Search

Impaired insulin secretion is one of the main causes of type 2 diabetes. Cholesterol accumulation-induced lipotoxicity contributes to impaired insulin secretion in pancreatic beta cells. However, the detailed mechanism in this process remains unclear. In this study, we proved that oxidized low-density lipoprotein (OxLDL) reduced insulin content, decreased PDX-1 expression, and impaired glucose-stimulated insulin secretion (GSIS) in INS-1 cells, which were rescued by addition of high-density lipoprotein (HDL). OxLDL receptors and cholesterol content were increased by OxLDL. Consistently, OxLDL suppressed cholesterol transporter ABCA1 expression and transcription in a dose-dependent and time-dependent manner. Inhibition of MEK by its specific inhibitor, PD98059, altered the effect of OxLDL on ABCA1 transcription and activation of ERK. Next, chromatin immunoprecipitation assay demonstrated that liver X receptor (LXR) could directly bind to ABCA1 promoter and this binding was inhibited by OxLDL. Furthermore, OxLDL decreased the nuclear LXR expression, which was prevented by HDL. LXR-enhanced ABCA1 transcription was suppressed by OxLDL, and the effect was cancelled by mutation of the LXR-binding sites. In summary, our study shows that OxLDL down-regulates ABCA1 expression by MEK/ERK/LXR pathway, leading to cholesterol accumulation in INS-1 cells, which may result in impaired insulin synthesis and GSIS.

Effective treatment regimens for triple-negative breast cancer (TNBC) are relatively scarce due to a lack of specific therapeutic targets. Epidermal growth factor receptor (EGFR) signaling is highly active in TNBC and is associated with poor prognosis. Most EGFR antagonists, which significantly improve outcome in lung and colon cancer, have shown limited clinical effects in breast cancer. However, limiting EGFR expression in TNBC is a potential strategy for improving the clinical efficacy of EGFR antagonists. Here, we found that the gamma-aminobutyric acid type A receptor π subunit (GABRP), as a membrane protein enriched in TNBC stem cells, interacted with EGFR and significantly sustained its expression, resulting in stemness maintenance and chemotherapy resistance. Silencing GABRP induced down-regulation of EGFR signaling, which hindered cell stemness and enhanced sensitivity to chemotherapies, including paclitaxel, doxorubicin, and cisplatin. We also identified that retigabine, an FDA-approved drug for adjunctive treatment of seizures, increased the sensitivity of EGFR to gefitinib in gefitinib-resistant cells. Our findings show that GABRP can sustain the stemness of TNBC via modulating EGFR expression, suggesting that GABRP may be a potential therapeutic target that can address EGFR inhibitor resistance in TNBC., (Copyright © 2021 Elsevier B.V. All rights reserved.)

MicroRNAs (miRNAs) are short endogenous noncoding RNAs regulating protein translation. However, the specific mechanism by which miR-181b influences sepsis via high-mobility group box-1 protein (HMGB1) still remains unknown. Thus, the aim of this study is to investigate the mechanism of miR-181b in regulating inflammatory response in sepsis-induced myocardial injury through targeting high-mobility group box-1 protein (HMGB1). Through cecal ligation and puncture (CLP), the rat model of sepsis was established. Then, the effect of altered expression of miR-181b and HMGB1 on cardiomyocytes was investigated. The positive expression rate of HMGB1, concentration of inflammatory factors, and serum myocardial enzyme of myocardial tissues were determined. Besides, the binding site between miR-181b and HMGB1 was determined by bioinformatics information and dual-luciferase reporter gene assay. The expression of related genes in cells of each group was determined by RT-qPCR and western blot analysis, and the apoptosis rate of transfected cells in each group was determined by TUNEL assay. HMGB1 expression and inflammatory factors were significantly increased in myocardial tissue of rats with sepsis. Cell morphology and the infiltration of inflammatory cells were significantly improved by overexpression of miR-181b. miR-181b directly targeted HMGB1, and downregulation of HMGB1 reduced inflammatory factors and myocardial injury and inhibited cardiomyocyte apoptosis in sepsis. This present study suggests that miR-181b decreased inflammatory factors and reduced myocardial injury in sepsis through downregulation of HMGB1. Thus, a better understanding of this process may aid in the development of novel therapeutic agents in sepsis., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC part of Springer Nature.)

Catechin, a flavonol belonging to the flavonoid group of polyphenols is present in many plant foods. The present study was done to evaluate the effect of catechin on various inflammatory mediators using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. The effect of catechin on total cyclooxygenase (COX) activity, 5-lipoxygenase (5-LOX), myeloperoxidase, nitrite and inducible nitric oxide synthase (iNOS) level, secretion of tumor necrosis factor-α (TNF-α) and interleukin-10 (IL-10) were assessed in LPS-stimulated RAW 264.7 cells. The expression of COX-2, iNOS, TNF-α, nuclear factor-ĸB (NF-κB) and p38 mitogen-activated protein kinase (MAPK) genes were also investigated. The effect was further analyzed using human PBMCs by assessing the level of TNF-α and IL-10. The study demonstrated that the inflammatory mediators such as COX, 5-LOX, nitrite, iNOS, and TNF-α were significantly inhibited by catechin in a concentration-dependent manner whereas IL-10 production was up-regulated in RAW 264.7 cells. Moreover, catechin down-regulated the mRNA level expression of COX-2, iNOS, TNF-α, NF-κB and p38 MAPK. The current study ratifies the beneficial effect of catechin as a dietary component in plant foods to provide protection against inflammatory diseases., (© 2021. The Author(s), under exclusive licence to Springer Nature Switzerland AG.)

Background: Alpha 2A -adrenergic (α 2A -AR) and cannabinoid CB 1 (CB 1 -R) receptors exert their functions modulating multiple signaling pathways, including MEK-ERK (extracellular signal-regulated kinases) and FADD (Fas-associated protein with death domain) cascades. These molecules are relevant in finding biased agonists with fewer side effects, but the mechanisms involving their modulations by α 2A -AR- and CB 1 -R in vivo are unclear. This study investigated the roles of Gαi 2 and Gαz proteins in mediating α 2A -AR- and CB 1 -R-induced alterations of MEK-ERK and FADD phosphorylation (p-) in mouse brain cortex., Methods: Gαi 2 or Gαz protein knockdown was induced in mice with selective antisense oligodeoxinucleotides (ODNs; 3 nmol/day, 5 days) prior to UK-14,304 (UK or brimonidine; 1 mg/kg) or WIN55212-2 (WIN; 8 mg/kg) acute treatments. Inactivated (p-T 286 ) MEK1, activated (p-S 217/221 ) MEK1/2, activated (p-T 202 /Y 204 ) ERK1/2, p-S 191 FADD, and the corresponding total forms of these proteins were quantified by immunoblotting., Results: Increased (+ 88%) p-T 286 MEK1 cortical density, with a concomitant reduction (-43%) of activated ERK was observed in UK-treated mice. Both effects were attenuated by Gαi 2 or Gαz antisense ODNs. Contrastingly, WIN induced Gαi 2 - and Gαz-independent upregulations of p-T 286 MEK1 (+ 63%), p-S 217/221 MEK1/2 (+ 86%), and activated ERK (+ 111%) in brain. Pro-apoptotic FADD was downregulated (- 34 to 39%) following UK and WIN administration, whereas the neuroprotective p-S 191 FADD was increased (+ 74%) in WIN-treated mice only. None of these latter effects required from Gαi 2 or Gαz protein integrity., Conclusion: The results indicate that α 2A -AR (UK), but not CB 1 -R (WIN), agonists use Gαi 2 and Gαz proteins to modulate MEK-ERK, but not FADD, pathway in mouse brain cortex., (© 2021. Maj Institute of Pharmacology Polish Academy of Sciences.)

Colorectal cancer (CRC) is the second leading cause of death of malignant tumors worldwide. Recent studies point to a role for the adiponectin-receptor axis in colorectal carcinogenesis, and in particular to the oncosuppressive properties of the T-cadherin receptor. In addition, the loss of T-cadherin expression in tumor tissues has been linked to cancer progression and attributed to aberrant methylation of its promoter. Recognizing the pivotal role of microRNAs in CRC, this study explores their possible contribution to the downregulation of T-cadherin. A systematic bioinformatics analysis, restricted by microRNA expression data in the colon or in cultured colorectal cell lines, predicted twelve top-ranking target miRNA sites within the 3' UTR of T-cadherin. Experimental validation analyses based on luciferase reporter constructs and miRNA mimic or miRNA inhibitor transfections toward colorectal adenocarcinoma cell lines indicated that miR-377-3p was able to directly bind to the T-cadherin sequence, and thus downregulating its expression. Given the oncogenic activity of miR-377 and the oncosuppressive activity of T-cadherin in CRC, the regulatory circuit highlighted in this study may add new insights into molecular mechanisms driving colorectal carcinogenesis, and perspectively it could be exploited to identify novel biomarkers and therapeutic targets., (© 2021 The Authors. Cell Biology International published by John Wiley & Sons Ltd on behalf of International Federation of Cell Biology.)

