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Background: Beetroot juice (BRJ) supplementation has been shown to increase sports performance under hypoxic conditions and to improve athletes’ recovery. Objectives: In the present study, we aimed to investigate the effect of acute BRJ supplementation on slalom (SL) run performance and muscle soreness (MS) in Alpine skiers at moderate to high altitudes. Methods: Ten male Alpine skiers received 220 mL of BRJ (8.9 mmol/L nitrate) or placebo (PLA) in 2 sessions with a 7-d wash out interval in a randomized, crossover, PLA-controlled, double-blind study. The 90-s box jump (BJ90), agility hexagonal obstacle jump (Hex Jump), and wall-sit tests were measured before on-hill SL runs in both sessions. After the functional tests, SL run performance was measured by time to complete 2 runs on the SL course; immediately after each SL run, the rating of perceived exertion (RPE) was recorded. In addition, perceived MS was recorded using the visual analog scale at 12, 24, and 48 h after the SL runs. Results: The data were meticulously analyzed using 2-way repeated measures analysis of variance and paired t tests with significance set at P < 0.05. The findings were significant, indicating that compared with PLA, BRJ notably improved wall-sit and BJ90 performances (P < 0.05), while a substantial reduction was observed in RPE, Hex Jump, and MS (P < 0.05). A 1.74% shorter time to complete SL runs was observed in the BRJ group compared with the PLA group; however, there were no significant differences between the PLA and BRJ groups (P > 0.05). Conclusions: These results underscore the potential of BRJ supplementation to enhance sports performance and reduce MS in Alpine skiers under hypoxic conditions.

Adipose-derived mesenchymal stem cells (ASCs) have the potential to differentiate into bone, cartilage, fat, and neural cells and promote tissue regeneration and healing. It is known that they can have variable responses to hypoxic conditions. In the present study, we aimed to explore diverse changes in the cells and secretome of ASCs under a hypoxic environment over time and to present the possibility of ASCs as therapeutic agents from a different perspective. The expression differences of proteins between normoxic and hypoxic conditions (6, 12, or 24 h) were specifically investigated in human ASCs using 2-DE combined with MALDI-TOF MS analysis, and secreted proteins in ASC-derived conditioned media (ASC-derived CM) were examined by an adipokine array. In addition, genetic and/or proteomic interactions were assessed using a DAVID and miRNet functional annotation bioinformatics analysis. We found that 64 and 5 proteins were differentially expressed in hypoxic ASCs and in hypoxic ASC-derived CM, respectively. Moreover, 7 proteins among the 64 markedly changed spots in hypoxic ASCs were associated with bone-related diseases. We found that two proteins, cathepsin D (CTSD) and cathepsin L (CTSL), identified through an adipokine array independently exhibited significant efficacy in promoting osteocyte differentiation in bone-marrow-derived mesenchymal stem cells (BM-MSCs). This finding introduces a promising avenue for utilizing hypoxia-preconditioned ASC-derived CM as a potential therapeutic approach for bone-related diseases. [ABSTRACT FROM AUTHOR]

Hypoxic environment is known as one of the critical factors in various physiological/pathological processes. It is imperative to recapitulate oxygen level in microscale for human physiology/pathology induced by hypoxia. Herein, we propose an oxygen-regulating system that can be applied to in vitro tissue models. We fabricated a microdevice with a gas-permeable membrane, allowing oxygen diffusion without direct contact to cells. We verified the formation of oxygen level less than 2% O₂ concentration inside the device through computational simulation and experiments. H9c2 heart myoblasts were exposed to hypoxic condition in the device, and their cell viability were investigated. We anticipate that our system will be integrated with a platform to study hypoxia-induced human physiology and pathology as an efficient oxygen-regulating system.

Hypoxic environment is known as one of the critical factors in various physiological/pathological processes. It is imperative to recapitulate oxygen level in microscale for human physiology/pathology induced by hypoxia. Herein, we propose an oxygen-regulating system that can be applied to in vitro tissue models. We fabricated a microdevice with a gas-permeable membrane, allowing oxygen diffusion without direct contact to cells. We verified the formation of oxygen level less than 2% O₂ concentration inside the device through computational simulation and experiments. H9c2 heart myoblasts were exposed to hypoxic condition in the device, and their cell viability were investigated. We anticipate that our system will be integrated with a platform to study hypoxia-induced human physiology and pathology as an efficient oxygen-regulating system.

