PPARγ Is Activated during Congenital Cytomegalovirus Infection and Inhibits Neuronogenesis from Human Neural Stem Cells

Abstract

Congenital infection by human cytomegalovirus (HCMV) is a leading cause of permanent sequelae of the central nervous system, including sensorineural deafness, cerebral palsies or devastating neurodevelopmental abnormalities (0.1% of all births). To gain insight on the impact of HCMV on neuronal development, we used both neural stem cells from human embryonic stem cells (NSC) and brain sections from infected fetuses and investigated the outcomes of infection on Peroxisome Proliferator-Activated Receptor gamma (PPARγ), a transcription factor critical in the developing brain. We observed that HCMV infection dramatically impaired the rate of neuronogenesis and strongly increased PPARγ levels and activity. Consistent with these findings, levels of 9-hydroxyoctadecadienoic acid (9-HODE), a known PPARγ agonist, were significantly increased in infected NSCs. Likewise, exposure of uninfected NSCs to 9-HODE recapitulated the effect of infection on PPARγ activity. It also increased the rate of cells expressing the IE antigen in HCMV-infected NSCs. Further, we demonstrated that (1) pharmacological activation of ectopically expressed PPARγ was sufficient to induce impaired neuronogenesis of uninfected NSCs, (2) treatment of uninfected NSCs with 9-HODE impaired NSC differentiation and (3) treatment of HCMV-infected NSCs with the PPARγ inhibitor T0070907 restored a normal rate of differentiation. The role of PPARγ in the disease phenotype was strongly supported by the immunodetection of nuclear PPARγ in brain germinative zones of congenitally infected fetuses (N = 20), but not in control samples. Altogether, our findings reveal a key role for PPARγ in neurogenesis and in the pathophysiology of HCMV congenital infection. They also pave the way to the identification of PPARγ gene targets in the infected brain.

Peroxisome proliferator-activated receptor γ (PPARγ) activation: A key determinant of neuropathogeny during congenital infection by cytomegalovirus

“…7 In our study, we explored the expression of the immediate early HCMV antigen (IE), a factor encoded by the HCMV genome and critical for virus replication, in histopathological slides from deceased infected or non-infected fetus. 8 In agreement with the studies in the mouse model, we observed cells clearly immunoreactive to IE in the ependymal and germinative zones of the brain of infected cases. However, no labeling was found in the white matter, whereas, as expected, brain vessels were positive to IE.…”

“…Last, we explored the relevance of these findings in vivo, through immunohistological examination of brain slices from a set of infected (N D 20) or uninfected control (N D 5) fetus of different gestational ages. Strinkingly, nuclear PPARg was immunodetected in the brain germinative zones of congenitally infected fetuses, but in none of the control samples 8 (Fig. 3), what strongly supported the role of PPARg in the disease phenotype.…”

“…Thus we demonstrated that (1) pharmacological activation of PPARg ectopically expressed through lentiviral transduction under the control of the EF1a minimal promoter was sufficient to induce impaired neuronogenesis of uninfected NSCs, (2) treatment of uninfected NSCs with 9-HODE impaired NSC differentiation and (3) treatment of HCMV-infected NSCs with the PPARg inhibitor T0070907 restored a normal rate of differentiation. 8 These findings revealed that PPARg activation was necessary and sufficient to impair NSC differentiation into neurons. Last, we explored the relevance of these findings in vivo, through immunohistological examination of brain slices from a set of infected (N D 20) or uninfected control (N D 5) fetus of different gestational ages.…”

“…8 Conversely, HCMV infection strongly increased PPARg levels and activity in NSCs, as shown by immunofluorescence analysis, western blot, luciferase reporter assays and chromatin immunoprecipitation. 8 Notably, we found that infected NSCs exerted a bystander effect on neighboring uninfected cells, which also showed enhanced PPARg levels. Levels of 9-hydroxyoctadecadienoic acid (9-HODE), a known PPARg agonist, were significantly increased in infected NSC cultures.…”

“…2) and that infection dramatically impaired NSC differentiation into neurons. 8 In the search for the molecular bases of such inhibition of differentiation, we next investigated the levels and activity of the nuclear receptor PPAR gamma (PPARg) in infected NSCs.…”