Pharmacological reactivation of developmentally silenced fetal hemoglobin (HbF) is an attractive approach to ameliorate the clinical manifestations of β-thalassemia and sickle cell anemia. Hydroxyurea, the only HbF inducer, has obtained regulatory approval. However, hydroxyurea non-responders and associated myelosuppression making its widespread use undesirable. A high level of HbF with safe and effective agents remains an elusive therapeutic goal for this global health burden. This study demonstrated the effect of acyclovir on γ-globin expression and erythropoiesis, associated with increased HbF production. In vitro, human erythroleukemia cells and human CD34 + erythroid progenitors, and in vivo β-YAC transgenic mice were used as experimental models. We found that acyclovir significantly induces expression of the γ-globin gene and HbF synthesis in CD34 + erythroid progenitors, without affecting terminal erythroid differentiation and erythroid cell proliferation. In contrast to other HbF inducers, no associated cytotoxicity with acyclovir was observed. Further, we reported the effect of acyclovir on γ-globin gene transcriptional regulators including BCL11A, FOP1, KLF1 SOX6, and GATA-1. Significant downregulation of the γ-globin repressors BCL11A and SOX6 was observed at both mRNA and protein levels. Whereas, GATA-1, a master erythroid transcription factor, was upregulated in acyclovir treated human CD34 + erythroid culture. Similarly, the HbF inducing effect of acyclovir in β-YAC transgenic mice revealed a good in vitro correlation, with a substantial increase in fetal globin mRNA, and F cells population. These findings collectively suggest acyclovir as an effective HbF inducer and pave the way to evaluate its clinical efficacy in treating β-globin disorders., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The imbalance between excess reactive oxygen species (ROS) generation and insufficient antioxidant defenses contribute to a range of neurodegenerative diseases. High ROS levels damage cellular macromolecules such as DNA, proteins and lipids, leading to neuron vulnerability and eventual death. However, the underlying molecular mechanism of the ROS regulation is not fully elucidated. Recently, an increasing number of studies suggest that microRNAs (miRNAs) emerge as the targets in regulating oxidative stress. We recently reported the neuroprotective effect of miR-137-3p for brachial plexus avulsion-induced motoneuron death. The present study is sought to investigate whether miR-137-3p also could protect PC12 cells against hydrogen peroxide (H 2 O 2 ) induced neurotoxicity. By using cell viability assay, ROS assay, gene and protein expression assay, we found that PC-12 cells exposed to H 2 O 2 exhibited decreased cell viability, increased expression levels of calpain-2 and neuronal nitric oxide synthase (nNOS), whereas a decreased miR-137-3p expression. Importantly, restoring the miR-137-3p levels in H 2 O 2 exposure robustly inhibited the elevated nNOS, calpain-2 and ROS expression levels, which subsequently improved the cell viability. Furthermore, the suppressive effect of miR-137-3p on the elevated ROS level under oxidative stress was considerably blunted when we mutated the binding site of calpain-2 targted by miR-137-3p, suggesting the critical role of calpain-2 involving the neuroprotective effect of miR-137-3p. Collectively, these findings highlight the neuroprotective role of miR-137-3p through down-regulating calpain and NOS activity, suggesting its potential role for combating oxidative stress insults in the neurodegenerative diseases.

The underlying mechanism of electroacupuncture (EA) in relieving obesity, anti-inflammation and the interaction with metabolic pathways in obese mice has not been elaborated. The aim of this study was to investigate the regulation of EA on macrophage polarization in obesity tissue of diet-induced obesity mice. Mice were divided in 6 groups: normal control group, model group, EA-7 group, EA-14 group, EA-21 group and EA-28 group. Low-frequency EA was applied at "Tianshu (ST 25)", "Guanyuan (CV 4)", "Zusanli (ST 36)" and "Sanyinjiao (SP 6)" for 10 min. Adipose tissue was assessed with hematoxylin and eosin staining. Adipocytokines and pro-inflammatory factors expression was measured by ELISA. The protein and mRNA levels of macrophage markers were examined by immumohistochemical staining and RT-PCR, respectively. EA treatment was associated with a decrease of adipose tissue and large adipocytes, and an increase of small adipocytes. After EA treatment, the levels of Leptin, Chemerin, TNF-α, F4/80, iNOS, and CD11c decreased obviously in adipose tissue, while IL-4, IL-10 and CD206 levels increased significantly. Besides, TNF-α in spleen tissue was also downregulated, but IL-4 and IL-10 were upregulated. EA prevents weight gain through modulation inflammatory response and macrophage polarization in obese adipose tissues., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Glioblastoma (GBM) has high mortality rates because of extreme therapeutic resistance. During surgical resection for GBM, 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PpIX) fluorescence is conventionally applied to distinguish GBM. However, surgical intervention is insufficient for high invasive GBM. Sonodynamic therapy (SDT) combined with low-intensity ultrasonication (US) and PpIX, as a sonosensitizer, is an emerging and promising approach, although its efficacy is limited. Based on our previous study that down-regulation of multidrug resistant protein (MDR1) in GBM augmented the anti-tumor effects of chemotherapy, we hypothesized that elevation of cellular PpIX levels by down-regulation of MDR1 enhances anti-tumor effects by SDT. In high invasive progeny cells from mouse glioma stem cells (GSCs) and a GSC-bearing mouse glioma model, we assessed the anti-tumor effects of SDT with a COX-2 inhibitor, celecoxib. Down-regulation of MDR1 by celecoxib increased cellular PpIX levels, as well as valspodar, an MDR1 inhibitor, and augmented anti-tumor effects of SDT. MDR1 down-regulation via the Akt/NF-κB pathway by celecoxib was confirmed, using an NF-κB inhibitor, CAPÉ. Thus, elevation of cellar PpIX by down-regulation of MDR1 via the Akt/NF-κB pathway may be crucial to potentiate the efficacy of SDT in a site-directed manner and provide a promising new therapeutic strategy for GBM., (© 2021. The Author(s).)

The expression of transporters on the apical and basal membranes of renal proximal tubular cells are down- or upregulated to various extents under cisplatin (CDDP)-induced acute kidney injury (AKI). However, little is known about the changes in transporters in tissues other than the kidney under CDDP-induced AKI. This study aimed to investigate the modulation of the expression/function of intestinal efflux transporters, P-glycoprotein (P-gp) and breast cancer resistance protein (Bcrp), in CDDP-induced AKI rats. On day 3 after the intraperitoneal administration of CDDP (5 mg/kg) to rats, the expression levels of P-gp and Bcrp were compared with those of normal rats. Further, the absorption of three P-gp substrates (6α-methylprednisolone, rhodamine 123, and gatifloxacin) was evaluated in both groups using conventional loop techniques. In the CDDP-induced AKI rats, P-gp expression in the ileum was markedly decreased to approximately 38% of that in the normal rats. However, no significant changes in Bcrp expression were observed in the AKI rats. In contrast with the reduction in P-gp expression in the AKI rats, the absorption of the three P-gp substrates remained almost the same or decreased in the AKI group. The addition of verapamil (a potent P-gp inhibitor) increased the absorption of the three P-gp substrates to the values obtained from the normal rats. In conclusion, our results suggested that P-gp expression is downregulated in rats with CDDP-induced AKI but that P-gp maintains its potency as a "gatekeeper" against the absorption of xenobiotics by amplifying its individual transport capacity under these conditions., (Copyright © 2021. Published by Elsevier Inc.)

In mice, a subset of cardiac macrophages and Kupffer cells derive from fetal precursors, seed the developing tissues, self-renew locally, and persist into adulthood. In this study we investigated how these cells survive acute systemic inflammation. In both tissues, early-derived subsets rapidly responded to acute systemic inflammation by assuming a temporary nonclassical activation state featuring upregulation of both proinflammatory ( Il1b, Tnf, Nfkb1 ), and anti-inflammatory ( Il10, Il4ra, Nfkbiz ) genes. During this process, transcription factor genes associated with myeloid identity ( Spi1 , Zeb2 ) were upregulated, whereas those associated with tissue specificity ( Nr1h3 for Kupffer cells and Nfatc2 and Irf4 for cardiac macrophages) were downregulated, suggesting that the cells reasserted their myeloid identity but renounced their tissue identity. Most of these changes in gene expression reverted to steady-state levels postresolution. We conclude that these early-derived macrophage subsets are resilient in the face of acute stress by temporary loss of adaptation to local tissue-specific niches while reasserting their generic myeloid identity., (Copyright © 2021 by The American Association of Immunologists, Inc.)

