YAP1 Recognizes Inflammatory and Mechanical Cues to Exacerbate Benign Prostatic Hyperplasia via Promoting Cell Survival and Fibrosis.

Chronic prostatic inflammation promotes cell survival and fibrosis, leading to benign prostatic hyperplasia (BPH) with aggravated urinary symptoms. It is investigated whether yes-associated protein 1 (YAP1), an organ size controller and mechanical transductor, is implicated in inflammation-induced BPH. The correlation between YAP1 expression and fibrosis in human and rat BPH specimens is analyzed. Furthermore, the effects of YAP1 activation on prostatic cell survival and fibrosis, as well as the underlying mechanism, are also studied. As a result, total and nuclear YAP1 expression, along with downstream genes are significantly upregulated in inflammation-associated human and rat specimens. There is a significant positive correlation between YAP1 expression and the severity of fibrosis or clinical performance. YAP1 silencing suppresses cell survival by decreasing cell proliferation and increasing apoptosis, and alleviates fibrosis by reversing epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) deposition in prostatic BPH-1 and WPMY-1 cells. Mechanistically, inflammatory stimulus and rigid matrix stiffness synergistically activate the RhoA/ROCK1 pathway to provoke cytoskeleton remodeling, thereby promoting YAP1 activation to exacerbate BPH development. Overall, inflammation-triggered mechanical stiffness reinforcement activates the RhoA/ROCK1/F-actin/YAP1 axis, thereby promoting prostatic cell survival and fibrosis to accelerate BPH progression.

The Effects of ROCK Inhibitor on Prevention of Dexamethasone-Induced Glaucoma Phenotype in Human Trabecular Meshwork Cells.

Purpose: This study investigated the effects of dexamethasone (Dex) on human trabecular meshwork (TM) cells, a model of glucocorticoid-induced glaucoma, and evaluated the impact of ripasudil (Rip) as a co-delivery or sequential dosing strategy. Methods: In vitro experiments were conducted to assess the effects of Dex and Rip on TM cells. Confocal microscopy was used to evaluate the impact of Dex and Rip on F-actin staining signals. Contractility of the TM cells upon Dex and Rip treatment mimicking co-delivery and sequential delivery was quantified using collagen gel contraction assay. Transepithelial electrical resistance (TEER) values and fluorescein isothiocyanate (FITC)-dextran permeability were also measured to assess the impact of Dex and Rip on TM cells. Results: Dex and Rip did not exhibit cytotoxicity at the maximum tested concentration (20 µM). Dex-treated TM cells exhibited higher F-actin staining signals compared to controls, which were reduced when co-treated with Rip. Rip inhibited Dex-induced collagen gel contraction activity in both co-delivery and sequential treatments. Dex resulted in increased TEER values as the dose increased, whereas TEER values were maintained when co-treated with Rip. Conclusions: Co-delivery of Rip has the potential to prevent glaucoma symptoms when patients are treated with Dex. This study highlights the importance of identifying strategies to reduce the side effects of prolonged use of glucocorticoids, such as Dex, in the treatment of various diseases. Translational Relevance: This study demonstrates the potential of co-delivering ripasudil with dexamethasone to mitigate glucocorticoid-induced ocular hypertension and a secondary glaucoma that resembles primary open-angle glaucoma, providing insights for the development of novel preventive strategies in clinical care.

Triptolide Reduces MDA-MB-231 Cell Metastasis by Attenuating Epithelial-Mesenchymal Transition through the ROCK/PTEN/Akt Axis.

Triple-negative breast cancer (TNBC) is a highly heterogeneous and invasive subtype of breast cancer. The prognosis of TNBC is poor because of its high distant metastasis rate. Triptolide is a type of diterpene trioxide natural compound with potential anti-tumor activities. This study explored the metastatic inhibitory effect of triptolide on MDA-MB-231 cells and its underlying mechanism. Triptolide suppressed cell proliferation and induced cell apoptosis in a time- and dose-dependent manner. Low doses of triptolide (0-8 nM) reduced the migration and invasion capabilities of MDA-MB-231 cells. Triptolide decreased ROCK1, p-Akt, N-cadherin, vimentin and MMP-9 expressions, but increased PTEN and E-cadherin expressions on protein and mRNA levels. Furthermore, the down-regulation of ROCK1 expression in MDA-MB-231 cells after being treated by triptolide could be rescued by ROCK1 specific inhibitor Y27632. Molecular docking showed that triptolide and Y27632 shared the same active center of ROCK1 protein. This article's findings taken together showed that ROCK1 is the primary target of triptolide, which can cause cell apoptosis and inhibit the epithelial-mesenchymal transition of MDA-MB-231 cells.

