PLOS ONE: [sortOrder=DATE_NEWEST_FIRST, from=editorLink, sort=Relevance, q=editor:"Partha Mukhopadhyay"]PLOShttps://journals.plos.org/plosone/webmaster@plos.orgaccelerating the publication of peer-reviewed sciencehttps://journals.plos.org/plosone/search/feed/atom?sortOrder=DATE_NEWEST_FIRST&unformattedQuery=editor:%22Partha%20Mukhopadhyay%22&from=editorLink&sort=RelevanceAll PLOS articles are Open Access.https://journals.plos.org/plosone/resource/img/favicon.icohttps://journals.plos.org/plosone/resource/img/favicon.ico2024-03-29T15:35:10ZUpregulation of Nrf2 in myocardial infarction and ischemia-reperfusion injury of the heartSahar ZuberiHira RafiAzhar HussainSatwat Hashmi10.1371/journal.pone.02995032024-03-15T14:00:00Z2024-03-15T14:00:00Z<p>by Sahar Zuberi, Hira Rafi, Azhar Hussain, Satwat Hashmi</p>
Myocardial infarction (MI) is a leading cause of morbidity and mortality in the world and is characterized by ischemic necrosis of an area of the myocardium permanently devoid of blood supply. During reperfusion, reactive oxygen species are released and this causes further insult to the myocardium, resulting in ischemia-reperfusion (IR) injury. Since Nrf2 is a key regulator of redox balance, it is essential to determine its contribution to these two disease processes. Conventionally Nrf2 levels have been shown to rise immediately after ischemia and reperfusion but its contribution to disease process a week after the injury remains uncertain. Mice were divided into MI, IR injury, and sham surgery groups and were sacrificed 1 week after surgery. Infarct was visualized using H&E and trichrome staining and expression of Nrf2 was assessed using immunohistochemistry, Western blot, and ELISA. MI displayed a higher infarct size than the IR group (MI: 31.02 ± 1.45%, IR: 13.03 ± 2.57%; p < 0.01). We observed a significantly higher expression of Nrf2 in the MI group compared to the IR model using immunohistochemistry, spot densitometry of Western blot (MI: 2.22 ± 0.16, IR: 1.81 ± 0.10, Sham: 1.52 ± 0.13; p = 0.001) and ELISA (MI: 80.78 ± 27.08, IR: 31.97 ± 4.35; p < 0.01). There is a significantly higher expression of Nrf2 in MI compared to the IR injury group. Modulation of Nrf2 could be a potential target for therapeutics in the future, and its role in cardioprotection can be further investigated.Melatonin serves as a novel treatment in bladder fibrosis through TGF-β1/Smad and EMTYang ZhangSun GongWeixin HeJie YuanDi DongJialong ZhangHaomin WangBinghai Chen10.1371/journal.pone.02951042024-03-13T14:00:00Z2024-03-13T14:00:00Z<p>by Yang Zhang, Sun Gong, Weixin He, Jie Yuan, Di Dong, Jialong Zhang, Haomin Wang, Binghai Chen</p>
Background <p>Melatonin (MEL) is an indole amine molecule primarily produced in the pineal gland. Melatonin has been shown in numerous studies to have antifibrotic effects on the kidney, liver, and other organs. However, it is still unclear how melatonin works in bladder fibrosis. We explored how melatonin affects animals with bladder fibrosis and the underlying mechanisms.</p> Materials and methods <p>MEL was used to treat human bladder smooth muscle cells (HBdSMCs) after they were stimulated with transforming growth factor-β1 (TGF-β1) <i>in vitro</i>. Proteomic analysis and bioinformatic analysis of the altered expression of these proteins were subsequently performed on HBdSMCs from the different processing methods. To construct an <i>in vivo</i> bladder fibrosis model, we injected protamine sulfate (PS) and lipopolysaccharide (LPS) twice a week into the rat bladder for six weeks. After two weeks of PS/LPS treatment, the mice in the treatment group were treated with MEL (20 mg/kg/d) for 4 weeks. Finally, we detected the expression of fibrosis markers from different perspectives. The TGF-β1/Smad pathway and epithelial–mesenchymal transition (EMT) in cell and bladder tissues were also identified. Further proteomic analysis was also performed.