Abstract Muscle atrophy is commonly caused by various diseases but still lacks effective treatment in clinical practice. Here, an artificial circular RNA (circRNA) named circmiR‐29b, which is designed to be a molecular sponge for miR‐29b containing 12 imperfect bulged miR‐29b binding sites is constructed. CircmiR‐29b shows a favorable functional effect with respect to miR‐29b repression in a specific and drastic manner. CircmiR‐29b can protect against in vitro muscle atrophy induced by dexamethasone (Dex), angiotensin II (Ang II), and tumor necrosis factor alpha (TNF‐α). Besides, circmiR‐29b attenuates in vivo muscle atrophy induced by denervation, immobilization, and Ang II. More importantly, skeletal muscle specific gene therapy using circmiR‐29b is able to attenuate established muscle atrophy induced by spiral wire immobilization. This work has developed an engineered circmiR‐29b acting as an miR‐29b sponge, and offers a promising therapeutic tool to prevent muscle atrophy.
Skeletal muscle atrophy is a common clinical feature of many acute and chronic conditions. Circular RNAs (circRNAs) are covalently closed RNA transcripts that are involved in various physiological and pathological processes, but their role in muscle atrophy remains unknown. Global circRNA expression profiling indicated that circRNAs are involved in the pathophysiological processes of muscle atrophy. circTmeff1 is identified as a potential circRNA candidate that influences muscle atrophy. It is further identified that circTmeff1 is highly expressed in multiple types of muscle atrophy in vivo and in vitro. Moreover, the overexpression of circTmeff1 triggers muscle atrophy in vitro and in vivo, while the knockdown of circTmeff1 expression rescues muscle atrophy in vitro and in vivo. In particular, the knockdown of circTmeff1 expression partially rescues muscle mass in mice during established atrophic settings. Mechanistically, circTmeff1 directly interacts with TAR DNA-binding protein 43 (TDP-43) and promotes aggregation of TDP-43 in mitochondria, which triggers the release of mitochondrial DNA (mtDNA) into cytosol and activation of the cyclic GMP-AMP synthase (cGAS)/ stimulator of interferon genes (STING) pathway. Unexpectedly, TMEFF1-339aa is identified as a novel protein encoded by circTmeff1 that mediates its pro-atrophic effects. Collectively, the inhibition of circTmeff1 represents a novel therapeutic approach for multiple types of skeletal muscle atrophy.
Pathological cardiac hypertrophy occurs in response to numerous stimuli and precedes heart failure (HF). Therapies that ameliorate pathological cardiac hypertrophy are highly needed.The expression level of miR-30d was analyzed in hypertrophy models and serum of patients with chronic heart failure by qRT-PCR. Gain and loss-of-function experiments of miR-30d were performed in vitro. miR-30d gain of function were performed in vivo. Bioinformatics, western blot, luciferase assay, qRT-PCR, and immunofluorescence were performed to examine the molecular mechanisms of miR-30d.miR-30d was decreased in both murine and neonatal rat cardiomyocytes (NRCMs) models of hypertrophy. miR-30d overexpression ameliorated phenylephrine (PE) and angiotensin II (Ang II) induced hypertrophy in NRCMs, whereas the opposite phenotype was observed when miR-30d was downregulated. Consistently, the miR-30d transgenic rat was found to protect against isoproterenol (ISO)-induced pathological hypertrophy. Mechanistically, methyltransferase EZH2 could promote H3K27me3 methylation in the promotor region of miR-30d and suppress its expression during the pathological cardiac hypertrophy. miR-30d prevented pathological cardiac hypertrophy via negatively regulating its target genes MAP4K4 and GRP78 and inhibiting pro-hypertrophic nuclear factor of activated T cells (NFAT). Adeno-associated virus (AAV) serotype 9 mediated-miR-30d overexpression exhibited beneficial effects in murine hypertrophic model. Notably, miR-30d was reduced in serum of patients with chronic heart failure and miR-30d overexpression could significantly ameliorate pathological hypertrophy in human embryonic stem cell-derived cardiomyocytes.Overexpression of miR-30d may be a potential approach to treat pathological cardiac hypertrophy.This work was supported by the grants from National Key Research and Development Project (2018YFE0113500 to J Xiao), National Natural Science Foundation of China (82020108002 to J Xiao, 81900359 to J Li), the grant from Science and Technology Commission of Shanghai Municipality (20DZ2255400 and 21XD1421300 to J Xiao, 22010500200 to J Li), Shanghai Sailing Program (19YF1416400 to J Li), the "Dawn" Program of Shanghai Education Commission (19SG34 to J Xiao), the "Chen Guang" project supported by the Shanghai Municipal Education Commission and Shanghai Education Development Foundation (19CG45 to J Li).
