Digital expression analysis of the genes associated with salinity resistance after overexpression of a stress-responsive small GTP-binding RabG protein in peanut.

The Rab protein family is the largest family of the small GTP-binding proteins. Among them, the RabG genes are known to be responsive to abiotic stresses, but the molecular mechanisms of the stress responses mediated by RabG genes in plants is poorly understood. To investigate the molecular mechanism of AhRabG gene in peanut, transgenic plants overexpressing the AhRabG gene (S6) with relatively higher salinity resistance than the non-transgenic plants (S7) were obtained. Digital gene expression (DGE) sequencing was performed with the leaves of S6 and S7 plants before and after salinity-stress treatment. The AhRabG gene in peanut was found to be involved in a few pathways such as "photosynthesis", "oxidative phosphorylation", "AMPK signaling pathway", "plant hormone signal transduction", etc. A total of 298 differentially expressed genes (DEGs) were found to be upregulated or downregulated at five sampling time points based on the comparison between S6 and S7 plants. Among them, 132 DEGs were responsive to salinity stress in S6 and/or S7 after salinity-stress treatment. These 132 DEGs included genes encoding various transcription factors and proteins involved in resistance to salinity stress such as MYB, AP2, RING-H2 zinc finger proteins, late embryogenesis abundant (LEA) proteins, dehydration-responsive protein RD22, peroxidases, CBL-interacting protein kinases, calcium-binding proteins, and others. The information from this study will be useful for further studies on elucidating the mechanism of salinity resistance conferred by RabG genein peanut.

Differential gene expression between the porcine morula and blastocyst.

The survival and development of pre-implantation embryos are determinant factors affecting the outcome of animal reproduction. It is essential to transfer the expression of the genetic material from maternal sources, that is the ovum to the zygote before implantation to ensure successful development. Differentiation and transformation of blastomeres initiated during the morula and blastocyst stages is an important step of the embryonic development prior to implantation. We collected morula and early blastocyst samples from pure-bred Landrace pigs in vivo to study the differential gene expression patterns at these two stages. Total RNA was extracted from individual embryos and two rounds of amplification were employed. Two micrograms of antisense RNA, targets, were prepared and hybridized with each of four custom made oligo microarrays representing 24,000 porcine genes. The analyses of replicate hybridizations showed that among the 24,000 genes, 162 genes were expressed fivefold or greater in the morula compared to early blastocysts and 2126 genes were expressed fivefold or greater in early blastocysts compared to the morula. Of these differentially expressed genes, 1429 genes were functionally annotated with related human Gene Ontology terms. In addition to basic metabolic processes, genes related to signal transduction, transportation and cell differentiation were found in both stages and were up-regulated as embryo development proceeded. Real time polymerase chain reaction was utilized to quantify 12 genes differentially expressed in the 2 embryonic stages and validated the reliability of major evidences shown in microarrays. In conclusion, we have obtained a preliminary landscape of genes differentially expressed during the transition from morula to early blastocysts in pigs and showed a generally increased transcriptional activity, perhaps in preparation for implantation. Our results provide an opportunity to study the functions of these genes in relation to the development and survival of pre-implantation porcine embryos.

Abundantly expressed hepatic genes and their differential expression in liver of prelaying and laying geese.

Geese have a short egg-laying period and a low egg production rate. To induce and maintain egg laying, genes related to generating hepatic lipid for yolk deposition should be adequately expressed. Liver mRNA from 6 laying geese was extracted and used for construction of a full-length enriched cDNA library. About 2,400 clones containing gene sequences were determined and National Center for Biotechnology Information Gallus gallus Gene Index databases were used to compare and analyze these sequences. Ten highly expressed genes were selected to determine the differential expression between laying and prelay goose liver. Tissue distribution data showed that very low density apolipoprotein II, liver type fatty acid binding protein, vitellogenin I, and vitellogenin II transcripts were specifically expressed in the liver of laying geese. Ovoinhibitor, preproalbumin, alpha-2-hs-glycoprotein, and vitamin D binding protein mRNA were highly expressed in the liver and to a lesser extent in other tissues. Ovotransferrin mRNA was expressed in liver, ovary, oviduct, shell gland, brain, and adipose tissues. The concentration of transthyretin mRNA was high in the liver and brain. The mRNA concentrations of liver type fatty acid binding protein, alpha-2-hs-glycoprotein, and transthyretin in the livers of laying and prelay geese were not different. The concentrations of hepatic ovotransferrin, ovoinhibitor, preproalbumin, very low density apolipoprotein II, vitellogenin I, vitellogenin II, and vitamin D binding protein mRNA were higher in the liver of laying geese than in prelay geese, suggesting that these genes may be involved in laying function or lipid metabolism related to egg formation.

