Cyclophosphamide-induced agenesis of cerebral aqueduct resulting in hydrocephalus in mice

The present work was undertaken to reveal the mechanism of cerebral aqueduct agenesis found to result in hydrocephalus following intrauterine exposure to model teratogen, cyclophosphamide, in murine fetuses. A single dose of 10-mg/kg body weight cyclophosphamide was injected intaperitoneally to pregnant mice on day 10, 11 or 12 of gestation. Fetuses were collected through abdominal incision on day 18 and studied for various malformations of brain and cranium including hydrocephalus. Incomplete development and failure of canalization of the cerebral aqueduct were detected when serial sections of brain in coronal and transverse planes were studied under the microscope. Biotechnological investigations such as % DNA fragmentation, % viable cell count and cell proliferation assay were carried out on brain cells for further studies. Agenesis and non-canalization of the cerebral aqueduct resulted in increased pressure of CSF, which led to rupture of the aqueduct complicated by leakage and accumulation of CSF in brain substance forming a cavity containing CSF parallel and lateral to the unopened part of the cerebral aqueduct. Incomplete development along with non-canalization of the cerebral aqueduct resulted in blockage of CSF flow through the ventricles that manifest as internal hydrocephalus. External hydrocephalus on the other hand was detected where the CSF accumulated in the cavity formed inside the brain substance and established communication with the CSF in the subarachnoid space. Cyclophosphamide induced inhibition of mitosis and cell differentiation of ependymal cells reflecting a decreased % viable cell count and cell proliferation assay along with augmentation of apoptosis of brain cells quantified as increased % DNA fragmentation count, which were identified as the contributing factors underlying the agenesis and incomplete development of the cerebral aqueduct. The study also suggests that cell survival, proliferation, migration or differentiation of ependymal cells might have been affected, and we speculate that CSF may have an inducing role in the development and canalization of the cerebral aqueduct.