| Intracerebral hemorrhage is a devastating stroke subtype with high mortality and morbidity, with harmful effects on human's health. There has no breakthrough medicine to the disease; especially neurological deficits caused by secondary brain injury still exist. The therapeutic object is to relieve secondary brain injury following ICH. Nevertheless, to better understanding the path-physiological mechanisms will be a good way to enhance the clinical therapeutic effect.Vascular reaction has attracted great attention of the medical science circle,which is related to pathologic lesion following ICH, and this academic trends partly coincides with the new rising hypothesis of'toxin hurts brain collaterals'and the therapeusis of'removing toxin and dredging collaterals'in Chinese medicine. Vascular endothelial cell damage exerted important effect on various pathological aspect of collaterals disease. A series of reaction arose as a compensative mechanisms,such as focal vasoconstriction, activation of the coagulation cascade, releasing of thrombin, erythrocatalysis,hemoglobin and its degradation products,which can lead to vascular reaction in hematoma and its surrounding. Vascular endothelial cells are activated and generate cytokine, which may affect neuron and focal microenvironment and lead to homeostasis disorder, cell surface receptor activated, and disorder of the signal transduction. Therefore, it is important to develop a formula to intervene vascular reaction following ICH, which probably enhances the therapeutic efficacy of stroke.An Gong Qing Kai Ling injection is a multicomponent injection derived from the study results of"973"and a latest Chinese drugs pharmaceutics for hemorrhagic stroke, which is developed according to the hypothesis of'toxin hurts brain collaterals'and the therapeusis of'removing toxin and dredging collaterals'. The components are include jasminoidin, baicalin, hyodeoxycholic acid, concha margaritifera usta digest and Radix Notoginseng. The research of"Study on functionary mechanisms of An Gong Qing Kai Ling injection in blocking inflammation following intracerebral hemorrhage"was funded by National High Technology Research Program of China (863 Program, 2002AA2Z3220).We copied experimental intracerebral hemorrhage model of spontaneous hypertensive rats using collagenase infusion in caudate nucleus method. And we used radioimmunoassay (RIA), enzyme linked immunosorbent assay (ELISA), immunohistochemistry stain, hematoxylin- eosine stain (HE) and Weatern Blotting to observe:①the time course of tumor necrosis factor-alpha(TNF-α), interleukin-1 beta(IL-1β) and intercellular adhesion molecule-1(ICAM-1) following intracerebral hemorrhage;②the relationship between tumor necrosis factor-alpha and inflammatory reaction, expression of interleukin-1 beta(IL-1β) and intercellular adhesion molecule-1(ICAM-1) and activation of neutrophil;③the pathomorphism changes in brain following ICH and the effect of An Gong Qing Kai Ling Injection;④the effection of An Gong Qing Kai Ling Injection on the expression of proinflamatory cytokine--TNF-αand IL-1β;⑤the effection of An Gong Qing Kai Ling Injection on the intercellular adhesion molecule-1 and activation of neutrophil;⑥the effection of An Gong Qing Kai Ling Injection on the expression of nuclear factor-κB and heat shock protein 70;⑦the effection of An Gong Qing Kai Ling Injection on the number of apoptotic cells. Results are as follows:1. The contents of TNF-α,IL-1βand ICAM-1 are significant elevated at 24 hours,48 hours, 72 hours and 7days following ICH.2. The concentration of IL-1βand sICAM-1 in brain or blood degrades after treated by exogenous anti- TNF-αmonoclonal antibody.3. Typical widespread hemorrhagic focus is formed at 48 hours after infusion, with small hematoma formation. The hemorrhagic focus is charactered by erythrocyte and inflammatory cell (neutrophil) infiltration, edema, nuclear structure of neurons are obscure, denaturation and necrosis, and sometimes neuronophagia can be seen. The tissues around hemorrhagic focus are loose, accompanied by infiltration of lymphocyte, cellular edema in neurons and glia cell, vascular endothelial cell swelling, cell aggregation leading to capillary blocked. An Gong Qing Kai Ling Injection can relieve the degree of looseness, The same as the inflammatory cell (neutrophil) infiltration, edema and vascular endothelial cell damage, without accumulation of blood cells, especially in middle and high dose groups. At 14 days following intracerebral hemorrhage, the hematoma was absorbed and its volume became small, and glial scar was formed with lots of microglial cells, vascular proliferation. Sometimes cavity was seen. There were microglial cells around hematoma which had been swallowing hemosiderin. The volume of glial scar and the number of cavity decreased in An Gong Qing Kai Ling Injection treated groups and control group, especially in middle and high dose groups.4. It has been confirmed that An Gong Qing Kai Ling Injection can prevent the expression of TNF-αat 48hours following ICH by RIA and immunohistochemistry stain, however the preventive effect diminished at 7days after ICH. The inhibitive effect of An Gong Qing Kai Ling Injection on the expression of IL-1βcan last from 48hours to 7 days following ICH.5. It has been found that An Gong Qing Kai Ling Injection can prevent the expression of ICAM-1 at 48 hours following ICH by ELISA and immunohistochemistry stain, and the effect can last from 48 hours to 7 days following ICH. The inhibitive effect of An Gong Qing Kai Ling Injection on the activation of neutrophil was significant at 48 hours following ICH, which weakened at 7 days.6. It has been found that An Gong Qing Kai Ling Injection can prevent the expression of NF-κB at 48 hours following ICH by immunohistochemistry stain. However it had no effect on the generation of HSP-70.7. It has been found that An Gong Qing Kai Ling Injection can decrease the number of apoptotic cells at 48 hours following ICH by immunohistochemistry stain.In summary, An Gong Qing Kai Ling Injection (low dose, middle dose and high dose) can prevent the expression of TNF-α, IL-1β, sICAM-1, MPO, NF-κB and decrease the number of apoptotic cells with dose-dependent manner at 48 hours and 7 days .following ICH. It is thus clear that An Gong Qing Kai Ling Injection can extensively suppress inflammatory reaction and decrease the number of apoptotic cells in acute stage.We can make such conclusion as follows:1. The content of cytokine was elevated and its peak between 24 hours and 72 hours following ICH, soon after descending and rising again ay 7days after ICH. Therefore, we select 48 hours and 7 days as the time window to administration.2. The content of IL-1β, sICAM-1 and MPO decreased after treated by TNF-αmonoclone antibody. It is thus clear that neutralization TNF-αin circulation can effectively suppress the expression of proinflamatary cytokine and adhesion molecule, as well as the activation of neutrophil.3. An Gong Qing Kai Ling Injection can effectively interfere the generation of TNF-αand IL-1β, thus can prevent the trigger of inflammatory reaction. Its action mechanism may be related with its prevention on the NF-κB.4. An Gong Qing Kai Ling Injection also can surppress the activation of neutrophil and the expression of ICAM-1. That is to say, it possess dual effect on cell adhersion, it can not only interfere VEC, neuron and glial cell in expression ICAM-1, but also suppress the actication of neutrophil, accordingly can decrease the neutrophil infiltration in hemorrhagic focus and exert neurprotective effect.5. An Gong Qing Kai Ling Injection also can decrease the number of apoptotic cells and prevent neuronal cell death, which can improve the outcome in clinic. Its mechanism may be related to its suppression on TNF-α, IL-1β, HSP-70 and NF-κB.
|
PDF Full Text Request