Ubiquitin ligase VvPUB26 in grapevine promotes proanthocyanidin synthesis and resistance to powdery mildew.

Proanthocyanidins (PAs) are an important group of flavonoids that contribute to astringency, color, and flavor in grapes (Vitis vinifera) and wines. They also play a crucial role in enhancing plant resistance to various stresses. However, the underlying regulatory mechanism governing PAs biosynthesis, particularly in relation to conferring resistance to powdery mildew, has not been extensively explored. This study focused on identifying a key player in PAs biosynthesis, namely the plant U-box (PUB) E3 ubiquitin ligase VvPUB26. We discovered that overexpression of VvPUB26 in grapes leads to a significant increase in PAs content, whereas interfering with VvPUB26 has the opposite effect. Additionally, our findings demonstrated that overexpression of VvPUB26 in transgenic grapevines enhances defense against powdery mildew while interfering with VvPUB26 results in increased susceptibility to the pathogen. Interestingly, we observed that VvPUB26 interacts with the WRKY transcription factor VvWRKY24, thereby facilitating ubiquitination and degradation processes. Through RNA-Seq analysis, we found that VvWRKY24 primarily participates in secondary metabolites biosynthesis, metabolic pathways, and plant-pathogen interaction. Notably, VvWRKY24 directly interacts with the promoters of dihydroflavonol-4-reductase (DFR) and leucoanthocyanidin reductase (LAR) to inhibit PAs biosynthesis. Meanwhile, VvWRKY24 also influences the expression of MYB transcription factor genes related to PAs synthesis. In conclusion, our results unveil a regulatory module involving VvPUB26-VvWRKY24-VvDFR/VvLAR that plays a fundamental role in governing PAs biosynthesis in grapevines. These findings enhance our understanding of the relationship between PAs biosynthesis and defense mechanisms against powdery mildew.

The putative ABCG transporter VviABCG20 from grapevine (Vitis vinifera) is strongly expressed in the seed coat of developing seeds and may participate in suberin biosynthesis.

Half-size ATP binding cassette G (ABCG) transporters participate in many biological processes by transporting specific substrates. Our previous study showed that VviABCG20 was strongly expressed in the seeds of seeded grape and the silencing of VviABCG20 homolog gene in tomato led to a reduction in seed number. To reveal the molecular mechanism of VviABCG20 gene involved in grape seed development/abortion, the gene expression and functional analysis of VviABCG20 were further carried out in the grapevine. It was shown that the gene expression of VviABCG20 was higher in seeds of seeded grapes compared with seedless. Further the expression of VviABCG20 in the seed coat was significantly higher than in ovules (young seeds) and endosperm. VviABCG20 was also induced by exogenous hormones (especially MeJA) in grape leaves. Subcellular localization analysis showed that VviABCG20 is a membrane protein. In overexpressed VviABCG20 transgenic callus of Thompson seedless, expression of genes GPAT5, FAR1 and FAR5 was increased significantly. After treatment with suberin precursors, the transgenic callus reduced the sensitivity to three cinnamic acid derivatives (cis-ferulic acid, caffeic acid, coumaric acid), succinic acid, and glycerol. In suspension cells, expression of VviABCG20 was increased significantly after treatment with suberin precursors. Our research suggested that VviABCG20 may function in seed development in grapevine, at least in part by participating in suberin biosynthesis in the seed coat.

Metformin use and associated risk of total joint replacement in patients with type 2 diabetes: a population-based matched cohort study.

BACKGROUND: It is uncertain whether metformin use is associated with reduced risk of joint replacement in patients with type 2 diabetes mellitus. We aimed to establish whether metformin use was associated with a reduced risk of total knee replacement (TKR) or total hip replacement (THR) among these patients. METHODS: We selected patients with type 2 diabetes mellitus that was diagnosed between 2000 and 2012 from the Taiwan National Health Insurance Research Database. We used prescription time-distribution matching and propensity-score matching to balance potential confounders between metformin users and nonusers. We assessed the risks of TKR or THR using Cox proportional hazards regression. RESULTS: We included 20 347 participants who were not treated with metformin and 20 347 who were treated with metformin, for a total of 40 694 participants (mean age 63 yr, standard deviation 11 yr; 49.8% were women) after prescription time-distribution matching. Compared with participants who did not use metformin, those who used metformin had lower risks of TKR or THR (adjusted hazard ratio [HR] 0.70, 95% confidence interval [CI] 0.60-0.81 for TKR or THR; adjusted HR 0.71, 95% CI 0.61-0.84 for TKR; adjusted HR 0.61, 95% CI 0.41-0.92 for THR) after adjustment for covariates. Propensity-score matching analyses (10 163 participants not treated with metformin v. 10 163 treated with metformin) and sensitivity analyses using inverse probability of treatment weighting and competing risk regression showed similar results. INTERPRETATION: Metformin use in patients with type 2 diabetes mellitus was associated with a significantly reduced risk of total joint replacement. Randomized controlled clinical trials in patients with osteoarthritis are warranted to determine whether metformin is effective in decreasing the need for joint replacement.

