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Specific saposin C deficiency: CNS impairment and acid beta-glucosidase effects in the mouse.

Sun Y, Ran H, Zamzow M, Kitatani K, Skelton MR, Williams MT, Vorhees CV, Witte DP, Hannun YA, Grabowski GA - Hum. Mol. Genet. (2009)

Bottom Line: Ultrastructural analyses revealed inclusions in axonal processes in the spinal cord, sciatic nerve and brain, but no excess of multivesicular bodies.Activated microglial cells and astrocytes were present in thalamus, brain stem, cerebellum and spinal cord, indicating regional pro-inflammatory responses.These results support the view that saposin C has multiple roles in glycosphingolipid (GSL) catabolism as well as a prominent function in CNS and axonal integrity independent of its role as an optimizer/stabilizer of GCase.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA.

ABSTRACT
Saposins A, B, C and D are derived from a common precursor, prosaposin (psap). The few patients with saposin C deficiency develop a Gaucher disease-like central nervous system (CNS) phenotype attributed to diminished glucosylceramide (GC) cleavage activity by acid beta-glucosidase (GCase). The in vivo effects of saposin C were examined by creating mice with selective absence of saposin C (C-/-) using a knock-in point mutation (cysteine-to-proline) in exon 11 of the psap gene. In C-/- mice, prosaposin and saposins A, B and D proteins were present at near wild-type levels, but the saposin C protein was absent. By 1 year, the C-/- mice exhibited weakness of the hind limbs and progressive ataxia. Decreased neuromotor activity and impaired hippocampal long-term potentiation were evident. Foamy storage cells were observed in dorsal root ganglion and there was progressive loss of cerebellar Purkinje cells and atrophy of cerebellar granule cells. Ultrastructural analyses revealed inclusions in axonal processes in the spinal cord, sciatic nerve and brain, but no excess of multivesicular bodies. Activated microglial cells and astrocytes were present in thalamus, brain stem, cerebellum and spinal cord, indicating regional pro-inflammatory responses. No storage cells were found in visceral organs of these mice. The absence of saposin C led to moderate increases in GC and lactosylceramide (LacCer) and their deacylated analogues. These results support the view that saposin C has multiple roles in glycosphingolipid (GSL) catabolism as well as a prominent function in CNS and axonal integrity independent of its role as an optimizer/stabilizer of GCase.

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CNS pathology in C−/− mice. (A) The paraffin sections of dorsal root ganglion and dorsal horn of spinal cord from 15-month-old saposin C+/− or C−/− mice were stained with H&E. Foamy storage material (arrows) was present in the dorsal root ganglion neurons of saposin C−/− mice. Larger axonal spheroids (arrows) were present in the spinal cord dorsal horn of these mice. As a control, saposin C+/− mice had normal morphology in dorsal root ganglion and spinal cord. (B) CNS proinflammation in saposin C−/− mice demonstrated by anti-GFAP (B1-B3) and anti-CD68 (B4-B6). (B1) WT spinal cord, (B2) saposin C−/− spinal cord and (B3) C−/− midbrain at 15 months showed astrogliosis with enhanced GFAP (green) signal. The nucleus was labeled by DAPI (blue). (B4) C−/− dorsal horn of spinal cord. (B5) C−/− thalamus. (B6) C−/− brain stem. Microglial cells were stained by anti-CD68 antibody (brown) in 24-month-old saposin C−/− mice. Tissue sections were counterstained with hematoxylin. (C) H&E stained paraffin sections of spinal cord and cerebellum from saposin C−/− mice at 3, 6, 12 and 18 months. Axonal spheroids were present in dorsal horn of spinal cord at 6, 12 and 18 months, but absent at 3 months. Purkinje cell layer (arrows) in cerebella lobule IV was normal at 3 months. Loss of Purkinje cells (*) was evident at 6 months and older.
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DDP531F6: CNS pathology in C−/− mice. (A) The paraffin sections of dorsal root ganglion and dorsal horn of spinal cord from 15-month-old saposin C+/− or C−/− mice were stained with H&E. Foamy storage material (arrows) was present in the dorsal root ganglion neurons of saposin C−/− mice. Larger axonal spheroids (arrows) were present in the spinal cord dorsal horn of these mice. As a control, saposin C+/− mice had normal morphology in dorsal root ganglion and spinal cord. (B) CNS proinflammation in saposin C−/− mice demonstrated by anti-GFAP (B1-B3) and anti-CD68 (B4-B6). (B1) WT spinal cord, (B2) saposin C−/− spinal cord and (B3) C−/− midbrain at 15 months showed astrogliosis with enhanced GFAP (green) signal. The nucleus was labeled by DAPI (blue). (B4) C−/− dorsal horn of spinal cord. (B5) C−/− thalamus. (B6) C−/− brain stem. Microglial cells were stained by anti-CD68 antibody (brown) in 24-month-old saposin C−/− mice. Tissue sections were counterstained with hematoxylin. (C) H&E stained paraffin sections of spinal cord and cerebellum from saposin C−/− mice at 3, 6, 12 and 18 months. Axonal spheroids were present in dorsal horn of spinal cord at 6, 12 and 18 months, but absent at 3 months. Purkinje cell layer (arrows) in cerebella lobule IV was normal at 3 months. Loss of Purkinje cells (*) was evident at 6 months and older.

