Responsiveness to adipogenic agents in stromal-vascular cultures derived from lean and preobese pig fetuses: an ontogeny study.

Primary cultures of stromal-vascular (S-V) cells from adipose tissue were used to evaluate characteristics of preadipocytes from lean and preobese fetuses at several ages (50, 75, and 110 d). In insulin-supplemented (1 microM) cultures (serum free) there was a significant age x fetal genotype interaction (P less than .01) for glycerol-phosphate dehydrogenase specific activity (GPDH); GPDH activity was genotype-dependent at 110 d (preobese greater than lean). The responses of S-V cultures (preadipocyte development) to 2% pig serum and to insulin (serum free) were similar. Main effects of genotype and age were significant (P less than .05) for protein levels in pig serum and insulin-treated cultures. There was a significant genotype x age (P less than .05) interaction for GPDH activity and protein levels in cultures treated with dexamethasone + 3-isobutyl-1-methylxanthine (DEX-IBMX). Treatment with DEX-IBMX induced more preadipocyte development in cultures from preobese fetuses than in cultures from lean fetuses at 110 d (P less than .05). The responsiveness of S-V cultures to DEX-IBMX (enhanced development) increased considerably between 50 and 75 d regardless of fetal genotype, but there was little response in cultures form 50-d fetuses. Preadipocyte development in lean and preobese fetuses diverged between 75 and 110 d, resulting in many more preadipocytes in preobese fetuses at 110 d. Therefore, S-V cells from preobese fetuses (late term) may be inherently more sensitive to adipogenic agents than S-V cells from lean fetuses.     

Fetal hypophysectomy causes a decrease in preadipocyte growth and insulin like growth factor-1 in pigs

Fetal pigs in one uterine horn of each of five gilts were hypophysectomized (HX) in utero by electrical cauterization at 72-74 days of gestation and sera collected at 110 days of gestation. Sera from HX fetuses had lower levels of insulin-like growth factor-1 compared to control littermates (P less than .05). Sera were tested for their effects on primary cultures of stromal-vascular cells from adipose tissue. The soluble protein concentration/dish was lower when pig cells were cultured in sera from HX fetuses compared to sera from control fetuses (P less than .01). Sera from HX fetuses inadequately supported growth of stromal-vascular cells so subsequent experiments utilized pooled sera from normal and HX adult pigs. Sera from HX and control fetuses were mixed with sera from the two adult pools and tested for incorporation of tritiated thymidine into rat preadipocytes and the appearance of adipocytes (determined histochemically) in pig stromal-vascular cultures. In cultures fed sera from HX fetuses there was a lower (P less than .05) number of pig fat cells/culture and a lower level (P less than .06) of preadipocyte proliferation in rat cell cultures when compared to control fetal sera. Fetal pig serum contains factors (adipogenic) which promote the proliferation and differentiation of adipocytes in culture. Serum from HX fetuses has a lower level of adipogenic factors.     

Patterns of gene expression in pig adipose tissue: insulin-like growth factor system proteins, neuropeptide Y (NPY), NPY receptors, neurotrophic factors and other secreted factors

Although cDNA microarray studies have examined gene expression in human and rodent adipose tissue, only one microarray study of adipose tissue from growing pigs has been reported. Total RNA was collected at slaughter from outer subcutaneous adipose tissue (OSQ) and middle subcutaneous adipose tissue (MSQ) from gilts at 90, 150, and 210 d (n=5 age(-1)). Dye labeled cDNA probes were hybridized to custom porcine microarrays (70-mer oligonucleotides). Gene expression of insulin-like growth factor binding proteins (IGFBPs), hormones, growth factors, neuropeptide Y (NPY) receptors (NPYRs) and other receptors in OSQ and MSQ changed little with age in growing pigs. Distinct patterns of relative gene expression were evident within NPYR and IGFBP family members in adipose tissue from growing pigs. Relative gene expression levels of NPY2R, NPY4R and angiopoietin 2 (ANG-2) distinguished OSQ and MSQ depots in growing pigs. We demonstrated, for the first time, the expression of IGFBP-7, IGFBP-5, NPY1R, NPY2R, NPY, connective tissue growth factor (CTGF), brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) genes in pig adipose tissue with microarray and RT-PCR assays. Furthermore, adipose tissue CTGF gene expression was upregulated while NPY and NPY2R gene expression were significantly down regulated by age. These studies demonstrate that expression of neuropeptides and neurotrophic factors in pig adipose tissue may be involved in regulation of leptin secretion. Many other regulatory factors were not influenced by age in growing pigs but may be influenced by location or depot.     