HA15 alleviates bone loss in ovariectomy-induced osteoporosis by targeting HSPA5.

The imbalance between osteogenesis and adipogenesis in the bone marrow is the main characteristic of osteoporosis (OP). Thus, exploring regulation of the differentiation of bone marrow stromal cells (BMSCs) into osteoblasts and adipocytes is important to identify novel targets for the treatment of OP. In the present study, the master regulator of endoplasmic reticulum (ER) stress, heat shock protein family A (Hsp70) member 5 (HSPA5) was shown to significantly accumulate in osteoblasts and adipocytes, but not in osteoclasts in bone sections from aged and postmenopausal OP mice. In vitro study revealed that HSPA5 negatively modulated osteogenic differentiation and positively promoted adipogenic differentiation, and that targeting HSPA5 with its inhibitor HA15 enhanced osteogenic differentiation and inhibited adipogenic differentiation. Also, HA15 treatment induces ER stress and autophagy, and decreases apoptosis in cells. We constructed a postmenopausal OP model in mice with ovariectomy surgery, and treated the mice with HA15. The results showed that HA15 treatment induced appropriate ER stress, activated autophagy and decreased apoptosis in osteoblasts, thereby alleviating bone loss in vivo. Our results indicated that HSPA5 participated in OP pathogenesis by regulating the differentiation of BMSCs. HSPA5 may serve as a new target for the treatment of OP, and targeting HSPA5 with HA15 prevents the progression of OP and provides a candidate therapeutic molecule for postmenopausal OP.

Notoginsenoside R1 counteracts mesenchymal stem cell-evoked oncogenesis and doxorubicin resistance in osteosarcoma cells by blocking IL-6 secretion-induced JAK2/STAT3 signaling.

Tumor microenvironment is a critical participant in the initiation, progression and drug resistance of carcinomas, including osteosarcoma. Notoginsenoside R1 (NGR1) is a proverbial active ingredient of the traditional Chinese medicine Panax notoginseng (PN) and possess undeniable roles in several cancers. Nevertheless, its function in osteosarcoma and tumor microenvironment remains elusive. In the current study, exposure to NGR1 dose-dependently inhibited osteosarcoma cell viability and migration, and induced apoptosis. Furthermore, osteosarcoma cells that were incubated with conditioned medium (CM) from bone marrow mesenchymal stem cells (BMSCs) exhibited greater proliferation, migration capacity and MMP-2 and MMP-9 expression relative to control cells, which was reversed when BMSCs were treated with NGR1. Notably, administration with NGR1 antagonized CM-evoked doxorubicin resistance in osteosarcoma cells by decreasing cell viability and increasing cell apoptosis and caspase-3/9 activity. Mechanically, NGR1 suppressed IL-6 secretion from BMSCs, as well as the subsequent activation of the JAK2/STAT3 signaling in osteosarcoma cells. In addition, blocking the JAK2 pathway by its antagonist AG490 reversed CM-induced osteosarcoma cell proliferation, migration and doxorubicin resistance. Moreover, exogenous supplementation with IL-6 engendered not only the reactivation of the JAK2/STAT3 signaling but also muted NGR1-mediated efficacy against osteosarcoma cell malignancy and doxorubicin resistance. Collectively, NGR1 may directly restrain osteosarcoma cell growth and migration, or indirectly antagonize MSC-evoked malignancy and drug resistance by interdicting IL-6 secretion-evoked activation of the JAK2/STAT3 pathway. Consequently, the current study may highlight a promising therapeutic strategy against osteosarcoma by regulating tumor cells and the tumor microenvironment.

