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Acute kidney injury and acute lung injury/acute respiratory distress syndrome are common in the pediatric intensive care unit.Lung-kidney interaction in critically ill patients is closely related to anoxia, fluid management, and inflammatory response in acute kidney injury and acute lung injury/acute respiratory distress syndrome patients.Strengthening the understanding of lung-kidney interaction can help clinicians to systematically manage critically ill patients.
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The vascular endothelial cells (VECs) hypoxia/reoxygenation(H/R) model is a classic cell model that simulates vascular endothelial ischemia/reperfusion (I/R) injury and related diseases. It has the advantages of convenient operation, intuitive image, and good stability, and can accurately reflect pathological changes at the cellular level of diseases. It is widely used in the study of molecular mechanisms of drugs and diseases.There are many similarities in the mechanism and formation between the H/R model and the I/R injury model, but the I/R model is more complex. Therefore, in recent years, many scholars have used the H/R model to simulate the I/R model for experimental research, and believe that the H/R model is also an ideal model for studying I/R. By implementing intervention measures on the established H/R model of VECs, the potential effects of the intervention measures in clinical practice can be verified, which has guiding significance for how to prevent, treat, and how to exacerbate I/R injury in clinical practice. This article introduces the different methods used by scholars in recent years, such as medium deoxygenation and mixed gas culture method, to construct H/R models using VECs cultured in vitro to simulate I/R models. The differences in methods used and the subtle differences between the same methods are also discussed. At the same time, due to the relatively single method of constructing H/R models at present, how to find new, more efficient and affordable methods based on scientific and reasonable experiments has also become a focus of attention.
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Hypoxia is a common phenomenon of solid tumor, which is closely related to the malignant proliferation, tumor progression, radiotherapy and chemotherapy resistance, treatment failure, and poor prognosis. At present, many researches focus on the application of medical imaging and nuclear medicine methods in detecting the hypoxic areas of tumors. This article focuses on the detection of hypoxia microenvironment and the application of PET tracers in tumor hypoxia imaging.
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[Abstract] China has a vast territory and often needs workers in a special environment. Working and training in a special environment (such as high temperature, plateau anoxia, cold, long voyage, and so on) will put forward higher requirements for the body, and the body will also have adaptive changes. However, once the compensation range of the body is exceeded, there will be a series of discomfort and even pathological changes. A small number of literatures have described the major changes of the whole body under various special conditions, but there is still a lack of related summary of the metabolic changes, compensatory changes and pathological changes of kidney under various extreme environments. Therefore, the compensatory modification of the kidney and the possible causes of injury in various special environments have been summarized and analyzed in present paper, and combined with the related pathophysiological mechanism, the related protective measures of kidney have also been introduced.
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Objective@#To investigate blood pressure and vascular remodeling of OSAS by establishing the chronic-intermittent hypoxia model in rat.@*Methods@#Experiments were performed on 35 adult male Sprague-Dawley rats. Animals were randomly divided into four groups: unhandled control group (with 5 rats in it), CIH group at 9/6/3 weeks (with 10 ratsin each group). Rats in CIH group went through 8-hour intermittent hypoxia everyday, and those in control group were raising normally. After 9-week experiment, blood pressure was measured. The changes of the following indexes were observed: pathological changes of aorta and the middle aorta thickness (HE staining), the collagen of aorta wall (Masson staining). The experimental data were analyzed by SPSS 24.0 statistical software. The variance was analyzed by one-way analysis of variance, and the irregularity was selected using the calibration t test.@*Results@#The systolic and diastolic blood pressures of the CIH9, 6, and 3 weeks groups and the control group were: (127±13) and (79±9), (124±11) and (81±7), (101±11) and (75±9), (91±10) and (65±9) mmHg (1 mmHg=0.133 kPa). The systolic blood pressure and diastolic blood pressure of the rats in the week of CIH 9 and 6 weeks were significantly higher than the control group (F=14.64, P=0.000; F=6.81, P=0.000). There was no significant difference in the mean blood pressure between the three groups of CIH and the control group. Membrane thickness in CIH9, 6 and 3 weeks and control group were: (20±2), (19±2), (14±2), (13±3) μm. Compared with the control group, the aortic pathology and thickness of the middle layer of the CIH9 and 6 weeks group were significantly thicker (F=20.24, P=0.000), but there was no significant difference between the CIH3 week group and the control group; the collagen deposition was unchanged compared with the control group.@*Conclusion@#Intermittent hypoxia for 6 weeks or more in rats resulted in the increasement of blood pressure, morphological changes of aorta and vascular remodeling in thickened media.
