Research In The Mechanism Of Neutrophils Functioning In Perfluoroisobutylene Inhaled Lung Injury

Perfluoroisobutylene (PFIB) is a gaseous hypmolecule organics with abundant fluorine in common temperature, and its molecular formula is (CF3)2C=CF2. The inhalation toxicity of colorless and odorless PFIB is ten times more than phosgene. Jot inhalation of PFIB will result in headache, cough, thoracalgia, dyspnea and fever, even more lead to acute lung injury (ALI). So far, the mechanism of ALI induced by PFIB is still incompletely clear, and the specific medicine and therapeutic measure are also totally unavailable yet. The research results from laboratories of both domestic and international in vivo on animals, including our laboratory have shown that polymorphonuclear leukocyte (PMN) play critical role in ALI induced by PFIB, especially in the prophase of ALI. The injury of PMN in the prophase is important for ALI even more in whole course of disease to death; In advanced stage of ALI induced by PFIB, other factors (such as alveolar macrophage, AM) are probably more important instead of PMN, AM may also play a certain role in the prophase of ALI. In order to elucidate the interaction of PMN, AM and pulmonary microvascular endothelial cell (PMVEC) in ALI induced by inhalation of PFIB, a series of researches basing on PMN isolated from blood, AM from bronchoalveolar larage fluid (BALF) and Rats pulmonary microvascular endothelial cell ( RPMVEC) as followed are made.First, Western Blot is used to examine the expression of ICAM-1 in the lung basing on the model of ALI in mice induced by PFIB inhalation. The results show that the expression of ICAM-1in the lung reach topmost at 1h and reduce to the normal level after the inhalation of PFIB; the expression of ICAM-1 in the lung of mouse in GdCL3 group in which AM is exhausted before the inhalation of PFIB by GdCl3 is similar to normal control group (physiological saline group) and decreased obviously compared with PFIB group. It is thus evident that the expression of ICAM-1is increased obviously in the prophase of ALI and the increased expression of ICAM-1 in the lung is related with AM in ALI induced by PFIB inhalation.Further, basing on RPMVEC subcultured stably, the effect of the culture supernatant of AM isolated from Rats at 1h after inhalation of PFIB on the expression of adhesion molecules (ICAM-1 and E- selectin) on RPMVEC. The results show that compared with normal control group(NCG), under the effect of the culture supernatant of AM isolated from Rats 1h after inhalation of PFIB, the expression of E-Selectin and ICAM-1 of RPMVEC is increased. The result indicates that the increased expression of ICAM-1 and E-Selectin on RPMVEC is related with the activation of AM in ALI induced by PFIB inhalation, which makes for the adhesion and sequestration of PMN in capillary.Following the researches above-mentioned, percoll density gradient centrifugation method is used to isolate the polymorphonuclear leukocyte (PMN) in Wistar Rats abdominal arterial blood and the effect of the culture supernatant of AM isolated from Rats 1h after inhalation of PFIB on the adherence of PMN on RPMVEC is studied. It is found that Compared with normal control group (NCG), the adherence of PMN on RPMVEC is increased under the effect of the culture supernatant of AM isolated from Rats 1h after inhalation of PFIB. At the same time, Superoxide dismutase (SOD) test box is used to detect the activity of SOD of RPMVEC in the culture supernatant and MTT colorimetric technique is used to examine the activity of RPMVEC after the adherence of PMN. The results show that Compared with normal control group (NCG), the activity of SOD and the survival activity of RPMVEC is decreased obviously under the effect of the culture supernatant of AM isolated from Rats 1h after inhalation of PFIB. The result shows that the increase in the adherence of PMN on RPMVEC and the increase in the damage of RPMVEC mediated by PMN in ALI induced by PFIB are related with the activation of AM.If the solution of N-acetylcysteine (N-AC) and protease inhibitor (PI) are given when the suspensions of PMN is added, the result shows that compared with the group only under the effect of the culture supernatant of AM isolated from Rats 1h after inhalation of PFIB, the activity of RPMVEC is increased in N-AC, PI and N-AC+PI group. Videlicet N-AC and PI can attenuate the damage of vascular endothelial cells induced by PMN.The experimental results above-mentioned show that in the ALI induced by PFIB inhalation, the activation of vascular endothelial cells, the adhesive attraction on vascular endothelial cells and activation of PMN and the injury of vascular endothelial cells mediated by PMN are enhanced. These processes are all related with the activation of AM. Proteinase Inhibitors and Reactive Oxygen Species scavenger (N- acetylcysteine) can produce a certain protective effect in the injury of vascular endothelial cells mediated by PMN. In a word, PMN and AM all play important role in ALI induced by PFIB. PMN mediate the direct injury of lung tissue; the activated AMs effects the function of PMN through the influence on the expression of adhesion molecules of RPMVEC, the adherence of PMN on RPMVEC and the activation of PMN and further effect the injury of tissue by PMN.