Osteoclast-derived exosomal miR-212-3p suppressed the anabolism and accelerated the catabolism of chondrocytes in osteoarthritis by targeting TGF-ß1/Smad2 signaling.

Osteoarthritis (OA) is a common aging-related disease affecting entire joint structures, encompassing articular cartilage and subchondral bone. Although senescence and dysfunction of chondrocytes are considered crucial factors in the occurrence of OA, the exact pathogenesis remains to be investigated. In our study, chondrocytes were incubated with a conditioned medium obtained from osteoclasts at different differentiation stages, suggesting that osteoclasts and osteoclast precursors suppressed anabolism and promoted the catabolism of chondrocytes in vitro. In contrast, the function of osteoclasts was more significant than osteoclast precursors. Further blocking of osteoclast exosome secretion by using GW4869 abolished the effect of osteoclasts on chondrocytes. Functionally, exosomal transfer of osteoclast-derived miR-212-3p inhibited Smad2 to mediate chondrocyte dysfunction, thus accelerating cartilage matrix degradation in OA via TGF-ß1/Smad2 signaling. The mechanism was also confirmed within the articular cartilage in OA patients and surgery-induced OA mice. Our study provides new information on intercellular interactions in the bone microenvironment within articular cartilage and subchondral bone during OA progression. The miR-212-3p/Smad2 axis is a potential target for the prevention and therapy of OA.

Antagonizing exosomal miR-18a-5p derived from prostate cancer cells ameliorates metastasis-induced osteoblastic lesions by targeting Hist1h2bc and activating Wnt/ß-catenin pathway.

More than 50% of prostate cancer (PCa) patients have bone metastasis with osteoblastic lesions. MiR-18a-5p is associated with the development and metastasis of PCa, but it remains unclear whether it is involved in osteoblastic lesions. We first found that miR-18a-5p was highly expressed in the bone microenvironment of patients with PCa bone metastases. To address how miR-18a-5p affects PCa osteoblastic lesions, antagonizing miR-18a-5p in PCa cells or pre-osteoblasts inhibited osteoblast differentiation in vitro. Moreover, injection of PCa cells with miR-18a-5p inhibition improved bone biomechanical properties and bone mineral mass in vivo. Furthermore, miR-18a-5p was transferred to osteoblasts by exosomes derived from PCa cells and targeted the Hist1h2bc gene, resulting in Ctnnb1 up-regulation in the Wnt/ß-catenin signaling pathway. Translationally, antagomir-18a-5p significantly improved bone biomechanical properties and alleviated sclerotic lesions from osteoblastic metastases in BALB/c nude mice. These data suggest that inhibition of exosome-delivered miR-18a-5p ameliorates PCa-induced osteoblastic lesions.

Inflammatory osteoclasts-derived exosomes promote bone formation by selectively transferring lncRNA LIOCE into osteoblasts to interact with and stabilize Osterix.

Bone loss is a hallmark of inflammatory bone diseases caused by aberrantly activated osteoclasts (OCLs). Studies have shown that OCLs exhibit various phenotypes and functions due to variations in the source(s) of precursor cells, cytokine expressions, and microenvironment-dependent factors. During these conditions, inflammatory osteoclasts (iOCLs) lose their immune-suppressive effect relative to OCLs under physiological conditions. This induces TNF α-producing CD4+ T cells in an antigen-dependent manner and finally leads to cascade amplification of iOCLs. OCL-derived exosomes have been reported to regulate OCL formation and inhibit the osteoblast activity. However, the specific function and mechanism of iOCL-derived exosomes on osteoblast have not been studied yet. In the present study, we compare the osteoblast promoting activities of iOCL-derived exosomes and OCL-derived exosomes. We found that iOCLs exosomes specifically target osteoblasts through ephrinA2/EphA2. Mechanistically, the lncRNA LIOCE is enriched in iOCL exosomes and promotes the osteoblast activity after being incorporated into osteoblasts. Furthermore, our results revealed that exosomal lncRNA LIOCE stabilizes osteogenic transcription factor Osterix by interacting and reducing the ubiquitination level of Osterix. This study demonstrated that the bone loss is alleviated in the inflammatory osteolysis mice model after injection of iOCL exosomes encapsulating lncRNA LIOCE. The role of exosomes encapsulating lncRNA LIOCE in promoting bone formation was well established in the rat bone repair model. Our results indicate that iOCL-derived exosomal lncRNA LIOCE promotes bone formation by upregulating Osx expression, and thus, the exosomes encapsulating lncRNA LIOCE may be an effective strategy to increase bone formation in osteoporosis and other bone metabolic disorders.

