Integration of cytokine biology and lipid metabolism in stroke

Abstract

Cytokines regulate the innate and adaptive immune responses and are pleiotropic, redundant and multifunctional. Expression of most cytokines, including TNF-alpha and IL-1alpha/beta, is very low in normal brain. Metabolism of lipids is of particular interest due to their high concentration in the brain. Inflammatory response after stroke suggests that cytokines (TNF-alpha, IL-1alpha/beta, IL-6), affect the phospholipid metabolism and subsequent production of eicosanoids, ceramide, and ROS that may potentiate brain injury. Phosphatidylcholine and sphingomyelin are source for lipid messengers. Sphingomyelin synthase serves as a bridge between metabolism of glycerolipids and sphingolipids. TNF-alpha and IL-1alpha/beta can induce phospholipases (A2, C, and D) and sphingomyelinases, and concomitantly proteolyse phosphatidylcholine and sphingomyelin synthesizing enzymes. Together, these alterations contribute to loss of phosphatidylcholine and sphingomyelin after stroke that can be attenuated by inhibiting TNF-alpha or IL-1alpha/beta signaling. Inflammatory responses are instrumental in the formation and destabilization of atherosclerotic plaques. Secretory PLA2 IIA is found in human atherosclerotic lesions and is implicated in initiation, progression and maturation of atherosclerosis, a risk factor for stroke. Lipoprotein-PLA2, part of apolipoprotein B-100 of LDL, plays a role in vascular inflammation and coronary endothelial dysfunction. Cytokine antagonism attenuated secretory PLA2 IIA actions, suggesting cytokine-lipid integration studies will lead to new concepts contributing to bench-to-bedside transition for stroke therapy.

Figure 1

Lipid metabolism in ischemic neuronal death. Activation of phospholipases (PLA₂, PC-PLC, PI-PLC, PLD) sphingomyelinases (N-SMase and A-SMase) following cerebral ischemia results in release of lipid second messengers DAG, phosphatidic acid (PA), docosahexaenoic acid (DHA), ArAc and ceramide. PI-PLC releases inositol-trisphosphate (IP₃) in addition to DAG; IP₃ stimulates intracellular Ca⁺⁺ release. PA and DAG can be readily interconverted by phosphatases and DAG-kinases. ArAc undergoes further metabolism by LOX to generate important vasoactive eicosanoids and ROS. Recent studies suggest that COX does not directly produce ROS during ArAc oxidative metabolism, but does form free radicals (i.e., carbon-centered radicals on ArAc) (148, 200). DHA (204) is metabolized to 10,17S-docosatriene (Neuroprotectin D1), an endogenous neuroprotectant. DAG and ArAc stimulate A-SMase and N-SMase, respectively, to generate pro-apoptotic ceramide (148).

Figure. 2

Asymmetric distribution of phospholipids (PL) in a plasma membrane

Figure. 3

PC synthesis through CDP-choline/Kennedy pathway (148). CCT is down-regulated after stroke, impeding PC synthesis (41). CCT:cytidine triphosphate:phosphocholine

Figure 4

The immediate products formed from PC by each class of phospholipase.

Figure 5

D609 (50 mg/kg in saline; i.p at the onset of reperfusion) attenuated ischemic injury volume (infarction) by 35 ± 5% after 1 hr MCAO/24 hr reperfusion (saline 270 ± 38 mm³).

Figure. 6

Cytokine mediated generation of lipid mediators DAG, ArAc and ceramide. This scheme also shows the relationship between glycerophospholipids and sphingolipids (148). TNF-α and IL-1 activate N-SMase and A-SMase through stimulation of PLA₂ and PC-PLC and release of ArAc and DAG, respectively. N-SMase and A-SMase hydrolyze SM to liberate ceramide. SM synthase transfers the phosphocholine head group of PC to ceramide to form SM and DAG.

Figure 7

Time course of changes related to cytokines, lipid metabolism and oxidative stress after stroke (148). ↑=increase, ↓=decrease, compared to control. HNE: 4-hydroxynonenal; MDA: malondialdehyde; •OH: hydroxyl radical. PLA₂ enzyme activity, sPLA₂ mRNA and protein expression, PC-PLC activity and PLD2 protein expression were increased after stroke. CCT catalyzes the rate-limiting step in the biosynthesis of PC. CCT activity and protein expression decreased following stroke. Activation of phospholipases and loss of CCT collectively resulted in the loss of PC (41).

Figure. 8

sPLA₂ protein expression increased in ipsi. cortex (IC) vs contra. cortex (CC); treatment with CDP-choline, TNF-α antibody (TNF-α ab), or IL-1 receptor antagonist (IL-1ra) attenuated this increase to CC levels. Relative sPLA₂ protein expression is given as IC/CC ratio.

Figure 9

D609 inhibited proliferation (75 ± 5%) of neurospheres. (A, B) bright field images; (C, D) BrdU incorporation (red); Hoechst nuclear stain (blue). (A, C) controls; (B, D) 3 days after exposure to 100 μM D609.

Figure. 10

Integration of cytokine response and lipid metabolism after stroke (148). Up-regulation of TNF-α and IL-1 differentially affects phospholipases, SMases; CCT and SM synthase in collectively causing loss of PC and SM and release of ArAc and ceramide.