Increasing evidence shows that inflammation plays a vital role in the occurrence and development of ischemia/reperfusion (I/R). Suppression of excessive inflammation can ameliorate impaired cardiac function, which shows therapeutic potential for clinical treatment of myocardial ischemia/reperfusion (MI/R) diseases. In this study, we investigated whether Ginkgolide C (GC), a potent anti-inflammatory flavone, extenuated MI/R injury through inhibition of inflammation. In vivo, rats with the occlusion of the left anterior descending (LAD) coronary artery were applied to mimic MI/R injury. In vitro, primary cultured neonatal ventricular myocytes exposed to hypoxia/reoxygenation (H/R) were applied to further discuss the anti-H/R injury property of GC. The results revealed that GC significantly improved the symptoms of MI/R injury, as evidenced by reducing infarct size, preventing myofibrillar degeneration and reversing the mitochondria dysfunction. Moreover, histological analysis and Myeloperoxidase (MPO) activity measurement showed that GC remarkably suppressed Polymorphonuclears (PMNs) infiltration and ameliorated the histopathological damage. Furthermore, GC pretreatment was shown to improve H/R-induced ventricular myocytes viability and enhance tolerance of inflammatory insult, as evidenced by suppressing expression of CD40, translocation of NF-κB p65 subunit, phosphorylation of IκB-α, as well as the activity of IKK-β. In addition, downstream inflammatory cytokines modulated by NF-κB signaling were effectively down-regulated both in vivo and in vitro, as determined by immunohistochemistry and ELISA. In conclusion, these results indicate that GC possesses a beneficial effect against MI/R injury via inflammation inhibition that may involve suppression of CD40-NF-κB signal pathway and downstream inflammatory cytokines expression, which may offer an alternative medication for MI/R diseases.

Post-cardiac arrest myocardial dysfunction significantly contributes to early mortality after the return of spontaneous circulation. However, no effective therapy is available now. Aldehyde dehydrogenase 2 (ALDH2) enzyme has been shown to protect the heart from aldehyde toxicity such as 4-hydroxy-2-nonenal (4-HNE) and oxidative stress. In this study, we evaluated the effect of enhanced activity or expression of ALDH2 on post-cardiac arrest myocardial dysfunction and survival in a rat cardiac arrest model. Furthermore, we elucidated the underlying mechanisms with a focus on mitochondrial reactive oxygen species (ROS) production in a cell hypoxia/reoxygenation model. A total of 126 rats were used for the ALDH2 activation or cardiac overexpression of ALDH2 studies. Randomization was done 10 min before the respective agonist injection or in vivo gene delivery. We showed that enhanced activity or expression of ALDH2 significantly improved contractile function of the left ventricle and survival rate in rats subjected to cardiac arrest-cardiopulmonary resuscitation procedure. Moreover, ALDH2 prevented cardiac arrest-induced cardiomyocyte death from apoptosis and mitochondrial damage. Mechanistically, 4-HNE, a representative substrate of ALDH2, was dominantly increased in the hypoxia/reoxygenation-exposed cardiomyocytes. Direct addition of 4-HNE led to significantly augmented succinate accumulation and mitochondrial ROS production. Through metabolizing 4-HNE, ALDH2 significantly inhibited mitochondrial ROS production. Our findings provide compelling evidence of the cardioprotective effects of ALDH2 and therapeutic targeting this enzyme would provide an important approach for treating post-cardiac arrest myocardial dysfunction.

Osteoarthritis (OA) is the most frequent and disabling disease in developed countries. The progressive degeneration of articular cartilage characterized as thinner and erosive. Inflammation is well-known to be involved in OA development. However, there are no effective therapeutic strategies to cure it. Xanthohumol (XH) is a natural prenylflavonoid isolated from hops and beer. The protective activity of XH against OA chondrocytes inflammation and ECM degradation is unclear. In this article, we found that XH significantly inhibited inflammatory responses, attenuated catabolic enzymes expression, and ameliorated ECM degradation, as showed by decreased production of NO, PGE2, TNFα, and IL-6, decreased expression of MMP-3/-13 and ADAMTS-4/-5, and increased expression of collagen-II and aggrecan. In addition, XH activated HO-1 signaling and attenuated IL-1β-induced C/EBPβ. XH promoted the interaction between HO-1 and C/EBPβ, inhibiting the nuclear translocation of C/EBPβ. HO-1 knockdown could abrogate the protective effects of XH in IL-1β-treated chondrocytes. Collectively, XH attenuated inflammatory responses and ECM degradation by mediating HO-1 and C/EBPβ signaling pathways in osteoarthritis chondrocytes.

