Itaconate ameliorates autoimmunity by modulating T cell imbalance via metabolic and epigenetic reprogramming.

Dysregulation of Th17 and Treg cells contributes to the pathophysiology of many autoimmune diseases. Herein, we show that itaconate, an immunomodulatory metabolite, inhibits Th17 cell differentiation and promotes Treg cell differentiation by orchestrating metabolic and epigenetic reprogramming. Mechanistically, itaconate suppresses glycolysis and oxidative phosphorylation in Th17- and Treg-polarizing T cells. Following treatment with itaconate, the S-adenosyl-L-methionine/S-adenosylhomocysteine ratio and 2-hydroxyglutarate levels are decreased by inhibiting the synthetic enzyme activities in Th17 and Treg cells, respectively. Consequently, these metabolic changes are associated with altered chromatin accessibility of essential transcription factors and key gene expression in Th17 and Treg cell differentiation, including decreased RORγt binding at the Il17a promoter. The adoptive transfer of itaconate-treated Th17-polarizing T cells ameliorates experimental autoimmune encephalomyelitis. These results indicate that itaconate is a crucial metabolic regulator for Th17/Treg cell balance and could be a potential therapeutic agent for autoimmune diseases.

Acquired factor V inhibitor with erythema and eosinophilia in a patient with end-stage renal disease.

Autoimmune factor V deficiency (AiFVD) is a rare bleeding disorder caused by factor V inhibitors. In this report, we present the case of an 89-year-old man who developed bleeding tendency during surgery to create arteriovenous fistula for hemodialysis. The bleeding tendency developed with prolongation of activated partial thromboplastin and prothrombin time, following drug-induced eruption and eosinophilia. Significant reduction in coagulation factor activity and inhibitory pattern in cross-mixing tests suggested the presence of inhibitors to coagulation factors. Subsequently, we detected a factor V inhibitor and anti-factor V autoantibodies was confirmed using enzyme-linked immunosorbent assay with purified human plasma factor V. Thus, the patient was 'definitely diagnosed' with AiFVD in accordance with the diagnostic criteria enacted by the Japanese Ministry of Health, Labor, and Welfare. The bleeding tendency improved after initiating oral prednisolone 50 mg (1 mg/kg) followed by normalization of activated partial thromboplastin time and prothrombin time at the 34th day. After improving the coagulation system prolongation, the inhibitor and autoantibodies has been eradicated. Since it is suggested that drug-induced immune response can cause AiFVD, AiFVD should be considered in patients who undergo hemodialysis and develop failure of hemostasis and drug-induced eruption.

Acute compartment syndrome of the lower limb after revision anterior cruciate ligament reconstruction: a case report.

Introduction: Acute compartment syndrome (ACS) is one of the most serious orthopedic diseases causing complications and requiring emergency surgery. Most cases of ACS are associated with fractures and crush injuries. However, surgical procedures can also cause ACS. Case presentation: We herein describe a 41-year-old man who underwent arthroscopic surgery for revision anterior cruciate ligament reconstruction with the semitendinosus tendon. Two days after the operation, the patient developed ACS of the left lower leg. This patient had undergone anterior cruciate ligament(ACL) reconstruction previously, and we assumed that the cause of the ACS was damage to the small blood vessels from the collateral circulation during hamstring tendon harvesting. Fasciotomy was performed under general anesthesia, and his postoperative course was uneventful. Conclusion: ACS after revision anterior cruciate ligament reconstruction is rare. We should keep in mind that patients who have undergone previous surgery may develop ACS due to damage to the collateral circulation. To avoid ACS, harvesting tendon from the healthy side can avoid damage to the collateral circulation.

Changes in patellar baja progress until 3 months after medial open-wedge high tibial osteotomy.

