Adipocytes are a known source of stem cells. They are easy to harvest, and are a suitable candidate for autogenous grafts. Adipose derived stem cells (ADSCs) have multiple target tissues which they can differentiate into, including bone and cartilage. In adipose tissue, ADSCs are able to differentiate, as well as providing energy and a supply of cytokines/hormones to manage the hypoxic and lipid/hormone saturated adipose environment. The plasminogen activation system (PAS) controls the majority of proteolytic activities in both adipose and wound healing environments, allowing for rapid cellular migration and tissue remodelling. While the primary activation pathway for PAS occurs through the urokinase plasminogen activator (uPA), which is highly expressed by endothelial cells, its function is not limited to enabling revascularisation. Proteolytic activity is dependent on protease activation, localisation, recycling mechanisms and substrate availability. uPA and uPA activated plasminogen allows pluripotent cells to arrive to new local environments and fulfil the niche demands. However, overstimulation, the acquisition of a migratory phenotype and constant protein turnover can be unconducive to the formation of structured hard and soft tissues. To maintain a suitable healing pattern, the proteolytic activity stimulated by uPA is modulated by plasminogen activator inhibitor 1. Depending on the physiological settings, different parts of the remodelling mechanism are activated with varying results. Utilising the differences within each microenvironment to recreate a desired niche is a valid therapeutic bio-engineering approach. By controlling the rate of protein turnover combined with a receptive stem cell lineage, such as ADSC, a novel avenue on the therapeutic opportunities may be identified, which can overcome limitations, such as scarcity of stem cells, low angiogenic potential or poor host tissue adaptation.
Epimorphosis, the regeneration of a specific part of an organism, such as a limb, does not occur in humans, and is limited to regrowth of the tips of the digits (
Adipocytes are present throughout the body within adipose tissue (
MSCs have been shown to serve an adjuvant role with beneficial effects as a clinical adjunct in non-healing ulcers (
Adipose derived stem cells (ADSCs) are easy to obtain due to the abundance of adipose tissue, as well as the fact that the isolation time is reported to be as short as 30 min (
Mature adipocytes can also be utilised clinically, for instance to form dedifferentiated fat (DFAT) cells, using a ceiling culture method, which provides a physical dedifferentiation stimulus on the cultured adipocytes. This physical stimulus activates the Wnt pathway, resulting in MSC-like protein expression and pluripotency (
ADSCs have been shown to be more versatile in adapting to surgical use than BMSCs (
PPARγ is considered the master regulator of proadipogenic differentiation since all stimuli of adipogenesis converge on it (
MSC sensitivity to PPARγ is reliant on a cascade of signalling factors. Bone morphogenic protein 4 signalling determines the adipose lineage, whereas CEBPβ stimulation by insulin like growth factor-1 or glucocorticoids stimulates the preadipocytes from a growth arrested state to re-entry back into the cell cycle, at which point PPARγ commits them to differentiate into terminal adipocytes (
Mature adipocytes have been shown to exhibit a plastic phenotype in a variety of conditions (
Inhibition of the PPARγ molecule is a method of forced dedifferentiation of mature adipocytes and has been achieved using tumour necrosis factor-α (TNF-α) (
MCP-1 stimulation of integrin mediated cell adhesion and migration has been shown to be abrogated by a naturally occurring truncated soluble urokinase plasminogen activator receptor (uPAR) lysis product termed D2D388-274, which inhibits the human formyl peptide receptor like-1 (FPRL-1) G-protein coupled receptor (
The αvβ3 vitronectin specific integrin has been found to allow MSC to activate Wnt signalling and maintain pluripotency (
The activation of the Wnt signalling pathway has an inhibitory effect on PPARγ production and therefore inhibited adipocytic differentiation (
TNF-α stimulated adipocyte dedifferentiation was found to be mediated by PAI-1, since PAI-1 deficiency caused an upregulation of PPARγ in TNF-α stimulated cells, resulting in abrogation of the dedifferentiation caused by TNF-α (
Due to the multiple sources indicating PAI-1 and Wnt involvement in dedifferentiation of adipocytes, or maintenance of MSC pluripotency, their interaction is discussed further below.