TNF-α evokes blood-brain barrier dysfunction through activation of Rho-kinase and neurokinin 1 receptor.

Ischaemic stroke, accompanied by neuroinflammation, impairs blood-brain barrier (BBB) integrity through a complex mechanism involving activation of both RhoA/Rho kinase/myosin light chain-2 and neurokinin 1 receptor (NK1R). Using an in vitro model of human BBB composed of brain microvascular endothelial cells (BMEC), astrocytes and pericytes, this study examined the potential contributions of these elements to BBB damage induced by elevated availability of pro-inflammatory cytokine, TNF-α. Treatment of human BMECs with TNF-α significantly enhanced RhoA activity and the protein expressions of Rho kinase and phosphorylated Ser19MLC-2 while decreasing that of NK1R. Pharmacological inhibition of Rho kinase by Y-27632 and NK1R by CP96345 neutralised the disruptive effects of TNF-α on BBB integrity and function as ascertained by reversal of decreases in transendothelial electrical resistance and increases in paracellular flux of low molecular weight permeability marker, sodium fluorescein, respectively. Suppression of RhoA activation, mitigation of actin stress fibre formation and restoration of plasma membrane localisation of tight junction protein zonula occludens-1 appeared to contribute to the barrier-protective effects of both Y-27632 and CP96345. Attenuation of TNF-α-mediated increases in NK1R protein expression in BMEC by Y-27632 suggests that RhoA/Rho kinase pathway acts upstream to NK1R. In conclusion, specific inhibition of Rho kinase in cerebrovascular conditions, accompanied by excessive release of pro-inflammatory cytokine TNF-α, helps preserve endothelial cell morphology and inter-endothelial cell barrier formation and may serve as an important therapeutic target.

Matrine Protects Intestinal Barrier Function via MicroRNA-155 Through ROCK1-Signaling Pathway.

BACKGROUND: The aim of this study was to investigate the protective effects on intestinal barrier function and tight junction by matrine via a small, non-coding RNA microRNA-155-signaling pathway. MATERIALS AND METHODS: Tight junction protein and target gene expressions were determined through microRNA-155 inhibition or overexpression of Caco-2 cell line with or without the treatment of matrine. In order to further verify the role of matrine, dextran sulfate sodium-induced mice colitis was treated by matrine. MicroRNA-155 and ROCK1 expressions were detected in clinical specimens of acute obstruction patients. RESULTS: Matrine could enhance the expression level of occludin, which might be inhibited by the overexpression of microRNA-155. After the transfection of the precursor of microRNA-155 into Caco-2 cells, ROCK1 expression was increased at both mRNA and protein levels. MicroRNA-155 inhibitor could decrease the ROCK1 expression after transfection. Furthermore, matrine could increase the permeability and decrease tight junction-associated proteins induced by dextran sulfate sodium-induced mice colitis. Clinical sample detection also found high levels of microRNA-155 in stercoral obstruction patients. CONCLUSIONS: Matrine could maintain the tight junction and protect the intestinal barrier from dysfunction. The molecular mechanism may be that matrine might inhibit microRNA-155 and increase the expression level of tight junction proteins.

Dietary protocatechuic acid redistributes tight junction proteins by targeting Rho-associated protein kinase to improve intestinal barrier function.

