Hypoxic Chamber Research Articles

Kaempferol ameliorates pulmonary vascular remodeling in chronic hypoxia-induced pulmonary hypertension rats via regulating Akt-GSK3β-cyclin axis

Excessive proliferation of pulmonary artery smooth muscle cells (PASMCs) is considered a major contributor to elevated pulmonary vascular resistance and a key mechanism of vascular remodeling in hypoxia-induced pulmonary hypertension (HPH). Kaempferol is a natural flavonoid compound and can be derived from numerous common medicinal herbs and vegetables, which exhibit antiproliferative and proapoptotic properties, however, the effects of kaempferol on vascular remodeling in HPH remain unexplored. In this study, SD rats were placed in a hypobaric hypoxia chamber for four weeks to establish a pulmonary hypertension model and given either kaempferol or sildenafil (an inhibitor of PDE-5) during days 1–28, after which the hemodynamic parameter and pulmonary vascular morphometry were assessed. Furthermore, primary rat PASMCs were exposed to hypoxic conditions to generate a cell proliferation model, then incubated with either kaempferol or LY294002 (an inhibitor of PI3K). Immunoblotting and real-time quantitative PCR assessed the protein and mRNA expression levels in HPH rat lungs and PASMCs. We found that kaempferol reduced pulmonary artery pressure and pulmonary vascular remodeling, and alleviated right ventricular hypertrophy in HPH rats. The mechanistic analysis demonstrated that kaempferol reduced the protein levels of phosphorylation of Akt and GSK3β, leading to decreased expression of pro-proliferation (CDK2, CDK4, Cyclin D1, and PCNA) and anti-apoptotic related proteins (Bcl-2) and increased expression of pro-apoptosis proteins (Bax and cleaved caspase 3). These results collectively demonstrate that kaempferol ameliorates HPH in rats by inhibiting PASMC proliferation and pro-apoptosis via modulation of the Akt/GSK3β/CyclinD axis.

Oxygen stress on age-stage, two-sex life tables and transcriptomic response of diamondback moth (Plutella xylostella).

Elucidating the genetic basis of local adaption is one of the important tasks in evolutionary biology. The Qinghai-Tibet Plateau has the highest biodiversity for an extreme environment worldwide, and provides an ideal natural laboratory to study adaptive evolution. The diamondback moth (DBM), Plutella xylostella, is one of the most devastating pests of the global Brassica industry. A highly heterozygous genome of this pest has facilitated its adaptation to a variety of complex environments, and so provides an ideal model to study fast adaptation. We conducted a pilot study combining RNA-seq with an age-stage, two-sex life table to study the effects of oxygen deprivation on DBM. The developmental periods of all instars were significantly shorter in the hypoxic environment. We compared the transcriptomes of DBM from Fuzhou, Fujian (low-altitude) and Lhasa, Tibet (high-altitude) under hypoxia treatment in a hypoxic chamber. Some DEGs are enriched in pathways associated with DNA replication, such as DNA repair, nucleotide excision repair, base excision repair, mismatch repair and homologous recombination. The pathways with significant changes were associated with metabolism process and cell development. Thus, we assumed that insects could adapt to different environments by regulating their metabolism. Our findings indicated that although adaptive mechanisms to hypoxia in different DBM strains could be similar, DBM individuals from Tibet had superior tolerance to hypoxia compared with those of Fuzhou. Local adaptation of the Tibetan colony was assumed to be responsible for this difference. Our research suggests novel mechanisms of insect responses to hypoxia stress.

Ferulic acid improves cognitive impairment by regulating jumonji C domain-containing protein 6 and synaptophysin in the hippocampus in neonatal and juvenile rats with intrauterine hypoxia during pregnancy.

To investigate the impacts of ferulic acid (FA) on jumonji C domain-containing protein 6 (JMJD6) and synaptophysin in the tissues of the hippocampus in neonatal and juvenile rats with intrauterine hypoxia-induced cognitive impairment. The Sprague-Dawley pregnant rats were randomly divided into three groups: control, hypoxia, and hypoxia + FA. On day 14 of pregnancy, the intrauterine hypoxia model was created by placing pregnant rats in the hypoxic and low-pressure experimental chamber for 2 hr a day for 3 days. In the hypoxia + FA group, pregnant rats were injected intraperitoneally with 4% FA, once a day for 7 days. The hypoxia group was treated with equal amounts of saline. After delivery, JMJD6 and synaptophysin mRNA and proteins in the hippocampus regions were detected by in situ hybridization and western blotting. The Morris water maze was used to evaluate cognitive function. The neonatal and juvenile rats in the hypoxia group had significantly increased expression of JMJD6 and decreased expression of synaptophysin protein and synaptophysin I mRNA in the hippocampus than those in the control group. Meanwhile, hypoxia also clearly prolonged the escape latency and shortened the stay time in the target quadrant. FA decreased the expression of JMJD6 and increased the expression of synaptophysin and improved cognitive function compared with those in the hypoxia group. FA probably ameliorated the cognitive impairment by regulating JMJD6 and synaptophysin in the hippocampus of neonatal and juvenile rats who had intrauterine hypoxia during pregnancy.