Glioblastoma multiforme (GBM) is one of the most malignant primary tumors in humans. Despite standard therapeutic strategy with tumor resection combined with radiochemotherapy, the prognosis remains disappointed. Recently, deubiquitinating enzymes (DUBs) has been reported as potential cancer therapy targets due to their multifunctions involved in the regulation of tumorigenesis, cell cycle, apoptosis, and autophagy. In this study, we found that knockdown of ubiquitin specific protease (USP5), a family member of DUB, could significantly suppress GBM cell line U251 and DBTRG-05MG proliferation and colony formation by inducing cell cycle G1/S arrest, which was correlated with downregulation of CyclinD1 protein level. CyclinD1 had been reported to play a critical role in the tumorigenesis and development of GBM via regulating cell cycle transition. Overexpression of USP5 could significantly extend the half-life of CyclinD1, while knockdown of USP5 decreased the protein level of CyclinD1, which could be restored by proteasome inhibitor MG-132. Indeed, USP5 was found to directly interact with CyclinD1, and decrease its K48-linked polyubiquitination level. Furthermore, knockdown of USP5 in U251 cells remarkably inhibited tumor growth in vivo. Taken together, these findings demonstrate that USP5 plays a critical role in tumorigenesis and progression of GBM by stabilizing CyclinD1 protein. Targeting USP5 could be a potential therapeutic strategy for GBM.

Osteoarthritis (OA), which is identified by chronic pain, impacts the quality of life. Cartilage degradation and inflammation are the most relevant aspects involved in its development. Signal transducer and activator of transcription 3(STAT3), a member of the STATs protein family, is associated with inflammation. Alantolactone (ALT), a sesquiterpene lactone compound, can selectively suppress the phosphorylation of STAT3. However, the pharmacological effect of ALT on OA is still imprecise. In this study, IL-1β (10 ng/ml) was applied to cartilage chondrocytes, which were treated with different concentrations of Alantolactone for 24 h. The expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2(COX2), matrix metalloproteinases (MMPs) and thrombospondin motifs-5 (ADAMTS5) were detected by western blot. Protein expression of Collagen Ⅱ was observed by western blot, safranin O staining and immunofluorescence. Manifestation of autophagy related proteins such as autophagy-related gene-5 (ATG5), P62, LC3Ⅱ/Ⅰ and PI3K/AKT/mTOR-related signaling molecules were measured by western blot and autophagic flux monitored by confocal microscopy. Expression of STAT3 and NF-κB-related signaling molecules were evaluated by western blot and immunofluorescence. In vivo, 2 mg/kg ALT or equal bulk of vehicle was engaged in the destabilization of medial meniscus (DMM) mouse models by intra-articular injection, the degree of cartilage destruction was classified by Safranin O/Fast green staining. Our findings reported that the enhance of inflammatory factors containing iNOS, COX2, MMPs and ADAMTS5 induced by IL-1β could be ameliorated by ALT. Additionally, the diminish of Collagen Ⅱ and autophagy which was stimulated by IL-1β could be alleviated by ALT. Mechanistically, STAT3, NF-κB and PI3K/AKT/mTOR signal pathways might be involved in the effect of ALT on IL-1β-induced mouse chondrocytes. In vivo, ALT protected cartilage in the DMM mouse model. Overall, this study illustrated that ALT attenuated IL-1β-induced inflammatory responses, relieved cartilage degeneration and promoted impaired autophagy via restraining of STAT3 and NF-κB signal pathways, implying its auspicious therapeutical effect for OA.

