LncRNA H19/miR-107 regulates endothelial progenitor cell pyroptosis and promotes flow recovery of lower extremity ischemia through targeting FADD.

BACKGROUND: Peripheral artery disease (PAD) is an ischemic disease with a rising incidence worldwide. The lncRNA H19 (H19) is enriched in endothelial progenitor cells (EPCs), and transplantation of pyroptosis-resistant H19-overexpressed EPCs (oe-H19-EPCs) may promote vasculogenesis and blood flow recovery in PAD, especially with critical limb ischemia (CLI). METHODS: EPCs isolated from human peripheral blood was characterized using immunofluorescence and flow cytometry. Cell proliferation was determined with CCK8 and EdU assays. Cell migration was assessed by Transwell and wound healing assays. The angiogenic potential was evaluated using tube formation assay. The pyroptosis pathway-related protein in EPCs was detected by western blot. The binding sites of H19 and FADD on miR-107 were analyzed using Luciferase assays. In vivo, oe-H19-EPCs were transplanted into a mouse ischemic limb model, and blood flow was detected by laser Doppler imaging. The transcriptional landscape behind the therapeutic effects of oe-H19-EPCs on ischemic limbs were examined with whole transcriptome sequencing. RESULTS: Overexpression of H19 in EPCs led to an increase in proliferation, migration, and tube formation abilities. These effects were mediated through pyroptosis pathway, which is regulated by the H19/miR-107/FADD axis. Transplantation of oe-H19-EPCs in a mouse ischemic limb model promoted vasculogenesis and blood flow recovery. Whole transcriptome sequencing indicated significant activation of vasculogenesis pathway in the ischemic limbs following treatment with oe-H19-EPCs. CONCLUSIONS: Overexpression of H19 increases FADD level by competitively binding to miR-107, leading to enhanced proliferation, migration, vasculogenesis, and inhibition of pyroptosis in EPCs. These effects ultimately promote the recovery of blood flow in CLI.

iPSCs-derived mesenchymal stromal cells mitigate anxiety and neuroinflammation in aging female mice.

Perimenopause is a natural transition to menopause, when hormone disturbance can result in both short-term mental disorders, such as anxiety, and long-term neuroinflammation due to blood-brain barrier (BBB) impairment, which may lead to more serious neurological disorders later on, such as dementia. Effective treatments may prevent both short-term and long-term neurological sequela, which formed the aim of this study. In aged female C57BL/6 mice (16-18 months of age), mesenchymal stromal cells (MSCs) differentiated from human-induced pluripotent stem cells (iPSCs), were administered via tail vein injection. Mice showed increased blood estrogen levels, alleviated anxiety and neuroinflammation, and improved BBB integrity. Interestingly, transplanted MSCs were located close to ovarian sympathetic nerves and decreased ovarian norepinephrine levels, which in turn increased ovarian estrogen secretion. Moreover, the administration of anastrozole, an inhibitor of estrogen synthesis, diminished the therapeutic effects of MSCs in vivo, suggesting the effect to be estrogen-dependent. In vitro study confirmed the impact of MSCs on sympathetic nerves via mitochondria exchange. In conclusion, iPSC-derived MSCs may provide a novel option to manage perimenopause-related hormonal dysregulation and neurological disorders during the female aging process.

Novel insight of N6-methyladenosine modified subtypes in abdominal aortic aneurysm.

Background: N6-methyladenosine (m6A) is the most prevalent non-cap reversible modification present in messenger RNAs and long non-coding RNAs, and its dysregulation has been linked to multiple cardiovascular diseases, including cardiac hypertrophy and atherosclerosis. Although limited studies have suggested that m6A modification contributes to abdominal aortic aneurysm (AAA) development, the full landscape of m6A regulators that mediate modification patterns has not been revealed. Methods: To distinguish the m6A methylation subtypes in AAA patients, an unsupervised clustering method was carried out, based on the mRNA levels of 17 m6A methylation regulators. Differentially expressed genes were identified by comparing clusters. An m6Ascore model was calculated using principal component analysis and structured to assess the m6A methylation patterns of single samples. Subsequently, the relationship between the m6Ascore and immune cells and the hallmark gene set was analyzed. Finally, pairs of circRNA-m6A regulators and m6A regulators-m6A related genes were used to establish a network. Results: We identified three m6A methylation subtypes in the AAA samples. The m6Acluster A and C were characterized as more immunologically activated because of the higher abundance of immune cells than that in m6Acluster B. The m6Acluster B was less enriched in inflammatory pathways and more prevalent in pathways related to extracellular matrix stability. Subsequently, we divided the individual samples into two groups according to the m6Ascore, which suggested that a high m6Ascore predicted more active inflammatory pathways and higher inflammatory cell infiltration. A network consisting of 9 m6A regulators and 37 circRNAs was constructed. Conclusion: This work highlighted that m6A methylation modification was highly correlated with immune infiltration of AAA, which may promote the progression of AAA. We constructed an individualized m6Ascore model to provide evidence for individualized treatments in the future.

