Effect and mechanism of miRNA-92a regulating SHH pathway on promoting vascular regeneration after myocardial I/R injury / 中华老年心脑血管病杂志

Objective To explore the effect and mechanism of miR-92a regulating sonic hedgehog(SHH)pathway on promoting vascular regeneration after myocardial ischemia-reperfusion(I/R)injury.Methods Primary cardiomyocytes were isolated from newborn SD rats(1 to 3 days old),and then cultured to establish a cellular model of hypoxia/reoxygenation injury.The cardiomyo-cytes were divided into cardiomyocyte normoxia group and cardiomyocyte I/R group.After miR-92a mimic and inhibitor were respectively transfected into primary cardiomyocytes to overex-press or lower its expression,the cells were then grouped into control,I/R,miR-92a mimic and in-hibitor groups.CCK-8 assay was used to determine cell viability,flow cytometry was employed to detect cell apoptosis,ELISA and QT-PCR were applied to detect the expression of VEGF,b-FGF and Ang-1,and Western blotting was performed to measure the expression of SHH signaling pathway related proteins.Results The expression level of miR-92a was significantly higher in the cardiomyocytes from the ischemia/reperfusion(I/R)group than the normoxia group(3.89±0.29 vs 1.53±0.19,P＜0.01).Statistical differences were observed among the control group,miR-92a inhibitor group,I/R group,and miR-92a mimic group in the protein levels of SHH(0.57±0.13 vs 0.51±0.11 vs 0.24±0.03 vs 0.14±0.02,P＜0.01),of Smoothened(SMO,0.53±0.12 vs 0.49± 0.10 vs 0.14±0.04 vs 0.09±0.01,P＜0.01),of glioma-associated oncogene homolog 1(Gli-1,0.56±0.14 vs 0.50±0.13 vs 0.15±0.03 vs 0.08±0.01,P＜0.01),and of glioma-associated onco-gene homolog 2(Gli-2,0.58±0.11 vs 0.49±0.12 vs 0.18±0.02 vs 0.11±0.03,P＜0.01).Conclu-sion MiR-92a is abnormally highly expressed in cardiomyocytes after I/R injury,and inhibition of miR-92a can activate SHH signaling pathway to promote the expression of angiogenesis factors effectively.

Role of autophagy in morphine preconditioning-induced reduction of OGD/R injury in primary cortical neurons of mice and the relationship with JNK / 中华麻醉学杂志

Objective:To evaluate the role of autophagy in morphine preconditioning-induced reduction of oxygen-glucose deprivation and restoration (OGD/R) injury in primary cortical neurons of mice and the relationship with c-Jun N-terminal kinase (JNK).Methods:Primary cortical neurons extracted from C57BL/6 neonatal mice within 24 h after birth were divided into 9 groups ( n=24 each) using a random number table method: control group (C group), OGD/R group, morphine preconditioning group (M group), autophagy inhibitor 3-methyladenine (3-MA) group (3-MA group), 3-MA+ morphine preconditioning group (3-MA+ M group), autophagy inhibitor chloroquine group (Ch group), chloroquine + morphine preconditioning group (Ch+ M group), JNK inhibitor SP600125 group (SP group) and SP600125 + morphine preconditioning group (SP+ M group). Morphine preconditioning: morphine was added at a final concentration of 3 μmol/L before OGD/R, and the cells were incubated for 2 h in OGD/R group. In 3-MA, Ch and SP groups, 3-MA 5 mmol/L, chloroquine 50 μmol/L and SP600125 25 μmol/L were added, respectively, and the cells were incubated for 150 min. In 3-MA+ M, Ch+ M and SP+ M groups, 3-MA 5 mmol/L, chloroquine 50 μmol/L and SP600125 25 μmol/L were added, respectively, at 30 min before morphine preconditioning. Then the cells were subjected to oxygen-glucose deprivation for 1 h followed by restoration of oxygen-glucose supply for 24 h. CCK-8 assay was used to detect the neuronal viability. The expression of JNK, phosphorylated JNK (p-JNK), microtubule-associated protein 1 light chain 3 (LC3), p62, Beclin1, caspase-3, and cleaved-caspase-3 was determined by Western blot. The autophagosomes and autolysosomes were counted using LC3-double fluorescent adenovirus transfection, and the neuronal apoptosis rate was determined by TUNEL staining. Results:Compared with C group, the neuronal viability was significantly decreased, the expression of Beclin1 was up-regulated, the expression of p62 was down-regulated, and the LC3Ⅱ/LC3Ⅰ ratio, p-JNK/JNK ratio, the number of autophagosomes and autolysosomes, cleaved-caspase-3/caspase-3 ratio and neuronal apoptosis rate were increased in OGD/R group ( P<0.001). Compared with OGD/R group, the neuronal viability, p-JNK/JNK ratio, LC3Ⅱ/LC3Ⅰ ratio and the number of autophagosomes and autolysosomes were significantly increased, the expression of Beclin1 was up-regulated, and the expression of p62 was down-regulated in M group, the LC3Ⅱ/LC3Ⅰ ratio was significantly decreased, and the expression of p62 was down-regulated in 3-MA group, the LC3Ⅱ/LC3Ⅰ ratio was significantly increased, and the expression of p62 was up-regulated in Ch group ( P<0.001), and no significant change was found in the parameters mentioned above in SP600125 group ( P>0.05). Compared with M group, the neuronal viability was significantly decreased, the LC3Ⅱ/LC3Ⅰ ratio was decreased, and the expression of p62 was up-regulated in M+ 3-MA group, the neuronal viability was significantly decreased, the LC3Ⅱ/LC3Ⅰ ratio was increased, and the expression of p62 was up-regulated in M+ Ch group, and the neuronal viability, LC3Ⅱ/LC3Ⅰ ratio and p-JNK/JNK ratio were significantly decreased, the expression of p62 was up-regulated, the number of autophagosomes and autolysosomes was decreased, the expression of Beclin1 was down-regulated, and the cleaved-caspase-3/caspase-3 ratio and neuronal apoptosis rate were increased in M+ SP group ( P<0.001). Conclusions:Morphine preconditioning can attenuate OGD/R injury by activating JNK, enhancing autophagy and inhibiting apoptosis in primary cortical neurons of mice.

