Effect of sulodexide on apoptosis of human dermal microvascular endothelial cells exposed to hypoxia

AIM: To investigate the effect of sulodexide (SDX) on the apoptosis of human dermal microvascular endothelial cells (HDMECs) exposed to hypoxia and its underlying mechanism. METHODS: The HDMECs were cultured and divided into normoxia control group cultured under normoxic condition; hypoxia control group cultured in a humid incubator maintained at 37℃ with 5% CO₂ and 1% O₂ for 24 h; treatment groups treated with SDX at 0.25, 0.5 and 1 LSU/mL for 24 h under hypoxic condition. The cell viability was measured by CCK-8 assay. The apoptotic rate of the HDMECs was analyzed by flow cytometry. The activity of caspase-3 in HDMECs was examined by caspase-3 activity assay kit. The expression of pro-apoptotic factor P53, caspase-3, Bax and anti-apoptotic factor Bcl-2 at mRNA and protein levels was determined by real-time PCR and Western blot. RESULTS: SDX increased the viability of HDMECs exposed to hypoxia, but also decreased the apoptosis. Furthermore, SDX down-regulated the expression of pro-apoptotic factor P53, Bax and caspase-3 at mRNA and protein levels as well as the activity of caspase-3, while the expression of anti-apoptotic factor Bcl-2 was up-regulated. CONCLUSION: SDX significantly increases the viability and decreases the apoptosis of HDMECs exposed to hypoxia. Inhibition of the mitochondrial apoptosis pathway may be involved in the underlying mechanism.     

Effects of tanshinone IIA on proliferation,apoptosis and expression of HIF-1α, VEGF and wild-type P53 in human hepatoma HepG2 cells under hypoxia

AIM:To investigate the effects of tanshinone IIA (Tan IIA) on proliferation, apoptosis and its molecular mechanism in human hepatoma HepG2 cells under hypoxic condition. METHODS:Hypoxia model was established by treatment with cobalt chloride (CoCl2). The cells were divided into normoxia control group, hypoxia control group and hypoxia combined at different concentrations of Tan IIA groups. After HepG2 cells were incubated with different concentrations of Tan IIA (0.5, 1.0, 2.0, 5.0 and 10.0 mg/L) for 24 h, 48 h and 72 h under hypoxic condition, the cell proliferation was determined by MTT assay. After Tan IIA was added to the media at different concentrations for 24 h and 48 h, the apoptotic cells were observed by Hoechst 33258 staining. The protein levels of hypoxia-inducible factor 1 alpha (HIF-1α), vascular endothelial growth factor (VEGF) and wild-type P53 were detected by Western blotting after cultured with different concentrations of Tan IIA for 48 h. RESULTS:Tan IIA inhibited the proliferation of HepG2 cells in a dose- and time-dependent manner. Tan IIA induced the typical morphology of apoptotic cells and increased the apoptotic rate in a dose- and time-dependent manner after treatment with 1.0 mg/L~5.0 mg/L for 24 h and 48 h under hypoxic condition. The protein levels of HIF-1α and VEGF were weakly expressed in HepG2 cells under normoxia but up-regulated after incubated under hypoxia for 48 h. The protein expression of HIF-1α and VEGF were decreased with the increase in the concentration of Tan IIA under hypoxia. The protein expression of wild-type P53 was increased with the increase in the concentrations of Tan IIA under hypoxia. CONCLUSION: Tan IIA significantly inhibits the proliferation and induces the apoptosis of human hepatoma HepG2 cells under hypoxia, which may be related to the down-regulation of HIF-1α and VEGF and up-regulation of wild-type P53.     

缺氧条件下‘黄富士’苹果挥发性成分变化趋势研究

通过测定‘黄富士’苹果在缺氧条件下挥发性产物含量的变化,分析挥发性成分变化与缺氧伤害的关系,为苹果低氧伤害的预警,保证气调贮藏品质提供基础方法。通过固相微萃取(SPME)技术萃取,采用气相色谱-质谱联用技术,对缺氧条件下2、4、6、8、10天挥发物进行成分分析。研究发现,随着缺氧时间的延长,苹果中共有18种挥发性成分含量下降,6种挥发性成分上升。筛选出的24种挥发性成分与缺氧伤害存在相关性,可为进一步筛选缺氧预警指标、研究缺氧伤害机理提供参考。     

Effects of acute and chronic hypobaric hypoxia on spatial learning and memory functions in adult rats

AIM: To explore the effects of acute and chronic hypobaric hypoxia on spatial learning and memory functions in adult rats. METHODS: Three separate experiments were carried out. In the first experiment, 30 adult male SD rats were divided into control group (group A) and acute hypoxia group (group B, 7 000 m, 72 h). Learning and memory functions were tested by Morris water maze. The spatial acquisition was performed 4 trails per day, 3 days to reach asymptotic performance. The platform was removed on the 4th day, using a novel start position during the probe trail. The data of escape latency, time in platform quadrant and times of passing platform were recorded. In the second experiment, 26 adult male SD rats were divided into control group (group C) and chronic hypoxia group (group D, 6 000 m, 35 d). The spatial acquisition was performed 4 trails per day, 5 days to reach asymptotic performance. The platform on the 6th day was removed, using a novel start position during the probe trail. The data of escape latency, time in platform quadrant and times of passing platform were recorded. In the third experiment, after trained with Morris water maze by using the method as the second experiment, 30 adult male SD rats were divided into control group (group E) and acute hypoxia group (group F, 7 000 m, 72 h), and 2 h later, the memory functions were reevaluated. RESULTS: In the first day after acute hypoxia exposure, the escape latency in group B was significantly shorter than that in group A. However, the escape latency in the following days and the and times of passing platform was not significantly different. No difference of the escape latency in the following days and the time of passing platform between group C and D was observed. Memory tests didn’t show any difference between group E and F. CONCLUSION: Either acute or chronic hypobaric hypoxia does not affect the spatial learning and memory functions in adult rats.     