The Long Non-coding RNA AC148477.2 Is a Novel Therapeutic Target Associated With Vascular Smooth Muscle Cells Proliferation of Femoral Atherosclerosis.

Arteriosclerosis obliterans (ASO) is a limb manifestation of large vessel atherosclerosis. Phenotype switching of vascular smooth muscle cells (VSMCs) occurs in the course of the pathological process. The underlying mechanism of SMCs proliferation remains unclear. Several studies have demonstrated that the dysregulation of long non-coding RNA (lncRNAs) plays a pivotal part in the progression of ASO by exacerbating the proliferation of VSMCs. Based on the endogenous competitive RNA (ceRNA) hypothesis, the mechanism of lncRNAs involved in the pathology of VSMCs was exposed, while the entire map of the regulatory network remains to be elucidated. In the current study, genes and the lncRNAs modules that are relevant to the clinical trait were confirmed through weighted gene co-expression network analysis (WGCNA). In this study, we comprehensively constructed a specific lncRNAs-mediated ceRNA and RBP network. The three lncRNAs, HMGA1P4, C5orf66, and AC148477.2, influenced the proliferation of VSMCs and were found to be associated with the immune landscape, thus they were ultimately screened out. Further verification revealed that AC147488.2 was significantly down-regulated in both ASO arteries and all stages of proliferative VSMCs, which implied that AC147488.2 might have a significant impact on ASO. This finding would improve our understanding of the epigenetic regulation of ASO and unravel novel diagnostic and therapeutic targets.

Human umbilical cord mesenchymal stem cell-derived exosomal microRNA-148a-3p inhibits neointimal hyperplasia by targeting Serpine1.

BACKGROUND: Restenosis is inevitable when patients undergo percutaneous transluminal angioplasty due to neointimal hyperplasia (NIH). Human umbilical cord mesenchymal stem cell-derived exosomes (hucMSC-Exos) have been studied in the field of cardiovascular diseases. However, the effects and mechanisms of hucMSC-Exos on NIH are unclear. We aimed to investigate whether MSC-Exos regulate vascular smooth muscle cell (VSMC) functions to inhibit NIH and explore the underlying mechanisms. METHODS: HucMSCs and mouse VSMCs were isolated and characterized by flow cytometry and immunofluorescence. HucMSC-Exos were identified by transmission electron microscopy, nanoparticle tracking analysis and western blots. Exosomes (Exos) were intravenously injected into mice with left common carotid artery ligation, and their effects on NIH were assessed by haematoxylin and eosin (H&E) and immunohistochemistry staining. The effects of hucMSC-Exos on VSMCs were evaluated by Cell Counting Kit-8, scratch wound, Transwell and Western blot assays. MicroRNA sequencing data in the Gene Expression Omnibus and mRNA sequencing results were used to identify potential molecules in hucMSC-Exos and target genes in VSMCs, respectively. We tested the regulatory effect of microRNAs in Exos and target genes in VSMCs using overexpression and knockdown experiments. RESULTS: Primary hucMSCs, VSMCs and hucMSC-Exos were isolated and characterized. Administration of hucMSC-Exos suppressed NIH after artery ligation. H&E and immunohistochemistry results showed that hucMSC-Exos decreased the intima and media area and intima/media ratio, increased the contractile phenotype protein SM22a in the media layer and downregulated Serpine1 expression in the carotid artery. Exos were ingested by VSMCs, which inhibited migration and upregulated SM22a expression by suppressing Serpine1 expression in vitro. MiR-148a-3p was enriched in hucMSC-Exos and repressed Serpine1 by targeting its 3' untranslated region. Moreover, exosomal miR-148a-3p suppressed VSMC phenotypic switching and migration by targeting Serpine1. CONCLUSIONS: We found that hucMSC-Exos inhibited NIH in a mouse carotid artery ligation model and that the inhibitory effects on VSMC phenotypic switching and migration were mediated by delivery of miR-148a-3p to VSMCs to target Serpine1.

Macrophage NFATc3 prevents foam cell formation and atherosclerosis: evidence and mechanisms.