Effect of TBK1 overexpression on hypoxia-reoxygenation injury in isolated mouse cardiomyocytes subjected to high glucose: relationship with mitochondrial autophagy / 中华麻醉学杂志

Objective:To evaluate the effect of TBK1 overexpression on hypoxia-reoxygenation (H/R) injury in isolated mouse cardiomyocytes subjected to high glucose and the relationship with mitochondrial autophagy.Methods:Normally cultured log-phase HL-1 mouse cardiomyocytes were inoculated in a 6-well plate at a density of 1×10 6 cells/ml and were divided into 4 groups ( n=10 each) using a random number table method: control group (group C), high glucose group (group HG), high glucose and H/R group (group HG+ H/R), and TBK1 overexpression group (group TBK1). The cells were incubated in culture medium with 1% fetal bovine serum and 1% double antibody for 24 h when the cell density reached 50%.When the cell density reached 80%, pcDNA3.1 (+ ) was used as a vector to achieve TBK1 overexpression.The cells were cultured with high glucose medium (33 mmol/L) for 24 h, exposed to 94% N 2+ 5% CO 2+ 1% O 2 for 24 h in an incubator at 37℃ followed by 12 h reoxygenation in an incubator containing 5% CO 2 at 37°C to establish the model of H/R injury to cardiomyocytes subjected to high glucose.After reoxygenation, CCK-8 assay was used to detect cell viability, the activity of lactic dehydrogenase (LDH) in supernatant was detected using LDH kit, mitochondrial contents were determined using Mito-Tracter green fluorescent probe, and the expression of TBK1 and mitophagy-related proteins PINK1, Parkin, LC3B and P62 was detected by Western blot. Results:Compared with group C, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, mitochondrial contents were decreased, the expression of TBK1, PINK1, Parkin and LC3B was down-regulated, and the expression of P62 was up-regulated in HG group and HG+ H/R group ( P<0.05). Compared with group HG, the cell viability was significantly decreased, the activity of LDH in supernatant was increased, mitochondrial contents were decreased, the expression of TBK1, PINK1, Parkin and LC3B was down-regulated, and the expression of P62 was up-regulated in group HG+ H/R ( P<0.05). Compared with group HG+ H/R, the the cell viability was significantly increased, the activity of LDH in supernatant was decreased, mitochondrial contents were increased, the expression of TBK1, PINK1, Parkin and LC3B was up-regulated, and the expression of P62 was down-regulated in group TBK1 ( P<0.05). Conclusion:The mechanism by which TBK1 overexpression reduces the H/R injury is related to restoring mitophagy in isolated mouse cardiomyocytes subjected to high glucose.

Relación entre el índice gradiente dióxido de carbono/gradiente arteriovenoso de oxígeno y lactato en el choque séptico por neumonía / Relationship between carbon dioxide gradient/arteriovenous oxygen gradient index and lactate in septic shock secondary to pneumonia

RESUMEN Objetivo Determinar la relación del índice gradiente de dióxido de carbono/gradiente arteriovenoso de oxígeno y lactato en pacientes con neumonía complicada y choque séptico en áreas críticas del Hospital III EsSalud Chimbote durante el periodo 2018 y 2019. Materiales y métodos Estudio de tipo cuantitativo, observacional, correlacional y longitudinal de tendencias. La unidad de análisis está constituida por los pacientes con neumonía complicada por choque séptico que presentan lecturas de índice de gradiente dióxido de carbono / gradiente arteriovenoso de oxígeno y lecturas de lactato. Resultados Se incluyeron 90 pacientes con neumonía complicada por choque séptico. El presente trabajo de investigación demuestra una fuerte relación entre el índice de gradiente de gases y el lactato sérico: al inicio, 3 horas, 6 horas y a las 12 horas, de iniciado y monitorizado estos pacientes. Conclusiones El índice del gradiente de dióxido de carbono/gradiente concentración arterio venoso de oxígeno > 1.4, se relaciona con el lactato sérico, en pacientes con shock séptico por neumonía.