Inflammatory bowel disease (IBD) is continuously increasing globally and caused by intestinal barrier dysfunction. Although protocatechuic acid (PCA) has a protective effect on colitis, the molecular mechanisms underlying its contribution to intestinal barrier function remain unknown. Transepithelial electrical resistance (TEER) and FITC-dextran permeability measurements reveled that PCA suppresses lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α-induced increase in intestinal permeability; zonula occludens (ZO)-1 and claudin-2 redistribution was also suppressed in the epithelial cell membranes of differentiated Caco-2 cells. PCA was found to directly bind Rho-associated coiled-coil containing protein kinase (ROCK), subsequently suppressing myosin light chain (MLC) phosphorylation. Notably, PCA binds ROCK to a similar degree as Y27632, a selective ROCK inhibitor. Orally administering PCA (5 or 25 mg per kg per day) to C57BL/6 mice alleviated the 3% dextran sulfate sodium (DSS)-induced colitis symptoms including reduced colon length, disrupted intestinal barrier structure, and increased proinflammatory cytokines expressions, such as interleukin (IL)-1ß, TNF-α, and IL-6. Furthermore, orally administering PCA suppressed DSS-induced ZO-1 and claudin-2/4 redistribution in mice colon membrane fractions. Therefore, PCA may serve as a promising nutraceutical to improve gut health and alleviate IBD by maintaining intestinal barrier function in vitro and in vivo.

Effects of Ripasudil, a Rho-Kinase Inhibitor, on Scar Formation in a Mouse Model of Filtration Surgery.

PURPOSE: Glaucoma is a leading cause of blindness worldwide. Characteristic changes occur in the optic nerve and visual field of patients with glaucoma; optic nerve damage can be mitigated by lowering intraocular pressure. Treatment modalities include drugs and lasers; filtration surgery is necessary for patients with insufficient intraocular pressure reduction. Scar formation often contributes to glaucoma filtration surgery failure by increasing fibroblast proliferation and activation. Here, we examined the effects of ripasudil, a Rho-associated protein kinase (ROCK) inhibitor, on postoperative scar formation in human Tenon's fibroblasts. METHODS: Collagen gel contraction assays were used to compare contractility activity among ripasudil and other anti-glaucoma drugs. The effect of Ripasudil in combination with other anti-glaucoma drugs and transforming growth factor-ß (TGF-ß), latanoprost and timolol-induce contractions were also tested in this study. Immunofluorescence and Western blotting were used to study the expression of factors relating scarring formation. RESULTS: Ripasudil inhibited contraction in collagen gel assay and reduced α-smooth muscle actin (SMA) and vimentin (scar formation-related factors) expression, which was inversely promoted by latanoprost, timolol or TGF-ß. Ripasudil also inhibited contraction on TGF-ß, latanoprost and timolol-induced contraction. Furthermore, we investigated the effects of ripasudil on postoperative scarring in a mouse model; ripasudil suppressed postoperative scar formation by altering the expression of α-SMA and vimentin. CONCLUSIONS: These results suggest that ripasudil, ROCK inhibitor may inhibit excessive fibrosis after glaucoma filtering surgery vis inhibition the transdifferentiation of tenon fibroblast into myofibroblast and may have a potential effect as anti-scarring for glaucoma filtration surgery.

ROCK inhibitors in ophthalmology: A critical review of the existing clinical evidence.

Rho kinase (ROCK) inhibitors have emerged as a key therapeutic class of interest in ophthalmology over the last decade. Promising in vitro studies laid the foundations for the development of novel therapeutic agents that target the ROCK signalling pathway in ocular disease, with subsequent clinical trials supporting their use. Corneal endothelial disease, glaucoma, and vitreoretinal disease are the major pathologies in which ROCK inhibitors have been investigated to date. Ripasudil and netarsudil represent the current leaders in this pharmaceutical group, having been extensively validated and approved for use in glaucoma in some countries. Less substantial evidence exists for fasudil in ophthalmic use. ROCK inhibitors are also increasingly used in cultured endothelial cell grafting and as an adjunct to aid in endothelial cell migration and replication in Descemet's stripping procedures or Descemet's membrane injuries. This review has synthesised both established and emerging research to provide a practical guide to prescribing in this drug class. Drug efficacies, side effect profiles, and the demographic and clinical characteristics of appropriate drug candidates are discussed.

Effect of the Rho-Kinase/ROCK Signaling Pathway on Cytoskeleton Components.