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Objective@#To understand the mechanism of chemotherapy resistance in nasopharyngeal carcinoma under hypoxic conditions through the perspective of protein SUMOylation modification.@*Methods@#Cobalt chloride (CoCl2) was used to establish the hypoxic model of human nasopharyngeal carcinoma CNE1 cells. Then, the cell cycle was detected by flow cytometry, and the expression level of small ubiquitin-related modifier(SUMO) and cyclin-dependent kinase 6 (CDK6) proteins were detected by western blotting. MTT assay was used to determine the median lethal dose (IC50) of cancer cells against cisplatin, and enzyme-linked immunosorbent assay (ELISA) was used to determine lactate dehydrogenase (LDH) level.@*Results@#The cell cycle of CNE1 induced by hypoxia was arrested in G0/G1 phase.The results of Western blot showed that the protein expression level of CDK6 in CNE1 cells was lower than that in the control group (0.83±0.25 vs. 0.43±0.21, t=14.67, P=0.003). The protein level of conjugated SUMO1 was significantly lower than that in the control group (2.69±0.48 vs. 1.38±0.31, t=17.22, P=0.001), while the level of free SUMO1 protein was significantly higher than that in the control group (2.01±0.43 vs. 2.60±0.59, t=15.45, P=0.002).The LC50 of CNE1 cells in the control group was significantly lower than that in the hypoxic group (29.44 μg/ml vs. 97.72 μg/ml, t=12.79, P=0.001). After CNE1 cells received 50 μg/ml cisplatin for 48 h, the LDH content in the supernatant of the control group was significantly higher than that in the hypoxic group ((541.49±64.59) ng/ml vs. (234.67±41.03) ng/ml, t=11.94, P=0.007)). The apoptosis rate of CNE1 cells in the control group was significantly higher than that in the hypoxic group ((76.64±5.37)% vs. (32.84±4.77) ng/ml, t=8.49, P=0.003)).@*Conclusion@#Hypoxia can dissociate the covalent modification of CDK6 and SUMO1, inhibit cell cycle and increase the chemotherapy resistance of nasopharyngeal carcinoma.
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Objective@#To explore the effects of transient receptor potential vanilloid 1 (TRPV1) on autophagy in early hypoxic mouse cardiomyocytes and the mechanism in vitro.@*Methods@#The hearts of 120 C57BL/6 mice aged 1-2 days, no matter male or female, were isolated, and then primary cardiomyocytes were cultured and used for the following experiments, the random number table was used for grouping. (1) The cells were divided into normoxia group and hypoxia 3, 6, and 9 h groups, with one well in each group. The cells in normoxia group were routinely cultured (the same below), the cells in hypoxia 3, 6, and 9 h groups were treated with fetal bovine serum-free and glucose-free Dulbecco′ s modified Eagle medium under low oxygen condition in a volume fraction of 1% oxygen, 5% carbon dioxide, and 94% nitrogen for 3, 6, and 9 h, respectively. The protein expressions of microtubule-associated protein 1 light chain 3 (LC3), Beclin-1, TRPV1 were determined with Western botting. (2) The cells were divided into normoxia group and hypoxia group, with two coverslips in each group. The cells in hypoxia group were treated with hypoxia for 6 h as above. The positive expression of TRPV1 was detected by immunofluorescence assay. (3) The cells were divided into 4 groups, with one well in each group. The cells in simple hypoxia group were treated with hypoxia for 6 h as above, and the cells in hypoxia+ 0.1 μmol/L capsaicin group, hypoxia+ 1.0 μmol/L capsaicin group, and hypoxia+ 10.0 μmol/L capsaicin group were respectively treated with 0.1, 1.0, 10.0 μmol/L capsaicin for 30 min before hypoxia for 6 h. The protein expressions of LC3, Beclin-1, and TRPV1 were detected by Western blotting. (4) The cells were divided into 5 groups, with 5 wells in each group. The cells in hypoxia group were treated with hypoxia for 6 h as above, the cells in hypoxia+ chloroquine group, hypoxia+ capsaicin group, and hypoxia+ capsaicin+ chloroquine group were treated with hypoxia for 6 h after being cultured with 50 μmol/L chloroquine, 10.0 μmol/L capsaicin, and 50 μmol/L chloroquine+ 10.0 μmol/L capsaicin for 30 min, respectively. Viability of cells was detected by cell counting kit 8 assay. (5) The cells were divided into simple hypoxia group and hypoxia+ 10.0 μmol/L capsaicin group, with one well in each group. The cells in hypoxia group were treated with hypoxia for 6 h as above, the cells in hypoxia+ 10.0 μmol/L capsaicin group were treated with 10.0 μmol/L capsaicin for 30 minutes and then with hypoxia for 6 h. The protein expressions of lysosomal associated membrane protein 1 (LAMP-1) and LAMP-2 were detected by Western blotting. Each experiment was repeated for 3 or 5 times. Data were processed with one-way analysis of variance, least significant difference t test, and Bonferroni correction.@*Results@#(1) Compared with those of normoxia group, the protein expressions of LC3, Beclin-1, and TRPV1 were significantly increased in cardiomyocytes of hypoxia 3, 6, and 9 h groups (t3 h=4.891, 5.890, 4.928; t6 h=9.790, 6.750, 10.590; t9 h=6.948, 6.764, 5.049, P<0.05 or P<0.01), which of hypoxia 6 h group were the highest (1.08±0.05, 1.12±0.10, 0.953±0.071, respectively). (2) The density of TRPV1 in cell membrane and inside the cardiomyocytes in hypoxia group was significantly increased with lump-like distribution, and the expression of TRPV1 was higher than that in normoxia group. (3) Compared with those of simple hypoxia group, the protein expression of Beclin-1 in cardiomyocytes of hypoxia+ 0.1 μmol/L capsaicin group was increased (t=10.488, P<0.01), while the protein expressions of LC3 and TRPV1 were increased without statistically significant differences (t=4.372, 3.026, P>0.05); the protein expressions of LC3, TRPV1, and Beclin-1 in cardiomyocytes of hypoxia+ 1.0 μmol/L capsaicin group and hypoxia+ 10.0 μmol/L capsaicin group were significantly increased (t=15.505, 5.773, 13.430; 20.915, 8.054, 16.384; P<0.05 or P<0.01), which of hypoxia+ 10.0 μmol/L capsaicin group were the highest (2.33±0.09, 1.34±0.07, 1.246±0.053, respectively). (4) Compared with 0.585±0.045 in normoxia group, the cardiomyocyte viability in hypoxia group was significantly decreased (0.471±0.037, t=4.365, P<0.05). Compared with that in hypoxia group, the cardiomyocyte viability in hypoxia+ chloroquine group was further decreased (0.350±0.023, t=6.216, P<0.01), while 0.564±0.047 in hypoxia+ capsaicin group was significantly increased (t=3.489, P<0.05). Compared with that in hypoxia+ chloroquine group, the cardiomyocyte viability in hypoxia+ capsaicin+ chloroquine group did not significantly change (0.364±0.050, t=0.545, P>0.05). (5) Compared with 0.99±0.04 and 0.54±0.04 in simple hypoxia group, the protein expressions of LAMP-1 and LAMP-2 in hypoxia+ 10.0 μmol/L capsaicin group were significantly increased (1.49±0.06, 0.81±0.05, t=12.550, 7.442, P<0.01).@*Conclusions@#TRPV1 can further promote the expression of autophagy-related proteins in hypoxic cardiomyocytes through autophagy-lysosomal pathway, enhance autophagy activity, and improve autophagic flow for alleviating early hypoxic cardiomyocyte injury.