ABSTRACT Objective To determine the relationship between carbon dioxide gradient/arteriovenous oxygen gradient index and lactate in patients with pneumonia complicated by septic shock in areas delivering care for critically ill or injured patients of the Hospital III EsSalud Chimbote between 2018 and 2019. Materials and methods A quantitative, observational, correlational, longitudinal trend study. The study population consisted of patients with pneumonia complicated by septic shock who had carbon dioxide gradient/arteriovenous oxygen gradient index and lactate readings. Results Ninety (90) patients with pneumonia complicated by septic shock participated in the research. The present study showed a robust relationship between gas gradient index and serum lactate when monitoring patients at baseline, 3 hours, 6 hours and 12 hours. Conclusions There is a relationship between carbon dioxide gradient/arteriovenous oxygen gradient index (> 1.4) and serum lactate levels in patients with septic shock secondary to pneumonia.

Oxygen impact on genome structure and chemistry / 药学学报

Oxygen is an essential element for life, which is mostly consumed at mitochondria for energy metabolism. For the genome inside nucleus, oxygen conducts structural regulations and chemical modifications through multiple pathways, where reactive oxygen species (ROS) serve as important messenger molecules. The highly activated ROS have the ability to produce different kinds of DNA lesions, while ferrous ions provide supports in many forms. Under the combinatorial action of oxygen and iron, almost all the genomic biochemical processes, such as replication, transcription and DNA damage repair are affected. Moreover, the variation of environmental oxygen concentration, particularly hypoxia that presents in many major diseases and critical physiological stages, provokes the responds at the genomic level. While the factors that lead to these genomic alterations are potential drug targets and deserve systematic investigations, herein, we collect the existing knowledge in the effects of ROS, ferrous ion and cell hypoxia on genome, along with brief discussions of the related drug molecules.

Preliminary exploration of 18 F-FMISO PET/CT SUVmax with relevant factors for nasopharyngeal carcinoma / 中华核医学与分子影像杂志

Objective To explore the relationship between the maximum standardized uptake value ( SUVmax ) of 18 F-fluoromisonidazole ( FMISO) PET/CT and the pathological classification, differentiation, T stage and primary tumor volume of nasopharyngeal carcinoma ( NPC) . Methods A retrospective analysis was performed on 41 patients with NPC (31 males, age 18-74 years;10 females, age 35-67 years) who underwent head and neck 18 F-FMISO PET/CT from 2012 to 2015. The relationship between the clinicopath-ological parameters (pathological classification, differentiation, T stage, tumor volume) of primary lesion and SUVmax were analyzed. Mann-Whitney u test, approximate t test and Spearman correlation were used for data analysis. Results There was no significant difference in SUVmax between non-keratinizing carcinoma and squamous cell carcinoma ( u=183.5, P>0.05) , nor between the differentiated carcinoma and undiffer-entiated carcinoma( t'=-1.23, P>0.05) . SUVmax of T1-T2 primary tumor was significantly lower than that of T3-T4 tumor (1.52±0.43 vs 2.05±0.85; t'=-2.60, P<0.05), and SUVmax was correlated with primary tumor volume ( rs=0.488, P<0.05) . Conclusions The hypoxic degree is related with T stage and primary tumor volume in NPC. The combination analysis of T stage and tumor size will contribute to the assessment of oxygen level and prognosis of primary NPC.

Role of mitochondrial calcium uniporter in mitophagy in SH-SY5Y cells subjected to oxygen-glucose deprivation and restoration / 中华麻醉学杂志

Objective To evaluate the role of mitochondrial calcium uniporter ( MCU) in mitoph-agy in SH-SY5Y cells subjected to oxygen-glucose deprivation and restoration (OGD∕R). Methods SH-SY5Y cells were cultured in vitro, seeded in 96-well plates at a density of 2×105cells∕ml, and randomly di-vided into 4 groups (n=6 each) using a random number table method: control group (group C), group OGD∕R, OGD∕R plus MCU inhibitor group ( group OGD∕R + Ru360) and MCU inhibitor group ( group Ru360) . Cells were cultured in normal culture medium in group C. Cells were subjected to O2-glucose dep-rivation for 6 h followed by restoration of O2-glucose supply for 24 h in group OGD∕R. In group OGD∕R+Ru360, Ru360 at a final concentration of 10 μmol∕L was added at 30 min before O2-glucose deprivation, and the other treatments were similar to those previously described in group OGD∕R. Ru360 was added at a final concentration of 10 μmol∕L, and 30 min later cells were cultured under normoxic conditions in group Ru360. At 24 h of restoration of O2-glucose supply, cell counting kit-8 assay was used to detect the cell survival rate, JC-1 assay was used to detect mitochondrial membrane potential ( MMP ) , the ultrastructure of cells was observed with a transmission electron microscope, and the expression of p62, Tom20 and Bec-lin-1 was detected by Western blot. Results Compared with group C, no significant change was found in each parameter in group Ru360 ( P>0. 05) , the cell survival rate and MMP were significantly decreased, the expression of Tom20 and p62 was down-regulated, Beclin-1 expression was up-regulated (P<0. 01), the mitochondria swelled, and mitochondrial autophagosomes were increased in group OGD∕R. Compared with group OGD∕R, the cell survival rate and MMP were significantly increased, the expression of Tom20 and p62 was up-regulated, Beclin-1 expression was down-regulated (P<0. 01), the mitochondrial mor-phology kept well, and mitochondrial autophagosomes were decreased. Conclusion MCU is involved in the process of mitophagy in SH-SY5Y cells subjected to OGD∕R.