A comparison of thiazolidinedione-induced adipogenesis and myogenesis in stromal-vascular cells from subcutaneous adipose tissue or semitendinosus muscle of postnatal pigs

This study compared the adipogenic potential of porcine stromal-vascular (S-V) cells from semitendinosus muscles and s.c. adipose tissue using thiazolidinediones. Stromal-vascular cells were obtained from s.c. adipose tissue and both semitendinosus muscles from 5- to 7-d-old pigs after collagenase digestion. Preadipocyte recruitment was measured using immunohistological evaluation for AD-3, a preadipocyte antibody. Ciglitazone increased the number of preadipocytes in adipose tissue but not semitendinosus muscle S-V cell cultures, whereas 10 microM troglitazone increased preadipocyte abundance in both adipose and muscle S-V cultures by approximately 3-fold (P < 0.05). Increasing troglitazone doses did not further increase preadipocyte number. Increases in preadipocytes were paralleled by increases in CCAAT/enhancer-binding protein alpha (C/EBPalpha) and peroxisome proliferator-activated receptor gamma (PPARgamma) positive cells in adipose tissue S-V cultures, whereas PPARgamma-reactive but not C/EBPalpha-reactive cells were increased in muscle S-V cultures treated with 10 microM troglitazone. Additionally, troglitazone treatment did not increase lipid content in s.c. adipose tissue or muscle S-V cell cultures. Cells plated on laminin-precoated culture dishes were used to determine whether troglitazone influenced adipogenesis or myogenesis in cocultures from muscle S-V cells. There was no effect on the number of myotubes or the average number of nuclei per myotube, suggesting myogenesis was not impaired by troglitazone treatment. These results suggest that regulation of intramuscular adipogenesis differs from that of subcutaneous adipogenesis.     

Lipogenesis and pancreatic insulin release in fetal pigs

Body composition, liver and adipose lipogenesis, and pancreatic insulin release were examined in intact and decapitated fetal pigs on d 110 of gestation. Decapitation was on d 45 of gestation. Decapitated fetuses deposited more body lipid and less body ash compared with intact fetuses. Body weight, water, dry matter and protein remained similar in intact and decapitated fetuses. Hepatic fatty acid esterification and synthesis were two- and threefold greater, respectively, in decapitated than in intact fetuses. Fatty acid synthesis in subcutaneous adipose tissue of decapitated fetuses was three times greater than values obtained in intact fetuses. The data supported the concept that substrate availability from the dam was not the rate-limiting step in fetal pig lipid synthesis and storage. High growth hormone levels in normal fetal pigs may be responsible for inhibiting lipogenesis, while fetal decapitation would remove this inhibition and be associated with greater lipid deposition. However, pancreatic insulin release was greater in decapitated than in intact fetuses; an indication that elevated lipid deposition may also be due to greater fetal insulin secretion.     

