Medicinal signaling cells niche in stromal vascular fraction from lipoaspirate and microfragmented counterpart

AimTo expand our previous findings by increasing the number of patients in a study characterizing medicinal signaling cells (MSC) of stromal vascular fraction from lipoaspirate (SVF-LA) and from microfragmented lipoaspirate (SVF-MLA) applied for the treatment of osteoarthritis (OA).MethodsTwenty OA patients, including 8 new patients, acquiring autologous microfragmented adipose tissue were enrolled. In-parallel immunophenotyping of SVF-LA and SVF-MLA was performed. The samples were incubated in a DuraClone SC prototype tube targeting the CD31, CD34, CD45, CD73, CD90, CD105, and CD146 surface markers, stained with the DRAQ7 cell nuclear dye and Live/Dead Yellow Fixable Stain, and analyzed by flow cytometry.ResultsThe population phenotypes in SVF-LA and SVF-MLA samples included CD31⁺CD34⁺CD73^±CD90^±CD105^±CD146^± endothelial progenitors (EP), CD31⁺CD34^-CD73^±CD90^±CD105^-CD146^± mature endothelial cells, CD31^-CD34^-CD73^±CD90⁺CD105^-CD146⁺ pericytes, CD31^-CD34⁺CD73^±CD90⁺CD105^-CD146⁺ transitional pericytes, and CD31^-CD34⁺CD73^highCD90⁺CD105^-CD146^- supra-adventitial-adipose stromal cells. Compared with the autologous SVF-LA samples, the prevailing cell type in SVF-MLA were EP, which outnumbered leukocytes and supra-adventitial-adipose stromal cells (SA-ASC). The ratio of progenitor cells in SVF-MLA samples differed between female and male patients, showing a higher EP-pericyte and pericyte-SA-ASC ratio in men.ConclusionOur results, hallmarked by EP-enriched anti-inflammatory features and indicating a possible sex-specific impact, contribute to defining the cellular composition of the clinically applied MSC serving as a regenerative cell therapy in OA.

Mesenchymal stromal/stem cells (MSC) are well known for their capability of differentiating into mesenchymal cell types. Caplan has recently suggested that they are renamed into medicinal signaling cells with the same acronym (1). The name ought to be adapted as our knowledge of the biological concept has expanded: during tissue regeneration, MSC perform their function via signaling rather by than differentiating – as they do under cell culture conditions (2-4). MSC comprise a heterogeneous population of stromal and stem cells with additional immunosuppressive and trophic properties, which upon injury or inflammation modulate the local environment by secreting numerous anti-apoptotic, anti-scaring, angiogenic, and mitotic factors (1,5). This paradigm for tissue regeneration has been brought up by the studies of MSC in osteoarthritis (OA), the most common joint disorder (6). MSC from adipose tissue are now widely investigated as a novel therapeutic method in the treatment of OA (7).Although 20 years have passed since the first characterization of the multipotent MSC from adult adipose tissue, ie lipoaspirate (3), a complete characterization of this heterogeneous cell type remains elusive. This tremendous discovery opened up unprecedented possibilities in clinical application, however, undefined cellular heterogeneity and non-standardized protocols represent the main obstacle to the MSC usage in regenerative medicine. Together with other cell types, MSC are found in the stromal vascular fraction (SVF), which is obtained from adipose tissue upon collagenase treatment. Besides being a fruitful source of MSC, adipose tissue seems to potentiate the MSC-mediated tissue regeneration if it previously undergoes microfragmentation. The secretome of microfragmented adipose tissue more abundantly harbors cytokines and angiogenic factors, accompanied by immunomodulation, angiogenesis, and tissue reparation benefits (8). In this new light of paracrine activity of microfragmented adipose tissue, the Lipogems® device brings innovative technology for processing autologous adipose tissue, producing small intact clusters of perivascular microfragments with a high therapeutic potential (9,10). In a prospective non-randomized study, an intra-articular injection of such a product led to a successful outcome, as revealed by an increased glycosaminoglycan content in the hyaline cartilage of the knee joint (11,12). Several studies of knee OA treatment demonstrated the efficacy of adipose MSC in tissue repair, and even a low-dose MSC application yielded significant functional improvement with pain relief (13-15).Although clinical implementation of microfragmented adipose tissue has brought an impressive step forward in orthopedics, standardization of clinical application requires a better understanding of MSC heterogeneity and the cellular subset characterization. The flow cytometry analysis of human lipoaspirate has shown that a heterogeneous SVF mixture contains endothelial progenitor (EP) cells (CD31⁺CD34⁺CD146⁺), endothelial mature (EM) cells (CD31⁺CD34^-CD146^±), pericytes (CD31^-CD34^-CD146⁺), supra-adventitial-adipose stromal cells (SA-ASC) (CD31^-CD34⁺CD146^-), and transitional pericytes (TP) (CD31^-CD34⁺CD146⁺), with differential expression of the CD73, CD90, and CD105 mesenchymal markers (16,17). The aim of this study was to expand the number of patients from our previous immunophenotyping analysis of SVF from lipoaspirate (SVF-LA) or microfragmented lipoaspirate (SVF-MLA) by means of polychromatic flow cytometry (18). Since SVF-MLA is used therapeutically in OA patients, the results contribute to the biological understanding of the cartilage regeneration.