AIMS: Our previous study demonstrated that Ca2+ influx through the Orai1 store-operated Ca2+ channel in macrophages contributes to foam cell formation and atherosclerosis via the calcineurin-ASK1 pathway, not the classical calcineurin-nuclear factor of activated T-cell (NFAT) pathway. Moreover, up-regulation of NFATc3 in macrophages inhibits foam cell formation, suggesting that macrophage NFATc3 is a negative regulator of atherogenesis. Hence, this study investigated the precise role of macrophage NFATc3 in atherogenesis. METHODS AND RESULTS: Macrophage-specific NFATc3 knockout mice were generated to determine the effect of NFATc3 on atherosclerosis in a mouse model of adeno-associated virus-mutant PCSK9-induced atherosclerosis. NFATc3 expression was decreased in macrophages within human and mouse atherosclerotic lesions. Moreover, NFATc3 levels in peripheral blood mononuclear cells from atherosclerotic patients were negatively associated with plaque instability. Furthermore, macrophage-specific ablation of NFATc3 in mice led to the atherosclerotic plaque formation, whereas macrophage-specific NFATc3 transgenic mice exhibited the opposite phenotype. NFATc3 deficiency in macrophages promoted foam cell formation by potentiating SR-A- and CD36-meditated lipid uptake. NFATc3 directly targeted and transcriptionally up-regulated miR-204 levels. Mature miR-204-5p suppressed SR-A expression via canonical regulation. Unexpectedly, miR-204-3p localized in the nucleus and inhibited CD36 transcription. Restoration of miR-204 abolished the proatherogenic phenotype observed in the macrophage-specific NFATc3 knockout mice, and blockade of miR-204 function reversed the beneficial effects of NFATc3 in macrophages. CONCLUSION: Macrophage NFATc3 up-regulates miR-204 to reduce SR-A and CD36 levels, thereby preventing foam cell formation and atherosclerosis, indicating that the NFATc3/miR-204 axis may be a potential therapeutic target against atherosclerosis.

Curcumin exerts a protective effect on murine knee chondrocytes treated with IL-1ß through blocking the NF-κB/HIF-2α signaling pathway.

BACKGROUND: Osteoarthritis (OA) is characterized by erosion and degradation of articular cartilage. This study assessed the effects of curcumin on mouse knee cartilage chondrocytes. METHODS: Chondrocytes were treated for 24 hours with interleukin IL-1ß (10 ng/mL) alone, or the combination of curcumin (10, 20, and 50 µM) and IL-1ß. The proliferation, viability, and cytotoxicity of the chondrocytes were evaluated by the MTS assay. Expression of SOX9, AGG, Col2α, MMP9, ADAMTS5, COX2, iNOS, pIκB-α, pNF-κB, and hypoxia-inducible factor-2α (HIF-2α) were detected by western blotting or quantitative polymerase chain reaction (q-PCR). Nuclear translocation of NF-κB and HIF-2α were investigated by immunofluorescence and immunohistochemistry. In in vivo experiments, mice were subjected to destabilization of the medial meniscus (DMM) and given curcumin orally for 6 weeks. Cartilage integrity was evaluated by OARSI (Osteoarthritic Research Society International) scores. RESULTS: Curcumin significantly inhibited the IL-1ß-induced reduction of cell viability, degradation of ECM, and the expression of SOX9, Col2α, and AGG (P<0.01). Western blotting, immunofluorescence and immunohistochemistry experiments demonstrated that curcumin dramatically inhibited the activation of NF-κB/HIF-2α in chondrocytes treated with IL-1ß (P<0.01). The articular scores were significantly lower in the DMM-induced OA mice compared to OA mice treated with curcumin (P<0.01). CONCLUSIONS: Curcumin may have the potential to inhibit OA development, partly through suppressing the activation of the NF-κB/HIF-2α pathway.

Characteristics and Prognosis of Abdominal or Thoracic Aortic Aneurysm Patients Admitted to Intensive Care Units After Surgical Treatment: A Multicenter Retrospective Observational Study.

OBJECTIVE: To investigate the characteristics and prognosis of abdominal or thoracic aortic aneurysm (AAA or TAA) patients admitted to intensive care unit (ICU) postoperatively. METHODS: Patients admitted to ICU postoperatively with a primary diagnosis of AAA or TAA were screened in the eICU Collaborative Research Database, which contained data from multiple ICUs throughout the continental United States in 2014 and 2015. Baseline characteristics and comorbidities and were investigated and factors associated with ICU mortality were explored using univariable logistic regression. Receiver operating characteristic (ROC) curve analysis was performed to evaluate the prognosis predictive performance of the widely used severity scoring system APACHE IVa. RESULTS: A total of 974 patients including 677 AAA and 297 TAA patients admitted to ICU postoperatively were included. Compared with TAA, AAA patients had a significantly higher median age (72 versus 64 years, P<0.001). 89.07% AAA and 84.51% TAA patients underwent elective surgery (P=0.046), 8.71% AAA and 31.99% TAA patients were with aortic dissection (P<0.001), and 10.19% AAA and 2.36% TAA patients suffered from rupture of aortic aneurysm (P<0.001). Hypertension requiring treatment was the most common comorbidity (57.31% for AAA and 61.95% for TAA). TAA patients had significantly higher ICU mortality (9.43% versus 2.36%, P<0.001) than AAA. Several factors were found to be significantly associated with ICU mortality, including urgent surgery, with aortic dissection, rupture of aortic aneurysm, TAA, and a higher APACHE IVa score on ICU admission. APACHE IVa showed a good predictive performance for ICU mortality with an area under the ROC curve of 0.9176 (95% CI 0.8789-0.9390). CONCLUSION: The prognosis of aortic aneurysm patients admitted to ICU postoperatively is yet to improve, and factors associated with prognosis are mainly related to the condition itself. APACHE IVa can be used for prognosis prediction.