PAI-1 is a member of the PAS. The primary protease of this system is plasmin, which is responsible for catalysing the lysis of fibrin, glycoproteins and other components of the extracellular matrix (ECM), but requires activation from a precursor state (
The cleavage of the plasmin precursor, plasminogen, is mediated by uPA and the tissue-type plasminogen activator (tPA), activity of both of which is regulated by the PA-I family of proteins, of which PAI-1 is the most rapidly acting and abundant (
Under hypoxic conditions, the adipose tissue actively utilises the PAS, both in physiological (
uPA activates plasminogen, and can itself initiate the extracellular remodelling process (
MMPs are secreted either as membrane anchored or free proteins, and in both instances as inactive zymogens, requiring lysis for activation (
In the context of adipose organs, continuous remodelling is important physiologically to maintain stability during adaptation to storage capacities of dietary nutrients, and this puts pressure on the vasculature and connective tissues (
Hypoxia primed neutrophils start adhering and migrating towards the affected ECs (
PAI-1 is found bound to vitronectin, in a latent state which prevents its autolysis (
The uPA/uPAR/integrin complex allows for directional endocytosis and proteolysis along the path of ECM degradation. The presence of PAI-1 at the uPA/uPAR/integrin complex can cause LRP-mediated intracellular recycling. This severs the extracellular connection of uPA/uPAR/integrin and vitronectin to the degrading ECM, and thus reduces the mobility caused by uPA/uPAR/integrin association (
Vitronectin cellular attachment is mediated by integrin αvβ3, which can also act in concert with αvβ5 to bind fibronectin (
Atherosclerotic vessels are a common complication of hypoxia inducing conditions, such as obesity and diabetes (
Integrin binding to collagen and fibronectin can increase the secretion of uPA, uPAR and PAI-1; however, αvβ3 binding to vitronectin was found to downregulate uPA and uPAR antigen levels and upregulate PAI-1(
PAI-1 is secreted by ADSCs and osteoblasts derived from ADSCs (
The integrin binding site on vitronectin is shared by PAI-1, suggesting an interaction between signalling pathways (
The uPA/PAI-1 complex on the uPAR causes clathrin/LRP1-mediated recycling (
Wnt-1 inducible signalling pathway protein 1 (WISP1) is a downstream mediator of Wnt signalling (
The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signalling pathway is necessary to stimulate adipocyte differentiation from 3T3-L1 preadipocytes in the absence of other inputs, as AKT1 can stimulate PPARγ production (
The change in uPAR colocalization to integrins from α3β1 to αvβ5 blocks the uPA signalling and activation of ERK or AKT (
Wound closure is initiated by thrombin, which activates fibrinogen to create a fibrin clot (
Fully differentiated adipocytes were reported to lower PAI-1 concentration, and the subsequent addition of PAI-1 to an osteoblast/fully differentiated adipocyte coculture did not cause spontaneous transition of adipocytes to osteoblasts (
Whilst detection of increased secretion of PAS components within the tissue can be indicative of oncogenesis (
Instances where establishing a sufficient blood supply is of high concern, such as an osseoinductive transplant, a collagenous cellular carrier implant, or for procedures such as bone distraction, extensive surgical flaps, or any other surgical intervention where scarring and grafting is an issue, may benefit from the possible therapeutic applications of receptive stem cells. When the protraction of healing is necessary to allow for complete angiogenesis, and adequate deposition of ECM to support the unformed tissue, uPA can ensure that the microenvironment is maintained in a state of turnover (
The ability for adipocytes to secrete uPA and PAI-1 to modulate and maintain their pluripotent microenvironment has been explored, and this may be conducive in wound healing or bio-engineering stents (
The MSC like fraction can be boosted by synthetically overloading harvested adipocytes with PAI-1 or any of the other Wnt activating PPARγ inhibitors and re-introducing this population into a wound or surgical microenvironment in order to become more susceptible to local differentiation factors and conduit healing or growth (
