Effectiveness of modified Buzhong Yiqi decoction in treating myasthenia gravis: study protocol for a series of N-of-1 trials.

BACKGROUND: Myasthenia gravis (MG) is an acquired autoimmune disease with high heterogeneity. The disease is chronic, relapsing repeatedly and progressive with acute exacerbation occasionally. Although the treatment of MG has developed, it is still unsatisfactory and has some unexpected side effects. Traditional Chinese medicine (TCM) has shown great potential in MG treatment, including relief of muscle weakness syndrome, improvement of patient's quality of life, and reduction of side effects of western medicine. The purpose of this study is to evaluate the effectiveness of modified Buzhong Yiqi decoction (MBYD) as an add-on therapy for MG through a small series of N-of-1 trials. METHODS: Single-centre, randomized, double-blind, 3 crossover N-of-1 trials will be conducted to enroll patients with MG diagnosed as spleen-stomach deficiency syndrome or spleen-kidney deficiency syndrome in TCM. Each N-of-1 trial has 3 cycles of two 4-week periods containing the MBYD period and placebo period. The wash-out interval of 1 week is prior to switching each period. PRIMARY OUTCOME: quantitative myasthenia gravis (QMG). SECONDARY OUTCOMES: the following scales: myasthenia gravis composite (MGC), myasthenia gravis activities of daily living profile (MG-ADL), myasthenia gravis quality of life (MG-QOL); the level of CD4+FoxP3+Treg cells and cytokines (IL-4, IL-17A, INF-γ, TGF-ß) in the peripheral blood; the alterations of the composition of gut microbiota; reduction of the side effects of western medicine. DISCUSSION: Used by WinBUGS software, we will conduct a hierarchical Bayesian statistical method to analyze the efficacy of MBYD in treating MG in individuals and populations. Some confounding variables such as TCM syndrome type and potential carryover effect of TCM will be introduced into the hierarchical Bayesian statistical method to improve the sensitivity and applicability of the trials, and the use of prior available information within the analysis may improve the sensitivity of the results of a series of N-of-1 trials, from both the individual and population level to study the efficacy of TCM syndrome differentiation. We assumed that this study would reveal that MBYD is effective for MG and provide robust evidence of the efficacy of TCM to treat MG. TRIAL REGISTRATION: Chinese Clinical Trial Register, ID: ChiCTR2000040477 , registration on 29 November 2020.

Effect of DLT-SML on Chronic Stable Angina Through Ameliorating Inflammation, Correcting Dyslipidemia, and Regulating Gut Microbiota.

ABSTRACT: Chronic stable angina (CSA) is caused by coronary atherosclerosis. The gut microbiota (GM) and their metabolite trimethylamine-N-oxide (TMAO) levels are associated with atherosclerosis. Danlou tablet (DLT) combined with Salvia miltiorrhiza ligustrazine (SML) injection has been used to treat CSA. This study aims to investigate how DLT combined with SML (DLT-SML) regulates serum lipids, inflammatory cytokines, GM community, and microbial metabolite in patients with CSA. In this study, 30 patients with CSA were enrolled in the DLT-SML group, and 10 healthy volunteers were included in the healthy control group. The patients in the DLT-SML group were subdivided as the normal total cholesterol (TC) group and high-TC group according to their serum TC level before treatment. Blood samples were collected to investigate the (1) lipid content, including triglyceride (TG), TC, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol, (2) fasting blood glucose (Glu), (3) inflammatory cytokines, including interleukin-1 beta (IL-1ß), interleukin-6 (IL-6), interleukin-10 (IL-10), and tumor necrosis factor-α (TNF-α), and (4) gut-derived metabolite, including lipopolysaccharides and TMAO level. GM composition was analyzed by sequencing 16S rRNA of fecal samples. Results showed that DLT-SML significantly decreased serum TG, TC, low-density lipoprotein cholesterol, IL-1ß, TNF-α, and TMAO levels of patients with CSA. DLT-SML increased the abundance of Firmicutes and decreased Proteobacteria, which were significantly lower or higher in patients with CSA, respectively, compared with the healthy control group. In particular, DLT-SML increased the microbial diversity and decreased Firmicutes/Bacteroidetes ratio of patients with high-TC. The abundance of Sarcina, Anaerostipes, Streptococcus, Weissella, and Erysipelatoclostridium was decreased, whereas Romboutsia, Faecalibacterium, and Subdoligranulum were increased by DLT-SML treatment in patients with CSA. These findings indicated that DLT-SML improved patients with CSA by ameliorating dyslipidemia profile, decreasing the circulating inflammatory cytokines, and regulating the GM composition and their metabolites.

