Circulating endothelial microparticles (EMPs) are emerging as biomarkers of chronic obstructive pulmonary disease (COPD) in individuals exposed to cigarette smoke (CS), but their mechanism of release and function remain unknown. We assessed biochemical and functional characteristics of EMPs and circulating microparticles (cMPs) released by CS. CS exposure was sufficient to increase microparticle levels in plasma of humans and mice, and in supernatants of primary human lung microvascular endothelial cells. CS-released EMPs contained predominantly exosomes that were significantly enriched in let-7d, miR-191; miR-126; and miR125a, microRNAs that reciprocally decreased intracellular in CS-exposed endothelium. CS-released EMPs and cMPs were ceramide-rich and required the ceramide-synthesis enzyme acid sphingomyelinase (aSMase) for their release, an enzyme which was found to exhibit significantly higher activity in plasma of COPD patients or of CS-exposed mice. The ex vivo or in vivo engulfment of EMPs or cMPs by peripheral blood monocytes-derived macrophages was associated with significant inhibition of efferocytosis. Our results indicate that CS, via aSMase, releases circulating EMPs with distinct microRNA cargo and that EMPs affect the clearance of apoptotic cells by specialized macrophages. These targetable effects may be important in the pathogenesis of diseases linked to endothelial injury and inflammation in smokers.

Recent studies suggest that peroxisome proliferator-activated receptor gamma (PPARγ) agonists may have cancer chemopreventive activity. Other studies have shown that loss of epidermal PPARγ results in enhanced chemical carcinogenesis in mice via unknown mechanisms. However, ultraviolet B (UVB) exposure represents the primary etiological agent for skin cancer formation and the role of PPARγ in photobiology and photocarcinogenesis is unknown. In previous studies, we demonstrated that UVB irradiation of cells results in the formation of oxidized glycerophosphocholines that exhibit PPARγ ligand activity. We therefore hypothesized that PPARγ would prove to be a chemopreventive target in photocarcinogenesis. We first showed that UVB irradiation of mouse skin causes generation of PPARγ agonist species in vivo. We then generated SKH-1 hairless, albino mice deficient in epidermal Pparg (Pparg−/−epi) using a cytokeratin 14 driven Cre-LoxP strategy. Using a chronic model of UVB photocarcinogenesis, we next showed that Pparg−/−epi mice exhibit an earlier onset of tumor formation, increased tumor burden, and tumor progression. Increased tumor burden in Pparg−/−epi mice was accompanied by a significant increase in epidermal hyperplasia and p53 positive epidermal cells in surrounding skin lacking tumors. Following acute UVB irradiation, Pparg−/−epi mice exhibited an augmentation of both UVB-induced caspase 3/7 activity and inflammation. Increased apoptosis and inflammation was also observed following treatment with the PPARγ antagonist GW9662. With chronic UVB irradiation, Pparg−/−epi mice exhibited a sustained increase in erythema and transepidermal water loss relative to wildtype littermates. This suggests that PPARγ agonists could have possible chemopreventive activity in non-melanoma skin cancer.

Background: Deficiency of the nucleotide excision repair protein xeroderma pigmentosum type A (XPA) is characterized by photosensitivity. Results: XPA-deficient cells and mice generate increased platelet-activating factor (PAF), oxidized glycerophosphocholines, and skin inflammation following ultraviolet B radiation with the skin inflammation blocked by PAF antagonists. Conclusion: PAF-R signaling mediates photosensitivity seen in XPA deficiency. Significance: These studies provide important insights into the mechanisms of photosensitivity.

Our understanding of the pathophysiological basis of chronic thromboembolic pulmonary hypertension (CTEPH) will be accelerated by an animal model that replicates the phenotype of human CTEPH. Sprague-Dawley rats were administered a combination of a single dose each of plastic microspheres and vascular endothelial growth factor receptor antagonist in polystyrene microspheres (PE) + tyrosine kinase inhibitor SU5416 (SU) group. Shams received volume-matched saline; PE and SU groups received only microspheres or SU5416, respectively. PE + SU rats exhibited sustained pulmonary hypertension (62 ± 13 and 53 ± 14 mmHg at 3 and 6 weeks, respectively) with reduction of the ventriculoarterial coupling in vivo coincident with a large decrement in peak rate of oxygen consumption during aerobic exercise, respectively. PE + SU produced right ventricular hypokinesis, dilation, and hypertrophy observed on echocardiography, and 40% reduction in right ventricular contractile function in isolated perfused hearts. High-resolution computed tomographic pulmonary angiography and Ki-67 immunohistochemistry revealed abundant lung neovascularization and cellular proliferation in PE that was distinctly absent in the PE + SU group. We present a novel rodent model to reproduce much of the known phenotype of CTEPH, including the pivotal pathophysiological role of impaired vascular endothelial growth factor-dependent vascular remodeling. This model may reveal a better pathophysiological understanding of how PE transitions to CTEPH in human treatments.

Although platelet-activating factor (PAF) is a well-known acute inflammatory mediator, little is known regarding the role of PAF in chronic inflammation. Phorbol esters are known to stimulate PAF production. Moreover, the ability of repeated applications of phorbol esters to induce a sustained inflammatory response is crucial to their tumorigenic activity. We therefore examined whether PAF acts as a mediator of phorbol ester-induced inflammation and tumorigenesis. While PAF receptor knockout mice (PAFR(-/-)) showed an expected but modest reduction in the acute inflammatory response to phorbol 12-myristate 13-acetate (PMA), these mice exhibited a surprising increase in inflammation following chronic PMA application. This increased inflammation was documented by a number of findings that included: increased skin thickness, increased myeloperoxidase activity and expression and increased expression of known inflammatory mediators. Interestingly, vehicle-treated PAFR(-/-) mice also exhibited modest increases in levels of inflammatory markers. This suggests that the platelet activating factor receptor (PAFR) acts to suppress chronic inflammation in response to other stimuli, such as barrier disruption. The idea that chronic PAFR activation is anti-inflammatory was documented by repetitive topical PAFR agonist administration that resulted in reduced myeloperoxidase activity in skin. We next utilized a 7,12-dimethylbenz(a)anthracene/PMA carcinogenesis protocol to demonstrate that PAFR(-/-) mice exhibit significantly increased tumor formation and malignant progression compared with wild-type control mice. These studies provide evidence for two important, unexpected and possibly interrelated pathological roles for the PAFR: first, the PAFR acts to suppress PMA-induced chronic inflammation; secondly, the PAFR acts to suppress neoplastic development in response to chemical carcinogens.