Nzm2410 mice

Mice were maintained at the Ralph H. Johnson VAMC Animal Facility (Charleston, SC). NZM2410 mice were acquired from The Jackson Laboratory (Bar Harbor, ME, USA). ERαKO mice (ERα short) on the C57BL/6 background were a kind gift of Dr. Ken Korach (NIEHS) and are also available commercially (Stock No. 004744, The Jackson Laboratory). Mice carrying this Esr1^tm1Ksk allele express a truncated form of ERα (ERα short), the result of alternate splicing of the transcript [14 (link)]. In contrast, mice carrying the Esr1^tm4.2Ksk allele (Stock No. 026176, The Jackson Laboratory) are ERα null, and have no tissue responses to estrogen or estrogen receptor alpha activity [16 (link)]. All experimental mice (n=44) were female and littermates when possible. All mice were ovariectomized (OVX) pre-disease at 4–8 weeks of age (peri-puberty). Two groups subsequently received 0.25 mg, 90-day sustained release 17β-estradiol pellet, implanted sub-dermally x 2 occurrences (180d) to ensure continuous systemic E2 levels in adult mice (Innovative Research of America, Sarasota, FL, USA).

ERα deficient (ERαKO) mice on the C57BL/6 background (kind gift of Dr. Ken Korach, NIEHS, RTP, NC) were backcrossed for 12 generations to NZM2410 mice (Jackson Laboratory, Bar Harbor, ME) and congenic status was verified as previously described (21 (link)). Female mice between 10 and 14 weeks of age were used for these experiments. All mice were maintained at the Ralph H. Johnson VAMC Animal Care Facility (Charleston, SC) using Institutional Animal Care and Use Committee approved protocols #421 and #498 originally approved August 2008 and 2011 respectively.

Mice were maintained at the Ralph H. Johnson VAMC Animal Facility (Charleston, SC). Animal protocols followed the principles outlined in the Guide for the Care and Use of Laboratory Animals, and were approved by MUSC’s and the VA’s IACUC. The NZM2410 mice were acquired from Jackson Laboratory (Bar Harbor, ME, USA), the ERα−/− C57BL/6 mouse strain was a kind gift of Dr. Ken Korach. The two strains were backcrossed for >10 generations to create the ERα−/− NZM2410 mouse (NZM ERα −/−). They were maintained on a 12 hr. light/dark cycle with access to food and water ad libitum. All experimental mice (n= 51) were female and were littermates when possible. Two cohorts (NZM WT and NZM ERα −/−) were unmanipulated. All other mice were ovariectomized (OVX) at 4 weeks of age, before puberty, and 2 groups subsequently received 0.1 mg, 90-day sustained release 17β-estradiol pellet, implanted sub-dermally (Innovative Research of America, Sarasota, FL, USA). Mice were sacrificed by cervical dislocation following induction of anesthesia by isoflurane at 32 weeks of age or when they reached pre-determined sacrifice requirements (>10% loss of weight, >500mg urine protein as assessed by dipstick, or upon recommendation by the animal facility veterinarian).