Biology subjects

Gearhart, P. J.

Publications and source records attributed to Gearhart, P. J..

2 recordsLinked to original sources

bioRxiv2025

Generation and characterization of a tamoxifen-inducible, Cre driver rat for transgene expression in microglia

Microglia are the resident immune cells of the central nervous system (CNS) and display diverse functions under both physiological and pathological conditions. The past decade has seen burgeoning interest in microglia function, with a variety of transgenic tools developed for specific genetic manipulation of microglia in various injury, disease, and developmental models. Although the majority of models have been developed in mice, the ability to manipulate microglia in rats provides additional advantages to studying microglial function in the brain especially related to complex behavior. Using BAC transgenesis, our lab has created a transgenic rat (Cx3cr1-CreERT2) that expresses a tamoxifen inducible Cre recombinase (CreERT2) under control of the microglial/macrophage specific fractalkine C-X3-C Motif Chemokine Receptor 1 (Cx3cr1) promoter. In mice, CreERT2 and other transgenes have been expressed in microglia using the Cx3cr1 promoter, however, this is the first demonstration in rats. Importantly, these rats exhibit similar cognitive behaviors compared to their wildtype (WT) controls. Microglial specificity of inducible Cre expression was confirmed by breeding the novel Cx3cr1-CreERT2+/- rat with a previously reported double floxed inverse open reading frame (DIO)-mCherry+/- reporter rat to show tamoxifen inducible mCherry expression that colocalizes with the microglial marker Iba1. In addition, we utilize flow cytometry to demonstrate time and Cre dependent differences in recombination of Cx3cr1+ cells in the spleen, peripheral blood, and brain at two- and eight-weeks post-tamoxifen treatment. Overall, we have created a novel transgenic rat model for researchers to employ in understanding microglial and peripheral immune cell function in rats.

neuroscience↗

bioRxiv2022

Greater breadth of vaccine-induced immunity in females than males is mediated by increased antibody diversity in germinal center B cells

Inactivated influenza vaccines induce greater antibody responses in females than males among both humans and mice. To test the breadth of protection, we used recombinant mouse-adapted A/California/2009 (maA/Cal/09) H1N1 viruses containing mutations at one (1M), two (2M), or three (3M) antigenic sites, in addition to a virus containing the 1M mutation and a substitution of the Ca2 antigenic site (Sub) with one derived from an H5 hemagglutinin (HA) to challenge mice of both sexes. Following maA/Cal/09 vaccination, females produced greater virus-specific class-switched IgG and IgG2c antibodies against the vaccine and all mutant viruses, and antibodies from females recognized more unique, linear HA epitopes than antibodies from males. While females had greater neutralizing antibody (nAb) titers against the vaccine virus, both sexes showed lower neutralization capacity against mutant viruses. After virus challenge, vaccinated females had lower pulmonary virus titers and reduced morbidity than males against the 1M and 2M viruses, but not the Sub virus. Females generated greater numbers of germinal center (GC) B cells containing superior somatic hypermutation frequencies than vaccinated males. Deletion of activation-induced cytidine deaminase (Aicda) eliminated female-biased immunity and protection against the 2M virus. Harnessing methods to improve GC B cell responses and frequencies of somatic hypermutations, especially in males, should be considered in the development of universal influenza vaccines. SummaryCompared with males, inactivated influenza vaccination of female mice causes greater production of class-switched, somatically-hypermutated antibodies and a more robust germinal center B cell response, leading to more diverse H1N1 antigen recognition and better protection against mutant influenza A viruses.

immunology↗