Hypoxia-Induced Kidney Injury in Newborn Rats

Exposure to hypoxia during the early postnatal period can have adverse effects on vital organs. Neonatal Sprague-Dawley rats housed in a hypoxic chamber were compared to those in a normoxic chamber from postnatal days 0 to 7. Arterial blood was collected to evaluate renal function and hypoxia. Kidney morphology and fibrosis were evaluated using staining methods and immunoblotting. In the kidneys of the hypoxic group, protein expressions of hypoxia-inducible factor-1 were higher than those in the normoxic group. Hypoxic rats had higher levels of hematocrit, serum creatinine, and lactate than normoxic rats. Body weight was reduced, and protein loss of kidney tissue was observed in hypoxic rats compared to normoxic rats. Histologically, hypoxic rats showed glomerular atrophy and tubular injury. Renal fibrosis with collagen fiber deposition was observed in the hypoxic group. The expression of nicotinamide adenine dinucleotide phosphate oxidases was enhanced in the kidneys of hypoxic rats. Proteins involved in apoptosis were upregulated in the kidneys of hypoxic rats. An increase in the expression of pro-inflammatory cytokines was also observed in the kidneys of hypoxic rats. Hypoxic kidney injury in neonatal rats was associated with oxidative stress, inflammation, apoptosis, and fibrosis.

Loss of Blood-Brain Barrier Integrity in an In Vitro Model Subjected to Intermittent Hypoxia: Is Reversion Possible with a HIF-1α Pathway Inhibitor?

Several sleep-related breathing disorders provoke repeated hypoxia stresses, which potentially lead to neurological diseases, such as cognitive impairment. Nevertheless, consequences of repeated intermittent hypoxia on the blood-brain barrier (BBB) are less recognized. This study compared two methods of intermittent hypoxia induction on the cerebral endothelium of the BBB: one using hydralazine and the other using a hypoxia chamber. These cycles were performed on an endothelial cell and astrocyte coculture model. Na-Fl permeability, tight junction protein, and ABC transporters (P-gp and MRP-1) content were evaluated with or without HIF-1 inhibitors YC-1. Our results demonstrated that hydralazine as well as intermittent physical hypoxia progressively altered BBB integrity, as shown by an increase in Na-Fl permeability. This alteration was accompanied by a decrease in concentration of tight junction proteins ZO-1 and claudin-5. In turn, microvascular endothelial cells up-regulated the expression of P-gp and MRP-1. An alteration was also found under hydralazine after the third cycle. On the other hand, the third intermittent hypoxia exposure showed a preservation of BBB characteristics. Furthermore, inhibition of HIF-1α with YC-1 prevented BBB dysfunction after hydralazine treatment. In the case of physical intermittent hypoxia, we observed an incomplete reversion suggesting that other biological mechanisms may be involved in BBB dysfunction. In conclusion, intermittent hypoxia led to an alteration of the BBB model with an adaptation observed after the third cycle.

Hypobaric hypoxia induced renal injury in rats: Prophylactic amelioration by quercetin supplementation.