The mechanical properties of cells are important in tissue homeostasis and enable cell growth, division, migration and the epithelial-mesenchymal transition. Mechanical properties are determined to a large extent by the cytoskeleton. The cytoskeleton is a complex and dynamic network composed of microfilaments, intermediate filaments and microtubules. These cellular structures confer both cell shape and mechanical properties. The architecture of the networks formed by the cytoskeleton is regulated by several pathways, a key one being the Rho-kinase/ROCK signaling pathway. This review describes the role of ROCK (Rho-associated coiled-coil forming kinase) and how it mediates effects on the key components of the cytoskeleton that are critical for cell behaviour.

Rho Kinase Inhibitor AR-12286 Reverses Steroid-Induced Changes in Intraocular Pressure, Effective Filtration Areas, and Morphology in Mouse Eyes.

Purpose: We investigated mechanisms of reduction of intraocular pressure (IOP) by Rho kinase inhibitor AR-12286 in steroid-induced ocular hypertension (SIOH). Methods: C57BL/6 mice (N = 56) were randomly divided into Saline, dexamethasone (DEX), DEX + AR-12286, and DEX-discontinuation (DEX-DC) groups. IOP was measured weekly during the first four weeks in all groups. Beginning at week 5, the DEX-DC group was followed without treatment until IOP returned to normal, and the other groups were treated as assigned with IOP measured every other day for another week. Fluorescent tracer was injected into the anterior chamber to visualize the outflow pattern in the trabecular meshwork (TM) and TM effective filtration area (EFA) was determined. Radial sections from both high- and low-tracer regions were processed for electron microscopy. Results: AR-12286 reduced IOP in SIOH mouse eyes in one day (P < 0.01). At the end of week 5, mean IOP in the DEX + AR-12286 group was ∼4 mm Hg lower than DEX group (P < 0.001) and ∼2 mm Hg lower than DEX-DC group (P < 0.05). After one-week AR-12286 treatment (P < 0.05) or five-week DC of DEX (P < 0.01), DEX-induced reduction of EFA was rescued and DEX-induced morphological changes in the TM were partially reversed. Conclusions: AR-12286 reversed steroid-induced morphological changes in the TM and reduced EFA, which correlated with reduced IOP in SIOH eyes. AR-12286 reduced IOP elevation in SIOH eyes more effectively than discontinuing DEX treatment even when accompanied by continuous DEX treatment. Therefore Rho kinase inhibitors may lower SIOH in patients who rely on steroid treatment.

Upregulation of PCSK9, rho kinase and cardiac troponin by Eucalyptus globulus leaf extract improves fructose-streptozotocin-induced diabetic cardiac dysfunction in rats.

CONTEXT: The effect of Eucalyptus globulus in diabetic cardiac dysfunction and the possible mechanisms involved have not been explored. OBJECTIVE: To evaluate the effect of ethanol leaf extract of E. globulus (NEE) on the cardiac function of fructose/streptozotocin-induced diabetic rats. MATERIALS AND METHODS: Type-2 diabetes was induced in rats with 10% fructose feeding for 14 days and an intraperitoneal administration of 40 mg/kg streptozotocin. Diabetic animals were treated with NEE (100-400 mg/kg) or 5 mg/kg glibenclamide orally for 21 days. Biochemical assays, histopathological examination and analyses of PCSK9, Rho kinase and Cardiac troponin expression were performed. RESULTS: The untreated diabetic group showed decreased expression of the genes, oxidative stress, dyslipidemia, increased activities of creatine kinase MB and lactate dehydrogenase, reduced nitric oxide level, and depletion of cardiomyocytes, which were reversed in NEE treated groups. CONCLUSIONS: Eucalyptus globulus ameliorated diabetic cardiac dysfunction through increased PCSK9, Rho kinase and Cardiac troponin expression.

Photodynamic therapy with zinc phthalocyanine enhances the anti-cancer effect of tamoxifen in breast cancer cell line: Promising combination treatment against triple-negative breast cancer?

