Progressive accumulation of
lipid-laden macrophages is a hallmark of the
acid sphingomyelinase (ASM)-deficient forms of
Niemann-Pick disease (i.e. Types A and B
NPD). To investigate the mechanisms underlying
enzyme replacement therapy for this disorder, we studied the uptake of recombinant, human ASM (rhASM) by alveolar macrophages from ASM knock-out (ASMKO) mice. The recombinant
enzyme used for these studies was produced in Chinese hamster ovary cells and contained complex type, N-linked
oligosaccharides. Binding of radiolabeled, rhASM to the ASMKO macrophages was enhanced as compared with normal macrophages, consistent with their larger size and increased surface area. However, internalization of the
enzyme by the ASMKO cells was markedly reduced when compared with normal cells. Studies using receptor-specific
ligands to inhibit
enzyme uptake revealed that in normal cells rhASM was taken up by a combination of
mannose and
mannose 6-phosphate receptors (MR and M6PR, respectively), whereas in the ASMKO cells the M6PR had a minimal role in rhASM uptake. Expression of M6PR
mRNA was normal in the ASMKO cells, although Western blotting revealed more receptors in these cells when compared with normal. We therefore hypothesized that
lipid accumulation in ASMKO macrophages led to abnormalities in M6PR trafficking and/or degradation, resulting in reduced
enzyme uptake. Consistent with this hypothesis, we also found that, when rhASM was modified to expose terminal
mannose residues and target
mannose receptors, the uptake of this modified
enzyme form by ASMKO cells was approximately 10-fold greater when compared with the "complex" type rhASM. These findings have important implications for
NPD enzyme replacement therapy, particularly in the lung.