Therapeutic efficacy and immunoregulatory effect of Qiangji Jianli Capsule for patients with myasthenia gravis: Study protocol for a series of randomized, controlled N-of-1 trials.

INTRODUCTION: Myasthenia gravis (MG) is an autoimmune disease in which antibodies directly target components of the neuromuscular junction, causing neuromuscular conduction damage that leads to muscle weakness. The current pharmaceutical treatment for MG is still not ideal to address the problems of disease progression, high recurrence rate, and drug side effects. Clinical observations suggest that traditional Chinese medicine (TCM) can strengthen immunity and improve symptoms of MG patients, delay the progression of the disease, reduce or even prevent the need for immunosuppressive therapy when used in combination with acetylcholinesterase inhibitors or low-dose prednisone, as well as improve the quality of life of patients. The Qiangji Jianli Capsule (QJC) is a combination of medicinal herbs which is used in traditional Chinese medicine. Since MG is a rare disorder, randomized controlled trials comparing large cohorts are difficult to conduct. Therefore, we proposed to aggregate data from a small series of N-of-1 trials to assess the effect of the Chinese medical prescription QJC, which strengthens the spleen and nourishes Qi, as an add-on treatment for MG with spleen and stomach Qi deficiency syndrome. METHODS AND ANALYSIS: Single-center, randomized, double-blind, multiple crossover N-of-1 studies will compare QJC versus placebo in 5 adult MG patients with spleen and stomach Qi deficiency syndrome. Patients will undergo 3 cycles of two 4-week intervention periods. According to the treatment schedule, patients will continue to be treated with pyridine bromide tablets, prednisone acetate, tablets and/or tacrolimus capsules throughout the entire trial. Each period consisting of 4-week oral add-on treatment with QJC will be compared with 4-week add-on treatment with a placebo. The primary endpoints are quantitative myasthenia gravis (QMG) test; measurement of the amount of Treg cells and cytokines such as interferon-γ (IFN-γ), interleukin-4 (IL-4), interleukin-17A (IL-17A), and transforming growth factor-ß (TGF-ß); and corticosteroid or immunosuppressive agent dosage. Secondary outcome measures: Clinical: Evaluation of the effect of TCM syndromes; MG-activities of daily living (MG-ADL) scales; adverse events. ETHICS AND DISSEMINATION: This study was approved by The First Affiliated Hospital of Guangzhou University of Chinese Medicine (GZUCM), No. ZYYECK[2019]038. The results will be published in a peer-reviewed publication. Regulatory stakeholders will comment on the suitability of the trial for market authorization and reimbursement purposes. Trial registration: Chinese Clinical Trial Register, ID: ChiCTR2000033516. Registered on 3 June 2020, http://www.chictr.org.cn/showprojen.aspx?proj=54618.

Shenmai Injection Protects Against Doxorubicin-Induced Cardiotoxicity via Maintaining Mitochondrial Homeostasis.