Availability of oxygen plays an important role in tissue organization and cell-type specific metabolism. It is, however, difficult to analyze hypoxia-related adaptations in vitro because of inherent limitations of experimental model systems. In this study, we establish a microfluidic tissue culture protocol to generate hypoxic gradients in vitro, mimicking the conditions found in the liver acinus. To accomplish this, four microfluidic chips, each containing two chambers, were serially connected to obtain eight interconnected chambers. HepG2 hepatocytes were uniformly seeded in each chamber and cultivated under a constant media flow of 50 µL/h for 72 h. HepG2 oxygen consumption under flowing media conditions established a normoxia to hypoxia gradient within the chambers, which was confirmed by oxygen sensors located at the inlet and outlet of the connected microfluidic chips. Expression of Hif1α mRNA and protein was used to indicate hypoxic conditions in the cells and albumin mRNA and protein expression served as a marker for liver acinus-like zonation. Oxygen measurements performed over 72 h showed a change from 17.5% to 15.9% of atmospheric oxygen, which corresponded with a 9.2% oxygen reduction in the medium between chamber1 (inlet) and 8 (outlet) in the connected microfluidic chips after 72 h. Analysis of Hif1α expression and nuclear translocation in HepG2 cells additionally confirmed the hypoxic gradient from chamber1 to chamber8. Moreover, albumin mRNA and protein levels were significantly reduced from chamber1 to chamber8, indicating liver acinus zonation along the oxygen gradient. Taken together, microfluidic cultivation in interconnected chambers provides a new model for analyzing cells in a normoxic to hypoxic gradient in vitro. By using a well-characterized cancer cell line as a homogenous hepatocyte population, we also demonstrate that an approximate 10% reduction in oxygen triggers translocation of Hif1α to the nucleus and reduces albumin production. [ABSTRACT FROM AUTHOR]

The hypoxic environment is a substantial factor in maintenance, proliferation, and differentiation of the cell cultures. Low oxygen is known as a potent chondrogenesis stimulus in stem cells that is important for clinical application and engineering of functional cartilage. Hypoxia can potentially induce angiogenesis process by secretion of cytokines. This systematic review goal is to discover the effect of hypoxic condition on tendon/ ligament culture and the best oxygen level of hypoxia for in vitro and in vivo studies. We included 21 articles. A comprehensive review of this database confirms that the hypoxic condition is a substantial factor in the maintenance, proliferation, and differentiation of ligament/tendon cultures. Cell proliferation in the severe hypoxic (oxygen concentration of 1%) group at 24 h postcultivation was considered significant, but cell proliferation was markedly inhibited in the severe hypoxic group after 48 h.

Abstract Background Improved chondrogenic differentiation of mesenchymal stem cells (MSCs) by genetic regulation is a potential method for regenerating articular cartilage. MiR-127-5p has been reported to promote cartilage differentiation of rat bone marrow MSCs (rMSCs); however, the regulatory mechanisms underlying hypoxia-stimulated chondrogenic differentiation remain unknown. Methods rMSCs were induced to undergo chondrogenic differentiation under normoxic or hypoxic conditions. Expression of lncRNA DNM3OS, miR-127-5p, and GREM2 was detected by quantitative real-time PCR. Proteoglycans were detected by Alcian blue staining. Western blot assays were performed to examine the relative levels of GREM2 and chondrogenic differentiation related proteins. Luciferase reporter assays were performed to assess the association among DNM3OS, miR-127-5p, and GREM2. Results MiR-127-5p levels were upregulated, while DNM3OS and GREM2 levels were downregulated in rMSCs induced to undergo chondrogenic differentiation, and those changes were attenuated by hypoxic conditions (1% O2). Further in vitro experiments revealed that downregulation of miR-127-5p reduced the production of proteoglycans and expression of chondrogenic differentiation markers (COL1A1, COL2A1, SOX9, and ACAN) and osteo/chondrogenic markers (BMP-2, p-SMAD1/2). MiR-127-5p overexpression produced the opposite results in rMSCs induced to undergo chondrogenic differentiation under hypoxic conditions. GREM2 was found to be a direct target of miR-127-5p, which was suppressed in rMSCs undergoing chondrogenic differentiation. Moreover, DNM3OS could directly bind to miR-127-5p and inhibit chondrogenic differentiation of rMSCs via regulating GREM2. Conclusions Our study revealed a novel molecular pathway (DNM3OS/miR-127-5p/GREM2) that may be involved in hypoxic chondrogenic differentiation.

Neuronal death is the main cause of nerve function impairment after spinal cord injury (SCI). Exosome-based therapy has become a novel strategy for tissue injury repair. We designed a method to treat SCI using exosomes secreted by adipose tissue–derived stromal cells (ADSCs) under hypoxic conditions. We established a neuronal oxygen-glucose deprivation and reperfusion (OGD/R) model in vitro to simulate the hypoxic environment after SCI. We observed that exosomes derived from hypoxia-conditioned ADSCs (Hypo-exo) significantly reduced neuronal apoptosis after OGD. By establishing a rat SCI model, we found that Hypo-exo can significantly reduce the formation of cavities in the injured area and improve the functional recovery of the hindlimbs of rats after injury. To explore the molecular mechanism, we conducted microRNA sequencing analysis of exosomes. Through real-time polymerase chain reaction, dual luciferase reporter assays and signaling pathway chip analysis, we determined that miR-499a-5p regulates the JNK3/c-jun-apoptotic signaling pathway by targeting JNK3. Further, we verified the expression of the key proteins in the JNK3/c-jun-apoptotic signaling pathway by immunofluorescence and Western blotting. These results support the hypothesis that Hypo-exo can reduce neuronal apoptosis after SCI and may provide new methods to treat SCI. [ABSTRACT FROM AUTHOR]