</p> Results <p><i>In vitro</i>, we found that TGF-β1 treatment enhanced the expression of the fibrosis markers collagen III and α-SMA in HBdSMCs. E-cadherin expression decreased while the TGF-β1/Smad pathway was activated. Vimentin and N-cadherin expression was also elevated at the same time. Similar findings were observed in the LPS group. After MEL treatment, the expression of collagen III and α-SMA decreased, the expression of E-cadherin increased, and the expression of vimentin and N-cadherin also decreased. According to our quantitative proteomics analysis, CCN1 and SQLE may be important proteins involved in the development of bladder fibrosis. MEL decreased the expression of these genes, leading to the relief of bladder fibrosis. Bioinformatics analysis revealed that the extracellular space structure related to metabolic pathways, actin filament binding, and stress fibers can serve as a pivotal focus in the management of fibrosis.</p> Conclusion <p>Melatonin attenuates bladder fibrosis by blocking the TGF-β1/Smad pathway and EMT. CCN1 appears to be a possible therapeutic target for bladder fibrosis.</p>miR-328-3p targets TLR2 to ameliorate oxygen-glucose deprivation injury and neutrophil extracellular trap formation in HUVECs via inhibition of the NF-κB signaling pathwayMengting YaoChucun FangZilong WangTianting GuoDongwen WuJiacheng MaJian WuJianwen Mo10.1371/journal.pone.02993822024-02-23T14:00:00Z2024-02-23T14:00:00Z<p>by Mengting Yao, Chucun Fang, Zilong Wang, Tianting Guo, Dongwen Wu, Jiacheng Ma, Jian Wu, Jianwen Mo</p>
Background <p>Endothelial cell injury is one of the important pathogenic mechanisms in thrombotic diseases, and also neutrophils are involved. MicroRNAs (miRNAs) have been demonstrated to act as essential players in endothelial cell injury, but the potential molecular processes are unknown. In this study, we used cellular tests to ascertain the protective effect of miR-328-3p on human umbilical vein endothelial cells (HUVECs) treated with oxygen-glucose deprivation (OGD).</p> Methods <p>In our study, an OGD-induced HUVECs model was established, and we constructed lentiviral vectors to establish stable HUVECs cell lines. miR-328-3p and Toll-like receptor 2 (TLR2) interacted, as demonstrated by the dual luciferase reporter assay. We used the CCK8, LDH release, and EdU assays to evaluate the proliferative capacity of each group of cells. To investigate the expression of TLR2, p-P65 NF-κB, P65 NF-κB, NLRP3, IL-1β, and IL-18, we employed Western blot and ELISA. Following OGD, each group’s cell supernatants were gathered and co-cultured with neutrophils. An immunofluorescence assay and Transwell assay have been performed to determine whether miR-328-3p/TLR2 interferes with neutrophil migration and neutrophil extracellular traps (NETs) formation.</p> Results <p>In OGD-treated HUVECs, the expression of miR-328-3p is downregulated. miR-328-3p directly targets TLR2, inhibits the NF-κB signaling pathway, and reverses the proliferative capacity of OGD-treated HUVECs, while inhibiting neutrophil migration and neutrophil extracellular trap formation.</p> Conclusions <p>miR-328-3p inhibits the NF-κB signaling pathway in OGD-treated HUVECs while inhibiting neutrophil migration and NETs formation, and ameliorating endothelial cell injury, which provides new ideas for the pathogenesis of thrombotic diseases.</p>Palmitoleic acid protects microglia from palmitate-induced neurotoxicity in vitroQingting YuYanzhuo YangTing XuYinsheng CaiZuisu YangFalei Yuan10.1371/journal.pone.02970312024-01-19T14:00:00Z2024-01-19T14:00:00Z<p>by Qingting Yu, Yanzhuo Yang, Ting Xu, Yinsheng Cai, Zuisu Yang, Falei Yuan</p>
