Title: ERα Signaling Is Required for TrkB-Mediated Hippocampal Neuroprotection in Female Neonatal Mice after Hypoxic Ischemic Encephalopathy
Legend: Increases in p-TrkB immuno and protein expressions are sexually differentiated and ERα dependent post-HI. A. Changes in MAP2 and p-TrkBY515 immunoexpression in the CL and IL hippocampi of ERα+/+ and ERα-/- P9 male and female mice were examined 3 days post HI. MAP2 (red), p-TrkBY515 (green) and DAPI (blue). Arrow: increased p-TrkBY515 staining Arrowhead: decreased p-TrkBY515 in ERα-/- hippocampi. Inset: primary antibody control. B. Summary figure showing the percent change in IL/CL ratio of hippocampal mean p-TrkBY515 fluorescent intensities in ERα+/+ and ERα-/- male and female mice hippocampi 3 days post-HI. IL/CL ratio of p-TrkBY515 obtained from uninjured naïve mouse hippocampus was assumed to be 1:1. Three hippocampal slices per brain were analyzed. Data are mean ± SEM. n = 8 for ERα+/+ and n=3 for ERα-/-. n = number of pups. ERα+/+ and ERα-/- mice were subjected to HI without (C) and with (D) 7,8-DHF therapy. Hippocampi from the CL and IL sides were obtained 3 days post-HI and blots were probed for p-TrkBY705 expression (upper), f-TrkB, t-TrkB, and i-TrkB. E. Summary figure of the hippocampal p-TrkB/f-TrkB ratio in ERα+/+ and ERα-/- mice with and without 7,8-DHF therapy 3 days post-HI. Data are mean IL adjusted for CL ± SEM, n = 3. n = number blots per group. p = 0.018 for *, p= 0.035 for +, p= 0.334 for ε, p = 0.0488 for τ versus ERα+/+.
Citation: Cikla U, Chanana V, Kintner DB, Udho E, Eickhoff J, Sun W, Marquez S, Covert L, Otles A, Shapiro RA, Ferrazzano P, Vemuganti R, Levine JE, Cengiz P (2016) ERα Signaling is required for TrkB-Mediated hippocampal neuroprotection in female neonatal mice after Hypoxic Ischemic Encephalopathy. eNeuro.28;3(1). pii: ENEURO.0025-15.2015. doi:10.1523/ENEURO.0025-15.2015.
Abstract: Male neonate brains are more susceptible to the effects of perinatal asphyxia resulting in hypoxia and ischemia (HI)-related brain injury. The relative resistance of female neonatal brains to adverse consequences of HI suggests that there are sex-specific mechanisms that afford females greater neuroprotection and/or facilitate recovery post-HI. We hypothesized that HI preferentially induces estrogen receptor α (ERα) expression in female neonatal hippocampi and that ERα is coupled to Src family kinase (SFK) activation that in turn augments phosphorylation of TrkB and thereby results in decreased apoptosis. After inducing the Vannucci’s HI model on P9 (C57BL/6J) mice, female and male ERα wild-type (ERα+/+) or ERα null mutant (ERα-/-) mice received vehicle control or the selective TrkB agonist 7,8-dihydroxyflavone (7,8-DHF). Hippocampi were collected for analysis of ERα and BDNF mRNA, and protein levels of ERα, p-TrkB, p-src, and cleaved caspase 3 (c-caspase-3) post-HI. Our results demonstrate that: (1) HI differentially induces ERα expression in the hippocampus of the female versus male neonate, (2) src and TrkB phosphorylation post-HI is greater in females than in males after 7,8-DHF therapy, (3) src and TrkB phosphorylation post-HI depend on the presence of ERα, and (4) TrkB agonist therapy decreases c-caspase-3 levels only in ERα+/+ female mice hippocampus. Together, these observations provide evidence that female-specific induction of ERα expression confers neuroprotection with TrkB agonist therapy via SFK activation and accounts for improved functional outcomes in female neonates post-HI.
About the investigator: Cengiz’s research focuses on finding a novel therapy for neonatal encephalopathy subsequent to hypoxia ischemia. Hypoxia ischemia (HI)-related brain injury is associated with high mortality and morbidity. One of every three neonates with HI develops chronic morbidity including cerebral palsy, intellectual and developmental disabilities, learning disabilities or epilepsy. Moderate hypothermia has been shown to improve the neurological outcome of term neonates. However, term neonates after 6 hours of life and pre-term neonates do not qualify for hypothermia therapy. Her research continues to focus on exploring a pharmacological therapy as an adjunct to hypothermia and for those neonates who are not eligible for hypothermia therapy. Cengiz is particularly interested in targeting membrane proteins that are either detrimental or beneficial to neuronal survival after hypoxia and ischemia. She is working both with in vitro and in vivo models of hypoxia and ischemia in order to identify the role of these proteins and their effects on neuronal degeneration. Using Vannucci-Rice’s neonatal HI model in nine-day-old mice, she has shown neuroprotection at 72 hours after HI by pharmacologically inhibiting the membrane protein Na+/H+ exchanger isoform-1. Her goal is to establish the mechanism(s) that leads to hippocampal neuroprotection by characterizing the function of Na+/H+ exchanger protein isoform-1 after HI. Her ultimate goal is to translate these preclinical findings into clinical studies that will benefit patients with neonatal encephalopathy after hypoxia and ischemia.