Double-Edged Effect of Hydroxychloroquine on Human Umbilical Cord-Derived Mesenchymal Stem Cells Treating Lupus Nephritis in MRL/lpr Mice.

Hydroxychloroquine (HCQ) and human umbilical cord-derived mesenchymal stem cells (UC-MSCs) were used to treat systemic lupus erythematosus (SLE), respectively. However, the effect of HCQ on UC-MSCs in lupus nephritis (LN) has not been investigated. In this study, HCQ and UC-MSCs were used in MRL/lpr mice. Surprisingly, although the treatment of both HCQ and UC-MSCs could ameliorate renal damage separately, the presence of HCQ decreased unexpectedly the therapeutic effects of UC-MSCs through interfering expression of IFN-γ. However, HCQ-pretreated UC-MSCs showed significant improvements of renal morphology and function more rapidly than that of UC-MSCs and HCQ alone. To test the role of HCQ in UC-MSCs, MRL/lpr mice and SLE patients' peripheral blood were used in vivo and in vitro. Results showed that after administration of UC-MSCs pretreated by HCQ, CXCR3 expression in renal tissues, serum IL-2, and IgM levels decreased significantly, and serum IL-10 level increased significantly. HCQ pretreatment caused a significant decrease of TNF-α and MCP-1 secretion and an increase of IL-1ß and CXCL10 release from UC-MSCs. Our results indicate that HCQ plays a double-edged role in UC-MSCs. It is necessary for clinical treatment to pre-evaluated concomitant application of UC-MSCs with HCQ. More importantly, the alterative expression of IFN-γ, the improvement of migration ability of UC-MSCs, the regulation of Th1/Th2 balance, and the changes of antibodies secretion in B cell might be involved in its mechanisms.

5-lipoxygenase activation is involved in the mechanisms of chronic hepatic injury in a rat model of chronic aluminum overload exposure.

We previously confirmed that rats overloaded with aluminum exhibited hepatic function damage and increased susceptibility to hepatic inflammation. However, the mechanism of liver toxicity by chronic aluminum overload is poorly understood. In this study, we investigated changes in the 5-lipoxygenase (5-LO) signaling pathway and its effect on liver injury in aluminum-overloaded rats. A rat hepatic injury model of chronic aluminum injury was established via the intragastric administration of aluminum gluconate (Al(3+) 200mg/kg per day, 5days a week for 20weeks). The 5-LO inhibitor, caffeic acid (10 and 30mg/kg), was intragastrically administered 1h after aluminum administration. Hematoxylin and eosin staining was used to visualize pathological changes in rat liver tissue. A series of biochemical indicators were measured with biochemistry assay or ELISAs. Immunochemistry and RT-PCR methods were used to detect 5-LO protein and mRNA expression in the liver, respectively. Caffeic acid administration protected livers against histopathological injury, decreased plasma ALT, AST, and ALP levels, decreased TNF-α, IL-6, IL-1ß and LTs levels, increased the reactive oxygen species content, and down-regulated the mRNA and protein expressions of 5-LO in aluminum overloaded rats. Our results indicate that 5-lipoxygenase activation is mechanistically involved in chronic hepatic injury in a rat model of chronic aluminum overload exposure and that the 5-LO signaling pathway, which associated with inflammation and oxidative stress, is a potential therapeutic target for chronic non-infection liver diseases.

Reduning injection combined with western medicine for pneumonia: A protocol for systematic review and meta-analysis.

BACKGROUND: Pneumonia is a common respiratory disease. In severe cases, it can induce cardiovascular disease and even life-threatening. In particular, pneumonia caused by the new coronavirus (SARS-CoV-2) that broke out at the end of 2019 has seriously affected the health of people in all countries. In recent years, it has been treated with the combination of traditional Chinese medicine (TCM) (such as Reduning injection) and Western medicine, and its mortality has decreased significantly. But their efficacy has not been scientifically and systematically assessed. Accordingly, it is essential to provide a systematized review program to estimate the efficacy and safety of Reduning injection combined with Western medicine to treat pneumonia. METHODS: The following databases are retrieved from start to September 2020: Pubmed, Cochrane Library, EMBASE, Web of Science, Chinese National Knowledge Infrastructure (CNKI), Wanfang database, the Chongqing VIP Chinese Science and Technology Periodical Database (VIP) databases, Chinese Biomedical Literature Database (CBM), and other databases, which are absorbed into clinical RCTs of pneumonia using western medicine alone or plus Reduning injections. The selection of studies, data extraction, and assessment of risk of bias will be performed independently by 2 reviewers. At the same time, Review Manager V.5.3.5 (Rev Man V.5.3.5) was used for bias risk assessment and data synthesis. RESULTS: The efficacy and safety of Reduning injection combined with western medicine in the treatment of pneumonia were evaluated in terms of overall effective rate, the patient's antipyretic time, antitussive time, rales disappearing time, X-ray recovery time, and the incidence of adverse reactions. CONCLUSIONS: This study provides reliable evidence-based support for the clinical application of Reduning injection combined with western medicine for pneumonia. ETHICS AND DISSEMINATION: Ethical approval is not required in this secondary research evidence, and we will publish the results of this study in a journal or relevant conferences. REGISTRATION NUMBER: DOI 10.17605/OSF.IO/VS75Y.