Although palmitoleic acid (POA) is a lipokine with beneficial effects on obesity and is produced as a byproduct from the manufacture of prescription omega-3 fatty acids, its role in nervous system inflammation is still unknown. This study aims to examine the mechanisms and protective effects of POA against palmitic acid (PA)-induced microglial death. PA-induced microglial death was used as a model for POA intervention. Various inhibitors were employed to suppress potential routes of PA entry into the cell. Immunofluorescence staining and Western blotting were conducted to elucidate the protective pathways involved. The results suggest POA has the potential to eliminate PA-induced lactate dehydrogenase (LDH) release, which decreases the overall number of propidium iodide (PI)-positive cells compared with control. Moreover, POA has the potential to significantly increase lipid droplets (LDs) in the cytoplasm, without causing any lysosomal damage. POA inhibited both canonical and non-canonical gasdermin D (GSDMD)-mediated pyroptosis and gasdermin E (GSDME)-mediated pyroptosis, which PA typically induces. Additionally, POA inhibited the endoplasmic reticulum (ER) stress and apoptosis-related proteins induced by PA. Based on the findings, POA can exert a protective effect on microglial death induced by PA via pathways related to pyroptosis, apoptosis, ER stress, and LDs.Regular exercise attenuates alcoholic myopathy in zebrafish by modulating mitochondrial homeostasisWei WenCheng GuoZhanglin ChenDong YangDanting ZhuQuwen JingLan ZhengChenchen SunChangfa Tang10.1371/journal.pone.02947002023-11-30T14:00:00Z2023-11-30T14:00:00Z<p>by Wei Wen, Cheng Guo, Zhanglin Chen, Dong Yang, Danting Zhu, Quwen Jing, Lan Zheng, Chenchen Sun, Changfa Tang</p>
Alcoholic myopathy is caused by chronic consumption of alcohol (ethanol) and is characterized by weakness and atrophy of skeletal muscle. Regular exercise is one of the important ways to prevent or alleviate skeletal muscle myopathy. However, the beneficial effects and the exact mechanisms underlying regular exercise on alcohol myopathy remain unclear. In this study, a model of alcoholic myopathy was established using zebrafish soaked in 0.5% ethanol. Additionally, these zebrafish were intervened to swim for 8 weeks at an exercise intensity of 30% of the absolute critical swimming speed (Ucrit), aiming to explore the beneficial effects and underlying mechanisms of regular exercise on alcoholic myopathy. This study found that regular exercise inhibited protein degradation, improved locomotion ability, and increased muscle fiber cross-sectional area (CSA) in ethanol-treated zebrafish. In addition, regular exercise increases the functional activity of mitochondrial respiratory chain (MRC) complexes and upregulates the expression levels of MRC complexes. Regular exercise can also improve oxidative stress and mitochondrial dynamics in zebrafish skeletal muscle induced by ethanol. Additionally, regular exercise can activate mitochondrial biogenesis and inhibit mitochondrial unfolded protein response (UPRmt). Together, our results suggest regular exercise is an effective intervention strategy to improve mitochondrial homeostasis to attenuate alcoholic myopathy.Cinnamaldehyde alleviates doxorubicin-induced cardiotoxicity by decreasing oxidative stress and ferroptosis in cardiomyocytesMeijiao MaoWang ZhengBin DengYouhua WangDuan ZhouLin ShenWankang NikuNa Zhang10.1371/journal.pone.02921242023-10-12T14:00:00Z2023-10-12T14:00:00Z<p>by Meijiao Mao, Wang Zheng, Bin Deng, Youhua Wang, Duan Zhou, Lin Shen, Wankang Niku, Na Zhang</p>
