The Roles of RhoA/ROCK/NF-κB Pathway in Microglia Polarization Following Ischemic Stroke.

Ischemic stroke is the leading cause of death and disability worldwide. Nevertheless, there still lacks the effective therapies for ischemic stroke. Microglia are resident macrophages of the central nervous system (CNS) and can initiate immune responses and monitor the microenvironment. Microglia are activated and polarize into proinflammatory or anti­inflammatory phenotype in response to various brain injuries, including ischemic stroke. Proinflammatory microglia could generate immunomodulatory mediators, containing cytokines and chemokines, these mediators are closely associated with secondary brain damage following ischemic stroke. On the contrary, anti-inflammatory microglia facilitate recovery following stroke. Regulating the activation and the function of microglia is crucial in exploring the novel treatments for ischemic stroke patients. Accumulating studies have revealed that RhoA/ROCK pathway and NF-κB are famous modulators in the process of microglia activation and polarization. Inhibiting these key modulators can promote the polarization of microglia to anti-inflammatory phenotype. In this review, we aimed to provide a comprehensive overview on the role of RhoA/ROCK pathway and NF-κB in the microglia activation and polarization, reveal the relationship between RhoA/ROCK pathway and NF-κB in the pathological process of ischemic stroke. In addition, we likewise discussed the drug modulators targeting microglia polarization.

Neuroinflammation and Post-Stroke Depression: Focus on the Microglia and Astrocytes.

Post-stroke depression (PSD), a frequent and disabling complication of stroke, has a strong impact on almost thirty percent of stroke survivors. The pathogenesis of PSD is not completely clear so far. Neuroinflammation following stroke is one of underlying mechanisms that involves in the pathophysiology of PSD and plays an important function in the development of depression and is regarded as a sign of depression. During the neuroinflammation after ischemic stroke onset, both astrocytes and microglia undergo a series of morphological and functional changes and play pro-inflammatory or anti-inflammatory effect in the pathological process of stroke. Importantly, astrocytes and microglia exert dual roles in the pathological process of PSD due to the phenotypic transformation. We summarize the latest evidence of neuroinflammation involving in PSD in this review, focus on the phenotypic transformation of microglia and astrocytes following ischemic stroke and reveal the dual roles of both microglia and astrocytes in the PSD via modulating the neuroinflammation.

Crosstalk Among Glial Cells in the Blood-Brain Barrier Injury After Ischemic Stroke.

Blood-brain barrier (BBB) is comprised of brain microvascular endothelial cells (ECs), astrocytes, perivascular microglia, pericytes, neuronal processes, and the basal lamina. As a complex and dynamic interface between the blood and the central nervous system (CNS), BBB is responsible for transporting nutrients essential for the normal metabolism of brain cells and hinders many toxic compounds entering into the CNS. The loss of BBB integrity following stroke induces tissue damage, inflammation, edema, and neural dysfunction. Thus, BBB disruption is an important pathophysiological process of acute ischemic stroke. Understanding the mechanism underlying BBB disruption can uncover more promising biological targets for developing treatments for ischemic stroke. Ischemic stroke-induced activation of microglia and astrocytes leads to increased production of inflammatory mediators, containing chemokines, cytokines, matrix metalloproteinases (MMPs), etc., which are important factors in the pathological process of BBB breakdown. In this review, we discussed the current knowledges about the vital and dual roles of astrocytes and microglia on the BBB breakdown during ischemic stroke. Specifically, we provided an updated overview of phenotypic transformation of microglia and astrocytes, as well as uncovered the crosstalk among astrocyte, microglia, and oligodendrocyte in the BBB disruption following ischemic stroke.

Flavonoids and ischemic stroke-induced neuroinflammation: Focus on the glial cells.