To control asparagus harvest timing, we investigated the effects of short-term low (5%) oxygen (O2) treatment in the cultivation area on asparagus growth and yield using a closed cultivation system. During 120 days of cultivation, low O2 treatments were initiated at 0 to 4, 20 to 24, and 40 to 44 days after planting (DAP). The sprouting spears and control crown yield gradually decreased with increasing DAP. However, low O2 treatment at 0 to 4 DAP significantly delayed the decrease until 80 DAP, although the total yield did not change during cultivation. In contrast, low O2 treatments at 20 to 24 and 40 to 44 DAP did not affect yield performance. Taken together, short-term low O2 treatment immediately after planting can change the harvest timing of white asparagus and can be used for effective asparagus culturing in a closed system, such as a plant factory.

The hypoxic environment is a substantial factor in maintenance, proliferation, and differentiation of the cell cultures. Low oxygen is known as a potent chondrogenesis stimulus in stem cells that is important for clinical application and engineering of functional cartilage. Hypoxia can potentially induce angiogenesis process by secretion of cytokines. This systematic review goal is to discover the effect of hypoxic condition on tendon/ ligament culture and the best oxygen level of hypoxia for in vitro and in vivo studies. We included 21 articles. A comprehensive review of this database confirms that the hypoxic condition is a substantial factor in the maintenance, proliferation, and differentiation of ligament/tendon cultures. Cell proliferation in the severe hypoxic (oxygen concentration of 1%) group at 24 h postcultivation was considered significant, but cell proliferation was markedly inhibited in the severe hypoxic group after 48 h. [ABSTRACT FROM AUTHOR]

Background: Improved chondrogenic differentiation of mesenchymal stem cells (MSCs) by genetic regulation is a potential method for regenerating articular cartilage. MiR-127-5p has been reported to promote cartilage differentiation of rat bone marrow MSCs (rMSCs); however, the regulatory mechanisms underlying hypoxia-stimulated chondrogenic differentiation remain unknown. Methods: rMSCs were induced to undergo chondrogenic differentiation under normoxic or hypoxic conditions. Expression of lncRNA DNM3OS, miR-127-5p, and GREM2 was detected by quantitative real-time PCR. Proteoglycans were detected by Alcian blue staining. Western blot assays were performed to examine the relative levels of GREM2 and chondrogenic differentiation related proteins. Luciferase reporter assays were performed to assess the association among DNM3OS, miR-127-5p, and GREM2. Results: MiR-127-5p levels were upregulated, while DNM3OS and GREM2 levels were downregulated in rMSCs induced to undergo chondrogenic differentiation, and those changes were attenuated by hypoxic conditions (1% O2). Further in vitro experiments revealed that downregulation of miR-127-5p reduced the production of proteoglycans and expression of chondrogenic differentiation markers (COL1A1, COL2A1, SOX9, and ACAN) and osteo/chondrogenic markers (BMP-2, p-SMAD1/2). MiR-127-5p overexpression produced the opposite results in rMSCs induced to undergo chondrogenic differentiation under hypoxic conditions. GREM2 was found to be a direct target of miR-127-5p, which was suppressed in rMSCs undergoing chondrogenic differentiation. Moreover, DNM3OS could directly bind to miR-127-5p and inhibit chondrogenic differentiation of rMSCs via regulating GREM2. Conclusions: Our study revealed a novel molecular pathway (DNM3OS/miR-127-5p/GREM2) that may be involved in hypoxic chondrogenic differentiation. [ABSTRACT FROM AUTHOR]

Declining oxygen concentrations in aquatic habitats represent extreme conditions that threaten benthic life. Hypoxia has recently become an important research topic, as areas affected by these phenomena are spreading globally. Omura Bay is one of the most highly enclosed seas in Japan, and severely hypoxic conditions occur at the bottom every summer. We conducted a preliminary study in the center of the bay to evaluate how seasonal hypoxia affects the abundance and community composition of benthic meiofauna, with particular reference to copepods. The copepod densities and their nauplii differed significantly among seasonal categories (before, during, and after hypoxia). Furthermore, the degree of the seasonal decline in copepods during hypoxia seemed much more severe than that in nematodes, the most abundant meiofauna. The assemblages of adult copepods had the simplest composition during hypoxia, when harpacticoid copepods in the family Cletodidae, which have smaller and slender bodies, occurred at significantly higher frequencies (a contribution of 84% to the mean similarity among seasons). After hypoxia, the relative abundance of copepods in the families Ectinosomatidae and Longipediidae increased, which may likely be attributed to their higher swimming abilities and rapid recruitment via specific planktonic nauplius stages, respectively. High frequencies of copepods in the Cletodidae family have also been observed under hypoxic conditions in the Mediterranean Sea, suggesting that similar processes affect benthic copepod communities, which work to the advantage of cletodid species with small and slender forms in the subtidal sediment bottom under severe hypoxia in Omura Bay and other regions. [ABSTRACT FROM AUTHOR]

