How leptin regulates the activity, cell adhesion and chemotaxis of ischemic neutrophils to protect brain from ischemic injury

• Main Authors: An-Jen Chang, 張安蓁
• Other Authors: 葛其梅
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/55289382353583723949

碩士 === 國立中興大學 === 生命科學系所 === 101 === Brain injection with leptin has been demonstrated to be able to reduce the permeability of blood brain barrier (BBB), brain infiltration of neutrophils and the brain infarction after ischemia/reperfusion (I/R) insult. The underlying mechanisms responsible for leptin-prevented brain infiltration of neutrophils and the consequent brain infarction (inflammation) remain still unclear and to be investigated. The primary goal of this thesis study was to know the direct impact of leptin upon neutrophils under an in vitro ischemic condition, based on changes in their survival, cell adhesion capability, degree of chemotaxis and inflammatory potential. In the study, an in vitro ischemia model we have previously developed was applied by growing neutrophils or endothelial cells (ECs) in culture medium deprived with glucose, oxygen and serum, so called GOSD to mimic the in vivo ischemic condition. Neutrophils to be analyzed were isolated from naive or ischemic Spraque-Dawley (S.D) rats at 24 h after reperfusion started. The results showed that GOSD-induced cell death in neutrophils was further exacerbated by leptin likely due to the increased expression of p-ERK. Neutrophils isolated from the ischemic rats survived better than that from the naive animals after GOSD/leptin co-treatment. Leptin also inhibited the cell-cell interaction between neutrophils and endothelial cells under GOSD stress and the interruption may result from the reduced expression of endothelial ICAM-1. Protein expression of beta2 integrin (counter part of ICAM-1) in GOSD neutrophils was not altered by leptin at present. Mobility of the neutrophils moving toward n-formyl-Methionin-Leucine-Phenoalamin (fMLP; known as a pro-inflammatory classical chemotaxin to neutrophils) was significantly increased by GOSD but inhibited by leptin, which likely was due to the increased expression of P38 and PPAR-gamma but reduction of p-Rho A by leptin. Leptin also revealed a “non-classical chemotaxin” role to naive and ischemia-primed neutrophils under normal or GOSD condition, by attracting neutrophils but without aggravating (in fact inhibiting) their inflammatory activities (release of NO and superoxides). In addition, leptin can also inhibit the movement of neutrophils toward fMLP but the significant inhibition was only seen in the naive neutrophils under GOSD condition and the ischemia-primed neutrophils under normal condition. In overall, under the GOSD condition, leptin was able to decrease the number of surviving neutrophils, inhibit the cell adhesion capability of neutrophils toward vascular endothelial cells, suppress the pro-inflammatory potential of neutrophils, attract neutrophils and prevent their chemotaxis to the inflammatory fMLP. Through these mechanisms leptin was able to lower down the number of neutrophils within the ischemic brain that subsequently can reduce the degree of brain infarction (inflammation) and serve its brain protection role in stroke. The results have uncovered the fact about how leptin can protect ischemic brain by limiting the survival, activity and mobility of neutrophils; also provided valuable information for future screening or developing of new anti-stroke drugs. Therapeutic potential of PPAR-gamma agonist, Rho A inhibitor or neutrophil depletion, are all worthy of note.