Background Revefenacin (REV) is a novel once-daily long-acting muscarinic antagonist (LAMA) in the treatment of moderate to very severe chronic obstructive pulmonary disease (COPD). This systematic review incorporating a dose-response meta-analysis aimed to assess the efficacy and safety of REV. Methods PubMed, Embase, Cochrane Library, China National Knowledge Infrastructure, VIP database, and Wanfang database were searched from their inception to April 2020. We included randomized controlled trials (RCTs) which evaluated the efficacy and safety of REV in COPD patients. Two reviewers independently performed study screening, data extraction, and risk of bias assessment. Outcomes consisted of the mean change in trough Forced Expiratory Volume in 1 second (FEV1) from baseline, adverse events (AEs), and serious adverse events (SAEs). A dose-response meta-analysis using the robust error meta-regression method was conducted. We used Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach to assess the quality of evidence. Results Nine RCTs (3,121 participants) were included in this systematic review. The meta-analyses indicated that 175 μg/day REV could significantly improve the trough FEV1 (MD=143.67, 95%CI: 129.67 to 157.68; I2=96%; 809 participants; studies=4; low quality) without increasing the risk of AEs (OR=0.98, 95%CI: 0.81 to 1.18; I2=34%; 2,286 participants; studies=7; low quality) or SAEs (OR=0.89, 95%CI: 0.55 to 1.46; I2=0%; 2,318 participants; studies=7; very low quality) compared to placebo. Furthermore, the effect of REV in increasing trough FEV1 was dose-dependent with an effective threshold of 88 μg/day (R2 = 0.7017). Nevertheless, only very low-quality to low-quality evidence showed that REV at a dose of 175 μg/day was inferior to tiotropium regarding the long-term efficacy, and its safety profile was not superior to tiotropium or ipratropium. Conclusion Current evidence shows that REV is a promising option for the treatment of moderate to very severe COPD. Due to most evidence graded as low quality, further studies are required to compare the efficacy, long-term safety and cost-effectiveness between REV and other LAMAs in different populations. Clinical Trial Registration: [PROSPERO], identifier [CRD42020182793].

Jinmaitong (JMT) is a compound prescription of traditional Chinese medicine that has been used to treat diabetic neuropathic pain (DNP) for many years. Here, we investigated the effects of JMT on the activation of NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome and pyroptosis in Dorsal root ganglia (DRG) of diabetic rats. Streptozotocin (STZ)-induced diabetic rats were gavaged with JMT (0.88 g/kg/d) or alpha-lipoic acid (ALA, positive control, 0.48 mmol/kg/d) for 12 weeks. Distilled water was administered as a vehicle control to both diabetic and non-affected control rats. Blood glucose levels and body weights were measured. Behavioral changes were tested with mechanical withdrawal threshold (MWT) and tail-flick latency (TFL) tests. Morphological injury associated with DRG was observed with hematoxylin and eosin (H&E) and Nissl's staining. mRNA and protein levels of NLRP3 inflammasome components (NLRP3, ASC, caspase-1), downstream IL-1β and gasdermin D (GSDMD) were evaluated by immunohistochemistry, quantitative real time-PCR and western blot. The results showed that JMT had no effect on blood glucose levels and body weights, but significantly improved MWT and TFL behavior in diabetic rats, and attenuated morphological damage in the DRG tissues. Importantly, JMT decreased the mRNA and protein levels of components of NLRP3 inflammasome, including NLRP3, ASC and caspase-1. JMT also down-regulated the expression of IL-1β and GSDMD in the DRG of DNP rats. In addition, ALA treatment did not perform better than JMT. In conclusion, JMT effectively relieved DNP by decreasing NLRP3 inflammasome activation and pyroptosis, providing new evidence supporting JMT as an alternative treatment for DNP.

The prognosis of cardiorenal dysfunction induced by diabetes mellitus (DM), which belongs to cardiorenal syndrome type 5, is poor and its pathogenesis remains elusive. We have reported that CX3CL1 exacerbated heart failure and direct inhibition of CX3CL1 improved cardiac function. Emerging evidence supports that CX3CL1 is involved in renal impairment. Here we attempt to clarify whether CX3CL1 might be a therapeutic target for cardiorenal dysfunction in diabetes. We found that cardiac and renal CX3CL1 protein levels were significantly increased in both streptozotocin-induced diabetic mice and in non-obese diabetic mice, and that hyperglycemia led to persistent CX3CL1 expression in the heart and kidneys even after it was controlled by insulin. In cultured cardiac and renal cells, soluble CX3CL1 accelerated mitochondrial-dependent apoptosis via activation of the RhoA/ROCK1-Bax signaling pathway and promoted fibrosis through cellular phenotypic trans-differentiation mediated by the TGF-β/Smad pathway. In the two diabetic mouse models, knockout of CX3CL1 receptor CX3CR1 or treatment with an CX3CL1 neutralizing antibody significantly improved cardiorenal dysfunction by inhibiting apoptosis, mitochondrial dysfunction, and fibrosis. Moreover, sodium glucose cotransporter 2 inhibitor canagliflozin significantly downregulated cardiac and renal CX3CL1 expression and improved cardiorenal dysfunction. These findings indicate that CX3CL1 could be a new therapeutic target for diabetes-induced cardiorenal dysfunction.