Abstract Background Treatment with bone marrow mesenchymal stem cells (BM-MSCs) has been demonstrated to be an excellent cellular-based therapeutic strategy for treating myocardial infarction (MI). However, most of the patients suffering with MI are elderly. Hypoxic conditions can cause apoptosis of BM-MSCs, and this type of apoptosis is more prevalent in aged BM-MSCs. Decreased autophagy is one of the mechanisms underlying aging. The aim of this study is to uncover whether the increased hypoxic injury of aged BM-MSCs is due to autophagy and whether reducing autophagy diminishes the tolerance of hypoxia in aged BM-MSCs. Methods Young and aged BM-MSCs were isolated from male young and aged GFP/Fluc transgenic C57BL/6 mice respectively and then exposed to hypoxia and serum deprivation (H/SD) injury. The apoptosis level induced by H/SD was measured by terminal deoxynucleotidy transferase-mediated dUTP nick end-labeling (TUNEL) assay. Additionally, autophagy was analyzed via transfection with plasmids encoding green fluorescent protein-microtubule-associated protein lightchain3 (GFP-LC3), and autophagic vacuoles were visualized with transmission electron microscopy. Meanwhile, protein expression was measured by western blot analysis. Autophagic activity was manipulated by the administration of IGF-1 (insulin-like growth factor siRNA) and 3-methyladenine (3MA). Furthermore, young, aged, and the IGF-1 siRNA-transfected aged BM-MSCs were transplanted to myocardial infarcted adult C57BL/6 mice respectively. In vivo longitudinal in vivo bioluminescence imaging (BLI) of transplanted BM-MSCs was performed to monitor the survival of transplanted BM-MSCs in each groups. Results Aged BM-MSCs exhibited a higher rate of apoptosis compared with young BM-MSCs under hypoxic conditions. Additionally, the level of autophagy was lower in aged BM-MSCs compared with young BM-MSCs under normoxic and hypoxic conditions. Meanwhile, hypoxia decreased the activity of the protein kinase B (Akt) and mammalian target of rapamycin (mTOR) signaling pathway in young and aged BM-MSCs, but aged BM-MSCs exhibited a relatively stronger Akt/mTOR activity compared with young BM-MSCs. In addition, IGF-1 knockdown significantly decreased the level of apoptosis in aged BM-MSCs under normoxic and hypoxic conditions. IGF-1 knockdown also decreased the activity of the Akt/mTOR signaling pathway and increased the level of autophagy in aged BM-MSCs under hypoxic condition. Furthermore, IGF-1 knockdown protected aged BM-MSCs from hypoxic injury by increasing the level of autophagy, thereby promoting the survival of aged BM-MSCs after myocardial infarction transplantation. Conclusion This study demonstrates that reducing autophagy decreases the hypoxia tolerance of aged BM-MSCs. Maintaining optimal levels of autophagy may serve as a new strategy in treating MI by BM-MSC transplantation in aged patients.

The Effect of Intermittent Hypoxic Breathing on Blood Pressure and Nitric Oxide in Hypertensive Patients with Excess Weight. JEPonline 2020;23(3):63- 76. The purpose of this study was to investigate the effect of intermittent hypoxic exposure (IHE) on blood pressure (BP), nitric oxide metabolites (NOx), malondialdehyde (MDA) levels and the 6-min walk distance (6MWD) test in hypertensive patients with excess weight. Twenty-seven patients on medications with a BMI of 25 to 31.6 kg·m-2 were randomized into the Control Group (CON) or the IHE Group. The IHE Group was exposed to 8 sets of a 3-min on-off of hypoxic gas (FIO2 = 0.14) interspersed with normoxic gas (FIO2 = 0.21) for 48 min·d-1, 2 d·wk-1 for 6 wks during rest; whereas, the patients in the CON Group had no intervention. All the parameters were measured before and after the experimental period. Relative to the CON Group, the IHE Group showed a significant decrease in all BP values [systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial pressure, (MAP)] and plasma MDA levels, and an increase in plasma NOx levels and 6MWD. The findings suggest that IHE during rest revealed substantial improvement in BP levels. The mechanisms behind the changes may relate to NO elevation and MDA reduction. This training program helped improve exercise capacity in patients with excess weight and hypertension. [ABSTRACT FROM AUTHOR]

Scrippsiella trochoidea produces a calcareous walled cyst with spines during its life history. It has been recently reported that the calcareous wall can be decalcified under acidified conditions in coastal areas linked to hypoxia caused by bacterial activities. In this study, in order to determine whether the calcareous cysts can be a biological tool for indicating an in situ hypoxic environment, the morphology of S. trochoidea cysts in the surface sediments of Ise Bay, Japan, was examined in relation to dissolved oxygen concentrations in the bottom water. The surface sediments were collected from the inner to outer parts of the bay in May 2014 and June 2018. The living cysts of S. trochoidea were counted separately into two morphotypes: cyst with calcareous wall (calcareous walled-type cyst) and without the wall (naked-type cyst). The proportions (%) of naked-type cyst abundance in the total living cyst population showed an increasing tendency in the hypoxic environment of the inner and central parts of Ise Bay, but not in the normoxic environment at the mouth. This result supports the supposition that the naked-type cyst of S. trochoidea can be used as a biological indicator for the assessment of hypoxia in coastal areas. [ABSTRACT FROM AUTHOR]

