Background An inflammatory environment around the vessel wall caused by leukocyte infiltration is one of the characteristic histopathological features of microscopic polyangiitis (MPA); however, the pathogenic mechanisms are not fully understood. Studies have found that circulating microRNA (miRNA) can be used as potential biomarkers for the diagnosis and classification of anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitides (AAV), and the E3 ubiquitin ligase casitas B-lineage lymphoma (CBL) seems to be associated with inflammation. In addition, evidence indicates that miRNA can be tracked into exosomes and transferred into recipient cells to mediate the process of vascular endothelial injury. Herein, we aimed to identify the profiles of exosomal miRNA, and determine the effect of exosomal miR-1287-5p and its target gene CBL on vascular endothelial cells in MPA. Method We isolated plasma exosomes from patients with MPA (MPA-exo) and healthy controls (HC-exo) by ultracentrifugation and conducted exosome small-RNA sequencing to screen differential miRNA expression in MPA-exo (n = 3) compared to HC-exo (n = 3). We measured the expression levels of miR-1303, miR-1287-5p, and miR-129-1-3p using quantitative reverse transcription-polymerase chain reaction (qRT-PCR, n = 6) and performed dual luciferase reporter gene assays to confirm the downstream target gene of miR-1287-5p. In addition, we treated human umbilical vein endothelial cell (HUVEC) with MPA-exo, or transfected them with miR-1287-5p mimic/inhibitor or with CBL-siRNA/CBL-siRNA+ miR-1287-5p inhibitor. After cell culture, we evaluated the effects on vascular endothelial cells by examining the mRNA levels of IL-6, IL-8, MCP-1, ICAM-1 and E-selectin using qRT-PCR and performed neutrophil adhesion assay with haematoxylin staining. Result Transmission electron microscopy, Western blot and nanoparticle tracking analysis showed that we successfully purified exosomes and MPA-exo could be absorbed into HUVEC. We screened a total of 1,077 miRNA by sequencing and observed a high abundance of miR-1287-5p in the exosomes obtained from MPA plasma. The dual luciferase reporter assay identified CBL as a downstream target gene of miR-1287-5p, and the results revealed that MPA-exo decreased CBL protein expression in HUVEC. In addition, treatment with MPA-exo, up-regulating miR-1287-5p or silencing of CBL in HUVEC significantly increased the mRNA expression of inflammatory factors (including IL-6, IL-8, and MCP-1) and adhesion molecules (including ICAM-1 and E-selection) and promoted the adhesion of neutrophils to HUVEC. However, down-regulating miR-1287-5p had the opposite effect. Conclusion Our study revealed that MPA-exo was involved in the intercellular transfer of miR-1287-5p and subsequently promote the development of acute endothelial injury in MPA. MiR-1287-5p and CBL agonists may be promising therapeutic approach for MPA-induced vascular inflammatory injury.

Microscopic polyangiitis (MPA) is an autoimmune disease, characterized by ANCA in blood and necrotizing inflammation of small and medium-sized vessels, one of the three clinical phenotypes of ANCA-associated vasculitis (AAV). Autophagy has been confirmed to be involved in the pathogenesis of AAV. AKT1 is one of the autophagy-regulated proteins. Its single nucleotide polymorphisms (SNPs) are associated with multiple immune-related diseases, but there are rarely studies in AAV. The incidence rate of AAV has a notable geographic difference, and MPA is predominant in China. The aim of this study was to investigate the association between AKT1 SNP and MPA risk. Genotypes of 8 loci in AKT1 were evaluated by multiplex polymerase chain reaction (PCR) and high-throughput sequencing in 416 people, including 208 MPA patients and 208 healthy volunteers from Guangxi in China. Additionally, data of 387 healthy volunteers from China were obtained from the 1000Genomes Project on public database. Differences were observed between the loci (rs2498786, rs2494752, and rs5811155) genotypes in AKT1 and MPA risk (P = 7.0 × 10–4, P = 3.0 × 10–4, and P = 5.9 × 10–5, respectively). A negative association was detected in the Dominant model (P = 1.2 × 10–3, P = 2.0 × 10–4 and P = 3.6 × 10–5, respectively). A haplotype (G-G-T) was associated with MPA risk negatively (P = 7.0 × 10–4). This study suggests that alleles (rs2498786 G, rs2494752 G and rs5811155 insT) are protective factors for MPA and alleles (rs2494752 G and rs5811155 insT) for MPO-ANCA in patients with MPA. There is a haplotype (G-G-T), which is a protective factor for MPA. It suggests that the role of AKT1 in MPA/AAV needs further study to provide more intervention targets for MPA/AAV.

Background Microscopic polyangiitis (MPA) is a systemic autoimmune disease characterized by inflammation of small- and medium-sized blood vessels. Autophagy-related protein polymorphisms are involved in autoimmune disease. The aim of this study was to evaluate the effects of single-nucleotide polymorphisms (SNPs) in the ULK1 and PIK3CA genes on the risk of MPA. Method A total of 208 patients with MPA and 211 controls in the Guangxi Zhuang Autonomous Region were recruited and analyzed. The SNPs selected were detected by polymerase chain reaction and high-throughput sequencing. The differences in allele and genotype frequency, various genetic models, and stratification analyses were evaluated, haplotype evaluation was performed after linkage disequilibrium analysis, and the interaction between gene alleles was analyzed. Results A statistically significant difference was detected in the genotypic distribution of two SNPs between the two groups: ULK1 rs4964879 (p = 0.019) and PIK3CA rs1607237 (p = 0.002). The results of the genetic models revealed that ULK1 rs4964879 and rs9481 were statistically significantly associated with an increased risk of MPA, whereas PIK3CA rs1607237 was associated with a reduced risk. The association between SNPs and MPA risk was affected by age, sex, and ethnicity. The ULK1 haplotype (G-T-A-C-G-A) and PIK3CA haplotype (T-G) were associated with a reduced risk of MPA, while the PIK3CA haplotype (C-G) was associated with an increased risk. Conclusion In this study, polymorphisms in the autophagy-related genes ULK1 and PIK3CA and their association with MPA were examined. The results showed that the polymorphisms in ULK1 (rs4964879 and rs9481) and PIK3CA (rs1607237) were significantly associated with MPA risk in the Guangxi population. However, the molecular mechanisms are still unclear; basic science research and studies with larger samples are needed to confirm our conclusions and explore the underlying mechanisms.

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