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Objective@#To investigate the role of hexokinase Ⅱ in the changes of autophagic flow in cardiomyocytes of mice with ischemia-hypoxia in vitro.@*Methods@#The hearts of totally six male and female C57BL/6 mice aged from 1 to 2 days were isolated to culture primary cardiomyocytes which were used for the following experiments. (1) The cells were divided into 6 groups according to the random number table (the same grouping method below), i. e., normal control 3, 6, and 9 h groups and ischemia-hypoxia 3, 6, and 9 h groups, with 4 wells in each group. After being regularly cultured for 48 h with Dulbecco′s modified Eagle medium/nutrient mixture F12 (DMEM/F12) medium (the same regular culture condition below), the cells in normal control 3, 6, and 9 h groups were cultured with replaced fresh DMEM/F12 medium for 3, 6, and 9 h, respectively, and the cells in ischemia-hypoxia 3, 6, and 9 h groups were cultured with replaced sugar-free serum-free medium in the low-oxygen incubator with a volume fraction of 1% oxygen and a volume fraction of 5% carbon dioxide at 37 ℃ (the same hypoxic culture condition below) for 3, 6, and 9 h, respectively. Cell viability was measured by the cell counting kit 8 (CCK-8) method. (2) The cells were grouped and treated the same as those in experiment (1), with 1 well in each group. Western blotting was used to detect the protein expressions of microtubule-associated protein 1 light chain 3 Ⅰ (LC3Ⅰ), LC3Ⅱ, p62, and hexokinase Ⅱ. (3) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, and ischemia-hypoxia 9 h+ 2-deoxyglucose (2-DG) group, with 4 wells in each group. After a regular culture for 48 h, the cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h; the cells in simple ischemia-hypoxia 9 h group were replaced with sugar-free serum-free medium, and the cells in ischemia-hypoxia 9 h+ 2-DG group were replaced with sugar-free serum-free medium in which 2-DG was dissolved in a concentration of 10 mmol/L (20 μmol), and then they were cultured with hypoxia for 9 h. Cell viability was measured by CCK-8 method. (4) The cells were grouped and treated the same as those in experiment (3), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, and p62. (5) The cells were grouped and treated the same as those in experiment (3), with 2 wells in each group. Transmission electron microscope was used to observe autophagosomes/autolysosomes in cardiomyocytes. (6) The cells were divided into normal control group, simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ hexosinase Ⅱ small interfering RNA1 (HK-ⅡsiRNA1) group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group, with 4 wells in each group. The cells in normal control group and simple ischemia-hypoxia 9 h group were regularly cultured for 48 h, and the cells in ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were respectively transfected with 200 nmol/L HK-ⅡsiRNA1 and HK-ⅡsiRNA2 and then also cultured for 48 h. The cells in normal control group were cultured with replaced fresh DMEM/F12 medium for 9 h, and the cells in simple ischemia-hypoxia 9 h group, ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group, and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were cultured with replaced sugar-free serum-free medium and hypoxia for 9 h. Cell viability was measured by CCK-8 method. (7) The cells were grouped and treated the same as those in experiment (6), with 1 well in each group. Western blotting was used to detect the protein expressions of LC3Ⅰ, LC3Ⅱ, p62, and hexokinase Ⅱ. Except for experiment (5), each experiment was repeated 3 times. Data were processed with one-way analysis of variance and lest significant difference t test, and Bonferroni correction.@*Results@#(1) The viabilities of cardiomyocytes in ischemia-hypoxia 3, 6, and 9 h groups were 0.450±0.022, 0.385±0.010, and 0.335±0.015, respectively, which were significantly lower than 0.662±0.026, 0.656±0.028, and 0.661±0.021 of the corresponding normal control 3, 6, and 9 h groups, respectively (t=6.21, 9.12, 12.48, P<0.01). (2) Compared with those of corresponding normal control 3, 6, and 9 h groups, the LC3Ⅱ/Ⅰ ratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of ischemia-hypoxia 3, 6, and 9 h groups were significantly increased (t3 h=16.15, 10.99, 5.30, t6 h=6.79, 10.42, 9.42, t9 h=15.76, 16.51, 7.20, P<0.05 or P<0.01). (3) The viability of cardiomyocytes in simple ischemia-hypoxia 9 h group was 0.353±0.022, which was significantly lower than 0.673±0.027 of normal control group (t=9.29, P<0.01). The viability of cardiomyocytes in ischemia-hypoxia 9 h+ 2-DG group was 0.472±0.025, which was significantly higher than that of simple ischemia-hypoxia 9 h group (t=3.60, P<0.05). (4) Compared with those of normal control group, the LC3Ⅱ/Ⅰ ratio and protein expression of p62 in cardiomyocytes of simple ischemia-hypoxia 9 h group were significantly increased (t=9.45, 8.40, P<0.01). Compared with those of simple ischemia-hypoxia 9 h