Edta containing tubes

All animal experiments were approved by the animal welfare committee of Wageningen University (2016.W-0093.015). Hilpda^ΔLysM and Hilpda^fl/fl mice were bred as described before (16 (link)). Mice had unlimited access to food (standard chow) and water and were housed under normal light–dark cycles in humidity- and temperature-controlled specific pathogen-free conditions. Both male and female mice were used for the isolation of primary cell cultures. For the in vivo study, male Hilpda^ΔLysM and Hilpda^fl/fl mice, aged 9 to 12 wk, were housed with two or three littermates per cage and randomly allocated to the different time points. Mice were weighed prior to injection to calculate specific concentrations of LPS. At each time point, eight mice per genotype received an intraperitoneal injection of LPS (from E. coli O55:B5; L6529; Sigma-Aldrich) at a concentration of 2 mg/kg body weight in an end volume of 200 µL sterile saline solution. Control groups were injected with 200 µL sterile saline solution. After the indicated time points, mice were anesthetized with isoflurane and blood was collected via orbital puncture in ethylenediaminetetraacetate (EDTA)-containing tubes (Sarstedt). After blood collection, mice were immediately killed by cervical dislocation. Subsequently, peritoneal macrophages and tissues were harvested.

Rats were single‐housed in BIODAQ cages as described earlier. Food intake and body weight were measured daily and tumour growth was induced in all animals. Blood was obtained by puncturing the sublingual vein under mild isoflurane anaesthesia. Sampling was conducted 3 days before tumour induction (baseline) and 11 days and 17 days after induction, that is, shortly after the onset of anorexia and during fully developed anorexia, respectively. Blood was collected in EDTA containing tubes (Sarstedt, Nümbrecht, Germany) and centrifuged at 7000 × g (4°C, 7 min) to obtain plasma, which was stored in aliquots at −80°C for subsequent analysis. The levels MIC‐1 were measured with using an ELISA (R&D Systems, USA) according to the manufacturers' instructions.

This experimental study was conducted at the cellular and molecular research center of Shahrekord University of Medical Sciences from February 2011 to January 2012. In the present study, we investigated the mutations of the TMC1 gene, locus DFNB7/11, in a cohort of 100 patients with NSHL in Iran. The 890 blood samples of families with Iranian origin was obtained in ethylene diamine tetra-acetic acid (EDTA)-containing tubes (Sarstedt) from 10 provinces of Iran, namely Semnan, Sistan & Baloochestan, Fars, Khozestan, Kohgilooye Va Boyer Ahmad, Kordestan, Chaharmahal & Bakhtiari, Booshehr, Golestan, and Gilan. Finally, 100 patients (one proband from each family) were selected (Table 1). NSHL informational questionnaires were filled out for all families. In previous work, these patients had no mutations in the cx26 gene (10 ). The blood samples were stored at -20°C until further processing. Known environmental risk factors such as head trauma and use of ototoxic drugs could affect the study, so families with the possibility of exposure to these factors were excluded from the research.