RNA Extraction from Cartilage: Issues, Methods, Tips

Abstract

The increase in degenerative diseases involving articular cartilage has pushed research to focus on their pathogenesis and treatment, exploiting increasingly complex techniques. Gene expression analyses from tissue are representative of the in vivo situation, but the protocols to be applied to obtain a reliable analysis are not completely cleared through customs. Thus, RNA extraction from fresh samples and specifically from musculoskeletal tissue such as cartilage is still a challenging issue. The aim of the review is to provide an overview of the techniques described in the literature for RNA extraction, highlighting limits and possibilities. The research retrieved 65 papers suitable for the purposes. The results highlighted the great difficulty in comparing the different studies, both for the sources of tissue used and for the techniques employed, as well as the details about protocols. Few papers compared different RNA extraction methods or homogenization techniques; the case study reported by authors about RNA extraction from sheep cartilage has not found an analog in the literature, confirming the existence of a relevant blank on studies about RNA extraction from cartilage tissue. However, the state of the art depicted can be used as a starting point to improve and expand studies on this topic.

Decellularized biological matrices for the repair of rotator cuff lesions: a systematic review of preclinical in vivo studies

“…Despite the requirement for sample harvest, these analyses allow for the determination of a variety of aspects related to the biological response of tissues to decellularized patches ( Maglio et al, 2020 ), such as the formation of new fibrocartilage tissue at the insertion site, enthesis maturation, implant degradation, organization, and orientation of newly deposited collagen fibers, as well as the presence of inflammatory infiltrates or the formation of new blood vessels. In addition, the evaluation of the expression levels of genes such as COL1 , COL2 , COL3 , ACAN , and TNC after RNA extraction from tissues may provide additional useful information on RC healing before and after treatment ( Yuan et al, 2022a ; Yuan et al, 2022b ; Pagani et al, 2023 ). No gait analysis is performed, except for a paper assessing the degree of lameness and functionality of the forelimbs ( Smith et al, 2020 ); this test is performed for locomotion analysis and allows longitudinal repeated measurements, sequentially or at different time points, without the need to restrain the animal, thus ethically complying with the principles of animal reduction and refinement.…”

RNA Extraction from Cumulus Cells for Gene Analysis: The Current Strategy and Review of Literature

“…Inefficient homogenization may cause the release of endogenous ribonucleases, leading to RNA degradation [40]. Subsequently, these enzymatic activities reduce RNA integrity and compromise downstream applications, such as gene expression analysis [16,41]. Thus, identifying the best homogenizing method for different cell types is crucial.…”

Potential Therapeutic Targets for Leishmaniasis: Modulating Cytokine Production and ECM Organization

“…It should be noted that DEPC is toxic, and all work must be done under a hood with gloves and a mask. Standard laboratory kits and materials are RNase-free and do not require treatment (5).…”