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Plasma membrane calcium ATPase deficiency causes neuronal pathology in the spinal cord: a potential mechanism for neurodegeneration in multiple sclerosis and spinal cord injury.

ABSTRACT: Dysfunction and death of spinal cord neurons are critical determinants of neurological deficits in various pathological conditions, including multiple sclerosis (MS) and spinal cord injury. Yet, the molecular mechanisms underlying neuronal/axonal damage remain undefined. Our previous studies raised the possibility that a decrease in the levels of plasma membrane calcium ATPase isoform 2 (PMCA2), a major pump extruding calcium from neurons, promotes neuronal pathology in the spinal cord during experimental autoimmune encephalomyelitis (EAE), an animal model of MS, and after spinal cord trauma. However, the causal relationship between alterations in PMCA2 levels and neuronal injury was not well established. We now report that inhibition of PMCA activity in purified spinal cord neuronal cultures delays calcium clearance, increases the number of nonphosphorylated neurofilament H (SMI-32) immunoreactive cells, and induces swelling and beading of SMI-32-positive neurites. These changes are followed by activation of caspase-3 and neuronal loss. Importantly, the number of spinal cord motor neurons is significantly decreased in PMCA2-deficient mice and the deafwaddler(2J), a mouse with a functionally null mutation in the PMCA2 gene. Our findings suggest that a reduction in PMCA2 level or activity leading to delays in calcium clearance may cause neuronal damage and loss in the spinal cord.

SUBMITTER: Kurnellas MP

PROVIDER: S-EPMC2896328 | biostudies-literature | 2005 Feb

REPOSITORIES: biostudies-literature

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Plasma membrane calcium ATPase deficiency causes neuronal pathology in the spinal cord: a potential mechanism for neurodegeneration in multiple sclerosis and spinal cord injury.

Kurnellas Michael P MP Nicot Arnaud A Shull Gary E GE Elkabes Stella S

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 20041202 2

Dysfunction and death of spinal cord neurons are critical determinants of neurological deficits in various pathological conditions, including multiple sclerosis (MS) and spinal cord injury. Yet, the molecular mechanisms underlying neuronal/axonal damage remain undefined. Our previous studies raised the possibility that a decrease in the levels of plasma membrane calcium ATPase isoform 2 (PMCA2), a major pump extruding calcium from neurons, promotes neuronal pathology in the spinal cord during ex ...[more]

PMID: 15576480

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Project description:BACKGROUND: Gangliosides, sialic acid-containing glycosphingolipids, are highly expressed in nervous systems of vertebrates and have been considered to be involved in the development, differentiation, and function of nervous tissues. Recent studies with gene-engineered animals have revealed that they play roles in the maintenance and repair of nervous tissues. In particular, knockout (KO) mice of various ganglioside synthase genes have exhibited progressive neurodegeneration with aging. However, neurological disorders and pathological changes in the spinal cord of these KO mice have not been reported to date. Therefore, we examined neurodegeneration in double knockout (DKO) mice of ganglioside GM2/GD2 synthase (B4GANLT1) and GD3 synthase (ST8SIA1) genes to clarify roles of gangliosides in the spinal cord. METHODS: Motor neuron function was examined by gait analysis, and sensory function was analyzed by von Frey test. Pathological changes were analyzed by staining tissue sections with Klüver-Barrera staining and by immunohistochemistry with F4/80 and glial fibrillary acidic protein (GFAP). Gene expression profiles were examined by using DNA micro-array of RNAs from the spinal cord of mice. Triple knockout mice were generated by mating DKO and complement component 3 (C3)-KO mice. Gene expression of the complement system and cytokines was examined by reverse transcription-polymerase chain reaction (RT-PCR) as a function of age. RESULTS: DKO mice showed progressive deterioration with aging. Correspondingly, they exhibited shrunk spinal cord, reduced thickness of spinal lamina II and III, and reduced neuronal numbers in spinal lamina IX, spinal lamina II, and spinal lamina I. Complement-related genes were upregulated in DKO spinal cord. Moreover, complement activation and inflammatory reactions were detected by GFAP-active astrocyte, microglial accumulation, and increased inflammatory cytokines such as tumor necrosis factor-alpha (TNF?) and interleukin-1-beta (IL-1?). Triple knockout mice showed restoration of reduced neuron numbers in the spinal cord of DKO mice, getting close to levels of wild-type mice. CONCLUSIONS: Disruption in the architecture of lipid rafts in the spinal cord was not so prominent, suggesting that mechanisms distinct from those reported might be involved in the complement activation in the spinal cord of DKO mice. Gene profiling revealed that inflammation and neurodegeneration in the spinal cord of DKO mice are, at least partly, dependent on complement activation.

