G M2 gangliosidoses are severe neurodegenerative disorders resulting from a deficiency in β-hexosaminidase A activity and lacking effective therapies. Using a Sandhoff disease (SD) mouse model (Hexb −/− ) of the G M2 gangliosidoses, we tested the potential of systemically delivered adeno-associated virus 9 (AAV9) expressing Hexb cDNA to correct the neurological phenotype. Neonatal or adult SD and normal mice were intravenously injected with AAV9-HexB or -LacZ and monitored for serum β-hexosaminidase activity, motor function, and survival. Brain G M2 ganglioside, β-hexosaminidase activity, and inflammation were assessed at experimental week 43, or an earlier humane end point. SD mice injected with AAV9-LacZ died by 17 weeks of age, whereas all neonatal AAV9-HexB-treated SD mice survived until 43 weeks (P < 0.0001) with only three exhibiting neurological dysfunction. SD mice treated as adults with AAV9-HexB died between 17 and 35 weeks. Neonatal SD-HexB-treated mice had a significant increase in brain β-hexosaminidase activity, and a reduction in G M2 ganglioside storage and neuroinflammation compared to adult SD-HexB-and SD-LacZ-treated groups. However, at 43 weeks, 8 of 10 neonatal-HexB injected control and SD mice exhibited liver or lung tumors. This study demonstrates the potential for long-term correction of SD and other G M2 gangliosidoses through early rAAV9 based systemic gene therapy.
Background:The role of hyaluronidase 2 in hyaluronan degradation and cardiopulmonary function is largely uncharacterized. Results: Hyaluronidase 2-deficient mice accumulate extracellular hyaluronan, leading to defective cardiopulmonary function. Conclusion: Hyaluronidase 2 is essential for hyaluronan degradation and normal heart and lung health. Significance: Our studies suggest that hyaluronidase 2 deficiency should be considered as a contributor to cardiac pathologies in humans.
Hyaluronidases are endoglycosidases that hydrolyze hyaluronan (HA), an abundant component of the extracellular matrix of vertebrate connective tissues. Six human hyaluronidase-related genes have been identified to date. Mutations in one of these genes cause a deficiency of hyaluronidase 1 (HYAL1) resulting in a lysosomal storage disorder, mucopolysaccharidosis (MPS) IX. We have characterized a mouse model of MPS IX and compared its phenotype with the human disease. The targeted Hyal1 allele in this model had a neomycin resistance cassette in exon 2 that replaced 753 bp of the coding region containing the predicted enzyme active site. As a result, Hyal1(-/-) animals had no detectable wild-type Hyal1 transcript, protein or serum activity. Hyal1 null animals were viable, fertile and showed no gross abnormalities at 1 year and 8 months of age. Histological studies of the knee joint showed a loss of proteoglycans occurring as early as 3 months that progressed with age. An increased number of chondrocytes displaying intense pericellular and/or cytoplasmic HA staining were detected in the epiphyseal and articular cartilage of null mice, demonstrating an accumulation of HA. Elevations of HA were not detected in the serum or non-skeletal tissues, indicating that osteoarthritis is the key disease feature in a Hyal1 deficiency. Hyal3 expression was elevated in Hyal1 null mice, suggesting that Hyal3 may compensate in HA degradation in non-skeletal tissues. Overall, the murine MPS IX model displays the key features of the human disease.
Hyaluronidases are enzymes that mediate the breakdown of hyaluronan (HA), a large polysaccharide abundant in the extracellular matrix of vertebrate tissues. Six genes have been predicted to encode hyaluronidases in humans, but the protein products of only SPAM1, HYAL1, and HYAL2 have been characterized. We have now expressed the mouse Hyal3 gene product, hyaluronidase 3 (Hyal3), in Baby Hamster Kidney (BHK) cells and demonstrated the presence of multiple forms of Hyal3 ranging from approximately 45 to 56 kDa in expression lysates. Complete and partial digestions of the expressed protein with PNGase F showed three N-linked oligosaccharides accounted for all forms of Hyal3 detected in expression lysates. Most of these oligosaccharides were Endo H sensitive, indicating that they were high mannose or hybrid N-linked oligosaccharides. Subcellular fractionation of Hyal3-expressing BHK cells by density gradient centrifugation revealed most Hyal3 in a low-density vesicular population. Low levels of Hyal3 were detected in higher density vesicles, but no colocalization with the late endosomal/lysosomal marker Lamp1 was found by immunofluorescence microscopy. BHK cells stably expressing Hyal3 had increased acid-active hyaluronidase activity, but no such activity was detected when Hyal3 was transfected into Hyaluronidase 1 (Hyal1)-deficient fibroblasts. Overexpression of Hyal3 in BHK cells increased the Hyal1 protein and mRNA levels, suggesting that the increased hyaluronidase activity in these cells was due to Hyal1 rather than Hyal3. The results indicate that Hyal3 overexpressed in cultured cells lacks intrinsic hyaluronidase activity and that Hyal3 may contribute to HA metabolism by augmenting the activity of Hyal1.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.