Regulation Of Prostaglandin D2 And Angiogenesis-Related Factors From Human Skin Mast Cells By Interleukin-6 And Resveratrol

Mast cells are classically recognized as the effector cells of Immunoglobulin E (IgE)-mediated hypersensitivity reactions (i.e. allergic reactions). Mast cells also play an important role in the innate immune response to parasitic helminth infection. Allergic disease including allergic rhinitis, asthma, atopic dermatitis, and anaphylaxis, is a major health concern in the United States with greater than 60 million Americans suffering from allergy and asthma. In addition, mast cells have more recently been implicated in non-allergic disease including various human cancers, which will affect approximately 39.6% of U.S. men and women. Therefore, understanding the immunological and molecular mechanisms that regulate mast cells is vital to our understanding of their role in both allergic and non-allergic disease in humans. In this study, we identified novel mechanisms that regulate FcεRI-induced biosynthesis of inflammatory Prostaglandin D2 (PGD2) from in situ-matured mast cells from human skin. Specifically, we demonstrated that interleukin-6 (IL-6), a classical pro-inflammatory cytokine, is a positive regulator, whereas Resveratrol, a natural polyphenol found in the skin of red grapes, is a negative regulator of PGD2 biosynthesis. We demonstrate that Resveratrol at relatively low concentrations specifically inhibited the production of PGD2 without affecting degranulation. Mechanistically, Resveratrol inhibited the expression of cyclooxygenase-2 (COX-2), an enzyme in the arachidonic acid pathway that is directly involved in PGD2 biosynthesis, but had no effect on phosphorylation of Syk kinase, which is a critical protein in mast cell degranulation. In contrast, IL-6 enhanced FcεRI-induced COX-2 expression leading to increased PGD2 biosynthesis. In support of this, we showed that FcεRI and gp130 (the signaling portion of the IL-6 receptor) signals cooperate to enhance STAT-3 phosphorylation, a key event in the IL-6 receptor signal pathway. Moreover, inhibition of IL-6-induced phosphorylation of STAT-3 with the specific inhibitor C188-9 also attenuated the increased production of PGD2. Thus, IL-6 potentiates PGD2 production by a STAT-3-dependent mechanism. Jointly, these data demonstrate that Resveratrol is a negative regulator, and IL-6 a positive regulator of FcεRI-induced PGD2 biosynthesis from human skin mast cells. We further demonstrate that IL-6 induced the expression of vascular endothelial growth factor (VEGF), a major regulator of angiogenesis, from human skin mast cells also by a STAT-3-dependent mechanism. Lastly, we demonstrated by proteome profiling analysis that human skin mast cells spontaneously secrete several angiogenesis-related proteins – CXCL16, DPPIV, Endothelin-1, GM-CSF, IL-8, MCP-1, Pentraxin 3, Serpin E1, Serpin F1, TIMP-1, Thrombospondin-1, uPA, and VEGF – at relatively high concentrations. The array data was verified by specific enzyme linked immunoassay (ELISA). This novel data suggests an innate role for mast cells in both human tumorigenesis and normal development of human vasculature. Together, these novel findings enhance our understanding of the mechanisms that regulate the release of allergic mediators from mast cells, and suggest a possible role for mast cells in oncogenesis and normal vascular development

Nuevas variantes clínicas y moleculares de mastocitosis. Caracterización clínico-biológica e implicaciones terapéuticas

INTRODUCCIÓN La mastocitosis sistémica (SM) constituye un grupo heterogéneo de enfermedades caracterizadas por la presencia de mastocitos (MCs) patológicos en diferentes órganos y tejidos, entre los que destaca la médula ósea (MO). La Organización de la Salud (OMS) reconoce actualmente diferentes subtipos de SM con implicaciones pronósticas, cuyo diagnóstico se basa en criterios bien establecidos. HIPÓTESIS DE TRABAJO Y OBJETIVOS A pesar de la utilidad de la clasificación de la OMS de mastocitosis para identificar subtipos de SM con mal pronóstico, los factores asociados a riesgo de progresión de las formas indolentes (ISM) no han sido aún establecidos. Además, en los últimos años han surgido nuevas variantes de SM con particularidades clínicas y biológicas diferentes al resto de subtipos de SM, que pueden pasar inadvertidas de acuerdo a los actuales criterios diagnósticos de la OMS; por este motivo, hasta la fecha estas nuevas variantes de la enfermedad no han sido bien caracterizadas.En la presente tesis doctoral nos planteamos como objetivos identificar los factores de riesgo de progresión de ISM, y caracterizar desde el punto de vista clínico y biológico la ISM sin lesión cutánea asociada a anafilaxia (ISMs-) y la SM bien diferenciada (WDSM), y establecer sus implicaciones pronósticas y terapéuticas. MATERIALES, MÉTODOS Y RESULTADOS Esta tesis doctoral está constituida por ocho trabajos llevados a cabo en pacientes con mastocitosis, cuyos principales resultados son: 1) la demostración de la afectación multilineal de la hematopoyesis por la mutación de KIT como principal factor de riesgo de progresión de la ISM, 2) el desarrollo de un modelo predictivo con elevada sensibilidad y especificidad para ISMs-, y 3) el establecimiento de criterios diagnósticos específicos complementarios a los de la OMS para la identificación de WDSM. CONCLUSIONES1. La identificación de pacientes con ISM de alto riesgo de progresión permite un seguimiento más estrecho y la adopción de medidas terapéuticas precoces.2. El diagnóstico de ISMs- y WDSM requiere de técnicas diagnósticas altamente sensibles y de criterios diagnósticos específicos, respectivamente

Mast Cell Proteases 6 and 7 Stimulate Angiogenesis by Inducing Endothelial Cells to Release Angiogenic Factors.

Mast cell proteases are thought to be involved with tumor progression and neo-vascularization. However, their exact role is still unclear. The present study was undertaken to further elucidate the function of specific subtypes of recombinant mouse mast cell proteases (rmMCP-6 and 7) in neo-vascularization. SVEC4-10 cells were cultured on Geltrex® with either rmMCP-6 or 7 and tube formation was analyzed by fluorescence microscopy and scanning electron microscopy. Additionally, the capacity of these proteases to induce the release of angiogenic factors and pro and anti-angiogenic proteins was analyzed. Both rmMCP-6 and 7 were able to stimulate tube formation. Scanning electron microscopy showed that incubation with the proteases induced SVEC4-10 cells to invade the gel matrix. However, the expression and activity of metalloproteases were not altered by incubation with the mast cell proteases. Furthermore, rmMCP-6 and rmMCP-7 were able to induce the differential release of angiogenic factors from the SVEC4-10 cells. rmMCP-7 was more efficient in stimulating tube formation and release of angiogenic factors than rmMCP-6. These results suggest that the subtypes of proteases released by mast cells may influence endothelial cells during in vivo neo-vascularization