Genome-wide map of R-loops reveals its interplay with transcription and genome integrity during germ cell meiosis.

INTRODUCTION: The R-loop is a naturally formed three-strand nucleic acid structure that recently has been reported to participate in multiple biological processes and helped answer some previously unexplained scientific questions. Meiosis process involves multiple chromatin-related events such as DNA double-stranded breaks (DSB) formation, repairing and transcriptional dynamics. OBJECTIVES: Explore the regulatory roles and physiological functions of R-loops in the mammalian meiosis process. METHODS: In our study, using genome-wide S9.6 CUT & Tag seq, we first mapped the genomic distribution and dynamic changes of R-loop during the meiotic process in mice, from spermatogonia to secondary spermatocytes. And we further explore the role of R-loop in physiological conditions by constructing conditional knockout mice of Rnaseh1, which deleted the R-loop endonuclease before meiosis entry. RESULTS: R-loop predominantly distributes at promoter-related regions and varies across different meiotic stages. By joint analysis with the corresponding transcriptome, we found that the R-loop was closely related to transcription during the meiotic process. The high frequency of promoter-related R-loop in meiotic cells is usually accompanied by high transcription activity, and we further verified this in the leptotene/zygotene to the pachytene transition process. Moreover, the lack of RNase H1 caused sterility in male mice with R-loop accumulation and abnormal DSB repair in spermatocytes. Further analysis showed that abnormal R-loop accumulation in the leptotene/zygotene stages influenced transcriptional regulation in the pachytene stage. CONCLUSION: The mutual regulation of the R-loop and transcription plays an essential role in spermatogenesis. And R-loop is also important for the normal repair process of DSB during meiosis.

Injection route affects intra-articular hyaluronic acid distribution and clinical outcome in viscosupplementation treatment for knee osteoarthritis: a combined cadaver study and randomized clinical trial.

The coverage of hyaluronic acid (HA) on the impaired cartilage should be the precondition to exert its beneficial effect on knee osteoarthritis (KOA) according to the pharmacological mechanism. However, the intra-articular distribution of HA might be correlated with the route of drug delivery. Forty-two cadaver knees with radiographic evidence of osteoarthritis were given anteromedial (AM) or medial midpatellar (MMP) injection of HA (molecular weight 600-1500 kD) followed by gait stimulation. Although 2.5 ml HA delivered through both routes failed to cover the entire cartilage, HA covered 96.12% cartilage of patellofemoral joint (PFJ) and 71.44% of medial femorotibial joint (FTJ) through MMP route, whereas mainly distributed into FTJ and posterior condyles through AM route. HA in the MMP group distributed more in PFJ than that in the AM group (P < 0.001), but no significant difference presented in medial FTJ (P = 0.084). The clinical efficacy was also associated with the route of drug delivery. One hundred patients with unilateral mild-to-moderate KOA were recruited and randomly assigned to receive five weekly HA injections with AM route (n = 50) or MMP route (n = 50). Patients in the MMP group obtained better improvement in WOMAC index total score, pain score, stiffness score, and Lequesne index total score over the entire follow-up period, as compared to patients in the AM group (all P < 0.01). More patients in the MMP group claimed pain relief (71.7%, P = 0.024) and felt satisfying (63.1%, P = 0.007) than in the AM group at the end of follow-up. Therefore, intra-articular HA injection through MMP route is recommended in treating mild-to-moderate KOA. Graphical Abstract .