Lipid Composition Of Different Adipose Tissues Of Pig And Regulatory Mechanisms Of Key Differential Lipid On Animal Glucose And Lipid Metabolism

China is a large producer and consumer of meat,and pork occupies a major position in meat production and consumption.Muscle development and fat deposition in animals is a global biological process.Under the current standardized feeding conditions,the lean percentage of Duroc × Landrace × Yorkshire(DLY)pig at finishing weight is between 40% and 60%.The fat percentage of DLY pig is around 30%,but local breed pig can reach 50%,indicating that adipose tissue is a major by-product of pork production.Feed energy is deposited as fat in adipose tissue,which consumes about five times as much energy as is deposited by an equal mass of muscle.Lard is traditionally used oil in the diets of Chinese residents and is one of the main ways of consuming pig fat.Since lard is rich in saturated fatty acids,which is contrary to the low saturated fatty acid intake advocated by modern dietary health,it has been gradually replaced by other fats and oils,and its economic value has been reduced.Balancing the economic value of adipose tissue with feed inputs is the key to increasing quality and efficiency in pork production.Adipose tissue is distributed throughout the body and its occurrence,development and progression have heterogeneous.In recent years,a large number of studies have reported that adipose tissue serves as an important metabolic organ and plays an important role in the homeostasis of glucose and lipid metabolism.Among them,more and more studies have reported the regulatory effects of the key functional lipids,such as fatty acids(linoleic acid,cardiolipin,oxylipins),phospholipids and lipokines on lipid metabolism and energy metabolism in the body.However,there are few studies on the lipid composition and nutritional function of different parts in pig adipose tissue.Therefore,in this study,we served the subcutaneous adipose tissue(SAT)and visceral adipose tissue(VAT)and their corresponding lipids of DLY pig and the local breed of Heigai pig as subjects,analyzed and compared the fatty acid composition and lipid composition of adipose tissues.We verified the nutritional function of different pig adipose by combining with animal experiments.We further screened the key functional lipids through multi-modeling,and investigated the effects and regulatory mechanisms of key functional lipids(phosphatidylserine)on adipocyte lipid metabolism and energy metabolism from the whole animal to the cellular level by combining with in vivo and in vitro experiments.The main research results are as follows:1.Comparative study on the fatty acid composition in different parts of pig adipose tissue and its lardIn this experiment,based on GC-MS targeted metabolomics technology,we comparatively investigated the differences in fatty acid composition of SAT and VAT between the exotic breed DLY pigs,and the excellent local breed Hei Gai pig,from the dimensions of deposition site and breed.It was found that,in DLY pigs,the content of saturated fatty acids(SFA)was significantly higher(C10:0,C12:0,C14:0,etc.)and the content of polyunsaturated fatty acids(PUFA)was significantly lower(C20:2 n-6,C18:2 n-6,etc.)in SAT compared to VAT.And the deposition site had no significant effect on the composition and total amount of monounsaturated fatty acid(MUFA)in adipose tissue.The results of the inter-breed comparison showed that the fatty acid compositions of SAT and VAT were significantly different between DLY pig and Heigai pig,with the contents of MUFA in SAT and VAT of Heigai pig being significantly higher than DLY pig(P<0.001),and the content of PUFA being significantly lower than DLY pig(P<0.001).The differences in fatty acid composition between SAT and VAT lard were further compared.Firstly,the process of hydro-enzymatic extraction of lard was optimized by response surface methodology,and then the medium-and long-chain fatty acid contents of lard from different sources were determined.The aqueous enzyme method was found to retain the fatty acid compositional properties of the original adipose tissue to a large extent.In DLY pigs,VAT lard contained significantly(P< 0.05)higher levels of SFAs(such as C14:0,C16:0,and C20:0)and significantly(P< 0.05)lower levels of PUFAs(e.g.C20:2 n-6)than SAT,and there was no significant difference in the composition of MUFAs when compared to the SAT fats.The results of inter-breed comparisons showed that the fatty acid composition of SAT and VAT lards significant different between DLY and Heigai pigs,the contents of SFA and MUFA in SAT and VAT lards of Heigai pigs were significantly higher and the contents of PUFA were significantly lower compared with those of DLY pigs(P<0.01).The above results suggest that breed and deposition site can affect the fatty acid composition of porcine adipose tissue and its lards,and that the optimized hydro-enzymatic lipid extraction process has less effect on the fatty acid composition and lipid nutritional properties of porcine adipose tissue.2.Effects of ketogenic diets with different sources of lard on glucose-lipid metabolism in miceIn this experiment,we synthesized ketogenic diet(KD)feeds using SAT and VAT fats of DLY pigs,VAT fats of Heigai pigs and high-PUFA commercial lard as lard sources,and investigated the nutritional characteristics of lipids in different porcine adipose tissues and their functions in glucose and lipid metabolism through a short-term KD feeding test,using healthy mice and Diet induced obese(DIO)mice as the research subjects.The results showed that the DLY SAT fat ketogenic diet(KD-SAT)modulated adipose tissue browning to alleviate KD-induced lipid deposition in adipose and liver tissues compared with the DLY VAT fat ketogenic diet(KD-VAT).High-PUFA commercial lard(KD-PUFA)attenuates KD-induced lipid deposition in adipose and liver tissues by maintaining glucose-lipid homeostasis(preserving glucose tolerance and improving insulin sensitivity)in mice compared with commercial lard ketogenic diet(KD-Con).Heigai VAT ketogenic diet(KD-H),through with lower