Adipose-derived Stem Cells (ASCs)
By boosting the body's natural regeneration ability, regenerative medicine holds tremendous promise for repairing damaged tissues and organs and restoring function. Tissue engineering, medicine, and molecular biology are some of the techniques and specialties used in this interdisciplinary area to replace, engineer, or regenerate cells, tissues, or organs with the goal of restoring or regaining normal function.
In this regard, numerous cell types, including adult mesenchymal stem cells, have been examined for their applicability in regenerative medicine. The ability of MSCs to differentiate into cells of mesenchymal origin such as osteoblasts, adipocytes, myocytes, and chondrocytes led to an interest in their potential for future cell-based therapies. Immune compatible stem cells produced from differentiated tissues, unlike embryonic stem cells, are not vulnerable to ethical concerns. MSCs are often extracted from the bone marrow and constitute an appealing cell source for regenerative medicine due to their extensive availability.
Bone marrow-derived MSCs, on the other hand, are unsuitable for clinical usage due to the highly invasive aspiration method required and the decline in both their proliferation and differentiation capability as they age. Zuk et al. introduced a multipotent, undifferentiated, self-renewing progenitor cell population obtained from adipose tissue that is morphologically and phenotypically comparable to MSCs in the early twenty-first century in seeking an alternative stem cell source. According to the Internal Fat Applied Technology Society, these so-called adipose-derived stem cells (ASCs) have a differentiation capacity similar to MSCs and express unique stem cell markers. Importantly, the ASCs' superiority as an alternative clinical cell source is highlighted by the ease of repeated access to subcutaneous adipose tissue via a minimally invasive method, the simple isolation procedure, and a stem cell quality and proliferation capacity that does not decline with age of the patient.
Isolation and Characterization
Mature adipocytes and a heterogeneous stromal vascular fraction (composed of fibroblasts, endothelial cells, pre-adipocytes, vascular smooth muscle cells, lymphocytes, monocytes, and ASCs) make up the bulk of subcutaneous adipose tissue. Collagenase digestion followed by centrifugal density gradient separation is the most extensively used method for isolating ASCs from fat tissue. ASCs have a spindle-shaped morphology in vitro and lack the intracellular lipid droplets that adipocytes have. Monolayer cultures on standard tissue dishes with a basal medium containing 10% fetal bovine serum are frequently used to grow isolated ASCs. In terms of clinical translation, however, the in vitro proliferation of ASCs must adhere to good manufacturing practice standards. Xenogeneic components for cell cultivation must be avoided in this scenario. Human platelet lysate, which is superior in terms of proliferation speed and allogenicity, has recently been discovered to be a promising alternative.
Therapeutic Application of Adipose-derived Stem Cells
Because of their potential to differentiate into a range of cell lineages, stem cells, including ASCs, have emerged as a significant component of regenerative medicine therapy. Furthermore, their ability to secrete a wide range of cytokines, chemokines, and growth factors in a paracrine manner makes them extremely clinically interesting. The anti-apoptotic, anti-inflammatory, proangiogenic, immunomodulatory, and anti-scarring actions that have been documented for ASCs are of specific importance, making these cells ideal candidates for cellular treatment in regenerative medicine.
On the US National Institutes of Health website, there are roughly 130 active clinical studies exploring the potential of ASCs. Soft-tissue regeneration, skeletal tissue repair, ischemic injuries, myocardial infarction, and immunological illnesses (including lupus, arthritis, Crohn's disease, multiple sclerosis, diabetes mellitus, and graft-versus-host disease) are all included in these clinical studies. Intervertebral disc degeneration and pulmonary disease, to name a few, are other therapeutic targets being investigated in clinical studies.
Mesodermal Potential of Adipose-derived Stem Cells
Because of their origins, ASCs were initially used in regenerative medicine for mesodermal regeneration, with an emphasis on their potential for osteogenic, adipogenic, chondrogenic, and subsequently cardiovascular applications
Cartilage and Intervertebral Disc Regeneration
Skeletal tissue is being increasingly impaired as the population ages. As a result, musculoskeletal problems are now a leading cause of disability and morbidity around the world, resulting in massive costs for health and social care systems. In geriatrics, chronic and inflammatory disorders of the joints and spine, such as osteoarthritis and low back pain caused by intervertebral disc degeneration, are major sources of disabilities. By interacting with local chondrocytes or cartilage explants in cartilage defects, transplanted ASCs play a significant role in the success of cellular therapies for cartilage repair.