Construction of a Novel Ferroptosis-Related Gene Signature of Atherosclerosis.

Atheroclerosis refers to a chronic inflammatory disease featured by the accumulation of fibrofatty lesions in the intima of arteries. Cardiovasular events associated with atherosclerosis remain the major causes of mortality worldwide. Recent studies have indicated that ferroptosis, a novel programmed cell death, might participate in the process of atherosclerosis. However, the ferroptosis landscape is still not clear. In this study, 59 genes associated with ferroptosis were ultimately identified in atherosclerosis in the intima. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed for functional annotation. Through the construction of protein-protein interaction (PPI) network, five hub genes (TP53, MAPK1, STAT3, HMOX1, and PTGS2) were then validated histologically. The competing endogenous RNA (ceRNA) network of hub genes was ultimately constructed to explore the regulatory mechanism between lncRNAs, miRNAs, and hub genes. The findings provide more insights into the ferroptosis landscape and, potentially, the therapeutic targets of atherosclerosis.

Restenosis Prevention With Drug Eluting or Covered Stents in Femoropopliteal Arterial Occlusive Disease: Evidence From a Comprehensive Network Meta-analysis.

BACKGROUND/OBJECTIVE: Endovascular interventions for femoropopliteal (FP) arterial diseases are limited by the development of restenosis. Current drug coated devices are capable of preventing restenosis by releasing antiproliferative agents to the vessel wall. However, default strategies for the treatment of FP diseases remain controversial. The aim of this study was to investigate the efficacy differences between drug eluting stents (DES), covered stents (CS), and other commonly used endovascular treatments in FP lesions, including drug coated balloons (DCBs), bare metal stents (BMS), and percutaneous transluminal angioplasty (PTA). METHODS: A comprehensive network meta-analysis was conducted using data from relevant randomised control trials published up to 16 December 2018. Primary patency and target lesion revascularisation (TLR) at 12 months were set as the primary and secondary end points, respectively. RESULTS: Twenty-eight eligible trials including 4728 patients were selected. DES was ranked as the most effective treatment in the multidimensional analysis of primary patency; however, there was no significant difference in the efficacy of DES and that of CS, DCB, and BMS. However, in short lesions (<10 cm), DES was significantly more effective than DCB (odds ratio 0.35; 95% confidence interval 0.15-0.83). Primary patency at 12 months was significantly lower with PTA. In terms of preventing TLR, DCB was ranked first, followed by DES, CS, BMS, and PTA. TLR was significantly higher with PTA than with other treatment strategies. CONCLUSION: The findings of this network meta-analysis suggest that this is not the appropriate time to identify the best endovascular treatment strategy for the FP segment. DES is effective in maintaining mid-term patency, especially in short lesions, whereas DCB seems more suitable for clinical use.

Presence of an Agrobacterium-Type Tumor-Inducing Plasmid in Neorhizobium sp. NCHU2750 and the Link to Phytopathogenicity.

The genus Agrobacterium contains a group of plant-pathogenic bacteria that have been developed into an important tool for genetic transformation of eukaryotes. To further improve this biotechnology application, a better understanding of the natural genetic variation is critical. During the process of isolation and characterization of wild-type strains, we found a novel strain (i.e., NCHU2750) that resembles Agrobacterium phenotypically but exhibits high sequence divergence in several marker genes. For more comprehensive characterization of this strain, we determined its complete genome sequence for comparative analysis and performed pathogenicity assays on plants. The results demonstrated that this strain is closely related to Neorhizobium in chromosomal organization, gene content, and molecular phylogeny. However, unlike the characterized species within Neorhizobium, which all form root nodules with legume hosts and are potentially nitrogen-fixing mutualists, NCHU2750 is a gall-forming pathogen capable of infecting plant hosts across multiple families. Intriguingly, this pathogenicity phenotype could be attributed to the presence of an Agrobacterium-type tumor-inducing plasmid in the genome of NCHU2750. These findings suggest that these different lineages within the family Rhizobiaceae are capable of transitioning between ecological niches by having novel combinations of replicons. In summary, this work expanded the genomic resources available within Rhizobiaceae and provided a strong foundation for future studies of this novel lineage. With an infectivity profile that is different from several representative Agrobacterium strains, this strain may be useful for comparative analysis to better investigate the genetic determinants of host range among these bacteria.