SIGIRR has been described as a negative regulator of several IL-1R/TLR family members and has been implicated in several inflammatory disease conditions. However, it is unknown whether it can suppress IL-36 family cytokines, which are members of the broader IL-1 superfamily that have emerged as critical orchestrators of psoriatic inflammation in both humans and mice. In this study, we demonstrate that SIGIRR is downregulated in psoriatic lesions in humans and mice, and this correlates with increased expression of IL-36 family cytokines. Using Sigirr -/- mice, we identify, for the first time (to our knowledge), SIGIRR as a negative regulator of IL-36 responses in the skin. Mechanistically, we identify dendritic cells and keratinocytes as the primary cell subsets in which IL-36 proinflammatory responses are regulated by SIGIRR. Both cell types displayed elevated IL-36 responsiveness in absence of SIGIRR activity, characterized by enhanced expression of neutrophil chemoattractants, leading to increased neutrophil infiltration to the inflamed skin. Blockade of IL-36R signaling ameliorated exacerbated psoriasiform inflammation in Sigirr -/- mice and inhibited neutrophil infiltration. These data identify SIGIRR activity as an important regulatory node in suppressing IL-36-dependent psoriatic inflammation in humans and mice., (Copyright © 2021 by The American Association of Immunologists, Inc.)

Valproic acid (VPA) is a powerful antiepileptic drug that was associated with several neurological and hepatic problems especially with increasing its dose and duration. These problems may be metabolic in origin and related to glucose homeostasis. So, the present study investigated the effect of different doses and durations of VPA on the expression of glucose transporters (Glut1 and Glut4), oxidative stress and inflammatory cytokine (IL-6) in the liver and specific brain regions. Seventy-two male Sprague-Dawley rats were randomly allocated into three equal groups: (1) saline group, (2) 200 mg VPA group and (3) 400 mg VPA group. By the end of experiments, the expressions of Glut1, Glut4 nuclear factor erythroid-like 2 related factor (Nrf2), IL-6 and oxidative stress markers [malondialdehyde (MDA) and glutathione (GSH)] in the liver, corpus striatum, prefrontal cortex (PFC) and cerebellum were assessed. We found that administration of VPA (200 mg and 400 mg) caused a significant decrease in the Glut1 and Glut4 expression in different tissues in a dose- and time-dependent manner (P < 0.01). Also, VPA (200 and 400 mg) caused a significant increase in MDA with a decrease in GSH in tissues at different times. Moreover, VPA (200 and 400 mg) caused significant upregulation in IL-6 expression and downregulation in Nrf2 expression (P < 0.01). The results suggest that increasing the dose and time of VPA therapy downregulates Glut1 and Glut4 in the liver and brain which may impair glucose uptake in these tissues. This effect was associated with enhanced oxidative stress, downregulation of nrf2 and upregulation of IL-6 in liver and brain tissues., (Copyright © 2021 Wolters Kluwer Health, Inc. All rights reserved.)

Platelets can modulate cancer through budding of platelet microparticles (PMPs) that can transfer a plethora of bioactive molecules to cancer cells upon internalization. In acute myelogenous leukemia (AML) this can induce chemoresistance, partially through a decrease in cell activity. Here we investigated if the internalization of PMPs protected the monocytic AML cell line, THP-1, from apoptosis by decreasing the initial cellular damage inflicted by treatment with daunorubicin, or via direct modulation of the apoptotic response. We examined whether PMPs could protect against apoptosis after treatment with a selection of inducers, primarily associated with either the intrinsic or the extrinsic apoptotic pathway, and protection was restricted to the agents targeting intrinsic apoptosis. Furthermore, levels of daunorubicin-induced DNA damage, assessed by measuring gH2AX, were reduced in both 2N and 4N cells after PMP co-incubation. Measuring different BCL2-family proteins before and after treatment with daunorubicin revealed that PMPs downregulated the pro-apoptotic PUMA protein. Thus, our findings indicated that PMPs may protect AML cells against apoptosis by reducing DNA damage both dependent and independent of cell cycle phase, and via direct modulation of the intrinsic apoptotic pathway by downregulating PUMA. These findings further support the clinical relevance of platelets and PMPs in AML.

Background: The gonadotropin-releasing hormone (GnRH) antagonist protocol for in vitro fertilization (IVF) often leads to lower pregnancy rates compared to the GnRH agonist protocol. Decreased endometrial receptivity is one reason for the lower success rate, but the mechanisms underlying this phenomenon remain poorly understood. The S100 calcium protein P (S100P) is a biomarker for endometrial receptivity. Both GnRH antagonist and S100P are involved in mediating cell apoptosis. However, the involvement of S100P in reduced endometrial receptivity during the GnRH antagonist protocol remains unclear., Methods: Endometrial tissue was collected at the time of implantation window from patients undergoing the GnRH agonist (GnRH-a) or GnRH antagonist (GnRH-ant) protocols, as well as from patients on their natural cycles. Endometrial cell apoptosis and expression levels of S100P, HOXA10, Bax, and Bcl-2 were assessed. Ishikawa cells were cultured to evaluate the effects that GnRH antagonist exposure or S100P up- or down- regulation had on apoptosis., Results: Endometrial tissue from patients in the GnRH-ant group showed elevated apoptosis and decreased expression of the anti-apoptotic marker Bcl-2. In addition, endometrial expression of S100P was significantly reduced in the GnRH-ant group, and expression of HOXA10 was lower. Immunofluorescence colocalization analysis revealed that S100P was mainly distributed in the epithelium. In vitro experiments showed that knockdown of S100P in Ishikawa cells induced apoptosis, decreased expression of Bcl-2, while overexpression of S100P caused the opposite effects and decreased expression of Bax. Furthermore, endometrial epithelial cells exposed to GnRH antagonist expressed lower levels of S100P and Bcl-2, increased expression of Bax, and had higher rates of apoptosis. The increased apoptosis induced by GnRH antagonist treatment could be rescued by overexpression of S100P., Conclusions: We found that GnRH antagonist treatment induced endometrial epithelial cell apoptosis by down-regulating S100P, which was detrimental to endometrial receptivity. These results further define a mechanistic role for S100P in contributing to endometrial apoptosis during GnRH antagonist treatment, and suggest that S100P is a potential clinical target to improve the success of IVF using the GnRH antagonist protocol.

The amygdala plays an important role in the integration of responses to noxious and fearful stimuli. Sensory information from many systems is integrated in the lateral and basolateral amygdala and transmitted to the central amygdala, the major output nucleus of the amygdala regulating both motor and emotional responses. The network of intercalated cells (ITC) which surrounds the lateral and basolateral amygdala and serves to modulate information flow from the lateral amygdala to the central nucleus, express a very high local concentration of mu-type opioid receptors. Loss of the ITC neurons impairs fear extinction. We demonstrate here that exposure of rats to a severe stress experience resulted in a marked downregulation of the level of expression of mu opioid receptors in the ITC nuclei over a period of at least 24 h after the end of the stress exposure. The endogenous opioid dynorphin is also expressed in the central and ITC nuclei of the amygdala. Following stress exposure, we also observed an increase in the expression in the more lateral regions of the central amygdala of pro-dynorphin mRNA and a peptide product of pro-dynorphin with known affinity for mu opioid receptors. It is possible that the downregulation of mu receptors in ITC neurons after stress may result from sustained activation and internalization of mu receptors following a stress-induced increase in the release of endogenous opioid peptides.

Increasing numbers of patients with spontaneous subarachnoid hemorrhage(SAH) who recover from surgery and intensive care management still live with cognitive impairment after discharge, indicating the importance of white matter injury at the acute stage of SAH. In the present study, standard endovascular perforation was employed to establish an SAH mouse model, and a microRNA (miRNA) chip was used to analyze the changes in gene expression in white matter tissue after SAH. The data indicate that 17 miRNAs were downregulated, including miR-706, miR-669a-5p, miR-669p-5p, miR-7116-5p and miR-195a-3p, while 13 miRNAs were upregulated, including miR-6907-5p, miR-5135, miR-6982-5p, miR-668-5p, miR-8119. Strikingly, miR-706 was significantly downregulated with the highest fold change. Further experiments confirmed that miR-706 could alleviate white matter injury and improve neurological behavior, at least partially by inhibiting the PKCα/MST1/NF-κB pathway and the release of inflammatory cytokines. These results might provide a deeper understanding of the pathophysiological processes in white matter after SAH, as well as potential therapeutic strategies for the translational research., (Copyright © 2021 Elsevier Inc. All rights reserved.)