However, the mechanism underlying ASCs' paracrine action on chondrocytes is still unknown. In vivo healing effects are achieved when mixed ASCs and chondrocytes are implanted into cartilage lesions. After co-culture, the BMP family members (BMP-2, BMP-4, and BMP-5) are down-regulated, while VEGF B, HIF-1, FGF-2, and PDGF are dramatically reduced. These findings imply that interaction between ASCs and chondrocytes could help with cartilage repair and regeneration, as well as cartilage tissue engineering.
Fat Regeneration
The first indications for fat transplantation were the repair of cosmetic deformities and breast reconstruction following mastectomy. Because breast cancer is still the most common malignancy among women, innovative approaches for postsurgical breast reconstruction are continually being developed. Enrichment with autologous SVF, platelet-derived growth factors, hormones, and/or insulin is used in these modern approaches to improve autologous fat grafting. The presence of ASCs in the SVF is likely to be a factor in the reported improvement in fat graft viability with these procedures. It is also known that fibroblast growth factor-2 (FGF-2) increases ASCs proliferation and hepatocyte growth factor (HGF) secretion through a c-Jun N-terminal kinase (JNK) signaling pathway in the process of adipose tissue regeneration following injury, among other injury-associated growth factors.
Although breast reconstruction has been the most common clinical objective for ASCs therapies, the field of plastic and reconstructive surgery now has a wide range of uses. The secretive qualities of ASCs, which allow them to create cytokines, chemokines, and growth factors, may help to develop a successful and novel anti-scarring therapy. Hypertrophic scars develop after an injury as a result of aberrant extracellular matrix (collagen) deposition, remodeling, or inflammation. Although a range of therapies for the treatment of hypertrophic scars is now available (e.g., excision, laser, IFN injection, and so on), none of them has been proven to be completely efficient in preventing excessive scar tissue formation or regeneration of healthy dermal tissue. The ability of ADSCs to prevent hypertrophic scar formation by secreting anti-fibrosis cytokines and lowering -SMA and collagen type I gene expression was proven in a rabbit ear hypertrophic scar model by injecting ASCs in the lesion locally.
Bone Regeneration
The regenerative efficacy of ASCs transplanted from rats, rabbits, and humans on bone repair in a range of defect systems has been studied in several of researches. For example, a painful hip disease caused by avascular necrosis of the femoral head frequently progresses to osteoarthritis and the need for total hip replacement. Autologous ASCs were injected directly into the femoral head in small animal trials. The ASCs improved the osteogenesis and microstructure of vascular deprivation-induced osteonecrotic tissue two months after surgery. In a case study, a mix of autologous ASCs and spongy cells from the iliac crest were employed to rebuild a young girl's calvarial defects following a serious head injury. While a chronic infection following reconstructive surgery resulted in an unstable skull with visible bone abnormalities, new bone development and nearly perfect calvarial continuity were found 3 months after cell injection.
Cardiovascular and Myocardial Regeneration
The Molecular analysis of ASCs not only revealed their osteogenic, adipogenic, and chondrogenic development in vitro but also suggests that they have the ability to expand the mesodermal lineage through myogenic proliferation. Cell therapies for cardiovascular and cardiac tissue regeneration have attracted a lot of attention in recent years. A wide range of cell types has been demonstrated to be advantageous, particularly in the regeneration of the ischemic myocardium. By lowering cardiomyocyte apoptosis or activating cardiac stem cells to enhance cardio-myogenesis, paracrine factors can improve heart function. The therapeutic potential of ASCs in the setting of chronic heart failure or acute myocardial infarction has been investigated by injecting them intracoronary. The ASCs engrafted in the infarct region 4 weeks after cell transplantation and improved cardiac function, perfusion, and remodeling after acute myocardial infarction.
Furthermore, ASCs have attracted attention as a source of interstitial cells to populate heart valve structures and have been studied for use in cardiovascular tissue engineering. It was shown that ASCs may create matrix components like collagen and elastin, as well as secrete matrix-enhancing or -degrading chemicals, to replicate the key tissue structures essential for correct valve performance. ASCs can differentiate into cells with phenotypic and functional characteristics of endothelial cells, which is important for the anti-thrombogenicity of heart valve constructions.