The entomopathogenic fungi encounter hypoxic conditions in both nature and artificial culture. Alcohol dehydrogenases (ADHs) are a group of oxidoreductases that occur in many organisms. Here we demonstrate that an alcohol dehydrogenase I, MaADH1, in the locust-specific fungal pathogen, Metarhizium acridum, functions in acetaldehyde detoxification mechanism under hypoxic conditions in growth and sporulation. The MaADH1 was highly expressed in sporulation stage under hypoxic conditions. Compared with a wild-type strain, the ΔMaADH1 mutant showed inhibited growth and sporulation under hypoxic conditions, but no impairment under normal conditions. Under hypoxic conditions, ΔMaADH1 mutant produced significant decreased alcohol, but significant increased acetaldehyde compared to wild type. M. acridum was sensitive to exogenous acetaldehyde, exhibiting an inhibited growth and sporulation with acetaldehyde added in the medium. MaADH1 did not affect virulence. Our results indicated that the MaADH1 was critical to growth and sporulation under hypoxic stress by detoxification of acetaldehyde in M. acridum.

Diabetes mellitus (DM) is a group of diseases characterized by abnormally high levels of glucose in the blood stream. In developing a potential therapy for diabetic patients, pancreatic cells transplantation has drawn great attention. However, the hinder of cell transplantation for diabetes treatment is insufficient sources of insulin-producing cells. Therefore, new cell based therapy need to be developed. In this regard, human embryonic stem cells (hESCs) may serve as good candidates for this based on their capability of differentiation into various cell types. In this study, we designed a new differentiation protocol that can generate hESC-derived insulin-producing cells (hES-DIPCs) in a hypoxic condition. We also emphasized on the induction of definitive endoderm during embryoid bodies (EBs) formation. After induction of hESCs differentiation into insulin-producing cells (IPCs), the cells obtained from the cultures exhibited pancreas-related genes such as Pdx1, Ngn3, Nkx6.1, GLUT2, and insulin. These cells also showed positive for DTZ-stained cellular clusters and contained ability of insulin secretion in a glucose-dependent manner. After achievement to generated functional hES-DIPCs in vitro, some of the hES-DIPCs were then encapsulated named encapsulated hES-DIPCs. The data showed that the encapsulated cells could possess the function of insulin secretion in a time-dependent manner. The hES-DIPCs and encapsulated hES-DIPCs were then separately transplanted into STZ-induced diabetic mice. The findings showed the significant blood glucose levels regulation capacity and declination of IL-1β concentration in all transplanted mice. These results indicated that both hES-DIPCs and encapsulated hES-DIPCs contained the ability to sustain hyperglycemia condition as well as decrease inflammatory cytokine level in vivo. The findings of this study may apply for generation of a large number of hES-DIPCs in vitro. In addition, the implication of this work is therapeutic value in type I diabetes treatment in the future. The application for type II diabetes treatment remain to be investigated.

The entomopathogenic fungi encounter hypoxic conditions in both nature and artificial culture. Alcohol dehydrogenases (ADHs) are a group of oxidoreductases that occur in many organisms. Here we demonstrate that an alcohol dehydrogenase I, MaADH1 , in the locust-specific fungal pathogen, Metarhizium acridum , functions in acetaldehyde detoxification mechanism under hypoxic conditions in growth and sporulation. The MaADH1 was highly expressed in sporulation stage under hypoxic conditions. Compared with a wild-type strain, the Δ MaADH1 mutant showed inhibited growth and sporulation under hypoxic conditions, but no impairment under normal conditions. Under hypoxic conditions, Δ MaADH1 mutant produced significant decreased alcohol, but significant increased acetaldehyde compared to wild type. M. acridum was sensitive to exogenous acetaldehyde, exhibiting an inhibited growth and sporulation with acetaldehyde added in the medium. MaADH1 did not affect virulence. Our results indicated that the MaADH1 was critical to growth and sporulation under hypoxic stress by detoxification of acetaldehyde in M. acridum. [ABSTRACT FROM AUTHOR]

Hypoxic environment is known as one of the critical factors in various physiological/pathological processes. It is imperative to recapitulate oxygen level in microscale for human physiology/pathology induced by hypoxia. Herein, we propose an oxygen-regulating system that can be applied to in vitro tissue models. We fabricated a microdevice with a gas-permeable membrane, allowing oxygen diffusion without direct contact to cells. We verified the formation of oxygen level less than 2% O2 concentration inside the device through computational simulation and experiments. H9c2 heart myoblasts were exposed to hypoxic condition in the device, and their cell viability were investigated. We anticipate that our system will be integrated with a platform to study hypoxia-induced human physiology and pathology as an efficient oxygen-regulating system.