Liver X receptors agonist promotes differentiation of rat bone marrow derived mesenchymal stem cells into dopaminergic neuron-like cells.

Dopaminergic (DA) neurons derived from bone marrow derived mesenchymal stem cells (BMSCs) maybe a valuable source for cell replacement therapy in Parkinson disease. Recent studies showed that new functions of LXR and their ligands have been proposed to prevent PD in the adult nervous system. The present study was designed to observe the effect of liver X receptors (LXR) agonist on differentiation of rat BMSCs into DA neurons. Expressions of the neuronal markers (Tuj1 and Nestin), the specific marker of DA neurons (tyrosine hydroxylase, TH), LXR α and LXR ß were measured by immunocytochemical assay and TH/Tuj1 positive cells were determined by quantitative cell count analyses. mRNA expressions of LXR α, LXR ß, TH, DAT, Nurr1, Pitx3, En1 and Lmx1b were measured by qPCR. Compared with growth factors (GF) treated group, combined use of LXR and GF induced rat BMSCs to TH-expressing cells with 87.42% of efficiency in 6 days of period of induction. LXR agonist alone did not induce the differentiation. Compared with GF alone, combined use of LXR and GF increased expressions of LXR α and LXR ß protein and mRNA and TH, DAT, Nurr1, and Pitx3 mRNA, decreased expressions of En1 and Lmx1b mRNA. Our experimental results indicated that LXR activation leads to improve induction efficiency and shorten induction period of rat BMSCs into DA neuron-like cells through regulating DA development-related genes expressions and that LXR can be considered as a candidate target for drug development to improve differentiation of BMSCs into DA neurons.

cAMP/PKA-CREB-BDNF signaling pathway in hippocampus mediates cyclooxygenase 2-induced learning/memory deficits of rats subjected to chronic unpredictable mild stress.

To investigate the mechanism of cyclooxygenase 2 (COX2) in learning and memory impairments in rats subjected to chronic unpredictable mild stress (CUMS), meloxicam was used intragastrically to inhibit the activity of cyclooxygenase 2. Moreover, cyclooxygenase 2 over-expressing or RNA interfere lentivirus was injected intraventricularly to increase or decrease the enzyme's expression, respectively. The body weights and sucrose consumption were used to analyze depressive behaviors, while the Morris water maze and step-down-type passive avoidance tests were carried out to evaluate the learning-memory functions. The levels of inflammatory cytokines were measured to estimate inflammation and the contents of cyclic adenosine monophosphate (cAMP) were used to measure the levels of the second messenger. Changes in cyclooxygenase 2 mRNA levels were analyzed using reverse transcription polymerase chain reaction. Moreover, the expression of cyclooxygenase 2, brain-derived neurotrophic factor (BDNF), prostaglandins receptor 3 (EP3), protein kinase A (PKA), cAMP response element binding protein (CREB), and phosphorylated CREB were estimated using immunohistochemical staining or western blotting. The results showed that CUMS led to significant depressive-like behaviors and learning and memory dysfunctions. Also, the cAMP levels decreased significantly, while levels of inflammatory cytokines and prostaglandins E2 increased significantly. The expressions of PKA, BDNF, phosphorylated CREB/CREB declined and cyclooxygenase 2 was increased. Meloxicam and cyclooxygenase 2 RNA interfere lentivirus reversed the changes caused by CUMS while cyclooxygenase 2-overexpressing lentivirus worsened these abnormalities. The findings also showed that CUMS increased cyclooxygenase 2 expression, which can cause learning and memory impairments, mainly through activating the hippocampal neuronal cAMP/PKA-CREB-BDNF signaling pathways.

Cyclooxygenase-2 Signalling Pathway in the Cortex is Involved in the Pathophysiological Mechanisms in the Rat Model of Depression.

