Studies on cytokines and chemokines in cerebrovascular diseases and experimental cerebral ischemia

Ischemic brain injury secondary to arterial occlusion, a major cause of stroke, is characterized by acute local inflammation, which involves the accumulation of polymorphonuclear neutrophils and other immune cells. Tissue damage resulting from ischemic stroke can be reduced in experimental animal models by blocking leukocyte accumulation. Cytokines and chemokines are factors that regulate leukocyte trafficking, and could play a pivotal role in the accumulation of leukocytes at sites of brain ischemia. The factors responsible for secondary brain damage in cerebral ischemia are incompletely defined, and studies in human stroke on cytokines and chemokines have been few.

Aims of the study: 1. To investigate mRNA expression of the chemokines IL-8, MCP-1, MIP-1alpha and MIP-1beta by blood mononuclear cells (MNC) in patients with acute ischemic stroke and healthy controls; 2. to study mRNA expression of IL-8 and MIP-1alpha in parallel with the pro-inflammatory IL- I beta and IL- 17 by blood MNC in a prospective study of ischemic stroke; 3. to investigate cytokines with anti-inflammatory properties, like IL-4 and IL-10 secreted by blood N/fNC from patients with acute stroke vs. healthy controls, using ELISPOT assays; 4. to examine leukocyte and cytokine alterations both systemically and locally in the brain over the course of an experimental rat model of permanent middle cerebral artery occlusion (pMCAO); 5. to analyze involvement of dendritic cells (DC) in pMCAO in the rat.

Results: Ischemic stroke in the human is associated with high numbers of IL-8 mRNA expressing blood MNC over the time interval of 1-7 days between onset of symptoms and examination. IL-8 concentrations in plasma correlated positively to IL-8 mRNA expressing blood MNC in examined patients. In the prospective ischemic stroke study, most patients with ischemic stroke had clearly elevated numbers of IL- I beta, IL-8 and IL- 17 mRNA expressing blood MNC 1-3 days after onset of symptoms compared to healthy controls. At follow-up after 20-31 days, numbers of IL-8 mRNA expressing blood MNC were lower than during the acute stage, and numbers of IL- I beta and IL- 17 mRNA expressing blood MNC had returned to levels in healthy controls. Numbers of MIP-1alpha mRNA expressing blood MNC did not differ between patients with ischemic stroke and healthy controls at any timepoint examined. IL-10-secreting blood MNC are elevated in patients with ischemic stroke as well as in intracerebral hemorrhage when examined up to 10 days after symptom onset. Levels of IL-4 secreting blood MNC are not different in stroke compared to healthy controls. Elevated levels of IL-17 and IFN-gamma mRNA are observed both in the ischemic hemispheres vs. sham, and systemically at 1 h to 6 days after experimental pMCAO. Increased levels of T cells expressing IL-17, IFN-gamma IL-8, MIP-2 and IP-10 mRNA in the ischemic vs. sham hemispheres are present in cerebral ischemia. OX 62+ DC are present at 1 h in the ischemic hemispheres, peaking at 6 days after pMCAO. Activated DC, expressing MHC class II (OX 62+OX 6), peak at 6 days in the ischemic vs. sham hemispheres. Microglia develop into OX 62+ DC in the ischemic hemispheres during the course of cerebral ischemia.

Conclusions: Systemic increases of pro-inflammatory IL-8, IL-1beta and IL-17, and the anti-inflammatory cytokine IL-10 early after human ischemic stroke indicate a role for these mediators in potentiating the inflammation secondary to brain ischemia. This is further supported by upregulated mRNA levels of these and other cytokines and chemokines in the ischemic hemispheres and systemically of pMCAO-operated rats. Increased levels of T cells expressing cytokines and presence of DC indicate a role for these immune cells in cerebral ischemia.

List of scientific papers

I. Kostulas N, Kivisakk P, Huang Y, Matusevicius D, Kostulas V, Link H (1998). "Ischemic stroke is associated with a systemic increase of blood mononuclear cells expressing interleukin-8 mRNA. " Stroke 29(2): 462-6
https://pubmed.ncbi.nlm.nih.gov/9472890

II. Kostulas N, Pelidou SH, Kivisakk P, Kostulas V, Link H (1999). "Increased IL-1beta, IL-8, and IL-17 mRNA expression in blood mononuclear cells observed in a prospective ischemic stroke study. " Stroke 30(10): 2174-9
https://pubmed.ncbi.nlm.nih.gov/10512924

III. Pelidou SH, Kostulas N, Matusevicius D, Kivisakk P, Kostulas V, Link H (1999). "High levels of IL-10 secreting cells are present in blood in cerebrovascular diseases. " Eur J Neurol 6(4): 437-42
https://pubmed.ncbi.nlm.nih.gov/10362896

IV. Li H, Kostulas N, Huang Y, Xiao B, van der Meide P, Kostulas V, Giedraitas V, Link H (2001). "IL-17 and IFN-gamma mRNA expression is increased in the brain and systemically after permanent middle cerebral artery occlusion in the rat" J Neuroimmunol 116(1): 5-14 (In Print)
https://pubmed.ncbi.nlm.nih.gov/11311324

V. Kosulas N, Li HL, Huang YM, Xiao BG, Kostulas V, Link H (2001). "Dentric cells are present in the ischemic brain after permanent middle cerebral artery occlusion." (Submitted)