The present study aims at assessing the effect of hypobaric hypoxia induced renal damage and associated renal functions in male SD rats. Further, this study was extended to explore the protective efficacy of quercetin in ameliorating the functional impairment in kidneys of rats under hypobaric hypoxia. Rats were exposed to 7620m (25000 ft.) at 25°C ±2 in a simulated hypobaric hypoxia chamber for different time durations (0h,1h, 3h, 6h, 12h, 24h and 48h) in order to optimize the time at which maximum renal damage would occur. The rats were exposed to hypoxia for 12h duration was considered as the optimum time, due to significant increase in oxidative stress (ROS, MDA) and renal metabolites (creatinine, BUN and uric acid) with remarkable reduction (p<0.001) in antioxidants (GSH) in plasma, as compared to other tested durations. Moreover, these findings were in support with the histopathology analysis of renal tissues. For optimum quercetin dose selection, the rats were administered with different doses of quercetin (25mg, 50mg, 100mg and 200mg/Kg BW) for 12h at 7620 m, 25°C ±2, 1h prior to hypoxia exposure. Quercetin 50mg/kg BW was considered as the optimum dose at which significant (p<0.001) reduction in oxidative stress levels followed by reduction in creatinine and BUN levels were obtained in plasma of the rats compared to hypoxia control rats. Quercetin prophylaxis (50mg/kg BW) stabilized the HIF-1α protein expression followed by reduced VEGF protein expression along with reduced levels of LDH (p<0.001) in the kidneys of rats compared to hypoxia control. Histopathological observations further substantiated these findings in reducing the renal tissue injury. The study findings revealed that, quercetin prophylaxis abrogates the possibility of hypobaric hypoxia induced renal injury by reducing the oxidative stress in rats.

TRAP-induced PAR1 expression with its mechanism during AMI in a rat model

Background and objectiveProtease-activated receptor 1 (PAR1) is crucial in individuals with acute myocardial infarction (AMI). The continuous and prompt PAR1 activation mainly dependent on PAR1 trafficking is essential for the role of PAR1 during AMI in which cardiomyocytes are in hypoxia. However, the PAR1 trafficking in cardiomyocytes specially during the hypoxia is still unclear.Methods and resultA rat AMI model was created. PAR1 activation with thrombin-receptor activated peptide (TRAP) had a transient effect on cardiac function in normal rats but persistent improvement in rats with AMI. Cardiomyocytes from neonatal rats were cultured in a normal CO2 incubator and a hypoxic modular incubator chamber. The cells were then subjected to western blot for the total protein expression and staining with fluorescent reagent and antibody for PAR1 localization. No change in total PAR1 expression following TRAP stimulation was observed; however, it led to increased PAR1 expression in the early endosomes in normoxic cells and decreased expression in the early endosomes in hypoxic cells. Under hypoxic conditions, TRAP restored the PAR1 expression on both cell and endosomal surfaces within an hour by decreasing Rab11A (8.5-fold; 179.93 ± 9.82% of the normoxic control group, n = 5) and increasing Rab11B (15.5-fold) expression after 4 h of hypoxia. Similarly, Rab11A knockdown upregulated PAR1 expression under normoxia, and Rab11B knockdown downregulated PAR1 expression under both normoxic and hypoxic conditions. Cardiomyocytes knocked out of both Rab11A, and Rad11B lost the TRAP-induced PAR1 expression but still exhibited the early endosomal TRAP-induced PAR1 expression under hypoxia.ConclusionsTRAP-mediated activation of PAR1 in cardiomyocytes did not alter the total PAR1 expression under normoxic conditions. Instead, it triggers a redistribution of PAR1 levels under normoxic and hypoxic conditions. TRAP reverses the hypoxia-inhibited PAR1 expression in cardiomyocytes by downregulating Rab11A expression and upregulating Rab11B expression.

Systemic Hypoxemia Induces Cardiomyocyte Hypertrophy and Right Ventricular Specific Induction of Proliferation.

A recent study suggests that systemic hypoxemia in adult male mice can induce cardiac myocytes to proliferate. The goal of the present experiments was to confirm these results, provide new insights on the mechanisms that induce adult cardiomyocyte cell cycle reentry, and to determine if hypoxemia also induces cardiomyocyte proliferation in female mice. EdU-containing mini pumps were implanted in 3-month-old, male and female C57BL/6 mice. Mice were placed in a hypoxia chamber, and the oxygen was lowered by 1% every day for 14 days to reach 7% oxygen. The animals remained in 7% oxygen for 2 weeks before terminal studies. Myocyte proliferation was also studied with a mosaic analysis with double markers mouse model. Hypoxia induced cardiac hypertrophy in both left ventricular (LV) and right ventricular (RV) myocytes, with LV myocytes lengthening and RV myocytes widening and lengthening. Hypoxia induced an increase (0.01±0.01% in normoxia to 0.11±0.09% in hypoxia) in the number of EdU+ RV cardiomyocytes, with no effect on LV myocytes in male C57BL/6 mice. Similar results were observed in female mice. Furthermore, in mosaic analysis with double markers mice, hypoxia induced a significant increase in RV myocyte proliferation (0.03±0.03% in normoxia to 0.32±0.15% in hypoxia of RFP+ myocytes), with no significant change in LV myocyte proliferation. RNA sequencing showed upregulation of mitotic cell cycle genes and a downregulation of Cullin genes, which promote the G1 to S phase transition in hypoxic mice. There was significant proliferation of nonmyocytes and mild cardiac fibrosis in hypoxic mice that did not disrupt cardiac function. Male and female mice exhibited similar gene expression following hypoxia. Systemic hypoxia induces a global hypertrophic stress response that was associated with increased RV proliferation, and while LV myocytes did not show increased proliferation, our results minimally confirm previous reports that hypoxia can induce cardiomyocyte cell cycle activity in vivo.

