MutL homolog 1 participates in interference-sensitive meiotic crossover formation in soybean.

MutL homolog 1 (MLH1), a member of the MutL-homolog family, is required for normal recombination in most organisms. However, its role in soybean (Glycine max) remains unclear to date. Here, we characterized the Glycine max female and male sterility 1 (Gmfms1) mutation that reduces pollen grain viability and increases embryo sac abortion in soybean. Map-based cloning revealed that the causal gene of Gmfms1 is Glycine max MutL homolog 1 (GmMLH1), and CRISPR/Cas9 knockout approach further validated that disruption of GmMLH1 confers the female-male sterility phenotype in soybean. Loss of GmMLH1 function disrupted bivalent formation, leading to univalent mis-segregation during meiosis and ultimately to female-male sterility. The Gmmlh1 mutant showed about a 78.16% decrease in meiotic crossover frequency compared to the wild type. The residual chiasmata followed a Poisson distribution, suggesting that interference-sensitive crossover formation was affected in the Gmmlh1 mutant. Furthermore, GmMLH1 could interact with GmMLH3A and GmMLH3B both in vivo and in vitro. Overall, our work demonstrates that GmMLH1 participates in interference-sensitive crossover formation in soybean, and provides additional information about the conserved functions of MLH1 across plant species.

High femoral anteversion in osteoarthritic knees, particularly for severe valgus deformity.

OBJECTIVE: Increased femoral anteversion (FA) has been correlated with less varus deformities in osteoarthritic (OA) knees, but the relationship between FA and the degree of valgus deformity in osteoarthritic (OA) knees is still largely unknown. We aimed to thoroughly analyze the distribution of FA in relation to varus or valgus deformities of the lower extremity in OA knees, and to further clarify the relationship between FA and trochlear morphology. METHODS: 235 lower extremities with OA knees were divided into five groups according to the mechanical tibiofemoral angle: excessive valgus (< - 10°), moderate valgus (- 10° to - 3°), neutral (- 3° to 3°), moderate varus (3° to 10°), and excessive varus (> 10°). FA (measured using the posterior condylar axis [pFA] and the transepicondylar axis [tFA]) was measured, and the relationships of FA to the mechanical tibiofemoral angle and femoral trochlear morphology were identified. RESULTS: Excessive FA (pFA ≥ 20°) was observed in 30.2% of all patients and in 58.8% of patients in the excessive valgus group. pFA showed a strong correlation with mechanical tibiofemoral angle (p = 0.018). Both the pFA and the tFA of patients in the excessive valgus group were greater than those in other four groups (all p ≤ 0.037). There were significant correlations between tFA and trochlear parameters, including the sulcus angle (SA), lateral trochlear slope (LTS), and medial trochlear slope (MTS) (all p ≤ 0.028). CONCLUSION: High FA is prevalent, particularly in severe valgus knees, and FA is significantly related to the femoral trochlear morphology in OA knees. With the aim of improving the patellofemoral prognosis and complications, high FA should be considered during total knee arthroplasty.

Genetic Mapping of the Gmpgl3 Mutant Reveals the Function of GmTic110a in Soybean Chloroplast Development.

The generation of oxygen and organic matter in plants mainly depends on photosynthesis, which directly affects plant growth and development. The chloroplast is the main organelle in which photosynthesis occurs. In this study, a Glycine max pale green leaf 3-1 (Gmpgl3-1) mutant was isolated from the soybean mutagenized population. The Gmpgl3-1 mutant presented with decreased chlorophyll contents, reduced chloroplast stroma thylakoids, reduced yields, and decreased numbers of pods per plant. Bulked segregant analysis (BSA) together with map-based cloning revealed a single-nucleotide non-synonymous mutation at the 341st nucleotide of the first exon of the chloroplast development-related GmTic110a gene. The phenotype of the knockout plants was the same as that of the mutant. The GmTic110a gene was highly expressed in the leaves at various developmental stages, and its protein was localized to the inner chloroplast membrane. Split luciferase complementation assays and coimmunoprecipitation (co-IP) experiments revealed that GmTic110a interacted with GmTic20, GmTic40a, and GmTic40b in tobacco leaves. These results indicated that the GmTic110a gene plays an important role in chloroplast development.

miR-125a-5p Regulates Osteogenic Differentiation of Human Adipose-Derived Mesenchymal Stem Cells under Oxidative Stress.