Project description:BACKGROUND:Neuropathic pain is often observed in individuals with multiple sclerosis (MS) and spinal cord injury (SCI) and is not adequately alleviated by current pharmacotherapies. A better understanding of underlying mechanisms could facilitate the discovery of novel targets for therapeutic interventions. We previously reported that decreased plasma membrane calcium ATPase 2 (PMCA2) expression in the dorsal horn (DH) of healthy PMCA2+/- mice is paralleled by increased sensitivity to evoked nociceptive pain. These studies suggested that PMCA2, a calcium extrusion pump expressed in spinal cord neurons, plays a role in pain mechanisms. However, the contribution of PMCA2 to neuropathic pain processing remains undefined. The present studies investigated the role of PMCA2 in neuropathic pain processing in the DH of wild-type mice affected by experimental autoimmune encephalomyelitis (EAE), an animal model of MS, and following SCI. METHODS:EAE was induced in female and male C57Bl/6N mice via inoculation with myelin oligodendrocyte glycoprotein fragment 35-55 (MOG35-55) emulsified in Complete Freund's Adjuvant (CFA). CFA-inoculated mice were used as controls. A severe SC contusion injury was induced at thoracic (T8) level in female C57Bl/6N mice. Pain was evaluated by the Hargreaves and von Frey filament tests. PMCA2 levels in the lumbar DH were analyzed by Western blotting. The effectors that decrease PMCA2 expression were identified in SC neuronal cultures. RESULTS:Increased pain in EAE and SCI was paralleled by a significant decrease in PMCA2 levels in the DH. In contrast, PMCA2 levels remained unaltered in the DH of mice with EAE that manifested motor deficits but not increased pain. Interleukin-1? (IL-1?), tumor necrosis factor ? (TNF?), and IL-6 expression were robustly increased in the DH of mice with EAE manifesting pain, whereas these cytokines showed a modest increase or no change in mice with EAE in the absence of pain. Only IL-1? decreased PMCA2 levels in pure SC neuronal cultures through direct actions. CONCLUSIONS:PMCA2 is a contributor to neuropathic pain mechanisms in the DH. A decrease in PMCA2 in DH neurons is paralleled by increased pain sensitivity, most likely through perturbations in calcium signaling. Interleukin-1? is one of the effectors that downregulates PMCA2 by acting directly on neurons.

Project description:Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder caused by progressive loss of motor neurons and there is currently no effective therapy. Cytoplasmic mislocalization and aggregation of TAR DNA-binding protein 43 kDa (TDP-43) within the CNS is a pathological hallmark in sporadic ALS and prion-like propagation of pathogenic TDP-43 is thought to be implicated in disease progression. However, cell-to-cell transmission of pathogenic TDP-43 in the human CNS has not been confirmed experimentally. Here we used induced pluripotent stem cells (iPSCs)-derived cerebral organoids as recipient CNS tissue model that are anatomically relevant human brain. We injected postmortem spinal cord protein extracts individually from three non-ALS or five sporadic ALS patients containing pathogenic TDP-43 into the cerebral organoids to validate the templated propagation and spreading of TDP-43 pathology in human CNS tissue. We first demonstrated that the administration of spinal cord extracts from an ALS patient induced the formation of TDP-43 pathology that progressively spread in a time-dependent manner in cerebral organoids, suggesting that pathogenic TDP-43 from ALS functioned as seeds and propagated cell-to-cell to form de novo TDP-43 pathology. We also reported that the administration of ALS patient-derived protein extracts caused astrocyte proliferation to form astrogliosis in cerebral organoids, reproducing the pathological feature seen in ALS. Moreover, we showed pathogenic TDP-43 induced cellular apoptosis and that TDP-43 pathology correlated with genomic damage due to DNA double-strand breaks. Thus, our results provide evidence that patient-derived pathogenic TDP-43 can mimic the prion-like propagation of TDP-43 pathology in human CNS tissue. Our findings indicate that our assays with human cerebral organoids that replicate ALS pathophysiology have a promising strategy for creating readouts that could be used in future drug discovery efforts against ALS.

Project description:BackgroundTraumatic spinal cord injury (SCI)-induced neuroinflammation results in secondary neurological destruction and functional disorder. Previous findings showed that microglial pyroptosis plays a crucial role in neuroinflammation. Thus, it is necessary to conduct a comprehensive investigation of the mechanisms associated with post-SCI microglial pyroptosis. The Fanconi Anemia Group C complementation group gene (FANCC) has been previously reported to have an anti-inflammation effect; however, whether it can regulate microglial pyroptosis remains unknown. Therefore, we probed the mechanism associated with FANCC-mediated microglial pyroptosis and neuroinflammation in vitro and in vivo in SCI mice.MethodsMicroglial pyroptosis was assessed by western blotting (WB) and immunofluorescence (IF), whereas microglial-induced neuroinflammation was evaluated by WB, Enzyme-linked immunosorbent assays and IF. Besides, flow cytometry, TdT-mediated dUTP Nick-End Labeling staining and WB were employed to examine the level of neuronal apoptosis. Morphological changes in neurons were assessed by hematoxylin-eosin and Luxol Fast Blue staining. Finally, locomotor function rehabilitation was analyzed using the Basso Mouse Scale and Louisville Swim Scale.ResultsOverexpression of FANCC suppressed microglial pyroptosis via inhibiting p38/NLRP3 expression, which in turn reduced neuronal apoptosis. By contrast, knockdown of FANCC increased the degree of neuronal apoptosis by aggravating microglial pyroptosis. Besides, increased glial scar formation, severe myelin sheath destruction and poor axon outgrowth were observed in the mice transfected with short hairpin RNA of FANCC post SCI, which caused reduced locomotor function recovery.ConclusionsTaken together, a previously unknown role of FANCC was identified in SCI, where its deficiency led to microglia pyroptosis, neuronal apoptosis and neurological damage. Mechanistically, FANCC mediated microglia pyroptosis and the inflammatory response via regulating the p38/NLRP3 pathway.

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Plasma membrane calcium ATPase deficiency causes neuronal pathology in the spinal cord: a potential mechanism for neurodegeneration in multiple sclerosis and spinal cord injury.

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Plasma membrane calcium ATPase deficiency causes neuronal pathology in the spinal cord: a potential mechanism for neurodegeneration in multiple sclerosis and spinal cord injury.

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