Ginsenoside Rb2 (Rb2), a fundamental saponin produced and isolated from ginseng (Panax ginseng C.A. Meyer), has a wide range of biological actions. The objective of this investigation was to see if ginsenoside Rb2 has any immunomodulatory properties against cyclophosphamide (CTX)-induced immunosuppression. For the positive control group, levamisole hydrochloride (LD) was used. We discovered that intraperitoneal injection of Rb2 (5, 10, 20 mg/kg) could relieve CTX-induced immunosuppression by enhanced immune organ index, reduced the pathological characteristics of immunosuppression, promoted natural killer (NK) cells viability, improved cell-mediated immune response, boosted the IFN-γ (Interferon-gamma), TNF-α (Tumor necrosis factor-alpha), IL-2 (Interleukin-2), and IgG (Immunoglobulin G), as well as macrophage activity like carbon clearance and phagocytic index. Rb2 significantly elevated the mRNA expression of IL-4 (Interleukin-4), SYK (Tyrosine-protein kinase-SYK), IL-2, TNF-α, and IL-6 (Interleukin-6) in the spleen of CTX-injected animals. Molecular docking results showed that Rb2 had excellent binding properties with IL-4, SYK, IL-2, TNF, and IL-6, indicating the target protein might be strongly correlated with the immunomodulatory effect of Rb2. Taken together, ginsenoside Rb2 can improve the immune function that is declined in CTX-induced immunosuppressed mice, the efficacy maybe due to the regulation of related cytokine and mRNA expression.

The purpose of this study was to evaluate the protective effect of acidic fibroblast growth factor targeted mediated by novel nanoparticles-cationic lipid microbubbles complex (aFGF-NP + CPMBs) combined with ultrasound targeted microbubble destruction (UTMD)on doxorubicin-induced heart failure (HF)and its mechanism. Heart failure rats induced by intraperitoneal injection with doxorubicin (DOX) to achieve cummulative dose of 15mg/kg for continuous 6 weeks showed left ventricular dysfunction, seriously oxidative stress, cardiomyocyte apoptosis, and decrease of myocardial vascular density. In contrast, aFGF-NP + CPMBs combined with UTMD therapy (3ug/kg, caudal vein injection, twice a week, 6weeks)prominently ameliorated left ventricular dysfunction by increased ejection fraction (EF) and fractional shortening (FS), decreased brain natriuretic peptide (BNP); strengthened the ability of antioxidant stress confirmed by increasing the activity of SOD and reducing the production of MDA; exerted the effect of anti-cardiomyocyte apoptosis and promotion angiogenesis by inhibited Bax expression and increased Bcl-2 expression and platelet endothelial cell adhesion molecule (CD31) expression. Taken together, the research suggested that aFGF targeted mediated by novel nanoparticles-cationic lipid microbubbles complex combined with UTMD should be a promising targeted treatment for heart failure.