group, the LC3Ⅱ/Ⅰratio and protein expression of p62 in cardiomyocytes of ischemia-hypoxia 9 h+ 2-DG group were significantly decreased (t=4.39, 4.74, P<0.05). (5) In cardiomyocytes of normal control group, only single autophagosome/autolysosome with bilayer membrane structure was observed. Compared with that of normal control group, the number of autophagosome/autolysosome with bilayer membrane structure in cardiomyocytes of simple ischemia-hypoxia 9 h group was increased significantly. Compared with that of simple ischemia-hypoxia 9 h group, the number of autophagosome/autolysosome with bilayer membrane structure in cardiomyocytes of ischemia-hypoxia 9 h+ 2-DG group was significantly decreased. (6) The viability of cardiomyocytes in simple ischemia-hypoxia 9 h group was 0.358±0.023, which was significantly lower than 0.673±0.026 in normal control group (t=9.12, P<0.01). The viabilities of cardiomyocytes in ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were 0.487±0.027 and 0.493±0.022, respectively, which were significantly higher than the viability in simple ischemia-hypoxia 9 h group (t=3.63, 4.28, P<0.05). (7) Compared with those of normal control group, the LC3Ⅱ/Ⅰratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of simple ischemia-hypoxia 9 h group were significantly increased (t=6.08, 6.31, 4.83, P<0.05 or P<0.01). Compared with those of simple ischemia-hypoxia 9 h group, the LC3Ⅱ/Ⅰ ratio and protein expressions of p62 and hexokinase Ⅱ in cardiomyocytes of ischemia-hypoxia 9 h+ HK-ⅡsiRNA1 group and ischemia-hypoxia 9 h+ HK-ⅡsiRNA2 group were significantly decreased (t=5.10, 7.76, 15.33, 4.17, 8.42, 12.11, P<0.05 or P<0.01).@*Conclusions@#Ischemia-hypoxia upregulates the expression level of hexokinase Ⅱ protein in mouse cardiomyocytes cultured in vitro, which decreases the viability of cardiomyocytes by impairing autophagic flow. To inhibit the activity of hexokinase Ⅱ or its expression can alleviate the ischemia-hypoxia damage of cardiomyocytes.
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Objective@#To investigate the protective effects and mechanism of keratinocyte growth factor (KGF) combined with hypoxia inducible factor-1α (HIF-1α) on intestinal crypt epithelial cells (IEC-6) of rats with hypoxia stress.@*Methods@#(1) The routinely cultured IEC-6 of rats were collected and divided into normoxia blank group, normoxia KGF group, normoxia HIF-1α group, and normoxia combine group, according to the random number table, and then the previous mediums were respectively replaced with dulbecco′s modified eagle medium (DMEM), medium with 0.5 ng/mL KGF, medium with 10.0 ng/mL HIF-1α, and medium with 0.5 ng/mL KGF and 30.0 ng/mL HIF-1α. And the cells were cultured in cell incubator with oxygen volume fraction of 21% for 24 hours. (2) Another batch of routinely cultured IEC-6 were collected and divided into normoxia control group, hypoxia control group, hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group, according to the random number table. The previous mediums were replaced with DMEM, DMEM, medium with 0.5 ng/mL KGF, medium with 10.0 ng/mL HIF-1α, and medium with 0.5 ng/mL KGF and 30.0 ng/mL HIF-1α respectively. And then, the cells in normoxia control group were cultured routinely for 24 hours, and cells in the other 4 groups were cultured in cells incubator of 3 gases, with oxygen volume fraction of 5% for 24 hours. Cells cultured in normoxic and hypoxic incubators were collected, with 3 samples in each group, and morphological changes of cells were observed with optical microscope. Cells cultured in normoxic and hypoxic incubators were collected, with 3 samples in each group, and survival rates of cells were detected by cell count kit 8. Cells in normoxia control group and cells cultured in hypoxic incubator were collected, with 3 samples in each group. The cell cycle changes and apoptosis rates were detected by flow cytometer, the content of adenosine triphosphate (ATP) was detected by ultraviolet spectrophotometer, and protein expression of p53 was detected by Western blotting. Data were processed with one-way analysis of variance and least significant difference test.@*Results@#(1) After being cultured for 24 h, cells cultured in normoxic incubator grew well with oval or round shapes and clear cytoplasm, and cells cultured in hypoxic incubator showed irregular shapes such as fusiform or starlike shape, with black particle in cytoplasm. (2) After being cultured for 24 h, cell survival rates of normoxia blank group, normoxia KGF group, normoxia HIF-1α group, and normoxia combine group were (107.4±8.7)%, (109.8±2.9)%, (115.8±7.4)%, and (112.8±10.6)% respectively. There was no significantly statistical difference in general comparison of cell survival rates among the above groups (F=0.685, P=0.586). After being cultured for 24 h, cell survival rates of hypoxia control group, hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group were (35.1±4.6)%, (52.9±6.8)%, (56.2±3.1)%, and (71.2±9.6)% respectively, which