Intracerebral Transplantation of Adult Mouse Neural Progenitor Cells into the Niemann-Pick-A Mouse Leads to a Marked Decrease in Lysosomal Storage Pathology

Abstract

Niemann-Pick disease is caused by a genetic deficiency in acid sphingomyelinase (ASM) leading to the intracellular accumulation of sphingomyelin and cholesterol in lysosomes. In the present study, we evaluated the effects of direct intracerebral transplantation of neural progenitor cells (NPCs) on the brain storage pathology in the ASM knock-out (ASMKO) mouse model of Type A Niemann-Pick disease. NPCs derived from adult mouse brain were genetically modified to express human ASM (hASM) and were transplanted into multiple regions of the ASMKO mouse brain. Transplanted NPCs survived, migrated, and showed region-specific differentiation in the host brain up to 10 weeks after transplantation (the longest time point examined). In vitro, gene-modified NPCs expressed up to 10 times more and released five times more ASM activity into the culture media compared with nontransduced NPCs. In vivo, transplanted cells expressed hASM at levels that were barely detectable by immunostaining but were sufficient for uptake and cross-correction of host cells, leading to reversal of distended lysosomal pathology and regional clearance of sphingomyelin and cholesterol storage. Within the host brain, the area of correction closely overlapped with the distribution of the hASM-modified NPCs. No correction of pathology occurred in brain regions that received transplants of nontransduced NPCs. These results indicate that the presence of transduced NPCs releasing low levels of hASM within the ASMKO mouse brain is necessary and sufficient to reverse lysosomal storage pathology. Potentially, NPCs may serve as a useful gene transfer vehicle for the treatment of CNS pathology in other lysosomal storage diseases and neurodegenerative disorders.

Gene transfer of human acid sphingomyelinase corrects neuropathology and motor deficits in a mouse model of Niemann-Pick type A disease

“…7 and 8, which are published as supporting information on the PNAS web site) overlapped with areas that were positive for hASM immunostaining, indicating that each serotype vector was capable of generating a functional transgene product. As previously demonstrated (3,18), cholesterol clearance also occurred in areas anatomically connected with the injection site that did not stain positively for hASM. METAMORPH analysis indicated that a reduction in filipin staining occurred throughout the entire rostral-caudal axis.…”

“…Mice that were unilaterally injected with different AAV serotypes encoding for hASM were killed 7 weeks after injection (14 weeks of age) with euthasol (150 mg͞kg i.p.) and then either decapitated (n ϭ 5 per group) or transcardially perfused (n ϭ 3 per group) with fixative as described (18). Brains from decapitated mice were rapidly removed, snapfrozen in liquid nitrogen, dissected into three sections [right cerebral hemisphere, left cerebral hemisphere, and cerebellum (including the brainstem)], homogenized, and analyzed for hASM by ELISA (18).…”

“…Fifty-micrometer Vibratome sections of brains and spinal cords from perfused mice were processed for hASM expression, cholesterol accumulation with filipin staining, and Purkinje cell survival with calbindin staining as described (3,18). Sections stained with filipin were evaluated for the percent reduction in fluorescent deposits using a METAMORPH Image Analysis System (Universal Imaging, Downington, PA) as described (18). A second cohort of ASMKO mice received bilateral injections (4 l per site) aimed at the DCN to determine whether increasing hASM distribution within the cerebellum leads to further improvement in rotarod performance.…”

“…Section 5 contained the cerebellum (i.e., injection site) and brainstem. The right hemisphere was used to quantify brain SPM levels (19) and the left hemisphere was used to determine hASM levels (18).…”

High sphingomyelin levels induce lysosomal damage and autophagy dysfunction in Niemann Pick disease type A

“…Thus, SM has a strong affinity for cholesterol, reflected in its tight association in signaling platforms in the plasma membrane and intracellular organelles. Moreover, ASMko mice brains show cholesterol deposits 29,30 and the accumulation of this lipid impairs lysosomal function in different LSDs. 21,22,24 To test cholesterol involvement, NPA fibroblasts were incubated with methyl-b-cyclodextrin, which sequesters this lipid without affecting SM.…”