Hypoxia-induced YAP activation and focal adhesion turnover to promote cell migration in mesenchymal TNBC cells.

Hypoxia is commonly characterized by malignant tumors that promote the aggressiveness and metastatic potential of cancer. Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer, with approximately 46% capacity related to distant metastasis. Transcriptional factor yes-associated protein (YAP), a core component of the Hippo pathway, is associated with poor prognosis and outcome in cancer metastasis. Here, we explored the effect of hypoxia-mediated YAP activation and focal adhesions (FAs) turnover in mesenchymal TNBC cell migration. We characterized the effect of hypoxia on YAP in different breast cancer cell lines using a hypoxia chamber and CoCl2 . Hypoxia-induced YAP nuclear translocation is significantly observed in normal breast epithelial cells, non-TNBC cells, mesenchymal TNBC cells, but not in basal-like TNBC cells. Functionally, we demonstrated that YAP activation was required for hypoxia to promote mesenchymal TNBC cell migration. Furthermore, hypoxia induced the localization of FAs at the leading edge of mesenchymal TNBC cells. In contrast, verteporfin (VP), a YAP inhibitor, significantly reduced the migration and the recruitment of nascent FAs at the cell periphery under hypoxia conditions, which only showed in mesenchymal TNBC cells. Our data support the hypothesis that YAP is novel factor and positively responsible for hypoxia-promoting mesenchymal TNBC cell migration. Our findings provide further evidence and outcomes to help prevent the progression of TNBC.

Nicotinamide mononucleotide attenuates HIF-1α activation and fibrosis in hypoxic adipose tissue via NAD+/SIRT1 axis.

Fibrosis is increasingly considered as a major contributor in adipose tissue dysfunction. Hypoxic activation of hypoxia-inducible factor 1α (HIF-1α) induces a profibrotic transcription, leading to adipose fibrosis. Nicotinamide mononucleotide (NMN), a member of the vitamin B3 family, has been shown to relieve hepatic and cardiac fibrosis, but its effects on hypoxic adipose fibrosis and the underlying mechanism remain unclear. We aimed to elucidate the roles of NMN in regulating HIF-1α and fibrosis in hypoxic adipose tissue. Mice were placed in a hypobaric chamber for four weeks to induce adipose fibrosis. NMN (500 mg/kg, every three days) was administered by intraperitoneal injection. In vitro, Stromal vascular fractions (SVF) cells were treated by hypoxia with or without NMN (200μM), sirtinol (25μM, a SIRT1 inhibitor) and CoCl2 (100μM, a HIF1α enhancer). The effects of NMN on hypoxia-associated adipose fibrosis, inflammation, NAD+/SIRT1 axis alteration, and HIF-1α activation were evaluated by real-time polymerase chain reaction (PCR), western blots, immunohistochemistry staining, immunoprecipitation, and assay kits. Mice placed in a hypoxic chamber for four weeks showed obvious adipose fibrosis and inflammation, which were attenuated by NMN. NMN also restore the compromised NAD+/SIRT1 axis and inhibited the activation of HIF-1α induced by hypoxia. In hypoxia-induced SVFs, the SIRT1 inhibitor sirtinol blocked the anti-fibrotic and anti-inflammatory effects of NMN, upregulated the HIF-1α and its acetylation level. The HIF1α stabilizer CoCl2 showed similar effects as sirtinol. NMN effectively attenuated HIF-1α activation-induced adipose fibrosis and inflammation by restoring the compromised NAD+/SIRT1 axis.

Apolipoprotein A1 suppresses the hypoxia-induced angiogenesis of human retinal endothelial cells by targeting PlGF.