Mesenchymal stem cells (MSCs) are accepted as a promising tool for therapeutic purposes. However, low proliferation and early senescence are still main obstacles of MSCs expansion for using as cell-based therapy. Thus, clinical scale of cell expansion is needed to obtain a large number of cells serving for further applications. In this study, we investigated the value of embryonic stem cells conditioned medium (ESCM) for in vitro expansion of Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) as compared to typical culture medium for MSCs, Dulbecco's modified Eagle's medium with 1.0 g/l glucose (DMEM-LG) supplemented with 10 % FBS, under hypoxic condition. The expanded cells from ESCM (ESCM-MSCs) and DMEM-LG (DMEM-MSCs) were characterized for both phenotype and biological activities including proliferation rate, population doubling time, cell cycle distribution and MSCs characteristics. ESCM and DMEM-LG could enhance WJ-MSCs proliferation as 204.66 ± 10.39 and 113.77 ± 7.89 fold increase at day 12, respectively. ESCM-MSCs could express pluripotency genes including Oct- 4, Oct- 3/4, Nanog, Klf- 4, C- Myc and Sox- 2 both in early and late passages whereas the downregulations of Oct- 4 and Nanog were detected in late passage cells of DMEM-MSCs. The 2 cell populations also showed common MSCs characteristics including normal cell cycle, fibroblastic morphology, cell surface markers expressions (CD29, CD44, CD90, CD34, CD45) and differentiation capacities into adipogenic, chondrogenic and osteogenic lineages. Moreover, our results revealed that ESCM exhibited as a rich source of several factors which are required for supportive WJ-MSCs proliferation. In conclusion, ESCM under hypoxic condition could accelerate WJ-MSCs expansion while maintaining their pluripotency properties. Our knowledge provide short term and cost-saving in WJ-MSCs expansion which has benefit to overcome insufficient cell numbers for clinical applications by reusing the discarded cell culture supernates from human ES culture system. Moreover, these findings can also apply for stem cell banking, regenerative medicine and pharmacological applications. [ABSTRACT FROM AUTHOR]

The design, implementation, and verification of a signal simulator for the generation of patho‐physiological records of foetal electrocardiograms (fECGs) during the prenatal period are briefly reported. The simulator enables users to model the patho‐physiological changes that occur within the foetus' myocardium under hypoxic conditions (hypoxemia, hypoxia, asphyxia, etc.) during the 20th to 42nd week of pregnancy. The simulator deploys a dynamic fECG model including an actual fECG record taken from clinical practice, patho‐physiological cardiotocography (CTG), and ST‐analysis (STAN) records along with the ratio of T waves to the QRS complex; as well as clinical recommendations by FIGO (International Federation of Gynecology and Obstetrics) for classifying these records. By comparing synthesised and real patho‐physiological CTG and STAN records, the functionality of the simulator, which effectively captured significant indicators of the foetus' condition during the prenatal period including fECG morphology, dynamic fECG characteristics, and others is evaluated and validated. The simulator enables users to test both current and emerging approaches in a very challenging area of gynaecology, namely the identification/classification of hypoxic conditions in the foetus during labour. Obstetricians can also use the simulator as a reference tool during the evaluation of suspect fECG abnormalities. [ABSTRACT FROM AUTHOR]

Long urethral strictures are often treated with autologous genital skin and buccal mucosa grafts; however, risk of hair ingrowth and donor site morbidity, restrict their application. To overcome this, we introduced a tissue-engineered human urethra comprising adipose-derived stem cell (ASC)-based self-assembled scaffold, human urothelial cells (UCs) and smooth muscle cells (SMCs). ASCs were cultured with ascorbic acid to stimulate extracellular matrix (ECM) production. The scaffold (ECM) was stained with collagen type-I antibody and the thickness was measured under a confocal microscope. Results showed that the thickest scaffold (28.06 ± 0.59 μm) was achieved with 3 × 104 cells/cm2 seeding density, 100 μg/mL ascorbic acid concentration under hypoxic and dynamic culture condition. The biocompatibility assessment showed that UCs and SMCs seeded on the scaffold could proliferate and maintain the expression of their markers (CK7, CK20, UPIa, and UPII) and (α-SMA, MHC and Smootheline), respectively, after 14 days of in vitro culture. ECM gene expression analysis showed that the ASC and dermal fibroblast-based scaffolds (control) were comparable. The ASC-based scaffold can be handled and removed from the plate. This suggests that multiple layers of scaffold can be stacked to form the urothelium (seeded with UCs), submucosal layer (ASCs only), and smooth muscle layer (seeded with SMCs) and has the potential to be developed into a fully functional human urethra for urethral reconstructive surgeries. [ABSTRACT FROM AUTHOR]

