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Identifying deer antler proliferation and mineralization genes using comparative RNA-seq

ABSTRACT: Purpose: The goal of this study is to compare (RNA-seq) transcriptomes of in vitro cultured human bone marrow-derived mesenchymal stem cells (hMSCs) and fallow deer antler-derived skeletal progenitors (FD RM Cells) under multiple conditions to identify candidate proliferation and mineralization genes responsible for fast antler regeneration Methods: hMSCs and FD RM Cells were cultured in vitro under 1) serum-free (0% serum) or serum (10% serum) conditions for 2.5 days or 2) Control (0 ng/mL BMP-2 and 0 nM dexamethasone) and osteogenic (100 ng/mL BMP-2 and 100 nM dexamethasone) media for 24 days. mRNA profiles were generated by deep sequencing, in duplicate, using Illumina HiSeq 2000. The sequence reads were analyzed at the transcript isoform level STARS followed by Cufflinks. Validation for genes of interest was performed using immunofluorescence staining. Results: Comparison of human and fallow deer skeletal progenitor datasets yielded proliferation and mineralization gene candidates Conclusions: Our study represents the first detailed analysis of human and fallow deer transcriptomes of skeletal progenitor cells under proliferation and mineralization conditions, with biologic replicates, generated by RNA-seq technology. Our in vitro comparative approach circumvent some of the logistical and technical challenges in identifying candidate proliferation and mineralization genes responsible for rapid deer antler regeneration. We conclude that in vitro comparison of RNA-seq based transcriptomes identified candidate proliferaiton and mineralization genes to advance bone biology and holds promise to rapidly regenerate large bone volumes for regenerative medicine. The comparative approach utilized here can be adapted for almost any tissue to study a specific phenomenon of interest.

ORGANISM(S): Homo sapiens Dama dama

PROVIDER: GSE89796 | GEO | 2018/11/16

REPOSITORIES: GEO

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Project description:Rib bone growth in red deer stags - Abstract: In 'The Bone and Joint Decade' interest is focused on genetic factors causing bone disorders. Osteoporosis, attacking 10% of the population worldwide, is the most common metabolic bone disease, which is mimiced by several ovarectomised or genetically modified 'cascadeur' animal species, but none of them is able to remedy its pathologically porous bone tissue. Regeneration in skeletal elements is the curiosity of our newly investigated osteoporosis animal model, red deer (Cervus elaphus). The cyclic physiological osteoporosis in red deer stag is a consequence of the annual antler cycle. This phenomenon raises the possibility to explore new genes involved in regulating bone mineral density (BMD) and recovery of bone resorption on the basis of comparative genomics between deer and human. Here we compared the gene expression activities of osteoporotic and regenerating flying rib bone samples versus late autumn dwell control in red deer by heterologous microarray hybridization. Identified genes were tested on human femoral bone tissue from postmenopausal osteoporotic and non-osteoporotic patients. Expression data were evaluated by Principal Components Analysis and Discriminant Analysis. Keywords: Gene Expression experiment Approximately 2-3 g flying rib bone pieces in the entire cross section of bony rib were surgically removed from 3 anaesthetized [SBH-Ketamine (2.5 mg/kg live weight) combined with Xylazine (0.2 mg/kg live weight) i.m. injection] 6, 7 and 8 year old Cervus elaphus stags. (Cast antler pairs weighed 7-8 kg for each animal.) Removed rib pieces were extensively washed in PBS for eliminating blood and marrow contamination, than immediately frozen in liquid nitrogen. The time of tissue collections were (i) within the period of the active mineralization of antler, at the beginning of June when skeletal osteoporosis takes place, (ii) in the fitness improvement period with velvet shedding in late July, that is the 'regenerating time' and (iii) in the period of late autumn dwell at the end of November when in the skeleton the mineral mobilization and deposition are dynamically equilibrated (BMD is in steady state). Each comparison performed on Platforms GPL4052 and GPL5352.

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Identifying deer antler proliferation and mineralization genes using comparative RNA-seq

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