This study was designed to investigate the effect of the cortical cyclooxygenase-2 (COX2) pathway on depressive behaviour in rats. Meloxicam, COX2 overexpressed lentivirus and COX2 RNAi lentivirus were administered to Sprague-Dawley rats subjected to chronic unpredictable mild stress (CUMS). Behaviour tests, biochemistry and immunohistochemistry methods, enzyme-linked immunosorbent assays, western blotting and reverse transcription polymerase chain reactions were used to evaluate the changes in rat behaviour and the cortical COX2 pathway. CUMS rats showed depressive-like behaviours. The superoxide dismutase activity and cyclic adenosine monophosphate (cAMP) contents were significantly decreased, the contents of malondialdehyde, prostaglandin E2 (PGE2) and inflammatory cytokines were significantly increased. The expressions of protein kinase A (PKA) and cAMP response element-binding protein (CREB) were decreased, and the levels of brain-derived neurotrophic factor (BDNF) and COX2 were significantly increased. Meloxicam and COX2 RNAi lentivirus significantly alleviated the abnormalities induced by CUMS, while COX2 overexpressed lentivirus aggravated these abnormalities. Our results indicated that the cortical COX2 pathway was activated in CUMS rats. Inhibition of COX2 activity/expression can obviously improve depressive behaviours in CUMS rats. Upregulating COX2 expression can increase the susceptibility of rats to CUMS. An imbalance in the cortical COX2-PGE2-cAMP/PKA-CREB-BDNF signalling pathway participates in the pathogenic mechanism of depression.

Re-evaluation of the interrelationships among the behavioral tests in rats exposed to chronic unpredictable mild stress.

The chronic unpredictable mild stress model of depression has been widely used as an experimental tool to investigate human psychopathology. Our objective was to provide an update on the validity and reliability of the chronic unpredictable mild stress model, by analyzing the interrelationships among the indexes using stepwise discriminant analysis and Pearson correlation coefficient to examine the possible combinations. We evaluated the depressive rats in both the presence and the absence of chronic unpredictable mild stress, using weight change, percentage of sucrose preference, coat state, splash test, open-field test, elevated plus-maze test, forced swimming test, and Morris water maze test. The results showed that 6-week-long chronic unpredictable mild stress produces significant depression and anxiety-like behavior. The combination of body weight change, percentage of sucrose preference, coat state score, open-field score, grooming latency of splash test, immobility time in force swimming test, and platform crossing in the Morris water maze test can effectively discriminate between normal and chronic unpredictable mild stress rats. Strong interrelationships were noted among these indexes in both open-field test and elevated plus-maze test. In conclusion, there might be certain criteria for the combination of behavioral endpoints, which is advantageous to more effectively and reliably assess the chronic unpredictable mild stress induced depression model.

The protection of meloxicam against chronic aluminium overload-induced liver injury in rats.

The present study was designed to observe the protective effect and mechanisms of meloxicam on liver injury caused by chronic aluminium exposure in rats. The histopathology was detected by hematoxylin-eosin staining. The levels of prostaglandin E2, cyclic adenosine monophosphate and inflammatory cytokines were detected by enzyme linked immunosorbent assay. The expressions of cyclooxygenases-2, prostaglandin E2 receptors and protein kinase A were measured by western blotting and immunohistochemistry. Our experimental results showed that aluminium overload significantly damaged the liver. Aluminium also significantly increased the expressions of cyclooxygenases-2, prostaglandin E2, cyclic adenosine monophosphate, protein kinase A and the prostaglandin E2 receptors (EP1,2,4) and the levels of inflammation and oxidative stress, while significantly decreased the EP3 expression in liver. The administration of meloxicam significantly improved the impairment of liver. The contents of prostaglandin E2 and cyclic adenosine monophosphate were significantly decreased by administration of meloxicam. The administration of meloxicam also significantly decreased the expressions of cyclooxygenases-2 and protein kinase A and the levels of inflammation and oxidative stress, while significantly increased the EP1,2,3,4 expressions in rat liver. Our results suggested that the imbalance of cyclooxygenases-2 and downstream prostaglandin E2 signaling pathway is involved in the injury of chronic aluminium-overload rat liver. The protective mechanism of meloxicam on aluminium-overload liver injury is attributed to reconstruct the balance of cyclooxygenases-2 and downstream prostaglandin E2 signaling pathway.

The mechanism of 5-lipoxygenase in the impairment of learning and memory in rats subjected to chronic unpredictable mild stress.