Maternal obesity upregulates fatty acid and glucose transporters and increases expression of enzymes mediating fatty acid biosynthesis in fetal adipose tissue depots

Maternal nutrient restriction leads to alteration in fetal adipose tissue, and offspring from obese mothers have an increased risk of developing obesity. We hypothesized that maternal obesity increases fetal adipogenesis. Multiparous ewes (Columbia/Rambouillet cross 3 to 5 yr of age) carrying twins were assigned to a diet of 100% (Control; CON; n = 4) or 150% (Obese; OB, n = 7) of NRC maintenance requirements from 60 d before conception until necropsy on d 135 of gestation. Maternal and fetal plasma were collected and stored at -80°C for glucose and hormone analyses. Fetal measurements were made at necropsy, and perirenal, pericardial, and subcutaneous adipose tissues were collected from 7 male twin fetuses per group and snap frozen at -80°C. Protein and mRNA expression of fatty acid translocase [cluster of differentiation (CD) 36], fatty acid transport proteins (FATP) 1 and 4, insulin-sensitive glucose transporter (GLUT-4), fatty acid synthase (FASN), and acetyl-coA carboxylase (ACC) was evaluated. Fetal weight was similar, but fetal carcass weight (FCW) was reduced (P < 0.05) in OB versus CON fetuses. Pericardial and perirenal adipose tissue weights were increased (P < 0.05) as a percentage of FCW in OB versus CON fetuses, as was subcutaneous fat thickness (P < 0.001). Average adipocyte diameter was greater (P < 0.01) in the perirenal fat and the pericardial fat (P = 0.06) in OB fetuses compared with CON fetuses. Maternal plasma showed no difference (P > 0.05) in glucose or other hormones, fetal plasma glucose was similar (P = 0.42), and cortisol, IGF-1, and thyroxine were reduced (P ≤ 0.05) in OB fetuses compared with CON fetuses. Protein and mRNA expression of CD 36, FATP 1 and 4, and GLUT-4 were increased (P ≤ 0.05) in all fetal adipose depots in OB versus CON fetuses. The mRNA expression of FASN and ACC was increased (P < 0.05) in OB vs. CON fetuses in all 3 fetal adipose tissue depots. Fatty acid concentrations were increased (P = 0.01) in the perirenal depot of OB versus CON fetuses, and specific fatty acid concentrations were altered (P < 0.05) in subcutaneous and pericardial adipose tissue because of maternal obesity. In conclusion, maternal obesity was associated with increased fetal adiposity, increased fatty acid and glucose transporters, and increased expression of enzymes mediating fatty acid biosynthesis in adipose depots. These alterations, if maintained into the postnatal period, could predispose the offspring to later obesity and metabolic disease.     

Influence of maternal obesity on fetal development in pigs

Fetuses from linebred lean (L) and linebred obese (O) and reciprocal crossmatings were examined at 110 d of gestation for line, maternal and heterotic effects. There was no significant heterotic effect for any trait measured. A significant maternal effect was observed for adipose tissue lipoprotein lipase (LPL) activity and for serum triglycerides. The enzyme activity and triglycerides concentration were higher in fetuses from O dams than in fetuses from L dams. In a lipid clearance test, no maternal effect was observed for changes in serum concentrations of triglycerides and free fatty acids or in optical density (associated with the disappearance of injected Liposyn). Linebred O fetuses exhibited higher LPL activity in both the biceps femoris muscle and sc adipose tissue compared with linebred L fetuses. The LPL activity of the adipose tissue was higher than that of the skeletal muscle. The percentage of dry matter, percentage of triglycerides and protein/DNA were higher in the muscle of linebred O fetuses than in that of linebred L fetuses. Based on tissue LPL activity and on muscle compositional traits, linebred O fetuses were more mature at 110 d of gestation than were the linebred L fetuses. Maternal obesity had little detectable influence on fetal development of the pig when measured at 110 d of gestation.     