Aims: To compare the efficacy of five kinds of antiangiogenic drugs in the treatment of diabetic macular edema Methods: A comprehensive search of seven databases without language restrictions includes PubMed, EMBASE, Web of Science, CBM, the Cochrane Library, CNKI, and WanFang date. All literature used was published before October 2020. Eligible randomized trials were screened for inclusion in this study, and Bayesian framework was used to perform a network meta-analysis (NMA). Data on the mean change of best-corrected visual acuity (BCVA), central macular thickness (CMT) and intraocular pressure (IOP) at 6 months were extracted. Results: 25 randomized controlled trials (RCTs) that covered 2214 eyes, which received treatment of more than 3 months durations were included. In the pooled pair-wise meta-analysis, there was no statistically significant difference between all treatments. The same result was observed in the network meta-analysis with 0-37.82% Global I-squared. For BCVA at 6 months, conbercept and ranibizumab may be favorable than bevacizumab, aflibercept, triamcinolone acetonide and sham injections according to the ranking probabilities. As for CMT at 6 months, ranibizumab may be the most effective compared to bevacizumab, aflibercept and triamcinolone acetonide. In terms of IOP at 6 months, ranibizumab have better effect than bevacizumab, triamcinolone acetonide and sham injections. The results of sensitivity analysis also confirm it. Conclusion: The analysis confirms that ranibizumab may be the most favorable for BCVA improvement and have a stronger efficacy in decreasing CMT and IOP than other drugs when taking all the indicators into consideration. This conclusion may provide clinical evidence to guide treatment decisions. However, more high-quality randomized controlled trials will be necessary to further confirm this.

Temporal lobe epilepsy (TLE) is characterized as an impaired ability of learning and memory with periodic and unpredictable seizures. Status epilepticus (SE) is one of the main causes of TLE. Neuroinflammation and oxidative stress are directly involved in epileptogenesis and neurodegeneration, promoting chronic epilepsy and cognitive deficit. Previous studies have shown that ursolic acid (UA) represses inflammation and oxidative stress, contributing to neuroprotection. Herein, we demonstrated that UA treatment alleviated seizure behavior and cognitive impairment induced by epilepsy. Moreover, UA treatment rescued hippocampal neuronal damage, aberrant neurogenesis, and ectopic migration, which are commonly accompanied by epilepsy occurrence. Our study also demonstrated that UA treatment remarkably suppressed the SE-induced neuroinflammation, evidenced by activated microglial cells and decreased inflammation factors, including TNF-α and IL-1β. Likewise, the expression levels of oxidative stress damage markers and oxidative phosphorylation (OXPHOS) enzyme complexes of mitochondria were also remarkably downregulated following the UA treatment, suggesting that UA suppressed the damage caused by the high oxidative stress and the defect mitochondrial function induced by SE. Furthermore, UA treatment attenuated GABAergic interneuron loss. In summary, our study clarified the notable anti-seizure and neuroprotective properties of UA in pilocarpine-induced epileptic rats, which is mainly achieved by abilities of anti-inflammation and anti-oxidation. Our study indicates the potential advantage of UA application in ameliorating epileptic sequelae.

Perioperative neurocognitive disorders (PND) is a common postoperative neurological complication. Neuroinflammation is a major cause that leads to PND. Autophagy, an intracellular process of lysosomal degradation, plays an important role in the development and maintenance of nervous system. PK11195 is a classic translocator protein (TSPO) ligand, which can improve the cognitive function of rats. In this study, we evaluate the protective effect of PK11195 on the learning and memory of rats. A rat model of lipopolysaccharide (LPS)-induced cognitive dysfunction was established by intraperitoneal injection of LPS. Morris Water Maze (MWM), Western blot, qRT-PCR, confocal microscopy and transmission electron microscopy (TEM) were used to study the role of TSPO-specific ligand PK11195 in LPS-activated mitochondrial autophagy in rat hippocampus. We found that PK11195 ameliorated LPS-induced learning and memory impairment, as indicated by decreased escape latencies, swimming distances and increased target quadrant platform crossing times and swimming times during MWM tests. TSPO, ATG7, ATG5, LC3B and p62 protein and mRNA expression increased in the hippocampus of PND model rats. The hippocampal microglia of PND model rats also have severe mitochondrial damage, and a large number of autophagosomes and phagocytic vesicles can be seen. PK11195 pretreatment significantly decreased the expression of TSPO, ATG7, ATG5, LC3B and p62 protein and mRNA, as well as mitochondrial damage. These findings suggested that PK11195 may alleviate the damage of LPS-induced cognitive dysfunction of rats by inhibiting microglia activation and autophagy.