To investigate the anti-angiogenic effect of apolipoprotein A1 (apoA1) on primary human retinal vascular endothelial cells (HRECs) and explore the possible mechanism. The primary HRECs were transfected with apoA1-GFP recombinant lentiviral and were compared with cells undergoing transfection with empty lentiviral vectors. Hypoxia chambers were used to simulate the anoxic environment of cells under pathological condition. The concentrations of secreted vascular endothelial growth factor (VEGF) and placental growth factor (PlGF) were measured by enzyme-linked immunosorbent assay (ELISA). Cell migration ability was detected by wound healing assay. The sprouting of HRECs was determined by tube formation assay. The protein levels of extracellular signal regulated kinase 1/2 (ERK1/2) and phosphorylated ERK1/2 (p-ERK1/2) were measured by Western blot. Overexpressed apoA1 in hypoxia-induced HRECs significantly suppressed PlGF (0.67±0.10 folds, P=0.007). Overexpressed apoA1 also attenuated hypoxia-induced cell migration (0.32±0.11 folds, P<0.0001), tube formation (0.66±0.01 folds, P<0.0001) and the phosphorylation levels of ERK (0.6±0.11 folds, P=0.025). Pretreatment of mitogen-activated protein kinase kinase (MEK) inhibitor (U0126) further reduced the PlGF and angiogenesis in hypoxia-induced HRECs. ApoA1 inhibits the angiogenesis at least in part by inactivating ERK1/2 in hypoxia-induced HRECs. Moreover, apoA1 suppresses the PlGF expression, which selectively associated with pathological angiogenesis.

Intermittent hypoxia in sport nutrition, performance, health status and body composition.

Intermittent hypoxia refers to the discontinuous use of low oxygen levels in normobaric environment. These conditions can be reproduced in hypoxic tents or chambers while the individual is training in different physical activity protocols. Intermittent hypoxia can affect several body systems, impacting nutrition, physical performance, health status and body composition. Therefore, it is necessary to assess protocols, regarding time and frequency of exposure, passive exposure or training in hypoxia, and the simulated altitude. At the molecular level, the hypoxia-inducible factor-1α is the primary factor mediating induction of target genes, including vascular endothelial growth factor and erythropoietin. The goal of these molecular changes is to preserve oxygen supply for cardiac and neuronal function. In addition, hypoxia produces a sympathetic adrenal activation that can increase the resting metabolic rate. Altogether, these changes are instrumental in protocols designed to improve physical performance as well as functional parameters for certain pathological disorders. In addition, nutrition must adapt to the increased energy expenditure. In this last context, performing physical activity in intermittent hypoxia improves insulin sensitivity by increasing the presence of the glucose transporter GLUT-4 in muscle membranes. These changes could also be relevant for obesity and type 2 diabetes treatment. Also, the anorectic effect of intermittent hypoxia modulates serotonin and circulating leptin levels, which may contribute to regulate food intake and favor body weight adaptation for optimal sport performance and health. All these actions suggest that intermittent hypoxia can be a very effective tool in sports training as well as in certain clinical protocols.

PS-B08-4: CHRONIC INTERMITTENT HYPOXIA INDUCES RENAL FIBROSIS BY MR ACTIVATION

Purpose: Obstructive sleep apnea is mainly characterized by chronic intermittent hypoxia (CIH), which is associated with chronic kidney disease. Macrophage is known to play an important role in renal fibrosis. However, little is known of the potential role of macrophage-to-myofibroblast transition (MMT) in hypoxic conditions. The aim of this study was to understand how infiltrating macrophages may contribute to renal fibrosis in hypoxic conditions. Methods: In this study, 30 adult male sprague dawley (SD) rats were randomly divided into normoxic control(NC), chronic intermittent hypoxia(CIH), and eplerenone(EPL) groups-each containing ten rats. The CIH and EPL groups were exposed to hypoxia in a commercial hypoxic chamber for 5 weeks. Rats in the EPL group were administered EPL orally via diet for 5 weeks. All rats were then sacrificed and serum, 24-hour urine and tissue specimens were collected for renal function, histopathology, western blot, immunohistochemistry. The RAW264.7 macrophages were subjected to 24 hours of hypoxia and 12 hours of reoxygenation. Cells were collected for immunofluorescence staining. Results: In addition to blood pressure, renal aldosterone level was also increased in CIH rats. Massons trichrome and Sirius red staining showed significantly higher collagen deposition and fibrosis in CIH rats. A markedly increased co-expression of α-SMA and CD68 in vitro and in vivo after CIH exposure. MR nuclear translocation was seen in hypoxic macrophages. Western blot showed that the expression of SGK-1 and p-SGK-1 was upregulated. These effects were reversed by MR blocker (EPL). Conclusions: CIH-induced macrophage infiltration, an important source of myofibroblasts, might participate in the development of renal interstitial fibrosis. These effects were reversed by EPL. Therefore, we speculate MR activation may be related to CIH-induced renal fibrosis.

