Glycosylated queuosines in tRNAs optimize translational rate and post-embryonic growth.

Transfer RNA (tRNA) modifications are critical for protein synthesis. Queuosine (Q), a 7-deaza-guanosine derivative, is present in tRNA anticodons. In vertebrate tRNAs for Tyr and Asp, Q is further glycosylated with galactose and mannose to generate galQ and manQ, respectively. However, biogenesis and physiological relevance of Q-glycosylation remain poorly understood. Here, we biochemically identified two RNA glycosylases, QTGAL and QTMAN, and successfully reconstituted Q-glycosylation of tRNAs using nucleotide diphosphate sugars. Ribosome profiling of knockout cells revealed that Q-glycosylation slowed down elongation at cognate codons, UAC and GAC (GAU), respectively. We also found that galactosylation of Q suppresses stop codon readthrough. Moreover, protein aggregates increased in cells lacking Q-glycosylation, indicating that Q-glycosylation contributes to proteostasis. Cryo-EM of human ribosome-tRNA complex revealed the molecular basis of codon recognition regulated by Q-glycosylations. Furthermore, zebrafish qtgal and qtman knockout lines displayed shortened body length, implying that Q-glycosylation is required for post-embryonic growth in vertebrates.

Celecoxib does not affect the release of hyaluronic acid in end stage osteoarthritic joints.

OBJECTIVES: Non-steroidal anti-inflammatory drugs play a major role in the management of osteoarthritis (OA). However, it remains unknown whether these drugs affect cartilage and synovial metabolism in osteoarthritic joints. The aim of this study was to evaluate the effects of a selective cyclooxygenase (COX)-2 inhibitor, celecoxib, on synovial fluids and tissues in severely osteoarthritic knees. METHODS: Patients were randomized into three groups and medicated two weeks prior to total knee arthroplasty with celecoxib, diclofenac sodium, or no medication (control). We checked for the presence of matrix metalloproteinase-3 (MMP-3), tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and hyaluronic acid (HA) in the synovial fluids of all three groups. RESULTS: MMP-3 significantly decreased in the celecoxib-treated patients (p = 0.0031). On the other hand, there were no significant differences among the three groups in their TNF-α and IL-1ß levels. HA in the joint fluid was significantly increased in the diclofenac-treated group, while HA was not changed in the celecoxib-treated patients. CONCLUSIONS: Our study suggests that celecoxib did not affect the level of HA in the joint fluid in the knee in severe OA, in contrast to the effect of the dual COX inhibitor.

Effect of selective estrogen receptor modulator/raloxifene analogue on proliferation and collagen metabolism of tendon fibroblast.

The selective estrogen receptor modulator raloxifene is therapeutically beneficial for postmenopausal connective tissue degradation, such as osteoporosis, vascular sclerosis, and dermal degradation; however, the effects of raloxifene on postmenopausal tendon metabolism have not been clarified. In this study, we investigated the effects of raloxifene analogue (LY117018) on cell proliferation and collagen metabolism using cultured rat Achilles tendon fibroblasts. 17beta-Estradiol (E(2); 10(-11)-10(-9) M) and LY117018 (10(-9)-10(-7) M) had no significant effects on tendon fibroblast proliferation, based on a BrdU (5-bromo-2'-deoxyuridine) incorporation assay (24 hr) and a WST-8 colorimetric assay (2 or 6 days). Neither E(2) nor LY117018 significantly altered the expression of type I collagen, which is a main component of the tendon extracellular matrix (ECM), whereas both E(2) and LY117018 significantly increased the expression of matrix metalloproteinase (MMP)-13, which is responsible for tendon collagen degradation in rat. Also, both E(2) and LY117018 increased the expression of type III collagen and elastin, which are minor components of tendon ECM, but are considered to govern the elastic properties of tendons. These changes in collagen and MMP induced by either E(2) or LY117018 were attenuated by the estrogen receptor alpha blocker ICI 182,780. The results of this study suggest that postmenopausal estrogen deficiency might downregulate tendon collagen turnover and decrease tendon elasticity. Further, raloxifene treatment might restore these changes to premenopausal levels.

Biomechanical and histologic evaluation of tissue engineered ligaments using chitosan and hyaluronan hybrid polymer fibers: a rabbit medial collateral ligament reconstruction model.

In this study, we used a rabbit medial collateral ligament reconstruction model to evaluate a novel chitosan-based hyaluronan hybrid polymer fiber scaffold for ligament tissue engineering and to examine whether mechanical forces exerted in an in vivo model increased extracellular matrix production by seeded fibroblasts. Scaffolds were used 2 weeks after incubation with fibroblasts obtained from the same rabbit in a cell-seeded scaffold (CSS) group and without cells in a noncell-seeded scaffold (NCSS) group. At 3, 6, and 12 weeks after surgery, the failure loads of the engineered ligaments in the CSS groups were significantly greater than those in the NCSS groups. At 6 weeks after surgery, the reconstructed tissue of the CSS group was positive for type I collagen, whereas that in the NCSS group was negative for type I collagen. At 12 weeks after surgery, the reconstructed tissue stained positive for type I collagen in the CSS group, but negative in the NCSS group. Our results indicate that the scaffold material enhanced the production of type I collagen and led to improved mechanical strength in the engineered ligament in vivo.