Ischemic stroke is one of the most cases worldwide, with high rate of morbidity and mortality. In the pathological process of ischemic stroke, neuroinflammation is an essential process that defines the functional prognosis. After stroke onset, microglia, astrocytes and the infiltrating immune cells contribute to a complicated neuroinflammation cascade and play the complicated roles in the pathophysiological variations of ischemic stroke. Both microglia and astrocytes undergo both morphological and functional changes, thereby deeply participate in the neuronal inflammation via releasing pro-inflammatory or anti-inflammatory factors. Flavonoids are plant-specific secondary metabolites and can protect against cerebral ischemia injury via modulating the inflammatory responses. For instances, quercetin can inhibit the expression and release of pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α, IL-6 and IL-1ß, in the cerebral nervous system (CNS). Apigenin and rutin can promote the polarization of microglia to anti-inflammatory genotype and then inhibit neuroinflammation. In this review, we focused on the dual roles of activated microglia and reactive astrocyte in the neuroinflammation following ischemic stroke and discussed the anti-neuroinflammation of some flavonoids. Importantly, we aimed to reveal the new strategies for alleviating the cerebral ischemic stroke.

The role of RhoA/ROCK pathway in the ischemic stroke-induced neuroinflammation.

It is widely known that ischemic stroke is the prominent cause of death and disability. To date, neuroinflammation following ischemic stroke represents a complex event, which is an essential process and affects the prognosis of both experimental stroke animals and stroke patients. Intense neuroinflammation occurring during the acute phase of stroke contributes to neuronal injury, BBB breakdown, and worse neurological outcomes. Inhibition of neuroinflammation may be a promising target in the development of new therapeutic strategies. RhoA is a small GTPase protein that activates a downstream effector, ROCK. The up-regulation of RhoA/ROCK pathway possesses important roles in promoting the neuroinflammation and mediating brain injury. In addition, nuclear factor-kappa B (NF-κB) is another vital regulator of ischemic stroke-induced neuroinflammation through regulating the functions of microglial cells and astrocytes. After stroke onset, the microglial cells and astrocytes are activated and undergo the morphological and functional changes, thereby deeply participate in a complicated neuroinflammation cascade. In this review, we focused on the relationship among RhoA/ROCK pathway, NF-κB and glial cells in the neuroinflammation following ischemic stroke to reveal new strategies for preventing the intense neuroinflammation.

H2S-RhoA/ROCK Pathway and Glial Cells in Axonal Remyelination After Ischemic Stroke.

Ischemic stroke is one of the main reasons of disability and death. Stroke-induced functional deficits are mainly due to the secondary degeneration of the white matter characterized by axonal demyelination and injury of axon-glial integrity. Enhancement of the axonal regeneration and remyelination could promote the neural functional recovery. However, cerebral ischemia-induced activation of RhoA/Rho kinase (ROCK) pathway plays a crucial and harmful role in the process of axonal recovery and regeneration. Inhibition of this pathway could promote the axonal regeneration and remyelination. In addition, hydrogen sulfide (H2S) has the significant neuroprotective role during the recovery of ischemic stroke via inhibiting the inflammatory response and oxidative stress, regulating astrocyte function, promoting the differentiation of endogenous oligodendrocyte precursor cells (OPCs) to mature oligodendrocyte. Among all of these effects, promoting the formation of mature oligodendrocyte is a crucial part of axonal regeneration and remyelination. Furthermore, numerous studies have uncovered the crosstalk between astrocytes and oligodendrocyte, microglial cells and oligodendrocyte in the axonal remyelination following ischemic stroke. The purpose of this review was to discuss the relationship among H2S, RhoA/ROCK pathway, astrocytes, and microglial cells in the axonal remyelination following ischemic stroke to reveal new strategies for preventing and treating this devastating disease.

H2S-mediated inhibition of RhoA/ROCK pathway and noncoding RNAs in ischemic stroke.

Ischemic stroke is one of major causes of disability. In the pathological process of ischemic stroke, the up-regulation of Ras homolog gene family, member A (RhoA) and its downstream effector, Ras homolog gene family (Rho)-associated coiled coil-containing kinase (ROCK), contribute to the neuroinflammation, blood-brain barrier (BBB) dysfunction, neuronal apoptosis, axon growth inhibition and astrogliosis. Accumulating evidences have revealed that hydrogen sulphide (H2S) could reduce brain injury in animal model of ischemic stroke via inhibiting the RhoA/ROCK pathway. Recently, noncoding RNAs (ncRNAs) such as circular RNAs (circRNAs), long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) have attracted much attention because of their essential role in adjusting gene expression both in physiological and pathological conditions. Numerous studies have uncovered the role of RhoA/ROCK pathway and ncRNAs in ischemic stroke. In this review, we focused on the role of H2S, RhoA/ROCK pathway and ncRNAs in ischemic stroke and aimed to reveal new strategies for preventing and treating this devastating disease.