were significantly lower than (106.3±12.3)% of normoxia control group (P<0.001). Survival rates of cells in hypoxia KGF group, hypoxia HIF-1α group, and hypoxia combine group were significantly higher than the rate of cells in hypoxia control group (P=0.023, 0.009, <0.001). Survival rate of cells in hypoxia combine group was significantly higher than the rates of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.017, 0.045). (3) After being cultured for 24 h, percentage of cells in G1 phase in hypoxia control group was significantly higher than that of cells in normoxia control group (P=0.030), percentages of cells in S phase in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously lower than the percentage of cells in normoxia control group (P=0.020, 0.031, 0.026), and percentages of cells in different phases in other groups were close to those of cells in normoxia control group (P=0.516, 0.107, 0.052, 0.985, 0.637, 0.465, 0.314, 0.591). After being cultured for 24 h, percentages of cells in G1 phase in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously higher than the percentage of cells in hypoxia combine group (P=0.001, 0.030, 0.014), and percentages of cells in S phase in the above 3 groups were obviously lower than the percentage of cells in hypoxia combine group (P=0.001, 0.012, 0.010). (4) After being cultured for 24 h, compared with that of cells in normoxia control group, apoptosis rate of cells in hypoxia control group obviously increased (P=0.018), and apoptosis rate of cells in hypoxia combine group obviously decreased (P=0.008). After being cultured for 24 h, compared with that of cells in hypoxia control group, apoptosis rates of cells in hypoxia KGF group and hypoxia combine group obviously decreased (P=0.004, 0.001). Apoptosis rate of cells in hypoxia combine group was obviously lower than those of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.032, 0.002). (5) After being cultured for 24 h, compared with that of cells in normoxia control group, the content of ATP of cells in hypoxia combine group changed unobviously (P=0.209), and content of ATP of cells in the other groups obviously decreased (P= <0.001, 0.001, 0.002). Content of ATP of cells in hypoxia HIF-1α group and hypoxia combine group was obviously higher than that of cells in hypoxia control group (P=0.044, 0.001). Content of ATP of cells in hypoxia combine group was obviously higher than that of cells in hypoxia KGF group and hypoxia HIF-1α group (P=0.011, 0.020). (6) After being cultured for 24 h, protein expressions of p53 of cells in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group were obviously higher than that of cells in normoxia control group (P<0.001), and protein expression of p53 of cells in hypoxia combine group was obviously lower than those of cells in hypoxia control group, hypoxia KGF group, and hypoxia HIF-1α group (P=0.001, 0.001, 0.002).@*Conclusions@#KGF combined with HIF-1α have significant protective effects on IEC-6 of rats with hypoxia stress, and can improve its survival in hypoxic environment by inhibiting cell cycle arrest, reducing the level of apoptosis, and increasing level of energy metabolism.
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Objective@#To explore the risk stratification of pulse oxygen saturation (SpO2) in patients with emergency hypoxemia patients, and to provide evidence for the identification of critical illness.@*Methods@#Self-designed clinical data registration form for patients with emergency hypoxemia, and prospective collection of 344 hypoxemia patients in the emergency department of Peking Union Medical College Hospital from March to April in 2018, including baseline data (name, gender, age, ID number, date, registration time), hospitalization method, past history, patient complaint and diagnosis, triage level, SpO2, whether to enter the rescue room, etc.@*Results@#All of 344 emergency hypoxemia patients, there were 163 cases (21.2%) of ambulances, and 107 cases (31.1%) of patients requiring immediate rescue. There were 54 cases (25.7%) and 53 cases (39.6%) in need of immediate rescue in day shift (8:00-20:00) and night shift (20:00-8:00 next day), with 9:00-10:00, 14:00-15:00, 20:00-24:00 in the majority. There was a statistical difference in the way of hospitalization, triage, and SpO2 (χ2=29.537, 25.780, t=4.722, all P<0.05) . SpO2 risk stratification was 0.905 in patients without pulmonary disease, and SpO2 risk stratification in patients with lung disease was 0.765.@*Conclusions@#Patients with hypoxemia account for a certain proportion in the emergency department and are in critical condition. The degree of critical condition of patients can be evaluated based on whether they have lung diseases, and the risk stratification of patients can be accurately determined with the help of SpO2, so as to further guide the hierarchical treatment measures for patients with emergency hypoxemia and rationally optimize the allocation of emergency resources.