LINGO-1 may be involved in the pathogenesis of cerebral ischemia. However, its biological function and underlying molecular mechanism in cerebral ischemia remain to be further defined. In our study, middle cerebral artery occlusion/reperfusion (MACO/R) mice model and HT22 cell oxygen-glucose deprivation/reperfusion (OGD/R) were established to simulate the pathological process of cerebral ischemia in vivo and in vitro and to detect the relevant mechanism. We found that LINGO-1 mRNA and protein were upregulated in mice and cell models. Down-regulation LINGO-1 improved the neurological symptoms and reduced pathological changes and the infarct size of the mice after MACO/R. In addition, LINGO-1 interference alleviated apoptosis and promoted cell proliferation in HT22 of OGD/R. Moreover, down-regulation of LINGO-1 proved to inhibit nuclear translocation of p-NF-κB and reduce the expression level of p-JAK2 and p-STAT3. In conclusion, our data suggest that shLINGO-1 attenuated ischemic injury by negatively regulating NF-KB and JAK2/STAT3 pathways, highlighting a novel therapeutic target for ischemic stroke.

Histone deacetylase (HDAC) inhibitors can protect the brain from ischemic injury. This study aimed to identify the regulation of HDAC3 in cerebral ischemic injury. Middle cerebral artery occlusion (MCAO) was performed to establish a mouse model with cerebral ischemic injury, in which expression of HDAC3 and miR-19a was evaluated using RT-qPCR. In MCAO mice with silencing of HDAC3, infarct volume was determined using 2,3,5-triphenyl tetrazolium chloride (TTC) staining, and serum levels of TNF-α, IL-6, and IL-8 were measured using ELISA. An in vitro model was constructed in human umbilical vein endothelial cells (HUVECs) with oxygen-glucose deprivation/reoxygenation (OGD/R), followed by gain- and loss-of-function experiments. Relationships among miR-19a, HDAC3, and syndecan-1 (SDC1) were explored using RIP, ChIP, and dual-luciferase reporter assays. The expression of HDAC3, SDC1, JAK1, and STAT3 along with the extent of JAK1 and STAT3 phosphorylation was measured by Western blot analysis. HUVEC viability, apoptosis, and angiogenesis were assessed by CCK-8, flow cytometry, and angiogenesis assays in vitro separately. We found elevated HDAC3 and downregulated miR-19a expression in the MCAO mice. Decreased TNF-α, IL-6, and IL-8 serum levels were observed in response to silencing of HDAC3. HDAC3 inhibited the expression of miR-19a, which in turn targeted SDC1, leading to JAK1/STAT3 signaling pathway activation. HDAC3 overexpression or miR-19a inhibition repressed HUVEC viability and angiogenesis but enhanced HUVEC apoptosis. Our data unraveled the mechanism whereby HDAC3 inhibition ameliorated cerebral ischemic injury by activating the JAK1/STAT3 signaling pathway through miR-19a-mediated SDC1 inhibition.

This study was designed to investigate the role of miR-671-5p in in vitro and in vivo models of ischemic stroke (IS). Middle cerebral artery occlusion and reperfusion (MCAO/R) in C57BL/6 mice as well as oxygen-glucose deprivation and reoxygenation (OGD/R) in a mouse hippocampal HT22 neuron line were used as in vivo and in vitro models of IS injury, respectively. miR-671-5p agomir, miR-671-5p antagomir, pcDNA3.1-NF-κB, and negative controls were transfected into cells using riboFECT CP reagent. miR-671-5p agomir, pcDNA3.1-NF-κB, and negative vectors were administered into MCAO/R mice via intracerebroventricular injection. The results showed that miR-671-5p was significantly downregulated and that miR-671-5p agomir alleviated injury and neuroinflammation induced by ischemic reperfusion. A dual-luciferase reporter assay confirmed that NF-κB is a direct target of miR-671-5p. Reverse experiments showed that miR-671-5p agomir reduced neuroinflammation via suppression of NF-κB expression in both in vitro and in vivo models of IS. Our data suggest that miR-671-5p may be a viable therapeutic target for diminishing neuroinflammation in patients with IS.

Hepatic fibrosis is a disease characterized by excessive deposition of extracellular matrix (ECM) in the liver. Activation of hepatic stellate cells (HSCs) is responsible for most of ECM production. Oxidative stress and reactive oxygen species (ROS) may be important factors leading to liver fibrosis. NADPH oxidase 4 (NOX4) is the main source of ROS in hepatic fibrosis, but the mechanism by which NOX4 regulates oxidative stress is not fully understood. β-Arrestin2 is a multifunctional scaffold protein that regulates receptor endocytosis, signaling and trafficking. In this study, we investigated whether β-arrestin2 regulated oxidative stress in hepatic fibrosis. Both β-arrestin2 knockout (Arrb2 KO) mice and wild-type mice were intraperitoneally injected with carbon tetrachloride (CCl 4 ) to induce hepatic fibrosis. Arrb2 KO mice showed significantly attenuated liver fibrosis, decreased ROS levels and NOX4 expression, and reduced collagen levels in their livers. In vitro, NOX4 knockdown significantly inhibited ROS production, and decreased expression of alpha-smooth muscle actin in angiotensin II-stimulated human HSC cell line LX-2. Through overexpression or depletion of β-arrestin2 in LX-2 cells, we revealed that decreased β-arrestin2 inhibited ROS levels and NOX4 expression, and reduced collagen production; it also inhibited activation of ERK and JNK signaling pathways. These results demonstrate that β-arrestin2 deficiency protects against liver fibrosis by downregulating ROS production through NOX4. This effect appears to be mediated by ERK and JNK signaling pathways. Thus, targeted inhibition of β-arrestin2 might reduce oxidative stress and inhibit the progression of liver fibrosis.

Neuromuscular junctions (NMJ) regulate cholinergic exocytosis through the M 1 and M 2 muscarinic acetylcholine autoreceptors (mAChR), involving the crosstalk between receptors and downstream pathways. Protein kinase C (PKC) regulates neurotransmission but how it associates with the mAChRs remains unknown. Here, we investigate whether mAChRs recruit the classical PKCβI and the novel PKCε isoforms and modulate their priming by PDK1, translocation and activity on neurosecretion targets. We show that each M 1 and M 2 mAChR activates the master kinase PDK1 and promotes a particular priming of the presynaptic PKCβI and ε isoforms. M 1 recruits both primed-PKCs to the membrane and promotes Munc18-1, SNAP-25, and MARCKS phosphorylation. In contrast, M 2 downregulates PKCε through a PKA-dependent pathway, which inhibits Munc18-1 synthesis and PKC phosphorylation. In summary, our results discover a co-dependent balance between muscarinic autoreceptors which orchestrates the presynaptic PKC and their action on ACh release SNARE-SM mechanism. Altogether, this molecular signaling explains previous functional studies at the NMJ and guide toward potential therapeutic targets., (©2021 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Peripheral neuropathy is the most frequent dose-limiting adverse effect of oxaliplatin. Acute pain symptoms that are induced or exacerbated by cold occur in almost all patients immediately following the first infusions. Evidence has shown that oxaliplatin causes ion channel expression modulations in dorsal root ganglia neurons, which are thought to contribute to peripheral hypersensitivity. Most dysregulated genes encode ion channels involved in cold and mechanical perception, noteworthy members of a sub-group of potassium channels of the K2P family, TREK and TRAAK. Downregulation of these K2P channels has been identified as an important tuner of acute oxaliplatin-induced hypersensitivity. We investigated the molecular mechanisms underlying this peripheral dysregulation in a murine model of neuropathic pain triggered by a single oxaliplatin administration. We found that oxaliplatin-mediated TREK-TRAAK downregulation, as well as downregulation of other K + channels of the K2P and Kv families, involves a transcription factor known as the neuron-restrictive silencer factor (NRSF) and its epigenetic co-repressors histone deacetylases (HDACs). NRSF knockdown was able to prevent most of these K + channel mRNA downregulation in mice dorsal root ganglion neurons as well as oxaliplatin-induced acute cold and mechanical hypersensitivity. Interestingly, pharmacological inhibition of class I HDAC reproduces the antinociceptive effects of NRSF knockdown and leads to an increased K + channel expression in oxaliplatin-treated mice.

