Surgical injury induces local and distant adipose tissue browning

Abstract

The adipose organ, which comprises brown, white and beige adipocytes, possesses remarkable plasticity in response to feeding and cold exposure. The development of beige adipocytes in white adipose tissue (WAT), a process called browning, represents a promising route to treat metabolic disorders. While surgical procedures constantly traumatize adipose tissue, its impact on adipocyte phenotype remains to be established. Herein, we studied the effect of trauma on adipocyte phenotype one day after sham, incision control, or surgical injury to the left inguinal adipose compartment. Caloric restriction was used to control for surgery-associated body temperature changes and weight loss. We characterized the trauma-induced cellular and molecular changes in subcutaneous, visceral, interscapular, and perivascular adipose tissue using histology, immunohistochemistry, gene expression, and flow cytometry analysis. After one day, surgical trauma stimulated adipose tissue browning at the site of injury and, importantly, in the contralateral inguinal depot. Browning was not present after incision only, and was largely independent of surgery-associated body temperature and weight loss. Adipose trauma rapidly recruited monocytes to the injured site and promoted alternatively activated macrophages. Conversely, PDGF receptor-positive beige progenitors were reduced. In this study, we identify adipose trauma as an unexpected driver of selected local and remote adipose tissue browning, holding important implications for the biologic response to surgical injury.

Keywords: adipose; beige adipose; browning; dietary restriction; surgery; trauma.

Figures

Figure 1.

Surgical trauma induces adipose tissue browning. (A) Representative macroscopic morphology from paraffin embedded inguinal adipose tissues. From the left to the right: baseline, sham, incision and trauma group. (B) Representative H&E-stained inguinal adipose tissue sections which differentiated lipid droplets (white) from connective tissue (red). From the left to the right: baseline, sham, incision and trauma group. (C) Representative characterization of brown/beige adipocyte marker UCP1 by immunohistochemistry in inguinal (top), epididymal (middle), and interscapular (bottom) depots. Epididymal or white visceral and interscapular or brown fat represent negative and positive controls respectively. Data are representative of 10 independent animals per group. Bars represent 100 μm.

Figure 2.

Trauma induces distant browning. (A, upper panel) Representative H&E stained inguinal fat from baseline and one day post trauma (site and contralateral). Trauma site and contralateral showed reduce lipid droplets to form packed multilocular adipocytes. (A, lower panel) Immunostaining using antibody against UCP1 showed protein expression in the traumatized (trauma Site) and contralateral adipose tissue. (B-D) Q-PCR against UCP1, UCP2, PGC1α, CD137 and TBX1 gene in the inguinal adipose depot (B) and against UCP1, UCP2 and Elovl3 gene in the interscapular (C) and perivascular fat depot (D) at baseline and one day after left inguinal trauma (trauma site and contralateral). Expression values were normalized to their respective baseline. Data are mean±SEM of 5–6 independent animals per group. Bars represent 20 μm. *p

Figure 3.

Trauma induces adipose browning up to day 5 following surgery. Representative H&E and UCP1-stained inguinal adipose tissue sections at different time points after surgical trauma as indicated. Data are representative of 4 independent animals per group. Bars represent 20 μm.

Figure 4.

Short-term caloric restriction minimally modulates adipose browning relative to surgical trauma. (A, B) Caloric intake (A) and weight (B) were measured in animal fed with a NC or CR one day before diet randomization (Pre-CR, weight only), one, 2 and 3 days after (CR1, 2 and 3) and at one day post-op (Post-op). Caloric intake is expressed as Kcal/mouse/day and weight as gram. (C) Body temperature was continuously monitored during procedures (15 min) with a rectal probe. T1 (2 ∼ 3 min after the rectal probe insertion), T2 (15 min after the rectal probe insertion), and T3 (the highest temperature during measurement) and is expressed in degrees Celsius (°C). Data are mean±SEM of 6 independent animals per group. *p

Figure 5.

PDGFRα+ cells decrease after surgical injury. (A, B) Cells were isolated from inguinal adipose tissue at baseline and one day post left inguinal injury (trauma site) and analyzed for the expression of PDGFRα, CD34 (A) and Sca1 (B). The percentages of cells are indicated on the flow profile. The graphs show percentage of cells in inguinal depot at baseline and one day after left inguinal trauma (trauma site). Data are mean±SEM of 5–10 independent animals per group. *p < 0.05 versus baseline.

Figure 6.

Surgical trauma promotes local monocyte recruitment and alternatively activated macrophages, but the remote browning can occur without increased macrophages in the browning adipose. (A-D) Cells were isolated from adipose tissue at baseline and one day post left inguinal injury (trauma site) and labeled for flow cytometry. (A) FACS analysis on cell surface marker expression of F4/80 and CD11b at baseline and one day after surgery. The graphs show percentage of F4/80+CD11b+ macrophages in inguinal depot at baseline and one day after left inguinal trauma (trauma site). (B-D) Macrophages were then assessed for the expression of CD11c (B), CD206 (C) and CD301 (D). Data are mean±SEM of 10 independent animals per group. *p < 0.05, **p < 0.01 vs. baseline. (E) Q-PCR against alternatively activated Arg1, Ym1, Fizz1 and against classically activated iNOS, IL-10, TNF macrophages genes in inguinal depot at baseline and one day after left inguinal trauma (trauma site and contralateral). Expression values were normalized to their respective baseline. Data are mean±SEM of 6 independent animals per group. *p < 0.05, **p < 0.01, ***p < 0.001 versus baseline.

Figure 7.

Surgical trauma increases plasma norepinephrine levels. Left panel: Physiological modulation of the sympathetic system in mice one, 2, 8 hours and one day after surgery or incision. NE circulating plasma levels peaked after 8 hours in the surgical trauma group but remained unaltered after incision compared to baseline (0 hour). Right panel: Quantitative assessment of plasma NE expressed as the area under the curve (A.U.C.). Data are mean±SEM of 5 independent animals per group and time point. *p