05 or 0.01). In the antioxidant preprocessing group, the contractile response was reduced significantly from heat stress to 2h after heat stress (P<0.05 or 0.01). There was no statistically significant difference in contractile response between different NE concentrations in the control group and heat stress group (P>0.05), 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.