ABSTRACT
Background and aim: Inflammatory myopathies are heterogeneous in terms of etiology, (immuno)pathology, and clinical findings. Endothelial cell injury, as it occurs in DM, is a common feature of numerous inflammatory and non-inflammatory vascular diseases. Vascular regeneration is mediated by both local and blood-derived mechanisms, such as the mobilization and activation of so-called proangiogenic cells (PACs) or early endothelial progenitor cells (eEPCs). The current study aimed to evaluate parameters of eEPC integrity in dermatomyositis (DM), compared to necrotizing myopathy (NM) and to non-myopathic controls. Methods: Blood samples from DM and NM patients were compared to non-myositis controls and analyzed for the following parameters: circulating CD133+/VEGFR-2+ cells, number of colony-forming unit endothelial cells (CFU-ECs), concentrations of angiopoietin 1, vascular endothelial growth factor (VEGF), and CXCL-16. Muscle biopsies from DM and NM subjects underwent immunofluorescence analysis for CXCR6, nestin, and CD31 (PECAM-1). Finally, myotubes, derived from healthy donors, were stimulated with serum samples from DM and NM patients, subsequently followed by RT-PCR for the following candidates: IL-1ß, IL-6, nestin, and CD31. Results: Seventeen (17) DM patients, 7 NM patients, and 40 non-myositis controls were included. CD133+/VEGFR-2+ cells did not differ between the groups. Both DM and NM patients showed lower CFU-ECs than controls. In DM, intramuscular CD31 abundances were significantly reduced, which indicated vascular rarefaction. Muscular CXCR6 was elevated in both diseases. Circulating CXCL-16 was higher in DM and NM in contrast, compared to controls. Serum from patients with DM but not NM induced a profound upregulation of mRNS expression of CD31 and IL-6 in cultured myotubes. Conclusion: Our study demonstrates the loss of intramuscular microvessels in DM, accompanied by endothelial activation in DM and NM. Vascular regeneration was impaired in DM and NM. The findings suggest a role for inflammation-associated vascular damage in the pathogenesis of DM.
ABSTRACT
Coccidioides immitis and Coccidioides posadasii are soil fungi endemic to desert regions of the southwestern United States, and the causative agents of valley fever, or coccidioidomycosis. Studies have shown that the distribution of Coccidioides in soils is sporadic and cannot be explained by soil characteristics alone, suggesting that biotic and other abiotic factors should be examined. However, tools to reliably and robustly screen the large number of soils needed to investigate these potential associations have not been available. Thus, we developed a real-time polymerase chain reaction (PCR) assay for testing environmental samples by modifying CocciDx, an assay validated for testing clinical specimens to facilitate coccidioidomycosis diagnosis. For this study, we collected soil samples from previously established locations of C. posadasii in Arizona and new locations in fall 2013 and spring 2014, and screened the extracted DNA with the new assay known as CocciEnv. To verify the presence of Coccidioides in soil using an alternate method, we employed next generation amplicon sequencing targeting the ITS2 region. Results show our modified assay, CocciEnv, is a rapid and robust method for detecting Coccidioides DNA in complex environmental samples. The ability to test a large number of soils for the presence of Coccidioides is a much-needed tool in the understanding of the ecology of the organism and epidemiology of the disease and will greatly improve our understanding of this human pathogen.
