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Next-generation sequencing facilitates quantitative analysis of wild-type peritoneal murine mast cells (PMCs) and their comparison to PMC lines

ABSTRACT: Spontaneously developed murine PMC lines were compared to naive murine PMCs to elucidate genomic changes and correlate those to phenotypes observed with respect to mast cell effector functions, characteristics and activation of siganling pathways. Moreover, control PMCs will be used to analyze gender differences.

ORGANISM(S): Mus musculus

PROVIDER: GSE227065 | GEO | 2023/03/14

REPOSITORIES: GEO

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Project description:Investigations of teriparatide (rPTH) as a potential treatment for critical defects have demonstrated the predicted anabolic effects on bone formation, and significant non-anabolic effects on healing via undefined mechanisms. Specifically, studies in murine models of structural allograft healing demonstrated that rPTH treatment increased angiogenesis (vessels <30μm), and decreased arteriogenesis (>30 μm) and mast cell numbers, which lead to decreased fibrosis and accelerated healing. To better understand these non-anabolic effects, we interrogated osteogenesis, vasculogenesis and mast cell accumulation in mice randomized to placebo (saline), rPTH (20µg/kg/2days), or the mast cell inhibitor sodium cromolyn (SC) (24µg/kg/2days), via longitudinal micro-CT and multiphoton laser scanning microscopy (MPLSM), in a critical calvaria defect model. Micro-CT demonstrated that SC significantly increased defect window closure and new bone volume versus placebo (p<0.05), although these effects were not as great as rPTH. Interestingly, both rPTH and SC has similar inhibitory effects on arteriogenesis versus placebo (p<0.05) without affecting total vascular volume. MPLSM time course studies in untreated mice revealed that large numbers of mast cells were detected 1 day post-op (43 +/- 17), peaked at 6 days (76 +/- 6), and were still present in the critical defect at the end of the experiment on day 30 (20 +/- 12). In contrast, angiogenesis was not observed until day 4, and functional vessels were first observed on 6 days, demonstrating that mast cell accumulation precedes vasculogenesis. To confirm a direct role of mast cells on osteogenesis and vasculogenesis, we demonstrated that specific diphtheria toxin-α deletion in Mcpt5-Cre-iDTR mice results in similar affects as SC treatment in WT mice. Collectively, these findings demonstrate that mast cells inhibit bone defect healing by stimulating arteriogenesis associated with fibrotic scaring, and that an efficacious non-anabolic effect of rPTH therapy on bone repair is suppression of arteriogenesis and fibrosis secondary to mast cell inhibition.And then, we need to figure out how rPTH works on mast cells. So we used the co-culture of osteoblast and mast cells, and perform RNAseq, by which we aim to screen out the factors in mast cells that are potential mediators for rPTH effects in bone formation, blood vessel formation, and fibrosis formation.

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Next-generation sequencing facilitates quantitative analysis of wild-type peritoneal murine mast cells (PMCs) and their comparison to PMC lines

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