RhoA/ROCK signaling pathway and astrocytes in ischemic stroke.

Ischemic stroke is one of the most common and undertreated cerebral diseases with high mortality and disability rate. Various intrinsic and extrinsic factors regulate the onset, severity, and progression of ischemic stroke. As an integral part of the neuronal glia system, astrocytes provide many housekeeping functions in nervous system, and perform multiple functions both beneficial and detrimental for neuronal survival after ischemic stroke. In addition, the small GTPase Rho and its downstream Rho kinase (ROCK) are associated with various neuronal functions such as dendrite development, migration and axonal extension, and numerous central nervous system (CNS) diseases. The aim of this review is to summarize the role of RhoA/ROCK signaling pathway and astrocytes on neurological function after ischemic stroke. We also discuss the interaction of RhoA/ROCK signaling pathway and astrocytes on the tissue repair after brain injury.

Neuroprotective roles of total flavones of Camellia on early brain injury andcognitive dysfunction following subarachnoid hemorrhage in rats.

The present study was undertaken to explore the role of total flavones of Camellia (TFC) on cerebral injury following subarachnoid hemorrhage (SAH) in rats. We showed that the increase of malondialdehyde (MDA) level in brain tissues, leakages of neuron-specifc enolase (NSE) and lactate dehydrogenase (LDH) from brain tissues to serum at 48 h after SAH were significantly blocked by TFC treatment. Besides, TFC treatment could reduce brain edema and the Bax/Bcl-2 ratio in hippocampal tissues at mRNA and protein levels at 48 h after SAH. In addition, and the reduction of neurological scores at 7d after SAH were significantly inhibited by TFC treatment. We next sought to demonstrate the role of TFC on cognitive rehabilitation and the tau phosphorylation in hippocampal tissues at 30d after SAH. Not surprisingly, cognitive dysfunction and the upregulation of tau phosphorylation at Ser262 (p-tau-Ser262) in hippocampal tissues were markedly reduced by TFC treatment. These findings suggested that TFC has protective effect on SAH-induced EBI and subsequent cognitive dysfunction, which may be related to downregulating the Bax/Bcl-2-related apoptosis pathway and inhibition of tau phosphorylation.

Distinct Roles of ROCK1 and ROCK2 on the Cerebral Ischemia Injury and Subsequently Neurodegenerative Changes.

Cerebral ischemic injury is one of the main causes of adult disability and death. Although significant progress has been made, cerebral ischemia continues to be a major risk to public health worldwide. The Rho kinase (ROCK) signaling pathway has been reported to be significantly involved in many mechanisms of cerebral injury. Although ROCK is ubiquitously expressed in all tissues, ROCK2 subtype expression in brain and the spinal cord is more abundant and improves with age. This makes it a promising target for new therapeutic approaches. In this article, we review the current knowledge on the involvement of ROCK in cerebral ischemia injury and neurodegenerative changes after cerebral injury. After a detailed description of the mechanism of ROCK involvement in axonal regeneration and synaptic function, different roles of ROCK1 and ROCK2 in neurons under physiological and pathological conditions are compared and discussed. In addition, different functions of genetic and pharmacological inhibitions of ROCK1 and ROCK2 on cerebral injury are discussed.

Determining the environmental impact of material hauling with wheel loaders during earthmoving operations.