Ectodermal Potential of ASCs
When cultured in the presence of the differentiation agents valproic acid, butylated hydroxy-anisole, insulin, and hydrocortisone, ASCs have been shown to display distinct markers of both the neuronal (NSE, nestin, MAP2, -tubulin III) and glial lineages (GFAP, NG2, p75 NGF receptor). ASCs' neuronal differentiation capacity could be useful in the cellular therapy of neuronal disorders like stroke or Parkinson's disease. Stroke is caused by cerebral ischemia, which sets off a series of physiological and biochemical events.
At this time, there is no effective treatment for stroke. Stem cell therapies, on the other hand, have the potential to reverse the effects of stroke. The potential of ASCs to differentiate into neuron-like cells, as well as other recent findings indicating that human ASCs protect endogenous neuron survival, suggest that this cell source has a high therapeutic benefit for stroke therapy. Direct cell replacement, angiogenesis, increased immunosuppression, and an improvement in the survivability of endogenous neurons are all known to help with stroke symptoms. More study is needed, however, to improve the efficacy of transplanted ASCs as a treatment for stroke and other neurological disorders.
Hepatic Regeneration
Despite the fact that the molecular pathways behind hepatic differentiation are yet unknown, ASCs treated with HGF, oncostatin M, and dimethyl sulfoxide (DMSO) have the ability to form a hepatocyte-like phenotype that expresses albumin and -fetoprotein. These hepatocyte-like cells also have the ability to absorb low-density lipoprotein and generate urea. In addition, when ASCs were transplanted into a CCl4-injured SCID animal model, they were able to develop into hepatocytes and express albumin in vivo.
Pancreatic Regeneration
Aside from ASCs' ability to differentiate into the hepatic lineage, in vitro endodermal differentiation into the pancreatic lineage has also been demonstrated. H human DSCs not only express pancreatic markers such as PDX1, CK19, IPF-1, and nestin after induction with regenerating pancreatic extract or co-culture with pancreatic adult stem cells in vitro, but also produce the pancreatic hormones insulin, somatostatin, and glucagon. ADSCs have the ability to develop into pancreatic cell lineages phenotypically in response to regenerating pancreas extract or by co-culture with pancreatic adult stem cells.
Cryopreservation of ASCs
The preservation and banking of ASCs are critical for clinical practice and logistics in the future use of autologous cells based on patient hope. To do this, the cells must be cryopreserved without losing their ability to proliferate and differentiate, as well as their functionality. In the meanwhile, new regimens for clinic use have been devised. The decrease of the cytotoxic cryoprotectant DMSO, as well as a xenofree and chemically described technique, are the major goals. Adipose tissue cryopreservation for more than two years has also been demonstrated.
Methods to Improve the Therapeutic Effect of ASCs
ASCs' ability to secrete large amounts of particular growth factors, cytokines, and other paracrine substances in their target environment can be used for therapeutic regenerative applications requiring regulated drug release. Efforts are currently being made to elucidate, augment, and utilize stem cell paracrine processes for tissue regeneration in order to support this hypothesis. For example, a number of various techniques focusing on genetic alteration and in vitro preconditioning of ASCs have been investigated in an attempt to increase the amount of released trophic factors upon cell distribution in vivo.
This concept's potential for osteochondral regeneration has already been established. The osteogenic characteristics of ASCs were boosted in an in vitro investigation by transducing these cells with an adenovirus, which resulted in increased synthesis of bone morphogenetic proteins such as BMP2 and BMP4. Similarly, ASCs were engineered to secrete significant levels of VEGF in order to increase their angiogenic properties in ischemic tissue. Surprisingly, the regeneration activity is dependent not only on soluble substances produced in vivo by ASCs but also on the activation of the recipient's own secretomes.
ASCs as All-Rounder of Regenerative Medicine
ASCs have good prerequisites for a wide range of therapeutic applications in regenerative medicine due to their tri-germ lineage differentiation capacity. Furthermore, ASCs' immunosuppressive qualities make them an appealing and clinically relevant cell population for the treatment of a variety of immune-mediated disorders, such as graft-versus-host disease, Crohn's disease, and rheumatoid arthritis.