Genome-wide identification, phylogenetic analysis, and expression profiling of glycine-rich RNA-binding protein (GRPs) genes in seeded and seedless grapes (Vitis vinifera).

Glycine-rich RNA-binding proteins (GRPs) are essential for many physiological and biochemical processes in plants, especially the response to environmental stresses. GRPs exist widely in angiosperms and gymnosperms plant species; however, their roles in Vitis vinifera are still poorly understood. To characterize VviGRP gene family, we performed a genomic survey, bioinformatics and expression analysis of VviGRPs in grape. We identified nineteen VviGRPs gene family members. The result of bioinformatics analysis showed their motif distribution, gene structure characteristics and chromosomal locations. Then we carried out synteny and phylogenetic analysis to study the origin and evolutionary relationship of GRPs. Tissue-specific expression analysis showed that VviGRPs have different expression patterns. Meanwhile, we studied expression profiles of seventeen ovule-expressed genes during seed development of stenospermocarpic seedless and seeded grapes, and the result showed that most of them have much higher relative expression levels in stenospermocarpic seedless grapes than that of seeded one before 25 days after full bloom (DAFB). It is suggested that VviGRPs may involve in the seed development process. Taken together, our research indicated that VviGRPs are related to seed development and will be beneficial for further investigations into the seed abortion mechanism under stenospermocarpic grapes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01082-3.

Fibronectin 1 activates WNT/ß-catenin signaling to induce osteogenic differentiation via integrin ß1 interaction.

Osteoporosis (OP) is a systemic skeletal disease leading to fragility fractures and is a major health issue globally. WNT/ß-catenin signaling regulates bone-remodeling processes and plays vital roles in OP development. However, the underlying regulatory mechanisms behind WNT/ß-catenin signaling in OP requires clarification, as further studies are required to identify novel alternate therapeutic agents to improve OP. Here we report that fibronectin 1 (FN-1) promoted differentiation and mineralization of osteoblasts by activating WNT/ß-catenin pathway, in cultured pre-osteoblasts. With isobaric tags for relative and absolute quantitation labeling proteomics analysis, we investigated protein changes in bone samples from OP patients and normal controls. FN-1 accumulated in osteoblasts in bone samples from OP patients and age-related OP mice compared to control group. In addition, we observed that integrin ß1 (ITGB1) acts as an indispensable signaling molecule for the interplay between FN-1 and ß-catenin, and that FN-1 expression increased, but ITGB1 expression decreased in osteoblasts during OP progression. Therefore, our study reveals a novel explanation for WNT/ß-catenin pathway inactivation in OP pathology. Supplying of FN-1 and ITGB1 may provide a potential therapeutic strategy in improving bone formation during OP.

COVID-19: asymptomatic carrier transmission is an underestimated problem.

At the present time, COVID-19 is spreading rapidly [1]. The global prevention and control of COVID-19 is focused on the estimation of the relevant incubation period, basic reproduction number (R0), effective reproduction number (Rt) and death risk. Although the prevention and control of COVID-19 requires a reliable estimation of the relevant incubation period, R0, Rt and death risk. Another key epidemiological parameter-asymptomatic ratio that provides strength and range for social alienation strategies of COVID-19, which is widely defined as the proportion of asymptomatic infections among all disease infections. In fact, the ratio of asymptomatic infection is a useful indicator of the burden of disease and a better measurement of the transmissibility of the virus. So far, people have not paid enough attention to asymptomatic carriers. The asymptomatic carriers discussed in this study are recessive infections, that is, those who have never shown symptoms after onset of infection. We will discuss three aspects: detection, infectivity and proportion of healthy carriers.

Molecular cloning and functional characterization of a seed-specific VvßVPE gene promoter from Vitis vinifera.

