| AIM:As the basic structural and functional unit of the blood vessels, ECs play an important role in physiological metabolism. It acts as a barrier to regulate the substance exchange of the blood vessels, and to maintain the stabilization of the innercellular environment. The regulational mechanism of ECs turned abnormal in harmful conditions, which would first lead to stress reaction in order to defend injury. Meanwhile, the change of phenotype in ECs takes part in tissue repair like injuried basement membrane's repair. All the abnormal regulation will induce the changes of cellular morphology and function and may become one of the factors leading to various diseases. Acute lung injury contains two phases-alveolitis and tissue recovery. However, vascular reaction is the key event in inflammation, among which the opening of ECs' tight junction and the elevation of vascular permeability are the primary manifestations Along with the recovery , the functions of PMVECs become normal. However, previous research show that BLM-induced PMVECs'tight junction persistent opening while the ECM were deposited excessively. This cellular barrier's dysfunction provides a pathway for monocyte emigration which releases various fibrotic factors. Upon PF, Fb's proliferation and transformation into Mfb become a key event, which needs a lot of cytokine. Our study demonstrated that different causative agent could lead to similar cytokines's expression up-regulation. Other reserches prove that important fribotic cytokines, like TGF-β1, CTGF, IGF etc., are aslo expressed in normal physiological metabolism. These indicate that relative cytokines are not only expressed in fibrosis, but aslo regulate the normal metabolism. It is confirmed that cytokines' expression are relevant to cellular function. For instance, as one of the primary blood cells, mononuclear cells don't have excretive function until they migrate into interstitial tissue and transform into microphages. The changed phenotype of mononuclear cells plays an important role in resisting tissue damage. CTGF excreted by Ecs can promote the Fb's transformation into MFb. In harmful conditions, the opening of ECs'TJ means that cellular morphological and function have changed. But BLM-induced damage seems to indicat that the regulational mechanism were wrong. Based on that, ECs of different phenotypes have incorporated abnormal functions of secreting CTGF, which is one of the evoked agent in PF. This idea wasn't reported. In this research I observed the microstructure changes in pulmonary microvascular endothelial cells(PMVEC) under bleomycin induced lung fibrosis and streptococcus pneumoniae(SP)-induced pneumonia. I also observed the concurrent variations in CTGF levels. The results could, in part, explain the relationship between the PMVECs' phenotype and lung fibrosis.METHODS:45 SD rats, female and male, weigh 150±20 g(provided by the experimental animal center of the Fourth Military Medical University), were divided randomly into 3 groups, which were bleomycin group, pneumococcal pneumonia group and control group(15/group). BLM-A5(Tianjin Taihe pharmaceuticals, Inc. 8mg, batch number 041205) was diluted to 4 g/L by physiological saline solution. The BLM group received BLM (5mg/kg) treatment intratracheally. The rats was then erected and spun during above procedure for full distribution of BLM in the lungs. The rats in the pneumococcal pneumonia group received activated pneumococcus 0.15 mL (CFU 3×108) and the rats in the control group received saline with the same procedures. Rats from each group were randomly chosen on day 1, 3, 7, 14, 21and 28 and sampled to determine the changes in PMVECs' morphological features. Rats from each group were executed by saline perfusion via right ventricle, the lung was then perfused and stabilized with 4% paraformaldehyde, then paraffin slices were stained by HE/VG; a small piece of the remaining lung tissue from each group was stabilized with 3% glutaraldehyde. Connective tissue growth factor(CTGF) was determined by ELISA and immunohistochemistry.RESULTS:1. Light microscopy showed obvious changes between the pneumonia group and BLM group. In pneumonia group, the histology of the lung manifested a typical acute inflammatory changes with no permanent damages. In the BLM group, the destruction proceeded progressively. The inflammatory cell started to effuse into septa and alveolar space from the 3rd day, homever, the inflammatory changes went on progressively. Fb bengan to accumulate in lung interstitial tissue at the 14th-21st day and collagen deposition in the septa between the alveoli was evident at the 28th day in the end.2. Electron microscopy showed that the proliferation of PMVECs in pnernonia group started from the 7th day. PMVECs'TJ was openned and micropinocytic vesicles significantly decreased at that stage. Then the morphology of PMVECs returned to normal at the 14th day in this group. In BLM group, PMVECs had digitations and platelets adhered to the luminal surface. PMVECs were in proliferative stage with increased volume. Micropinocytic vesicles increased and TJ kept opening from the 7th day to the 21st day, and a large quantity of collagen fibers deposited surrouding PMVECs at 28th day.3. SP challenge which was in great contrast with those with BLM challenge which began to increase at the 7th day and reach the peak (19.45±0.41pg/mL) at the 21st day after BLM challenge(P<0.05,n=15).4,CTGF expression elevated in lung and PMVECs under BLM challenge compared with those under SP. (P<0.05, n=15).CONCLUSION:1,BLM induces the changes of morphological and dysfunction in PMVECs. The phenotype of PMVECs might contribute to the pathogenesis of bleomycin- induced pulmonary fibosis, which is one of the key agent for CTGF's abnormal expression.2,The changes of phenotype, from the barrier type to the contraction type, might be closely related to PMVECs'metabolism. Metabolic disturbances of [Ca2+]i may contribute to the changes of phenotype and the abnormal secretion in PMVECs.
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