Cannabinoid receptor 2 (CB2) has been reported to produce a cardio-protective effect in cardiovascular diseases such as myocardial infarction. Here in this study, we investigated the role of CB2 in diabetic cardiomyopathy (DCM) and its underlying mechanisms. HU308 was used for the selective activation of CB2. Bafilomycin A1 was used for the blockade of autophagy and compound C was used to inhibit AMPK signaling. An streptozotocin (STZ)-induced mice model and high glucose (HG)-challenged cardiomyocytes were applied for study. Cardiac function was detected by echocardiography and Western blot for the detection of autophagy-related and its signaling-related proteins. Transmission electron microscopy was used for the analysis of autophagosome number. Cell viability was detected by Cell Counting Kit-8 (CCK-8) and lactate dehydrogenase (LDH) release assays. We found that activating CB2 by HU308 improved cardiac function in DCM as well as cell viability in cardiomyocytes under HG challenge, while the administration of bafilomycin A1 attenuated the protective effects. HU308 enhanced the level of autophagy in the heart tissues from DCM mice as well as cardiomyocytes under HG challenge. HU308 triggered the AMPK-mTOR-p70S6K signaling pathway, while the administration of compound C attenuated the cardio-protective effect of HU308 in cardiomyocytes under HG challenge. In conclusion, we initially demonstrated that activating CB2 produced a cardio-protective effect in DCM as well as cardiomyocytes under HG challenge through inducing the AMPK-mTOR-p70S6K signaling-mediated autophagy.

Objectives: Lotus leaf is a kind of traditional Chinese medicine. We aimed to explore the effects of lotus leaf aqueous extract (LLAE) on peroxisome proliferative activated receptor γ2 (PPARγ2) expression in preadipocytes and adipocytes and further investigate its effects on high fat diet (HFD)-induced obese rats. Methods: pGL3-Enhancer-PPARγ2 (625 bp)-Luc plasmid, a luciferase reporter gene expression plasmid containing PPARγ2 promoter, was stably transfected into 3T3-L1 preadipocytes. PPARγ2 promoter activities were determined by assaying the luciferase activities. Then PPARγ2 promoter activities in preadipocytes and PPARγ2 mRNA levels in human subcutaneous adipocytes were measured after the administration with LLAE. Additionally, the effects of LLAE on body weight, fat mass, glucose and lipid metabolism and the expression of PPARγ2, insulin receptor substrate 1 and glucose transporter 4 (GLUT4) in visceral adipose tissue (VAT) were measured in HFD-induced obese rats treated with low or high dose [0.5 or 3.0 g crude drug/(kg.d)] LLAE for 6 weeks. Results: Ten μg/ml LLAE significantly increased the luciferase activities in 3T3-L1 cells and the stimulatory action reached 2.51 folds of controls when LLAE was 1000 μg/ml (P < 0.01). After treating 3T3-L1 cells with 100 μg/ml LLAE, the stimulatory role peaked at 32 h where it was 2.58 folds of controls (P < 0.01). Besides, 100 μg/ml LLAE significantly increased PPARγ2 mRNA levels in human adipocytes to 1.91 folds of controls (P < 0.01). In HFD-induced obese rats, administration with both low and high dose LLAE notably reduced visceral fat mass by 45.5 and 58.4%, respectively, and significantly decreased fasting serum insulin levels (P < 0.05). The high dose LLAE also significantly decreased homeostasis model assessment of insulin resistance in obese rats (P < 0.05). Furthermore, the mRNA levels of PPARγ2 and GLUT4 in VAT of obese rats were significantly increased when compared with control rats, and were notably suppressed by LLAE intervention for 6 weeks (P < 0.05). Conclusion: LLAE significantly reduces visceral fat mass and ameliorates insulin resistance in HFD-induced obese rats. These beneficial effects of LLAE may associate with its role in stimulating PPARγ2 expression in preadipocytes and subcutaneous adipocytes and suppressing PPARγ2 and GLUT4 expression in VAT.