Although doxorubicin (DOX) is an efficient chemotherapeutic drug for human tumors, severe cardiotoxicity restricts its clinical use. Cinnamaldehyde (CA), a bioactive component isolated from Cinnamonum cassia, possesses potent anti-oxidative and anti-apoptotic potentials. The major aim of this study was to evaluate the protective role of CA against DOX-induced cardiotoxicity. To this end, cardiomyocyte injury models were developed using DOX-treated H9c2 cells and DOX-treated rats, respectively. Herein, we found that CA treatment increased cardiomyocyte viability and attenuated DOX-induced cardiomyocyte death <i>in vitro</i>. CA further protected rats against DOX-induced cardiotoxicity, as indicated by elevated creatine kinase (CK) and lactate dehydrogenase (LDH) levels, myocardium injury, and myocardial fibrosis. CA alleviated DOX-induced myocardial oxidative stress by regulating reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH) levels. Mechanistically, CA markedly accelerated nuclear translocation of nuclear erythroid factor 2-related factor 2 (Nrf2) and increased heme oxygenase-1 (HO-1) expression. Consequently, CA decreased DOX-induced cardiomyocyte ferroptosis, while Erastin (a ferroptosis agonist) treatment destroyed the effect of CA on increasing cardiomyocyte viability. Taken together, the current results demonstrate that CA alleviates DOX-induced cardiotoxicity, providing a promising opportunity to increase the clinical application of DOX.Race and sex differences in ROS production and SOD activity in HUVECsSara E. MasconeKatherine I. KimWilliam S. EvansSteven J. PriorMarc D. CookSushant M. Ranadive10.1371/journal.pone.02921122023-10-04T14:00:00Z2023-10-04T14:00:00Z<p>by Sara E. Mascone, Katherine I. Kim, William S. Evans, Steven J. Prior, Marc D. Cook, Sushant M. Ranadive</p>
Black individuals and men are predisposed to an earlier onset and higher prevalence of hypertension, compared with White individuals and women, respectively. Therefore, the influence of race and sex on reactive oxygen species (ROS) production and superoxide dismutase (SOD) activity following induced inflammation was evaluated in female and male human umbilical vein endothelial cells (HUVECs) from Black and White individuals. It was hypothesized that HUVECs from Black individuals and male HUVECs would exhibit greater ROS production and impaired SOD activity. Inflammation was induced in HUVEC cell lines (n = 4/group) using tumor necrosis factor-alpha (TNF-α, 50ng/ml). There were no between group differences in ROS production or SOD activity in HUVECs from Black and White individuals, and HUVECs from Black individuals exhibited similar SOD activity at 24hr compared with 4hr of TNF-α treatment (p>0.05). However, HUVECs from White individuals exhibited significantly greater SOD Activity (p<0.05) at 24hr as compared to 4hr in the control condition but not with TNF-α treatment (p>0.05). Female HUVECs exhibited significantly lower ROS production than male HUVECs in the control condition and following TNF-α induced inflammation (p<0.05). Only female HUVECs exhibited significant increases in SOD activity with increased exposure time to TNF-α induced inflammation (p<0.05). HUVECs from White individuals alone exhibit blunted SOD activity when comparing control and TNF-α conditions. Further, compared to female HUVECs, male HUVECs exhibit a pro-inflammatory state.Elastin-targeted nanoparticles delivering doxycycline mitigate cytokine storm and reduce immune cell infiltration in LPS-mediated lung inflammationShivani AroraNarendra Vyavahare10.1371/journal.pone.02862112023-06-02T14:00:00Z2023-06-02T14:00:00Z<p>by Shivani Arora, Narendra Vyavahare</p>