Carbonic anhydrases (CAs) are acid–base regulatory proteins that modulate a variety of physiological functions. Recent findings have shown that CAIX is particularly upregulated in glioblastoma multiforme (GBM) and is associated with a poor patient outcome and survival rate. An analysis of the GSE4290 dataset of patients with gliomas showed that CAIX was highly expressed in GBM and was negatively associated with prognosis. The expression of CAIX under hypoxic conditions in GBM significantly increased in protein, mRNA, and transcriptional activity. Importantly, CAIX upregulation also regulated GBM motility, monocyte adhesion to GBM, and the polarization of tumor-associated monocytes/macrophages (TAM). Furthermore, the overexpression of CAIX was observed in intracranial GBM cells. Additionally, epidermal growth factor receptor/signal transducer and activator of transcription 3 regulated CAIX expression under hypoxic conditions by affecting the stability of hypoxia-inducible factor 1α. In contrast, the knockdown of CAIX dramatically abrogated the change in GBM motility and monocyte adhesion to GBM under hypoxic conditions. Our results provide a comprehensive understanding of the mechanisms of CAIX in the GBM microenvironment. Hence, novel therapeutic targets of GBM progression are possibly developed. [ABSTRACT FROM AUTHOR]

The direct electron transfer between human cytoglobin (Cygb) and the electrode surface, which would allow manipulating the oxidation states of the heme iron in Cygb, was first observed by immobilizing Cygb on a nanoporous gold (NPG) electrode via a carboxy-terminated alkanethiol. The voltammetric performances of the wild type and mutated Cygb-immobilized NPG electrodes were evaluated in the absence or presence of potential substrates. The obtained results demonstrated that the usefulness of the proposed method in understanding the function of Cygb in molecular basis.

Background: Mesenchymal stem cells (MSCs) can be easily expanded. They can be acquired from medical waste such as adipose and umbilical cord tissues, are influenced by culturing conditions, and exert anti-inflammatory, antioxidant, anti-fibrotic, and angiogenic effects. We analyzed the multi-directional effects of MSCs cultured under hypoxic conditions and their underlying mechanisms in the treatment of liver cirrhosis in a mouse model., Methods: Human bone marrow-derived MSCs cultured under hypoxic (5% O 2 ; hypoMSCs) and normoxic (21% O 2 ; norMSCs) conditions were compared by cap analysis of gene expression (CAGE) with or without serum from liver cirrhosis patients. The therapeutic effects of MSCs, including serum liver enzyme induction, fibrosis regression, and hepatic oxidative stress, were evaluated by injecting 1 × 10 6 , 2 × 10 5 , or 4 × 10 4 MSCs/mouse into the tail veins of mice with carbon tetrachloride (CCl 4 )-induced liver cirrhosis. Intravital imaging was performed with a two-photon excitation microscope to confirm the various MSC migration paths to the liver., Results: CAGE analysis revealed that the RNA expression levels of prostaglandin E synthase ( Ptges ) and miR210 were significantly higher in hypoMSCs than in norMSCs. In vivo analysis revealed that both hypoMSCs and norMSCs reduced serum alanine aminotransferase, oxidative stress, and fibrosis compared to that in control mice in a dose-dependent manner. However, hypoMSCs had stronger therapeutic effects than norMSCs. We confirmed this observation by an in vitro study in which hypoMSCs changed macrophage polarity to an anti-inflammatory phenotype via prostaglandin E2 (PGE2) stimulation. In addition, miR210 reduced the rate of hepatocyte apoptosis. Intravital imaging after MSC administration showed that both cell types were primarily trapped in the lungs. Relatively a few hypoMSCs and norMSCs migrated to the liver. There were no significant differences in their distributions., Conclusion: The therapeutic effect of hypoMSCs was mediated by PGE2 and miR210 production and was greater than that of norMSCs. Therefore, MSCs can be manipulated to improve their therapeutic efficacy in the treatment of liver cirrhosis and could potentially serve in effective cell therapy. MSCs produce several factors with multidirectional effects and function as "conducting cells" in liver cirrhosis., (© 2019 The Japanese Society for Regenerative Medicine. Production and hosting by Elsevier B.V.)