Our pilot studies have shown that clemastine fumarate (CLE) can protect against myocardial ischemia-reperfusion injury (MIRI) through regulation of toll like receptor 4 (TLR4). However, the protective mechanism of CLE and related signaling pathways for MIRI remains unclear. The objective of this study is to determine the mechanism by which CLE relieves MIRI in cardiomyocytes and its relationship with the TLR4/PI3K/Akt signaling pathway. CCK8 analysis was used to test the optimal concentration of TLR4 inhibitor CLI-095 and TLR4 agonist lipopolysaccharide (LPS) on MIRI. The expression of inflammatory factors, oxidative stress response, cell damage, and intracellular calcium redistribution of cardiomyocytes were examined using the ELISA kits, Total Superoxide Dismutase Assay Kit with WST-8 and Lipid Peroxidation MDA Assay Kit, LDH Cytotoxicity Assay Kit, and laser scanning confocal microscope. The expression of TLR4/PI3K/Akt and cleaved caspase-3 were determined by Western blotting and immunofluorescent staining. Our results showed that MIRI aggravated the inflammatory response, oxidative stress, cellular damage of cardiomyocytes, and caused redistribution of intracellular calcium, upregulated the expression of TLR4 protein, cleaved caspase-3 protein, and down-regulated the expression of PI3K/Akt protein. After treatment with CLE, the inflammatory response, oxidative stress, and cellular damage of cardiomyocytes were alleviated, and intracellular calcium ion accumulation decreased. The expression of TLR4 protein, cleaved caspase-3 protein declined, but PI3K/Akt protein expression increased in cardiomyocytes treated with CLE. In addition, after treatment with the TLR4 inhibitor CLI-095, the results were similar to those of CLE treatment. The TLR4 agonist LPS aggravated the reactions caused by MIRI. The role of LPS was reversed after CLE treatment. These results suggested that CLE can attenuate MIRI by activating the TLR4/PI3K/Akt signaling pathway.

Sepsis commonly leads to acute and long-term cognitive and affective impairments which are associated with increased mortality in patients. Neuroinflammation characterized by excessive cytokine release and immune cell activation underlies the behavioral changes associated with sepsis. We previously reported that the administration of a traditional Chinese herbal Qiang Xin 1 (QX1) formula improves survival in septic mice. This study was performed to better understand the effects and the mechanisms of QX1 formula treatment on behavioral changes in a preclinical septic model induced by cecal ligation and puncture. Oral administration of QX1 formula significantly improved survival, alleviated overall cognitive impairment and emotional dysfunction as assessed by the Morris water maze, novel object recognition testing, elevated plus maze and open field testing in septic mice. QX1 formula administration dramatically inhibited short and long-term excessive pro-inflammatory cytokine production both peripherally and centrally, and was accompanied by diminished microglial activation in septic mice. Biological processes including synaptic transmission, microglia cell activation, cytokine production, microglia cell polarization, as well as inflammatory responses related to signaling pathways including the MAPK signaling pathway and the NF-κB signaling pathway were altered prominently by QX1 formula treatment in the hippocampus of septic mice. In addition, QX1 formula administration decreased the expression of the M1 phenotype microglia gene markers such as Cd32, Socs3, and Cd68, while up-regulated M2 phenotype marker genes including Myc, Arg-1, and Cd206 as revealed by microarray analysis and Real-time PCR. In conclusion, QX1 formula administration attenuates cognitive deficits, emotional dysfunction, and reduces neuroinflammatory responses to improve survival in septic mice. Diminished microglial activation and altered microglial polarization are involved in the neuroprotective mechanism of QX1 formula.

Bone marrow mesenchymal stem cells (BMSCs) can effectively alleviate liver fibrosis, but the efficacy of cell therapy alone is insufficient. In recent years, a combination of traditional Chinese medicine (TCM) and cell therapy has been increasingly used to treat diseases in clinical trials. Ferulic acid (FA) is highly effective in treating liver fibrosis, and a combination of cells and drugs is being tested in clinical trials. Therefore, we combined BMSCs and Ferulic acid to treat CCl4-induced fibrosis and determine whether this combination was more effective than single treatment. We used BMSCs and FA to treat CCl4-induced fibrosis in rat models, observed their therapeutic effects, and investigated the specific mechanism of this combination therapy in liver fibrosis. We created a BMSC/hepatic stellate cell (HSC) coculture system and used FA to treat activated HSCs to verify the specific mechanism. Then, we used cytochalasin D and angiotensin II to investigate whether BMSCs and FA inactivate HSCs through cytoskeletal rearrangement. MiR-19b-3p was enriched in BMSCs and targeted TGF-β receptor II (TGF-βR2). We separately transfected miR-19b-3p into HSCs and BMSCs and detected hepatic stellate cell activation. We found that the expression of the profibrotic markers α-SMA and COL1-A1 was significantly decreased in the combination group of rats. α-SMA and COL1-A1 levels were also significantly decreased in the HSCs with the combination treatment. Cytoskeletal rearrangement of HSCs was inhibited in the combination group, and RhoA/ROCK pathway gene expression was decreased. Following angiotensin II treatment, COL1-A1 and α-SMA expression increased, while with cytochalasin D treatment, profibrotic gene expression decreased in HSCs. The expression of COL1-A1, α-SMA and RhoA/ROCK pathway genes was decreased in the activated HSCs treated with a miR-19b-3p mimic, indicating that miR-19b-3p inactivated HSCs by suppressing RhoA/ROCK signalling. In contrast, profibrotic gene expression was significantly decreased in the BMSCs treated with the miR-19b-3p mimic and FA or a miR-19b-3p inhibitor and FA compared with the BMSCs treated with the miR-19b-3p mimic alone. In conclusion, the combination therapy had better effects than FA or BMSCs alone. BMSC and FA treatment attenuated HSC activation and liver fibrosis by inhibiting cytoskeletal rearrangement and delivering miR-19b-3p to activated HSCs, inactivating RhoA/ROCK signalling. FA-based combination therapy showed better inhibitory effects on HSC activation.