Adipose-derived mesenchymal stem cells (ADSCs) are a well-recognized multilineage stem cell with vital clinical feasibility for tissue regeneration. Extensive evidence indicates that oxidative stress and microRNAs (miRNAs/miRs) play an important role in the osteoinduction of adipose-derived mesenchymal stem cells. In this study, we investigated the mechanism of miR-125a-5p in regulating the osteogenesis of human adipose-derived mesenchymal stem cells (hADSCs) under oxidative stress. The expression of miR-125a-5p lessened gradually during the osteogenic differentiation of hADSCs. Relative to the negative group, the expression levels of runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP), osteocalcin (OCN), and osterix in the miR-125a-5p group were marked lower than those in the miR-125a-5p inhibitor group. The levels of p16, p21, p53, miR-125a-5p, and ROS during osteoinduction of hADSCs were assessed in vitro under oxidative stress and were observed to be upregulated. Further experiments showed that oxidative stress and miR-125a-5p together suppressed the expression of VEGF during osteogenic differentiation of hADSCs and that the inhibition of miR-125a-5p reversed the effect of oxidative stress. In short, our study indicated that miR-125a-5p is induced under oxidative stress and inhibits the expression of VEGF, leading to the reduction of osteogenic differentiation of hADSCs. Our outcomes showed that miR-125a-5p could be a potential clinical target for bone repairing.

Urinary miRNAs as biomarkers for idiopathic osteonecrosis of femoral head: A multicentre study.

OBJECTIVES: Urinary microRNAs (miRNAs) have shown great diagnostic and prognostic values for multiple diseases. The profile of urinary miRNAs in patients with idiopathic osteonecrosis of femoral head (ONFH) is currently unclear. METHODS: We first randomly chose ten patients with each Association Research Circulation Osseous (ARCO) stage (I, II, III and IV) and ten healthy participants from the entire cohorts for initial screening. The miRNA polymerase chain reaction (PCR) array was then performed to identify the differentially abundant miRNAs in urine of these participants. We then verified the findings in the entire cohort. Clinical features including age, gender, bone mass index (BMI), lesion size and stages were recorded. We then analysed the association between the level of urinary miRNAs and clinical features. RESULTS: Our data indicated that there were 13 differentially abundant miRNAs among all groups. Urinary miR-150 demonstrated the highest diagnostic value among all candidates. Urinary miR-185 and miR-133a increased by ARCO staging. The levels of urinary miR-4824 abruptly decreased after femoral head collapse (ARCO stage III and IV). Urinary miR-144 was the only marker that correlated with lesion size. CONCLUSIONS: The levels of urinary miRNAs are valuable biomarkers for idiopathic ONFH. Given the noninvasive nature of this test, it is potentially useful for diagnosis and monitoring of idiopathic ONFH progression. THE TRANSLATIONAL POTENTIAL OF THIS ARTICLE: This article gives novel methods for ONFH diagnosis and progression monitoring in a convenient and non-invasive way.

PSMA-Targeting Reduction-Cleavable Hyperbranched Polyamide-Amine Gene Delivery System to Treat the Bone Metastases of Prostate Cancer.

OBJECTIVE: This study aimed to develop aptamer-anchored hyperbranched poly(amido amine) (HPAA) for the systemic delivery of miRNA-133a-3p and to evaluate its therapeutic potential against bone metastasis of prostate cancer in vivo and in vitro. METHODS: A glutathione (GSH)-responsive cationic HPAA was prepared by the Michael addition reaction. Furthermore, HPAA-PEG was produced by PEGylation, and then the aptamer targeted to prostate-specific membrane antigen (PSMA) was conjugated to the HPAA-PEG. The obtained HPAA-PEG-APT could form nanocomplexes with miRNA-133a-3p through electrostatic adsorption. RESULTS: The results of immunocytochemistry indicated that the complexes could target PSMA-expressing LNCaP cells. The ability of HPAA-PEG-APT to facilitate the delivery of miRNA-133a-3p into LNCaP cells was proven, and HPAA-PEG-APT/miRNA-133a-3p demonstrated enhanced antitumor activity, lower cytotoxicity and better biocompatibility in vitro. Moreover, in a mouse tibial injection tumor model, the intravenous injection of the HPAA-PEG-APT/miRNA-133a-3p complex significantly inhibited cancer growth and extended the survival time. CONCLUSION: This study provided an aptamer-anchored HPAA-loaded gene system to deliver miRNA-133a-3p for better therapeutic efficacy of bone metastasis of prostate cancer.