MAIN CONCLUSION: The grapevine VvßVPE promoter is specifically expressed in the seed. The - 1306~- 1045 bp core region restricts expression in other tissues and organs. Vacuolar processing enzyme (VPE) is a cysteine proteinase regulating vacuolar protein maturation and executing programmed cell death (PCD) in plants. Vitis vinifera (Vv)ßVPE is a ß-type VPE showing seed-specific expression that processes seed proteins during ovule development. However, the regulation of the seed-specific gene expression is far from understood. In this study, we characterize VvßVPE promoter (pVvßVPE) from 12 seeded and seedless grape genotypes. 94.56% of the pVvßVPE coding sequence is consistent. Two ßVPE promoters were constructed and transformed into Arabidopsis thaliana via ß-glucuronidase (GUS) fused expression vectors, using cv. Pinot Noir and cv. Thompson as seed and seedless candidates. GUS staining in different tissues and organs revealed that VvßVPE expresses specifically in the embryo, including the cotyledon, hypocotyl and suspensor, but not in the leaf, stem, root or flowers of the seedling. Using promoter deletion analysis, we created four incomplete VvßVPE promoters and found each pVvßVPE deletion could drive GUS gene to express in seeds. Interestingly, seed specificity disappeared when the promoter missed the core - 1306~- 1045 bp region. All deletion promoters presenting various quantified GUS activities indicate that the region - 1704~- 1306 bp inhibits, and the region - 705~- 861 bp promotes gene expression of VvßVPE. Our results demonstrate that pVvßVPE is a seed-specific promoter in both seeded and seedless grapes. Moreover, the core region of pVvßVPE (- 1306~- 1045 bp) is the key one responsible for seed-specific expression.

Association between functional polymorphisms in IL-33/ST2 pathway and risk of osteosarcoma.

Interleukin (IL)-33/ST2 pathway plays crucial roles in tumour growth and metastasis. The aim of this study was to investigate the association of two functional polymorphisms (IL-33 rs7025417 and ST2 rs3821204) with osteosarcoma (OS) risk. The rs7025417 and rs3821204 were genotyped by Taqman assay. IL-33mRNA and protein levels were measured by real-time PCR or enzyme-linked immunosorbent assay. The luciferase activity was measured by a dual luciferase reporter gene assay. The allele-specific transcription factor binding for rs7025417 was examined by ChIP-seq. The IL-33 rs7025417 CC genotype was significantly associated with a decreased risk of OS (CC vs TT: OR = 0.59, 95% CI, 0.41-0.85; recessive model: OR = 0.68, 95% CI, 0.49-0.94; C vs T: OR = 0.76, 95% CI, 0.63-0.91). Combined analysis showed that the IL-33 rs7025417CT/CC-ST2 rs3821204CG/CC and the IL-33 rs7025417CT/CC-ST2 rs3821204GG genotypes also had a decreased risk of OS. IL-33mRNA and protein levels in OS patients were significantly higher than controls. Patients with the rs7025417 CC genotype exhibited lower levels of IL-33 (P = .03). The rs7025417 C allele presented a lower transcriptional activity by disrupting the binding site to c-Myb (P < .01). Moreover, the rs3821204 G/C influences the transcriptional activity and ST2mRNA expression by altering the binding site of miR-202-3p. These findings suggest that the rs7025417 and rs3821204 may have a combined effect to protect against the development of OS by decreasing the expression levels of IL-33 or ST2.

Polymorphisms in the precursor microRNAs and aflatoxin B1-related hepatocellular carcinoma.

The altered expression of some microRNAs (miRNAs) is observed in hepatocellular carcinoma (HCC); however, the genetic polymorphisms in the precursor miRNAs (pre-miRNAs) in aflatoxin B1 (AFB1)-related HCC have not yet been investigated. A hospital-based case-control study, including 1,706 HCC cases and 2,270 controls without any liver diseases or tumors, was conducted in a high AFB1 exposure area of China to assess the relationship between 48 polymorphisms in the pre-miRNAs and AFB1-related HCC risk and prognosis. Among 48 polymorphisms, only rs28599926 (in the miRNA 1268a) affected HCC risk. Compared with the homozygote of rs28599926C alleles (rs28599926-CC), the genotypes of rs28599926 T alleles (namely rs28599926-CT or -TT) increased HCC risk (odds ratio [OR]: 1.63 and 5.52, 95% confidence interval [CI]: 1.40-1.90 and 4.27-7.14, respectively). Significant interactive effects between risk genotypes and AFB1 exposure status were also observed in the joint effects analysis. This polymorphism was associated not only with larger tumor size, higher portal vein tumor risk, and tumor dedifferentiation, but also with higher AFB1 adducts levels and increasing the mutation risk of TP53 gene. Furthermore, rs28599926 modified the tumor recurrence-free survival (hazard ratio [HR]: 2.86, 95% CI: 2.36-3.43) and overall survival (HR: 2.12, 95% CI: 1.86-2.41) of cases. Additionally, one target of miR-1268a was show to be the ADAMTS4 mRNA and rs28599926 polymorphism might modify ADAMTS4 expression. These findings indicate that polymorphisms in the pre-miRNAs may be risk and prognostic biomarkers of AFB1-related HCC, and rs28599926 in miR-1268a is such a potential candidate. © 2015 Wiley Periodicals, Inc.

