Quantitative Proteomic Approach Reveals Altered Metabolic Pathways in Response to the Inhibition of Lysine Deacetylases in A549 Cells under Normoxia and Hypoxia

Growing evidence is showing that acetylation plays an essential role in cancer, but studies on the impact of KDAC inhibition (KDACi) on the metabolic profile are still in their infancy. Here, we analyzed, by using an iTRAQ-based quantitative proteomics approach, the changes in the proteome of KRAS-mutated non-small cell lung cancer (NSCLC) A549 cells in response to trichostatin-A (TSA) and nicotinamide (NAM) under normoxia and hypoxia. Part of this response was further validated by molecular and biochemical analyses and correlated with the proliferation rates, apoptotic cell death, and activation of ROS scavenging mechanisms in opposition to the ROS production. Despite the differences among the KDAC inhibitors, up-regulation of glycolysis, TCA cycle, oxidative phosphorylation and fatty acid synthesis emerged as a common metabolic response underlying KDACi. We also observed that some of the KDACi effects at metabolic levels are enhanced under hypoxia. Furthermore, we used a drug repositioning machine learning approach to list candidate metabolic therapeutic agents for KRAS mutated NSCLC. Together, these results allow us to better understand the metabolic regulations underlying KDACi in NSCLC, taking into account the microenvironment of tumors related to hypoxia, and bring new insights for the future rational design of new therapies.     

Superior foetal electrocardiogram signal elicitation using a novel artificial intelligent Bayesian methodology

Innumerable casualties due to intrauterine hypoxia are a major worry during prenatal phase besides advanced patient monitoring with latest science and technology. Hence, the analysis of foetal electrocardiogram (fECG) signals is very vital in order to evaluate the foetal heart status for timely recognition of cardiac abnormalities. Regrettably, the latest technology in the cutting edge field of biomedical signal processing does not seem to yield the desired quality of fECG signals required by physicians, which is the major cause for the pathetic condition. The focus of this work is to extort non-invasive fECG signal with highest possible quality with a motive to support physicians in utilizing the methodology for the latest intrapartum monitoring technique called STAN (ST analysis) for forecasting intrapartum foetal hypoxia. However, the critical quandary is that the non-invasive fECG signals recorded from the maternal abdomen are affected by several interferences like power line interference, baseline drift interference, electrode motion interference, muscle movement interference and the maternal electrocardiogram (mECG) being the dominant interference. A novel hybrid methodology called BANFIS (Bayesian adaptive neuro fuzzy inference system) is proposed. The BANFIS includes a Bayesian filter and an adaptive neuro fuzzy filter for mECG elimination and non-linear artefacts removal to yield high quality fECG signal. Kalman filtering frame work has been utilized to estimate the nonlinear transformed mECG component in the abdominal electrocardiogram (aECG). The adaptive neuro fuzzy filter is employed to discover the nonlinearity of the nonlinear transformed version of mECG and to align the estimated mECG signal with the maternal component in the aECG signal for annulment. The outcomes of the investigation by the proposed BANFIS system proved valuable for STAN system for efficient prediction of foetal hypoxia.     

Tangeretin Ameliorates Glucose-Induced Podocyte Injury through Blocking Epithelial to Mesenchymal Transition Caused by Oxidative Stress and Hypoxia

Podocyte injury inevitably results in leakage of proteins from the glomerular filter and is vital in the pathogenesis of diabetic nephropathy (DN). The underlying mechanisms of podocyte injury facilitate finding of new therapeutic targets for DN treatment and prevention. Tangeretin is an O-polymethoxylated flavone present in citrus peels with anti-inflammatory and antioxidant properties. This study investigated the renoprotective effects of tangeretin on epithelial-to-mesenchymal transition-mediated podocyte injury and fibrosis through oxidative stress and hypoxia caused by hyperglycemia. Mouse podocytes were incubated in media containing 33 mM glucose in the absence and presence of 1–20 μM tangeretin for up to 6 days. The in vivo animal model employed db/db mice orally administrated with 10 mg/kg tangeretin for 8 weeks. Non-toxic tangeretin inhibited glucose-induced expression of the mesenchymal markers of N-cadherin and α-smooth muscle actin in podocytes. However, the reduced induction of the epithelial markers of E-cadherin and P-cadherin was restored by tangeretin in diabetic podocytes. Further, tangeretin enhanced the expression of the podocyte slit diaphragm proteins of nephrin and podocin down-regulated by glucose stimulation. The transmission electron microscopic images revealed that foot process effacement and loss of podocytes occurred in diabetic mouse glomeruli. However, oral administration of 10 mg/kg tangeretin reduced urine albumin excretion and improved foot process effacement of diabetic podocytes through inhibiting loss of slit junction and adherenes junction proteins. Glucose enhanced ROS production and HIF-1α induction in podocytes, leading to induction of oxidative stress and hypoxia. Similarly, in diabetic glomeruli reactive oxygen species (ROS) production and HIF-1α induction were observed. Furthermore, hypoxia-evoking cobalt chloride induced epithelial-to-mesenchymal transition (EMT) process and loss of slit diaphragm proteins and junction proteins in podocytes, which was inhibited by treating submicromolar tangeretin. Collectively, these results demonstrate that tangeretin inhibited podocyte injury and fibrosis through blocking podocyte EMT caused by glucose-induced oxidative stress and hypoxia.     

