FTO-mediated RNA m6A methylation regulates synovial aggression and inflammation in rheumatoid arthritis.

Fibroblast-like synoviocytes (FLS) plays an important role in synovial inflammation and joint damage in rheumatoid arthritis (RA). As the most abundant mRNA modification, N6-methyladenosine (m6A) is involved in the development of various diseases; however, its role in RA remains to be defined. In this study, we reported the elevated expression of the m6A demethylase fat mass and obesity-associated protein (FTO) in FLS and synovium from RA patients. Functionally, FTO knockdown or treatment with FB23-2, an inhibitor of the mRNA m6A demethylase FTO, inhibited the migration, invasion and inflammatory response of RA FLS, however, FTO-overexpressed RA FLS exhibited increased migration, invasion and inflammatory response. We further demonstrated that FTO promoted ADAMTS15 mRNA stability in an m6A-IGF2BP1 dependent manner. Notably, the severity of arthritis was significantly reduced in CIA mice with FB23-2 administration or CIA rats with intra-articular injection of FTO shRNA. Our results illustrate the contribution of FTO-mediated m6A modification to joint damage and inflammation in RA and suggest that FTO might be a potential therapeutic target in RA.

Transcriptome analysis reveals ADAMTS15 is a potential inflammation-related gene in remote ischemic postconditioning.

Background: Remote ischemic postconditioning (RIPostC) induced by brief episodes of the limb ischemia is a potential therapeutic strategy for myocardial ischemia/reperfusion injury, achieved by reducing cardiomyocyte death, inflammation and so on. The actual mechanisms underlying cardioprotection conferred by RIPostC remain unclear. Exploring gene expression profiles in myocardium at transcriptional level is helpful to deepen the understanding on the cardioprotective mechanisms of RIPostC. This study aims to investigate the effect of RIPostC on gene expressions in rat myocardium using transcriptome sequencing. Methods: Rat myocardium samples from the RIPostC group, the control group (myocardial ischemia/reperfusion group) and the sham group were performed transcriptome analysis using RNA sequencing. The levels of cardiac IL-1ß, IL-6, IL-10 and TNFα were analyzed by Elisa. The expression levels of candidate genes were verified by qRT-PCR technique. Infarct size was measured by Evans blue and TTC staining. Apoptosis was assessed by TUNEL assays and caspase-3 levels were detected using western blotting. Results: RIPostC can markedly decrease infarct size and reduce the levels of cardiac IL-1ß, IL-6 and increase the level of cardiac IL-10. This transcriptome analysis showed that 2 genes were up-regulated (Prodh1 and ADAMTS15) and 5 genes (Caspase-6, Claudin-5, Sccpdh, Robo4 and AABR07011951.1) were down-regulated in the RIPostC group. Go annotation analysis showed that Go terms mainly included cellular process, metabolic process, cell part, organelle, catalytic activity and binding. The KEGG annotation analysis of DEGs found only one pathway, amino acid metabolism, was up-regulated. The relative mRNA expression levels of ADAMTS15, Caspase-6, Claudin-5 and Prodh1 were verified by qRT-PCR, which were consistent with the RNA-seq results. In addition, the relative expression of ADAMTS15 was negatively correlated with the level of cardiac IL-1ß (r = -0.748, P = 0.005) and positively correlated with the level of cardiac IL-10 (r = 0.698, P = 0.012). A negative correlation statistical trend was found between the relative expression of ADAMTS15 and the level of cardiac IL-6 (r = -0.545, P = 0.067). Conclusions: ADAMTS15 may be a potential inflammation-related gene in regulation of cardioprotection conferred by remote ischemic postconditioning and a possible therapeutic target for myocardial ischemia reperfusion injury in the future.

Biallelic variants in ADAMTS15 cause a novel form of distal arthrogryposis.

PURPOSE: We aimed to identify the underlying genetic cause for a novel form of distal arthrogryposis. METHODS: Rare variant family-based genomics, exome sequencing, and disease-specific panel sequencing were used to detect ADAMTS15 variants in affected individuals. Adamts15 expression was analyzed at the single-cell level during murine embryogenesis. Expression patterns were characterized using in situ hybridization and RNAscope. RESULTS: We identified homozygous rare variant alleles of ADAMTS15 in 5 affected individuals from 4 unrelated consanguineous families presenting with congenital flexion contractures of the interphalangeal joints and hypoplastic or absent palmar creases. Radiographic investigations showed physiological interphalangeal joint morphology. Additional features included knee, Achilles tendon, and toe contractures, spinal stiffness, scoliosis, and orthodontic abnormalities. Analysis of mouse whole-embryo single-cell sequencing data revealed a tightly regulated Adamts15 expression in the limb mesenchyme between embryonic stages E11.5 and E15.0. A perimuscular and peritendinous expression was evident in in situ hybridization in the developing mouse limb. In accordance, RNAscope analysis detected a significant coexpression with Osr1, but not with markers for skeletal muscle or joint formation. CONCLUSION: In aggregate, our findings provide evidence that rare biallelic recessive trait variants in ADAMTS15 cause a novel autosomal recessive connective tissue disorder, resulting in a distal arthrogryposis syndrome.

Transcriptome Analysis of Hypoxic Lymphatic Endothelial Cells Indicates Their Potential to Contribute to Extracellular Matrix Rearrangement.

Lymphedema (LE) affects millions of people worldwide. It is a chronic progressive disease with massive development of fibrosclerosis when untreated. There is no pharmacological treatment of lymphedema. The disease is associated with swelling of the interstitium of the affected organ, mostly arm or leg, impressive development of adipose tissue, fibrosis and sclerosis with accumulation of huge amounts of collagen, and Papillomatosis cutis. Malnutrition and reduced oxygenation of the affected tissues is a hallmark of lymphedema. Here, we investigated if the hypoxia of lymphatic endothelial cells (LECs) might contribute to fibrosis. We applied RNASeq and qPCR to study the concordant changes of the exome of three human foreskin-derived LEC isolates after 4 days of hypoxia (1% O2) vs. normoxia (21% O2). Of the approximately 16,000 genes expressed in LECs, 162 (1%) were up- or down-regulated by hypoxia. Of these, 21 genes have important functions in the production or modification of the extracellular matrix (ECM). In addition to the down-regulation of elastin, we found up-regulation of druggable enzymes and regulators such as the long non-coding RNA H19, inter-alpha-trypsin inhibitor heavy chain family member 5 (ITIH5), lysyl-oxidase (LOX), prolyl 4-hydroxylase subunit alpha 1 (P4HA1), procollagen-lysine 2-oxoglutarate 5-dioxygenase 2 (PLOD2), and others that are discussed in the paper. Initial lymphatics do not produce a continuous basement membrane; however, our study shows that hypoxic LECs have an unexpectedly high ability to alter the ECM.