Dynamic m6A mRNA methylation reveals the involvement of AcALKBH10 in ripening-related quality regulation in kiwifruit.

Kiwifruit ripening is a complex and highly coordinated process that occurs in conjunction with the formation of fruit edible quality. The significance of epigenetic changes, particularly the impact of N6-methyladenosine (m6A) RNA modification on fruit ripening and quality formation, has been largely overlooked. We monitored m6A levels and gene expression changes in kiwifruit at four different stages using LC-MS/MS, MeRIP, RNA-seq, and validated the function of AcALKBH10 through heterologous transgenic expression in tomato. Notable m6A modifications occurred predominantly at the stop codons and the 3' UTRs and exhibited a gradual reduction in m6A levels during the fruit ripening process. Moreover, these m6A modifications in the aforementioned sites demonstrated a discernible inverse relationship with the levels of mRNA abundance throughout the ripening process, suggesting a repression effect of m6A modification in the modulation of kiwifruit ripening. We further demonstrated that AcALKBH10 rather than AcECT9 predominantly regulates m6A levels in ripening-related genes, thereby exerting the regulatory control over the ripening process and the accumulation of soluble sugars and organic acids, ultimately influencing fruit ripening and quality formation. In conclusion, our findings illuminate the epi-regulatory mechanism involving m6A in kiwifruit ripening, offering a fresh perspective for cultivating high-quality kiwifruit with enhanced nutritional attributes.

Diagnostic Role of Metagenomic Next-Generation Sequencing in Tubercular Orthopedic Implant-Associated Infection.

Objective: The diagnosis of tubercular orthopedic implant-associated infection (TB-IAI) is challenging. This study evaluated the value of metagenomic next-generation sequencing (mNGS) for the diagnosis of TB-IAI and developed a standardized diagnostic procedure for TB-IAI. Methods: The records of all patients with TB-IAI diagnosed and treated at our institution between December 2018 and September 2022 were retrospectively reviewed. Patient demographic characteristics, medical history, laboratory test, microbial culture, histopathology, and mNGS results, and time to diagnosis were recorded. The diagnostic efficiency of mNGS for TB-IAI was assessed by comparing the results and diagnostic time with that of other diagnostic modalities. Results: Ten patients were included in the analysis, including eight with prosthetic joint infections and two with fracture-related infections. The mNGS positivity rate was 100% (10/10), which was higher than that of TB-antibody (11%, 1/9), real-time quantitative polymerase chain reaction (22%, 2/9), T-SPOT.TB (25%, 2/8), purified protein derivative (50%, 4/8), microbial culture (50%, 5/10), and histopathology (20%, 2/10). mNGS shortened the time to diagnosis of TB-IAI. A standardized diagnostic procedure for TB-IAI was developed based on the findings. Conclusion: mNGS is useful for the diagnosis of TB-IAI. mNGS is recommended in cases where it is difficult to identify a pathogen using routine diagnostic tests. The standardized diagnostic procedure might improve TB-IAI diagnosis. Importance: TB-IAI is a rare infection, which occurs after orthopedic surgery and hard to diagnose microbiologically. mNGS is a new detection technique not yet discussed in current literature as a means for TB-IAI diagnostics. Here we describe a cohort of patients with TB-IAI diagnosed by mNGS show high efficiency of mNGS for detection of this pathology and present a clinical algorithm supplementing conventional methods for TB-IAI assessment.

Methylation of AcGST1 Is Associated with Anthocyanin Accumulation in the Kiwifruit Outer Pericarp.

