The Study On The Regulation And Mechanism Of3T3-L1Preadipocyte Differentiation By Type â…  Collagen

Part1. Different expression of Type Ⅰ Collagen in Adipose TissueObjective:. Type ⅠCollagen (Col Ⅰ) is one of the most important components ofExtracellular matrix (ECM), it is highly expressed in adipose tissue. But there are noreports about the different expression of Type Ⅰ Collagen in adipose tissue. The study aimsto analyze the different expression of Type Ⅰ Collagen in subcutaneous and visceral adiposetissue of normal weight human, c57/bl6mice, obese patients and mice to elucidate itsrelationship with obesity.Methods:10human’s subcutaneous and visceral adipose tissue from normal weight andobese patients undergoing elective cholecystectomy operation.12male c57/bl6mice wererandomly divided into2groups, normal diet group (NC, n=6), high fat diet group (HFD,n=6). Western blot was used to assay the expression of Type Ⅰ collagen in subcutaneous andvisceral adipose tissues, and analysis its relationship with body weight.Results:1. Type Ⅰcollagen was expressed in subcutaneous and visceral adipose tissue ofhuman and mice.2. The expression of Type Ⅰ collagen in visceral adipose tissue was lowerthan subcutaneous adipose tissue of obese human, but they were all lower than normalweight human. They had significant differences (P<0.05).3. The expression of Type Ⅰcollagen in visceral adipose tissue was lower than subcutaneous adipose tissue of obesemice, but they were all lower than normal weight mouse. They had significant differences(P<0.05). Type Ⅰcollagen had negative relationship with body weight.Conclusion:1. Type Ⅰcollagen was expressed in subcutaneous and visceral adiposetissue of human and mice，which was lower in visceral adipose tissue than in subcutaneousadipose tissue.2. Expression of Type Ⅰ collagen in subcutaneous and visceral adipose tissueof obese human were all lower than normal weight human.3. Expression of Type Ⅰ collagenin subcutaneous and visceral adipose tissue of obese mice were all lower than normalweight mice. Type Ⅰcollagen had negative relationship with body weight. Part2. Effect of Type Ⅰ Collagen on the differentiation of3T3-L1preadipocyte and its underlying molecular mechanismsObjective: To study the effect of Type Ⅰ collagen (Col Ⅰ) on3T3-L1preadipocytesdifferentiation into mature adipocytes and the underlying.molecular mechanisms.Methods: During the process of3T3-L1preadipocytes differentiation, we added Type Ⅰcollagen and cathepsin k. We established control group (CON), cathepsin k group（CTSK），Type Ⅰ collagen group (Col Ⅰ), Type Ⅰ collagen and cathepsin k group (ColI+CTSK). The proliferation of3T3-L1preadipocytes was detected by MTT methods.Through Oil Red O staining, we observed the morphological of mature adipocytes.Western blot and RT-PCR were used to detect the expression of Type Ⅰ collagen,peroxisome proliferators-activated receptor γ (PPARγ) and CCAAT/enhancer-bindingprotein ɑ (C/EBPɑ) after3T3-L1pre-adipocytes differentiation.Results:1. Different concentration of Type Ⅰ collagen did not affect the proliferation of3T3-L1preadipocytes compared with the control.2. With the increased concentration ofType Ⅰ collagen, Oil Red O staing became shallow suggesting that mature adipocytesgradually decreased. But when added Type Ⅰ collagen with cathepsin k during the processof3T3-L1cells differentiation, Oil Red O staing became deep, suggesting matureadipocytes increased.3. The expression of Type Ⅰ collagen mRNA and protein weregradually increased in a dose-dependent manner with the increased concentration of Type Ⅰcollagen, but PPARγ and C/EBPɑ mRNA and protein were gradually decreased in adose-dependent manner, and significantly when the concentrations of Type Ⅰ Collagen were10-5mol/l and10-4mol/l respectively(P＜0.05).4. The expression of Type Ⅰ collagen mRNAand protein were significantly decreased in CTSK group than in control, PPARγ andC/EBPɑ mRNA and protein were increased than the control group. When added cathepsink with Type Ⅰ collagen during the process of3T3-L1cells differentiation, Type Ⅰ collagenmRNA and protein were significantly decreased than the Type Ⅰ collagen group, PPARγ andC/EBPɑmRNA and protein were increased, there were significant differences (P<0.05).Conclusion: Type Ⅰcollagen did not affect the proliferation of3T3-L1preadipocytes, butit did dose-dependent inhibit adipogenesis of3T3-L1preadipocytes, decreased theexpression of PPARγ and C/EBPɑ mRNA and protein levels, and increased the expressionof Type Ⅰ collagen mRNA and protein levels. Cathepsin k promotes adipoctes differentiaton.When added Type Ⅰ collagen with cathepsin k during the process of preadipocyte differentiation, PPARγ and C/EBPɑ in mature adipocytes were increased than type Icollagen group, suggesting that type I collagen may inhibit adipocyte differentiation bydecreasing the expression of PPARγ and C/EBPɑ.Conclusion1. Type Ⅰcollagen was expressed in subcutaneous and visceral adipose tissue of human andmice. It is higher in subcutaneous adipose tissue than in visceral adipose tissue.2. Expression of type I collagen in subcutaneous and visceral adipose tissue of obese humaand mice were all lower than normal weight human and mice.3. Type Ⅰcollagen had negative relationship with body weight.4. Type Ⅰcollagen did not affect the proliferation of3T3-L1preadipocyte,5. Type Ⅰcollagen could dose-dependent inhibit adipogenesis of3T3-L1preadipocytes, itmay inhibit adipocyte differentiation by decreasing the expression of PPARγ and C/EBPɑ.