The differential expression of hepatic genes between prelaying and laying geese.

Suppression subtractive hybridization was used to detect differential expression of genes in the livers of laying and prelaying geese. Liver tissues from prelaying and laying geese were dissected for mRNA extraction. The cDNA, reverse transcribed from liver mRNA of prelaying geese, was subtracted from the cDNA generated from the laying geese (forward subtraction). Five hundred seventy-six clones with possible differentially expressed gene fragments were observed by forward subtraction hybridization. After differential screening using the reverse and forward subtraction cDNA, 164 clones were subjected to gene sequence determination and further analysis. Using Northern analysis, 5 known and 8 unknown genes were shown to be highly expressed in the livers of laying geese compared with prelaying geese. Vitellogenin I, apoVLDL-II, ethanolamine kinase, G-protein gamma-5 subunit, and leucyl-tRNA synthase were highly expressed in the livers of laying geese compared with that from the prelaying geese (P<0.05). The expression of these known genes suggests that their function in the liver of laying geese is primarily involved in lipid and lipoprotein metabolism. Several of these differentially expressed genes were found to be responsive to estrogen stimulation, confirming the involvement of these genes in the egg-laying function of the goose.

The expression of pituitary gland genes in laying geese.

The purpose of this study was to detect differential expression of genes in the pituitary gland in laying geese by suppression subtractive hybridization (SSH). Pituitary glands from prelaying and laying geese were dissected for mRNA extraction. The cDNA from pituitary glands of prelaying geese was subtracted from the cDNA from the pituitary glands of laying geese (forward subtraction); the reverse subtraction was also performed. We screened 384 clones with possible differentially expressed gene fragments by differential screening. Sixty-five clones from the differential screening results were subjected to gene sequencing and further analysis. We found that at least 19 genes were highly expressed in the pituitary glands of laying geese compared with prelaying geese. Among these, 6 genes (including 4 novel genes) were confirmed by virtual Northern analysis. We found that prolactin and visinin-like protein were highly expressed in the pituitary glands of laying geese compared with prelaying geese (P < 0.05). Further investigation is needed to demonstrate specific functions of the novel genes discovered in the current study.

Cloning and expression of the genes associated with lipid metabolism in Tsaiya ducks.

Sterol regulatory element binding protein 1 (SREBP1) drives the expression of several lipogenic genes, whereas SREBP2 dictates the expression of every gene involved in cholesterolgenesis in mammals. In the current study, we cloned the cDNA fragments for SREBP1, SREBP2, fatty acid synthase (FAS), 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CoA reductase), and very low density apolipoprotein-II (apoVLDL-II), the genes associated with lipid metabolism. Fifteen ducks immediately before the first egg was laid (18 wk old) and 15 ducks from the same population at an egg production rate of 80% were killed. Total RNA was extracted from liver and used to amplify the targeted genes by reverse transcription-PCR and screening of a cDNA library. The sequence data showed that Tsaiya duck SREBP1, SREBP2, FAS, and HMG-CoA reductase were highly homologous to that of chicken. Tsaiya duck SREBP1 mRNA was expressed in adipose tissue, cardiac muscle, skeletal muscle, liver, and ovary. The SREBP2 mRNA concentration was highest in liver and ovary. Concentrations of FAS and HMG-CoA reductase mRNA were high in liver and lower in other tissues. The apoVLDL-II mRNA was specifically expressed in the liver. The differences between mRNA concentrations of SREBP1, SREBP2, and FAS in the livers of laying and prelay ducks were not significant. However, the concentrations of hepatic HMG-CoA reductase and apoVLDL-II mRNA were higher in the laying ducks than in prelay ducks. Therefore, laying may affect particular aspects of lipid metabolism, especially biochemical pathways that involved apoVLDL-II and HMG-CoA reductase.

Effects of atosiban on stress-related neuroendocrine factors.

