ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
@#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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
ABSTRACT
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.
Subject(s)
Cardiovascular Physiological Phenomena , Cell Physiological Phenomena/physiology , Cardiopulmonary Resuscitation/statistics & numerical data , Heart Arrest/physiopathology , Cell Hypoxia/physiologyABSTRACT
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.
ABSTRACT
Objective To evaluate the role of sirtuin 1 ( SIRT1)∕nuclear factor kappa B ( NF-κB) signaling pathway in oxygen-glucose deprivation and restoration ( OGD∕R) injury to hippocampal neurons of mice. Methods The HT22 hippocampal neurons were seeded in a culture plate ( 96-well plate, 100 μl∕well; 6-well plate, 2 ml∕well) at the density of 5×104 cells∕ml or in a culture dish (6 cm in diameter), and then divided into 4 groups ( n=24 each) using a random number table method: control group ( group C) , OGD∕R group ( group OGD) , SIRT1 inhibitor EX-527 preconditioning group ( group EX) and SIRT1 agonist SRT1720 preconditioning group ( group SRT) . Neurons were cultured in normal culture atmosphere at 37 ℃ in group C. In OGD, EX and SRT groups, the culture medium was replaced with oxygen-poor flu-id, and neurons were exposed to 5％ CO2-95％ N2 for 12 h in an incubator at 37℃, oxygen-poor fluid was replaced with the culture medium, and neurons were cultured for 24 h in normal culture atmosphere at 37℃. SIRT1 inhibitor EX-5271 μmol∕L and SIRT1 agonist SRT172010 μmol∕L were added at 12 h beforeOGD in EX and SRT groups, respectively. The cell viability was measured by CCK8 assay, the activity of LDH was detected by chemical colorimetry, cell apoptosis rate was determined by flow cytometry, the ex-pression of SIRT1, NF-κB, IκBα, Bcl-2 and Bax was detected by Western blot, and the Bcl-2∕Bax ratio was calculated. Results Compared with group C, the cell viability was significantly decreased, LDH ac-tivity and cell apoptosis rate were increased, the expression of SIRT1, IκBα and Bcl-2 was down-regula-ted, the expression of NF-κB and Bax was up-regulated, and the Bcl-2∕Bax ratio was decreased in OGD, EX and SRT groups ( P<0. 05) . Compared with group OGD, the cell viability was significantly increased, the LDH activity and cell apoptosis rate were decreased, the expression of SIRT1, IκBαand Bcl-2 was up-regulated, the expression of NF-κB and Bax was down-regulated, and the Bcl-2∕Bax ratio was increased in group SRT, and the cell viability was significantly decreased, LDH activity and cell apoptosis rate were in-creased, the expression of SIRT1, IκBαand Bcl-2 was down-regulated, the expression of NF-κB and Bax was up-regulated, and the Bcl-2∕Bax ratio was decreased in group EX (P<0. 05). Conclusion SIRT1∕NF-κB signaling pathway inhibition is involved in OGD∕R injury to hippocampal neurons of mice.
ABSTRACT
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.