Electrophysiological effects of lidocaine on myocardial tissue in guinea-pig left ventricular outflow tract under conditions of hypoxia,acidosis and treatment with epinephrine

AIM：To study the electrophysiological effects of lidocaine on the myocardial tissue in guinea-pig left ventricular outflow tract under the conditions of hypoxia, acidosis and treatment with epinephrine. METHODS：The action potentials of pacemaker cells in guinea-pig left ventricular outflow tract were recorded by conventional technique with intracellular microelectrodes. The effects of lidocaine on the spontaneous slow response potentials were investigated under the conditions of hypoxia, acidosis and treatment with epinephrine (EPI). RESULTS：Lidocaine markedly decreased the rate of pacemaker firing (RPF), the velocity of diastolic depolarization (VDD), the maximal rate of depolarization (Vmax), the maximal diastolic potential (MDP) and the amplitude of action potential (APA). Lidocaine also shortened the 50% and 80% of duration of action potential (APD50 and APD80). At the concentrations from 0.1 μmol/L to 10 μmol/L, the effects of lidocaine were more significant. Under the condition of hypoxia and perfusion with deprived glucose content for 15 min, VDD, RPF, Vmax, MDP and APA significantly decreased, and APD50 notably shortened. Under the condition of hypoxia, lidocaine at 1 μmol/L significantly decreased VDD, RPF, Vmax and APA as compared with the cells treated with hypoxia only. Perfusion with pH 6.8 solution for 10 min, VDD, RPF, Vmax and APA significantly decreased, MDP notably increased, and APD50 and APD80 markedly shortened. Under the condition of acidosis for 10 min, lidocaine significantly decreased VDD, RPF and Vmax, and lengthened APD50 and APD80 as compared with the cells under the condition of acidosis alone. Perfusion with EPI at 10 μmol/L for 10 min resulted in significant increases in VDD, RPF, Vmax, MDP and APA, and notable shortenings of APD50 and APD80 were also observed. Compared with 10 μmol/L EPI group, 1 μmol/L lidocaine+10 μmol/L EPI significantly reduced VDD, RPF, MDP and APA, and lengthened APD50 and APD80. CONCLUSION：Lidocaine markedly decreases the autorhythmicity of the pacemaker cells in guinea-pig left ventricular outflow tract and influences the electrophysiological effects of hypoxia, acidosis and EPI.     

Hypoxia facilitates rapid pacing-induced calcium transient alternations in ventricular myocytes

AIM:To explore the effect of hypoxia on rapid pacing-induced calcium transient alternations in ventricular myocytes.METHODS:Ventricular myocytes were isolated from the heart of adult SD rats and cultured in serum-free hypoxic fluid to set up an in vitro model of hypoxia-induced cardiac injury. The calcium transient and its alternations were investigated under confocal laser scanning microscope. The mitochondrial function was also examined by WST-8 kit. RESULTS:Under normoxic condition, the ventricular myocytes were claviform. Low frequency of pacing, ranging from 60 to 240 min^-1, induced calcium transient, but not calcium transient alternations, which was elicited by the pacing over a threshold frequency of(288 ±27)min^-1. Exposure of the ventricular myocytes to hypoxia did not obviously affect the morphology of the cells, but reduced the threshold frequency of pacing to(227±26) min^-1(P<0.05). Additionally, exposure of the cells to hypoxia repressed the activity of mitochondrial dehydrogenase from(100.2±8.7)%(control group) to(57.6±7.5)%, which was partially blocked by L-type Ca²⁺ channel inhibitor. CONCLUSION:Hypoxia facilitates calcium transient alternations induced by rapid pacing, and the calcium transient alternations are involved in the hypoxia-injured mitochondria function.     

Cloning and sequencing of encoding region of heme oxygenase-1 gene from Tibetan antelope

AIM: To clone and analyze the encoding region of heme oxygenase-1(HO-1) gene from Tibetan antelope(Pantholops hodgsonii). METHODS:The total RNA was isolated from the liver of Tibetan antelope and the HO-1 gene was amplified by RT-PCR. The PCR product was cloned into pGEM-T vector and sequenced. The nucleotide sequences were compared with the data from GenBank by BLAST method. RESULTS:The encoding region of HO-1 gene from Tibetan antelope was obtained and deposited in GenBank as accession number JQ809687. The encoding sequence was 897 bp in full length, which encoded 298 amino acids. Sequence similarity analysis showed that the HO-1 gene cloned from Tibetan antelope shared 98% and 96% in the nucleotides, and shared 92% and 97% in the amino acids with those of Capra hircusand Bos taurus, respectively. The sequence similarity of nucleotides and amino acids also shared 86% and 87% with other vertebrates, which were highly conservative. The molecular phylogenetic tree based on the amino sequence of HO-1 showed that Tibetan antelope and Capra hircus was classified to one cluster, which was basically concerted to the evolutionary relationship of traditional species. CONCLUSION:The encoding region of Tibetan antelope HO-1 gene is successfully cloned, which provides a foundation for exploring the molecular and biological mechanisms of high altitude adaptation and cell protection in Qinghai-Tibet Plateau species under hypoxic condition.