Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2019 Apr;60(4):856-868.
doi: 10.1194/jlr.M091173. Epub 2019 Feb 19.

Rosiglitazone remodels the lipid droplet and britens human visceral and subcutaneous adipocytes ex vivo

Mi-Jeong Lee  1 , Sukanta Jash  2 , Jessica E C Jones  3 , Vishwajeet Puri  2 , Susan K Fried  4
Affiliations

Rosiglitazone remodels the lipid droplet and britens human visceral and subcutaneous adipocytes ex vivo

Mi-Jeong Lee et al. J Lipid Res. 2019 Apr.

Abstract

Treatment with PPARγ agonists in vivo improves human adipocyte metabolism, but the cellular mechanisms and possible depot differences in responsiveness to their effects are poorly understood. To examine the ex vivo metabolic effects of rosiglitazone (Rosi), we cultured explants of human visceral (omental) and abdominal subcutaneous adipose tissues for 7 days. Rosi increased mRNA levels of transcriptional regulators of brite/beige adipocytes (PGC1α, PRDM16), triglyceride synthesis (GPAT3, DGAT1), and lipolysis (ATGL) similarly in adipose tissues from both depots. In parallel, Rosi increased key modulators of FA oxidation (UCP1, FABP3, PLIN5 protein), rates of FA oxidation, and protein levels of electron transport complexes, suggesting an enhanced respiratory capacity as confirmed in newly differentiated adipocytes. Rosi led to the formation of small lipid droplets (SLDs) around the adipocyte central lipid droplet; each SLD was decorated with redistributed mitochondria that colocalized with PLIN5. SLD maintenance required lipolysis and FA reesterification. Rosi thus coordinated a structural and metabolic remodeling in adipocytes from both visceral and subcutaneous depots that enhanced oxidative capacity. Selective targeting of these cellular mechanisms to improve adipocyte FA handling may provide a new approach to treat metabolic complications of obesity and diabetes.

Keywords: adipose depots; fatty acid oxidation; perilipins; protein carbonylation; thiazolidinedione.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing financial interests.