Podocyte injury is a major determinant of focal segmental glomerular sclerosis (FSGS) and the identification of potential therapeutic targets for preventing podocyte injury has clinical importance for the treatment of FSGS. CLEC14A is a single-pass transmembrane glycoprotein belonging to the vascular expressed C-type lectin family. CLEC14A is found to be expressed in vascular endothelial cells during embryogenesis and is also implicated in tumor angiogenesis. However, the current understanding of the biological functions of CLEC14A in podocyte is very limited. In this study, we found that CLEC14A was expressed in podocyte and protected against podocyte injury in mice with Adriamycin (ADR)-induced FSGS. First, we observed that CLEC14A was downregulated in mice with ADR nephropathy and renal biopsies from individuals with FSGS and other forms of podocytopathies. Moreover, CLEC14A deficiency exacerbated podocyte injury and proteinuria in mice with ADR nephropathy accompanied by enhanced inflammatory cell infiltration and inflammatory responses. In vitro, overexpression of CLEC14A in podocyte had pleiotropic protective actions, including anti-inflammatory and anti-apoptosis effects. Mechanistically, CLEC14A inhibited high-mobility group box 1 protein (HMGB1) release, at least in part by directly binding HMGB1, and suppressed HMGB1-mediated signaling, including NF-κB signaling and early growth response protein 1 (EGR1) signaling. Taken together, our findings provide new insights into the pivotal role of CLEC14A in maintaining podocyte function, indicating that CLEC14A may be an innovative therapeutic target in FSGS., (© 2021 Federation of American Societies for Experimental Biology.)

The therapeutic effect of gemcitabine (GEM) in pancreatic ductal adenocarcinoma (PDAC) is limited due to low drug sensitivity and high drug resistance. Tissue inhibitor of matrix metalloprotease 1 (TIMP1) is reportedly associated with GEM resistance in PDAC. However, the effect of TIMP1 down-regulation in combination with GEM treatment is unknown. We analyzed the expression of TIMP1 in human PDAC tissue using western blot, quantitative real-time polymerase chain reaction (qRT-PCR), and immunohistochemistry. TIMP1 was highly expressed in PDAC specimens. Kaplan-Meier survival analysis suggested that a higher level of TIMP1 was correlated with poorer overall survival in 103 PDAC patients. The mRNA and protein expression profiles of TIMP1 were explored in the HTERT-HPNE human pancreatic ductal epithelium cell line, five PDAC cell lines (MIA PaCa-2, PANC-1, BxPC-3, Capan2, and SW1990), and two GEM-resistant PDAC cell lines (MIA PaCa-2 R and PANC-1 R ). Compared with HTERT-HPNE, TIMP1 was highly expressed in the PDAC cell lines. In addition, TIMP1 was upregulated in GEM-resistant PDAC cell lines compared with their parental cells. When TIMP1 was knocked-down using short hairpin RNA, GEM-induced cytotoxicity and apoptosis were increased, while colony formation was repressed in MIA PaCa-2, PANC-1, and their GEM-resistant cells. When Bax was activated by BAM7 or Bcl-2 was inhibited by venetoclax, CCK-8 assays demonstrated that GEM sensitivity was restored in GEM-resistant cells. When Bax was down-regulated by siRNA, CCK-8 assays verified that GEM sensitivity was decreased in PDAC cells. The observations that TIMP1 knockdown enhanced GEM sensitivity and reversed chemoresistance by inducing cells apoptosis indicated cooperative antitumor effects of shTIMP1 and GEM therapy on PDAC cells. The combination may be a potential strategy for PDAC therapy., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Background/aim: Heparanase (HPSE) is relevant to therapy resistance in many malignancies yet is largely unstudied in Hodgkin's lymphoma. Here, we investigated links between HPSE, cancer stem cell (CSC) features and radioresistance in KM-H2 and L428 Hodgkin's lymphoma cells., Materials and Methods: Firstly, HPSE expression in unsorted and sorted CSCs was assessed. Post-irradiation, HPSE and CSC-related gene expression changes were then quantified. Clonogenic ability was investigated with and without artificial changes in HPSE expression pre and post irradiation., Results: HPSE was highly expressed in L428 but barely present in KM-H2 cells. HPSE was overexpressed in sorted L428 CSCs. Irradiation induced HPSE and expression of CSC markers. High HPSE-expressing L428 cells showed higher clonogenic ability than low HPSE-expressing KM-H2 cells after irradiation. Down-regulation of HPSE in L428 cells reduced their clonogenic capability post-radiation, whilst overexpression of HPSE in KM-H2 cells increased colony formation., Conclusion: HPSE expression is associated with CSC features and contributes to radioresistance in Hodgkin's lymphoma cells., (Copyright © 2021 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Tripartite motif protein 21 (TRIM21) is an E3 ubiquitin ligase and cytosolic antibody receptor of the TRIM family. Previous reports have indicated that TRIM21 plays an important role during viral infection. This study aimed at examining the role of TRIM21 in the replication of porcine epidemic diarrhea virus (PEDV) and showed that TRIM21 inhibits PEDV proliferation by targeting and degrading the nucleocapsid (N) protein through the proteasomal pathway. Furthermore, the endogenous expression of TRIM21 was found to be downregulated by PEDV infection in Vero and LLC-PK1 cells. Overexpression of TRIM21 inhibited PEDV replication, whereas knockdown of TRIM21 increased viral titers and N protein levels. TRIM21 was found to interact and colocalize with the N protein, and the TRIM21-mediated antiviral effect was dependent on its ubiquitin ligase activity, which engages in polyubiquitination and degradation of the N protein in a proteasome-dependent manner. Taken together, these findings provide information about the role of TRIM21 in PEDV proliferation and increase our understanding of host-virus interactions.

Iron release from macrophages is closely regulated by the interaction of hepcidin, a peptide hormone produced by hepatocytes, with the macrophage iron exporter ferroportin (FPN1). However, the functions of FPN1 in hepatocyte secretion and macrophage polarization remain unknown. CD68 immunohistochemical staining and double immunofluorescence staining for F4/80 and Ki67 in transgenic mouse livers showed that the number of macrophages in FPN1 -/+ and FPN1 -/- mouse livers was significantly increased compared to that in WT (FPN +/+ ) mice. FPN1 downregulation in hepatic cells increased the levels of the M2 markers CD206, TGF- β, VEGF, MMP-9, Laminin, Collagen, IL-4 and IL-10. Furthermore, the expression of CD16/32 and iNOS, as M1 markers, exhibited the opposite trend. Meanwhile, α-SMA immunohistochemistry and Sirius red staining showed that the trend of liver fibrosis in FPN1 -/- mice was more significant than that in control mice. Similarly, in vitro FPN1 knockdown in L02-Sh/L02-SCR liver cell lines yielded similar results. Taken together, we demonstrated that downregulated FPN1 expression in hepatocytes can promote the proliferation and polarization of macrophages, leading to hepatic fibrosis. Above all, the FPN1 axis might provide a potential target for hepatic fibrosis.

We have addressed in the current study the potential of L-carnitine (LC) to extenuate the reproductive toxic insults of carbendazim (CBZ) in male rats, and the molecular mechanisms whereby carnitine would modify the spermatogenic and steroidogenic derangements invoked by the endocrine disruptor. Herein, animals received daily doses of carbendazim (100 mg/kg) by gavage for 8 weeks. Another CBZ-challenged group was co-supplemented with LC (500 mg/kg, IP) twice weekly for 8 weeks. Sperm quantity and quality (morphology, motility and viability), serum testosterone and gonadotropins, and thyroid hormone levels were assessed. Serum tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and interleukin-10 (IL-10) concentrations were determined by ELISA. Oxidant/antioxidant status in rat testis was investigated via measuring testicular contents of malondialdehyde (MDA) and reduced glutathione (GSH), as well as the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPx). Immunohistochemical localizations of the junctional protein; occludin, and inflammatory markers; inducible nitric oxide synthase (iNOS) and nuclear factor kappa beta (NF-κB) were further analyzed. A host of transduction genes that regulate spermatogenic and steroidogenic pathways, and their encoded proteins namely, Steroidogenic Acute Regulatory Protein (StAR), Fatty acid binding protein 9 (FABP9) and P38-mitogen activated protein kinase (P38-MAPK) were assessed by real time quantitative (RT-qPCR) and Western blot. LC improved rat spermiogram, testicular histological alterations and endocrine perturbances, and modulated genes' expressions and their respective proteins. In conclusion, LC effects appear to reside for the most part on its endocrine-preserving, anti-oxidant and anti-inflammatory properties through a myriad of interlaced signal transductions that ultimately recapitulated its beneficial effects on spermatogenesis and steroidogenesis., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Background: Recurrent miscarriage (RM) is a very frustrating problem for both couples and clinicians. To date, the etiology of RM remains poorly understood. Decidualization plays a critical role in implantation and the maintenance of pregnancy, and its deficiency is closely correlated with RM. The F-box protein S-phase kinase associated protein 2 (SKP2) is a key component of the SCF-type E3 ubiquitin ligase complex, which is critically involved in ErbB family-induced Akt ubiquitination, aerobic glycolysis and tumorigenesis. SKP2 is pivotal for reproduction, and SKP2-deficient mice show impaired ovarian development and reduced fertility., Methods: Here, we investigated the expression and function of SKP2 in human decidualization and its relation with RM. A total of 40 decidual samples were collected. Quantitative PCR analysis, western blot analysis and immunohistochemistry analysis were performed to analyze the differential expression of SKP2 between RM and control cells. For in vitro induction of decidualization, both HESCs (human endometrial stromal cells) cell line and primary ESCs (endometrial stromal cells) were used to analyze the effects of SKP2 on decidualization via siRNA transfection., Results: Compared to normal pregnant women, the expression of SKP2 was reduced in the decidual tissues from individuals with RM. After in vitro induction of decidualization, knockdown of SKP2 apparently attenuated the decidualization of HESCs and resulted in the downregulation of HOXA10 and FOXM1, which are essential for normal human decidualization. Moreover, our experiments demonstrated that SKP2 silencing reduced the expression of its downstream target GLUT1., Conclusions: Our study indicates a functional role of SKP2 in RM: downregulation of SKP2 in RM leads to impaired decidualization and downregulation of GLUT1 and consequently predisposes individuals to RM.

