Acute transcriptional up-regulation specific to osteoblasts/osteoclasts in medaka fish immediately after exposure to microgravity.

Masahiro Chatani,Akira Kudo,Yoshiro Takano,Hiromi Suzuki,Kazuhiro Takeyama,Koji Kubota,Hiroya Morimoto,Naoki Tanigawa,Oleg Gusev,Akiko Mantoku,Fumiaki Tanigaki,Takehiko Itoh,Vladimir Sychev,Satoko Uchida,Masaki Shirakawa

doi:10.1038/srep39545

Abstract

Bone loss is a serious problem in spaceflight; however, the initial action of microgravity has not been identified. To examine this action, we performed live-imaging of animals during a space mission followed by transcriptome analysis using medaka transgenic lines expressing osteoblast and osteoclast-specific promoter-driven GFP and DsRed. In live-imaging for osteoblasts, the intensity of osterix- or osteocalcin-DsRed fluorescence in pharyngeal bones was significantly enhanced 1 day after launch; and this enhancement continued for 8 or 5 days. In osteoclasts, the signals of TRAP-GFP and MMP9-DsRed were highly increased at days 4 and 6 after launch in flight. HiSeq from pharyngeal bones of juvenile fish at day 2 after launch showed up-regulation of 2 osteoblast- and 3 osteoclast- related genes. Gene ontology analysis for the whole-body showed that transcription of genes in the category “nucleus” was significantly enhanced; particularly, transcription-regulators were more up-regulated at day 2 than at day 6. Lastly, we identified 5 genes, c-fos, jun-B-like, pai-1, ddit4 and tsc22d3, which were up-regulated commonly in the whole-body at days 2 and 6, and in the pharyngeal bone at day 2. Our results suggested that exposure to microgravity immediately induced dynamic alteration of gene expression levels in osteoblasts and osteoclasts.

Highlights

Of bone quality revealed a loss of bone in short-duration spaceflight for 20 days[9]
The experimental schedule for live-imaging of the 4 double transgenic lines, osterix-DsRed/the former and acp5 (TRAP)-GFP, osteocalcin-DsRed/TRAP-GFP, MMP9-DsRed/RANKL-GFP, and cox2-GFP/TRAPDsRed during 8 days in flight and on the ground is shown in Supplementary Table S1
We focused on the pharyngeal bone region, in which bone turnover is high and sensitive to microgravity[21], and observed ground and flight samples at high magnification with a 20x lens (Fig. 3a,b) to examine the details of fluorescent signals in osteoblasts

Summary

Introduction

Of bone quality revealed a loss of bone in short-duration spaceflight for 20 days[9]. To examine the early effects of microgravity on bone cells, we embedded transgenic medaka larvae in a gel for a live-imaging study in space in 2014, and observed signals by fluorescence microscopy at the ISS via remote operation from Tsukuba Space Center. For this experiment, we utilized 4 different double medaka transgenic lines and, in particular, investigated up-regulation of fluorescent signals of osteoblasts and osteoclasts in these double transgenic lines as an important way to study osteoblast-osteoclast interaction under microgravity. Our results about live-imaging and transcriptome analysis may prompt the establishment of a new field in gravitational biology

Methods

Results

Conclusion