Atosiban, an oxytocin/vasopressin receptor antagonist, is used to decrease preterm uterine activity. The risk of preterm delivery is undoubtedly associated with stress, but potential side effects of atosiban on neuroendocrine functions and stress-related pathways are mostly unknown. These studies were designed to test the hypothesis that the chronic treatment of rats with atosiban modulates neuroendocrine functions under stress conditions. Male rats were treated (osmotic minipumps) with atosiban (600 µg/kg per day) or vehicle and were restrained for 120 min/day for 14 days. All animals were treated with a marker of cell proliferation 5-bromo-2-deoxyuridine. Anxiety-like behavior was measured using an elevated plus-maze. Treatment with atosiban failed to modify plasma concentrations of the stress hormones ACTH and corticosterone, but led to a rise in circulating copeptin. Atosiban increased prolactin levels in the non-stressed group. Oxytocin receptor mRNA levels were increased in rats exposed to stress. Treatment with atosiban, in both control and stressed animals, resulted in a decrease in oxytocin receptor gene expression in the hypothalamus. No changes were observed in vasopressin receptor 1A and 1B gene expression. The decrease in hippocampal cell proliferation induced by stress exposure was not modified by atosiban treatment. This study provides the first data, to our knowledge, revealing the effect of atosiban on gene expression of oxytocin receptors in the brain. Atosiban-induced enhancement of plasma copeptin indicates an elevation in vasopressinergic tone with potential influence on water-electrolyte balance.

Serum responsiveness of the rat PCNA promoter involves the proximal ATF and AP-1 sites.

We have previously shown that the rat PCNA (proliferating cell nuclear antigen) gene promoter is responsive to serum stimulation. In this study, the sequence of the promoter responsive to serum stimulation has been localized in the region between nucleotides -70 and +125 relative to the transcription initiation site. This region contains an ATF site (nucleotides -51 to -44) and an AP-1 site (nucleotides -64 to -58). Mutation at either the ATF or the AP-1 site reduced the serum responsiveness of the promoter. In gel mobility shift assays, nuclear extracts from serum stimulated cells, compared to those from quiescent cells, exhibit an increasing binding activity toward a promoter related oligonucleotide (-70 to -42) which includes the ATF site and the AP-1 site. Formation of the DNA:protein complexes requires the simultaneous involvement of ATF and AP-1 sites as either element can abrogate the complexes in the competition experiment. Both the distance and sequence are essential to complex formation. Moreover, ATF-1 but not ATF-2 (or CREB) has been identified as a major component of the complexes in the antibody supershift or interference experiment. The results of this study suggest that ATF-1 in association with other factors is involved in regulating the serum stimulation of the rat PCNA promoter activity via the proximal ATF and AP-1 sites.

Expression of porcine adipocyte transcripts: tissue distribution and differentiation in vitro and in vivo.

Transcription factor transcripts implicated in adipocyte differentiation (peroxisome proliferator-activated receptor gamma (PPAR gamma), retinoid x receptor alpha (RXR alpha), adipocyte determination and differentiation-dependent factor 1 (ADD1), and CCAAT/enhancer binding protein alpha (C/EBP alpha)) and adipocyte-characteristic protein transcripts (lipoprotein lipase (LPL) and adipocyte fatty acid binding protein (aP2)) were measured in pig tissues. Transcripts for PPAR gamma, ADD1, and aP2 were localized in porcine subcutaneous and perirenal adipose tissues; transcripts for C/EBP alpha and LPL were detected in other tissues, but the greatest concentrations were in the adipose tissues. In porcine stromal-vascular cells (S/V cells) differentiating in vitro, transcripts for PPAR gamma and aP2 increased gradually, transcripts for ADD1, and LPL increased early and transcripts for C/EBP alpha increased late. In pigs, adipose tissue transcripts for PPAR gamma, ADD1, and LPL were minimal at birth and increased to 28 days postpartum, transcripts for C/EBP alpha were low until 28 days and transcripts for aP2 were at high levels, regardless of age. Although transcript development was somewhat different in vitro and in vivo, the data suggest PPAR gamma (and ADD1 are involved in regulation of transcripts for LPL and that there may be more partially differentiated precursor cells in S/V cells at day 0 than in adipose tissue at birth.

Expression of porcine adipocyte transcripts during differentiation in vitro and in vivo.

