10 research outputs found
Dedifferentiating human mature adipocytes.
<p>A) Confocal microscopy of human mature adipocytes undergoing dedifferentiation at various stages (10x) B) Oil red O staining of OM mature adipocytes undergoing dedifferentiation in ceiling culture (4x). Days 4, 7 and 12 are shown. C) Lipid droplet size measurement of dedifferentiating mature adipocyte cells at days 4, 7 and 12 (n = 6, 4M; 2F, BMI range: 39–56.9 kg/m<sup>2</sup>, Age range: 29–59 years). Left panel: Statistical MANOVA analysis: Time p<0.05. Middle and right panels: Student T-tests *p<0.05.</p
FAP and DPP4 protein expression in mature adipocytes and whole adipose tissue.
<p>A) FAP and DPP4 proteins are induced during adipocyte dedifferentiation in both OM and SC fat depots at various time-points. β-tubulin was used as loading control. B) Immunofluorescent FAP and DPP4 proteins were detected in 4 days-dedifferentiating mature adipocytes. (D: day, OM: omental, SC: subcutaneous)</p
Effects of a DPP4 inhibitor on gene expression of dedifferentiating SC adipocytes.
<p>A) C/EBPα, PPARγ2 and adiponectin gene expression were quantified in mature adipocytes undergoing dedifferentiation (day 5) after a 24-hour treatment with a DPP4 inhibitor (50 μM) or vehicle (Control). N = 13 patients, Matched pair t-test: *p<0.05, § p≤0.10, δ p≤0.15. B) Dose response of DPP4 inhibitor on C/EBPα expression levels in dedifferentiating adipocytes. Cells were treated at days 4 and 6 of ceiling culture in a 6-well plate and harvested at day 6 for qPCR analysis. Repeated measures analysis p-value = 0.06.</p
FAP and DPP4 protein detection in whole adipose tissue by immunohistochemistry from a 59-year old man with a BMI of 35.5 kg/m<sup>2</sup>.
<p>A) Negative control-mouse antiserum, B) DPP4 antibody 4x, C) DPP4 antibody 20x, E) Negative control-rabbit antiserum, F) FAP antibody 4x, G) FAP antibody 20x. Scale bar: 250 ÎĽM for panels A, B, D and E, 50 ÎĽM for C and F.</p
Expression levels of transcripts associated with mature adipocyte functions over a dedifferentiation time-course.
<p>PPARγ2, C/EBPα, LPL and Adiponectin expression levels were measured in mature adipocytes (D0) and in dedifferentiating adipocytes at days 4 and 7 (D4-D7) of ceiling culture. Data are expressed as ΔΔCt relative to G6PD expression (Mean value ± SEM, n = 4). Student t-test * p≤0.01, ** p<0.001.</p
Cytokines secreted by dedifferentiating OM and SC mature adipocytes.
<p>A) Culture media from OM and SC adipocytes in dedifferentiation were blotted on cytokine membranes (representative results shown). Positive cytokines appear as black dots, in duplicate. +: membrane positive control B) Graphs of IL-6 (top panel), IL-8 (middle panel) and VEGF (bottom panel) values measured by Bioplex in media of dedifferentiating adipocytes at days 7 and 12.</p
Relative gene expression of matrix remodeling genes in dedifferentiating adipocytes.
<p>A) FAP, DPP4, MMP1 and TGFβ1 expression levels were measured in mature adipocytes (D0) and in dedifferentiating adipocytes at days 4 and 7 (D4-D7) of ceiling culture. B) Data are expressed with depot-specific expression at days 0, 4 and 7 for omental (OM) and subcutaneous (SC) cells. C) Comparison of gene expression between adipose tissues, preadipocytes and DFAT cells from the OM and SC fat depots. Data are expressed as mean value of ΔΔCt relative to G6PD expression ± SEM (n = 4). Matched pair t-test: *p<0.05, ** p<0.01, § p<0.10.</p
Cytokine detection in dedifferentiating mature adipocyte culture media by bioplex analysis.
<p>Cytokine detection in dedifferentiating mature adipocyte culture media by bioplex analysis.</p
<b>IL-6 trans-signaling is increased in diabetes, impacted by glucolipotoxicity and associated with liver stiffness and fibrosis in fatty liver disease</b>
Many people living with diabetes also have non-alcoholic fatty liver disease (NAFLD). Interleukin-6 (IL-6) is involved in both diseases, interacting with both membrane-bound (classical) and circulating soluble receptors (trans-signaling). We investigated whether secretion of IL-6 trans-signaling co-receptors are altered in NAFLD by diabetes, and whether this might associate with the severity of fatty liver disease. Secretion patterns were investigated using human hepatocyte, stellate and monocyte cell lines. Associations with liver pathology were investigated in two patient cohorts: 1) biopsy-confirmed NASH and 2) class 3 obesity. We found that exposure of stellate cells to high glucose and palmitate increased IL-6 and sgp130 secretion. In line with this, plasma sgp130 in both patient cohorts positively correlated with HbA1c, and subjects with diabetes had higher circulating levels of IL-6 and trans-signaling co-receptors. Plasma sgp130 strongly correlated with liver stiffness and was significantly increased in subjects with F4 fibrosis stage. Monocyte activation was associated with reduced sIL-6R secretion. These data suggest that the hyperglycemia and hyperlipidemia can directly impact IL-6 trans-signaling, and that this may be linked to enhanced severity of NAFLD in patients with concomitant diabetes.</p