ABSTRACT
@#[Abstract] Objective: To investigate the effects of FPR1 (formyl peptide receptor 1) antagonist Boc2 on migration, invasion, proliferation and tumorigenicity of human lung adenocarcinoma cells under hypoxia conditions. Methods: The protein expressions of hypoxia inducible factor 1α (HIF-1α) and FPR1 in human lung adenocarcinoma A549 cells induced by hypoxia was detected by Western blotting. FPR1 antagonist Boc2 was used to treat the hypoxia-induced A549 cells in vitro. The cells were divided into three groups: control group (cultured under normoxic condition), hypoxia group and hypoxia+Boc2 treatment group. Cell scratch test, transwell matrigel invasion assay and MTT method were used to detect the migration, invasion and proliferation of each group of cells, respectively. The A549 cells of each group were inoculated into nude mice to prepare xenograft model.After 4 weeks, the nude mice were sacrificed, and the differences in average tumor volume and mass, tumor formation rate, the expression of migration-related protein-E-cadherin (E-cad) and invasion-related protein-matrix metalloproteinase 9 (MMP-9) were analyzed. Results: Hypoxia induction can promote the expression of FPR1 protein in A549 cells in a time-dependent manner (P<0.05). The results of cell experiments showed that the ability of migration, invasion and proliferation of cells in hypoxia group were significantly higher than those in control group (P<0.01); while compared with hypoxia group, Boc2 treatment significantly inhibited the migration, invasion and proliferation of A549 cells (P<0.05). The results of nude mice experiments showed that the average volume and mass of nude mice in hypoxia group were significantly higher than those in the control group (all P<0.01). But the mean volume and mass of nude mice in hypoxia+Boc2 treatment group were significantly lower than those in the hypoxia group (all P<0.01). The rate of tumor formation in nude mice of hypoxia group was 100.0% (15/ 15), which was significantly higher than 60.0% (9/15) in the control group (χ2=7.500, P=0.006) and 73.3% (11/15) in the hypoxia + Boc2 treatment group (χ2=4.615, P=0.032). The expression of E-cad and MMP-9 protein in hypoxia group was significantly higher than that in control group (P<0.01), while Boc2 treatment significantly decreased the expression of E-cad and MMP-9 protein in hypoxia group (P<0.05). Conclusions: FPR1 antagonist Boc2 can significantly inhibit the migration, invasion, proliferation and tumorigenicity of hypoxia-induced human lung adenocarcinoma A549 cells, indicating that FPR1 plays an important role in the development and progression of human lung adenocarcinoma and may become a potential target of human lung adenocarcinoma treatment.
ABSTRACT
RESUMEN: Objetivo: Evaluar la influencia de la variación en la presión de oxígeno ambiental sobre la regeneración ósea guiada en cobayos. Material y método: Treinta y dos cobayos machos de 750±50g de peso fueron asignados en 2 grupos de 16 integrantes cada uno (grupo A: trabajado a 150msnm en la ciudad de Lima y a presión de oxígeno de 157mmHg; grupo B: trabajado a 3.230msnm en la ciudad de Jauja y a presión de oxígeno de 107mmHg). En ambos grupos se indujeron defectos óseos mandibulares de 5×6mm a través de un acceso quirúrgico extraoral; a 8 cobayos de cada grupo se les recubrió el defecto con una membrana de colágeno reabsorbible de origen porcino, mas al resto de animales no. Se evaluó el conteo celular de osteoblastos y osteocitos a los 15 y 30 días postoperatoriamente. Resultados: A los 15 y a los 30 días, en los grupos trabajados en altura y en los que se aplicó la membrana, la cantidad de osteoblastos fue 71±12,1 cél/camp y 83±7,2 cél/camp respectivamente, y la de osteocitos fue 34,5±6,6 cél/camp y 25±7,6 respectivamente; siendo estos grupos, en todas las comparaciones, los que tuvieron mayor cantidad de células óseas, aunque sin ser diferencias estadísticamente significativas (p>0,05). Conclusión: Se encontró tendencia a formar mayor cantidad de células óseas en las muestras tratadas con regeneración ósea y expuestas a la altitud comparados con el nivel del mar.
ABSTRACT: Objective: To evaluate the influence of the variation in ambient oxygen pressure on guided bone regeneration in guinea pigs. Material and method: A total of 32 male guinea pigs weighing 750±50g were assigned into two groups of 16 (group A: studied at 150 metres above sea level in the city of Lima and oxygen pressure of 157mmHg; group B: was at 3230 meters above sea level in the city of Jauja and an oxygen pressure of 107mmHg). Bone defects of 5×6mm were induced in the jaw in both groups through extra-oral surgical access. The defect in 8 guinea pigs of each group were covered with a porcine resorbable collagen membrane, but not in the other animals. The osteoblast and osteocyte cell counts were evaluated at 15 and 30 days post-operatively. Results: At 15 and 30 days, in the groups studied at height and with the membrane applied, the osteoblast count was 71±12.1 cells/field, and 83±7.2 cells/field, respectively, and an osteocyte count of 34.5±6.6 cells/field, and 25±7.6 cells/field, respectively. These groups had a higher number of bone cells in all the comparisons, but there were no statistically significant differences (P>.05). Conclusion: There was a tendency to form a greater amount of bone cells was found in the samples treated with bone regeneration and exposed to altitude compared to those at sea level.