Transcript concentrations for the transcription factors, CCAAT enhancer binding protein beta and alpha (C/EBPbeta and C/EBPalpha), plus the adipocyte-characteristic proteins, fatty acid synthase (FAS), glucose transporter 4 (Glut 4), hormone-sensitive lipase (HSL), insulin receptor (InsR), lipoprotein lipase (LPL), and leptin were measured during differentiation of porcine stromal-vascular (S/V) cells in vitro. These same transcripts, excluding FAS and InsR, were measured in porcine adipose tissue from birth to 7 weeks of age. In S/V cells, C/EBPbeta and InsR were continuously elevated. At day 0, C/EBPalpha was approximately 20% of the day 9 value. The LPL increased gradually from day 0 to 9, whereas most other transcripts had a lag period of several days. In tissue, C/EBPbeta was substantial at birth and increased gradually. The C/EBPalpha was relatively low at birth and increased at day 17. The LPL and leptin increased continuously. The Glut 4 was low at birth and increased at day 28. The HSL was relatively low at birth, increased at day 10, and plateaued at day 28. Transcripts in porcine S/V cells develop somewhat differently from adipocyte differentiation models established in clonal cells, but the porcine cells represent a model that should be more applicable to pigs.

Dietary fat has minimal effects on fatty acid metabolism transcript concentrations in pigs.

Young, crossbred pigs were fed either a low-fat, corn-based diet; a high-fat, tallow-based diet with a considerable saturated fatty acid (FA) content; or a high-fat, fish oil-based diet with a considerable polyunsaturated FA content, for 14 d. There were six pigs per dietary group (approximately 4-wk-old with a body weight of 6.16 kg). The plasma and adipose tissue FA composition reflected the composition of the diet to a large extent, but also reflected de novo FA synthesis coupled with chain elongation and desaturation. The liver and skeletal muscle FA composition reflected the diet and endogenous synthesis, but the indications for preferential incorporation or exclusion of specific FA were greater in these tissues than in plasma or adipose tissue. An important transcription factor for adipocyte differentiation and other aspects of lipid metabolism is adipocyte determination and differentiation-dependent factor 1 (ADD1). Liver ADD1 messenger RNA (mRNA) tended to be decreased (P = 0.06) in the fish oil-fed group, as well as in the combined high-fat-fed groups (tallow + fish oil) compared to the low-fat-fed group (P = 0.06). The muscle acyl-CoA oxidase mRNA tended to be increased in the tallow-fed group and decreased in fish oil-fed groups (P = 0.06). The muscle carnitine palmitoyltransferase mRNA tended to be elevated in both fat-fed groups (P = 0.07). None of the adipose tissue mRNA were changed by the diet (P > 0.20). The observations suggest there are major differences between rodents and pigs in modulation of transcripts associated with lipid metabolism by the dietary FA composition or concentration. Also, in porcine adipose tissue, as well as in liver and skeletal muscle, these transcripts are rather refractory to modification by dietary FA.

Expression of porcine transcription factors and genes related to fatty acid metabolism in different tissues and genetic populations.

Several transcription factors are involved in regulating lipid metabolism in various tissues of animals. Adipocyte determination and differentiation-dependent factor 1 (ADD1), peroxisome proliferator activated receptor alpha (PPAR alpha), and peroxisome proliferator activated receptor gamma (PPAR gamma) regulate both lipogenesis and fatty acid oxidation. We determined the tissue distribution and genetic difference in mRNA concentrations of these transcription factors in two genetic populations of pigs (Newsham XL-sired Newsham Landrace x Large White Duroc and Duroc-sired US Yorkshire x Duroc-Landrace). We also determined the tissue distribution and genetic difference in the mRNA concentration of fatty acid synthase (FAS) and acyl-CoA oxidase (ACO). Our data showed that ADD1 was highly expressed in adipose tissue and liver and that mRNA concentrations of ADD1 were similar between the two genotypes. The PPAR alpha mRNA concentration was high in adipose tissue and was similar between the two genotypes. In both populations, PPAR gamma mRNA was detected only in adipose tissue. There was no difference between the two genotypes in PPAR gamma mRNA concentration. The ACO mRNA was expressed in adipose tissue, skeletal muscle, and liver with no difference between genotypes. The FAS mRNA concentration in adipose tissue was seven times higher than that in the liver. There was no detectable FAS mRNA in skeletal muscle. These data support the concept that pig adipose tissue has considerable capability for fatty acid oxidation and synthesis. The uniqueness of expression patterns for FAS and ADD1 mRNA further indicates that adipose tissue is significantly involved in fatty acid and triacylglycerol synthesis in pigs.