The role of hypoxia-related molecules in the pathogenesis of osteoarthritis / 中华创伤骨科杂志

Osteoarthritis is one of the most common chronic diseases in orthopedics.With increasing populations of aging and obese people,its incidence has risen year by year and become a major public health problem.The hallmark of osteoarthritis is cartilage destruction,the main cause of which is degradation of extracellular matrix by catabolic enzymes and death of chondrocytes caused by apoptosis or autophagy.Articular cartilage is a hypoxic environment because it lacks blood supply and the joint cavity is relatively closed.A hypoxic environment induces chondrocytes to produce a series of hypoxia-related molecules which can regulate the expression of catabolic enzymes,autophagy and apoptosis of chondrocytes for osteoarthritis.This paper aims to review recent reports on the relationship between hypoxic-related molecules and pathogenesis of osteoarthritis and discuss the role of hypoxia-related molecules in the pathogenesis of osteoarthritis.

Hypoxic condition monitoring and treatment evaluation for non-small cell lung cancer before and after radiotherapy by 18F-FMISO PET/CT / 中华核医学与分子影像杂志

Objective To evaluate the changes of hypoxic conditions in non-small cell lung cancer (NSCLC) patients before and after radiotherapy and assess the value of 18F-fluoromisonidzaole (FMISO)PET/CT for radiotherapy efficacy evaluation.Methods A total of 21 NSCLC patients (15 males,6 females,age 30-74 years) from January 2014 to October 2016 were prospectively enrolled.18F-FMISO PET/CT was performed before and after radiotherapy,and all patients underwent 18F-fluorodeoxyglucose (FDG)PET/CT before radiotherapy.Routine chest CT was performed at the 3rd and 6th month after radiotherapy.The maximum standardized uptake value (SUVmax) of tumor and muscle,tumor volume and hypoxic volume (HV) were measured.Tumor-to-muscle (T/M) value of 18F-FMISO was calculated,and T/M ≥ 1.3 was considered as the hypoxia cut-off value.Data were analyzed using Pearson correlation,paired t test,signed rank sum test and Wilcoxon rank sum test.Results Totally 81.0％(17/21) of NSCLC patients had hypoxia.There were significant positive correlations between 18F-FMISO T/M value and tumor volume or 18F-FDG SUVmax(r:0.72,0.60,both P＜0.05).The T/M value after radiotherapy was significantly lower than that before radiotherapy (1.42± 1.12 vs 2.08±0.71;t =3.62,P＜0.05),and median HV was also significantly lower than that before radiotherapy (6.53 vs 12.41 cm3;z =-3.83,P＜0.05).The median T/M values of effective group (n =14) and ineffective group (n =7) before radiotherapy were significantly different (2.14 vs 2.87;z=-2.27,P＜0.05),and the median HV of 2 groups before radiotherapy was also significantly different (6.43 vs 10.20 cm3;z=-2.14,P＜0.05).Conclusions Most NSCLC patients have hypoxia before radiotherapy.The larger tumor volume,the higher degree of hypoxia.Radiotherapy can alleviate the hypoxia of tumors.18F-FMISO PET/CT imaging before radiotherapy can be used to predict the efficacy of patients with NSCLC.

Protective effects of a Yiqi Huatan Quyu formulation on liver injury in chronic intermittent hypoxia rats / 中华老年医学杂志

Objective To investigate the protective effects of a Yiqi Huatan Quyu formulation on liver injury in chronic intermittent hypoxia rats.Methods Thirty healthy male Sprague-Dawley rats of SPF grade were randomly divided into 3 groups:Group A(normoxia and normal saline gavage),Group B(chronic intermittent hypoxia and normal saline gavage)and Group C(chronic intermittent hypoxia and Yiqi Huatan Quyu formulation gavage)(n =10 in each group).After 8 weeks of treatment,the liver tissue was extracted for protein and RNA.Western blot was used to test the protein levels of B-cellymphoma-2 associated X protein (BAX) and B-cellymphoma-2 (BCL-2),and Realtime PCR was used to test gene expression of BAX and BCL-2.Transferase-mediated dUTP nick end-labeling(TUNEL)was used to assay the apoptotic rate of liver cells.Results The expression of the BAX protein and BAX gene was higher and the expression of the BCL-2 protein and BCL-2 gene was lower in Group B than in Group A(t =13.490 and 16.480,P =0.005 and 0.002;t =5.142 and 9.776,P =0.036 and 0.001,respectively).The levels of the BAX protein and BAX gene were lower and the levels of the BCL-2 protein and BCL-2 gene were higher in Group C than in Group B(t =6.088 and 15.240,P =0.026 and 0.005;t =5.296 and 16.380,P =0.034 and 0.004,respectively).The apoptosis rate of hepatocytes was higher in Group B than in Group A(t =7.698,P =0.002),and was lower in Group C than in Group B(t=4.345,P =0.012).Conclusions The Yiqi Huatan Quyu formulation may alleviate the apoptosis of liver cells in chronic intermittent hypoxia rats by upregulating BCL-2 and down-regulating BAX levels,thus exerting protective effects on the liver.

