Corrigendum: Human abdominal subcutaneous-derived active beige adipocytes carrying FTO rs1421085 obesity-risk alleles exert lower thermogenic capacity.

[This corrects the article DOI: 10.3389/fcell.2023.1155673.].

Human abdominal subcutaneous-derived active beige adipocytes carrying FTO rs1421085 obesity-risk alleles exert lower thermogenic capacity.

Introduction: White adipocytes store lipids, have a large lipid droplet and few mitochondria. Brown and beige adipocytes, which produce heat, are characterized by high expression of uncoupling protein (UCP) 1, multilocular lipid droplets, and large amounts of mitochondria. The rs1421085 T-to-C single-nucleotide polymorphism (SNP) of the human FTO gene interrupts a conserved motif for ARID5B repressor, resulting in adipocyte type shift from beige to white. Methods: We obtained abdominal subcutaneous adipose tissue from donors carrying FTO rs1421085 TT (risk-free) or CC (obesity-risk) genotypes, isolated and differentiated their preadipocytes into beige adipocytes (driven by the PPARγ agonist rosiglitazone for 14 days), and activated them with dibutyryl-cAMP for 4 hours. Then, either the same culture conditions were applied for additional 14 days (active beige adipocytes) or it was replaced by a white differentiation medium (inactive beige adipocytes). White adipocytes were differentiated by their medium for 28 days. Results and Discussion: RNA-sequencing was performed to investigate the gene expression pattern of adipocytes carrying different FTO alleles and found that active beige adipocytes had higher brown adipocyte content and browning capacity compared to white or inactive beige ones when the cells were obtained from risk-free TT but not from obesity-risk CC genotype carriers. Active beige adipocytes carrying FTO CC had lower thermogenic gene (e.g., UCP1, PM20D1, CIDEA) expression and thermogenesis measured by proton leak respiration as compared to TT carriers. In addition, active beige adipocytes with CC alleles exerted lower expression of ASC-1 neutral amino acid transporter (encoded by SLC7A10) and less consumption of Ala, Ser, Cys, and Gly as compared to risk-free carriers. We did not observe any influence of the FTO rs1421085 SNP on white and inactive beige adipocytes highlighting its exclusive and critical effect when adipocytes were activated for thermogenesis.

Mitophagy Mediates the Beige to White Transition of Human Primary Subcutaneous Adipocytes Ex Vivo.

Brown and beige adipocytes have multilocular lipid droplets, express uncoupling protein (UCP) 1, and promote energy expenditure. In rodents, when the stimulus of browning subsides, parkin-dependent mitophagy is activated and dormant beige adipocytes persist. In humans, however, the molecular events during the beige to white transition have not been studied in detail. In this study, human primary subcutaneous abdominal preadipocytes were differentiated to beige for 14 days, then either the beige culture conditions were applied for an additional 14 days or it was replaced by a white medium. Control white adipocytes were differentiated by their specific cocktail for 28 days. Peroxisome proliferator-activated receptor γ-driven beige differentiation resulted in increased mitochondrial biogenesis, UCP1 expression, fragmentation, and respiration as compared to white. Morphology, UCP1 content, mitochondrial fragmentation, and basal respiration of the adipocytes that underwent transition, along with the induction of mitophagy, were similar to control white adipocytes. However, white converted beige adipocytes had a stronger responsiveness to dibutyril-cAMP, which mimics adrenergic stimulus, than the control white ones. Gene expression patterns showed that the removal of mitochondria in transitioning adipocytes may involve both parkin-dependent and -independent pathways. Preventing the entry of beige adipocytes into white transition can be a feasible way to maintain elevated thermogenesis and energy expenditure.

BMP7 Increases UCP1-Dependent and Independent Thermogenesis with a Unique Gene Expression Program in Human Neck Area Derived Adipocytes.

White adipocytes contribute to energy storage, accumulating lipid droplets, whereas brown and beige adipocytes mainly function in dissipating energy as heat primarily via the action of uncoupling protein 1 (UCP1). Bone morphogenic protein 7 (BMP7) was shown to drive brown adipocyte differentiation in murine interscapular adipose tissue. Here, we performed global RNA-sequencing and functional assays on adipocytes obtained from subcutaneous (SC) and deep-neck (DN) depots of human neck and differentiated with or without BMP7. We found that BMP7 did not influence differentiation but upregulated browning markers, including UCP1 mRNA and protein in SC and DN derived adipocytes. BMP7 also enhanced mitochondrial DNA content, levels of oxidative phosphorylation complex subunits, along with PGC1α and p-CREB upregulation, and fragmentation of mitochondria. Furthermore, both UCP1-dependent proton leak and UCP1-independent, creatine-driven substrate cycle coupled thermogenesis were augmented upon BMP7 addition. The gene expression analysis also shed light on the possible role of genes unrelated to thermogenesis thus far, including ACAN, CRYAB, and ID1, which were among the highest upregulated ones by BMP7 treatment in both types of adipocytes. Together, our study shows that BMP7 strongly upregulates thermogenesis in human neck area derived adipocytes, along with genes, which might have a supporting role in energy expenditure.

Irisin Stimulates the Release of CXCL1 From Differentiating Human Subcutaneous and Deep-Neck Derived Adipocytes via Upregulation of NFκB Pathway.

