Progesterone treatment normalizes the levels of cell proliferation and cell death in the dentate gyrus of the hippocampus after traumatic brain injury

Abstract

Traumatic brain injury (TBI) increases cell death in the hippocampus and impairs hippocampus-dependent cognition. The hippocampus is also the site of ongoing neurogenesis throughout the lifespan. Progesterone treatment improves behavioral recovery and reduces inflammation, apoptosis, lesion volume, and edema, when given after TBI. The aim of the present study was to determine whether progesterone altered cell proliferation and short-term survival in the dentate gyrus after TBI. Male Sprague-Dawley rats with bilateral contusions of the frontal cortex or sham operations received progesterone or vehicle at 1 and 6 h post-surgery and daily through post-surgery Day 7, and a single injection of bromodeoxyuridine (BrdU) 48 h after injury. Brains were then processed for Ki67 (endogenous marker of cell proliferation), BrdU (short-term cell survival), doublecortin (endogenous marker of immature neurons), and Fluoro-Jade B (marker of degenerating neurons). TBI increased cell proliferation compared to shams and progesterone normalized cell proliferation in injured rats. Progesterone alone increased cell proliferation in intact rats. Interestingly, injury and/or progesterone treatment did not influence short-term cell survival of BrdU-ir cells. All treatments increased the percentage of BrdU-ir cells that were co-labeled with doublecortin (an immature neuronal marker in this case labeling new neurons that survived 5 days), indicating that cell fate is influenced independently by TBI and progesterone treatment. The number of immature neurons that survived 5 days was increased following TBI, but progesterone treatment reduced this effect. Furthermore, TBI increased cell death and progesterone treatment reduced cell death to levels seen in intact rats. Together these findings suggest that progesterone treatment after TBI normalizes the levels of cell proliferation and cell death in the dentate gyrus of the hippocampus.

Figure 1

Depicts the procedural timeline used in this experiment.

Figure 2

Panel A shows Ki67-ir cells in the granule cell layer. Panel B shows BrdU-ir cells in the granule cell layer. Panel C shows a BrdU (red) and doublecortin (green) double labelled cell in the granule cell layer. Photomicrographs are magnified 1000x. 1 cm scale bar = 10 μm.

Figure 3

A) Mean (+ SEM) total number of Ki67-ir cells in the granule cell layer (GCL) and hilus of the dentate gyrus. The sham group had significantly fewer Ki67-ir cells compared to the sham + progesterone group and the traumatic brain injury (TBI) group and had significantly more Ki67-ir cells compared to the TBI + progesterone group. The TBI + progesterone group had significantly fewer Ki67-ir cells than the TBI group and the sham + progesterone group. There were no significant differences between groups in the hilus. B) Mean (+ SEM) total number of BrdU-ir cells in the GCL and hilus of the dentate gyrus. Groups did not differ in the total number of BrdU-ir cells in the GCL or the hilus. C) Mean (+ SEM) total number of immature neurons surviving for 6 days. The TBI group had significantly more immature neurons compared to the sham group. All other groups did not differ from the sham group. a denotes significantly different from the sham group (p < 0.05), b denotes significantly different from the TBI group and the sham + progesterone group (p < 0.05).

Figure 4

Mean (+ SEM) total number of Fluoro-Jade B labelled cells in the granule cell layer (GCL) of the dentate gyrus and the CA1 region of the hippocampus. The traumatic brain injury (TBI) group had significantly more Fluoro-Jade B labelled cells in the GCL compared to the sham group, the sham + progesterone group, and the TBI + progesterone group. The TBI group tended to have more Fluoro-Jade B labelled cells in the CA1 region compared to the sham group and the sham + progesterone group. a denotes significantly different from the TBI group (p < 0.05).