The Effect Of Choline On Growth Performance And Lipid Metabolism In Pekin Ducks And Its Regulation Mechanism

Five experiments in four parts were conducted to study the effect of dietary choline level on growth performance and lipid metabolism in Pekin ducks and its regulation mechanism.Expt.1was conducted to study the effect of dietary choline on growth performance, carcass quality, and liver lipid content of starter and growing Pekin ducks in order to obtain choline requirements. In starter Pekin ducks, the test was conducted to study the effect of dietary choline level on growth performance and carcass quality in Pekin duck from1to21days of age. Three hundred eighty-four1-d-old male White Pekin ducks were randomly allotted to eight dietary treatments with8dietary choline levels (302,496,778,990,1182,1414,1625, and1832mg/kg), each containing6replicate pens with8birds per pen. The basal diet was formulated using corn, cron starch, and isolated soybean protein. The results showed that the body weight gain and feed intake were improved with the increase of dietary choline level (P<0.05) and this response reached a plateau when dietary choline was above990mg/kg. Weight gain and feed intake of ducks at21day of age increased from32.8g/d and46.9g/d to47.3g/d and69.4g/d, increased by44.2%and47.9%, respectively. The yield of breast meat and abdominal fat increased gradually with increasing choline level (P<0.05), but no significant differences in leg meat yield were observed between treatments (P>0.05). According to broken-line regression, the choline requirement of Pekin ducks from1to21d of age for weight gain, feed intake, and abdominal fat yield was810mg/kg (P=0.0116, R2=0.832),823mg/kg (P=0.019, R2=0.792), and1048mg/kg (P=0.0020, R2=0.916), respectively. In growing Pekin ducks, A dose-response experiment with6dietary choline levels (0,342,779,1285,1662, and1962mg/kg) was conducted to study the effect of dietary choline on growth performance, carcass quality, and liver lipid content in Pekin duck from21to42d of age. Ninety21-d-old male White Pekin ducks were allotted to six dietary treatments, each containing5replicate pens with3birds per pen. The basal diet was formulated using corn starch and isolated soybean protein. The results showed that significant positive effects of dietary choline on weight gain, and feed intake were observed (P<0.05). Dietary choline supplementation significantly decreased liver lipid and triglyceride content and increased liver phospholipids of Pekin ducks (P<0.05). In addition, dietary choline level significantly affected the yield of abdominal fat, leg, and liver. Liver lipid and triglyceride content were highest at the choline level of0mg/kg and improved10.09%and15.79%compared with that at the choline level of1285mg/kg. According to broken-line regression analysis, the choline requirements of Pekin ducks from21to42d of age for weight gain, feed intake, abdominal fat yield, liver phospholipids, and total liver lipid were980,950,1010,888, and1130mg/kg. Pekin ducks needed more choline to prevent excess liver lipid deposition than to maintain growth. Liver phospholipids content was a new sensitive indicator for evaluating choline requirement of Pekin ducks.On the basis of the expt.l, expt.2was conducted to study the effect of choline deficiency on bone development and related metabolism index of plasma. Four choline levels (302,496,778,990mg/kg) were selected to finish it. The results suggested that choline-deficient ducks showed various degrees of choline deficiency disease on the leg and the ducks showed the symptom with mental fatigue, skelasthenia, hock enlargement, bowed leg, and perosis. The symptom reduced gradually with increasing choline level and was prevented completely when dietary choline was990mg/kg. The ducks with bowed leg were accompanied with slipped tendon. Choline deficiency significantly decreased tibia weight, length, width, and backbone length (P<0.05). Dietary choline level affected significantly the plasma HCY, ALP, and TrACP levels(P<0.05), but no significant differences in plasma HOP, Cys, Ca, and P level were observed (P>0.05). In a word, choline deficiency model could be built by the diet based corn, corn starch, and isolated soy protein. Choline deficiency inhibited growth, osteoblast, and osteoclast activity, and decreased the tibia parameters to aggravate bone dysplasia.On the basis of the expt.1, Expt.3was conducted to study the effect of dietary choline on lipid metabolism of plasma and liver in Pekin duck at21days of age. The results showed that dietary choline supplementation significantly increased plasma TG, LDL-CHO, VLDL, and FFA level and decreased GPT activity (P＜0.05). Plasma TG and VLDL levels increased from0.62mmol/L (302mg/kg) to0.80mmol/L (1414mg/kg) and from0.63mmol/L (302mg/kg) to1.13mmol/L (1414mg/kg), respectively, and improved by29.0%and79.4%. However, plasma GPT activity decreased from38.40U/L (302mg/kg) to27.75U/L (1414mg/kg) and declined by27.8%. Liver yield, total lipid, total TG reduced gradually with increasing choline level and phospholipids content had an opposite tendency (P<0.05). At1182mg/kg group, total liver lipid and triglyceride content decreased by22.6%and19.0%, and phospholipids content improved by35.1%compared to302mg/kg group. Dietary choline supplementation significantly increased liver LPL, HL, and TL activity (P<0.05). Histology observation showed that swelling of hepatocyte was observed and the cytoplasm was full of fat at302mg/kg group compared to the normal liver cell at the1414mg/kg group. According to broken-line regression analysis, the choline requirements of Pekin ducks from1to21d of age for total lipid, total TG, and total phospholipids were833,1311, and1230mg/kg. Compared to the requirements in expt.1, Pekin ducks needed more choline to prevent excess liver lipid deposition than to maintain growth. So we suggested the appropriate choline requirements of Pekin ducks from1to21d of age were1300mg/kg.In Expt.4, RNA-seq was used to study molecular mechanisms and signal pathways of fat metabolism regulated by choline. Three ducks were selected from choline-deficient (CD) group (302mg/kg) and choline-sufficient (CS) group (1414mg/kg), respectively. Their livers were collected for RNA isolation, transcriptome sequencing, and analysis. The results show that six different liver transcriptome were sequenced using the Illumina HiSeqTM2500sequencing platform.5.4-6.7million reads were obtained for single liver sample, and more than36.5million reads were produced in the six libraries, of which89.72%of the clean sequence were high-quality sequence and can be used for subsequent analysis. Coverage evaluation and RNA-seq data validation showed that more than90%of the clean reads were matched to the reference genome exon, and qRT-PCR validation results were consistent with RNA-seq data.363genes (233up-regulated,130down-regulated) were characterized as the most differentially expressed genes between CD and CS group, and a lot of them were involved in fat metabolism, energy balance, choline metabolism, phospholipids metabolism, immune stress and redox. Additionally, GO functional enrichment analysis indicated that candidate genes acted as cellular Component in cell membrane integrity, cytoplasm, nucleus, and transcription factors, and they were involved in many biological processes, such as transcriptional regulation, signal transduction pathways, redox, electron transfer, DNA connection, ATP binding and so on. PPAR signaling pathways, fat digestion and absorption pathways, and glycerolipid metabolism were acutely affected by dietary choline level. Long-term choline deficiency reduced PPAR signaling genes (PPARa, RXR, SCD, CYP7A1, DBI) mRNA expression as well as increased FABP, ACBP, and HMGCS1mRNA expression, which indicated choline deficiency decreased liver fat decomposition and β-oxidation and led to fatty liver disease.In conclusion, choline played a role in promoting growth and development as well as the lipotropic activity in Pekin ducks. Choline deficiency led to a significant increase in liver fat content and its underlying mechanisms was that the expression of genes related to lipodieresis and lipid oxidation were down-regulated, which led to fatty liver. Pekin ducks needed more choline to prevent excess liver lipid deposition than to maintain growth.