Environment-responsive fluorescence probes in image-guided surgery / 中华核医学与分子影像杂志

Unlike conventional imaging technologies, fluorescent imaging benefits from its safety, high-spatial resolution and real-time capability, which make it a highly adoptable imaging method for tumor detection and image-guided surgery in clinics. There are two types of fluorescent probes, including always-on type and environment-responsive type, wherein environment-responsive probes are preferred due to higher target-to-background ratios, which can improve sensitivity and specificity. The environment-responsive probes include enzyme-reactive probes, pH-sensitive probes and hypoxia responsive probes. This review summarizes recent progress in environment-responsive probes, and discusses their potentials in tumor detection and image-guided surgery.

Compensatory role of wild-type p53-induced phosphatase in trophoblastic apoptosis in response to hypoxia / 中华围产医学杂志

Objective To investigate the mechanism of wild-type p53-induced phosphatase (Wip1) in regulating p53-dependent apoptosis of trophoblasts for further understanding the etiology of preeclampsia (PE). Methods Placenta tissues were collected from normal (n=15) and PE (n=13) gravidas who underwent caesarean section in the First Affiliated Hospital of Chongqing Medical University from June 2017 to December 2018. Chorionic villus and decidua tissues were collected from another 10 women who aborted in early pregnancy. Two in vitro trophoblastic hypoxia cultures were established by subjecting human chorionic trophoblast cells (HTR8/SVneo) to either hypoxia intervention in incubator (HII) or simulated ischemic buffer (SIB). Wip1 expressions at the transcriptional and protein levels were determined by real-time quantitative polymerase chain reaction and Western blotting, respectively. The localization of Wip1 in placental tissues and HTR8/SVneo cells was determined by immunohistochemistry and immunofluorescence. Cell apoptosis was assessed by flow cytometry after viral infection and hypoxia. And the changes of pathway-related molecules including p53, phospho-p53 (p-p53), mouse double minute 2 homolog (Mdm2) and cleaved caspase3 (cl-cas3) were measured by Western blotting. The impact of Wip1 on Mdm2-p53 interaction was examined by co-immunoprecipitation. NVP-CGM097, an Mdm2-p53 specific inhibitor, was administered in PE cell models to verify the regulation of Wip1 on trophoblastic apoptosis through Mdm2-p53 pathway. Independent student's t-test, Welch's t-test and one-way analysis of variance were used as statistical methods. Results (1) Wip1 expression, which was mainly in trophoblast cells, was significantly elevated in human PE placentas (mRNA: 1.711±0.141 vs 0.860±0.126, t=4.496; protein: 0.449±0.027 vs 0.192±0.019, t=7.902) and in both in vitro trophoblastic PE models (protein in HII: 1.376±0.086 vs 0.977±0.114, t=2.792; SIB: 1.243±0.057 vs 0.381±0.045, t=11.910) compared with the corresponding control groups (all P<0.05). (2) Compared with corresponding control groups, overexpression of Wip1 suppressed the hypoxia-induced upregulation of p53 (HII: 0.185±0.024 vs 0.572±0.072; SIB: 0.400±0.067 vs 0.803±0.064), cl-cas3 (HII: 0.243±0.034 vs 0.529±0.072; SIB:0.179±0.011 vs 0.368±0.025) and p-p53/p53 protein expression (HII: 1.326±0.129 vs 2.100±0.187; SIB:0.473±0.028 vs 0.925±0.036) and also reduced the apoptosis rate [HII: (8.925±1.092)% vs (17.610±1.980)%;SIB: (13.910±1.886)% vs (24.650±1.622)%], which in turn promoted Mdm2-p53 binding (all P<0.05). However, knockdown of Wip1 gene expression in HTR8/SVneo cells brought about opposite effects (all P<0.05). (3) Neither overexpression nor knockdown of Wip1 influenced p53 or cl-cas3 expression when Mdm2-p53 interaction was blocked by NVP-CGM097. Conclusions Mdm2-p53 interaction promoted by Wip1 upregulation could compensate for the trophoblastic p53 accumulation in response to hypoxia, while exogenous upregulation of Wip1 in trophoblasts may reverse hypoxia-induced apoptosis. Therefore, this might provide a new therapeutic target for PE.

Effect of miR-34a on Proliferation of Pulmonary Artery Smooth Muscle Cells in Rats Induced by Hypoxia / 中山大学学报(医学科学版)

@#To observe the effect of miR-34a on the proliferation of pulmonary artery smooth muscle cells in rats induced by hypoxia and explore its possible mechanism.【Methods】Rat pulmonary artery smooth muscle cells were primarily isolated from pulmonary arteriole and cultured. After 3% O2 treatment，the expression of miR- 34a and Notch1 mRNA in rat PASMC were detected by real time PCR. The cell proliferation was detected by EDU after over-expression and inhibition of miR-34a and silencing Notch1 by cell transfection under hypoxia，and the expression of PCNA was detected by real time PCR and western blot method.【Results】We successfully isolated and cultured rat PASMC. And after 3% O2 treatment，the expression of miR-34a in rat PASMC was significantly decreased after 48 h compared with 24 h（P < 0.05）. However，the expression of Notch1 mRNA increased significantly after 48 h compared with 24 h（P < 0.05）. In addition， over-expression of miR-34a and silencing Notch1 significantly inhibited hypoxia-induced cell proliferation ，while inhibition of miR-34a significantly promoted the PASMC proliferation（P < 0.05）.【Conclusion】miR-34a participates in the proliferation of PASMC induced by hypoxia，and it may be through up-regulation of Notch1 to induce cell proliferation.