Figures

Fig. 1.
Fig. 1.
Rosi induced brite phenotypes in both Om adipose tissues and subcutaneous human adipose tissues. A, B: mRNA and protein levels of genes known to be more abundantly expressed in brown and brite than white adipocytes were measured in Om adipose tissues and abdominal subcutaneous (SC) human adipose tissues that had been cultured without or with Rosi (1 μM) for 7 days. C, D: After treating Om adipose tissue without or with Rosi, fat cells (FC) and stromal vascular cells (SVC) were isolated with collagenase digestion and mRNA and protein levels were measured in tissue (T), FCs, and SVCs. Data are expressed as the mean ± SEM of five to nine experiments [BMI, 37.5 ± 3.7 kg/m2 (range 29–54 kg/m2); age, 50.8 ± 4.0 years (range 36–71 years); seven female/two male]. *P < 0.05, **P < 0.01, ***P < 0.001 (paired t-tests after one- or two-way ANOVA).
Fig. 2.
Fig. 2.
Effects of Rosi on lipolytic rates and FA oxidative capacity in human adipocytes. A: Glycerol accumulation in culture media during the final 24 h. *P < 0.05 with paired t-tests after two-way ANOVA, n = 6. B: Lipolytic capacity under basal and stimulated conditions was measured in isolated adipocytes from control or Rosi-treated Om tissues. Data are expressed as the mean ± SEM of seven experiments. *P < 0.05 (control vs. Rosi); #P < 0.05 compared with the basal within each condition (post hoc paired t-tests) after RM-ANOVA (F = 3.22, P = 0.004). C: FA oxidation rates (14C-oleate oxidation to 14CO2) in adipose tissues cultured with or without Rosi. Data are expressed as the mean ± SEM of four experiments. *P < 0.05, **P < 0.01 with paired t-tests. D: Expression levels of mitochondrial ETC proteins were measured in isolated mitochondria after 7 days of treatment with Rosi and a representative Western blot of three (Om) or four subcutaneous (SC) are shown.
Fig. 3.
Fig. 3.
Rosi increased expression levels of genes involved in TAG turnover while suppressing inflammatory markers in both Om adipose tissues and subcutaneous human adipose tissues. Expression levels of mRNAs in TAG breakdown and FA oxidation (A), synthesis of FA, TAG, and phospholipids (B), and secretory factors (C) were measured in control or Rosi-treated Om and abdominal subcutaneous (SC) adipose tissues with qPCR. Data are expressed as the mean ± SEM of values from five to nine subjects [BMI, 37.5 ± 3.7 kg/m2 (range 29–54 kg/m2); age, 50.8 ± 4.0 years (range 36–71 years); seven female/two male]. *P < 0.05, **P < 0.01, ***P < 0.001 (paired t-tests after two-way ANOVA).
Fig. 4.
Fig. 4.
Rosi induced remodeling of LDs and rearrangement of mitochondria in human adipocytes. Isolated adipocytes from control or Rosi-treated Om tissues were used for confocal imaging of LDs and mitochondria. Neutral lipids were labeled with LipidTOX-Deep Red, mitochondria were stained with Mitotracker-Green (A) or Mitotracker-Red (B), and nuclei were stained with DAPI. White scale bars = 10 μm.
Fig. 5.
Fig. 5.
LD remodeling was blocked by both orlistat and triacsin C while mitochondrial rearrangement was blocked by triacsin C only. After culturing Om adipose tissues with or without Rosi for 5 days, triacsin C or orlistat was added during the final 2 days of culture without or with Rosi, as described in the Materials and Methods. Adipocytes were isolated and stained with LipidTOX-Deep Red and Mitotracker-Red followed by confocal imaging. White scale bars = 10 μm.
Fig. 6.
Fig. 6.
PLIN5 coated the small droplets and colocalized with mitochondria in brite adipocytes. Adipocytes isolated from Om tissues after 7 day culture without or with Rosi were stained with LipidTOX-Deep Red and Mitotracker-Red and used for immunostaining of PLIN1 (A) and PLIN5 (B), as described in the Materials and Methods. White arrows indicate where PLIN5 colocalized with mitochondria. White scale bars = 10 μm.
Fig. 7.
Fig. 7.
Rosi increased oxidative capacity in newly differentiated subcutaneous adipocytes. A: Differentiated abdominal subcutaneous adipocytes (day 14) were treated with Rosi for 7 days and protein levels of UCP1, PLIN5, FABP3, and porin were measured. Blots from three different individuals are shown. B: OCRs were measured with a Seahorse extracellular flux analyzer. After measuring basal OCR, the following were injected sequentially: oligomycin (ATP synthase inhibitor), FCCP (uncoupler), and rotenone (complex I inhibitor)/antimycin A (complex III inhibitor) (Rot/AA). The mean ± SD of five wells per condition over time from a representative subject is shown. C: Average values for different components of mitochondrial respiration were calculated (43). Data are expressed as the mean ± SEM of values from three independent subjects (**P < 0.01, ***P < 0.001 by paired t-tests for Rosi effects).
Fig. 8.
Fig. 8.
Rosi increased ROS in human adipocytes. Differentiated abdominal subcutaneous adipocytes were treated with or without Rosi for 7 days with TNFα (50 pM) added during the final 24 h. Protein carbonylation was determined in cell lysates. A representative blot (A) and quantification four independent experiments using cells from different subjects (B) are presented. **Effects of Rosi (P < 0.01) and #effects of TNFα (P < 0.05) with paired t-tests after two-way ANOVA.
Fig. 9.
Fig. 9.
Rosi-mediated metabolic and structural remodeling of human adipocytes. Activation of PPARγ with Rosi drove transcriptional changes that reprogrammed lipid metabolism in mature white adipocytes from both visceral and subcutaneous adipose tissues. The high oxidative capacity in the brite adipocytes was accompanied by a structural remodeling of LDs and mitochondria. Clusters of SLDs were formed, decorated with PLIN5, and became closely associated with rearranged mitochondria. These small droplets were formed through reesterification of FAs released by lipolysis.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Gustafson B., Hedjazifar S., Gogg S., Hammarstedt A., and Smith U.. 2015. Insulin resistance and impaired adipogenesis. Trends Endocrinol. Metab. 26: 193–200. - PubMed
    1. Koutsari C., and Jensen M. D.. 2006. Free fatty acid metabolism in human obesity. J. Lipid Res. 47: 1643–1650. - PubMed
    1. Rydén M., and Arner P.. 2017. Subcutaneous adipocyte lipolysis contributes to circulating lipid levels. Arterioscler. Thromb. Vasc. Biol. 37: 1782–1787. - PMC - PubMed
    1. Das S. K., Chu W. S., Mondal A. K., Sharma N. K., Kern P. A., Rasouli N., and Elbein S. C.. 2008. Effect of pioglitazone treatment on endoplasmic reticulum stress response in human adipose and in palmitate-induced stress in human liver and adipose cell lines. Am. J. Physiol. Endocrinol. Metab. 295: E393–E400. - PMC - PubMed
    1. Frayn K. N., Langin D., and Karpe F.. 2008. Fatty acid-induced mitochondrial uncoupling in adipocytes is not a promising target for treatment of insulin resistance unless adipocyte oxidative capacity is increased. Diabetologia. 51: 394–397. - PubMed

Publication types