Mast Cell Supernatant Inhibit Bone Formation Markers Gene Expression

Abstract

Spontaneously hypertensive rats (SHR) present genetic alterations in bone metabolism and in adaptive immunity. Mast cells (MCs) play a fundamental role in inflammatory response, tissue repair and defense of the organism and may also be involved in the bone metabolism. Thus, in this study we aimed to evaluate the influence of mast cell mediators in the acquisition of the osteogenic phenotype in vitro and in the gene expression of bone markers from calvariae cells of SHR.MethodsPrimary culture from neonatal calvariae obtained by explant technique were expanded and plated in 24‐well culture plates where they received osteogenic medium (OM, ascorbic acid, β‐glycerophosphate and dexamethasone). The RBL‐2H3 MC lines were adapted to osteogenic medium, subsequently stimulated via FcɛRI, and culture supernatant was collected and was added directly into the calvariae culture in concentration of 15%, 10% and 5%. Cell viability was accessed by MTT assay, and gene expression of osteogenic markers was evaluated at day 7 by qRT‐PCR. The protocol was approved by Institutional Animal Care and Use Committees (School of Dentistry of Araçatuba; Process:00490‐2018).ResultsMTT assay did not show a cytotoxic potential from MC supernatant on calvariae culture. MC supernatant, in all concentration, significantly decreased gene expression of the transcription factors Runx2, Osterix, but not β‐catenin. Moreover, it decreased osteocalcin and bone alkaline phosphatase expression, in a dose‐dependent manner. Bone sialoprotein expression was decreased in response to the higher MC supernatant concentration, but the opposite was observed in response to the lower concentration. Osteopontin and type 1 collagen expression were decreased, in a similar manner by all MC supernatant concentrations.ConclusionOur current data allow us to suggest that MC‐derived factors can directly inhibit bone formation‐related markers expression. Further studies are being performed to better understand the mechanisms related to mast cell – bone cell crosstalk.Support or Funding InformationFinancial support and acknowledgments: FAPESP (Grant: #2015/03965‐2 and Grant: #2017/05873‐3)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.