Nanostring technology on Fibrous Dysplasia bone biopsies. A pilot study suggesting different histology-related molecular profiles

Abstract

Identifying the molecular networks that underlie Fibrous Dysplasia (FD) is key to understand the pathogenesis of the disease, to refine current diagnostic approaches and to develop efficacious therapies. In this study, we used the NanoString nCounter Analysis System to investigate the gene signature of a series of nine Formalin Fixed Decalcified and Paraffin-Embedded (FFDPE) bone biopsies from seven FD patients. We analyzed the expression level of 770 genes. Unsupervised clustering analysis demonstrated partitioning into two clusters with distinct patterns of gene expression. Differentially expressed genes included growth factors, components of the Wnt signaling system, interleukins and some of their cognate receptors, ephrin ligands, matrix metalloproteinases, neurotrophins and genes encoding components of the cAMP-dependent protein kinase. Interestingly, two tissue samples obtained from the same skeletal site of one patient one year apart failed to segregate in the same cluster. Retrospective histological review of the samples revealed different microscopic aspects in the two groups. The results of our pilot study suggest that the genetic signature of FD is heterogeneous and varies according to the histology and, likely, to the age of the lesion. In addition, they show that the Nanostring technology is a valuable tool for molecular translational studies on archival FFDPE material in FD and other rare bone diseases.

Transcriptomic signature and pro-osteoclastic secreted factors of abnormal bone marrow stromal cells in fibrous dysplasia

“…Although our understanding of the lesional cell population dynamics in FD pathogenesis has advanced significantly in recent years, we still have limited knowledge of the transcriptional effects of hyperactive Gα s and cAMP excess in BMSCs, the underlying cause of the lesions. Previous differential gene expression analyses of bulk FD tissue fail to capture FD BMSCs-specific transcriptomic changes (9,14,15) , and attempts seeking to characterize the effects of Gα s R201C expression in human BMSCs by lentiviral transduction of wildtype BMSCs involved high cell manipulation, limiting the model’s validity to emulate the transcriptomic profile of FD BMSCs (17,18) . In the present study, to analyze the cell-intrinsic effects of GNAS gain-of-function in FD BMSCs unaffected by the lesional microenvironment, we performed a comprehensive exploration of the transcriptome and secretome of FD BMSCs cultured in the absence of other cell types.…”

“…On the other hand, we and others carried out efforts to characterize the transcriptomic profile of FD, but our approach involved the isolation of mRNA in bulk FD tissue, which does not allow for determination of the contribution of BMSCs bearing gain-of-function variants of GNAS to the differential expression genes identified (9,(14)(15)(16) . In addition, previous attempts were done to characterize the transcriptomic changes caused by GNAS activation in FD BMSCs using cultured human BMSCs transduced with Gαs R201C , though the cellular manipulation of this technique may limit its reliability to capture the transcriptomic effects of Gαs gain of function in lesional BMSCs (17,18) .…”

Transcriptomic Signature and Pro-Osteoclastic Secreted Factors of Abnormal Bone-Marrow Stromal Cells in Fibrous Dysplasia

“…Although our understanding of lesional cell population dynamics in FD pathogenesis has advanced significantly in recent years, we still have limited knowledge of the transcriptional effects of hyperactive Gα s and cAMP excess in BMSCs, the underlying causes of the lesions. Previous differential gene expression analyses of bulk FD tissue fail to capture BMSC-specific transcriptomic changes [11,16,17], and attempts to characterize the effects of Gα s R201C expression in human BMSCs through lentiviral transduction involved high cell manipulation, limiting the models' validity to emulate the transcriptomic profile of FD BMSCs [19,20]. In the present study, we performed a comprehensive exploration of the transcriptome and secretome of FD BMSCs cultured in the absence of other cell types to determine the cell-intrinsic effects of Gα s activation.…”

“…On the other hand, we and others carried out efforts to characterize the transcriptomic profile of FD. Our approach involved isolating bulk mRNA from FD tissue; while informative, this did not allow us to determine how GNAS variants independently affect gene expression in BMSCs [11,[16][17][18]. In addition, previous studies characterized the transcriptomic changes caused by Gα s activation using cultured human BMSCs transduced with Gα s R201C , though the substantial cellular manipulation of this technique may limit its reliability to capture the transcriptomic effects of the Gα s gain of function in lesional BMSCs [19,20].…”

Clinical Value of RANKL, OPG, IL-6 and Sclerostin as Biomarkers for Fibrous Dysplasia/McCune-Albright Syndrome

“…Research on the sclerostin/Wnt pathway in FD/MAS is scarce. Some patients are described to exhibit gene upregulation of components of the Wnt signaling pathway [17,18] and in a mice model of FD, diminished expression of sclerostin was observed by immunohistochemistry, although mRNA expression was comparable to controls [19]. In summary, this study investigated the role of RANKL, OPG, IL-6 and sclerostin as potential biomarkers.…”