Shenmai injection (SMI), as a patented traditional Chinese medicine, is extracted from Panax ginseng and Ophiopogon japonicus. It commonly used in the treatment of cardiovascular disease and in the control of cardiac toxicity induced by doxorubicin (DOX) treatment. However, its anti-cardiotoxicity mechanism remains unknown. The purpose of this study was to investigate the underlying mitochondrial protective mechanisms of SMI on DOX-induced myocardial injury. The cardioprotective effect of SMI against DOX-induced myocardial damage was evaluated in C57BL/6 mice and H9c2 cardiomyocytes. In vivo, myocardial injury, apoptosis and phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB/Akt)/glycogen synthase kinase 3 beta (GSK-3ß) signaling pathway related proteins were measured. In vitro, apoptosis, mitochondrial superoxide, mitochondrial membrane potential, mitochondrial morphology, levels of mitochondrial fission/fusion associated proteins, mitochondrial respiratory function, and AMP-activated protein kinase (AMPK) activity were assessed. To further elucidate the regulating effects of SMI on AMPK and PI3K/Akt/GSK-3ß signaling pathway, compound C and LY294002 were utilized. In vivo, SMI decreased mortality rate, levels of creatine kinase, and creatine kinase-MB. SMI significantly prevented DOX-induced cardiac dysfunction and apoptosis, decreased levels of Bax/Bcl-2 and cleaved-Caspase3, increased levels of PI3K, p-Akt, and p-GSK-3ß. In vitro, SMI rescued DOX-injured H9c2 cardiomyocytes from apoptosis, excessive mitochondrial reactive oxygen species production and descending mitochondrial membrane potential, which were markedly suppressed by LY294002. SMI increased ratio of L-OPA1 to S-OPA1, levels of AMPK phosphorylation, and DRP1 phosphorylation (Ser637) in order to prevent DOX-induced excessive mitochondrial fission and insufficient mitochondrial fusion. In conclusion, SMI prevents DOX-induced cardiotoxicity, inhibits mitochondrial oxidative stress and mitochondrial fragmentation through activation of AMPK and PI3K/Akt/GSK-3ß signaling pathway.

Investigation of cardiovascular protective effect of Shenmai injection by network pharmacology and pharmacological evaluation.

BACKGROUND: Shenmai injection (SMI) has been used in the treatment of cardiovascular disease (CVD), such as heart failure, myocardial ischemia and coronary heart disease. It has been found to have efficacy on doxorubicin (DOX)-induced cardiomyopathy. The aims of this study were to explore the underlying molecular mechanisms of SMI treatment on CVD by using network pharmacology and its protective effect on DOX-induced cardiotoxicity by in vitro and in vivo experiment based on network pharmacology prediction. METHODS: Network pharmacology method was used to reveal the relationship between ingredient-target-disease and function-pathway of SMI on the treatment of CVD. Chemical ingredients of SMI were collected form TCMSP, BATMAN-TCM and HIT Database. Drugbank, DisGeNET and OMIM Database were used to obtain potential targets for CVD. Networks were visualized utilizing Cytoscape software, and the enrichment analysis was performed using IPA system. Finally, cardioprotective effects and predictive mechanism confirmation of SMI were investigated in H9c2 rat cardiomyocytes and DOX-injured C57BL/6 mice. RESULTS: An ingredient-target-disease & function-pathway network demonstrated that 28 ingredients derived from SMI modulated 132 common targets shared by SMI and CVD. The analysis of diseases & functions, top pathways and upstream regulators indicated that the cardioprotective effects of SMI might be associated with 28 potential ingredients, which regulated the 132 targets in cardiovascular disease through regulation of G protein-coupled receptor signaling. In DOX-injured H9c2 cardiomyocytes, SMI increased cardiomyocytes viability, prevented cell apoptosis and increased PI3K and p-Akt expression. This protective effect was markedly weakened by PI3K inhibitor LY294002. In DOX-treated mice, SMI treatment improved cardiac function, including enhancement of ejection fraction and fractional shortening. CONCLUSIONS: Collectively, the protective effects of SMI on DOX-induced cardiotoxicity are possibly related to the activation of the PI3K/Akt pathway, as the downstream of G protein-coupled receptor signaling pathway.