Myelin contains neutral sphingomyelinase activity that is stimulated by tumor necrosis factor-?

Abstract

Purified myelin from mouse brain was found to contain two forms of neutral sphingomyelinase, one Mg2+ dependent and the other Mg2+ independent. The former had a pH optimum of 7.5 and Km of 0.35 mM, whereas the corresponding values for the latter were pH 8.0 and Km 3.03 mM. Specific activity of the Mg(2+)-dependent enzyme showed a rostral-caudal gradient, ranging from 75 nmol/mg protein/hr in myelin from cerebral hemispheres to 21 nmol/mg protein/hr in myelin from spinal cord. Relative specific activity was approximately 20% that of brain stem or cerebral hemisphere homogenate. Treatment of myelin with taurocholate or high salt concentration did not significantly reduce activity of the Mg(2+)-dependent enzyme. The activity of that enzyme did not change with time or in the presence or absence of protease inhibitors; by contrast, that of Mg(2+)-independent enzyme decreased sharply in the absence of protease inhibitors but rose in their presence. To test for the effect of tumor necrosis factor-alpha (TNF alpha) on myelin sphingomyelinase, mouse brain myelin was labeled in vivo by intracerebral injection of [3H]acetate into 18-20-day-old mice. After 40 hr, brain stems were removed, minced, and treated with TNF alpha in Krebs-Ringer solution, after which myelin was immediately isolated. Separation and counting of individual lipids revealed TNF alpha treatment to cause increased labeling of myelin ceramide and cholesterol ester with concomitant decrease in myelin sphingomyelin. Western blotting of myelin proteins using antibodies to the two TNF alpha receptors as probes revealed the presence of the p75 receptor. Implications of these findings in relation to possible mechanisms of autoimmune demyelination are discussed.

Ceramide and neurodegeneration: Susceptibility of neurons and oligodendrocytes to cell damage and death

“…Such neurotoxic molecules as inflammatory cytokines, chemokines, excitotoxic molecules, and ROS, elaborated from activated glia increase in the brain and CSF of neurodegenerative disorders, and several of these couple the sphingomyelin-ceramide pathway with its prominent connection to apoptosis (and to other cellular processes as well). Early on, TNF-α released from astrocytes and macrophages was recognized to activate myelin associated sphingomyelinases to generate ceramide and other downstream products with consequent deleterious effects upon membrane integrity [101,102]. This is fully consistent with our own co-culture studies of genetic knockdown of NSMase in human primary neurons using an antisense strategy; finding that NSMase-impotent neurons resist activated glia-mediated cytotoxicity (unpublished data).…”

Sphingolipids in human lens membranes: an update on their composition and possible biological implications

“…Whereas ceramides produced after cleavage of the phosphorylcholine headgroup from SMs by sphingomyelinases (Andrieu-Abadie and Levade, 2002;Chakraborty et al, 1997;Chatterjee, 1993;Gatt, 1978;Hannun, 1996;Hannun and Obeid, 1995;Jayadev et al, 1995;Liu et al, 1997;Merrill and Jones, 1990) behave as potent lipid second messengers, the saturated dihydroceramides lack activity in most fundamental processes (Galadari et al, 1998;Simon and Gear, 1998;Wolff et al, 1994), and are often used as controls in functional studies of ceramides. It has been postulated that the inactivity of dihydroceramides is not due to their decreased uptake or increased metabolism, but probably arises from the lack of response by the ceramide-activated protein phosphatase, a potential cellular target for ceramide action during signal transduction (Galadari et al, 1998;Wolff et al, 1994).…”

Identification and evaluation of neutral sphingomyelinase 2 inhibitors

“…Several studies showed that many pathophysiological processes in the brain, such as Parkinson's disease (Hunot et al, 1997), Alzheimer's disease (Fiebich et al, 1995), ischemic brain injury (Kubota et al, 1989) and autoimmune demyelination (Chakraborty et al, 1997) are attributable to this ceramide-dependent signaling pathway. Neurotrophin-(p75NT) and TNF-α-induced signaling pathways are known to utilize this ceramide as a signaling molecule for downstream effectors in many neuronal cells, including oligodendrocytes (Casaccia-Bonnefil et al, 1996), PC12 cells (Dobrowsky et al, 1995) and T9 glioma cells (MacPhee 6.…”

“…nSMase is generally found in the brain (Liu et al, 1998) and its activation leads to the activation of the ceramide-dependent signaling pathway, which affects pathophysiological processes in brain such as ischemic brain injury (Kubota et al, 1989), autoimmune demyelination (Chakraborty et al, 1997), Parkinson's disease (Hunot et al, 1997), Alzheimer's disease (Fiebich et al, 1995) and other neurodegenerative diseases (Magen et al, 2008). Therefore, a specific inhibitor of nSMase can lead to the development of alternative therapies for these diseases.…”