Objective: In the present study, the regulation mechanism of NAA10 (N-Alpha-Acetyltransferase 10) in sevoflurane preconditioning induced neuroprotective effect was explored., Methods: Firstly, si-NAA10 or negative control (NC) were constructed for cell transfection and injected into intracerebroventricular of rats. Oxygen-glucose deprivation/reperfusion (OGD/R) model in vitro and middle cerebral artery occlusion (MCAO) model in vivo were established to simulate cerebral I/R injury. QRT-PCR analysis and western blotting assay were performed to assess the expression of NAA10. TTC staining, neurological evaluation and cell counting kit-8 (CCK-8) were performed to evaluate the effect of NAA10 on sevoflurane induced neuroprotection. TUNEL assay and flow cytometry were used to detect the apoptosis in vivo and in vitro., Results: It showed that sevoflurane preconditioning increased the expression of NAA10 in MCAO rats. TTC staining, TUNEL assay and neurological evaluation results suggested that si-NAA10 attenuated the neuroprotective effect of sevoflurane preconditioning against MCAO. CCK-8 assay, flow cytometry, qRT-PCR and western blot results showed that NAA10 mediated sevoflurane preconditioning-induced neuroprotection in vitro. Furthermore, western blot results showed that down-regulation of NAA10 could reverse the attenuation of ERK1/2 phosphorylation induced by sevoflurane preconditioning in vivo or in vitro., Conclusion: Down-regulation of NAA10 regulated ERK1/2 phosphorylation mediating sevoflurane preconditioning induced neuroprotective effects. The results revealed the regulatory mechanism of NAA10 in the neuroprotective effect of sevoflurane preconditioning., (Copyright © 2021. Published by Elsevier B.V.)

Chronic inflammation plays a central role in hepatocellular carcinoma (HCC), but the contribution of hepatocytes to tumor-associated inflammation is not clear. Here, we report that the zinc finger transcription factor Miz1 restricted hepatocyte-driven inflammation to suppress HCC, independently of its transcriptional activity. Miz1 was downregulated in HCC mouse models and a substantial fraction of HCC patients. Hepatocyte-specific Miz1 deletion in mice generated a distinct sub-group of hepatocytes that produced pro-inflammatory cytokines and chemokines, which skewed the polarization of the tumor-infiltrating macrophages toward pro-inflammatory phenotypes to promote HCC. Mechanistically, Miz1 sequestrated the oncoprotein metadherin (MTDH), preventing MTDH from promoting transcription factor nuclear factor κB (NF-κB) activation. A distinct sub-group of pro-inflammatory cytokine-producing hepatocytes was also seen in a subset of HCC patients. In addition, Miz1 expression inversely correated with disease recurrence and poor prognosis in HCC patients. Our findings identify Miz1 as a tumor suppressor that prevents hepatocytes from driving inflammation in HCC., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Cytochrome P450 (CYP) enzymes play critical roles in drug transformation, and the total CYPs are markedly decreased in alcoholic hepatitis (AH), a fatal alcoholic liver disease. miRNAs are endogenous small noncoding RNAs that regulate many essential biological processes. Knowledge concerning miRNA regulation of CYPs in AH disease is limited. Here we presented the changes of key CYPs in liver samples of AH patients retrieved from GEO database, performed in silico prediction of miRNAs potentially targeting the dysregulated CYP transcripts, and deciphered a novel mechanism underlying miRNA mediated CYPs expression in liver cells. Nine miRNAs were predicted to regulate CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C19, CYP2J2, and CYP3A4, among which hsa-miR-148a-3p was selected as a case study. Biochemical and molecular evidences demonstrated that miR-148a promoted CYP2B6 expression by increasing mRNA stability via directly binding to the 3'UTR sequence, and that this positive posttranscriptional regulation was AGO1/2-dependent. Further, luciferase reporter gene assay and RNA secondary structure analysis illustrated that the seedless target site, not the seed target site, controlled miR-148a-mediated CYP2B6 upregulation. Moreover, we identified HNF4A as a liver-specific transcription factor of MIR-148A through EMSA and chromatin immunoprecipitation experiments. In conclusion, ethanol downregulated miR-148a in hepatocytes through HNF4A regulation, which eventually decreased CYP2B6 expression. Our finding will benefit the understanding of dysregulated drug metabolism in AH patients and highlight an unconventional mechanism for epigenetic regulation of CYP gene expression., (Copyright © 2021 Elsevier Inc. All rights reserved.)

In the adult brain, sonic hedgehog acts on cerebral microvascular endothelial cells to stabilize the blood-brain barrier. The expression of sonic hedgehog by astrocytes is altered during brain injury, and this change has been shown to affect permeability of blood-brain barrier. However, much remains unknown about the regulation of astrocytic sonic hedgehog production. Our results showed that endothelin-1 reduced sonic hedgehog mRNA expression and extracellular protein release in mouse cerebral cultured astrocytes, but had no effect in bEnd.3, a mouse brain microvascular endothelial-derived cell line. The effect of endothelin-1 on astrocyte sonic hedgehog expression was suppressed by an ET B antagonist BQ788, but was unchanged by the ET A antagonist FR139317. In cultured astrocytes and bEnd.3, endothelin-1 did not affect the expression of the sonic hedgehog receptor-related molecules, patched-1 and smoothened. In an animal model of traumatic brain injury, fluid percussion injury on the mouse cerebrum increased the expression of sonic hedgehog, patched-1, and smoothened. Repeated administration of BQ788 enhanced sonic hedgehog expression at 5 days after fluid percussion injury. Histochemical examination revealed sonic hedgehog expression in glial fibrillary acidic protein-positive astrocytes in the cerebrum after fluid percussion injury. Administration of exogenous sonic hedgehog and BQ788 suppressed Evans blue extravasation, an indicator of blood vessel permeability, induced by fluid percussion injury. The effects of BQ788 on fluid percussion injury-induced Evans blue extravasation were reduced by the administration of jervine, a sonic hedgehog inhibitor. Altogether, these results suggest that endothelin-1 down-regulates astrocytic sonic hedgehog to promote disruption of the blood-brain barrier during traumatic brain injury., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Human tauopathies represent a group of neurodegenerative disorders, characterized by abnormal hyperphosphorylation and aggregation of tau protein, which ultimately cause neurodegeneration. The aberrant tau hyperphosphorylation is mostly attributed to the kinases/phosphatases imbalance, which is majorly contributed by the generation of reactive oxygen species (ROS). Globin(s) represent a well-conserved group of proteins which are involved in O 2 management, regulation of cellular ROS in different cell types. Similarly, Drosophila globin1 (a homologue of human globin) with its known roles in oxygen management and development of nervous system exhibits striking similarities with the mammalian neuroglobin. Several recent evidences support the hypothesis that neuroglobins are associated with Alzheimer's disease pathogenesis. We herein noted that targeted expression of human-tau induces the cellular level of Glob1 protein in Drosophila tauopathy models. Subsequently, RNAi mediated restored level of Glob1 restricts the pathogenic effect of human-tau by minimizing its hyperphosphorylation via GSK-3β/p-Akt and p-JNK pathways. In addition, it also activates the Nrf2-keap1-ARE cascade to stabilize the tau-mediated increased level of ROS. These two parallel cellular events provide a significant rescue against human tau-mediated neurotoxicity in the fly models. For the first time we report a direct involvement of an oxygen sensing globin gene in tau etiology. In view of the fact that human genome encodes for the multiple Globin proteins including a nervous system specific neuroglobin; and therefore, our findings may pave the way to investigate if the conserved oxygen sensing globin gene(s) can be exploited in devising novel therapeutic strategies against tauopathies., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Purpose: Hypothyroidism (HT) is associated with accelerated atherosclerosis (AS). The efficacy of traditional strategies of hypothyroid AS remains controversial. Here, we aimed to deepen the understanding of the HT-induced acceleration of AS, to decrease the residual risk of coronary artery disease (CAD) and to find a new therapeutic target., Methods: We collected peripheral venous blood samples from 20 patients and divided them into 4 groups, namely, the normal group, the HT group, the CAD group and the HT + CAD group. Then we performed mRNA microarray analysis and bioinformatics analysis to screen the differentially expressed genes and pathways, and we also conducted validations on ApoE knockout mice models and Raw264.7 cell models., Results: In short, (1) in the analysis between the CAD group and the HT + CAD group, we found a total of 1218 differentially expressed genes, 11 upregulated pathways and 40 downregulated pathways. (2) We validated that patients with HT and CAD had a significantly decreased expression of MAP3K7 (encoding transforming growth factor-β-activated kinase 1, TAK1) gene than normal subjects. (3) In animal and cell experiments, we found the decreased expression of TAK1 and the reduced phosphorylation of AMP-activated protein kinase (AMPK) under the hypothyroid and atherosclerotic condition. (4) Changes in the expressions of TAK1 may affect the progression of AS., Conclusion: Taken together, these data suggest that the accelerated AS in hypothyroid patients may be due to the suppression of TAK1-AMPK pathway in macrophages. This new finding may become a novel therapeutic target in hypothyroid AS.