A method has been developed to estimate the environmental impact of wheel loaders used in earthmoving operations. The impact is evaluated in terms of energy use and emissions of air pollutants (CO2, CO, NOx, CH4, VOC, and PM) based on the fuel consumption per cubic meter of hauled material. In addition, the effects of selected operational factors on emissions during earthmoving activities were investigated to provide better guidance for practitioners during the early planning stage of construction projects. The relationships between six independent parameters relating to wheel loaders and jobsite conditions (namely loader utilization rates, loading time, bucket payload, horsepower, load factor, and server capacity) were analyzed using artificial neural networks, machine performance data from manufacturer's handbooks, and discrete event simulations of selected earthmoving scenarios. A sensitivity analysis showed that the load factor is the largest contributor to air pollutant emissions, and that the best way to minimize environmental impact is to maximize the wheel loaders' effective utilization rates. The new method will enable planners and contractors to accurately assess the environmental impact of wheel loaders and/or hauling activities during earthmoving operations in the early stages of construction projects. Implications: There is an urgent need for effective ways of benchmarking and mitigating emissions due to construction operations, and particularly those due to construction equipment, during the pre-construction phase of construction projects. Artificial Neural Networks (ANN) are shown to be powerful tools for analyzing the complex relationships that determine the environmental impact of construction operations and for developing simple models that can be used in the early stages of project planning to select machine configurations and work plans that minimize emissions and energy consumption. Using such a model, it is shown that the fuel consumption and emissions of wheel loaders are primarily determined by their engine load, utilization rate, and bucket payload. Moreover, project planners can minimize the environmental impact of wheel loader operations by selecting work plans and equipment configurations that minimize wheel loaders' idle time and avoid bucket payloads that exceed the upper limits specified by the equipment manufacturer.

Establishment of competitive binding assay to detect and differentiate hepatitis E virus infection.

INTRODUCTION AND OBJECTIVES: This study was undertaken to demonstrate a promising approach for detection and differentiation the serum immunoglobulin G (IgG) against hepatitis E virus (anti-HEV IgG) using a competitive binding assay established with known genotype-specific monoclonal antibodies (mAbs) 2B1 and 4C5. MATERIALS AND METHODS: The mAb 2B1 derived from genotype 1 hepatitis E virus (HEV) antigen and specifically reacted with genotype 1, 2 antigens; 4C5 induced by genotype 4 HEV antigen was specific to genotypes 3, 4 antigens. The 2B1 and 4C5 were labeled with Horseradish peroxidase (HRP), respectively. Subsequently, the titers of coated antigens and HRP-conjugated mAbs for establishment of competitive binding assay were determined by enzyme linked immunosorbent assay (ELISA). And then, the competitive binding assay was performed to assess the inhibition percentage of mAbs binding to antigens inhibited by different genotypes anti-HEV IgG. RESULTS: The results of competitive binding assay revealed that genotype 1 anti-HEV IgG could inhibit the binding of mAb 2B1 to genotype 1 antigen more strongly than that of mAb 4C5 to genotype 4 antigen. Whereas, the genotype 3 or 4 anti-HEV IgG could inhibit the binding of mAb 4C5 to genotype 4 antigen more remarkably than that of mAb 2B1 to genotype 1 antigen. CONCLUSIONS: These findings provided us a valuable approach for detection and differentiation the HEV infection derived from genotypes 1, 2 (human) or genotypes 3, 4 (zoonosis).

Protective effect of extract of the Camellia japonica L. on cerebral ischemia-reperfusion injury in rats.

OBJECTIVE: We investigated the protective effect of the extract of the Camellia japonica L. flower on cerebral ischemia-reperfusion injury in rats. METHODS: The rat ischemia-reperfusion injury was induced by middle cerebral artery occlusion for 90 minutes and reperfusion for 48 hours. The animals received an intravenous injection once a day of 20, 40, 80 mg/kg extract of C. japonica for three consecutive days before the ischemia reperfusion. The learning and memory function, the infarct volume, serum malondialdehyde (MDA) level and lactate dehydrogenase activity, and extravasation of immunoglobulin G (IgG) into cerebral parenchyma were assessed as the cell damage index. RESULTS: Pretreatment with extract of C. japonica markedly reduced the infarct volume, serum malondialdehyde level and lactate dehydrogenase activity, and markedly inhibited the extravasation of IgG. Moreover, pretreatment with extract of C. japonica may also inhibit the learning and memory deficits induced by an ischemia-reperfusion injury. CONCLUSION: It was concluded that pretreatment with extract of C. japonica has a protective effect on cerebral ischemia-reperfusion injury in rats.