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Objective To explore the risk stratification of pulse oxygen saturation (SpO2) in patients with emergency hypoxemia patients, and to provide evidence for the identification of critical illness. Methods Self-designed clinical data registration form for patients with emergency hypoxemia , and prospective collection of 344 hypoxemia patients in the emergency department of Peking Union Medical College Hospital from March to April in 2018, including baseline data (name, gender, age, ID number, date, registration time), hospitalization method, past history, patient complaint and diagnosis, triage level, SpO2, whether to enter the rescue room, etc. Results All of 344 emergency hypoxemia patients, there were 163 cases (21.2% ) of ambulances, and 107 cases (31.1% ) of patients requiring immediate rescue. There were 54 cases (25.7%) and 53 cases (39.6%) in need of immediate rescue in day shift (8:00-20:00) and night shift (20:00-8:00 next day), with 9:00-10:00, 14:00-15:00, 20:00-24:00 in the majority. There was a statistical difference in the way of hospitalization, triage, and SpO(2 25.780, t=4.722, all P<0.05). SpO2 risk stratification was 0.905 in patients without pulmonary disease, and SpO2 risk stratification in patients with lung disease was 0.765. Conclusions Patients with hypoxemia account for a certain proportion in the emergency department and are in critical condition. The degree of critical condition of patients can be evaluated based on whether they have lung diseases, and the risk stratification of patients can be accurately determined with the help of SpO2, so as to further guide the hierarchical treatment measures for patients with emergency hypoxemia and rationally optimize the χ2=29.537, allocation of emergency resources.
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Objective To investigate the pathways by which hypoxia aggravates the neurotoxic effects of amyloid-beta protein (Aβ) on neurons.Methods Human neuroblastoma cells (SH-SY5Y cells) were cultured in vitro,and were divided into four groups:the control group,Aβ intervention group,hypoxia group,Aβ intervention plus hypoxia group.Quantitative real time polymerase chain reaction(qRT-PCR) was adopted to detect the mRNA expression levels of p21-activated kinase 1 (PAK1),LIM kinase 1 protein (LIMK1)and cofilin.Western blot was used to measure the protein levels of PAK1,LIMK1,phosphate-LIMK1 (P-LIMK1),cofilin and phosphate-cofilin (P-cofilin).Results After Aβ treatment,the activity of SH-SY5Y cells was decreased.Compared with the control group,the protein levels of PAK1,LIMK1,P-LIMK1,P-cofilin,and the mRNA expression levels of PAK1 and LIMK1 were decreased(all P＜0.05),but the protein and mRNA expression of cofilin had no significant changes after 24 h of treatment with 10μmol/L Aβ.Compared with the Aβ intervention group,the protein levels of PAK 1,LIMK1,P-LIMK 1 and P-cofilin were decreased (all P ＜ 0.05),and the mRNA expression levels of PAK1 and LIMK1 were decreased(both P＜0.05),but the protein and mRNA expression of cofilin had no significant changes after 24 h of treatment of SH-SY5Y cells with 10 μmol/L Aβ plus 2％ oxygen.Conclusions Aβ may reduce P-LIMK1 expression by inhibiting the activity of PAK1,thereby reducing the P-cofilin,increasing the formation of dephosphorylated cofilin,leading to neural cells damage,and hypoxia aggravates the neurotoxicity of Aβ through this pathway.
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Objective To study the effects of hypoxia on the expression of inflammatory factor high mobility group box-l(HMGB1) in the pulmonary arteriolae of neonatal SD rats.Method A total of 80 neonatal SD rats were randomly assigned into control group and hypoxia-induced persistent pulmonary hypertension of the newborn model (PPHN) group.The PPHN group was subdivided into 2 h,8 h,24 h,and 3 d post-PPHN subgroups according to the time of sacrifice.PPHN model was established on postnatal day 4 when rat pups in PPHN group were kept in low-oxygen box (10％ O2 and 90％ N2) for consecutively 7 days.Multi-channel physiological transducer RM-6280 was used recording the mean pulmonary artery pressure (mPAP) at the root to pulmonary artery of rat pups.ELISA method was used examining the serum level of HMGB1 of rat pups in each group.The pathology of the lung tissue was studied using optical microscope after HE staining,and MIAS-2000 medical image analysis software was used to calculate the ratio of the middle membrane thickness to the outer diameter of the pulmonary arteriolae wall (MT％).Protein level of HMGB1 in the lung was examined using Western Blot.Result The lung pathology in PPHN rats showed thickening of the middle membrane of the pulmonary arteriolae wall and stenosis of the pulmonary arteriolae.MT％ of control group and PPHN group were 5.3％ (3.7％,7.6％) and 7.1％ (4.6％,9.2％),respectively,without significant differences (P＞0.05).At 2 h,8 h,24 h,3 d post-PPHN timepoints,the serum levels of HMGB1 in PPHN group were (13.2±3.1),(15.4±3.6),(17.1±3.5),and (15.8±3.6) ng/ml,respectively,without intra-subgroup differences (F=2.134,P＞0.05),but significant differences existed when compared with control group at each timepoint (P＜0.01).Western Blot showed that HMGB1 protein expression in the lungs were significantly elevated soon after PPHN,peaked at 8～24 h,and reduced but still significantly elevated at 3 d after PPHN comparing with normal control.Significant differences existed at 2 h,8 h,and 24 h timepoints (P＜0.01,respectively).The HMGB1 protein of PPHN group declined significantly at 3 d timepoint without significant differences comparing with the control group (P＞0.05).Conclusion HMGB1 is closely related with the pathogenesis of PPHN,indicating the inflammatory response plays an important role in the mechanisms of PPHN.HMGB1 may be an indicator for the assessment of hypoxia-induced PPHN.