As the transmembrane receptor of Netrin-1, the tumor suppressor gene Unc-5 Netrin Receptor C (UNC5C) can trigger apoptosis. Although its tumor suppressor effects have been demonstrated in solid tumors such as colon cancer, there is still a lack of systematic research on its expression regulation mechanism. To address this need, we analyzed datasets from The Cancer Genome Atlas (TCGA) database, including multi-omics data for 32 types of cancers and 10,967 cases. Analysis of these data revealed a trend of significantly decreased UNC5C expression in 16 types of solid tumors. Additionally, low UNC5C expression is related to poor prognosis of five types of tumors and restoring the expression of UNC5C can effectively inhibit the proliferation potential of renal cancer cells. Promoter DNA methylation, chromatin remodeling-mediated epigenetic regulation, transcriptional inhibition, RNA-binding protein and miRNA-mediated post-transcriptional inhibition, genetic changes caused by deep deletion and truncated mutations, and ubiquitinating enzyme-mediated protein degradation can synergistically cause the down-regulation of UNC5C expression in solid tumors. This study is the first to analyze the comprehensive molecular mechanism of down-regulation of the tumor suppressor gene UNC5C from multiple dimensions using pan-cancer data. Our results suggest that analyses of gene expression regulation relying on computational biological methods may help guide the targeted therapy of tumor suppressor gene reactivation., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Pseudoephedrine (substituted phenethylamine) is well known as psychotic and bronchodilator. Numerous studies on phenethylamine derivatives indicated that these agents have the potential to abolish inflammatory responses in the non-biological and biological systems. These facts provided the basis to conduct a study on pseudoephedrine to explore its therapeutics in Complete Freund's Adjuvant (CFA)-induced arthritis. Furthermore, existing treatment approaches for RA associated with limited effect on chronic immunological models. Real-time polymerase chain reaction (q-PCR) was performed to execute the expression of pro and anti-inflammatory cytokines in treated and non-treated arthritic rats. These findings were further co investigate by histological observations. The paw volume, paw diameter, weight variations and arthritic score were determined at specific days throughout the experiment of 28 days. Pseudoephedrine at all doses significantly (p < 0.001) suppressed the expression of PGE2, TNF-α, IL-1β and IL-6. Moreover, pseudoephedrine (20 and 40 mg/kg) caused significant augmentation of IL-4 and IL-10. Similarly, the drug expressed a significant anti-arthritic effect by reducing the paw volume, paw diameter and arthritic score. Similarly, it also reverts the reduction in body weight of arthritic rats at all above-mentioned doses. These findings supported the anti-arthritic potential of pseudoephedrine and recommended it for clinical trials.

The extracellular matrix (ECM) is the principal structure of bone tissue. Long-term spaceflights lead to osteopenia, which may be a result of the changes in composition as well as remodeling of the ECM by osteogenic cells. To elucidate the cellular effects of microgravity, human mesenchymal stromal cells (MSCs) and their osteocommitted progeny were exposed to simulated microgravity (SMG) for 10 days using random positioning machine (RPM). After RPM exposure, an imbalance of MSC collagen/non-collagen ratio at the expense of a decreased level of collagenous proteins was detected. At the same time, the secretion of proteases (cathepsin A, cathepsin D, MMP3) was increased. No significant effects of SMG on the expression of stromal markers and cell adhesion molecules on the MSC surface were noted. Upregulation of COL11A1 , CTNND1 , TIMP3 , and TNC and downregulation of HAS1 , ITGA3 , ITGB1 , LAMA3 , MMP1 , and MMP11 were detected in RPM exposed MSCs. ECM-associated transcriptomic changes were more pronounced in osteocommitted progeny. Thus, 10 days of SMG provokes a decrease in the collagenous components of ECM, probably due to the decrease in collagen synthesis and activation of proteases. The presented data demonstrate that ECM-associated molecules of both native and osteocommitted MSCs may be involved in bone matrix reorganization during spaceflight.

Purpose: Proper aqueous humor (AH) dynamics is crucial for maintaining the intraocular pressure (IOP) in the eye. This study aims to investigate the function of Piezo2, a newly discovered mechanosensitive ion channel, in regulating AH dynamics., Methods: Immunohistochemistry (IHC) analysis and western blotting were performed to detect Piezo2 expression. The Cre-lox system was applied to create a conditional knockout model of Piezo2. IOP and aqueous humor outflow facility in live animals were recorded with a Tonometer and a syringe-pump system for up to 2 weeks., Results: We first detected Piezo2 with robust expression in the human trabecular meshwork (TM), Schlemm's canal (SC), the ciliary body's epithelium, and ciliary muscle. In addition, we found Piezo2 in human retinal ganglion cells (RGCs) and astrocytes in the optic nerve head (ONH). Through the Cre-lox system, Piezo2 can be successfully downregulated in mouse iridocorneal angle tissues. However, Piezo2 downregulation cannot significantly influence the IOP and outflow facility through the conventional pathway. Instead, we observed an effect of downregulated Piezo2 on decreasing the intercept in the flow rate versus pressure plot. According to the Goldmann equation, Piezo2 may function in regulating unconventional outflow, AH production, and episcleral venous pressure., Conclusions: These findings, for the first time, demonstrate that Piezo2 acts as an essential mechanosensor in maintaining the proper aqueous humor dynamics in the eye., (Copyright © 2021 Molecular Vision.)

Patients deprived of cigarettes exhibit increased pain sensitivity during perioperative periods, yet the underlying neuroanatomical and molecular bases of this hypersensitivity are unclear. The present study showed that both the mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were significantly decreased in a rat model of nicotine withdrawal. These rats showed less tryptophan hydroxylase 2 (TPH2) positive neurons and reduced TPH2 expression in the nucleus raphe magnus (NRM), and thus resulted in decreased 5-hydroxytryptamine (5-HT) levels in cerebrospinal fluid. Intrathecal injection of 5-HT or NRM microinjection of TPH-overexpression adeno-associated virus alleviated nicotine withdrawal-induced hyperalgesia, whereas 5-HT receptor pharmacological blockade by methysergide (a 5-HT receptor antagonist) exacerbated hypersensitivity and diminished the difference between the two groups. Together, these data indicate that hyperalgesia after nicotine withdrawal is mediated by declined descending serotonergic pathways in the NRM. This provides a new perspective to improve the postoperative pain management of patients, especially the smokers., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

A hallmark of aging is immunosenescence, a decline in immune functions, which appeared to be inevitable in living organisms, including Caenorhabditis elegans. Here, we show that genetic inhibition of the DAF-2/insulin/IGF-1 receptor drastically enhances immunocompetence in old age in C. elegans. We demonstrate that longevity-promoting DAF-16/FOXO and heat-shock transcription factor 1 (HSF-1) increase immunocompetence in old daf-2(-) animals. In contrast, p38 mitogen-activated protein kinase 1 (PMK-1), a key determinant of immunity, is only partially required for this rejuvenated immunity. The up-regulation of DAF-16/FOXO and HSF-1 decreases the expression of the zip-10/bZIP transcription factor, which in turn down-regulates INS-7, an agonistic insulin-like peptide, resulting in further reduction of insulin/IGF-1 signaling (IIS). Thus, reduced IIS prevents immune aging via the up-regulation of anti-aging transcription factors that modulate an endocrine insulin-like peptide through a feedforward mechanism. Because many functions of IIS are conserved across phyla, our study may lead to the development of strategies against immune aging in humans., (© 2021 Lee et al.)

