Prolidase deficiency (PD) is a rare autosomal recessive disorder caused by mutations in the prolidase gene, the PEPD, causing the reduction or the loss of the prolidase enzyme activity. PD patients present a variable onset, and severe skin ulcers mainly characterize the pathology. However, PD has a broad spectrum of phenotypes including mental impairment and developmental delay of variable degrees. Prolidase is a member of the matrix metalloproteinase (MMP) family. MMPs together with their inhibitors (tissue inhibitor of metalloproteinases, TIMPs) regulate the extracellular matrix maturation and remodelling life-long. Among them, prolidase is able to cleave dipeptides when prolin or hydroxyprolin residues are located at the C-terminal end. According to prolidase activity, its function has an impact on the metabolism of many biologically important molecules, particularly during the biosynthesis and degradation of collagen and procollagen. Therefore, prolidase indirectly has a role in the ECM remodelling. In particular, the ECM adjustments are essential in the brain, especially during the critical period of development: from passive structural property, to a direct influence on cell proliferation, migration, axonal guidance, synaptogenesis, homeostatic plasticity, learning and memory processes, and angiogenesis. In particular, the basement membrane beside the pial meninx (pBM) is a specialized structure of ECM whose integrity and proper assembly is essential for a correct cortical development and the collagen IV plays an essential role in pBM stability. Ruptures, even localized, in the pBM are accompanied by changes in the morphology of radial glia cells, subsequent cortical dysplasia, overmigration of neurons, decrease in the proliferation and migration of granule cell precursors, and reduction in Purkinje neuron dendrites. Recently, a mutant mouse with reduced prolidase activity has been identified with a spontaneous 4 bp deletion in the exon 14 of Pepd gene. The mutant mouse was named dark-like (dal) because of its darkened-coat color in homozygosis. The dal/dal phenotype includes small body size, reproductive degeneration, vacuolated cells at the cortical medullary junction of the adrenal gland, mild hydrocephalus, dark urine and altered bone phenotype. The prolidase activity was strongly reduced in cerebrum and cerebellum in dal mice. Moreover, they develop hypertrophic cardiomyopathy, but neither skin lesions nor recurrent infections were reported (in contrast to the reported human cases). The aim of this thesis was the study the brain development of dal/+ and dal/dal mice. Since no information were available, the analysis started with a morphological evaluation of the cerebellum, neocortex and hippocampus, through histological stainings. Then, immunohistochemistry reactions and western blotting analysis helped the anomalies characterizations. In particular, the attention has been mainly focused on the cerebellum, since it is the structure in which the ontogenetic events occurred also postnatally. The neocortex and hippocampal results were not described in details. Our results suggested that the absence of a full functional prolidase enzyme in the dal/dal mice results in a damage of the integrity of the pBM with an altered collagen, laminin and reelin profile. Such damage, could affect as a cascade of developmental events the proper lamination process of the cerebellum, leading to a cortical dysplasia together with the presence of degenerating and ectopic cells, defects in cerebellar lobulation and in the excitation/inhibition pattern of the cerebellar circuit.

Brain Development in Prolidase Deficient Mice

INSOLIA, VIOLETTA
2016-12-16

Abstract

Prolidase deficiency (PD) is a rare autosomal recessive disorder caused by mutations in the prolidase gene, the PEPD, causing the reduction or the loss of the prolidase enzyme activity. PD patients present a variable onset, and severe skin ulcers mainly characterize the pathology. However, PD has a broad spectrum of phenotypes including mental impairment and developmental delay of variable degrees. Prolidase is a member of the matrix metalloproteinase (MMP) family. MMPs together with their inhibitors (tissue inhibitor of metalloproteinases, TIMPs) regulate the extracellular matrix maturation and remodelling life-long. Among them, prolidase is able to cleave dipeptides when prolin or hydroxyprolin residues are located at the C-terminal end. According to prolidase activity, its function has an impact on the metabolism of many biologically important molecules, particularly during the biosynthesis and degradation of collagen and procollagen. Therefore, prolidase indirectly has a role in the ECM remodelling. In particular, the ECM adjustments are essential in the brain, especially during the critical period of development: from passive structural property, to a direct influence on cell proliferation, migration, axonal guidance, synaptogenesis, homeostatic plasticity, learning and memory processes, and angiogenesis. In particular, the basement membrane beside the pial meninx (pBM) is a specialized structure of ECM whose integrity and proper assembly is essential for a correct cortical development and the collagen IV plays an essential role in pBM stability. Ruptures, even localized, in the pBM are accompanied by changes in the morphology of radial glia cells, subsequent cortical dysplasia, overmigration of neurons, decrease in the proliferation and migration of granule cell precursors, and reduction in Purkinje neuron dendrites. Recently, a mutant mouse with reduced prolidase activity has been identified with a spontaneous 4 bp deletion in the exon 14 of Pepd gene. The mutant mouse was named dark-like (dal) because of its darkened-coat color in homozygosis. The dal/dal phenotype includes small body size, reproductive degeneration, vacuolated cells at the cortical medullary junction of the adrenal gland, mild hydrocephalus, dark urine and altered bone phenotype. The prolidase activity was strongly reduced in cerebrum and cerebellum in dal mice. Moreover, they develop hypertrophic cardiomyopathy, but neither skin lesions nor recurrent infections were reported (in contrast to the reported human cases). The aim of this thesis was the study the brain development of dal/+ and dal/dal mice. Since no information were available, the analysis started with a morphological evaluation of the cerebellum, neocortex and hippocampus, through histological stainings. Then, immunohistochemistry reactions and western blotting analysis helped the anomalies characterizations. In particular, the attention has been mainly focused on the cerebellum, since it is the structure in which the ontogenetic events occurred also postnatally. The neocortex and hippocampal results were not described in details. Our results suggested that the absence of a full functional prolidase enzyme in the dal/dal mice results in a damage of the integrity of the pBM with an altered collagen, laminin and reelin profile. Such damage, could affect as a cascade of developmental events the proper lamination process of the cerebellum, leading to a cortical dysplasia together with the presence of degenerating and ectopic cells, defects in cerebellar lobulation and in the excitation/inhibition pattern of the cerebellar circuit.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11571/1203288
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