Angelicin Alleviates Post-Trauma Osteoarthritis Progression by Regulating Macrophage Polarization via STAT3 Signaling Pathway.

Post-trauma osteoarthritis (PTOA) is the most common articular disease characterized by degeneration and destruction of articular cartilage (Bultink and Lems, Curr. Rheumatol Rep., 2013, 15, 328). Inflammatory response of local joint tissue induced by trauma is the most critical factor accelerating osteoarthritis (OA) progression (Sharma et al., 2019; Osteoarthritis. Cartilage, 28, 658-668). M1/M2 macrophages polarization and repolarization participates in local inflammation, which plays a major role in the progression of OA (Zhang et al., 2018; Ann. Rheum. Dis., 77, 1524-1534). The regulating effect of macrophage polarization has been reported as a potential therapy to alleviate OA progression. Synovitis induced by polarized macrophages could profoundly affect the chondrocyte and cartilage matrix (Zhang et al., 2018; Ann. Rheum. Dis., 77, 1524-1534). Generally, anti-inflammatory medications widely used in clinical practice have serious side effects. Therefore, we focus on exploring a new therapeutic strategy with fewer side effects to alleviate the synovitis. Angelicin (ANG) is traditional medicine used in various folk medicine. Previous studies have revealed that angelicin has an inhibitory effect on inflammation (Wei et al., 2016; Inflammation, 39, 1876-1882), tumor growth (Li et al., 2016; Oncology reports, 36, 3,504-3,512; Wang et al., 2017; Molecular Medicine Reports, 16, 5441-5449), DNA damage (Li et al., 2019; Exp. Ther. Med., 18, 1899-1906), and virus proliferation (Li et al., 2018; Front. Cell. Infect. Microbiol., 8, 178). But its specific effects on influencing the process of OA were rarely reported. In this study, the molecular mechanism of angelicin in vivo and in vitro was clearly investigated. Results showed that angelicin could regulate the M1/M2 ratio and function and alleviate the development of PTOA in the meanwhile. Bone marrow monocytes were isolated and induced by macrophage colony-stimulating factor (M-CSF), lipopolysaccharide (LPS) and interferon (IFN)-γ for M1 polarization and interleukin (IL)-4/IL-13 for M2 polarization. Subsequently, repolarization intervention was performed. The results indicate that angelicin can repolarize M1 toward M2 macrophages by upregulating the expression of CD9. Besides, angelicin can also protect and maintain M2 polarization in the presence of LPS/IFN-γ, and subsequently downregulate the expression of inflammatory mediators such as IL-1ß and TNF-α. Mechanistically, angelicin can activate the p-STAT3/STAT3 pathway by conducting CD9/gp130 to repolarize toward M2 macrophages. These results suggest angelicin can alleviate the progression of OA by regulating M1/M2 polarization via the STAT3/p-STAT3 pathway. Therefore, angelicin may have a promising application and potential therapeutic value in OA clinical treatment.

Epothilone B prevents lipopolysaccharide-induced inflammatory osteolysis through suppressing osteoclastogenesis via STAT3 signaling pathway.

