The TRPM2 ion channel regulates the browning process in human adipose tissue

Background. Adipose tissue (AT) is important for energy balance, glucose and lipid homeostasis and thermogenesis. During thermogenesis, AT becomes more active and enhances oxidative metabolism. This process mainly occurs in brown AT (BAT). The promotion of this process in white AT (WAT) is called “browning” and, together with BAT activation, is considered as a promising approach to counteract obesity and metabolic diseases. Transient receptor potential cation channel, subfamily M, member 2 (TRPM2) is an ion channel that allows Ca2+ influx into the cytosol from the extracellular space, and is gated mainly by ADPR, produced from NAD+ degradation by CD38 and/or poly-ADPR polymer degradation by poly-ADPR hydrolases (such as MacroD1 and MacroD2). In mice, Trpm2 is overexpressed and ADPR levels are higher in response to cold exposure and Trpm2-/- mice exposed to cold display lower levels of browning markers compared to WT mice, pointing to TRPM2 as a fundamental player in BAT activation and WAT browning in mice(1). Upon cold exposure, ADPR is likely produced in BAT and WAT by the up-regulation of poly ADPR hydrolases, and not by CD38(1). Aim. The aim of this study is to investigate the relevance of TRPM2 in human BAT activation and WAT “browning”. Methods. Human immortalized white pre-adipocytes (hTERT-WA) or brown pre adipocytes (hTERT-BA) were differentiated in vitro into white or brown adipocytes, respectively, for 12 days, using appropriate cocktails. For browning inducement, white adipocytes were cultured in the presence of rosiglitazone for 3 days. White adipocytes were stimulated with 100 nM isoprenaline or with 100 µM forskolin, in the presence or absence of the TRPM2 antagonist JNJ-28583113 (10 μM). Gene expression was evaluated by qPCR and changes in the [Ca2+]i were monitored in Fluo-3- or Fura2 loaded cells. Gene expression from human BAT and WAT biopsies was analyzed by next Generation sequencing. BAT glucose uptake rate was measured by PET-CT imaging. Fasting blood samples were utilized for measuring plasma concentrations of total cholesterol, High Density Lipoprotein (HDL) cholesterol and triglycerides (ModularP800, Roche Diagnostics GmbH). Human cohort is described in Mueez U-Din et al 2023(2). Results. TRPM2 expression increased in hTERT-WA and hTERT-BA following differentiation by 70% and 90%, respectively, and it was further increased by an additional 20% in hTERT-WA after browning inducement. TRPM2 expression was increased in differentiated hTERT-WA following stimulation with isoprenaline or with forskolin, which increases intracellular cAMP by activating adenylyl cyclase. This result indicates that TRPM2 expression occurs downstream of cAMP increase, upon β-adrenergic stimulation, controlling the browning process. The expression of the browning marker genes UCP1 and PGC1α was increased upon stimulation with forskolin and with isoprenaline; the presence of the TRPM2 inhibitor JNJ-28583113 significantly attenuated the browning markers overexpression (by approximately 40%) and the [Ca2+]i rise upon forskolin stimulation. Analysis conducted on human biopsies showed that TRPM2 was overexpressed in BAT compared to WAT. In BAT, the expression of MACROD1 and MACROD2, which are poly-ADPR hydrolases generating the TRPM2 agonist ADPR, positively correlated with the browning marker UCP1 and with glucose uptake upon cold stimulus, suggesting that poly-ADPR polymer degradation is a fundamental process also in human BAT activation. In BAT, TRPM2 expression inversely correlated with glucose uptake upon cold exposure, as well as with the expression of the glucose transporter GLUT4. These unexpected observations might suggest that TRPM2 is regulating lipid usage for energy production instead of glucose. Interestingly, TRPM2 expression in BAT positively correlated with serum concentrations of: triglycerides in HDL, whereas TRPM2 expression in WAT positively correlated with medium and large HDL particles, with total lipids, total cholesterol and triglycerides in medium and large HDL particles. Conclusion. Our data suggest that TRPM2 may play an important role in human BAT activation and WAT browning. Pharmacological activation of TRMP2 could present potential benefits for the treatment of obesity. References 1. Benzi A, Heine M, Spinelli S, Salis A, Worthmann A, Diercks B, Astigiano C, Pérez Mato R, Memushaj A, Sturla L, Vellone V, Damonte G, Jaeckstein MY, Koch-Nolte F, Mittrücker HW, Guse AH, De Flora A, Heeren J, Bruzzone S. Front Endocrinol (2024) 14:1251351 2. U-Din M, de Mello VD, Tuomainen M, Raiko J, Niemi T, Fromme T, Klåvus A, Gautier N, Haimilahti K, Lehtonen M, Kristiansen K, Newman JW, Pietiläinen KH, Pihlajamäki J, Amri EZ, Klingenspor M, Nuutila P, Pirinen E, Hanhineva K, Virtanen KA. Cell Rep. (2023) 42(9):113131.