[Effects of low temperature exposure on dermal microvascular endothelial cells function].

OBJECTIVE: To explore the damage effects and expression of vascular endothelial growth factor (VEGF) exposed with different low-temperatures on rat dermal microvascular endothelial cells (DMVECs). METHODS: Primary DMVECs were obtained by discontinuous Percoll gradient centrifugation. The DMVECs were identified by phase contrast microscope and immunofluorescence studies for CD31 antigen. Applied 28 degrees C, 12 degrees C and 0 degrees C to interfere with rat DMVECs as cold-exposure model. The changes of cells morphology were observed under invert microscope. The membrane integrity was determined by lactate dehydrogenase (LDH) activity. RT-PCR was used to examine the expression of vascular endothelial growth factor mRNA in cells. RESULTS: The monolayer of cultured PMVECs displayed the shape of pavingstone. CD31 antigen and binding BSI results by fluorescence microscope identified the cultured cells were DMVECs. After 24 h cold exposure, the cell morphology of 0 degrees C group was shrunken, the other groups were "Fibroblast-like". The LDH activity (U/L) in the medium of 28 degrees C, 12 degrees C and 0 degrees C groups was 54.17 +/- 3.02, 64.66 +/- 3.03, 82.13 +/- 10.91 respectively, which was significantly higher than that of 37 degrees C group (12.23 +/- 3.0, P < 0.01). The VEGF mRNA expression level was up-regulated in 28 degrees C group and 12 degrees C group versus control group (P < 0.05), but unchanged in 0 degrees C group. CONCLUSION: The rat DMVECs injury severity are deteriorated with temperature decreasing, and VEGF might be involved in the regulation of membrane permeability in this period.

[The damage effects of hypothermia combined with hypoxia on rat pulmonary microvascular endothelial cells].

OBJECTIVE: To study the synergistic effects of hypothermia and hypoxia on the damage of pulmonary microvascular endothelial cells (PMVEC) in rat. METHODS: Primary PMVECs were obtained by complex phosphoesterasum digesting from isolated lung tissues of Wistar rats, the PMVECs were identified by phase contrast microscope and immunofluorescence studies for CD31 antigen and bandeiraea simplicifolia isolectin (BSI) binding test. Factorial design was adopted in trial according to hypothermia and hypoxia existing or not. Using corresponding kit measured the levels of lactate dehydrogenase (LDH) activity in cell medium. Level of nitric oxide (NO) concentration was measured by Griess Assay. RT-PCR was used to examine the expression of vascular endothelial growth factor (VEGF), endothelin-1 (ET-1) mRNA in PMVECs. RESULTS: The monolayer of cultured PMVECs displayed the shape of pavingstone. CD31 antigen and binding BSI results by fluorescence microscope identified the cultured cells were PMVECs. Compared to the control group, LDH activity and VEGF, ET-1 expression levels were significantly increased in hypothermia group, hypoxia group and hypoxia combined with hypothermia group. And the levels of NO concentration were reduced in these three groups. The results of One-way ANOVA showed that there was a synergistic effect between hypothermia and hypoxia. CONCLUSION: Hypothermia and hypoxia both have an effect on PMVECs whether in altering the cell permeability or in releasing of vasoactive substances including NO and ET-1. In addition, there is a synergistic effect between hypothermia and hypoxia.

[Changes of protein kinase A expressions in central amygdaloid nuclei during the process of chronic morphine-induced conditioned place aversion in rats].

OBJECTIVE: To explore neurobiological mechanisms of the withdrawal-induced aversion. The changes of protein kinase A were measured in central amygdaloid nucleic (CeA) of conditioned place aversion (CPA) model rats. METHODS: (1) All 72 male SD rats were divided into three groups, model group (MN group), and control group (MS group and SN group). MN group was injected with morphine,6.5 days, 10 mg/kg, intraperitoneally (ip), twice per day, naloxone injection, 0.3 mg/kg, ip, along with conditioned place aversion training, to develop the CPA model. The MS group was administrated equivalent volume of morphine and saline. Also the SN group was injected with equivalent volume of saline and naloxone. (2) During the process of morphine-induced CPA, the expression of protein kinase A was assayed with immunohistochemistry in the CeA. RESULTS: In the MN group, protein kinase A expressions in the CeA occurred adaptive changes at different points of CPA (P < 0.05). Protein kinase A expressions after establishment(Day7,134.43 +/- 4.481, P < 0.05), and after extinction (Day 13, 141.01 +/- 3.360, P < 0.01), and after reinstatement (Day 14,137.18 +/- 40.330, P < 0.05) were also lower than those before the establishment of the CPA (Day 5, 124.48 +/- 6.722). However, PKA expressions were not significantly different both in MS group (P > 0.05)and SN group (P > 0.05). CONCLUSION: (1) Protein kinase A expression, in turn regulating the aversion expression, in the CeA probably is a key pathway contributing to the development of CPA. (2) The neuroadaptation mediated by protein kinase A may be one of the important molecular underpinnings of CPA.

[Protective effects of SUR2B/Kir6.1 potassium channel opener natakalim against RAVECs injuries induced by hypoxia].

OBJECTIVE: To investigate the protective effects of natakalim against rat aortic vascular endothelial cells (RAVECs) injuries induced by hypoxia and its mechanisms. METHODS: Selecting RAVECs as a cell model injured by hypoxia, these RAVECs were divided into 5 groups: i.e. control group, hypoxia group, natakalim low, medium and high group. The cell survival rate was determined by MTT assay, con was measured using Griess Assay, RT-PCR was used to examine t he expression of intercellular adhesion molecule-1 (ICAM-1), vascular endothelial growth factor (VEGF), endothelin-1 (ET-1) mRNA in RAVEC. RESULTS: Natakalim could reverse hypoxia-induced changes in endothelial cell function, including increased endothelial cell survival rate and level of NO concentration, significantly inhibited the hypoxia-induced endothelial ICAM-1, ET-1, VEGF mRNA expression levels increased. CONCLUSION: Natakalim have protective effects on hypoxia-induced changes in endothelial cell function, increasing of permeation, excess expression of cell adhesion molecules.