The effect of dietary docosahexaenoic acid on the expression of porcine lipid metabolism-related genes.

To study the effect of dietary docosahexaenoic acid (DHA) on the expression of adipocyte determination and differentiation-dependent factor 1 (ADD1) mRNA in pig tissues, weaned, crossbred pigs (30 d of age) were fed either 2% (as-fed basis) tallow or DHA oil for 18 d. Body weight of the pigs was not affected by different dietary fatty acid (FA) compositions. The plasma and liver FA composition reflected the composition of the diet. The adipose tissue and skeletal muscle FA composition only partially reflected the diet, indicating either a slower FA turnover or that a greater proportion of the FA in these tissues is from endogenous FA synthesis. The ADD1 is an important transcription factor that modulates transcription of FA synthase to regulate the endogenous FA synthesis in the liver and adipose tissue. The ADD1 mRNA was decreased (P < 0.05) in the liver of DHA-treated pigs compared with that of the tallow-treated pigs. The diets did not have an effect on the ADD1 mRNA in pig adipose tissue. The ADD1 transcript was not detected in pig skeletal muscle. These results indicate that significant enrichment of liver DHA content inhibits the expression of ADD1 mRNA. Such an effect is similar to that reported in porcine differentiating adipocytes cultured with DHA. The liver and muscle acyl CoA oxidase mRNA concentration was increased (P < 0.05) by DHA oil treatment, suggesting that DHA treatment may increase peroxisomal fatty acid oxidation in these two tissues. Our present observations demonstrate that dietary DHA enrichment not only affects tissue DHA concentration but also mildly modifies the expression of genes related to fatty acid metabolism in the porcine liver and skeletal muscle.

Modulation of porcine adipocyte beta-adrenergic receptors by a beta-adrenergic agonist.

Mammalian cells have several mechanisms to decrease the response to beta-adrenergic agonists. Agonists are metabolized or taken up by nerve endings. The beta-adrenergic receptors (betaAR) are inactivated by phosphorylation and removed from the cell membrane, and synthesis is decreased or degradation is increased. Knowledge about adipocyte betaAR desensitization is mostly from rodent adipocytes with > or = 90% beta3AR. Porcine adipocyte betaAR have functional and ligand-binding properties that are quite different from those in many other species. Furthermore, the predominant betaAR subtype in the porcine adipocyte is the beta1AR (70 to 80%). Given these species differences, it might be expected that desensitization in porcine adipocytes would not be totally concordant with the rodent-derived model. Isolated porcine adipocytes were incubated without or with a betaAR agonist, isoproterenol. The total betaAR number, measured by ligand-binding in a crude membrane fraction, tended to be lower after 6 h of incubation without isoproterenol. The addition of 10(-5) M isoproterenol during the incubation caused the betaAR number to decrease 43% compared to cells incubated without isoproterenol. The beta1AR and beta2AR transcript concentrations both decreased 45% after 6 h of incubation without isoproterenol. There was no decrease in mRNA when cells were incubated with isoproterenol. The results suggest the betaAR were desensitized by incubation with isoproterenol, perhaps by phosphorylation and removal from the membrane, but this was not accompanied by modulation of the concentration of transcripts for beta1AR or beta2AR.

Modulation of porcine adipocyte beta-adrenergic receptors by hormones and butyrate.

