Skeletal site-related variation in human trabecular bone transcriptome and signaling

Abstract

Background: The skeletal site-specific influence of multiple genes on bone morphology is recognised, but the question as to how these influences may be exerted at the molecular and cellular level has not been explored.

Methodology: To address this question, we have compared global gene expression profiles of human trabecular bone from two different skeletal sites that experience vastly different degrees of mechanical loading, namely biopsies from iliac crest and lumbar spinal lamina.

Principal findings: In the lumbar spine, compared to the iliac crest, the majority of the differentially expressed genes showed significantly increased levels of expression; 3406 transcripts were up- whilst 838 were down-regulated. Interestingly, all gene transcripts that have been recently demonstrated to be markers of osteocyte, as well as osteoblast and osteoclast-related genes, were markedly up-regulated in the spine. The transcriptome data is consistent with osteocyte numbers being almost identical at the two anatomical sites, but suggesting a relatively low osteocyte functional activity in the iliac crest. Similarly, osteoblast and osteoclast expression data suggested similar numbers of the cells, but presented with higher activity in the spine than iliac crest. This analysis has also led to the identification of expression of a number of transcripts, previously known and novel, which to our knowledge have never earlier been associated with bone growth and remodelling.

Conclusions and significance: This study provides molecular evidence explaining anatomical and micro-architectural site-related changes in bone cell function, which is predominantly attributable to alteration in cell transcriptional activity. A number of novel signaling molecules in critical pathways, which have been hitherto not known to be expressed in bone cells of mature vertebrates, were identified.

Figure 1. TNF receptor signaling in trabecular bone.

Direct interaction network models were constructed by Pathway Architect analysis of the differentially expressed transcript in the lumbar spine and iliac crest trabecular bone. Up-regulated transcripts are indicated red whilst the down-regulated are shown in green. Arrows link interacting genes and positive (+) and negative (−) associations are marked respectively. Green boxes denote regulation, blue boxes binding and orange circles indicating phosphorylation.

Figure 2. BMP signaling in trabecular bone.

In this interaction network model, constructed by Pathway Architect analysis software of differential expressed genes between iliac crest and the lumbar spine, up-regulated transcripts are indicated red whilst the down-regulated are shown in green. Arrows link interacting genes and positive (+) and negative (−) associations are marked respectively. Green boxes denote regulation, blue boxes binding and orange circles indicating phosphorylation.

Figure 3. Proteogylcan Syndecan-mediated signaling in trabecular bone.

Direct interaction network models constructed by Pathway Architect analysis of the differentially expressed transcript in the lumbar spine and iliac crest trabecular bone. Up-regulated transcripts are indicated red whilst the down-regulated are shown in green. Arrows link interacting genes and positive (+) and negative (−) associations are marked respectively. Green boxes denote regulation, blue boxes binding and orange circles indicating phosphorylation.

Figure 4. Network enriched for genes involved in skeletal system.

Molecules are represented as nodes, and the biological relationship between two nodes is represented as an edge (line). The intensity of the node color indicates the degree of down- (red) or up- (green) regulation where lumbar spine is compared with iliac crest. Nodes are displayed using various shapes that represent the functional class of the gene product. Direct relationships are shown in solid arrows, indirect relationships in dashed arrows. Genes with no colour are added to the network by Ingenuity Pathway Analysis as part of the network generation algorithm.

Figure 5. Role of osteocytes in mechanotransduction in trabecular bone.

The increased mechanical stress is detected by mechanosensors, a function which is primarily performed by the osteocytes (∼90% of the total cell number). The increased mechanical stress is mechano-transduced into intracellular biochemical signals by osteocytes and results in increased transcriptional activity of range of genes (SOST, MEPE and DMP1). Osteocytes also transmute mechanical stress into intercellular biochemical signals to modulate the respective activities of the osteoclasts and osteoblasts.