Preliminary Study Of The Mechanism Of High Fat/High Cholesterol Diet Induced Lipid Accumulation In C57BL/6J Mouse Lung

Background:It has been verified that disturbance of lipid metabolismcould result in multiple organ injuries, which include heart and brain bloodvessels, pancrea, liver, and kidney etc. Abnormal lipid accumulation inmacrophages, vascular smooth muscle cells, adipocytes, pancreatic β cells,and kidney mesangial cells were demonstrated asthe pathophysiologicalbasis of abnormal lipid metabolism caused multi-organ injuries.However, the relationship between lipid metabolism disorder andpulmonary injury were rarely studied. In recent years, a number of lipidmetabolism-related gene knockout mice studies prompted that abnormallipid metabolism could lead to lipid accumulation and pathologicalchanges in these gene knockout mice lungs, but not yet see the report inwild-type mice lungs.Objective: In vivo study: to investigate whether high fat/highcholesterol diet (Paigen Diet) could lead lipid accumulation in lungs ofC57BL/6J mice. In vitro study: to investigate whether alveolar type Ⅱepithelial cell is the target cell of lipid accumulation, and whether inflammatory stress aggravate cholesterol accumulation in alveolar typeⅡepithelial cellsand the underlying mechanisms.Methods:1. Sixty six C57BL/6J male mice were randomly dividedinto two groups and treated with either regular diet or high fat/highcholesterol diet, respectively. Fast serum lipid profiles including TotalCholesterol, Triglycerides, Low-density Lipoprotein Cholesterol andHigh-density Lipoprotein Cholesterol were detected at the time-point of12th,16th and20th week, respectively. The aortas atherosclerotic plaqueand lipidosis in lungs were determined by Oil Red O staining.2. Cultured human lung adenocarcinoma cell line A549cells weredivided into4groups, one group was chosen as control and the otherswere treated with high lipid [100μg/ml low-density lipoprotein (LDL)] orinflammatory stress [300ng/ml Lipopolysaccharide (LPS)] or combinationof both [100μg/ml LDL plus300ng/ml LPS] for24hours. Then theintracellular lipidosis was detected by Oil Red O staining. The mRNAsynthesis of sterol regulatory elememnt binding protein2(SREBP2),HMGCoA reductase and low-density lipoprotein receptor (LDLr) wereexamined by real-time PCR and the protein expressions of SREBP2, LDLrand ATP-binding cassette sub-family A member1(ABCA1) weredetermined by Western blot.Results:1. Hypercholesterolemia and aorta atherosclerosis plaqueswere observed in high fat/high cholesterol diet treated C57BL/6J mice. Oil Red O staining of lung at different time-point suggested that high fat/highcholesterol diet could lead to time-dependent lipid accumulation in lungsof C57BL/6J mice.2. Electron microscope results showed that majority of the lipiddeposit in alveolar type II epithelial cells.3. Oil Red O staining showed that lipidosis in the cooperative groupwas the most severe one among the four groups. Compared to the controlgroup, the mRNA synthesis of SREBP2, HMGCoA reductase and LDLrwere inhibited0.54±0.09、0.51±0.06and0.34±0.06times in the highlipid group, respectively. However, the suppressive effect was overriddenby LPS triggered inflammatory stress. The changes of SREBP2and LDLrprotein expression were similar. ABCA1protein expression in high lipidgroup was up-regulated2.78±0.38times compared to the control group,and the effect was overridden by inflammatory stress also.Conclusion:1. High fat/high cholesterol diet could lead totime-dependent lipid accumulation in lungs of C57BL/6J mice, whichpartly due to the disruption of lipid homeostasis in alveolar type Ⅱepithelial cells.2. Inflammation exacerbate cholesterol accumulation in alveolar typeⅡ epithelial cells partly due to the disruption ofSREBP2-LDLr/HMGCoA reductase pathway regulated intracellularcholesterol homeostasis.