The beta-adrenergic receptors (betaAR) on the surface of mammalian cells are desensitized when the cell is stimulated by betaAR agonists to eliminate excessive response by the cell. Investigations with adipocytes, primarily rodent-derived cells, indicate other hormones and substrates also can modulate the individual betaAR subtypes. For example, glucocorticoids decrease the beta1AR and the beta3AR but increase the beta2AR. Insulin decreases the beta3AR. Thyroid hormones increase the beta3AR and butyrate increases beta1AR and beta2AR but dramatically decreases beta3AR. Because porcine adipocytes have unique functional and ligand-binding properties compared to rodent adipocytes and because porcine adipocytes contain predominantly beta1AR, compared to rodent adipocytes with predominantly beta3AR, we expect the regulation of porcine adipocyte betaAR by hormones and substrates to be different from that in rodent adipocytes. Isolated porcine adipocytes were incubated for 6 and 21 h without and with dexamethasone, insulin, triiodothyronine, or butyrate. Cells incubated without hormone or butyrate had a decreased betaAR number at 21 h. The beta1AR and beta2AR transcript concentrations were decreased after 6 h and tended to rebound after 21 h of incubation. Dexamethasone did not change the total betaAR number but tended to increase the beta1AR and beta2AR transcript concentrations. Insulin increased the betaAR number and decreased both transcript concentrations at 21 h. Triiodothyronine and butyrate did not change the receptor number or transcript concentrations. The results indicate that betaAR transcript concentrations do not accurately predict the betaAR protein concentration (estimated by ligand binding). Results also indicate that betaAR in porcine adipocytes are relatively unresponsive to hormones and butyrate compared to rodent-derived adipocytes described in the literature.

Effect of feed restriction on adipose tissue transcript concentrations in genetically lean and obese pigs.

To determine possible genetic influences on the steady-state concentrations of several key transcription factor transcripts and the transcript concentrations for adipocyte-characteristic proteins, young, genetically obese and lean pigs were given ad libitum access or feed or were restrictively fed at 50% of ad libitum intake for 5 wk. Obese pigs were smaller and fatter than lean pigs, whether intake was ad libitum or restrictive. Plasma protein, albumin, and cholesterol concentrations were greater in obese than in lean pigs. Plasma NEFA, blood urea nitrogen, triacylglycerols, and postprandial glucose and insulin concentrations were less (P < .02) in pigs fed restrictively than in pigs with ad libitum access to feed, regardless of genetic group. The adipose tissue glucose transporter 4, fatty acid synthase, and leptin transcript concentrations were greater (P < .05) in obese than in lean pigs. The CCAAT/enhancer binding proteins beta and alpha, adipocyte fatty acid binding protein, hormone-sensitive lipase, and the beta1-adrenergic receptor transcript concentrations tended (P < . 10) to be greater in adipose tissue from obese than in that from lean pigs. Several other transcripts were numerically greater in obese than in lean pigs. The data collectively suggest that messenger RNA concentration for several adipose tissue proteins is a contributing factor to the excess fat deposition in these obese pigs. Restricted feeding did not change the concentration of any transcript except that for adipocyte fatty acid binding protein, which was reduced. The accretion of fat was markedly reduced in the restrictively fed pigs, but this diminution does not seem to be regulated by modulation of messenger RNA concentration.

Changes in fatty acid profiles in different lipid classes during late development of turkey embryos from two genetic lines.

Fatty acid (FA) profiles in embryonic yolk sacs (YS) and livers were studied in embryos from a randombred turkey line (RBC2) and a line selected for body weight at 16 wk (F line). There were no differences in FA profiles of fresh yolk lipids. During the course of incubation, oleic acid (C18:1) was higher and linoleic acid (C18:2) was lower in YS triglyceride (TG) and phospholipid (PL) subclasses in F line compared with RBC2 embryos. In both lines, the C18:1 content of YS cholesteryl esters (CE) increased from 58 to 63% during the last 6 d of incubation. From 22 to 28 d of incubation, there was a constant C18:1 concentration in hepatic CE, which was > 60% of total hepatic CE FA. As incubation proceeded, palmitic acid (C16:0) and C18:1 in hepatic TG decreased from 27 to 16% and 37 to 34%, respectively. The stearic acid (C18:0) in TG increased from 12% at Day 22 to 32% of total FA at hatch (Day 28) in RBC2 embryos compared with a lesser increase in the F line (11.8 to 18.6%). In hepatic PL, arachidonic acid (C20:4) decreased, whereas both C16:0 and C18:0 increased from 22 to 28 d of incubation. During this same time period, there was an overall decline in docosahexaenoic acid (C22:6) only in the RBC2. On Days 26 and 28, F line embryos had greater concentrations of C22:6 and C20:4 in hepatic PL than did RBC2. These results suggest that selection for increased BW has changed the proportional incorporation of different FA into embryonic lipids.

Characterization of changes in yolk sac and liver lipids during embryonic and early posthatch development of turkey poults.