OBJECTIVES: To examine the mechanism of 5-lipoxygenase (5-LO) in the learning and memory dysfunction in rats subjected to chronic unpredictable mild stress (CUMS). METHODS: Eighty rats were divided into eight groups: the 0.5% sodium carboxymethyl cellulose solution (NaCMC)-treated group, empty vector (LV-Mock)-treated group, CUMS+NaCMC-treated group, CUMS+sertraline-treated group, CUMS+caffeic acid (10mg/kg)-treated group, CUMS+caffeic acid (30mg/kg)-treated group, CUMS+LV-Mock-treated group, and CUMS+5-LO-silencers lentiviral vectors (LV-si-5-LO)-treated group, n=10. Sucrose preference tests were performed to assess depression-like behavior. The Morris water maze and step-down tests were used to evaluate learning and memory performance. The levels of inflammatory cytokines, malondialdehyde, and the activity of superoxide dismutase (SOD) were detected to estimate inflammation and oxidative stress. Changes in 5-LO mRNA and protein were detected using reverse transcription polymerase chain reaction and Western blotting. The expression of synaptophysin, postsynaptic density-95 (PSD-95), and brain-derived neurotrophic factor (BDNF) in the hippocampus were measured using immunohistochemical staining. RESULTS: Treatment with caffeic acid or LV-si-5-LO increased sucrose consumption, decreased escape latency and increased the number of platform crosses in the Morris water maze test, and decreased the number of errors and prolonged the latency in the step-down test. We observed a decreased expression of 5-LO, and levels of malondialdehyde, leukotriene-B4, tumor necrosis factor-α, and interleukin-6, while the protein levels of synaptophysin, PSD-95, BDNF, and the activity of SOD were increased in the hippocampus of the CUMS-treated rats. CONCLUSIONS: CUMS-induced impairment in learning and memory could be triggered by an inflammatory response in the rat hippocampus, which results in oxidative stress injury and impacts the synaptic plasticity of hippocampal neurons. Inhibition of the activity or expression of 5-LO could suppress hippocampal inflammation, enhance synaptic plasticity, and improve learning and memory function in depressed rats.

PGE2-EP3 signaling pathway contributes to protective effects of misoprostol on cerebral injury in APP/PS1 mice.

Epidemiological studies indicate chronic use of non-steroidal anti-inflammatory drugs (NSAIDs), which inhibit the enzymatic activity of the inflammatory cyclooxygenases (COX), reduces the risk of developing Alzheimer's disease (AD) in normal aging populations. Considering multiple adverse side effects of NSAIDs, findings suggest that COX downstream prostaglandin signaling function in the pre-clinical development of AD. Our previous study found that misoprostol, a synthetic prostaglandin E2 (PGE2) receptor agonist, has neuroprotection against brain injury induced by chronic aluminum overload. Here, we investigated the neuroprotective effects and mechanisms of misoprostol on neurodegeneration in overexpressing both amyloid precursor protein (APP) and mutant presenilin 1 (PS1) mice. Here were young group, elderly group, APP/PS1 group and misoprostol-treated group. Mice in misoprostol-treated group were administrated with misoprostol (200 µg·kg-1·d-1, p.o.) five days a week for 20 weeks. The spatial learning and memory function was impaired and karyopycnosis of hippocampal and cortical neurons was observed; amyloid beta (Aß) deposition was increased; superoxide dismutase (SOD) activity was decreased and malondialdehyde (MDA) content was increased in APP/PS1 mice. However, misoprostol could significantly blunte these changes in APP/PS1 mic. Moreover, the expressions of microsomal PGE2 synthase (mPGES-1), PGE2, PGE2 receptor (EP) 2 and EP4 were increased and EP3 expression was decreased in APP/PS1 mice, while misoprostol reversed these changes. Our present experimental results indicate that misoprostol has a neuroprotective effect on brain injury and neurodegeneration of APP/PS1 mice and that the activation of PGE2-EP3 signaling and inhibition of oxidative stress contribute to the neuroprotective mechanisms of misoprostol.

Effect of the PGD2-DP signaling pathway on primary cultured rat hippocampal neuron injury caused by aluminum overload.

In the present study, the agonists and antagonists of DP receptor were used to examine whether the PGD2-DP signaling pathway affects neuronal function. Primary cultured hippocampal neuron was prepared and treated with aluminum maltolate (100 µM) to establish the neuronal damage model. PGD2 and cAMP content was detected by ELISA. L-PGDS and DPs mRNA and protein expression were measured by RT-PCR and Western blotting, respectively. The aluminium-load neuron was treated with the DP1 agonist BW245C, the DP1 antagonist BWA868C, the DP2 agonist DK-PGD2, and the DP2 antagonist CAY10471, respectively. Neuronal pathomorphology was observed using H-E staining. The cell viability and the lactate dehydrogenase leakage rates of neurons were measured with MTT and LDH kit, respectively. Ca(2+) level was detected by Fluo-3/AM. In the model group, the MTT values obviously decreased; LDH leakage rates and PGD2 content increased significantly; L-PGDS, DP1 mRNA and protein expressions increased, and DP2 level decreased. BW245C reduced the Ca(2+) fluorescence intensity and protected the neurons. DK-PGD2 increased the intensity of Ca(2+) fluorescence, while CAY10471 had the opposite effect. In conclusion, contrary to the effect of DP2, the PGD2-DP1 signaling pathway protects against the primary cultured rat hippocampal neuronal injury caused by aluminum overload.