Inflammatory osteolysis is a common osteolytic specificity that occurs during infectious orthopaedic surgery and is characterized by an imbalance in bone homeostasis due to excessive osteoclast bone resorption activity. Epothilone B (Epo B) induced α-tubulin polymerization and enhanced microtubule stability, which also played an essential role in anti-inflammatory effect on the regulation of many diseases. However, its effects on skeletal system have rarely been investigated. Our study demonstrated that Epo B inhibited osteoclastogenesis in vitro and prevented inflammatory osteolysis in vivo. Further analysis showed that Epo B also markedly induced mature osteoclasts apoptosis during osteoclastogenesis. Mechanistically, Epo B directly suppressed osteoclastogenesis by the inhibitory regulation of the phosphorylation and activation of PI3K/Akt/STAT3 signaling directly, and the suppressive regulation of the CD9/gp130/STAT3 signaling pathway indirectly. The negative regulatory effect on STAT3 signaling further restrained the translocation of NF-κB p65 and NFATc1 from the cytosol to the nuclei during RANKL stimulation. Additionally, the expression of osteoclast specific genes was also significantly attenuated during osteoclast fusion and differentiation. Taken together, these findings illustrated that Epo B protected against LPS-induced bone destruction through inhibiting osteoclastogenesis via regulating the STAT3 dependent signaling pathway.

Engineered scaffolds based on mesenchymal stem cells/preosteoclasts extracellular matrix promote bone regeneration.

Recently, extracellular matrix-based tissue-engineered bone is a promising approach to repairing bone defects, and the seed cells are mostly mesenchymal stem cells. However, bone remodelling is a complex biological process, in which osteoclasts perform bone resorption and osteoblasts dominate bone formation. The interaction and coupling of these two kinds of cells is the key to bone repair. Therefore, the extracellular matrix secreted by the mesenchymal stem cells alone cannot mimic a complex bone regeneration microenvironment, and the addition of extracellular matrix by preosteoclasts may contribute as an effective strategy for bone regeneration. Here, we established the mesenchymal stem cell/preosteoclast extracellular matrix -based tissue-engineered bones and demonstrated that engineered-scaffolds based on mesenchymal stem cell/ preosteoclast extracellular matrix significantly enhanced osteogenesis in a 3 mm rat femur defect model compared with mesenchymal stem cell alone. The bioactive proteins released from the mesenchymal stem cell/ preosteoclast extracellular matrix based tissue-engineered bones also promoted the migration, adhesion, and osteogenic differentiation of mesenchymal stem cells in vitro. As for the mechanisms, the iTRAQ-labeled mass spectrometry was performed, and 608 differentially expressed proteins were found, including the IGFBP5 and CXCL12. Through in vitro studies, we proved that CXCL12 and IGFBP5 proteins, mainly released from the preosteoclasts, contributed to mesenchymal stem cells migration and osteogenic differentiation, respectively. Overall, our research, for the first time, introduce pre-osteoclast into the tissue engineering of bone and optimize the strategy of constructing extracellular matrix-based tissue-engineered bone using different cells to simulate the natural bone regeneration environment, which provides new sight for bone tissue engineering.

Osteoclast-derived exosomal let-7a-5p targets Smad2 to promote the hypertrophic differentiation of chondrocytes.

The invasion of osteoclasts into the cartilage via blood vessels advances the process of endochondral ossification, and dysregulation of dynamic intercellular interactions results in skeletal dysplasias. Although the regulation of osteoclasts by growth plate chondrocytes has been reported in detail, the effect of osteoclasts on chondrocytes remains to be determined. In this study, ATDC5 cells and bone marrow mesenchymal stem cells were differentiated into chondrocytes and treated with conditioned medium obtained from bone marrow macrophages differentiated to osteoclast precursors and osteoclasts. Exosomes were inhibited in conditioned medium or were isolated directly from osteoclasts to further determine whether osteoclast-derived exosomes play an important role in chondrocyte hypertrophy. Additionally, exosomal miRNAs were detected, and let-7a-5p was selected as an miRNA with significantly increased expression in osteoclast-derived exosomes. Experiments were performed to verify the potential target Smad2 and investigate how let-7a-5p affected chondrocytes. The results suggest that both osteoclast precursors and osteoclasts promote chondrocyte hypertrophy and that the promotive effect of osteoclasts is more significant than that of osteoclast precursors. Osteoclast-derived exosomes promote the hypertrophic differentiation of chondrocytes. Moreover, osteoclast-derived exosomal let-7a-5p inhibits Smad2 to decrease the transforming growth factor-ß-induced inhibition of chondrocyte hypertrophy. Our research reveals the role of osteoclasts in the regulation of chondrocytes and provides insights into the highly coordinated intercellular process of endochondral ossification.