Most red-fleshed kiwifruit cultivars, such as Hongyang, only accumulate anthocyanins in the inner pericarp; the trait of full red flesh becomes the goal pursued by breeders. In this study, we identified a mutant "H-16" showing a red color in both the inner and outer pericarps, and the underlying mechanism was explored. Through transcriptome analysis, a key differentially expressed gene AcGST1 was screened out, which was positively correlated with anthocyanin accumulation in the outer pericarp. The result of McrBC-PCR and bisulfite sequencing revealed that the SG3 region (-292 to -597 bp) of AcGST1 promoter in "H-16" had a significantly lower CHH cytosine methylation level than that in Hongyang, accompanied by low expression of methyltransferase genes (MET1 and CMT2) and high expression of demethylase genes (ROS1 and DML1). Transient calli transformation confirmed that demethylase gene DML1 can activate transcription of AcGST1 to enhance its expression. Overexpression of AcGST1 enhanced the anthocyanin accumulation in the fruit flesh and leaves of the transgenic lines. These results suggested that a decrease in the methylation level of the AcGST1 promoter may contribute to accumulation of anthocyanin in the outer pericarp of "H-16".

ITGB1 alleviates osteoarthritis by inhibiting cartilage inflammation and apoptosis via activating cAMP pathway.

OBJECTIVE: We aimed to screen novel biomarkers for osteoarthritis (OA) using bioinformatic methods and explore its regulatory mechanism in OA development. METHODS: Differentially expressed genes were screened out from GSE98918 and GSE82107 datasets. Protein-protein interaction network and enrichment analysis were employed to search for hub gene and regulatory pathway. Hematoxylin-eosin, Safranin O-Fast green staining, and immunohistochemistry were performed to assess pathological damage. TNF-α, IL-1ß, and IL-6 concentrations were determined by enzyme-linked immunosorbent assay. Real-time quantitative PCR was applied to verify expression of hub genes in OA model. The expression of key protein and pathway proteins was determined by western blot. Furthermore, Cell Counting Kit-8 and flow cytometry were conducted to explore the role of hub gene in chondrocytes. RESULTS: We identified 6 hub genes of OA, including ITGB1, COL5A1, COL1A1, THBS2, LAMA1, and COL12A1, with high prediction value. ITGB1 was screened as a pivotal regulator of OA and cAMP pathway was selected as the key regulatory pathway. ITGB1 was down-regulated in OA model. ITGB1 overexpression attenuated pathological damage and apoptosis in OA rats with the reduced levels of TNF-α, IL-1ß and IL-6. ITGB1 overexpression activated cAMP pathway in vivo and vitro models. In vitro model, ITGB1 overexpression promoted cell viability, while inhibited apoptosis. ITGB1 overexpression also caused a decrease of TNF-α, IL-1ß, and IL-6 concentrations. cAMP pathway inhibitor reversed the positive effect of ITGB1 on OA cell model. CONCLUSION: ITGB1 is a novel biomarker for OA, which inhibits OA development by activating the cAMP pathway.

Interactions between Macrophages and Biofilm during Staphylococcus aureus-Associated Implant Infection: Difficulties and Solutions.

Staphylococcus aureus (S. aureus) biofilm is the major cause of failure of implant infection treatment that results in heavy social and economic burden on individuals, families, and communities. Planktonic S. aureus attaches to medical implant surfaces where it proliferates and is wrapped by extracellular polymeric substances, forming a solid and complex biofilm. This provides a stable environment for bacterial growth, infection maintenance, and diffusion and protects the bacteria from antimicrobial agents and the immune system of the host. Macrophages are an important component of the innate immune system and resist pathogen invasion and infection through phagocytosis, antigen presentation, and cytokine secretion. The persistence, spread, or clearance of infection is determined by interplay between macrophages and S. aureus in the implant infection microenvironment. In this review, we discuss the interactions between S. aureus biofilm and macrophages, including the effects of biofilm-related bacteria on the macrophage immune response, roles of myeloid-derived suppressor cells during biofilm infection, regulation of immune cell metabolic patterns by the biofilm environment, and immune evasion strategies adopted by the biofilm against macrophages. Finally, we summarize the current methods that support macrophage-mediated removal of biofilms and emphasize the importance of considering multi-dimensions and factors related to implant-associated infection such as immunity, metabolism, the host, and the pathogen when developing new treatments.