On the other hand, vascularization is one of the most important prerequisites for tissue healing, aside from tissue restoration. ASCs have been shown to differentiate easily to endothelial cells and to create rapid and simple vessel-like structures in Matrigel substrates with presumed endothelial function, suggesting that they are major regulators of new blood vessel formation. Furthermore, vessel formation has been reported after injection of ASCs alone or in conjunction with other cell types in many studies, including myocardial infarction treatment, epithelial regeneration, and brain tissue healing.
Adipose-derived Stem Cells Banking
The AABB and the FACT, along with NetCord, have already adopted worldwide standards for quality management systems and technical requirements for umbilical cord blood (UCB) banking. While FACT-NetCord CB requirements are independent but consistent with their cellular therapy standards, they are incorporated into their general cellular therapy standards. Despite the fact that there is variety and differences in the ethical and legal processes governing the use of ASCs in therapies, such broad standards and precautions have been implemented. In the case of hematopoietic stem cells and CB, long-term storage of ASCs is now possible thanks to cryovials or cryo-bags used in manual operations as well as semi-closed or closed automated systems. Stem cell banks, on the other hand, primarily utilized specialized cryo-bags to facilitate culture and quality control methods such as those used for freezing and banking CB stem cells in accordance with GMP regulations. Containers necessary for storage and shipment to the point of care are largely the same as those used in UCB banking around the world.
Long-term storage of bone marrow and adipose-derived stem cells is also becoming more prevalent. According to a recent study, AT and ASCs can be cryopreserved for up to 44 months before being used. However, the perspectives on cell viability, functionality, and integrity are still lacking, and more research is needed to determine whether long-term banking is feasible. Low temperatures would not stop infectious agents from growing in the event of bacterial/viral infections, implying that with constant agents like the coronavirus, long-term quality control at different time points is becoming a significant challenge during cryopreservation.
Tumorigenesis
Wang et al. described the suitability of ASCs for the treatment of hepatocellular carcinoma, among other things. Through the downregulation of Akt signaling, a signal pathway that promotes cell survival and growth in response to extracellular signals, ASCs suppressed hepatocellular carcinoma cell proliferation and division and triggered hepatocellular carcinoma cell death.
When ASCs are used for cancer reconstruction, their significant qualities of being pivotal for tissue regeneration have been found to also generate the perilous potential for tumor development stimulation. The use of ASCs, and more broadly MSCs, causes tumor stimulation, which leads to cell engraftment in the tumor mass. In the instance of breast cancer, cancer cells induce MSCs to secrete the chemokine CCL5, which subsequently acts in a paracrine way on the cancer cells to increase cell motility, invasion, and metastasis.
In addition, human ASCs were found to interact with human squamous cell cancer cells. In co-culture with ASCs, the invasive propensity of squamous cell carcinoma cells was dramatically increased, implying an elevated oncological risk.
Several studies have demonstrated that ASCs increase cancer cell proliferation, implying that ASCs may promote the growth of pre-existing tumors even if they do not form tumors themselves. As a result, using ASCs to treat cancer disorders is not suggested because it may cause serious negative effects. Even though no malignant behavior of MSCs has been recorded in clinical research to date, these studies should not be overlooked when considering the clinical application of human ASCs for regenerative therapy.
Conclusion
ASCs have been regarded as advantageous in regenerative therapies for a variety of disease processes due to their promising therapeutic benefits. For various reasons, ASCs are thought to be excellent for use in regenerative therapies. They can be retrieved, handled, and expanded in a minimally invasive, simple, and effective manner, and they have a high potential for transformation into mature cells along the mesodermal, ectodermal, and endodermal lineages. In vitro stem cells have made significant progress in terms of isolation, morphological characteristics, molecular biology, and differentiation capacity in recent years, and it is now obvious that ASCs can mediate therapeutic benefits.
To be more specific, they not only serve as tissue-specific progenitor cells, but also work through a variety of mechanisms, including paracrine-mediated angiogenesis, inflammation, cell homing, cell survival, and other related processes. These fundamental insights have aided in gradually closing the gap between basic knowledge and clinical application, while ASCs have been tested in clinical trials all over the world, demonstrating that they are safe and practical in a range of scenarios.