Alternatively, whole tissue adequately prepared to stimulate activation of the PAS, such as in a wound healing or hypoxic environment, may create a DFAT rich graft which would be significantly more conducive to local cellular populations or ECM architectures in mesenchymal lineage use cases (
For directed mobility via extracellular degradation, there needs to be present a steady stream of uPA to counteract any baseline PAI-1 secretion, to overcome the background PAI-1 recycling of the uPA/uPAR/integrin complex, which undergoes intracellular recycling along with LRP and downstream targets. Such interventions are difficult to perform
PAI-1 is found to be stable for 145 h when bound to vitronectin, and for 2 h when expressed in isolation
Studies have shown that platelet rich plasma with ADSCs can significantly improve tissue incorporation of synthetic scaffolds and their neovascularisation (
Adipocytes can be seen as a versatile cell lineage which harbour mesenchymal stem potential in the form of MSCs, ADSCs and the induced DFAT cells. The formation of the latter has been found to rely on inhibition of PPARγ. Wnt signalling has been shown to be stimulated by TNF-α and integrins, notably αvβ3 via fibrin. The proteolytic cascade activated by PAS during inflammation is mediated by multiple Wnt activators, acting on Wnt to stimulate uPA release for ECM degradation, PAI-1 release for endocytosis, uPAR for localisation to LRP, and integrins to provide motility and directional specificity.
Since Wnt activation also inhibits PPARγ expression, there is a high chance that adipose cells exposed to a fibrin or vitronectin rich wound environment would undergo dedifferentiation. PAI-1 expression has also been linked to dedifferentiation, which could be explained by its ability to stimulate endocytotic clathrin basket mediated recycling of uPA/uPAR/integrin complexes. After wound resolution, in order to reach homeostasis and inhibit uPA mediated extracellular ECM degradation, PAI-1 needs to be released following its internalisation, suggesting that activation of Wnt signalling is necessary in wound healing in order to produce PAI-1, and consequently inhibit PPARγ. Coupled with Wnt mediated stimulation of PAS expression to allow for remodelling of the ECM and cellular motility, there is a strong suggestion that Wnt is central to the success and high versatility of adipose tissue and more importantly DFAT cells in pilot studies. An initial dedifferentiating priming of adipose to DFAT cells from a lipoaspirate harvest in a PRF could be followed by cytokine, mineral or ECM exposure to initiate target cell differentiation prior to clinical use, taking the DFAT cells one step closer to use in a clinically applicable environment.
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MWS, AE and conceived the topic of the review MN wrote the manuscript. MN and AE reviewed and revised the manuscript. All authors have read and approved the final manuscript. Data sharing is not applicable.
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The authors declare that they have no competing interests.
Stimuli mediating differentiation of MSCs to adipocytes. MSC, mesenchymal stem cell.
Significant molecules involved in regulation of PPARγ. PPARγ, peroxisome proliferator-activated receptor γ.
Primary pathways identified in the literature, which are either physiologically or pathologically involved in mediating adipocyte dedifferentiation.
The adipocyte tissue niche and hypoxia resulting from growth, hyperplasia or damage to the vasculature. The resulting response is magnified showing vascular ingress towards the hypoxic location, vascular perfusion, neutrophil infiltration and ECM degradation. The primary molecules regulated by the hypoxic environment are highlighted. ECM, extracellular matrix.
PAS activity is dependent on the microenvironment and available complex elements. PAS, plasminogen activation system.
PAI-1/LRP mediated clathrin recycling of the uPA/uPAR complex and the activation of the vitronectin/αvβ3 complex can be simultaneous due to the different binding sites between PAI-1/uPA and PAI-1/LRP, and uPA/uPAR itself also has a separate integrin binding domain. The resultant vacuole is internalised with the plasma membrane, and is isolated from the ECM, allowing only for non-ECM dependent signalling. PAI, plasminogen activator inhibitor 1; LRP, lipoprotein receptor-related protein; uPA, urokinase plasminogen activator; uPAR, uPA receptor.