Adipose-Derived Stem Cells

Adipose-derived stem cells are a population of cells found in adipose tissue that have adipogenic, myogenic, neurogenic, chondrogenic, and osteogenic differentiation potential. Additionally, they possess angiogenic and vasculogenic tendencies.⁸ Human adipose stem cells were first characterized by Zuk et al., in 2001.¹³ In adipose tissue, MSCs originate in perivascular locations and can be found in the stromal vascular fraction (SVF). The regenerative components of SVF include preadipocytes, endothelial progenitor cells, macrophages, monocytes, pericytes, and MSCs.²

Rodbell and Jones pioneered the initial methods to isolate stem cells from adipose tissue in the 1960s.¹⁴ Current methods of adipose stem cell isolation involve tumescent liposuction which produces the minced adipose tissue fragments, depending on the size of the harvesting cannula used. This is followed by cell washing to remove some red blood cells, enzymatic digestion, centrifugation, and further cell wash steps to isolate the SVF.

The regenerative capacity of the SVF is likely a result of the “sum of its parts.” This multi-component cellular result provides numerous pathways and mechanisms for regeneration to occur.² Preadipocytes are the most plentiful component of SVF and share many characteristics and markers of MSCs.¹⁵ Endothelial progenitor cells release VEGF and insulin-like growth factor-1 (IGF-1), which activate angiogenesis.¹⁶ Macrophages contained in SVF have demonstrated immunomodulatory properties via cytokine expression, as well as plastic adherence, multilineage differentiation, and immunosuppression via T-cell modulation.¹⁷ Pericytes produce extracellular matrix components capable of improving cellular adhesion, migration, and cell interaction.¹⁸

Cell surface markers, including clusters of differentiation (CD), are critical for cell-cell and cell-environment interactions and are used to define mixed cell populations. Despite an enormous amount of research, there is little consensus with regard to CD characterization of the SVF. Table 13.1 shows a comparison of SVF-defined cell surface markers.¹⁵¹⁹-²⁵ Table 13.2 provides a comparison of cell surface markers between bone marrow and adipose-derived stem cells.²⁰

It is important to note that, as of this publication, the FDA currently views the standard method of adipose-derived stem cell isolation as manufacturing a drug; therefore this is not permissible in clinical practice in the United States. Bone marrow aspirate concentration (BMAC) using density centrifugation methods is permissible so long as the procedure is carried out the same day.

MSCs in Pain Management

It is well-established that the effectiveness of MSCs is mainly due to their paracrine effects to promote a regenerative localized environment, rather than their direct effect or replacing tissue. For chronic pain and neuroinflammation, data suggest MSCs secrete the following noteworthy factors: transforming growth factor beta, interleukin-10, tumor necrosis factor-stimulated gene-6, hepato- cyte growth factor 1, and matrix metalloproteinases.

Transforming growth factor beta (TGF-(3) controls many functions in the cellular environment, including growth, propagation, differentiation, and apoptosis. TGF-p possesses both powerful immunosuppressive and neuromodulatory properties. It is thought that TGF-(3 may reduce chronic pain by inhibiting the activation and propagation of nerve-injury-induced microglia and astrocytes.²⁶ TGF-p also decreases the secretion of pro-inflammatory cytokines. However, the mechanisms of TGF-P inhibition of neuropathic as well as chronic pain need further elucidation.²⁶-²⁸

Interleukin-10 (IL-10) is known to be a powerful anti-inflammatory cytokine affecting inflammation and immunoregulation. The role of IL-10 remains elusive in pain management, with conflicting study results. A study by Li et al. demonstrated that the effects of MSCs were reversed by TGF-P and IL-10 antibody neutralization.²⁹ These data suggest that IL-10 plays a critical role in reducing pain and inflammation.

Tumor necrosis factor stimulated gene-6 (TSG-6) is an immunomodulatory protein secreted by MSCs. Animal studies using various disease models suggest that MSCs show dependence on TSG-6 for therapeutic effect. TSG-6 may shift cells such as macrophages from a pro-inflammatory state to an anti-inflammatory state. Injection of TSG-6 in mice with ulcerative colitis reduces the severity of colitis and the number of inflammatory cells.³⁰

TABLE 13.1

SVF Cell Surface Markers

Hepatocyte growth factor-1 (HGF-1) is secreted in large amounts by MSCs. HGF-1 plays a role in angiogenesis and possesses the ability to promote cell growth and proliferation, and resist apoptosis. Indeed it has a neuroprotective effect, and in vitro studies illustrate that MSCs secrete HGF-1 which may have a protective effect on excitotoxicity-injured PC12 cells.³¹ HGF-1 has demonstrated protective properties against autoimmune disease in vivo by activating regulatory T-cells, leading to the production of immunosuppressive cytokine IL-10.^{32 33}

Matrix metalloproteinases (MMPs) are enzymes that are capable of degrading extracellular matrix proteins and can process a number of bioactive molecules. MMPs play a major role in neuroinflammation and pain through the division and suppression of extracellular matrix proteins, cytokines, and chemokines.³⁴ Nerve injury-induced changes in the dorsal root ganglion are responsible for neuropathic pain.³⁵ MMP-9, produced by injured dorsal root ganglions, provokes spinal microglia activation and neuropathic pain development due to MMP-9 pathophysiology, including IL-1 beta cleavage and microglia p38 activation in early-phase neuropathic pain. MMP-2 activation

TABLE 13.2

Differences between Bone Marrow and Adipose Tissue

SVF = stromal vascular fraction

BM-MC = bone marrow mononucleated cells

ASC = adipose stem cells

MSC = mesenchymal stem cells

leads to late-phase neuropathic pain via IL-1 beta cleavage and astrocyte ERK activation.³⁵ Tissue inhibitor of metalloproteinases (TIMP) suppress neuropathic pain. TIMP-1 diminishes early-phase neuropathic pain, while TIMP-2 diminishes late-phase neuropathic pain.³⁵