CHEMORESISTANCE RELATED TO HYPOXIA ADAPTATION INMESOTHELIOMA CELLS FROM TUMOR SPHEROIDS.

Hypoxia has been noted as a key factor for induction and maintenance of cancer stemness thereby leading to therapy resistance. Three-dimensional (3D) spheroid models demonstrate a heterogeneity of hypoxic regions replicating the in vivo situation within tumors. Utilizing an established 3D spheroid model, we investigated whether extrinsic hypoxia reinforced chemoresistance in malignant pleural mesothelioma (MPM) spheroids. Tumor spheres were generated from Meso-1 (a typical human MPM cell line) cells having high spheroid-forming ability. To induce hypoxia condition, we utilized a hypoxia chamber with regulation of O2and CO2levels. Cell viability was estimated by a WST-8assay. Real-time polymerase chain reaction and Western blot were performed to evaluate the expression at mRNA and protein levels. Compared with cells cultured in the two-dimensional monolayer model, tumor sphere cells showed elevated mRNA levels of cancer stemness markers (CD26, CD44and ABCG2) and protein levels of the stemness and hypoxia adaptation markers (ABCG2, ALDH1A1and HIFs). Correlating with this, 3D spheroid cells were more resistant to permetrexed and topotecan than the two-dimensional cells, indicative of their potential for hypoxic adaptation. Furthermore, significantly stronger resistance to both chemotherapeutic agents was observed in spheroid cells upon hypoxic challenge compared to spheroid cells under normoxia. From the present data, it is concluded that hypoxia adaptation of MPM cells from tumor spheres could enhance their chemoresistance.

Trophoblast Migration with Different Oxygen Levels in a Gel-Patterned Microfluidic System.

In the placenta, substances such as nutrients, oxygen, and by-products are exchanged between the mother and the fetus, and the proper formation of the placenta determines the success of pregnancy, including the growth of the fetus. Preeclampsia is an obstetric disease in which the incomplete formation of the placenta occurs, which is known to occur when there is an abnormality in the invasion of trophoblast cells. The invasion of trophoblast cells is controlled by oxygen concentration, and HIF-1α changes according to oxygen concentration, showing a difference in cell mobility. MMP-2 and MMP-9 are observed to be high in the endometrium involved in trophoblast invasion, and the expression is regulated according to the oxygen concentration. In this experiment, cell culture was conducted using a gel-patterned system with a hypoxic chamber. Before the chip experiment, the difference in the expression of MMP-2 and MMP-9 according to the oxygen concentration was confirmed using a hypoxia chamber. After that, trophoblast cells (HTR8/SVneo) and endothelial cells (HUVECs) were separated and cultured through a physical barrier through a hydrogel on a microfluidic chip. Cells were cultured in a hypoxic chamber under controlled oxygen levels. It was confirmed that the mobility of trophoblast cells in culture on the chip was upregulated in a hypoxic environment through oxygen control. This suggests that the formation of a hypoxic environment in the endometrium where the invasion of trophoblast cells occurs plays a role in increasing cell mobility.

Development of cardioprotective effect of chronic continuous hypoxia in rats with induced metabolic syndrome