Effect of low-dose zearalenone exposure on reproductive capacity of male mice.

Zearalenone (ZEA), a kind of nonsteroidal mycotoxin with estrogenic effects, can influence animal reproductive capacity through interfering with estrogen signaling pathway. Previous studies have shown exposure to ZEA at high doses (higher than No-Observed Effect Level, NOEL) had a significant impact on mouse sperm quality and pregnant rate, but little is known about the effect of exposure to ZEA at low doses (lower than NOEL) on mouse reproductive capacity. This study evaluated the effects of exposure to low-dose ZEA on mouse spermatogenesis and semen quality. Male mice (CD-1) of 21days were exposed to ZEA at 20, or 40µg/kg body weight for 14, 28 or 42days. After exposure to ZEA for 14days, the spermatogenic cells in seminiferous tubules were declined dose-independently; however in groups treated by ZEA for 28days, the spermatogenic cells were declined dose-dependently. Moreover, after treatment for 28days or 42days, the DNA double stand break (DSB) in spermatogenic cells were increased in a dose-dependent manner in treated groups. Compared with the control group, the sperm concentration, viability, motility, and hyperactive rate in treated groups were decreased dose-dependently and time-dependently. Meanwhile, deformity and mortality rate of sperm in treated groups were increased remarkably dose-dependently too. In conclusion, low dose ZEA impaired male reproductive capacity especially in spermatogenesis and semen quality of mouse.

LTRtype, an Efficient Tool to Characterize Structurally Complex LTR Retrotransposons and Nested Insertions on Genomes.

The amplification and recombination of long terminal repeat (LTR) retrotransposons have proven to determine the size, organization, function, and evolution of most host genomes, especially very large plant genomes. However, the limitation of tools for an efficient discovery of structural complexity of LTR retrotransposons and the nested insertions is a great challenge to confront ever-growing amount of genomic sequences for many organisms. Here we developed a novel software, called as LTRtype, to characterize different types of structurally complex LTR retrotransposon elements as well as nested events. This system is capable of rapidly scanning large-scale genomic sequences and appropriately characterizing the five complex types of LTR retrotransposon elements. After testing on the Arabidopsis thaliana genome, we found that this program is able to properly annotate a large number of structurally complex elements as well as the nested insertions. Thus, LTRtype can be employed as an automatic and efficient tool that will help to reconstruct the evolutionary history of LTR retrotransposons and better understand the evolution of host genomes. LTRtype is publicly available at: http://www.plantkingdomgdb.com/LTRtype/index.html.

Full transcription of the chloroplast genome in photosynthetic eukaryotes.

Prokaryotes possess a simple genome transcription system that is different from that of eukaryotes. In chloroplasts (plastids), it is believed that the prokaryotic gene transcription features govern genome transcription. However, the polycistronic operon transcription model cannot account for all the chloroplast genome (plastome) transcription products at whole-genome level, especially regarding various RNA isoforms. By systematically analyzing transcriptomes of plastids of algae and higher plants, and cyanobacteria, we find that the entire plastome is transcribed in photosynthetic green plants, and that this pattern originated from prokaryotic cyanobacteria - ancestor of the chloroplast genomes that diverged about 1 billion years ago. We propose a multiple arrangement transcription model that multiple transcription initiations and terminations combine haphazardly to accomplish the genome transcription followed by subsequent RNA processing events, which explains the full chloroplast genome transcription phenomenon and numerous functional and/or aberrant pre-RNAs. Our findings indicate a complex prokaryotic genome regulation when processing primary transcripts.