Long non-coding RNA reprogramming (lncRNA-ROR) regulates cell apoptosis and autophagy in chondrocytes.

Long Non-Coding RNA Reprogramming (lncRNA-ROR) plays an important role in regulating various biologic processes, whereas the effect of lncRNA-ROR in osteoarthritis (OA) is little studied. This study aimed to explore lncRNA-ROR expression in articular cartilage and identify the functional mechanism of lncRNA-ROR in OA. OA cartilage tissues were obtained from 15 OA patients, and 6 normal cartilage tissues were set as controls. Chondrocytes were isolated from the collected cartilage tissues. lncRNA-ROR was knockdown in normal cells and overexpressed in OA cells. Cell viability was determined with Cell Counting Kit-8 assay, and apoptosis was measured using flow cytometric analysis. Moreover, proteins and mRNAs involved in this study were also measured using Western blotting and quantitative real-time PCR (qPCR). Level of lncRNA-ROR was decreased in OA compared with normal chondrocytes, and overexpression of lncRNA-ROR dramatically promoted cell viability of OA chondrocytes. In addition, knockdown lncRNA-ROR inhibited apoptosis and promoted autophagy of normal chondrocytes. Moreover, lncRNA-ROR inhibited the expression of p53 in both mRNA and protein levels. Furthermore, we revealed that lncRNA-ROR regulated apoptosis and autophagy of chondrocytes via HIF1α and p53. The results indicated that lncRNA-ROR played a critical role in the pathogenesis of OA, suggesting that lncRNA-ROR could serve as a new potential therapeutic target for OA.

CRMP4 and CRMP2 Interact to Coordinate Cytoskeleton Dynamics, Regulating Growth Cone Development and Axon Elongation.

Cytoskeleton dynamics are critical phenomena that underpin many fundamental cellular processes. Collapsin response mediator proteins (CRMPs) are highly expressed in the developing nervous system, mediating growth cone guidance, neuronal polarity, and axonal elongation. However, whether and how CRMPs associate with microtubules and actin coordinated cytoskeletal dynamics remain unknown. In this study, we demonstrated that CRMP2 and CRMP4 interacted with tubulin and actin in vitro and colocalized with the cytoskeleton in the transition-zone in developing growth cones. CRMP2 and CRMP4 also interacted with one another coordinately to promote growth cone development and axonal elongation. Genetic silencing of CRMP2 enhanced, whereas overexpression of CRMP2 suppressed, the inhibitory effects of CRMP4 knockdown on axonal development. In addition, knockdown of CRMP2 or overexpression of truncated CRMP2 reversed the promoting effect of CRMP4. With the overexpression of truncated CRMP2 or CRMP4 lacking the cytoskeleton interaction domain, the promoting effect of CRMP was suppressed. These data suggest a model in which CRMP2 and CRMP4 form complexes to bridge microtubules and actin and thus work cooperatively to regulate growth cone development and axonal elongation.

Katanin p60 promotes neurite growth and collateral formation in the hippocampus.

OBJECTIVE: This study aimed to investigate the effect of Katanin p60 on the neurite growth and collateral formation in the hippocampus. METHODS: Gene cloning was performed to construct the Katanin p60 eukaryotic vector. The microtubule cutting effect and protein expression of Katanin p60 were investigated in 293T cells. Then, these vectors were transfected into hippocampal neurons of rats, and the effects of Katanin p60 on the neurite growth and collateral formation were observed. RESULTS: In the present study, we successfully constructed Katanin p60-GFP recombinant plasmids. After transfecting into 293T cells, the Katanin p60 was over-expressed in these cells, the mesh-like structure of microtubules was disrupted, the residual microtubules circled the nucleus, the expression microtubule proteins reduced, and the tapered protrusions disappeared. In hippocampal neurons with Katanin p60 over-expression, the neural neurite growth was obvious, and a lot of dendrites arose from cell bodies. In cells without Katanin p60 expression, the neurites were small, and the number and length of dendrites reduced significantly when compared with Katanin p60 over-expressing cells (P < 0.05). In addition, in Katanin p60 over-expressing cells, the number of collaterals from the neurites and dendrites increased markedly when compared with cells without Katanin p60 expression (P < 0.05). CONCLUSION: Katanin p60 can promote the neurite growth and collateral formation of hippocampal neurons.