Effects Of Compound 48/80 On Plaque Induced By Perivascular Common Carotid Collar Placement In Apolipoprotein E-knock Out Mice

Objective Researches on the relationship between mast cells and atherosclerosisare mostly in pathological observation and cytology experiments. To investigate the effects of mast cells degranulation on plaque development and theirs possible mechanisms in animal experiments, we dealed apolipoprotein E-knock out mice which had been placed perivascular common carotid collars with mast cells degranulator - Compound 48/80.Methods 40 apoE mice were fed a western-type diet from 10 weeks old. After 2weeks,mice were operated with perivascular right common carotid collar placement. Mice were anesthetized by intraperitoneal injection with pentobarbital.The bilateral common carotids were dissected free from the surrounding connective tissue through a sagittal anterior neck incision.Silastic collar was placed around the right common carotid, and its axial edge was approximated by placement of 3 circumferential silk ties, which resulted in lumen stenosis.Subsequently, the entry wound was closed and the animal was returned to its cage for recovery from anesthesia. 4 weeks after surgery,mice were divided into 2 groups and treated for 7 days as follows: experimental mice were intraperitoneally injected with Compound 48/80,0.5mg/ kg; control mice were intraperitoneally injected with an equal volume of D-Hanks; every other day,together 4 times. 30 minutes after the fourth injection,blood was collected from the orbit for measurement of serum lipids and activity of tryptase .The bilateral common carotids were in situ perfusion fixed with 10% neutral buffered formalin and removed.Then fixed in neutral buffered formalin, and embedded in paraffin. Sections were routinely stained with hematoxylin and eosin.Corresponding sections on separateslides were stained with toluidine blue and immunohistochemically with antibodies against a macrophage-specific antigen(Mac3), a-smooth muscle actin, IL-lP,vWF and VE-cadherin. Simultaneously, bFGF was detected by in situ hybridization and immunofluorescence.Plaque area was quantified at maximum cross section.The degree of lumen stenosis was expressed by the intima-lumen ratio.The images were analyzed with a Olympus BX51 microscope and HMIAS2000 software to distinguish plaque area,the degree of lumen stenosis, the percentage of degranulated mast cells, the density of intraplaque neovessels, the cases of intraplaque haemorrhage and the expressions of Mac3,a-smooth muscle actin,IL-l 3 ,VE-cadherin and bFGF in plaque of common carotids collar placement between the two groups.Results There were not pathological change in common carotids non-collar placement but atherogenesis in common carotids collar placement of both groups.Significant increase in plaque area(5.85±0.75xl0Vm2 in Compound 48/80;0.86±0.28xl04/am2 in control group, i'<0.05),the degree of lumen stenosis (81 ±15% in Compound 48/80; 41±12% in control group, P<0.05) and the percentage of degranulated mast cells (80.6 ±17.8% in Compound 48/80; 13.5±4.1% in control group, P<0.05). The activity of tryptase was 0.54±0.29 u/L versus 0.36±0.13 u/L in Compound48/80 and control group,respectively (F<0.05).The expressions of Mac3,a-smooth muscle actin, bFGF, IL-1 P and VE-cadherin in plaque of common carotids collar placement were more in Compound 48/80 than in control group.The density of neovessels and the cases of intraplaque haemorrhage were increased in Compound 48/80 group.Conclusions1. Perivascular common carotid collar placement can accelerate atherosclerotic plaque formation in apolipoprotein E-knock out mice.2. Compound 48/80 increases plaque area of maximum cross section and the degree of lumen stenosis in common carotids collar placement. The mechanism is possibly that Compound 48/80 promotes proliferation of smooth muscle cells and accumulation of macrophages by stimulating mast cells degranulation.3. Compound 48/80 enhances the expression of IL-1 P in plaque.