Photodynamic therapy (PDT) is a light-based anti-neoplastic therapeutic approach. Growing evidence indicates that combining conventional anti-cancer therapies with PDT can be a promising approach to treat malignancies. Herein, we aimed to investigate anti-cancer effects of the combination treatment of zinc phthalocyanine (ZnPc)-PDT with tamoxifen (TA) on MDA-MB-231 cells (as a triple-negative breast cancer (TNBC) cell line). For this purpose, we investigated the cytotoxicity of TA and ZnPc-PDT on MDA-MB-231 cells performing the MTT assay. The effect of TA and ZnPc-PDT on the apoptosis of MDA-MB-231 cells was studied using Annexin V/PI and DAPI staining. The wound-healing assay, and colony formation assay were performed to study the effect of TA and ZnPc-PDT on the migration, and clonogenicity of MDA-MB-231 cells, respectively. The qRT-PCR was done to study the gene expression of caspase-8, caspase-9, caspase-3, ZEB1, ROCK1, SNAIL1, CD133, CD44, SOX2, and ABCG2 (ATP-binding cassette sub-family G member 2). Based on our results, monotherapies with TA and ZnPc-PDT can remarkably increase cell cytotoxicity effects, stimulate apoptosis via downregulating Bcl-2 and upregulating caspase-3 and caspase-9, inhibit migration via downregulating SNAIL1 and ZEB1, and suppress clonogenicity via downregulating SOX2 and CD44 in MDA-MB-231 cells. Besides, these monotherapies can downregulate the expression of ABCG2 in MDA-MB-231 cells. Nevertheless, the combination treatment can potentiate the above-mentioned anti-cancer effects compared to monotherapy with TA. Of interest, the combined treatment of TA with ZnPc-PDT can synergically increase cell cytotoxicity effects on MDA-MB-231 cells. In fact, synergistic effects were estimated by calculation of Combination Index (CI); that synergistic outcomes were observed in all groups. Also, this combination treatment can significantly upregulate the caspase-8 gene expression and downregulate ROCK1 and CD133 gene expression in MDA-MB-231 cells. Overall, our results show that ZnPc-PDT can more sensitize the MDA-MB-231 cells to TA treatment. Based on our knowledge and experiment, the synergistic effects of ZnPc-PDT and TA deserve further evaluation in cancer research.

The Therapeutic Role of Rho Kinase Inhibitor, Fasudil, on Pulmonary Hypertension; a Systematic Review and Meta-Analysis.

BACKGROUND: Pulmonary hypertension (PH) is a pathophysiological disorder, which involves multiple clinical conditions such as the upregulation of the Rho/ROCK signaling pathway. On the other hand, fasudil as a Rho kinase inhibitor has been investigated in the treatment of PH in some clinical studies. OBJECTIVES: The present systematic review and meta-analysis aimed to evaluate the human clinical trials regarding the efficacy of fasudil in the management of PH. METHODS: Databases were searched with pre-defined search terms, up to December 2021. Efficacy measures were such as mean pulmonary arterial pressure (mPAP), systolic PAP (sPAP), pulmonary vascular resistance (PVR), systolic vascular resistance (SVR) and cardiac index (CI). RESULTS: A total of 12 studies involving 575 PH patients were included in our research. Eight short-term trials and four mid-term trials were found (no clinical trials on the long-term effects). Short-term trials had a before-after study design and measuring pulmonary hemodynamic parameters' intervention revealed a statistically significant improvement of mPAP, sPAP, PVR, SVR, and CI in the meta-analysis of five eligible studies. Three mid-term trials also revealed improvement in some pulmonary hemodynamic parameters with fasudil and in another mid-term trial, fasudil significantly decreased rehospitalization and mortality in PH patients. No serious adverse effects with fasudil were reported in these trials. CONCLUSION: Fasudil therapy is efficacious and probably safe in the improvement of some hemodynamics in PH patients along short and mid-term periods. However, long-term randomized controlled trials comparing fasudil with placebo and other treatments are warranted for confirmation of these benefits.

Rho-kinase inhibitors: Role in corneal endothelial disorders.