Muscle atrophy is a common complication of many chronic diseases including heart failure, cancer cachexia, aging, etc. Unhealthy habits and usage of hormones such as dexamethasone can also lead to muscle atrophy. However, the underlying mechanisms of muscle atrophy are not completely understood. Non-coding RNAs (ncRNAs), such as microRNAs (miRNAs), long ncRNAs (lncRNAs), and circular RNAs (circRNAs), play vital roles in muscle atrophy. This review mainly discusses the regulation of ncRNAs in muscle atrophy induced by various factors such as heart failure, cancer cachexia, aging, chronic obstructive pulmonary disease (COPD), peripheral nerve injury (PNI), chronic kidney disease (CKD), unhealthy habits, and usage of hormones; highlights the findings of ncRNAs as common regulators in multiple types of muscle atrophy; and summarizes current therapies and underlying mechanisms for muscle atrophy. This review will deepen the understanding of skeletal muscle biology and provide new strategies and insights into gene therapy for muscle atrophy.
Abstract Checkpoint inhibitors have been approved for the treatment of solid tumor and hematological malignancies. While significant responses have been observed in a subset of patients, outcomes are variable and there is a need to identify additional predictive biomarkers beyond PD-L1 levels as measured by IHC. Tumor mutation burden (TMB) has been correlated with response to checkpoint inhibitors and is emerging as a key biomarker for predicting checkpoint inhibitor response. So far, the methods used to assess tumor mutation burden have included exome sequencing and multiple laboratory-developed targeted NGS panels (e.g., FoundationOne and MSK-IMPACT). In order to fully determine the value of TMB as a predictive biomarker for immunotherapy, a standardized panel, workflow and data analysis pipeline for TMB assessment are needed. In this study we evaluated the performance of a commercially available targeted NGS panel and workflow for TMB analysis. A set of 30 FFPE tumor samples including colon, renal, gastric, endometrial, and lung tissues was analyzed. DNA and RNA were extracted using the RecoverAll Total Nucleic Acid Isolation Kit. DNA quantity and quality were assessed using Qubit and qPCR, respectively. Samples were analyzed with the ThermoFisher Oncomine™ Mutation Load Research Assay (TML), which evaluates tumor mutation load (mutations/Mb) by interrogating 409 cancer-related genes, spanning ~1.7 megabases of the genome. TMB was measured by counting somatic single-base substitutions per Mb at ≥10% allele frequency in single, non-matched, tumor DNA samples. The impact of DNA quality on the TMB score was evaluated. Deamination errors (i.e. G>A and C>T) in poor quality FFPE samples was found to cause the overestimation of TMB. Therefore, a delta Ct cutoff was established to qualify DNA samples for TMB analysis. 12 of the 30 samples were also analyzed using a comparator NGS panel covering ~1.25 megabases. The correlation of TMB results between the two panels was 0.87. Overall, TMB was lowest in RCC (9-17/Mb) compared to NSCLC and CRC (16-37/Mb). MSI status was determined using the Promega MSI Analysis System v1.2. A correlation was observed between TMB and MSI status in a subset of samples. Reproducibility of the assay was also evaluated. To identify clinically relevant mutations and genetic alteration associated with high mutation burden, the Oncomine Comprehensive assay v3 (OCAv3) was also used to analyze the sample set. Mutations in genes involved in several DNA repair pathways were found to correlate with TMB. This study demonstrated the feasibility of utilizing a commercial targeted NGS panel and data analysis pipeline for TMB evaluation in clinical FFPE tumor samples. Standardization of TMB analysis will enable the clinical validation of TMB as a predictive biomarker for therapy selection. Citation Format: Peng Fang, Zhenyu Yan, Quyen Vu, David Smith, Chad Galderisi, Cynthia S. Spittle, Jin Li. Evaluation of a commercial targeted NGS panel for tumor mutation burden assessment in FFPE tissue [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3614.