Multicenter analysis of gastrointestinal stromal tumors in inner Mongolia of China: A study of 804 cases.

INTRODUCTION: To analyze the clinicopathological characteristics, immunohistochemistry, genotyping and prognosis of patients in the multicenter GIST data in Inner Mongolia, China. METHODS: Retrospective analysis was performed on GIST data from January 2013 to January 2018 in Inner Mongolia. Descriptive statistics were used to analyze the clinical characteristics of GIST patients. The Chi-square test was performed on the modified NIH criteria by age distribution, and Kaplan-Merie method was used for survival analysis. RESULTS: A total of 804 patients were included in the GIST database in Inner Mongolia, with a male to female ratio of 1.1102:1. The most common location was the gastric (465). Mitotic count ≤5/50HPFs was found in 67.3 % patients. There were 276 patients with tumor diameter of 2-5 cm and 354 patients with tumor diameter of 5.1-10 cm.The modified NIH criteria was mainly of intermediate risk (210) and high risk (342). The recurrence and metastasis of patients were related to the tumor location, mitotic index, tumor size, and modified NIH criteria. All patients were followed up for 1-10 years, in which 63.1 % of them were followed up for at least three years. The 3-year survival rates of patients with modified NIH criteria of very low risk, low risk, intermediate risk, and high risk were 100 %, 100 %, 100 %, and 96.3 %, respectively. CONCLUSIONS: The incidence of GIST in middle-aged and elder people in Inner Mongolia is high, and the long-term prognosis of patients after surgical treatment is good, which can objectively reflect the incidence, diagnosis and treatment of GIST in Inner Mongolia.

Ethylene response factor AcERF91 affects ascorbate metabolism via regulation of GDP-galactose phosphorylase encoding gene (AcGGP3) in kiwifruit.

Kiwifruit is known as 'the king of vitamin C' because of the high content of ascorbic acid (AsA) in the fruit. Deciphering the regulatory network and identification of the key regulators mediating AsA biosynthesis is vital for fruit nutrition and quality improvement. To date, however, the key transcription factors regulating AsA metabolism during kiwifruit developmental and ripening processes remains largely unknown. Here, we generated a putative transcriptional regulatory network mediating ascorbate metabolism by transcriptome co-expression analysis. Further studies identified an ethylene response factor AcERF91 from this regulatory network, which is highly co-expressed with a GDP-galactose phosphorylase encoding gene (AcGGP3) during fruit developmental and ripening processes. Through dual-luciferase reporter and yeast one-hybrid assays, it was shown that AcERF91 is able to bind and directly activate the activity of the AcGGP3 promoter. Furthermore, transient expression of AcERF91 in kiwifruit fruits resulted in a significant increase in AsA content and AcGGP3 transcript level, indicating a positive role of AcERF91 in controlling AsA accumulation via regulation of the expression of AcGGP3. Overall, our results provide a new insight into the regulation of AsA metabolism in kiwifruit.

SIRT3-mediated mitochondrial unfolded protein response weakens breast cancer sensitivity to cisplatin.

BACKGROUND: Mitochondrial unfolded protein response plays an important role in the occurrence and development of breast cancer. However, the role of mitochondrial unfolded protein response (UPRmt) in the sensitivity of breast cancer to cisplatin chemotherapy has not yet been cleared. OBJECTIVES: The purpose of this study is to explore the role of mitochondrial unfolded protein response in breast cancer sensitivity to cisplatin. METHODS: In this study, qRT-PCR, Western blotting, Immunofluorescence, CCK-8, Colony formation, Transwell assay and TUNEL staining assay were used to confirm the role of UPRmt in breast cancer cells treated with cisplatin. RESULTS: Cisplatin increased the levels of UPRmt including CLPP, HSP60, LONP1 in MCF7 and MDA-MB-231 cells. UPRmt inducer Nicotinamide ribose (NR) could promote the proliferation and invasion of breast cancer cells treated with cisplatin. Importantly, SIRT3 was discovered to increase UPRmt in breast cancer cells and silencing of SIRT3 could inhibit the effect of NR in breast cancer. CONCLUSIONS: UPRmt regulated by SIRT3 could protect breast cancer cell from cisplatin. Controlling SIRT3-induced UPR may be a potential therapeutic target to increase the sensitivity of breast cancer chemotherapy.