PUFA content,alleviated KD-induced adipose tissue lipid deposition compared with KD-VAT,suggesting that Nutrients other than PUFA in lipids play a role in maintaining glucose and lipid homeostasis in KD mice.The above results suggest that fatty acid composition influences the nutritional properties of lard diets,and that consuming SAT lards with higher PUFA content,or increasing the PUFA content of commercial lard is more metabolically beneficial.3.Analysis of differential lipids in lard from different sourcesIn order to further elucidate the differences in lipid composition in lard from different sources,the present experiment examined the differences in lipid composition of lards by LC-MS non-targeted lipidomics.The results of the study showed that there were significant differences in the lipid composition of lard from different sources.Analyzing the proportions of lipid composition,in addition to the highest proportion of glycerol lipids(GLs),DSAT lard,HSAT lard and HVAT lard had higher proportions of glycerophospholipids(GPs)and sphingolipids(SPs);and DVAT lard had higher proportions of SPs and sterol lipids(SLs).In terms of the composition of lipid subclasses,the content of lipid subclasses such as monoglyceride(MG),sulfatide(ST),phosphatidylglycerol(PG),and phosphatidylinositol(PI)was significantly higher(P<0.05)and Phosphatidylserine(PS),phosphatidylcholine(PC)tended to increase(P<0.1)in the DSAT lard compared to the DVAT lard.The results of comparison between species showed that compared with DSAT lard,HSAT lard contained significantly higher(P < 0.01)content of Sitosterol ester(Si E),significantly lower(P <0.05)content of lipid subclasses such as MG,Diglyceride(DG),Triglyceride(TG),Ceramide(Cer),ST,cardiolipin(CL),PI and PS lipid subclasses were significantly decreased(P<0.05).Lipid subclasses such as cholesteryl ester(Ch E),sphingosine(SPH),ST and PI were significantly higher(P<0.05)in HVAT lard compared to DVAT lard(P<0.05),and there was a trend for PC content to increase(P=0.06).Compared with DVAT lard,the content of lipid subclasses such as cholesteryl ester(Ch E),sphingosine(SPH),ST,and PI was significantly higher in HVAT lard(P<0.05),and there was a tendency for the content of PC to increase(P=0.06);the content of lipid subclasses such as DG,MG,Cer,and phosphoceramide(Cer P)and other lipid subclasses were significantly decreased(P<0.05).We found that the transcriptional activities of adipose tissue in different parts of pigs and the blood metabolic activities under cold stimulation were closely associated with glycerophospholipid metabolism,suggesting that the role of DSAT lard in regulating adipose tissue browning may be related to glycerophospholipids in its lipid composition.Glycerophospholipid-related data from lard lipidomics were then analyzed in combination with published data from animal fat and mitochondrial lipidomics and differential lipid PS was identified as a candidate lipid factor for metabolic health benefits in lard.4.Effect of differential lipid phosphatidylserine on glucose and lipid metabolism in miceThe effects of PS on glucose and lipid metabolism in mice were further investigated using healthy mice and DIO mice.The results showed that PS gavage for 1 week increased the thermogenic capacity of healthy mice in a cold-stimulated environment,and gavage for 2 weeks significantly reduced e WAT deposition and promoted UCP1 protein expression in BAT and i WAT in mice.The results of the PS gavage test in DIO mice showed that PS supplementation for 4 weeks significantly reduced body weight,significantly reduced adipose and liver tissue weight and lipid deposition,and promoted UCP1 protein expression in BAT.Moreover,feeding mice with PS-HFD revealed that15 weeks of PS supplementation alleviated HFD-induced obesity in mice,significantly reduced lipid deposition in subcutaneous adipose tissue and liver as well as cholesterol levels in serum and liver,and promoted the expression of UCP1 in BAT.These results indicated that PS supplementation can promote the expression of UCP1 in mouse adipose tissue,promote lipid metabolism and thermogenesis in adipose tissue,and thus regulate lipid metabolism and inhibit obesity in mice.5.Molecular mechanisms of PS in regulating lipid metabolismThe molecular mechanisms by which PS regulates lipid metabolism and energy metabolism were revealed by cellular experiments.The results showed that exogenous PS promoted UCP1 expression in adipocytes,with enhanced lipolysis and energy metabolism process.PS treatment significantly increased the expression of mitochondrial proteins and PGC1α,which is a key transcription factor that regulates energy metabolism and lipid metabolism in adipocytes.The molecular docking results showed that the binding sites of PS with both UCP1 and PGC1α were associated with protein stability which might contribute to adipose thermogenesisis.Transcriptomics data showed that PS significantly increased the expression of ADCY3 and ADCY7(adenylate cyclase),the key enzymes for c AMP synthesis,and further studies revealed that PS regulated the expression of PGC1α and UCP1 through ADCY3-c AMP-PKA signaling to promote lipid metabolism and energy metabolism in adipocytes.In summary,in this study,we compared the lipid nutritional characteristics of porcine adipose tissue in different sites and their effects on the homeostasis of animal glucose-lipid metabolism through lipidomics,transcriptomics and molecular docking,etc.,and obtained the differential lipid PS,elucidated the effects of the differential lipid PS on fat deposition in animals,and revealed the molecular mechanism of its function of glucose-lipid metabolism through regulation of protein stability via PS-protein binding and ADCY3-c AMP-PKA-PGC1α signal pathway.The results provide a theoretical basis for elucidating the differences in lipid composition of different adipose tissues and the functions of differentiated lipids,and the revelation of the functions of differentiated lipids PS and its regulatory mechanism provides a certain reference for the prevention and treatment of human metabolic diseases such as obesity.