Abstract

Considering the increasing interest in adipose-derived stem cells (ASCs) in regenerative medicine, optimization of methods aimed at isolation, characterization, expansion and evaluation of differentiation potential is critical to ensure (a) the quality of stem cells also in terms of genetic stability; (b) the reproducibility of beneficial effects; and (c) the safety of their use. Numerous studies have been conducted to understand the mechanisms that regulate ASC proliferation, growth and differentiation, however standard protocols about harvesting and processing techniques are not yet defined. It is also important to note that some steps in the procedures of harvesting and/or processing have been reported to affect recovery and/or the physiology of ASCs. Even considering the great opportunity that the ASCs provide for the identification of novel molecular targets for new or old drugs, the definition of homogeneous preparation methods that ensure adequate quality assurance and control, in accordance with current GMPs (good manufacturing practices), is required. Here, we summarize the literature reports to provide a detailed overview of the methodological issues underlying human ASCs isolation, processing, characterization, expansion, differentiation techniques, recalling at the same time their basilar principles, advantages and limits, in particular focusing on how these procedures could affect the ASC quality, functionality and plasticity.

Highlights

  • The viability, yield, proliferative index and stemness of adipose-derived stem cells (ASCs) can be influenced by the type of harvesting procedure

  • With reference to the step of density-gradient centrifugation or spontaneous stratification, in a previous report [8], our group has analyzed the influence of different handling methods on stromal vascular fraction (SVF) cell and adipocyte yield and viability

  • The results show that the lipoaspirates enriched with enzymatic methods have a significantly higher number of cells with respect to non-enriched lipoaspirate or enriched with a mechanical procedure, while no difference has been reported in stemness marker expression

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Summary

Sources of ASCs and Isolation Procedures

The viability, yield, proliferative index and stemness of ASCs can be influenced by the type of harvesting procedure. With reference to the step of density-gradient centrifugation or spontaneous stratification, in a previous report [8], our group has analyzed the influence of different handling methods on SVF cell and adipocyte yield and viability. Clinical devices for SVF isolating and obtaining enriched fat grafts in stem cells are available with different performance primarily in terms of cell yield, effectiveness and safety. The advantages derived by a simple device able to fat harvest with minimal manipulation through soft mechanical action, are described [24] With this technology, cell components are preserved in the SVF, the enzyme absence maintains unaltered the exosome content, and the micro-fragmented tissue promotes the healing process of receiving tissue. Responding to clinical requirements, the non-enzymatic methods require large amounts of lipoaspirate and have lower efficiency in cell recovery when compared to enzymatic methods generally used in a research context [25]

Culture and Expansion of ASCs
ASC Differentiation Potential
Findings
Influence of Age and Obesity on ASC Isolation and Functions
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