Background. It is known that adaptation to chronic continuous hypoxia leads to a pronounced cardioprotective effect. The efficiency of acute adaptation to hypoxia is reduced in metabolic syndrome. However, the effectiveness of the myocardial infarct size-limiting effect of chronic continuous hypoxia in metabolic syndrome remains an understudied fact. The aim. To study the effectiveness of the development of the myocardial infarct size-limiting effect of chronic continuous hypoxia in rats with metabolic syndrome. Materials and methods. The study was carried out on 43 Wistar rats. Adaptation of animals to chronic hypoxia was performed during 21 days in a hypoxic chamber (12 % O2, 0.3 % CO2). Metabolic syndrome was modeled by keeping rats on a high-carbohydrate and high-fat diet (proteins 16 %, fats 21 %, carbohydrates 46 % (including fructose 17 %), cholesterol 0.125 %, cholic acid 0.5 %) for 12 weeks with replacement of drinking water with 20% fructose solution. Coronary occlusion-reperfusion was performed in vivo. The effect of chronic hypoxia and metabolic syndrome on myocardial infarct size was assessed. Results. It was found that myocardial infarct size in rats after chronic continuous hypoxia was 38 % less than in animals of the control group. In rats which were kept on a high-carbohydrate and high-fat diet we observed the obesity, decreased glucose tolerance, increased serum triglycerides level, and hypertension. Adaptation to chronic continuous hypoxia in animals on a high-carbohydrate and high-fat diet improved carbohydrate metabolism, but did not affect the severity of other metabolic disorders. At the same time, the myocardial infarct size-limiting effect of chronic hypoxia was not observed in rats with metabolic syndrome. Conclusion. Metabolic syndrome eliminated myocardial infarct size-limiting effect of chronic continuous hypoxia.

THE EFFECTS OF DEXMEDETOMIDINE ON OXYGEN-GLUCCOSE DEFICIENCY/REOXYGENIZATION-INDUCED DAMAGE IN MOUSE NEUROBLASTOMA CELLS

<h3>Aim</h3> In this study, it was aimed to evaluate effects of dexmedetomidine (DEX) on the damage model induced by hypoxia-ischemia in oxygen and glucose deprivation (OGD) through cell viability (MTT), inflammation (iNOS, eNOS, IL-6, and TGF-b), apoptosis (TUNEL) in the neuroblastoma cell line of mouse origin (NB2a). <h3>Materials and Methods</h3> The effects of DEX on cell viability and toxicity were analyzed with MTT and NTT. Four groups were established for observing the effects of hypoxia-ischemia in the oxygen and glucose deprivation (OGD) model. Control group (C); Cells are incubated with DMEM high glucose medium for 4 hours in normoxic condition. OGY group: Cells are incubated in the oxygen and glucose deprivation (OGD) medium for 4 hours in hypoxic condition which have a gas mixture of 5% CO2, 1% O2 and 92% N2 for 4 hours in hypoxic chamber. DEX+OGD group: Before the OGD procedure, cells are treated with DEX 10 µM for 3 hours. OGD+DEX group: After the OGD procedure, cells were treated with DEX 10 µM for 3 hours. Inflammatory effects were tested by immunohistochemistry and H-scores belonging to antibodies of iNOS, eNOS, and TGF-b were calculated. Apoptotic effects of DEX were tested by the TUNEL method. Data were analyzed one-way ANOVA and Kruskal-Wallis. Data were given mean±SD. p<0.05 is statically significant. <h3>Results</h3> Dexmedetomidine increased the cell viability in those all concentrations (0.1-45 µM) except 60 uM (p<0.01, Figure 1a). H-score value belongs to eNOS antibody decreased significantly with DEX 10 mm in DEX+OGD group and OGD+DEX group compared to OGD group (p<0.001, Figure 1b). H-score value belongs to IL-6 antibody did not show any difference between OGD, DEX+OGD, and OGD+DEX groups (Figure 1d). H-score value belongs to TGF-b antibody significantly decreased with DEX 10 mM in DEX+OGD group and OGD+DEX group compared to OGD group (p<0.0001, Figure 1d). The apoptotic index decreased significantly in the DEX+OGD group and OGD+DEX group compared to Group OGD (p<0.001, Figure 1f) <h3>Conclusions</h3> Dexmedetomidine increased the cell viability in those almost all concentrations (0.1-45 µM) except 60 µM. Either before or after injury treatment with DEX 10 mM decreased apoptosis induced by hypoxia-ischemia in NB2a cells. Treating with DEX 10 mM before OGD injury decreased both iNOS and eNOS antibodies as well. Neuroprotective effects of DEX against OGD injury might be due to antiapoptotic and antioxidative effects.