Association of interleukin-8 gene polymorphisms with the risk of hepatocellular carcinoma.

Interleukin-8 (IL8) polymorphisms have been implicated in several cancers, but their roles in the pathogenesis of hepatocellular carcinoma (HCC) are largely unknown. The present study was designed to explore the association between IL8 polymorphism and the risk of HCC in a Chinese population. Four single nucleotide polymorphisms (SNPs) of the IL8 gene -251A/T, +781C/T, -353A/T and +678T/C were analyzed in 205 HCC patients and 208 healthy controls in a Chinese population. Serum levels of IL8 were detected in HCC patients and healthy controls. The association between IL8 polymorphisms and HCC risk was measured using the adjusted odds ratios (OR) and their 95% confidence intervals (CI) from multiple logistic regression analysis. Haplotype analysis and gene-environment interaction analysis was also performed. The serum level of IL8 was significantly higher in HCC patients compared with healthy controls (P < 0.001). After adjusting for confounding factors, no significant associations were found between -251A/T, +781C/T, -353A/T and +678T/C and HCC risk (all P > 0.05). Haplotype analysis showed that A(251)-C(781)-A(353)-C(678) conferred decreased risk of HCC onset (adjusted OR 0.31, 95% CI 0.13-0.77). No significant interaction effects were found between the four SNPs and HBV infection, cirrhosis, gender smoking and alcohol consumption (all P > 0.05). No association between -251A/T, +781C/T, -353A/T and +678T/C of the IL8 gene and the risk of HCC was found in this Chinese population, and the SNPs did not display any interaction with several environmental factors with regard to HCC risk. However, it appears that A(251)-C(781)-A(353)-C(678) is perhaps a protective haplotype for HCC.

Practice effects of personalized interventions with interdisciplinary teamwork in type 2 diabetes remission: a retrospective study.

Objectives: A retrospective analysis of the clinical outcomes of personalized interventions for type 2 diabetes mellitus (T2DM) in an interdisciplinary team. Methods: Under the guidance of an interdisciplinary team, 40 patients with T2DM underwent a systematic examination at the beginning of the intervention, 3 months after the intervention, and 3 months of follow-up at the end of the intervention (i.e., at 6 months). Key indicators such as fasting plasma glucose (FPG), 2-hour postprandial glucose (2hPG), fasting insulin level (FINS), glycated hemoglobin (HbA1c), blood lipids, and body mass index (BMI) were measured. Results: After the 3-month intervention, participants' BMI, FPG, 2hPG, FINS, and HbA1c improved significantly, with statistically significant differences (P<0.05).These metrics remained essentially stable at the 3-month follow-up. Of all the participants, 92.5% (37 cases in total) successfully discontinued their medication after 3 months of intervention, of which 80% (32 cases) remained stable during the 3-month follow-up after discontinuation, fulfilling the criteria for remission of T2DM; 2 cases successfully reduced the dose of their medication, and only 1 case was maintained on the original treatment. Conclusions: Through an interdisciplinary team intervention strategy, we significantly optimized the glucose metabolism, lipid metabolism, and BMI status of patients with T2DM, making diabetes remission an achievable goal, which provides valuable experience for further optimization of diabetes prevention and control protocols.

Effect of cytotoxic T-lymphocyte antigen-4, TNF-alpha polymorphisms on osteosarcoma: evidences from a meta-analysis.