Mentions: Saposin C−/− mice had foamy storage material in the dorsal root ganglion starting at 25 weeks. Larger axonal spheroids were evident in the dorsal horn of the spinal cord (Fig. 6A). No storage cells were visible in the brain. Pro-inflammation in saposin C−/− mice was assessed using anti-CD68 and anti-GFAP antibody staining (Fig. 6B). CD68 is an intracellular membrane glycoprotein that is expressed in quiescent and activated tissue macrophages (23). GFAP is an astrocytic marker. The enhanced GFAP signals indicate astrogliosis. Clearly enhanced CD68 and GFAP signals were found in spinal cord, brainstem, hindbrain, cerebellum and thalamic regions in saposin C−/− mice compared with WT mice. Disease progression was evident in H&E stained paraffin sections of spinal cord and cerebellum in 3–19-month-old saposin C−/− mice (Fig. 6C). Axonal spheroids were present in spinal cord dorsal horn after 6 months, but absent at 3 months. The Purkinje cell layer in cerebellar lobule IV was normal at 3 months. Loss of Purkinje cells was observed after 6 months. TUNEL assays of paraffin sections were negative suggesting that loss of Purkinje cells was not by an apoptotic mechanism. Visceral organs in saposin C−/− mice were not distinguishable from WT mice.


Specific saposin C deficiency: CNS impairment and acid beta-glucosidase effects in the mouse.

Sun Y, Ran H, Zamzow M, Kitatani K, Skelton MR, Williams MT, Vorhees CV, Witte DP, Hannun YA, Grabowski GA - Hum. Mol. Genet. (2009)

CNS pathology in C−/− mice. (A) The paraffin sections of dorsal root ganglion and dorsal horn of spinal cord from 15-month-old saposin C+/− or C−/− mice were stained with H&E. Foamy storage material (arrows) was present in the dorsal root ganglion neurons of saposin C−/− mice. Larger axonal spheroids (arrows) were present in the spinal cord dorsal horn of these mice. As a control, saposin C+/− mice had normal morphology in dorsal root ganglion and spinal cord. (B) CNS proinflammation in saposin C−/− mice demonstrated by anti-GFAP (B1-B3) and anti-CD68 (B4-B6). (B1) WT spinal cord, (B2) saposin C−/− spinal cord and (B3) C−/− midbrain at 15 months showed astrogliosis with enhanced GFAP (green) signal. The nucleus was labeled by DAPI (blue). (B4) C−/− dorsal horn of spinal cord. (B5) C−/− thalamus. (B6) C−/− brain stem. Microglial cells were stained by anti-CD68 antibody (brown) in 24-month-old saposin C−/− mice. Tissue sections were counterstained with hematoxylin. (C) H&E stained paraffin sections of spinal cord and cerebellum from saposin C−/− mice at 3, 6, 12 and 18 months. Axonal spheroids were present in dorsal horn of spinal cord at 6, 12 and 18 months, but absent at 3 months. Purkinje cell layer (arrows) in cerebella lobule IV was normal at 3 months. Loss of Purkinje cells (*) was evident at 6 months and older.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