Background: Secondary plasma cell leukaemia (sPCL) is an aggressive form of multiple myeloma (MM), but the mechanism underlying MM progresses into PCL remains unknown., Methods: Gene expression profiling of MM patients and PCL patients was analysed to identify the molecular differences between the two diseases. Cox survival regression and Kaplan-Meier analysis were performed to illustrate the impact of integrin subunit alpha 6 (ITGA6) on prognosis of MM. Invasion assays were performed to assess whether ITGA6 regulated the progression of MM to PCL., Results: Gene expression profiling analyses showed that cell metastasis pathways were enriched in PCL and ITGA6 was differentially expressed between PCL and MM. ITGA6 expression was an independent prognostic factor for event-free survival (EFS) and overall survival (OS) of MM patients. Moreover, the stratification ability of the International Staging System (ISS) of MM was improved when including ITGA6 expression. Functional studies uncovered that increased ITGA6 reduced the myeloma cell invasion. Additionally, low expression of ITGA6 resulted from epigenetic downregulating of its anti-sense non-coding RNA, ITGA6-AS1., Conclusion: Our data reveal that ITGA6 gradually decreases during plasma cell dyscrasias progression and low expression of ITGA6 contributes to myeloma metastasis. Moreover, ITGA6 abundance might help develop MM prognostic stratification.

BACKGROUND Acute myocardial infarction is the leading cause of mortality among adults worldwide. The present study aimed to investigate the role and mechanism of thrombin and SIRT1 in hypoxia/reoxygenation (H/R) injury. MATERIAL AND METHODS H9c2 cardiomyocytes were used to create an H/R model to simulate in vivo ischemia/reperfusion injury. The MTT assay was used to measure cell viability, qRT-PCR was used to detect the level of SIRT1, thrombin, and PAR-1, and western blot analysis was conducted for evaluation of thrombin, PAR-1, SIRT1, LC3I, LC3II, and Beclin1. ELISA was applied for determination of IL-1ß, IL-6, TNF-alpha, MMP-9, and ICAM-1. After the establishment of the H/R model, superoxide dismutase (SOD) activity was evaluated by the xanthine oxidase method, malondialdehyde content was detected by thiobarbituric acid assay, and reactive oxygen species generation was measured by CM-H2DCFDA. RESULTS The findings showed that thrombin enhanced inflammatory factor secretion and oxidative stress but inhibited cell viability in H/R-injured cardiomyocytes. We also observed that thrombin promoted autophagy in H/R-injured cardiomyocytes. In addition, thrombin enhanced the upregulation of SIRT1 expression by H/R. However, it was found that inhibition of SIRT1 could suppress the effect of thrombin on inflammatory factor secretion, oxidative stress, and cell viability. Moreover, downregulation of SIRT1 suppressed the inhibitory effect of thrombin on autophagy in H/R injury. CONCLUSIONS Thrombin aggravates H/R injury of cardiomyocytes by activating an autophagy pathway mediated by SIRT1. These findings might provide a potential target therapy for the treatment of ischemia/reperfusion injury in future clinical work.

This study aimed to explore the effects and function of microRNA-101a-3p (miR-101a-3p) in epilepsy. Rat model of pilocarpine-induced epilepsy was established and the seizure frequency was recorded. Expression of miR-101a-3p and c-Fos in hippocampus tissues of Rat models were detected by qRT-PCR and western blot. Besides, we established a hippocampal neuronal culture model of acquired epilepsy using Mg 2+ free medium to evaluate the effects of miR-101a-3p and c-Fos in vitro. Cells were transfected with miR-101a-3p mimic, si-c-FOS, miR-101a-3p + c-FOS and its corresponding controls. MTT assay was used to detect cell viability upon transfection. Flow cytometry was performed to determine the apoptosis rate. Western blot was performed to measure the protein expression of apoptosis-related proteins (Bcl-2, Bax, and cleaved caspase 3), autophagy-related proteins (LC3 and Beclin1) and c-FOS. The targeting relationship between miR-101a-3p and c-FOS was predicted and verified by TargetScan software and dual-luciferase reporter assay. The role of miR-101a-3p was validated using epilepsy rat models in vivo. Another Rat models of pilocarpine-induced epilepsy with miR-NC or miR-101a-3p injection were established to evaluate the effect of miR-101a-3p overexpression on epilepsy in vivo. MiR-101a-3p was downregulated while c-FOS was increased in hippocampus tissues of Rat model of pilocarpine-induced epilepsy. Overexpression of miR-101a-3p or c-FOS depletion promoted cell viability, inhibited cell apoptosis and autophagy. C-FOS was a target of miR-101a-3p and miR-101a-3p negatively regulated c-FOS expression to function in epilepsy. Overexpression of miR-101a-3p attenuated pilocarpine-induced epilepsy in Rats in vivo. This study indicated that miR-101a-3p could attenuate pilocarpine-induced epilepsy by repressing c-Fos expression.

Exposure to specific doses of hypoxia can trigger endogenous neuroprotective and neuroplastic mechanisms of the central nervous system. These molecular mechanisms, together referred to as hypoxic preconditioning (HPC), remain poorly understood. In the present study, we applied RNA sequencing and bioinformatics analyses to study HPC in a whole-body HPC mouse model. The preconditioned (H4) and control (H0) groups showed 605 differentially expressed genes (DEGs), of which 263 were upregulated and 342 were downregulated. Gene Ontology enrichment analysis indicated that these DEGs were enriched in several biological processes, including metabolic stress and angiogenesis. The Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that the FOXO and Notch signaling pathways were involved in hypoxic tolerance and protection during HPC. Furthermore, 117 differential alternative splicing events (DASEs) were identified, with exon skipping being the dominant one (48.51%). Repeated exposure to systemic hypoxia promoted skipping of exon 7 in Edrf1 and exon 9 or 13 in Lrrc45. This study expands the understanding of the endogenous protective mechanisms of HPC and the DASEs that occur during HPC.

Inflammation is a self-protection mechanism that can be triggered when innate immune cells detect infection. Eradication of pathogen infection requires appropriate immune and inflammatory responses, but excessive inflammatory responses can cause uncontrolled inflammation, autoimmune diseases, or pathogen dissemination. Mounting evidence has shown that microRNAs (miRNAs) in mammals act as important and versatile regulators of innate immunity and inflammation. However, miRNA-mediated regulation networks are largely unknown in inflammatory responses in lower vertebrates. Here miR-144 and miR-217 are identified as negative regulators in teleost inflammatory responses. We find that Vibrio harveyi and lipopolysaccharide (LPS) treatment significantly upregulate the expression of fish miR-144 and miR-217. Upregulated miR-144 and miR-217 suppress LPS-induced inflammatory cytokine expression by targeting nucleotide-binding oligomerization domain-containing protein 1 (NOD1), thereby avoiding excessive inflammatory responses. In addition, miR-144 and miR-217 regulate inflammatory responses through NOD1-induced nuclear factor kappa (NF-kB) signaling pathways. These findings demonstrate that miR-144 and miR-217 play regulatory roles in inflammatory responses by modulating the NOD1-induced NF-κB signaling pathway.

Dysfunction of bone marrow mesenchymal stem cells (BMSCs), osteoblasts and osteocytes may be one of the main causes of bone loss in the elderly. In the present study, we found osteogenic cells from aged rats all exhibited senescence changes, with the most pronounced senescence changes in osteocytes. Meanwhile, the proliferative capacity and functional activity of osteogenic cells from aged rats were suppressed. Osteogenic differentiation capacity of BMSCs from aged rats decreased while adipogenic capacity increased. The mineralization capacity, ALP activity and osteogenic proteins expression of osteoblasts from aged rats decreased. Additionally, osteocytes from aged rats up-expressed sclerosteosis protein, a negative regulator of bone formation. To inhibit osteogenic cell senescence, we use low magnitude vibration (LMV) to eliminate the senescent osteogenic cells. After LMV treatment, the number of osteogenic cells staining positively for senescence-associated-β-galactosidase (SA-β-Gal) decreased significantly. Besides, the expression of anti-aging protein SIRT1 was upregulated significantly, while p53 and p21 were downregulated significantly after LMV treatment. Thus, the LMV can inhibit the senescence of osteogenic cells partly through the Sirt1/p53/p21 axis. Furthermore, LMV was found to promote bone formation of aged rats. These results suggest that the inhibition of osteogenic cell senescence by LMV is a valuable treatment to prevent or delay osteoporosis.