OBJECTIVE: Previous studies have investigated the role of cytotoxic T-lymphocyte antigen-4 (CTLA-4) and tumor necrosis factor-alpha (TNF-α) in carcinogenesis of osteosarcoma, but their results were inconsistent. We aimed to clarify the associations between CTLA-4, TNF-α polymorphism and osteosarcoma risk by using meta-analysis. METHODS: We searched relevant studies without language restriction in PubMed, EMbase, Cochrane Library, Google Scholar databases, Chinese National Knowledge Infrastructure (CNKI) and conference literature in humans published prior to March 2013. The strengths of the associations between genetic variants and osteosarcoma risk were estimated by odds ratio (OR) with 95% confidence interval (95% CI). RESULTS: A total of seven studies with 1,198 osteosarcoma patients and 1,493 controls were selected. Four studies were eligible for CTLA-4 (1,003 osteosarcoma and 1,162 controls), and three studies for TNF-α (195 osteosarcoma and 331 controls). Pooled results showed that rs231775 polymorphism of CTLA-4 was associated with osteosarcoma risk (GG vs. AA: OR=1.63, 95% CI=1.24-2.13; GG + GA vs. AA: OR=1.56, 95% CI=1.21-2.01; AA + GA vs. GG: OR=0.83, 95% CI=0.71-0.97; G vs. A: OR=1.21, 95% CI=1.08-1.36). No significant heterogeneity was observed across the studies. No significant associations were found between rs5742909 polymorphism of CTLA-4 or rs1800629 polymorphism of TNF-α and osteosarcoma risk. CONCLUSIONS: These results suggest that the rs231775 polymorphism of CTLA-4 may play an important role in carcinogenesis of osteosarcoma.

Framework Nucleic Acid-Based Multifunctional Tumor Theranostic Nanosystem for miRNA Fluorescence Imaging and Chemo/Gene Therapy.

Intelligent stimulus-responsive theranostic systems capable of specifically sensing low-abundance tumor-related biomarkers and efficiently killing tumors remain a pressing endeavor. Here, we report a multifunctional framework nucleic acid (FNA) nanosystem for simultaneous imaging of microRNA-21 (miR-21) and combined chemo/gene therapy. To achieve this, two FNA nanoarchitectures labeled with Cy5/BHQ2 signal tags were designed, each of which contained an AS1411 aptamer, two pairs of DNA/RNA hybrids, a pH-sensitive DNA catcher, and doxorubicin (DOX) intercalating between cytosine and guanine in the tetrahedral DNA nanostructure (TDN). In the acidic tumor microenvironment, the DNA catchers spontaneously triggered to form an i-motif and create an FNA dimer (dFNA) while releasing DOX molecules to exert a cytotoxic effect. In addition, the overexpressed miR-21 in tumor cells dismantled the DNA/RNA hybrids to produce vascular endothelial growth factor-associated siRNA via a toehold-mediated strand displacement reaction, thus enabling a potent RNA interfering. Also importantly, the liberated miR-21 could initiate cascade-reaction amplification to efficiently activate the Cy5 signal reporters, thereby realizing on-site fluorescence imaging of miR-21 in living cells. The exquisitely designed FNA-based nanosystem showed favorable biocompatibility and stability as well as acid-driven DOX release characteristics. Owing to the aptamer-guided targeting delivery, specific uptake of the FNA-based theranostic nanosystem by HepG2 cells was verified with confocal laser scanning microscopy and flow cytometry analyses, which therefore resulted in apoptosis of HepG2 cells while doing minimal damage to normal H9c2 and HL-7702 cells. Strikingly, both in vitro and in vivo experiments demonstrated the achievements of the FNA-enabled miR-21 imaging and synergistically enhanced chemo/gene therapy. This work thus represents a noteworthy advance on the FNA-based theranostic strategy that can effectively avoid the undesirable premature leakage of anticarcinogen and off-target of siRNA, and achieve on-demand reagents release for tumor diagnostics and treatment.

VviKFB07 F-box E3 ubiquitin ligase promotes stilbene accumulation by ubiquitinating and degrading VviCHSs protein in grape.

