Protocol for parallel proteomic and metabolomic analysis of mouse intervertebral disc tissues

Abstract

The comprehensiveness of data collected by "omics" modalities has demonstrated the ability to drastically transform our understanding of the molecular mechanisms of chronic, complex diseases such as musculoskeletal pathologies, how biomarkers are identified, and how therapeutic targets are developed. Standardization of protocols will enable comparisons between findings reported by multiple research groups and move the application of these technologies forward. Herein, we describe a protocol for parallel proteomic and metabolomic analysis of mouse intervertebral disc (IVD) tissues, building from the combined expertise of our collaborative team. This protocol covers dissection of murine IVD tissues, sample isolation, and data analysis for both proteomics and metabolomics applications. The protocol presented below was optimized to maximize the utility of a mouse model for "omics" applications, accounting for the challenges associated with the small starting quantity of sample due to small tissue size as well as the extracellular matrix-rich nature of the tissue.

Colonization of intervertebral discs byCutibacterium acnesin patients with low back pain: protocol for an analytical study with microbiological, phenotypic, genotypic, and multiomic techniques

“…(30) These techniques have already been used successfully to define molecular signatures in other clinical settings, (31)(32)(33)(34) especially in chronic pain, (35)(36)(37)(38)(39)(40) and can also provide integrative information on cell function at the molecular level about the role of C. acnes in LDD and LBP. (41)(42)(43)(44)…”

Improving reproducibility in spine research

“…These methods will be useful for the rigorous analysis of IVD cells and will aid in the understanding of intervertebral disc biology. While these methods were designed for mouse models they can also be adapted for other species within the community. Veras et al 3 reports a collaborative development of a consensus methodology for the parallel proteomic and metabolomic profiling of mouse intervertebral disc. The methodology covered describes the dissection, extraction of cells specifically for omics applications and data analysis, providing considerations for the small starting materials and successful extraction from the problematic ECM rich tissues of the IVD.…”

Colonization of intervertebral discs by Cutibacterium acnes in patients with low back pain: Protocol for an analytical study with microbiological, phenotypic, genotypic, and multiomic techniques

“…These techniques have already been used successfully to define molecular signatures in other clinical settings [ 31 – 34 ], especially in chronic pain [ 35 – 40 ], and can also provide integrative information on cell function at the molecular level about the role of C . acnes in LDD and LBP [ 41 – 44 ].…”

“…In this context, the combined use of proteomic and metabolomic techniques to analyze LIVD colonized by C. acnes would be more appropriate in order to assess the subclinical consequences of the colonization by this pathogen [30]. These techniques have already been used successfully to define molecular signatures in other clinical settings [31][32][33][34], especially in chronic pain [35][36][37][38][39][40], and can also provide integrative information on cell function at the molecular level about the role of C. acnes in LDD and LBP [41][42][43][44].…”