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Objective To explore the risk factors for postoperative hypoxemia in patients undergoing Stanford type A aortic dissection surgery.Methods The clinical data of 77 patients with Stanford type A aortic dissection surgery were analyzed retrospectively.Among the patients, 40 patients occurred hypoxemia(hypoxemia group),and 37 patients did not occur hypoxemia(non-hypoxemia group).The preoperative,intraoperative and postoperative clinical data were compared between 2 groups,and the independent risk factors for postoperative hypoxemia were analyzed by multiple Logistic regression analysis.Results The incidence of postoperative hypoxemia in patients with Stanford type A aortic dissection was 51.9% (40/77).The multiple Logistic regression analysis result showed that age (OR =1.088,95% CI 1.018-1.164,P=0.013),body mass index≥25 kg/m2(OR=6.495,95% CI 1.327-31.789,P=0.021),pericardial effusion(OR=6.384,95% CI 1.426-28.576,P=0.015),white blood cell count(OR=1.289,95% CI 1.033-1.609,P=0.024)and using recombinant human coagulationⅦa (OR = 23.757, 95% CI 2.849 - 198.085, P = 0.003) were the independent predictive factors for postoperative hypoxemia in patients with Stanford type A aortic dissection.Conclusions The postoperative hypoxemia in patients with Stanford type A aortic dissection is related with perioperative systemic inflammation, especially in obese patients who should be given anti-inflammatory treatment during perioperative period.Control of bleeding and reducing the recombinant human coagulationⅦa as far as possible can reduce the incidence of postoperative hypoxemia.
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It is an important clinical subject to illuminate the mechanisms of myocardial damage in the early stage post severe burn in prevention against and treatment of burn shock, which may offer a targeted " dynamic support" in the treatment of severe burn patients. In recent years, the role of autophagy in hypoxic myocardial injury has attracted much attention. Autophagy is a physiological phenomenon on intracellular digestion process of long-life proteins and the aging and damaged organelles through lysosomal system, and it is essential for maintaining the homeostasis of cells. Severe hypoxia/ischemia causes lysosome dysfunction, insufficient fusion between autophagosome and lysosome, accumulation of autophagosomes, and damaged autophagy flux, thus leading to cell dysfunction and cell death. To study the roles of autophagy and explore the potential signals in autophagy modulation will provide a new therapeutic target for alleviating cardiac dysfunction following severe burn.
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Objective To study the relationship between the expression of sonic hedgehog (Shh) and vascular endothelial growth factor (VEGF) in hypoxic human retinal pigment epithelial (hRPE) cells.Methods Cultured hRPE-19 cells (3rd-6th generations) were used in this experiment.hRPE-19 cells were divided into three groups including the control group,the hypoxia experimental group (100 μmol/L CoCl2) and the inhibition group (pretreatment with 20 μmol/L cyclopamine 1 hour before hypoxia).After culturing for 4,8,12 and 24 hours,the mRNA level of Shh and VEGF genes in these cells were measured by fluorescence quantitative polymerase chain reaction,and the protein level of Shh and VEGF in the supematants were measure by enzymelinked immunosorbent assay.The relationship between the expression of Shh and VEGF was analyzed by Pearson correlation analysis.Results The control group expressed low levels of Shh and VEGF mRNA/protein.The expression of Shh and VEGF mRNA/protein in the hypoxia experimental group was significantly higher than that in the control group (F=178.364,183.732,77.456,91.572;P＜0.01).The expression of Shh and VEGF mRNA in the inhibition group was significantly lower than that in the hypoxia experimental group (F=68.745,121.834;P＜0.01).In the hypoxia experimental group,the expression of VEGF protein was positively correlated with the expression of Shh protein (r=0.942,P＜ 0.05);and the expression of VEGF and Shh mRNA was positively correlated (r=0.970,P＜0.01).However,there was no significant correlation in the expression of VEGF and Shh mRNA in the inhibition group (r=0.915,P＞ 0.05).Conclusion There is a positive correlation between the expression of Shh and VEGF in hypoxic hRPE cells.
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Objective To evaluate effect of dexmedetomidine on mitochondrion-dependent apoptosis during hypoxia-reoxygenation (H/R) injury to hippocampal neurons of rats.Methods The primarily cultured hippocampal neurons of Sprague-Dawley rats were divided into 4 groups (n =40 each) using a random number table method:control group (C group),vehicle group (V group),H/R group and dexmedetomidine group (D group).Hippocampal neurons were subjected to oxygen-glucose deprivation followed by restoration of oxygen supply to establish the model of H/R injury.Dexmedetomidine 1 μmol/L was added at 6 h of reoxygenation in D group.The viability of neurons was measured by methyl thiazolyl tetrazolium assay at 20 h of reoxygenation.The ultrastructure of mitochondria was observed by transmission electron microscopy.The expression of cytochrome c (Cyt c),caspase-3,Fis1 and Drp1 was detected by Western blot.The neuronal apoptosis was detected by flow cytometry,and apoptosis rate was calculated.Results Compared with C group,no significant change was found in the viability of neurons in group V (P＞0.05),and the viability of neurons was significantly decreased,the apoptosis rate was increased,the expression of Cyt c,caspase-3,Fis1 and Drp1 was up-regulated (P＜0.05),and the damage to mitochondrial ultrastructure was accentuated in H/R and D groups.Compared with H/R group,the viability of neurons was significantly increased,the apoptosis rate was decreased,the expression of Cyt c,caspase-3,Fis1 and Drp1 was down-regulated (P＜0.05),and the damage to mitochondrial ultrastructure was significantly attenuated in D group.Conclusion The nechanism by which dexmedetomidine reduces the H/R injury to hippocampal neurons is related to inhibiting mitochondrion-dependent apoptosis in rats.