Stilbene and flavonoid are phytochemicals in plants and play an important role in plant disease resistance and human health. The regulation of stilbene and flavonoid synthesis in plants has been extensively studied at the transcriptional level, but translational and post-translational controls of stilbene and flavonoid biosynthesis are still poorly understood. In this study, a grape F-box E3 ubiquitin ligase VviKFB07 associated with the metabolism of stilbene and flavonoid was screened out with transcriptome. Overexpression of VviKFB07 in the Nicotiana tabacum resulted in a decrease in flavonol and anthocyanin content in corolla, and stable overexpression assays of VviKFB07 in grape callus promoted the accumulation of resveratrol. Subsequently, Yeast two-hybrid and bimolecular fluorescence complementation assays identified the physical interaction between VviKFB07 and VviCHSs proteins. In vivo experiments verified that VviKFB07 was involved in the ubiquitination and degradation of VviCHSs protein. Taken together, our findings clarify the role of ubiquitin ligase VviKFB07 in the synthesis of stilbene and flavonoid in grapes.

MYL3 protects chondrocytes from senescence by inhibiting clathrin-mediated endocytosis and activating of Notch signaling.

As the unique cell type in articular cartilage, chondrocyte senescence is a crucial cellular event contributing to osteoarthritis development. Here we show that clathrin-mediated endocytosis and activation of Notch signaling promotes chondrocyte senescence and osteoarthritis development, which is negatively regulated by myosin light chain 3. Myosin light chain 3 (MYL3) protein levels decline sharply in senescent chondrocytes of cartilages from model mice and osteoarthritis (OA) patients. Conditional deletion of Myl3 in chondrocytes significantly promoted, whereas intra-articular injection of adeno-associated virus overexpressing MYL3 delayed, OA progression in male mice. MYL3 deficiency led to enhanced clathrin-mediated endocytosis by promoting the interaction between myosin VI and clathrin, further inducing the internalization of Notch and resulting in activation of Notch signaling in chondrocytes. Pharmacologic blockade of clathrin-mediated endocytosis-Notch signaling prevented MYL3 loss-induced chondrocyte senescence and alleviated OA progression in male mice. Our results establish a previously unknown mechanism essential for cellular senescence and provide a potential therapeutic direction for OA.

Retraction: Acellularized spinal cord scaffolds incorporating bpV(pic)/PLGA microspheres promote axonal regeneration and functional recovery after spinal cord injury.

[This retracts the article DOI: 10.1039/D0RA02661A.].

Error assessment and correction for extrusion-based bioprinting using computer vision method.

299Bioprinting offers a new approach to addressing the organ shortage crisis. Despite recent technological advances, insufficient printing resolution continues to be one of the reasons that impede the development of bioprinting. Normally, machine axes movement cannot be reliably used to predict material placement, and the printing path tends to deviate from the predetermined designed reference trajectory in varying degrees. Therefore, a computer vision-based method was proposed in this study to correct trajectory deviation and improve printing accuracy. The image algorithm calculated the deviation between the printed trajectory and the reference trajectory to generate an error vector. Furthermore, the axes trajectory was modified according to the normal vector approach in the second printing to compensate for the deviation error. The highest correction efficiency that could be achieved was 91%. More significantly, we discovered that the correction results, for the first time, were in a normal distribution instead of a random distribution.

Double crosslinked biomimetic composite hydrogels containing topographical cues and WAY-316606 induce neural tissue regeneration and functional recovery after spinal cord injury.

Spinal cord injury (SCI) is an overwhelming and incurable disabling condition, for which increasing forms of multifunctional biomaterials are being tested, but with limited progression. The promising material should be able to fill SCI-induced cavities and direct the growth of new neurons, with effective drug loading to improve the local micro-organism environment and promote neural tissue regeneration. In this study, a double crosslinked biomimetic composite hydrogel comprised of acellularized spinal cord matrix (ASCM) and gelatin-acrylated-ß-cyclodextrin-polyethene glycol diacrylate (designated G-CD-PEGDA) hydrogel, loaded with WAY-316606 to activate canonical Wnt/ß-catenin signaling, and reinforced by a bundle of three-dimensionally printed aligned polycaprolactone (PCL) microfibers, was constructed. The G-CD-PEGDA component endowed the composite hydrogel with a dynamic structure with a self-healing capability which enabled cell migration, while the ASCM component promoted neural cell affinity and proliferation. The diffusion of WAY-316606 could recruit endogenous neural stem cells and improve neuronal differentiation. The aligned PCL microfibers guided neurite elongation in the longitudinal direction. Animal behavior studies further showed that the composite hydrogel could significantly recover the motor function of rats after SCI. This study provides a proficient approach to produce a multifunctional system with desirable physiological, chemical, and topographical cues for treating patients with SCI.