Transforming growth factor β receptor (TGF-β1R) and receptor tyrosine kinases (RTKs), such as VEGFRs, PDGFRs and FGFRs are considered important therapeutic targets in blocking myofibroblast migration and activation of idiopathic pulmonary fibrosis (IPF). To screen and design innovative prodrug to simultaneously target these four classes of receptors, we proposed an approach based on network pharmacology combining virtual screening and machine learning activity prediction, followed by efficient in vitro and in vivo models to evaluate drug activity. We first constructed Collagen1A2-A549 cells with type I collagen as the main biomarker and evaluated the activity of compounds to inhibit collagen expression at the cellular level. The data from the first round of Collagen1A2-A549 cell screening were substituted into the machine learning model, and the model was optimized accordingly. As a result, the false positive rate of the model was reduced from 85.0% to 66.7%, and two prospective compounds, Z103080500 and Z104578368, were finally selected. Collagen levels were reduced effectively by both Z103080500 (67.88% reduction) and Z104578368 (69.54% reduction). Moreover, these two compounds showed low cellular cytotoxicity. Subsequently, the effect of Z103080500 and Z104578368 was evaluated in a bleomycin-induced C57BL/6 mouse IPF model. These results showed that 50 mg/kg Z103080500 and Z104578368 could effectively reduce the number of inflammatory cells and the expression level of α-SMA. Meanwhile, Z103080500 and Z104578368 reduced the expression of major markers and inflammatory factors of IPF, such as collagen, IFN-γ, IL-17 and HYP, indicating that these screened Z103080500 and Z104578368 effectively delayed lung tissue inflammation and had a potential therapeutic effect on IPF. Our findings demonstrate that a screening and evaluation model for prodrug against IPF has been successfully established. It is of great significance to further modify these compounds to enhance their potency and activity.

Osteoarthritis (OA) is a major cause of cartilage pain and limited mobility in middle-aged and elderly individuals. The degeneration of cartilage induced by inflammation and cartilage anabolic and catabolic disorder plays a key role in OA. Shikimic acid (SA), a natural ingredient extracted from Illicium verum, has been shown to exert notable anti-inflammatory effects in previous studies, suggesting its potential effects in the treatment of OA. In this study, we revealed that the pretreatment of SW1353 human chondrocytes with SA before interleukin 1β (IL-1β) stimulation effectively decreased the expression of inducible nitric oxide synthase (iNOS), cyclooxygenase (Cox)-2, matrix metalloproteinases (MMPs; MMP3 and MMP13), a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-5, type X collagen, and p62; increased the expression of type II collagen, ATG7, Beclin-1, and LC3; and increased the autophagic flux. Mechanistically, we found that SA suppressed the IL-1β-induced activation of the mitogen-activated protein kinase (MAPK) and nuclear factor-kappaB (NF-κB) pathways. Furthermore, the results of safranin O staining and toluidine blue staining of primary rat cartilage chondrocytes and a trauma-induced rat model of OA showed that SA alleviated progression of OA in vivo. Collectively, our research enhances understanding of the mechanism of protective effect of SA against the progression of OA, which involves amelioration of cartilage degeneration, thereby providing new evidence for the use of SA as a therapy to prevent the development of OA.