PURPOSE: The purpose of this study was to evaluate sequential patellar height changes as well as the factors leading to these changes after medial open-wedge high tibial osteotomy (MOWHTO). METHODS: The study cohort constituted 37 knees from 36 patients who underwent MOWHTO for varus knee. The Caton-Deschamps index (CDI) for patellar height was measured preoperatively and at 2 weeks and 3, 6, and 12 months postoperatively. The factors related to sequential changes in patellar height were evaluated. RESULTS: Significant differences were observed between preoperative CDI and postoperative CDI at all time points (p < .05). Two-week postoperative CDI and postoperative CDI at 3,6,12-months was also significantly different (p < .05). The only significant factor for the change in patellar height between preoperative CDI and postoperative CDI at 2-weeks and 12-months was the change in the Δ medial proximal tibial angle (ΔMPTA) (postoperative MPTA-preoperative MPTA). We could not identify the factor that affected the change in patellar height change from 2-weeks postoperatively. CONCLUSION: The change in patellar height continued sequentially until at least 3 months postoperatively. ΔMPTA was associated with the change in patellar height at 2 weeks postoperatively compared with preoperatively; however, no factors associated with the change in patellar height from 2 weeks postoperatively to 3, 6, and 12-months postoperatively were identified.

Protective Role of Optineurin Against Joint Destruction in Rheumatoid Arthritis Synovial Fibroblasts.

OBJECTIVE: Optineurin (OPTN) is an autophagy adaptor/receptor that acts as an intrinsic negative regulator of osteoclast differentiation. RANKL expressed by rheumatoid arthritis synovial fibroblasts (RASFs) is primarily responsible for the development of bone erosions in patients with RA. The aim of the present study was to explore the role of OPTN in the pathogenesis of joint destruction in RA. METHODS: RASFs were left untreated or incubated with tumor necrosis factor (TNF) or interferon-γ (IFNγ), and expression of OPTN by RASFs was analyzed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and Western blotting. Expression of RANKL and osteoprotegerin (OPG) was evaluated in cultures of OPTN-reduced RASFs with or without TNF or IFNγ treatment. OPTN-reduced RASFs were cocultured with monocytes and stained for tartrate-resistant acid phosphatase (TRAP). IκBα, NF-κB1, and RelA protein levels were measured to evaluate NF-κB signaling. Expression of messenger RNA (mRNA) for matrix metalloproteinase 3 (MMP3), interleukin-6 (IL6), GATA binding protein 3 (GATA3), carbohydrate sulfotransferase 15 (CHST15), hyaluronan synthase 1 (HAS1), and GATA1 was analyzed by RT-qPCR. RESULTS: In RASFs incubated with TNF or IFNγ, OPTN expression was up-regulated and RANKL expression was increased, and these effects were further pronounced in OPTN-reduced RASFs (all P < 0.05 versus controls). OPG mRNA levels remained unchanged. Monocytes cocultured with OPTN-reduced RASFs differentiated to a greater extent into TRAP+ multinucleated cells compared to monocytes cocultured with control RASFs (P < 0.05). IκBα degradation and nuclear NF-κB1 expression following TNF treatment were both prolonged in OPTN-reduced RASFs (each P < 0.05 versus controls). MMP3 mRNA levels were up-regulated, while GATA3, CHST15, and HAS1 mRNA levels were down-regulated in OPTN-reduced RASFs (each P < 0.05 versus controls). CONCLUSION: OPTN plays a protective role in RA when it is up-regulated in RASFs in the presence of proinflammatory cytokines. Absence of OPTN might worsen RA by generating a joint-destructive state, as indicated by evidence of increased RANKL expression on RASFs and subsequent osteoclast differentiation.

MicroRNA-9 ameliorates destructive arthritis through down-regulation of NF-κB1-RANKL pathway in fibroblast-like synoviocytes.

We investigated the effect of miR-9 on fibroblast-like synoviocytes (FLS) from RA patients and animal arthritis model. The binding of miR-9 to NF-κB1 3'UTR was analyzed by luciferase reporter assay and immunoprecipitation. ChIP assay and luciferase promoter assay were performed to identify the binding of NF-κB1 to RANKL promoter and its activity. FLS were treated with miR-9/anti-miR-9 to evaluate cell proliferation and the expression of RANKL. Therapeutic effect of intra-articular miR-9 was evaluated in type-II collagen-induced arthritis in rats. miR-9 bound to the 3'-UTR of NF-κB1 and downregulated NF-κB1. NF-κB1 bound to RANKL promoter and increased the promoter activity of RANKL. RANKL was downregulated by miR-9. Proliferation of FLS was increased by miR-9 inhibitor. miR-9 dampened experimental arthritis by lowering inflammatory state, reducing RANKL and osteoclasts formation. Our findings revealed miR-9-NF-κB1-RANKL pathway in RA-FLS, further, miR-9 ameliorated inflammatory arthritis in vivo which propose therapeutic implications of miR- 9 in RA.

