Phospho-Rb Mediating Cell Cycle Reentry Induces Early Apoptosis Following Oxygen–Glucose Deprivation in Rat Cortical Neurons

Abstract

The aim of this study was to investigate the relationship between cell cycle reentry and apoptosis in cultured cortical neurons following oxygen-glucose deprivation (OGD). We found that the percentage of neurons with BrdU uptake, TUNEL staining, and colocalized BrdU uptake and TUNEL staining was increased relative to control 6, 12 and 24 h after 1 h of OGD. The number of neurons with colocalized BrdU and TUNEL staining was decreased relative to the number of TUNEL-positive neurons at 24 h. The expression of phosphorylated retinoblastoma protein (phospho-Rb) was significantly increased 6, 12 and 24 h after OGD, parallel with the changes in BrdU uptake. Phospho-Rb and TUNEL staining were colocalized in neurons 6 and 12 h after OGD. This colocalization was strikingly decreased 24 h after OGD. Treatment with the cyclin-dependent kinase inhibitor roscovitine (100 μM) decreased the expression of phospho-Rb and reduced neuronal apoptosis in vitro. These results demonstrated that attempted cell cycle reentry with phosphorylation of Rb induce early apoptosis in neurons after OGD and there must be other mechanisms involved in the later stages of neuronal apoptosis besides cell cycle reentry. Phosphoralated Rb may be an important factor which closely associates aberrant cell cycle reentry with the early stages of neuronal apoptosis following ischemia/hypoxia in vitro, and pharmacological interventions for neuroprotection may be useful directed at this keypoint.

RB controls growth, survival, and neuronal migration in human cerebral organoids

“…In addition to the increase in the NSC death, we also found more apoptotic neuroblasts and neurons, represented by AC3 + DCX + cells and AC3 + Tuj1 + cells at 15 and 28 DIV, respectively. Similar results were previously observed in mice (Yu et al, 2012;Vandenbosch et al, 2016). In addition, there was no significant change in S-phase entry in RB1-KO organoids compared with wild type at 15 DIV, which is in line with no difference in organoid size between wild type and RB1-KO at this stage.…”

“…What is surprising is that there was also increased cell death in RB1-KO organoids compared with wild type at 28 DIV, possibly due to an increase of the proapoptotic gene BAX (Drew et al, 2016). In line with our results, it has been described that loss of RB in mice promotes apoptosis in the developing and adult brain (Macleod et al, 1996;Vandenbosch et al, 2016;Yu et al, 2012). However, other studies have shown that an increase in cell proliferation due to the lack of RB is not followed by enhanced cell death (MacPherson et al, 2003).…”

“…Moreover, KO mouse models have revealed the role of RB during neuronal differentiation and migration, as well as in the regulation of cell death in the embryo and adult brain (Andrusiak et al, 2011;Ghanem et al, 2012;McClellan et al, 2007;Christie et al, 2014;Ferguson et al, 2005;Macleod et al, 1996;Vandenbosch et al, 2016;Yu et al, 2012). The finding that lack of the RB1 gene is associated with structural brain abnormalities in human (Mitter et al, 2011;Rodjan et al, 2010), and inactivation of RB family proteins affects cell cycle and cell death in human ESCs (Conklin et al, 2012), suggest that RB might have a role during human brain development.…”

“…RB loss has been related to an increase of cell death in different areas of the mouse brain during development (Macleod et al, 1996;Vandenbosch et al, 2016;Yu et al, 2012). We therefore analyzed whether the absence of RB promotes cell death using a marker of apoptosis (cleaved caspase 3, AC3 Fig.…”

“…Recently, studies of brain-specific Rb1-KO mice revealed other roles of the Rb1 gene that extend beyond cell cycle control, such as regulation of neuronal differentiation and migration (Andrusiak et al, 2011;Ghanem et al, 2012;McClellan et al, 2007;Christie et al, 2014;Ferguson et al, 2005). Moreover, the lack of RB promotes apoptosis in some cell types, but not in others, through p53-dependent or -independent pathways (Macleod et al, 1996;Vandenbosch et al, 2016;Yu et al, 2012).…”