Complete genome sequencing of Pseudomonas syringae pv. actinidiae Biovar 3, P155, kiwifruit pathogen originating from China / Sequenciamento completo do genoma Pseudomonas syringae pv. actinidiae Biovar 3, P155, agente patogênico do kiwi, originário da China

Pseudomonas syringae pv. actinidiae is a bacterial pathogen of kiwifruit. Based on the results of the pathogenicity assay, we sequenced the strain Pseudomonas syringae (Psa3) P155 which possesses a series of virulence and resistance genes, CRISPR candidate elements, prophage related sequences, methylation modifications, genomic islands as well as one plasmid. Most importantly, the copper resistance genes copA, copB, copC, copD, and copZ as well as aminoglycoside resistance gene ksgA were identified in strain P155, which would pose a threat to kiwifruit production. The complete sequence we reported here will provide valuable information for a better understanding of the genetic structure and pathogenic characteristics of the genome of P155.

Pseudomonas syringae pv. actinidiae agente causal do cancro bacteriano do kiwi. Com base nos resultados do teste de patogenicidade, foi sequenciado um isolado de Pseudomonas syringae (Psa3) P155, que abriga a uma série de genes de virulência e resistência, elementos candidatos CRISPR, sequências relacionadas a profagos, modificações na metilação, ilhas genômicas, e também um plasmídeo. O mais importante foram os genes de resistência ao cobre, copA, copB, copC, copD e copZ, bem como, o gene de resistência aminoglicosídea ksgA identificados na estirpe P155, os quais representariam uma ameaça à produção de kiwi. A sequência completa relatada fornecerá informações valiosas para uma melhor compreensão da estrutura genética e as características patogênicas do genoma de P155.

The Significance of Interleukin-6 in the Early Detection of Surgical Site Infections after Definitive Operation for Gastrointestinal Fistulae.

BACKGROUND: Surgical site infections (SSIs) are among the most common complications after definitive treatment for intestinal fistulae. Serum inflammatory markers including white blood cell count (WBC), C-reactive protein (CRP), interleukin-6 (IL-6), as well as procalcitonin (PCT) have been used to help diagnosis post-operative complications. OBJECTIVE: The goal of this study was to assess the clinical value of inflammatory markers, specifically IL-6, in predicting SSIs after intestinal fistulae resection. METHODS: A total of 184 consecutive patients who underwent elective intestinal fistula resection were enrolled prospectively. All patients were screened to exclude patients with existing clinical infection. Plasma IL-6 concentrations, serum PCT, and CRP concentrations were measured pre-operatively and on post-operative days one, three, and seven. The predictive value of each laboratory marker for SSI was calculated. RESULTS: The incidence of SSI after elective intestinal fistula resection was 26.7%. Interleukin-6, PCT, and CRP concentrations were higher in patients with SSIs compared with patients without. In contrast, there was no statistical difference for WBC counts between the two groups. Receiver operating characteristic curves demonstrated that IL-6 had the highest diagnostic effectiveness for post-operative SSI on post-operative day one, with an area under the curve of 0.77, and a sensitivity of 85.7% and specificity of 63.9%. CONCLUSION: A concentration of IL-6 above 95.6 ng/L on post-operative day one and 52.5 ng/L on post-operative day three, and a concentration of PCT exceeding 0.61 mcg/L predict the occurrence of SSI after definitive operations for gastrointestinal fistulae.