Hypoxia promotes migration and invasion of pancreatic cancer cells via inducing calpain 2 expression / 肿瘤

Objective: To investigate the expressions of Calpain 1 and Calpain 2 proteins, as well as the effects of regulating Calpain 2 expression on the migration and invasion of pancreatic cancer PANC-1 cells in hypoxia microenvironment. Methods: The expressions of Calpain 1 and Calpain 2 mRNAs in pancreatic cancer tissues and their survival prognosis value were analyzed using GEPIA (Gene Expression Profiling Interactive Analysis) database online tool. The pancreatic cancer PANC-1 cells were cultured in normoxia (21% O2) and hypoxic (1% O2) environment, then the expressions of Calpain 1 and Calpain 2 mRNAs and proteins were detected by real-time fluorescent quantitative PCR and Western blotting, respectively. The siRNA targeting CAPN2gene(encoding Calpain 2) was transfected into pancreatic cancer PANC-1 cells, then the silencing effect of CAPN2 gene was detected by real-time fluorescent quantitative PCR and Western blotting. The PANC-1 cells with Calpain 2 low expression were cultured under the normoxia and hypoxic conditions, then the migration and invasion abilities of these cells were detected by wound healing test and Transwell chamber assay, the expressions of E-cadherin and Vimentin were detected by Western blotting, and the cell morphology was observed by inverted microscopy. Results: The expressions of Calpain 1 and Calpain 2 mRNAs in human pancreatic cancer tissues were higher than those in normal pancreatic tissues (both P < 0.01), and the high expression of Calpain 2 was associated with the poor prognosis of pancreatic cancer patients (P = 0.013). Compared with the normoxia, the expression levels of Calpain 2 mRNA and protein were increased in hypoxia microenvironment (both P< 0.01), but the expressions of Calpain 1 mRNA and protein were not changed. The Calpain 2 low-expressed PANC-1 cells were established successfully. Under normoxia condition, the down-regulation of Calpain 2 expression inhibited the migration and invasion of PANC-1 cells (both P < 0.05); hypoxia promoted the migration and invasion of PANC-1 cells (both P < 0.05); but the down-regulation of Calpain 2 expression could reverse the promotion effects of hypoxia on the migration and invasion of PANC-1 cells (both P < 0.05). Furthermore, the expression of E-cadherin increased, the expression of Vimentin decreased in PANC-1 cells after the down-regualtion of Calpain 2 expression under normoxia condition (both P < 0.05), while the size of irregular cells increased, and the part of cells had apoptotic change; but the expression of E-cadherin decreased, the expression of Vimentin increased in PANC-1 cells cultured under hypoxia condition (both P < 0.05), while the most of cells had mesenchymal-like changes, showing long shuttle type; but the down-regulation of Calpain 2 expression could reverse the effect of hypoxia on the epithelial-mesenchymal transition of PANC-1 cells (both P< 0.05). Conclusion: Stimulated by hypoxia microenvironment, the expression of Calpain 2 was increased, so as to promote the migration and invasion of PANC-1 cells via regulating E-cadherin and Vimentin expressions.

Compensatory role of wild-type p53-induced phosphatase in trophoblastic apoptosis in response to hypoxia / 中华围产医学杂志

Objective@#To investigate the mechanism of wild-type p53-induced phosphatase (Wip1) in regulating p53-dependent apoptosis of trophoblasts for further understanding the etiology of preeclampsia (PE).@*Methods@#Placenta tissues were collected from normal (n=15) and PE (n=13) gravidas who underwent caesarean section in the First Affiliated Hospital of Chongqing Medical University from June 2017 to December 2018. Chorionic villus and decidua tissues were collected from another 10 women who aborted in early pregnancy. Two in vitro trophoblastic hypoxia cultures were established by subjecting human chorionic trophoblast cells (HTR8/SVneo) to either hypoxia intervention in incubator (HII) or simulated ischemic buffer (SIB). Wip1 expressions at the transcriptional and protein levels were determined by real-time quantitative polymerase chain reaction and Western blotting, respectively. The localization of Wip1 in placental tissues and HTR8/SVneo cells was determined by immunohistochemistry and immunofluorescence. Cell apoptosis was assessed by flow cytometry after viral infection and hypoxia. And the changes of pathway-related molecules including p53, phospho-p53 (p-p53), mouse double minute 2 homolog (Mdm2) and cleaved caspase3 (cl-cas3) were measured by Western blotting. The impact of Wip1 on Mdm2-p53 interaction was examined by co-immunoprecipitation. NVP-CGM097, an Mdm2-p53 specific inhibitor, was administered in PE cell models to verify the regulation of Wip1 on trophoblastic apoptosis through Mdm2-p53 pathway. Independent student's t-test, Welch's t-test and one-way analysis of variance were used as statistical methods.@*Results@#(1) Wip1 expression, which was mainly in trophoblast cells, was significantly elevated in human PE placentas (mRNA: 1.711±0.141 vs 0.860±0.126, t=4.496; protein: 0.449±0.027 vs 0.192±0.019, t=7.902) and in both in vitro trophoblastic PE models (protein in HII: 1.376±0.086 vs 0.977±0.114, t=2.792; SIB: 1.243±0.057 vs 0.381±0.045, t=11.910) compared with the corresponding control groups (all P<0.05). (2) Compared with corresponding control groups, overexpression of Wip1 suppressed the hypoxia-induced upregulation of p53 (HII: 0.185±0.024 vs 0.572±0.072; SIB: 0.400±0.067 vs 0.803±0.064), cl-cas3 (HII: 0.243±0.034 vs 0.529±0.072; SIB: 0.179±0.011 vs 0.368±0.025) and p-p53/p53 protein expression (HII: 1.326±0.129 vs 2.100±0.187; SIB: 0.473±0.028 vs 0.925±0.036) and also reduced the apoptosis rate [HII: (8.925±1.092)% vs (17.610±1.980)%; SIB: (13.910±1.886)% vs (24.650±1.622)%], which in turn promoted Mdm2-p53 binding (all P<0.05). However, knockdown of Wip1 gene expression in HTR8/SVneo cells brought about opposite effects (all P<0.05). (3) Neither overexpression nor knockdown of Wip1 influenced p53 or cl-cas3 expression when Mdm2-p53 interaction was blocked by NVP-CGM097.@*Conclusions@#Mdm2-p53 interaction promoted by Wip1 upregulation could compensate for the trophoblastic p53 accumulation in response to hypoxia, while exogenous upregulation of Wip1 in trophoblasts may reverse hypoxia-induced apoptosis. Therefore, this might provide a new therapeutic target for PE.