Background: Treatment with bone marrow mesenchymal stem cells (BM-MSCs) has been demonstrated to be an excellent cellular-based therapeutic strategy for treating myocardial infarction (MI). However, most of the patients suffering with MI are elderly. Hypoxic conditions can cause apoptosis of BM-MSCs, and this type of apoptosis is more prevalent in aged BM-MSCs. Decreased autophagy is one of the mechanisms underlying aging. The aim of this study is to uncover whether the increased hypoxic injury of aged BM-MSCs is due to autophagy and whether reducing autophagy diminishes the tolerance of hypoxia in aged BM-MSCs. Methods: Young and aged BM-MSCs were isolated from male young and aged GFP/Fluc transgenic C57BL/6 mice respectively and then exposed to hypoxia and serum deprivation (H/SD) injury. The apoptosis level induced by H/SD was measured by terminal deoxynucleotidy transferase-mediated dUTP nick end-labeling (TUNEL) assay. Additionally, autophagy was analyzed via transfection with plasmids encoding green fluorescent protein-microtubule-associated protein lightchain3 (GFP-LC3), and autophagic vacuoles were visualized with transmission electron microscopy. Meanwhile, protein expression was measured by western blot analysis. Autophagic activity was manipulated by the administration of IGF-1 (insulin-like growth factor siRNA) and 3-methyladenine (3MA). Furthermore, young, aged, and the IGF-1 siRNA-transfected aged BM-MSCs were transplanted to myocardial infarcted adult C57BL/6 mice respectively. In vivo longitudinal in vivo bioluminescence imaging (BLI) of transplanted BM-MSCs was performed to monitor the survival of transplanted BM-MSCs in each groups. Results: Aged BM-MSCs exhibited a higher rate of apoptosis compared with young BM-MSCs under hypoxic conditions. Additionally, the level of autophagy was lower in aged BM-MSCs compared with young BM-MSCs under normoxic and hypoxic conditions. Meanwhile, hypoxia decreased the activity of the protein kinase B (Akt) and mammalian target of rapamycin (mTOR) signaling pathway in young and aged BM-MSCs, but aged BM-MSCs exhibited a relatively stronger Akt/mTOR activity compared with young BM-MSCs. In addition, IGF-1 knockdown significantly decreased the level of apoptosis in aged BM-MSCs under normoxic and hypoxic conditions. IGF-1 knockdown also decreased the activity of the Akt/mTOR signaling pathway and increased the level of autophagy in aged BM-MSCs under hypoxic condition. Furthermore, IGF-1 knockdown protected aged BM-MSCs from hypoxic injury by increasing the level of autophagy, thereby promoting the survival of aged BM-MSCs after myocardial infarction transplantation. Conclusion: This study demonstrates that reducing autophagy decreases the hypoxia tolerance of aged BM-MSCs. Maintaining optimal levels of autophagy may serve as a new strategy in treating MI by BM-MSC transplantation in aged patients. [ABSTRACT FROM AUTHOR]

The suppressor of MEK null (sMEK1) protein possesses pro-apoptotic activities. In the current study, we reveal that sMEK1 functions as a novel anti-angiogenic factor by suppressing vascular endothelial growth factor (VEGF)-induced cell proliferation, migration, and capillary-like tubular structure in vitro. In addition, sMEK1 inhibited the phosphorylation of the signaling components up- and downstream of Akt, including phospholipase Cγ1 (PLC-γ1), 3-phosphoinositide-dependent protein kinase 1 (PDK1), endothelial nitric oxide synthetase (eNOS), and hypoxia-inducible factor 1 (HIF-1α) during ovarian tumor progression via binding with vascular endothelial growth factor receptor 2 (VEGFR-2). Furthermore, sMEK1 decreased tumor vascularity and inhibited tumor growth in a xenograft human ovarian tumor model. These results supply convincing evidence that sMEK1 controls endothelial cell function and subsequent angiogenesis by suppressing VEGFR-2-mediated PI3K/Akt/eNOS signaling pathway. Taken together, our results clearly suggest that sMEK1 might be a novel anti-angiogenic and anti-tumor agent for use in ovarian tumor.

Pancreatic cancer has become the fourth leading cause of cancer death in the last two decades. Only 3%-15% of patients diagnosed with pancreatic cancer had 5 year survival rate. Drug resistance, high metastasis, poor prognosis and tumour relapse contributed to the malignancies and difficulties in treating pancreatic cancer. The current standard chemotherapy for pancreatic cancer is gemcitabine, however its efficacy is far from satisfactory, one of the reasons is due to the complex tumour microenvironment which decreases effective drug delivery to target cancer cell. Studies of the molecular pathology of pancreatic cancer have revealed that activation of KRAS, overexpression of cyclooxygenase-2, inactivation of p16(INK4A) and loss of p53 activities occurred in pancreatic cancer. Co-administration of gemcitabine and targeting the molecular pathological events happened in pancreatic cancer has brought an enhanced therapeutic effectiveness of gemcitabine. Therefore, studies looking for novel targets in hindering pancreatic tumour growth are emerging rapidly. In order to give a better understanding of the current findings and to seek the direction in future pancreatic cancer research; in this review we will focus on targets suppressing tumour metastatsis and progression, KRAS activated downstream effectors, the relationship of Notch signaling and Nodal/Activin signaling with pancreatic cancer cells, the current findings of non-coding RNAs in inhibiting pancreatic cancer cell proliferation, brief discussion in transcription remodeling by epigenetic modifiers (e.g., HDAC, BMI1, EZH2) and the plausible therapeutic applications of cancer stem cell and hyaluronan in tumour environment.

Pancreatic cancer is one of the most aggressive and difficult cancers to treat. Despite numerous research efforts, limited success has been achieved in the therapeutic management of patients with this disease. In the current review, we focus on one component of morphogenesis signaling, Hedgehog (Hh), with the aim of developing novel, effective therapies for the treatment of pancreatic cancer. Hh signaling contributes to the induction of a malignant phenotype in pancreatic cancer and is responsible for maintaining pancreatic cancer stem cells. In addition, we propose a novel concept linking Hh signaling and tumor hypoxic conditions, and discuss the effects of Hh inhibitors in clinical trials. The Hh signaling pathway may represent a potential therapeutic target for patients with refractory pancreatic cancer.