Abstract Muscle atrophy is a frequently observed complication, characterized by the loss of muscle mass and strength, which diminishes the quality of life and survival. No effective therapy except exercise is currently available. In our previous study, repressing miR-29b has been shown to reduce muscle atrophy. In our current study, we have constructed artificially engineered extracellular vesicles for the delivery of CRISPR/Cas9 to target miR-29b (EVs-Cas9-29b). EVs-Cas9-29b has shown a favorable functional effect with respect to miR-29b repression in a specific and rapid manner by gene editing. In in vitro conditions, EVs-Cas9-29b could protect against muscle atrophy induced by dexamethasone (Dex), angiotensin II (AngII), and tumor necrosis factor-alpha (TNF-α). And EVs-Cas9-29b introduced in vivo preserved muscle function in the well-established immobilization and denervation-induced muscle atrophy mice model. Our work demonstrates an engineered extracellular vesicles delivery of the miR-29b editing system, which could be potentially used for muscle atrophy therapy.
Objective:Through the establishment of new rural cooperative based HIS single disease real-time settlement system,the new rural cooperative single disease patients in our hospital's real-time settlement.Methods:Based on the charge system in our hospital and medical center,established the data interaction,real-time settlement.Results:Real time settlement to realize the seamless connection and the new interface of HIS system in our hospital and medical single disease.Conclusion:Implementation of the system to some extent solve the farmers' difficult and expensive'problem,effectively reducing the burden on farmers in the medical aspects of the patient's farmers simplify the reimbursement process.
Vibrio vulnificus (V. vulnificus) is a facultative anaerobic, alkalophilic, halophilic, mesophilic, gram-negative Vibrio bacterium that can cause severe wound infection, sepsis, and diarrhoea. In summer, the seawater temperature is 23 ℃ ~ 29 ℃ for the high incidence period. Infections caused by V. vulnificus are most common in patients with chronic underlying medical conditions. The disease has a rapid onset and progression and rapidly develops into septic shock and multiple organ failure (MOF). We report a case of an elderly diabetic patient with a long history of alcoholism who was successfully treated in our hospital suffering from V. vulnificus sepsis complicated by septic shock. In this case, the patient's medical history was clear, the emergency doctor's clinical assessment of the patient was rapid and accurate, the aetiology was sent for examination in a timely manner, the evidence for the treatment plan was sufficient, and the clinical outcome was satisfactory.
Vibrio vulnificus is a facultative anaerobic, alkalophilic, halophilic, mesophilic, Gram-negative bacterium that can cause severe wound infection, sepsis and diarrhea. This paper reported a case of 85-year-old male patient infected with Vibrio vulnificus due to being stabbed by a sea shrimp. This patient also had diabetes with a long history of alcoholism. Due to bacterial pathogenicity and the patient's underlying diseases, his condition deteriorated rapidly. Based on the rapid diagnosis of Vibrio vulnificus using the next-generation sequencing（NGS）technology and blood culture method, as well as the selection of the most effective antibiotics via drug sensitivity test, this patient underwent precise antimicrobial treatment, thorough debridement and drainage within the shortest possible time, and thus the prognosis of this patient was greatly improved. In this paper, we have systematically explored the epidemiology, clinical features, diagnosis and treatment of Vibrio vulnificus infection, thus providing a practical reference for the clinicians to quickly identify and treat possible Vibrio vulnificus infection in diabetic patients after contacting with sea water or seafood.
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