Z gel 1.1 ml serum preparation tubes

Blood serum was isolated using Z-Gel 1.1 ml serum preparation tubes (41.1378.005, Sarstedt) according to the manufacturer’s instructions. Tissue for protein measurements was snap frozen in liquid nitrogen. 50–100 mg tissue were added to 10× the weight T-PER Tissue Protein Extraction Reagent (78510, ThermoFisher) containing Halt Proteinase and Phosphatase inhibitor cocktail (78440, ThermoFisher). Tissue was homogenized using a 5 mm steel bead (Qiagen) and a TissueLyser II (Qiagen) at 30 hz for 2 min (spleen, liver, adipose tissue, pancreas, thymus, kidney), 5 min (muscle, heart, lung, cecum, large intestine, small intestine) or 10 min (skin, stomach). Debris was pelleted and supernatant used for assays. Bone marrow was prepared by flushing 2 femurs with 500 ul PBS. After pelleting the cells, supernatant was used for BMEF measurements, while cells were taken up in T-PER lysis buffer, underwent one freeze-thaw cycle and debris was pelleted. For measurement of serum and tissue chemokine levels commercially available ELISA kits for CCL2/JE (MJE00, R&D) and CXCL12/SF-1 (MCX120, R&D) were used. For measurement of blood stress hormone corticosterone an ELISA kit was used (ADI-900–097, Enzo). Multiplex was performed using a kit for mouse metabolic hormones (MMHMAG-44K, Millipore) and human metabolic hormones (HMEMAG-34K, Millipore).