[Compound amino acid combined with vitamin E for idiopathic asthenospermia].

Objective: To study the therapeutic efficacy of compound amino acid combined with vitamin E in the treatment of idiopathic asthenospermia. Methods: This study included 120 cases of idiopathic asthenospermia treated in the Outpatient Department of our hospital between February 2014 and January 2015. We randomized the patients into a treatment group( n = 70,aged23- 43 [mean 32. 5] years) and a control group( n = 50,aged 23- 44 [mean 31. 7] years),the former treated with compound amino acid plus vitamin E while the latter with vitamin E only. After 90 days of medication, we evaluated the therapeutic effects by comparing the total sperm motility( progressive motility + non-progressive motility, PR + NP),the percentage of progressively motile sperm( PR),and the pregnancy rate between the two groups. Results: Before treatment, PR + NP and PR were(26. 24 ± 6. 56) %and(24. 65 ± 6. 43) % in the treatment group and(15. 13 ± 5. 68) % and(14. 73 ± 6. 16) in the control, with no statistically significant differences between the two groups( P > 0. 05). After 90 days of medication, PR + NP and PR were( 49. 63 ± 9. 78) % and(33. 33 ± 5. 64)% in the former and(37. 67 ± 7. 98)% and(27. 23 ± 6. 46)% in the latter, remarkably increased in both groups as compared with the baseline( P < 0. 05),but significantly more in the treatment than in the control group( P < 0. 05). Four pregnancies(5. 71%) were achieved in the former group but only 1(2. 00%) in the latter. The total rate of effectiveness was markedly higher in the treatment group than in the control(74. 28% vs 44. 00%,P < 0. 05). No adverse reactions were observed in either group. Conclusion: Compound amino acid combined with vitamin E can safely and effectively improve sperm motility in idiopathic asthenospermia patients.

The complete chloroplast genome sequence of American bird pepper (Capsicum annuum var. glabriusculum).

The complete chloroplast genome sequence of American bird pepper (Capsicum annuum var. glabriusculum) is reported and characterized in this study. The genome size is 156,612 bp, containing a pair of inverted repeats (IRs) of 25,776 bp separated by a large single-copy region of 87,213 bp and a small single-copy region of 17,851 bp. The chloroplast genome harbors 130 known genes, including 89 protein-coding genes, 8 ribosomal RNA genes, and 37 tRNA genes. A total of 18 of these genes are duplicated in the inverted repeat regions, 16 genes contain 1 intron, and 2 genes and one ycf have 2 introns.

Rapid diversification of five Oryza AA genomes associated with rice adaptation.

Comparative genomic analyses among closely related species can greatly enhance our understanding of plant gene and genome evolution. We report de novo-assembled AA-genome sequences for Oryza nivara, Oryza glaberrima, Oryza barthii, Oryza glumaepatula, and Oryza meridionalis. Our analyses reveal massive levels of genomic structural variation, including segmental duplication and rapid gene family turnover, with particularly high instability in defense-related genes. We show, on a genomic scale, how lineage-specific expansion or contraction of gene families has led to their morphological and reproductive diversification, thus enlightening the evolutionary process of speciation and adaptation. Despite strong purifying selective pressures on most Oryza genes, we documented a large number of positively selected genes, especially those genes involved in flower development, reproduction, and resistance-related processes. These diversifying genes are expected to have played key roles in adaptations to their ecological niches in Asia, South America, Africa and Australia. Extensive variation in noncoding RNA gene numbers, function enrichment, and rates of sequence divergence might also help account for the different genetic adaptations of these rice species. Collectively, these resources provide new opportunities for evolutionary genomics, numerous insights into recent speciation, a valuable database of functional variation for crop improvement, and tools for efficient conservation of wild rice germplasm.