Background and purpose <p>Cytokine storm invoked during acute and chronic lung injury promotes alveolar damage and remodeling. The current study shows that degraded elastin-targeted nanoparticles releasing doxycycline (Doxy NPs) are potent in mitigating cytokines storm, migration of immune cells in the lungs, and inhibiting inflammasome pathways in the LPS mouse model.</p> Experimental approach <p>Cytokine storm and lung injury were induced using LPS and elastase in C57BL/6 mice (rodent model for emphysema). The mice were then treated with I.V. Doxy NPs, blank NPs, or Doxy a day before LPS administration. Cytokine levels, immune cell population, and MMP activity were analyzed in broncheo-alveolar lavage fluid (BALF) 4 hours after LPS administration. Additionally, gene expression of IL-6, IL-1beta, MCP-1, NLRP3, Caspase 1 and MMPs were investigated in alveolar cells on day 3 after LPS administration.</p> Key results <p>Doxycycline NPs but not Doxycycline significantly decreased IL-6, TNF-α, IL-23 and were significantly more effective in decreasing the percentage of immune cells in the BALF. This is the first in-vivo study to demonstrate that Doxycycline can effectively inhibit inflammasome pathways in the lungs.</p> Conclusion and implications <p>IV administration of elastin antibody conjugated Doxycycline-loaded albumin NPs can effectively modulate the local immune environment in the lungs, which is not achieved by IV Doxycycline even at 100-fold higher dose. This novel method of drug delivery can effectively lead to the repurposing of traditional Doxycycline as a potential adjunct treatment for managing the cytokine storm in the lungs in COPD and viral infections.</p>miR-96-5p is involved in alcohol-induced apoptosis in PC12 cells via negatively regulating TAp73Bin YangQi WangYanzhong LiLin LiYanJie ZhangMohammad Farris Iman Leong Bin AbdullahWei HaoDuan LiRuiling Zhang10.1371/journal.pone.02824882023-04-26T14:00:00Z2023-04-26T14:00:00Z<p>by Bin Yang, Qi Wang, Yanzhong Li, Lin Li, YanJie Zhang, Mohammad Farris Iman Leong Bin Abdullah, Wei Hao, Duan Li, Ruiling Zhang</p>
Objective <p>The present study opted for the adrenal phaeochromocytoma (PC12) cell line to frame a neuronal injury model induced by alcohol exposure in vitro, aiming to probe whether TAp73 and miR-96-5p are involved in the neuronal injury process induced by alcohol and elucidate the regulatory relationship between miR-96-5p and TAp73.</p> Methods <p>Immunofluorescence staining was used to observe the structural features of PC12 cells after culturing in medium with nerve growth factor (NGF). After different doses and different durations of alcohol treatment, CCK-8 assay was performed to detect the viability of PC12 cells, flow cytometry assay was carried out to detect the apoptosis rate of PC12 cells, dual-luciferase reporter assay was used to definitude the regulatory relationship between miR-96-5p and Tp73, and western blot was used to detect the protein expression of TAp73.</p> Results <p>The result of immunofluorescence staining demonstrated that PC12 cells abundantly expressed Map2, CCK-8 assay illustrated alcohol exposure significantly downregulated the cell viability of PC12 cells, Treatment with miR-96-5p inhibitor induced apoptosis and upregulated the expression of TAp73 in PC12 cells. Contrastingly, miR-96-5p mimic reversed the above effects and downregulation of TAp73 inhibited the apoptosis of PC12 cells.</p> Conclusion <p>The present study demonstrated that miR-96-5p participates in alcohol-induced apoptosis in PC12 cells via negatively regulating TAp73.</p>Necrotic cardiac myocytes skew macrophage polarization towards a classically activated phenotypeWenlong JiangLuigi AdamoKenji LimScot J MatkovichSarah EvansCibele Rocha-ResendeDouglas L Mann10.1371/journal.pone.02829212023-03-30T14:00:00Z2023-03-30T14:00:00Z<p>by Wenlong Jiang, Luigi Adamo, Kenji Lim, Scot J Matkovich, Sarah Evans, Cibele Rocha-Resende, Douglas L Mann</p>