High Keratin Secretion of T47D Cell under Hypoxic Condition

BACKGROUND: Hypoxia adaptation in cancer cells is mediated by hypoxia inducible factor (HIF)-1 that induces the expression of various proteins. Up to date, the analysis of T47D breast cancer cells-secreted protein is still limited. The aim of this study was to compare the protein profile secreted by T47D breast cancer cells under hypoxic and to normoxic conditions and identify the proteins as candidate for hypoxia marker proteins in T47D breast cancer cells.METHODS: T47D breast cancer cells were cultured under standard conditions. Cells were subcultured in normoxic and hypoxic conditions. The normoxic group was incubated with 20% oxygen and the hypoxic groups were incubated in a hypoxic chamber with 0.5% and 5% oxygen for 6, 24, and 48 hours in serum free medium. Proteins in the culture media were isolated and precipitated. Protein concentrations released by the cells were then measured by bicinchoninic assay (BCA). Protein bands were visualized by the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) method. The bands that had differences between the hypoxic and normoxic groups were further analyzed by liquid chromatography-mass spectrophotometer (LC-MS).RESULTS: Hypoxic groups showed higher secretory protein than normoxic group. Protein bands were found in the 0.5% hypoxic group with a size of 50-75 kDa. Secretory proteins identified by LC-MS were keratin 1, 2, 9, and 10.CONCLUSION: The T47D cell line under 0.5% hypoxic treatment showed higher secretory proteins that identified as keratin 1, 2, 9, and 10.KEYWORDS: hypoxia, secretory protein, liquid chromatography-mass spectrophotometer

Rosiglitazone attenuates hypoxia-induced renal cell apoptosis by inhibiting NF-κB signaling pathway in a PPARγ-dependent manner

Background In recent years, peroxisome proliferator-activated receptor γ (PPARγ) has been found to be closely associated with hypoxia renal disease. The aim of this study was to investigate the relationship between rosiglitazone and mitochondrial apoptosis in renal tissue and its associated mechanisms. Methods Twenty-four male Sprague-Dawley rats were randomly divided into three groups (n = 8 in each): normal control group, hypoxia injury group (equal volume of 0.9% saline), and PPARγ agonist group (Rosiglitazone, 10 mg/kg · d, intraperitoneally). The hypoxia injury group and PPARγ agonist group were placed in a hypoxia chamber and the simulated altitude was set at 7,000 m for 7 days. Blood and kidney samples were collected after 7 days. The quantitative real-time polymerase chain reaction and Western blot methods were used to determine the expression of PPARγ, nuclear factor kappa-B (NF-κB), B-cell lymphoma-2 (Bcl-2), and Bax. Results The results showed that compared with the normal control group, the renal tissue of rats after hypoxia was severely damaged, as shown by massive renal tubular epithelial cell degeneration and detachment, and renal tubular dilation. The NF-κB protein expression significantly increased, the Bcl-2 protein and mRNA expression significantly decreased, and Bax protein and mRNA expression significantly increased (p < .05 for all). Renal injury was much less severe in the PPARγ agonist group compared to the hypoxia injury group. Conclusions Rosiglitazone can alleviate hypoxia renal injury, with the possible mechanism involving attenuation of apoptosis by inhibiting the activation of the NF-κB signaling pathway in a PPARγ-dependent manner and increasing Bcl-2 and decreasing Bax expression.

Sheng-Mai San extracts attenuate heart injury in chronic intermittent hypoxia via suppressing oxidative damage and regulating glucose metabolism

BackgroundSheng-Mai San extracts (SMSE), derived from a Chinese medicine formula Sheng-Mai San, benefits prevention and treatment of cardiovascular disease. However, it remains unknown how SMSE exerts its effects on chronic intermittent hypoxia (CIH)-induced heart injury. ObjectiveThis study aims to evaluate the efficacy of SMSE on CIH-induced heart injury by suppressing oxidative damage and promoting glucose energy metabolism. MethodsThe CIH-induced model was established through the mice being placed in a hypoxic chamber with 7 to 8% O2. Then, such indexes as cardiac function, histopathological changes, creatine kinase (CK), superoxide dismutase (SOD) and myeloperoxidase (MPO) and malondialdehyde (MDA) were measured. Lastly, the influence on energy metabolism was explored through determining the ratio of ATP/ADP, concentrations of pyruvate and lactic acid, ATPase activities and levels of proteins including pyruvate kinase M2 (PKM2), fructose 6-phosphate kinase (6-FPK) and pyruvate dehydrogenase kinase 4 (PDK4). ResultsSMSE could obviously improve cardiac function and histopathological changes of heart tissue; reduce CK and MPO activities as well as MDA and lactic acid contents; increase SOD and ATPase activities, the ratio of ATP/ADP and pyruvate concentration; and regulate the protein expression of PKM2, 6-FPK and PDK4. ConclusionSMSE could attenuate CIH-induced heart injury via suppressing oxidative damage and regulating glucose metabolism.