129s2 svpascrl mice

All animal procedures conformed to European Union laws and guidelines for animal care, and experimental procedures were approved by the ethics committee of the Universidad Rey Juan Carlos (Spain). Male 129S2/SvPasCrl mice (Charles River Laboratories) were used for all studies and were housed individually in climate‐controlled quarters (21ºC) with a 12‐hr light/dark cycle. Food (13% calories derived from fat; Research diet#2014C, Envigo) and water were available ad libitum unless otherwise stated. In some experiments, mice were randomly assigned to undergo 20% CR from 3 months of age, as described earlier (Sierra Rojas et al., 2016; Speakman & Mitchell, 2011). The restricted food was administrated at 19 hr just before the dark period starts (from 20 to 8 hr). For cold exposure, animals were housed in their individual cages in a cold chamber (4ºC) for 24 hr. Animals were sacrificed by cervical dislocation at fed state, and selected tissues were kept for analysis.

All experimental procedures were approved by the Institutional Animal Care and Use Committees of Boys Town National Research Hospital and the University of Nebraska Medical Center in accordance with NIH guidelines. Experiments were carried out in six- to ten-week-old female 129S2/SvPasCrl mice purchased from Charles River Laboratories (Wilmington, MA).

PodoPalldBL/6-/- mice were generated as described previously [23 (link)]. These mice have a C57BL/6 background and were backcrossed to the 129 background using 129S2/SvPasCrl mice (Charles River Laboratories, Wilmington, MA, USA) and the “speed congenics” approach [27 (link)]. After 8x backcrossing, the new mouse line was established.
Podocyte-specific palladin knockout mice with 129 background (PodoPalld129-/-) and heterozygous Cre-recombinase expression were used for experiments. Mice without Cre-recombinase expression were used as controls (PodoPalld129+/+). Experiments were done with 6 and 12 months old male PodoPalld mice (at least n = 3 of each group). Genotyping of mice was performed with Phire^® Animal Tissue Direct PCR Kit (Finnzymes/Thermo Fisher Scientific) in accordance to the manufacturer’s instructions using primers shown in S1 Table.
All prerequisites of the German animal protection law were met and experiments were performed in accordance with the guidelines of the federal agencies in Mecklenburg-Western Pomerania (LALLF M-V). The responsible ethics committee within the LALLF M-V approved the experiments with mice. For kidney removal, mice were sacrificed by the use of barbiturate.