外源乙烯对黄瓜幼苗根系生长及内源多胺含量的影响

精确控制营养液溶氧浓度和外源乙烯浓度,研究了黄瓜幼苗根系在低氧水培时的生长情况以及PAs含量的变化。结果表明,外源乙烯对根际低氧逆境下黄瓜幼苗根系的生长有抑制作用,虽然根径增粗,含水量增加,但根系的总根数、总表面积、总体积、干重和鲜重都有所减少,随着乙烯浓度的增加,抑制作用增强;外源乙烯对根际低氧逆境下黄瓜幼苗根系PAs含量有着明显的拮抗作用,外源乙烯使根际低氧逆境下黄瓜幼苗根系的伤害症状加重。     

The Effect of Hypoxia on the Expression of CXC Chemokines and CXC Chemokine Receptors—A Review of Literature

Hypoxia is an integral component of the tumor microenvironment. Either as chronic or cycling hypoxia, it exerts a similar effect on cancer processes by activating hypoxia-inducible factor-1 (HIF-1) and nuclear factor (NF-κB), with cycling hypoxia showing a stronger proinflammatory influence. One of the systems affected by hypoxia is the CXC chemokine system. This paper reviews all available information on hypoxia-induced changes in the expression of all CXC chemokines (CXCL1, CXCL2, CXCL3, CXCL4, CXCL5, CXCL6, CXCL7, CXCL8 (IL-8), CXCL9, CXCL10, CXCL11, CXCL12 (SDF-1), CXCL13, CXCL14, CXCL15, CXCL16, CXCL17) as well as CXC chemokine receptors—CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, CXCR7 and CXCR8. First, we present basic information on the effect of these chemoattractant cytokines on cancer processes. We then discuss the effect of hypoxia-induced changes on CXC chemokine expression on the angiogenesis, lymphangiogenesis and recruitment of various cells to the tumor niche, including myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), tumor-associated neutrophils (TANs), regulatory T cells (T_regs) and tumor-infiltrating lymphocytes (TILs). Finally, the review summarizes data on the use of drugs targeting the CXC chemokine system in cancer therapies.     

Inhalational Anesthetics Inhibit Neuroglioma Cell Proliferation and Migration via miR-138, -210 and -335

Inhalational anesthetics was previously reported to suppress glioma cell malignancy but underlying mechanisms remain unclear. The present study aims to investigate the effects of sevoflurane and desflurane on glioma cell malignancy changes via microRNA (miRNA) modulation. The cultured H4 cells were exposed to 3.6% sevoflurane or 10.3% desflurane for 2 h. The miR-138, -210 and -335 expression were determined with qRT-PCR. Cell proliferation and migration were assessed with wound healing assay, Ki67 staining and cell count kit 8 (CCK8) assay with/without miR-138/-210/-335 inhibitor transfections. The miRNA downstream proteins, hypoxia inducible factor-1α (HIF-1α) and matrix metalloproteinase 9 (MMP9), were also determined with immunofluorescent staining. Sevoflurane and desflurane exposure to glioma cells inhibited their proliferation and migration. Sevoflurane exposure increased miR-210 expression whereas desflurane exposure upregulated both miR-138 and miR-335 expressions. The administration of inhibitor of miR-138, -210 or -335 inhibited the suppressing effects of sevoflurane or desflurane on cell proliferation and migration, in line with the HIF-1α and MMP9 expression changes. These data indicated that inhalational anesthetics, sevoflurane and desflurane, inhibited glioma cell malignancy via miRNAs upregulation and their downstream effectors, HIF-1α and MMP9, downregulation. The implication of the current study warrants further study.     