Necrotic and dying cells release damage-associated molecular patterns (DAMPs) that can initiate sterile inflammatory responses in the heart. Although macrophages are essential for myocardial repair and regeneration, the effect of DAMPs on macrophage activation remains unclear. To address this gap in knowledge we studied the effect of necrotic cardiac myocyte extracts on primary peritoneal macrophage (PPM) cultures in vitro. We first performed unbiased transcriptomic profiling with RNA-sequencing of PPMs cultured for up to 72 hours in the presence and absence of: 1) necrotic cell extracts (NCEs) from necrotic cardiac myocytes in order to mimic the release of DAMPs; 2) lipopolysaccharide (LPS), which is known to polarize macrophages towards a classically activated phenotype and 3) Interleukin-4 (IL-4), which is known to promote polarization of macrophages towards an alternatively activated phenotype. NCEs provoke changes in differential gene expression (DEGs) that had considerable overlap with LPS-induced changes, suggesting that NCEs promote macrophage polarization towards a classically activated phenotype. Treating NCEs with proteinase-K abolished the effects of NCEs on macrophage activation, whereas NCE treatment with DNase and RNase did not affect macrophage activation. Stimulation of macrophage cultures with NCEs and LPS resulted in a significant increase in macrophage phagocytosis and interleukin-1β secretion, whereas treatment with IL-4 had no significant effect on phagocytosis and interleukin-1β. Taken together, our findings suggest that proteins released from necrotic cardiac myocytes are sufficient to skew the polarization of macrophages towards a classically activated phenotype.Anti-inflammatory effects of chlorogenic acid from <i>Taraxacum officinale</i> on LTA-stimulated bovine mammary epithelial cells via the TLR2/NF-κB pathwayPing XuXiaobo XuHanna FotinaTetiana Fotina10.1371/journal.pone.02823432023-03-22T14:00:00Z2023-03-22T14:00:00Z<p>by Ping Xu, Xiaobo Xu, Hanna Fotina, Tetiana Fotina</p>
Mastitis is an inflammatory disease caused by microbial infection. Chlorogenic acid (CGA), one of the major phenolic acids in <i>Taraxacum officinale</i>, has natural antioxidant and anti-inflammatory properties in various cell types; however, the effects of CGA on Lipoteichoic acid (LTA)-induced bovine mammary epithelial cells (BMECs) have not been investigated. In this study, the CGA content in <i>T</i>. <i>officinale</i> was determined by High-performance liquid chromatography (HPLC). BMECs were infected with LTA to induce the mastitis model. Different concentrations of CGA were administered after establishing the LTA infection. The results showed that the <i>T</i>. <i>officinale</i> contained CGA 1.36 mg/g. CGA significantly reduced the pro-inflammatory gene and protein expression of TNF-α, IL-6, and IL-1β. In addition, CGA downregulated the NO, TLR2, and NF-κB signaling pathways in LTA-infected bovine mammary epithelial cells. Our results indicate that CGA reduced the expression of TNF-α, IL-6, IL-1β, and TLR2 by inhibiting the phosphorylation of proteins in the NF-κB signaling pathways in a dose-dependent manner. This finding suggests that CGA may be a potential agent for the treatment of mastitis in dairy cows.Combined non-psychoactive Cannabis components cannabidiol and β-caryophyllene reduce chronic pain via CB1 interaction in a rat spinal cord injury modelAnjalika EeswaraAmanda Pacheco-SpiewakStanislava JergovaJacqueline Sagen10.1371/journal.pone.02829202023-03-13T14:00:00Z2023-03-13T14:00:00Z<p>by Anjalika Eeswara, Amanda Pacheco-Spiewak, Stanislava Jergova, Jacqueline Sagen</p>