Thermogenic brown and beige adipocytes might open up new strategies in combating obesity. Recent studies in rodents and humans have indicated that these adipocytes release cytokines, termed "batokines". Irisin was discovered as a polypeptide regulator of beige adipocytes released by myocytes, primarily during exercise. We performed global RNA sequencing on adipocytes derived from human subcutaneous and deep-neck precursors, which were differentiated in the presence or absence of irisin. Irisin did not exert an effect on the expression of characteristic thermogenic genes, while upregulated genes belonging to various cytokine signaling pathways. Out of the several upregulated cytokines, CXCL1, the highest upregulated, was released throughout the entire differentiation period, and predominantly by differentiated adipocytes. Deep-neck area tissue biopsies also showed a significant release of CXCL1 during 24 h irisin treatment. Gene expression data indicated upregulation of the NFκB pathway upon irisin treatment, which was validated by an increase of p50 and decrease of IκBα protein level, respectively. Continuous blocking of the NFκB pathway, using a cell permeable inhibitor of NFκB nuclear translocation, significantly reduced CXCL1 release. The released CXCL1 exerted a positive effect on the adhesion of endothelial cells. Together, our findings demonstrate that irisin stimulates the release of a novel adipokine, CXCL1, via upregulation of NFκB pathway in neck area derived adipocytes, which might play an important role in improving tissue vascularization.

ASC-1 transporter-dependent amino acid uptake is required for the efficient thermogenic response of human adipocytes to adrenergic stimulation.

Brown and beige adipocytes dissipate energy by uncoupling protein 1 (UCP1)-dependent and UCP1-independent thermogenesis, which may be utilized to develop treatments against obesity. We have found that mRNA and protein expression of the alanine/serine/cysteine transporter-1 (ASC-1) was induced during adipocyte differentiation of human brown-prone deep neck and beige-competent subcutaneous neck progenitors, and SGBS preadipocytes. cAMP stimulation of differentiated adipocytes led to elevated uptake of serine, cysteine, and glycine, in parallel with increased oxygen consumption, augmented UCP1-dependent proton leak, increased creatine-driven substrate cycle-coupled respiration, and upregulation of thermogenesis marker genes and several respiratory complex subunits; these outcomes were impeded in the presence of the specific ASC-1 inhibitor, BMS-466442. Our data suggest that ASC-1-dependent consumption of serine, cysteine, and glycine is required for efficient thermogenic stimulation of human adipocytes.

FTO Intronic SNP Strongly Influences Human Neck Adipocyte Browning Determined by Tissue and PPARγ Specific Regulation: A Transcriptome Analysis.

Brown adipocytes, abundant in deep-neck (DN) area in humans, are thermogenic with anti-obesity potential. FTO pro-obesity rs1421085 T-to-C single-nucleotide polymorphism (SNP) shifts differentiation program towards white adipocytes in subcutaneous fat. Human adipose-derived stromal cells were obtained from subcutaneous neck (SC) and DN fat of nine donors, of which 3-3 carried risk-free (T/T), heterozygous or obesity-risk (C/C) FTO genotypes. They were differentiated to white and brown (long-term Peroxisome proliferator-activated receptor gamma (PPARγ) stimulation) adipocytes; then, global RNA sequencing was performed and differentially expressed genes (DEGs) were compared. DN and SC progenitors had similar adipocyte differentiation potential but differed in DEGs. DN adipocytes displayed higher browning features according to ProFAT or BATLAS scores and characteristic DEG patterns revealing associated pathways which were highly expressed (thermogenesis, interferon, cytokine, and retinoic acid, with UCP1 and BMP4 as prominent network stabilizers) or downregulated (particularly extracellular matrix remodeling) compared to SC ones. Part of DEGs in either DN or SC browning was PPARγ-dependent. Presence of the FTO obesity-risk allele suppressed the expression of mitochondrial and thermogenesis genes with a striking resemblance between affected pathways and those appearing in ProFAT and BATLAS, underlining the importance of metabolic and mitochondrial pathways in thermogenesis. Among overlapping regulatory influences that determine browning and thermogenic potential of neck adipocytes, FTO genetic background has a thus far not recognized prominence.

Differentiating SGBS adipocytes respond to PPARγ stimulation, irisin and BMP7 by functional browning and beige characteristics.

Brown and beige adipocytes are enriched in mitochondria with uncoupling protein-1 (UCP1) to generate heat instead of ATP contributing to healthy energy balance. There are few human cellular models to reveal regulatory networks in adipocyte browning and key targets for enhancing thermogenesis in obesity. The Simpson-Golabi-Behmel syndrome (SGBS) preadipocyte line has been a useful tool to study human adipocyte biology. Here we report that SGBS cells, which are comparable to subcutaneous adipose-derived stem cells, carry an FTO risk allele. Upon sustained PPARγ stimulation or irisin (a myokine released in response to exercise) treatment, SGBS cells differentiated into beige adipocytes exhibiting multilocular lipid droplets, high UCP1 content with induction of typical browning genes (Cidea, Elovl3) and the beige marker Tbx1. The autocrine mediator BMP7 led to moderate browning with the upregulation of the classical brown marker Zic1 instead of Tbx1. Thermogenesis potential resulted from PPARγ stimulation, irisin and BMP7 can be activated in UCP1-dependent and the beige specific, creatine phosphate cycle mediated way. The beige phenotype, maintained under long-term (28 days) conditions, was partially reversed by withdrawal of PPARγ ligand. Thus, SGBS cells can serve as a cellular model for both white and sustainable beige adipocyte differentiation and function.