PURPOSE: This exhaustive review of published literature aims to put forth the current understanding, and experiences of the role Rho-kinase inhibitors play in the management of corneal endothelial disorders. METHODS: A meticulous search for PubMed and Medline databases was carried out using keywords 'rho-kinase inhibitors', 'corneal endothelial disorders', 'rho-kinase inhibitors in corneal endothelial disorders', and 'fuchs endothelial corneal dystrophy'. The articles published in last five years relevant to the area of interest were selected and compiled in a chronological order to write this review. RESULTS: Published research shows the proven role of rho-kinase inhibitors in promoting endothelial proliferation and suppressing its apoptosis. It has shown to be effective in the treatment of endothelial diseases and endothelial cell loss due to other causes in small clinical studies. It has also shown to promote growth of human corneal endothelial cell culture suitable for transplantation. The side effects of topical rho-kinase inhibitors are minimal and self-limiting. CONCLUSION: Rho-kinase inhibitors show immense promise in providing a non-surgical therapy for treatment of endothelial disorders. If consistent results are demonstrated through large clinical trials then this can be a major breakthrough in treating endothelial decompensation and reduce our dependence on endothelial keratoplasty for the same.

Lysophosphatidic Acid Induced Apoptosis, DNA Damage, and Oxidative Stress in Spinal Cord Neurons by Upregulating LPA4/LPA6 Receptors.

Lysophosphatidic acid (LPA) has disruptive effects on lumbar spinal stenosis (LSS). Recently, LPA has been reported to be involved in spinal cord neuronal injury and toxicity, promoting the pathogenesis of LSS. However, the exact effects of LPA on spinal cord neurons remain unknown. The purpose of this study is to investigate the effects of LPA (18 : 1) on spinal cord neuronal cytotoxicity, apoptosis, DNA damage, and oxidative stress. After clinical detection of LPA secretion, spinal cord neurons were treated with LPA (18 : 1); cell viability was analyzed by MTT assay, and LDH leakage was detected by LDH kit; cell apoptosis was detected by flow cytometry; ROS production was measured by DCFDA staining and MitoSOX Red Staining; the activation of the Gα12/Gα13 signaling pathway was detected by serum response factor response element (SRF-RE) luciferase reporter gene; the relationship among LPA, LPA4/6, and ROCK was examined by western blotting. In spinal cord neurons treated with LPA (18 : 1), cellular activity decreased and LDH release increased. The Rho kinase inhibitor (Y-27632) can attenuate LPA-induced apoptosis, DNA damage, and oxidative stress in spinal cord neurons. Moreover mechanistic investigation indicated that LPA (18 : 1) activates Gα12/13-Rho-ROCK2-induced apoptosis, DNA damage, and oxidative stress in spinal cord neurons by upregulating LPA4/LPA6 receptors. Further, the Rho kinase inhibitor Y-27632 attenuates the effects of LPA by downregulating LPA4/LPA6 receptors. Taken together, the possible mechanism by which LPA secretion in LSS patients aggravates patient injury was further elucidated using an LPA-induced spinal cord neuronal injury cell model in vitro.

Tetrandrine Attenuates Podocyte Injury by Inhibiting TRPC6-Mediated RhoA/ROCK1 Pathway.

Tetrandrine (Tet), a compound found in a traditional Chinese medicine, presents the protective effect for kidney function. Our study is aimed at clarifying the efficacy and underlying mechanism of Tet on podocyte injury. In this study, podocyte injury was induced in rats with adriamycin (ADR), and MPC5 podocytes were constructed with TRPC6 overexpression. We found that Tet treatment reduced the levels of proteinuria, serum creatinine, and blood urea nitrogen and increased plasma albumin levels in ADR-induced rats. Tet reduced intracellular Ca2+ influx and apoptosis in MPC5 podocytes overexpressing TRPC6. Tet downregulated the expression of renal TRPC6, RhoA, and ROCK1 and upregulated the expression of synaptopodin; meanwhile, it reduced calcineurin activity in vivo and in vitro. In conclusion, Tet protects against podocyte by affecting TRPC6 and its downstream RhoA/ROCK1 signaling pathway.

ROCK2 inhibition: A futuristic approach for the management of Alzheimer's disease.