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Show All Figures
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DDP531F6: CNS pathology in C−/− mice. (A) The paraffin sections of dorsal root ganglion and dorsal horn of spinal cord from 15-month-old saposin C+/− or C−/− mice were stained with H&E. Foamy storage material (arrows) was present in the dorsal root ganglion neurons of saposin C−/− mice. Larger axonal spheroids (arrows) were present in the spinal cord dorsal horn of these mice. As a control, saposin C+/− mice had normal morphology in dorsal root ganglion and spinal cord. (B) CNS proinflammation in saposin C−/− mice demonstrated by anti-GFAP (B1-B3) and anti-CD68 (B4-B6). (B1) WT spinal cord, (B2) saposin C−/− spinal cord and (B3) C−/− midbrain at 15 months showed astrogliosis with enhanced GFAP (green) signal. The nucleus was labeled by DAPI (blue). (B4) C−/− dorsal horn of spinal cord. (B5) C−/− thalamus. (B6) C−/− brain stem. Microglial cells were stained by anti-CD68 antibody (brown) in 24-month-old saposin C−/− mice. Tissue sections were counterstained with hematoxylin. (C) H&E stained paraffin sections of spinal cord and cerebellum from saposin C−/− mice at 3, 6, 12 and 18 months. Axonal spheroids were present in dorsal horn of spinal cord at 6, 12 and 18 months, but absent at 3 months. Purkinje cell layer (arrows) in cerebella lobule IV was normal at 3 months. Loss of Purkinje cells (*) was evident at 6 months and older.
Mentions: Saposin C−/− mice had foamy storage material in the dorsal root ganglion starting at 25 weeks. Larger axonal spheroids were evident in the dorsal horn of the spinal cord (Fig. 6A). No storage cells were visible in the brain. Pro-inflammation in saposin C−/− mice was assessed using anti-CD68 and anti-GFAP antibody staining (Fig. 6B). CD68 is an intracellular membrane glycoprotein that is expressed in quiescent and activated tissue macrophages (23). GFAP is an astrocytic marker. The enhanced GFAP signals indicate astrogliosis. Clearly enhanced CD68 and GFAP signals were found in spinal cord, brainstem, hindbrain, cerebellum and thalamic regions in saposin C−/− mice compared with WT mice. Disease progression was evident in H&E stained paraffin sections of spinal cord and cerebellum in 3–19-month-old saposin C−/− mice (Fig. 6C). Axonal spheroids were present in spinal cord dorsal horn after 6 months, but absent at 3 months. The Purkinje cell layer in cerebellar lobule IV was normal at 3 months. Loss of Purkinje cells was observed after 6 months. TUNEL assays of paraffin sections were negative suggesting that loss of Purkinje cells was not by an apoptotic mechanism. Visceral organs in saposin C−/− mice were not distinguishable from WT mice.

Bottom Line: Ultrastructural analyses revealed inclusions in axonal processes in the spinal cord, sciatic nerve and brain, but no excess of multivesicular bodies.Activated microglial cells and astrocytes were present in thalamus, brain stem, cerebellum and spinal cord, indicating regional pro-inflammatory responses.These results support the view that saposin C has multiple roles in glycosphingolipid (GSL) catabolism as well as a prominent function in CNS and axonal integrity independent of its role as an optimizer/stabilizer of GCase.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati, OH, USA.

ABSTRACT
Saposins A, B, C and D are derived from a common precursor, prosaposin (psap). The few patients with saposin C deficiency develop a Gaucher disease-like central nervous system (CNS) phenotype attributed to diminished glucosylceramide (GC) cleavage activity by acid beta-glucosidase (GCase). The in vivo effects of saposin C were examined by creating mice with selective absence of saposin C (C-/-) using a knock-in point mutation (cysteine-to-proline) in exon 11 of the psap gene. In C-/- mice, prosaposin and saposins A, B and D proteins were present at near wild-type levels, but the saposin C protein was absent. By 1 year, the C-/- mice exhibited weakness of the hind limbs and progressive ataxia. Decreased neuromotor activity and impaired hippocampal long-term potentiation were evident. Foamy storage cells were observed in dorsal root ganglion and there was progressive loss of cerebellar Purkinje cells and atrophy of cerebellar granule cells. Ultrastructural analyses revealed inclusions in axonal processes in the spinal cord, sciatic nerve and brain, but no excess of multivesicular bodies. Activated microglial cells and astrocytes were present in thalamus, brain stem, cerebellum and spinal cord, indicating regional pro-inflammatory responses. No storage cells were found in visceral organs of these mice. The absence of saposin C led to moderate increases in GC and lactosylceramide (LacCer) and their deacylated analogues. These results support the view that saposin C has multiple roles in glycosphingolipid (GSL) catabolism as well as a prominent function in CNS and axonal integrity independent of its role as an optimizer/stabilizer of GCase.

Show MeSH
Related in: MedlinePlus