The most frequently reported use of medical marijuana is for pain relief. However, its psychoactive component Δ9-tetrahydrocannabinol (THC) causes significant side effects. Cannabidiol (CBD) and β-caryophyllene (BCP), two other cannabis constituents, possess more benign side effect profiles and are also reported to reduce neuropathic and inflammatory pain. We evaluated the analgesic potential of CBD and BCP individually and in combination in a rat spinal cord injury (SCI) clip compression chronic pain model. Individually, both phytocannabinoids produced dose-dependent reduction in tactile and cold hypersensitivity in male and female rats with SCI. When co-administered at fixed ratios based on individual A50s, CBD and BCP produced enhanced dose-dependent reduction in allodynic responses with synergistic effects observed for cold hypersensitivity in both sexes and additive effects for tactile hypersensitivity in males. Antinociceptive effects of both individual and combined treatment were generally less robust in females than males. CBD:BCP co-administration also partially reduced morphine-seeking behavior in a conditioned place preference (CPP) test. Minimal cannabinoidergic side effects were observed with high doses of the combination. The antinociceptive effects of the CBD:BCP co-administration were not altered by either CB2 or μ-opioid receptor antagonist pretreatment but, were nearly completely blocked by CB1 antagonist AM251. Since neither CBD or BCP are thought to mediate antinociception via CB1 activity, these findings suggest a novel CB1 interactive mechanism between these two phytocannabinoids in the SCI pain state. Together, these findings suggest that CBD:BCP co-administration may provide a safe and effective treatment option for the management of chronic SCI pain.Mutation of regulatory phosphorylation sites in PFKFB2 does not affect the anti-fibrotic effect of metformin in the kidneyGeoff HarleyMarina KaterelosKurt GleichMardiana LeePeter F. MountDavid A. Power10.1371/journal.pone.02807922023-02-09T14:00:00Z2023-02-09T14:00:00Z<p>by Geoff Harley, Marina Katerelos, Kurt Gleich, Mardiana Lee, Peter F. Mount, David A. Power</p>
The anti-fibrotic effect of metformin has been widely demonstrated. Fibrosis in the kidney after injury is associated with reduced expression of genes involved in both fatty acid and glycolytic energy metabolism. We have previously reported that the anti-fibrotic effect of metformin requires phosphoregulation of fatty acid oxidation by AMP-activated protein kinase (AMPK). To determine whether metformin also acts via regulation of glycolysis, we mutated regulatory phosphosites in the PFKFB2 isoform of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase (PFKFB2), a key regulator of glycolysis in the kidney. Mice with inactivating knockin (KI) mutations of the phosphorylation sites in PFKFB2 (PFKFB2 KI mice), which reduces the ability to increase the rate of glycolysis following stimulation, were used. Metformin was administered via drinking water to mice with a unilateral ureteric obstruction (UUO) model of renal fibrosis. In the PFKFB2 KI mice treated with metformin, there was decreased fibrosis and macrophage infiltration following UUO as assessed by Western blot for fibronectin and RT-PCR for α-smooth muscle actin, collagen 3, and F4.80, and confirmed by histology. Expression of the inducible PFKFB3 isoform was increased with metformin in UUO in both WT and PFKFB2 KI mice. There was no significant difference between WT and PFKFB2 KI mice treated with metformin in the degree of fibrosis following UUO in any of the Western blot or RT-PCR parameters that were measured. These data show that inhibition of the regulation of glycolysis by PFKFB2 does not diminish the anti-fibrotic effect of metformin in a model of renal fibrosis.Inhibition of RIP1-RIP3-mediated necroptosis attenuates renal fibrosis via Wnt3α/β-catenin/GSK-3β signaling in unilateral ureteral obstructionShang Guo PiaoJun DingXue Jing LinQi Yan NanMei Ying XuanYu Ji JiangHai Lan ZhengJi Zhe JinCan Li10.1371/journal.pone.02741162022-10-12T14:00:00Z2022-10-12T14:00:00Z<p>by Shang Guo Piao, Jun Ding, Xue Jing Lin, Qi Yan Nan, Mei Ying Xuan, Yu Ji Jiang, Hai Lan Zheng, Ji Zhe Jin, Can Li</p>
