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Influence of glucose and insulin in human adipogenic differentiation models with adipose-derived stem cells.

ABSTRACT: Autologous fat grafting represents an attractive source for tissue engineering applications in the field of reconstructive medicine. However, in adipogenic differentiation protocols for human adipose-derived stem cells, the concentration of glucose and insulin varies considerably. With the intent to gain maximum tissue augmentation, we focused on the late phase of adipogenesis. In this study, we modified the differentiation protocol for adipose-derived stem cells by prolongation of the induction period and the application highly concentrated glucose and insulin. Human adipose-derived stem cells were isolated from subcutaneous depots and differentiated in a standard induction medium for the first two weeks, followed by two weeks with varying glucose and insulin concentrations. Morphological changes assessed using Oil-Red-O staining were examined for corresponding alterations in the expression of the adipogenic markers peroxisome proliferator-activated receptor gamma (PPAR?) and lipoprotein lipase (LPL). Furthermore, glucose and lactate levels in conditioned media were monitored over the period of differentiation. We found high-glucose media increasing the level of lipid accumulation and the size of single droplets whereas insulin significantly showed a dose-dependent negative effect on fat storage. However, whereas high glucose stimulated PPAR? transcription, expression levels in insulin-treated cells remained constant. Results permit assumptions that a high-glucose medium intensifies the degree of differentiation in mature adipocytes providing conditions to promote graft volume while we have identified highly concentrated insulin treatment as an inhibitor of lipid storage in the late adipogenic differentiation.

SUBMITTER: Kolodziej M

PROVIDER: S-EPMC6768274 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Influence of glucose and insulin in human adipogenic differentiation models with adipose-derived stem cells.

Kolodziej Michaela M Strauss Sarah S Lazaridis Andrea A Bucan Vesna V Kuhbier Jörn W JW Vogt Peter M PM Könneker Sören S

Adipocyte 20191201 1

Autologous fat grafting represents an attractive source for tissue engineering applications in the field of reconstructive medicine. However, in adipogenic differentiation protocols for human adipose-derived stem cells, the concentration of glucose and insulin varies considerably. With the intent to gain maximum tissue augmentation, we focused on the late phase of adipogenesis. In this study, we modified the differentiation protocol for adipose-derived stem cells by prolongation of the induction ...[more]

PMID: 31280651

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Project description:BackgroundAdipose-derived stem cells (ASC) and adipocytes are involved in numerous physiological and pathophysiological conditions, which have been extensively described in subcutaneous and visceral fat depots over the past two decades. However, much less is known about ASC and adipocytes outside classical fat tissue depots and their necessity in tissue remodeling after injury. Therefore, we investigated the etiology of adipocytes in human granulation tissue and define their possible role wound healing.MethodsIdentification of human wound tissue adipocytes was determined by immunohistochemical staining of granulation tissue sections from patients undergoing surgical debridement. Stromal cell fractions from granulation tissue and subcutaneous fat tissue were generated by collagenase type II-based protocols. Pro- and anti-inflammatory wound bed conditions were mimicked by THP1- and CD14+ monocyte-derived macrophage models in vitro. Effects of macrophage secretome on ASC differentiation and metabolism were determined by immunoblotting, flow cytometry, and microscopy assessing early and late adipocyte differentiation states. Functional rescuing experiments were conducted by lentiviral transduction of wildtype PPARG, IL1RA, and N-chlorotaurine (NCT) treatment.ResultsSingle and clustered adipocyte populations were detected in 11 out of 13 granulation tissue specimens and single-cell suspensions from granulation tissue showed adipogenic differentiation potential. Pro-inflammatory signaling by IFNG/LPS-stimulated macrophages (M (IFNG/LPS)) inhibited the maturation of lipid droplets in differentiated ASC. In contrast, anti-inflammatory IL4/IL13-activated macrophages (M (IL4/IL13)) revealed minor effects on adipocyte development. The M (IFNG/LPS)-induced phenotype was associated with a switch from endogenous fatty acid synthesis to glycolysis-dominated cell metabolism and increased pro-inflammatory cytokine production. Impaired adipogenesis was associated with increased, but seemingly non-functional, CEBPB levels, which failed to induce downstream PPARG and CEBPA. Neither transgenic PPARG overexpression, nor inhibition of IL1B was sufficient to rescue the anti-adipogenic effects induced by IFNG/LPS-activated macrophages. Instead, macrophage co-treatment during stimulation with NCT, a mild oxidant produced by activated granulocytes present in human wounds in vivo, significantly attenuated the anti-adipogenic effects.ConclusionsIn conclusion, the appearance of adipocytes in wound tissue indicates a prevailing anti-inflammatory environment that could be promoted by NCT treatment and may be associated with improved healing outcomes.

Dataset Information

Influence of glucose and insulin in human adipogenic differentiation models with adipose-derived stem cells.

Publications

Influence of glucose and insulin in human adipogenic differentiation models with adipose-derived stem cells.

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