Protective effect of extract of the Camellia japonica L. on cerebral ischemia-reperfusion injury in rats / Efeito protetor do extrato da Camellia japonica L. na lesão de reperfusão isquêmica cerebral em ratos

ABSTRACT Objective: We investigated the protective effect of the extract of the Camellia japonica L. flower on cerebral ischemia-reperfusion injury in rats. Methods: The rat ischemia-reperfusion injury was induced by middle cerebral artery occlusion for 90 minutes and reperfusion for 48 hours. The animals received an intravenous injection once a day of 20, 40, 80 mg/kg extract of C. japonica for three consecutive days before the ischemia reperfusion. The learning and memory function, the infarct volume, serum malondialdehyde (MDA) level and lactate dehydrogenase activity, and extravasation of immunoglobulin G (IgG) into cerebral parenchyma were assessed as the cell damage index. Results: Pretreatment with extract of C. japonica markedly reduced the infarct volume, serum malondialdehyde level and lactate dehydrogenase activity, and markedly inhibited the extravasation of IgG. Moreover, pretreatment with extract of C. japonica may also inhibit the learning and memory deficits induced by an ischemia-reperfusion injury. Conclusion: It was concluded that pretreatment with extract of C. japonica has a protective effect on cerebral ischemia-reperfusion injury in rats.

RESUMO Objetivo: Investigamos o efeito protetor do extrato da flor de Camellia japonica L. (ECJ) na lesão de reperfusão isquêmica cerebral (I/R) em ratos. Métodos: A lesão de I/R de rato foi induzida por uma oclusão da artéria cerebral média por 90 minutos e reperfusão por 48 horas. Os animais receberam uma injeção intravenosa uma vez ao dia de 20, 40, 80 mg/kg de ECJ por três dias consecutivos antes da I/R. A função de aprendizagem e memória, o volume do infarto, o nível sérico de malondialdeído (MDA), a atividade da desidrogenase láctica e o extravasamento de imunoglobulina (IgG) no parênquima cerebral foram avaliados como índices de dano celular. Resultados: O pré-tratamento com ECJ reduziu acentuadamente o volume do infarto, o nível sérico de MDA e a atividade da desidrogenase láctica, e inibiu marcadamente o extravasamento de IgG. Além disso, o pré-tratamento com ECJ também poderia inibir os déficits de aprendizado e memória induzidos pela lesão de I/R. Conclusão: O pré-tratamento com ECJ tem um efeito protetor contra lesão cerebral de I/R em ratos.

Establishment of a Competitive Binding Assay Identifying the Different Characteristics of Neutralizing Epitopes of Hepatitis E Virus.

AIMS: To confirm the different characteristics of genotype-specific and common neutralizing epitopes of hepatitis E virus (HEV). METHODS: A competitive binding assay was established with known genotype-common neutralizing monoclonal antibodies (mAbs) 3G1 and 5G5 as well as genotype-specific neutralizing mAbs 2B1 and 4C5. HEV ORF2 recombinant p166W01 derived from genotype 1 and p166Chn derived from genotype 4 were used as coated antigens, to determine whether the mAbs recognize independent, similar, or overlapping epitopes. mAbs were produced, purified, and conjugated with horseradish peroxidase (HRP). HRP-conjugated 2B1 could react only with p166W01 but not p166Chn, HRP-conjugated 4C5 could react only with p166Chn but not p166W01, while HRP-conjugated 3G1 and 5G5 could react both with p166W01 and p166Chn. Thus, competitive binding assays were performed successively using p166W01 and p166Chn antigen. RESULTS AND CONCLUSION: The results of competitive binding assays revealed that the binding of HRP-conjugated 2B1 to p166W01 could not be inhibited by 5G5 or 3G1. Similarly, the binding of HRP-conjugated 4C5 to p166Chn could not be inhibited by 5G5 or 3G1. However, the mAbs 5G5 and 3G1 blocked each other's binding to p166W01 and p166Chn, suggesting that common and genotype-specific neutralizing mAbs recognize independent epitopes.