Renal fibrosis represents the final common outcome of chronic kidney disease of virtually any etiology. However, the mechanism underlying the evolution of renal fibrosis remains to be addressed. This study sought to clarify whether RIP1-RIP3-mediated necroptosis is involved in renal fibrosis via Wnt3α/β-catenin/GSK-3β signaling in vitro and in a rat model of unilateral ureteral obstruction (UUO). Rats with UUO were administered RIP inhibitors (necrostatin-1 or GSK872) or β-catenin/TCF inhibitor ICG-001 daily for 7 consecutive days. UUO caused significant renal tubular necrosis and overexpression of RIP1-RIP3-MLKL axis proteins, and was accompanied by activation of the NLRP3 inflammasome and renal fibrosis. Oxidative stress caused by UUO was closely associated with endoplasmic reticulum stress and mitochondrial dysfunction, which resulted in apoptotic cell death via Wnt3α/β-catenin/GSK-3β signaling. All of these effects were abolished by an RIP inhibitor (necrostatin-1 or GSK872) or ICG-001. In H<sub>2</sub>O<sub>2</sub>-treated HK-2 cells, both RIP inhibitor and ICG-001 decreased intracellular reactive oxygen species production and apoptotic cells, but increased cell viability. Activated Wnt3α/β-catenin/GSK-3β signaling was decreased by either RIP inhibitor or ICG-001. Our findings suggest that RIP1-RIP3-mediated necroptosis contributes to the development of renal fibrosis via Wnt3α/β-catenin/GSK-3β signaling in UUO and may be a therapeutic target for protection against renal scarring of other origins.Resistin production does not affect outcomes in a mouse model of acute surgical sepsisAnthony S. BonaviaZissis C. ChroneosVictor Ruiz-VelascoCharles H. Lang10.1371/journal.pone.02652412022-03-14T14:00:00Z2022-03-14T14:00:00Z<p>by Anthony S. Bonavia, Zissis C. Chroneos, Victor Ruiz-Velasco, Charles H. Lang</p>
Introduction <p>Because of the strong correlation between the blood concentration of circulating resistin and the illness severity of septic patients, resistin has been proposed as a mediator of sepsis pathophysiology. <i>In vitro</i> data indicate that human resistin directly impairs neutrophil migration and intracellular bacterial killing, although the significance of these findings <i>in vivo</i> remain unclear.</p> Objective <p>The objectives of the present study were: (1) to validate the expression of human resistin in a clinically relevant, murine model of surgical sepsis, (2) to assess how sepsis-induced changes in resistin correlate with markers of infection and organ dysfunction, and (3) to investigate whether the expression of human resistin alters immune function or disease outcomes <i>in vivo</i>.</p> Methods <p>107 male, C57BL/6 mice transgenic for the human resistin gene and its promoter elements (<i>Retn</i><sup>+/−/−</sup>, or Retn+) were generated on a <i>Retn</i><sup>−/−</sup> (mouse resistin knockout, or Rko) background. Outcomes were compared between age-matched transgenic and knockout mice. Acute sepsis was defined as the initial 24 h following cecal ligation and puncture (CLP). Physiologic and laboratory parameters correlating to the human Sequential Organ Failure Assessment (SOFA) Score were measured in mice, and innate immune cell number/function in the blood and peritoneal cavity were assessed.</p> Results <p>CLP significantly increased circulating levels of human resistin. The severity of sepsis-induced leukopenia was comparable between Retn+ and Rko mice. Resistin was associated with increased production of neutrophil reactive oxygen species, a decrease in circulating neutrophils at 6 h and an increase in peritoneal Ly6C<sup>hi</sup> monocytes at 6 h and 24 h post-sepsis. However, intraperitoneal bacterial growth, organ dysfunction and mouse survival did not differ with resistin production in septic mice.</p> Significance <p><i>Ex vivo</i> resistin-induced impairment of neutrophil function do not appear to translate to increased sepsis severity or poorer outcomes <i>in vivo</i> following CLP.</p>