Hypoxia-inducible factor HIF-1α modulates drugs resistance in colon cancer cells / Factor inducible por hipoxia HIF-1α modula la resistencia a drogas en células de cáncer de colon

Abstract Introduction: Drug resistance mechanisms may be associated with decreased cell death and its induction may depend on the response to oxidative stress caused by hypoxia. The correlation between hypoxia-inducible factor HIF-1α, the number of reactive oxygen species and their effect on cell survival has not yet been evaluated. Objective: The purpose of this study was to evaluate the effect of HIF-1α activity and reactive oxygen species (ROS) accumulation in apoptosis of colon cancer cells. Materials and methods: HT29 colon cancer cells were treated with Cobalt(II) chloride (CoCl2) or doxorubicin and the activity of HIF-1α was determined by ELISA assay. ROS were determined using fluorescence probe carboxy-H2DFFDA. Apoptosis was assessed by caspase-3 activation analysis, and PUMA and BAX mRNA levels by qRT-PCR. The reduction of the antiapoptotic effect due to hypoxia was attenuated by use of the endonuclease APE-1 (E3330) inhibitor. The endonuclease E3330 APE-1 inhibitor allowed evaluating the effect of ROS generated by doxorubicin and CoCl2 on apoptosis. Results: Chemical hypoxia in combination with doxorubicin is an oxidative stressor in HT29 cells and induces a reduction in the apoptotic process in a time-dependent manner. Conclusion: Resistance to hypoxia and doxorubicin-mediated cell death could be controlled by a mechanism related to the activity of HIF-1α and the amount of reactive oxygen species generated.

Resumen Introducción. Los mecanismos de resistencia a drogas podrían asociarse con disminución en la muerte celular y su inducción podría depender de la respuesta al estrés oxidativo que origina la hipoxia. La correlación entre factor inducible por hipoxia HIF-1α, cantidad de especies reactivas de oxígeno y su efecto sobre la supervivencia celular aún no ha sido evaluada. Objetivo. Evaluar el efecto de la inducción de la actividad de HIF-1α y la cantidad de especies reactivas de oxígeno sobre la apoptosis en células de cáncer de colon. Materiales y métodos. Células de cáncer de colon HT29 fueron tratadas con cloruro de cobalto (CoCl2) o doxorrubicina; la actividad de HIF-1α se evaluó por ELISA. Las especies reactivas de oxígeno fueron determinadas con sonda fluorescente carboxi-H2DFFDA. La apoptosis fue evaluada por la actividad de caspasa-3 y los niveles de mRNA de los genes proapoptóticos PUMA y BAX por qRT-PCR. El inhibidor de la endonucleasa APE-1 E3330 permitió evaluar el efecto de las especies reactivas de oxígeno generadas por doxorubicina y CoCl2 sobre la apoptosis. Resultados. La hipoxia química combinada + doxorubicina es estresor oxidativo en células HT29 e induce una reducción en el proceso apoptótico de manera tiempo dependiente. Conclusión. La resistencia a la muerte celular mediada por hipoxia y doxorubicina podría estar controlada por un mecanismo relacionado con la actividad de HIF-1α y la cantidad de especies reactivas de oxígeno generadas.

Cambios morfofisiológicos celulares durante la reanimación cardiopulmocerebral / Morph-physiological cellular changes during cardiac-pulmonary-cerebral resuscitation

Las células realizan transformaciones estructurales y metabólicas ante situaciones de estrés, lo que les permite mantener una adecuada homeostasis y evitar la muerte. La presente revisión bibliográfica tuvo como objetivo describir los principales cambios morfofisiológicos celulares que acontecen en la parada cardiaca y reanimación cardiopulmocerebral. El método incluyó una revisión documental (bases de datos SciELO Regional, PubMed, Cochrane e Infomed), realizada durante el primer semestre del 2018. Fueron seleccionadas 28 referencias. Se concluye que existen cambios celulares durante el cese circulatorio, las maniobras de resucitación y en la reperfusión. En la parada cardiaca, los cambios celulares se expresan en todos los organelos y puede llevar a muerte por necrosis. Durante la reperfusión se producen nuevos cambios estructurales, por entrada de calcio, alteraciones en sodio, producción de radicales libres e inflamación. Los cambios morfofisiológicos dependerán del estado metabólico previo, el tiempo de parada cardiaca y la instauración eficaz de medidas de resucitación.