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Objective To evaluate the role of Toll-like receptor 4 (TLR4) in cardiomyocyte apoptosis induced by hypoxia-reoxygenation (H/R) in newborn rats.Methods Cardiomyocytes obtained from newborn Sprague-Dawley rats,aged 1-3 days,were primarily cultured in DMEM culture medium supplemented with 20％ fetal bovine serum and divided into 4 groups (n=10 each) using a random number table:control group (group C),H/R group,negative control group (con-siRNA group) and TLR4-siRNA group.Cardiomyocytes were routinely cultured for 24 h in group C.Cardiomyocytes were subjected to hypoxia for 2 h followed by 4 h of reoxygenation in group H/R.Cardiomyocytes were transfected with siRNA and TLR4-siRNA at the final concentration of 80 nmol/L in con-siRNA group and TLR4-siRNA group,respectively.After successful establishment of H/R injury model,the cell survival rate was measured by MTT assay,flow cytometry was used to detect apoptosis in cardiomyocytes,the expression of TLR4 mRNA was determined by real-time polymerase chain reaction,and the expression of TLR4,Bcl-2,Bax and caspase-3 was detected by Western blot.The apoptosis rate was calculated.Results Compared with group C,the cell survival rate was significantly decreased,the expression of TLR4 protein and mRNA,caspase-3 and Bax was up-regulated,the apoptosis rate was increased and the expression of Bcl-2 was down-regulated in the other three groups (P＜0.01).Compared with H/R and con-siRNA groups,the cell survival rate was significantly increased,the expression of TLR4 protein and mRNA,caspase-3 and Bax was down-regulated,the apoptosis rate was decreased and the expression of Bcl-2 was up-regulated in group TLR4-siRNA (P＜0.01).Conclusion TLR4 is involved in cardiomyocyte apoptosis induced by H/R in newborn rats.
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Objective To evaluate the effect of dexmedetomidine on the expression of NR1 subunit-containing NMDA receptors and GIuR2 subunit-containing AMPA receptors during hypoxic injury to rat hippocampal neurons.Methods The hippocampal neurons were isolated from Wistar rats within 24 h after birth and divided into 3 groups (n=24 each) using a random number table:control group (group C),hypoxia group (group H) and dexmedetomidine group (group D).The cells were subjected to hypoxia for 6 h to establish the model of neuronal hypoxic injury in H and D groups.In group D,0.1 μmol/L dexmedetomidine was added at 6 h of hypoxia and neurons were incubated for 3 h,and then the culture medium was replaced with a normal medium and neurons were incubated for 24 h.The neuronal viability was measured by CCK-8 assay,the leakage of LDH was detected,and the leakage rate was calculated.The expression of NR1 subunits-containing NMDA receptors and GluR2 subunits-containing AMPA receptors was detected by Western blot.The concentration of calcium ion in cytoplasm was measured using Fluo-3AM.Results Compared with group C,the neuronal viability was significantly decreased,the LDH leakage rate was increased,the expression of NR1 subunits-containing NMDA receptors in the membrane was up-regulated,the expression of GluR2 subunits-containing AMPA receptors was down-regulated,and the concentration of calcium ion in cytoplasm was increased in H and D groups (P＜0.05).Compared with group H,the neuronal viability was significantly increased,the LDH leakage rate was decreased,the expression of NR1 subunits-containing NMDA receptors in the membrane was down-regulated,the expression of GluR2 subunitscontaining AMPA receptors was up-regulated,and the concentration of calcium ion in cytoplasm was decreased in group D (P＜0.05).Conclusion The mechanism by which dexmedetomidine reduces hypoxic injury to rat hippocampal neurons may be related to inhibiting up-regulation of the expression of NR1 subunits-containing NMDA receptors in the membrane and down-regulation of the expression of GluR2 subunitscontaining AMPA receptors.
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Objective To study the effect of repetitive normobaric hypoxic preconditioning (RNHP) on white matter lesions (WMLs) and cognitive impairment in chronic cerebral ischemia rats.Methods Twenty-four healthy adult male SD rats were divided into sham operation group,model group,and RNHP group (8 in each group).The bilateral common carotid arteries in sham operation group were isolated but not ligated in ambient air,those in model group were ligated in ambient air,and those in RNHP group were preconditioned for 2 weeks before ligation.Their cognitive function was assessed in Morris water maze test,their WMLs were caluculated with KlüverBarrera staining.The astroglia,microglia and oligodendrocyte in cerebral white matter were stained with immunolabelling technique using antibodies to glial fibrillary acidic protein,Iba-1 and CNPase.Results The percentage of target quadrant swimming time was significantly higher in RNHP group and sham operation group than in model group (27.26％ ± 2.06％,29.06％ ± 1.72％ vs 20.58％±2.23％,P＜0.05,P＜0.01).The scores of WMLs in corpus callosum,caudate putamen and anterior commissure were significantly lower,the number of astrocytes and microglias was significantly smaller while that of oligodendrocytes was significantly greater in RNHP group and sham operation group than in model group (P＜0.05,P＜0.01).Conclusion RNHP can improve WMLs and cognitive impairment in chronic cerebral ischemia rats.