Neurons depend on mitochondrial functions for membrane excitability, neurotransmission, and plasticity. Mitochondrial dynamics are important for neural cell maintenance. To maintain mitochondrial homeostasis, lysosomes remove dysfunctional mitochondria through mitophagy. Mitophagy promotes mitochondrial turnover and prevents the accumulation of dysfunctional mitochondria. In many neurodegenerative diseases (NDDs), including Alzheimer's disease (AD), mitophagy is disrupted in neurons. Mitophagy is regulated by several proteins; recently, Rho-associated coiled-coil containing protein kinase 2 (ROCK2) has been suggested to negatively regulate the Parkin-dependent mitophagy pathway. Thus, ROCK2 inhibition may be a promising therapy for NDDs. This review summarizes the mitophagy pathway, the role of ROCK2 in Parkin-dependent mitophagy regulation, and mitophagy impairment in the pathology of AD. We further discuss different ROCK inhibitors (synthetic drugs, natural compounds, and gene therapy-based approaches) and examine their effects on triggering neuronal growth and neuroprotection in AD and other NDDs. This comprehensive overview of the role of ROCK in mitophagy inhibition provides a possible explanation for the significance of ROCK inhibitors in the therapeutic management of AD and other NDDs.

Apremilast exerts protective effects on stroke outcomes and blood-brain barrier (BBB) dysfunction through regulating Rho-associated protein kinase 2 expression.

AIMS: Stroke is a devastating event and a huge public health concern worldwide. Apremilast (APR) is a selective inhibitor of phosphodiesterase-4 involved in various neurological diseases, including stroke. However, the protective effects of APR on stroke have not been investigated. Here, we explored the effects of APR on stroke outcomes and blood-brain barrier (BBB) dysfunction using a middle cerebral artery occlusion (MCAO) stroke mice model. RESULTS: The results show that APR attenuated neurological injury in MCAO mice with decreased neurological deficit scores and infarct size, as well as increased hanging grip time. The increased BBB permeability and decreased expression of the tight junction protein Claudin-5 in MCAO mice were attenuated by APR treatment. APR treatment also mitigated neuroinflammation in MCAO mice, as shown by the decreased levels of inflammatory cytokines. In vitro assays also proved that APR ameliorated the oxygen/glucose deprivation/reoxygenation (OGD/R)-induced increase in endothelial permeability and restored the expression of Claudin-5 in bEnd.3 brain endothelial cells. Moreover, overexpression of ROCK2 in bEnd.3 cells abolished the protective effects of APR on endothelial permeability against OGD/R induction. CONCLUSION: Taken together, our results demonstrate that APR showed significant efficacy on ischemic stroke outcomes by alleviating enhanced BBB permeability and neuroinflammation by inhibiting ROCK2. These findings suggest a novel therapeutic window for ischemic stroke.

[Umbelliferone improves chronic hypoxia-induced pulmonary hypertension by inhibiting the RhoA/ROCK signaling pathway and autophagy].

This study aimed to investigate the effects of hypoxia on RhoA/Rho-kinase (ROCK) signaling pathway and autophagy in pulmonary artery smooth muscle cells (PASMCs), and to explore the underlying mechanism of Umbelliferone (Umb) in ameliorating chronic hypoxic pulmonary hypertension. PASMCs were cultured from Sprague-Dawley (SD) rats and randomly divided into control group, hypoxia group, hypoxia + Umb intervention group and normoxia + Umb intervention group. Alpha smooth muscle actin (α-SMA) and LC3 were assessed by immunofluorescence staining. Protein expression of RhoA, ROCK2, p-MYPT1, LC3-II, Beclin-1, p62, C-Caspase 3, Bax and Bcl-2 was analyzed by Western blotting. In in vivo study, SD rats were divided into control group, hypoxia group and hypoxia + Umb intervention group. Weight ratio of the right ventricle (RV)/left ventricle plus septum (LV+S) was detected, and pulmonary arterial morphological features were examined by HE staining. The results indicated that compared with the control group, the LC3-II/LC3-I ratio and expression of Beclin-1 were significantly increased, while p62 expression was significantly decreased, and the expressions of RhoA, ROCK2 and p-MYPT1 were significantly increased in PASMCs of hypoxia group (P < 0.05). The changes of LC3-II/LC3-I ratio, the expressions of Beclin-1, p62, RhoA, ROCK2 and p-MYPT1 in PASMCs were reversed by Umb treatment (P < 0.05). Consistently, the pulmonary arterial wall was thickened and the RV/(LV+S) ratio was increased in hypoxic rats, which were significantly improved by Umb treatment (P < 0.05). These results suggest that Umb can improve hypoxia-induced pulmonary hypertension by inhibiting the RhoA/ROCK signaling pathway and autophagy in PASMCs.