Cell suffer structural and metabolic changes in stress situations,which allow them to maintain an adequate homeostasis and avoid death . This bibliographic review had the objective of describing the main morph-physiological changes which occur in cardiac failure and cardiac-pulmonary-cerebral resuscitation. The method was documentary reviewing (database Regional SciELO, PubMed, Cochrane and Infomed), developed during the first semester of 2018. Twenty eight references were selected. It was concluded that there are cellular changes during circulatory stop, the procedures of resuscitation and re-perfusion. In cardiac failure, cellular changes are expressed in all the organelles. And may cause death due to necroses. During re-perfusion new structural changes occur, for calcium entrance, sodium disturbances, production of free radicals and swelling. Morph.physiological changes depend on previous metabolic condition, time of cardiac failure and the successful establishment of resuscitation measures.

Recent advance in hyperbaric oxygen adjunctive therapy for gliomas / 中华神经医学杂志

Hyperbaric oxygen therapy (HBOT) has been widely used in the treatment ofhypoxia,ischemia,and a series of diseases caused by hypoxia and ischemia.The treatment effect of HBOT on gliomas remains controversial.The presence of local hypoxia in gliomas is the main cause of tumor resistance to radiotherapy and chemotherapy;HBOT can improve the oxygen content in these areas,and thus,treatment should be as effective as other hypoxic diseases;however,most researchers believe that in the process of gliomas,the efficacy of HBOT alone is limited,and HBOT is usually not used alone.More commonly,HBOT is commonly used as adjuvant therapy in other treatments (such as radiotherapy and chemotherapy).Based on the above contents,we summarized as follows and put forward the views on the current debate.

Neuroprotective effects of brimonidine on retinal ganglion cells induced by oxidative stress mitochondrial dysfunction / 中华实验眼科杂志

Objective To explore whether brimonidine has a protective effect on retinal ganglion cells (RGCs) through improving mitochondrial function under the oxidative stress.Methods Mouse RGC-5 cells were cultured in DMEM medium containing low concentration of glucose (1 g/L),10％ fetal bovine serum and 100 U/ml penicillin-streptomycin solution.The cells were divided into normal control group,H2O2-treated group and brimonidine+ H2O2 group.H2O2 at the concentration of 800 μmol/L was added into the medium in the H2O2-treated group,and 1 μmol/L brimonidine was added into the medium for 2 hours prior to the addition of H2O2 in the brimonidine+H2O2 group.The cells were sequently cultured for 24 hours.The morphology of the cell nucleus was examined by Hoechst fluorscence staining.The expressions of apoptosis-related protein in the cells were detected by Western blot assay.Mitochondrial membrane potential was assessed by JC-1 staining.Results The cell nuclei showed round or oval in shape with consistent size in the normal control group.The pycnosis and karyorrhexis of the cell nuclei were seen in the H2O2-treated group,and less abnormal nuclei were found in the brimonidine+H2O2 group.The relative expression level of bcl-2 protein in the cells was 0.76±0.15,0.50±0.13 and 0.75±0.17 in the normal control group,H2O2-treated group and brimonidine + H2O2 group,respectively,and the expression of bcl-2 protein in the H2O2-treated group was significantly lower than that in the normal control group and brimonidine+H2O2 group (both at P＜0.05).The relative expression level of bax protein in the cells was 0.65±0.13,0.83±0.07 and 0.70±0.10 in the normal control group,H2O2-treated group and brimonidine+H2O2 group,respectively,and the expression of bax protein in the H2O2-treated group was significantly higher than that in the normal control group and brimonidine+H2O2 group (both at P＜0.05).A strong orange fluorescence was seen in the mitochondrial membrane of RGC-5 in the normal control group with a coexpression with the green fluorescence of cell membrane.In the H2O2-treated group,the orange fluorescence intensity in the cells was evidently weakened,and the number of JC-1 responsed cells was considerably increased and the orange fluorescence intensity was enhanced in the brimonidine + H2O2 group.Conclusions Brimonidine can prevent RGCs from oxidative-stress damage by improving the mitochondrial function and therefore play a potential neuroprotective effect on optic nerve.

Research advance on the correlation between gastrointestinal microenvironment and Parkinson's disease in hypoxia situations / 中华老年医学杂志

Recently,patients with Parkinson's disease are suffering from gastrointestinal diseases before the diagnosis.Both clinical and neuropathological evidences have indicated that Parkinson's disease is often accompanied with gastrointestinal symptoms.The hypoxia-related plateau environment shows that gastrointestinal microenvironment is closely related with gastrointestinal disorders.The hypoxia,low temperature and strong radiation on intestinal flora are three important environmental factors.The hypoxia environment may be related to the changed gastrointestinal microenvironment.In this article,we summarized the latest progress in the correlation between gastrointestinal microenvironment and Parkinson's disease in hypoxia situations.