In two studies, changes in lipids and fatty acids from the yolk sac and liver of turkeys were determined during incubation and early postnatal growth. In Experiment 1, embryos were collected from Day 13 through 6 d posthatch. Embryos weighed 4.25 g at Day 13, 52.8 g at hatch (Day 28), and 104 g at 6 d. Total yolk lipid (grams per yolk sac) decreased from 9.48 g at Day 13 to 2.52 g at hatch (Day 28). Almost half (47%) of the yolk lipid decreased during the last week of incubation. In Experiment 2, embryos were collected daily from Day 22 through 2 d posthatch. Embryonic weight increased from 32.5 g at Day 22 to 55.8 g at hatch (Day 28), 3 g more than in Experiment 1. On Days 22 and 25, total yolk lipid (grams per yolk sac) was slightly less in Experiment 2 than in Experiment 1, but at hatch (Day 28) there were considerable differences (Experiment 1, 2.52 g; Experiment 2, 0.63 g), concomitant with increased hatch weight in Experiment 2. Liver DM percentage and ether extractable lipid increased as incubation proceeded. Oleic acid comprised the largest proportion of total yolk fatty acids and of liver fatty acids. There was a small but significant increase in yolk sac oleic acid (percentage of total fatty acids) between Day 22 (40%) and hatch (Day 28; 45.4%) and a much larger increase in liver oleic acid (46.6% to 56.5%). The absolute amount of all yolk sac fatty acids declined greatly during the second half of incubation through 6 d posthatch.

The developmental expression of acyl-coenzyme A:cholesterol acyltransferase in the yolk sac membrane, liver, and intestine of developing embryos and posthatch turkeys.

Acyl-coenzyme A:cholesterol acyltransferase (ACAT) catalyzes the formation of cholesterol esters (CE) from free cholesterol and fatty acyl-coenzyme A. This experiment was conducted to study the ontogeny of ACAT activity in the yolk sac membrane, liver, and intestine during embryonic development and early posthatch growth of turkeys. The ACAT activity was measured on tissue samples collected at 3-d intervals from embryonic Day (ED 13) 13 through 6 d posthatch (PD 6). The ACAT activity (pmol/mg microsomal protein per min) in the yolk sac membrane increased form 840 pmol at ED 13 to 2,497 pmol at ED 22, and subsequently declined to a very low level by PD 3. The high level of enzyme activity at ED 22 is concomitant with the large quantity of CE formed within the yolk sac membrane at this developmental age. Liver ACAT activity increased from 60 pmol at ED 13 to 242 to 243 pmol at ED 25 and PD 3, followed by a decline to 130 pmol by PD 6, mirroring the peak in hepatic CE concentration. This suggests that even during incubation, the liver plays a significant role in lipid metabolism. Intestinal ACAT specific activity increased from 14 pmol (ED 16) to 44 pmol (ED 25), and then declined to 23 pmol by hatch (ED 28), with no further decline through PD 6. Total intestinal ACAT activity (pmol per intestine/min) increased, however, from ED 16 through PD 6. This increase in activity suggests that the total capacity for cholesterol esterification increases during the course of incubation and shortly after hatching.

The ontogeny of fatty acid-binding protein in turkey (Meleagridis gallopavo) intestine and yolk sac membrane during embryonic and early posthatch development.

Experiments were conducted to confirm the existence and ontogeny of fatty acid binding protein (FABP) in the intestine and yolk sac membrane of turkey poults (Meleagridis gallopavo) during embryonic and early posthatch development. Intestinal (I-) FABP was measured using an immunoblot procedure incorporating anti-chick liver (L-)FABP antisera. FABP activity in both tissues was also confirmed with a ligand-binding assay incorporating 14C-oleic acid. I-FABP did not cross-react with chick L-FABP antisera until hatch, embryonic day 28 (ED 28), after which there was a 39% increase in I-FABP concentration through the first 3 d posthatch (PD 3). FABP concentration calculated on a total intestinal basis (ng/intestine), however, increased 10-fold through PD 6. Specific activity [disintegrations per minute (dpm)/ mg cytosolic protein] was greatest at hatch and decreased slightly thereafter, whereas specific activity of FABP in the yolk sac membrane peaked between ED 16 and ED 19 and then declined. Total yolk sac activity (dpm/yolk sac membrane), however, plateaued at ED 22 before declining to low levels by PD 3, coincident with the period of maximal lipid transfer out of the yolk.