CHK1 cleavage in programmed cell death is intricately regulated by both caspase and non-caspase family proteases.

BACKGROUND: CHK1 is an important effector kinase that regulates the cell cycle checkpoint. Previously, we showed that CHK1 is cleaved in a caspase (CASP)-dependent manner during DNA damage-induced programmed cell death (PCD) and have examined its physiological roles. METHODS AND RESULTS: In this study, we investigated the behavior of CHK1 in PCD. Firstly, we found that CHK1 is cleaved at three sites in PCD, and all cleavages were inhibited by the co-treatment of a pan-CASP inhibitor or serine protease inhibitors. We also showed that CHK1 is cleaved by CASP3 and/or CASP7 recognizing at (296)SNLD(299) and (348)TCPD(351), and that the cleavage results in the enhancement of CHK1 kinase activity. Furthermore, as a result of the characterization of cleavage sites by site-directed mutagenesis and an analysis performed using deletion mutants, we identified (320)EPRT(323) as an additional cleavage recognition sequence. Considering the consensus sequence cleaved by CASP, it is likely that CHK1 is cleaved by non-CASP family protease(s) recognizing at (320)EPRT(323). Additionally, the cleavage catalyzed by the (320)EPRT(323) protease(s) markedly and specifically increased when U2OS cells synchronized into G1 phase were induced to PCD by cisplatin treatment. CONCLUSION: CHK1 cleavage is directly and indirectly regulated by CASP and non-CASP family proteases including serine protease(s) and the "(320)EPRT(323) protease(s)." Furthermore, (320)EPRT(323) cleavage of CHK1 occurs efficiently in PCD which is induced at the G1 phase by DNA damage. GENERAL SIGNIFICANCE: CASP and non-CASP family proteases intricately regulate cleavage for up-regulation of CHK1 kinase activity during PCD.

DCK is frequently inactivated in acquired gemcitabine-resistant human cancer cells.

Although gemcitabine is the most effective chemotherapeutic agent against pancreatic cancer, a growing concern is that a substantial number of patients acquire gemcitabine chemoresistance. To elucidate the mechanisms of acquisition of gemcitabine resistance, we developed gemcitabine-resistant cell lines from six human cancer cell lines; three pancreatic, one gastric, one colon, and one bile duct cancer. We first analyzed gemcitabine uptake using three paired parental and gemcitabine resistant pancreatic cancer cell lines (PK-1 and RPK-1, PK-9 and RPK-9, PK-59 and RPK-59) and found that uptake of gemcitabine was rapid. However, no DNA damage was induced in resistant cells. We further examined the microarray-based expression profiles of the cells to identify genes associated with gemcitabine resistance and found a remarkable reduction in the expression of deoxycytidine kinase (DCK). DCK is a key enzyme that activates gemcitabine by phosphorylation. Genetic alterations and expression of DCK were studied in these paired parental and derived gemcitabine-resistant cell lines, and inactivating mutations were found only in gemcitabine-resistant cell lines. Furthermore, siRNA-mediated knockdown of DCK in the parental cell lines yielded gemcitabine resistance, and introduction of DCK into gemcitabine-resistant cell lines invariably restored gemcitabine sensitivities. Mutation analyses were expanded to three other different paired cell lines, DLD-1 and RDLD-1 (colon cancer cell line), MKN-28 and RMKN-28 (gastric cancer cell line), and TFK-1 and RTFK -1 (cholangiocarcinoma cell line). We found inactivating mutations in RDLD-1 and RTFK-1 and decreased expression of DCK in RMKN-28. These results indicate that the inactivation of DCK is one of the crucial mechanisms in acquisition of gemcitabine resistance.