Real-Time Monitoring the Effect of Cytopathic Hypoxia on Retinal Pigment Epithelial Barrier Functionality Using Electric Cell-Substrate Impedance Sensing (ECIS) Biosensor Technology

Disruption of retinal pigment epithelial (RPE barrier integrity is a hallmark feature of various retinal blinding diseases, including diabetic macular edema and age-related macular degeneration, but the underlying causes and pathophysiology are not completely well-defined. One of the most conserved phenomena in biology is the progressive decline in mitochondrial function with aging leading to cytopathic hypoxia, where cells are unable to use oxygen for energy production. Therefore, this study aimed to thoroughly investigate the role of cytopathic hypoxia in compromising the barrier functionality of RPE cells. We used Electric Cell-Substrate Impedance Sensing (ECIS) system to monitor precisely in real time the barrier integrity of RPE cell line (ARPE-19) after treatment with various concentrations of cytopathic hypoxia-inducing agent, Cobalt(II) chloride (CoCl₂). We further investigated how the resistance across ARPE-19 cells changes across three separate parameters: R_b (the electrical resistance between ARPE-19 cells), α (the resistance between the ARPE-19 and its substrate), and C_m (the capacitance of the ARPE-19 cell membrane). The viability of the ARPE-19 cells and mitochondrial bioenergetics were quantified with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay and seahorse technology, respectively. ECIS measurement showed that CoCl₂ reduced the total impedance of ARPE-19 cells in a dose dependent manner across all tested frequencies. Specifically, the ECIS program’s modelling demonstrated that CoCl₂ affected R_b as it begins to drastically decrease earlier than α or C_m, although ARPE-19 cells’ viability was not compromised. Using seahorse technology, all three concentrations of CoCl₂ significantly impaired basal, maximal, and ATP-linked respirations of ARPE-19 cells but did not affect proton leak and non-mitochondrial bioenergetic. Concordantly, the expression of a major paracellular tight junction protein (ZO-1) was reduced significantly with CoCl_2-treatment in a dose-dependent manner. Our data demonstrate that the ARPE-19 cells have distinct dielectric properties in response to cytopathic hypoxia in which disruption of barrier integrity between ARPE-19 cells precedes any changes in cells’ viability, cell-substrate contacts, and cell membrane permeability. Such differences can be used in screening of selective agents that improve the assembly of RPE tight junction without compromising other RPE barrier parameters.     

Hypoxia/Temperature/pH Triple Stimuli–Responsive Block Copolymers: Synthesis,Self-Assembly,and Controlled Drug Release

The amphiphilic block copolymer poly(methacrylic acid-co-2-nitroimidazole acrylate)-b-poly(2-(N,N-dimethylamino)ethyl methacrylate) (P(MAA-co-NIMA)-b-PDMAEMA) with the hypoxia/temperature/pH triple responsiveness is synthesized by reversible addition-fragmentation chain transfer polymerization (RAFT), hydrolysis, and 3-(3-dimethylaminopropyl)-1-ethylcarbodiimide (EDC) reactions, and successfully self-assembled into micelles. The hypoxia response in vitro is realized, and then the sensitivity of the self-assembled micelles to the hypoxia condition is studied by controlling the grafting amount of aminated 2-nitroimidazole. Because 2-(N,N-dimethylamino) ethyl methacrylate (DMAEMA) is a typical material sensitive to temperature and pH conditions, the self-assembled micelles are also responsive to temperature and different acidic/basic conditions. In addition, the cumulative release rate of doxorubicin (DOX) at 42 °C, pH = 6.0, and hypoxic conditions increases significantly, and verifies the synergistic promotion effect of the above stimulations. This intelligent polymer with triple response mechanism improves the controllability and efficiency of drug release, and is expected to be a drug carrier for cancer treatment.     

Versatile Hypoxic Extracellular Vesicles Laden in an Injectable and Bioactive Hydrogel for Accelerated Bone Regeneration

Extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) have emerged as an appealing alternative to cell therapy in regenerative medicine. Unlike bone marrow MSCs (BMSCs) cultured in vitro with normoxia, bone marrow in vivo is exposed to a hypoxic environment. To date, it remains unclear whether hypoxia preconditioning can improve the function of BMSC-derived EVs and be more conducive to bone repair. Herein, it is found that hypoxia preconditioned BMSCs secrete more biglycan (Bgn)-rich EVs via proteomics analysis, and these hypoxic EVs (Hypo-EVs) significantly promote osteoblast proliferation, migration, differentiation, and mineralization by activating the phosphatidylinositide 3-kinase/protein kinase B pathway. Subsequently, an injectable bioactive hydrogel composed of poly(ethylene glycol)/polypeptide copolymers is developed to improve the stability and retention of Hypo-EVs in vivo. The Hypo-EVs-laden hydrogel shows continuous liberation of Hypo-EVs for 3 weeks and substantially accelerates bone regeneration in 5-mm rat cranial defects. Finally, it is confirmed that Bgn in EVs is a pivotal protein regulating osteoblast differentiation and mineralization and exerts its effects through paracrine mechanisms. Therefore, this study shows that hypoxia stimulation is an effective approach to optimize the therapeutic effects of BMSC-derived EVs and that injectable hydrogel-based EVs delivery is a promising strategy for tissue regeneration.