The histology of developing porcine adipose tissue

At each of the following days after conception (45, 60, 75, 90 and 105), pig fetuses were removed from sows representing lean and fat stains. From two additional litters, postnatal pigs were sacrificed at 1, 3, 6, 9, 12, 15, 18 and 21 d. Pelikan dye was injected into fetuses and pigs. The whole of the dorsal subcutaneous tissue, including some underlying muscle, was removed. Tissue was fixed into paraffin blocks or was frozen. Paraffin and frozen sections were stained and examined for stromal-vascular and cellular changes during growth. Organized stromal-vascular changes occurred during a period of adipocyte formation from 45 d gestation until 9 d postnatally. At 45 d gestation, the subcutaneous tissue contained many short unorganized connective tissue fibers. Gradually, these fibers became more organized in a ventral to dorsal and caudal to cranial gradient, so that by 1 d postnatally, they formed complete lobules around all existing fat cell clusters. The presumptive adipose space of the complete lobules contained delicate strands of connective tissue and reacted metachromatically for mucin. Connective tissue around lobules became progressively thinner throughout the remaining postnatal ages. Vascularity of the subcutaneous tissue increased as the stromal became organized. Lipid was not present in the subcutaneous tissue at 45 d gestation, but some deposition was apparent in the inner layer at 60 d. Between 60 d gestation and 9 d postnatally, fat cells filled both subcutaneous layers in a ventral to dorsal formation. Presumptive adipose lobules were the source of adipocytes and capillaries of developing fat cell clusters. Adipocytes from fetuses through 1-d postnatal pigs were multilocular, while unilocular fat cells were first observed at 3 d. At 9 d, multilocular adipocytes were found singly or in groups within unilocular fat cell lobules.     

Differentiation of adipose tissue and muscle in hypophysectomized pig fetuses

In the present study, fetuses were hypophysectomized (hypox) in utero on d 72 to 74 of gestation with an electrical cauterizing needle. One to six successfully hypox fetuses were removed on d 110 of gestation from each of five gilts. Subcutaneous adipose tissue samples and semitendinosus muscles were obtained from the hypox fetuses and an equal number of control fetuses. Body weights of control fetuses (n = 15; mean +/- SE, 1,195 +/- 33 g) were similar to weights of hypox fetuses (n = 15; 1,179 +/- 67 g). Fat cell size in the middle subcutaneous layer of adipose tissue was increased in hypox fetuses (P less than .01) compared with control fetuses. The number of obvious fat cell clusters (outer layer) in lipid stained sections was reduced (P less than .01) by 50% in hypox fetuses. Histochemical reactions for glucose-6-phosphate dehydrogenase, esterase and lipoprotein lipase (LPL) activities in middle layer cell clusters were considerably enhanced in sections from hypox fetuses compared with sections from controls. Quantitative analysis of percent light transmittance (Zeiss photometer) through LPL-stained cell clusters indicated an increase (P less than .001) in LPL staining in sections from hypox fetuses when compared with sections from control fetuses. Transverse muscle sections (cryostat) from hypox fetuses failed to show normal patterns (as seen in control muscles) of reactions for acid ATPase, malate dehydrogenase (NAD-dependent), NADH-TR and alpha-glycerol phosphate dehydrogenase (without NAD). The number of muscle fibers that were stained for these enzymes was greatly reduced in hypox fetuses compared with control fetuses. The number of lipid positive fibers was also reduced in hypox fetuses compared with control fetuses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Cellular and enzyme-histochemical aspects of adipose tissue development in obese (Ossabaw) and lean (crossbred) pig fetuses: an ontogeny study

The cellular and enzyme-histochemical differentiation of subcutaneous adipose tissue was studied in lean and obese pig fetuses at several ages. Positive reactions for a variety of cytosolic and organellar enzyme markers indicate metabolic competence of fetal adipocytes despite their small size (12 to 15 microns). Reactions for several enzymes decreased with fetal age and may be associated with a qualitative change in activity of adipocyte organelles. Age-associated increases in two lipogenic enzymes were observed in obese adipocytes. Observations on developing cells around hair follicles in the younger fetuses indicated significant temporal lags between the appearance of detectable enzyme activities in adipocytes. Enzyme activities in order of appearance were: dehydrogenases (cytosolic and mitochondrial), lipoprotein lipase and esterase. Esterase activity and several other enzymes were never observed in lipid positive cells that were not spherical. A proportion of hair follicle associated adipocytes in 110-d-old lean fetuses were histochemically and morphologically similar to brown adipocytes in the young rat. There was no evidence for brown adipocyte like cells in obese fetuses. Finally, comparison of the enzyme-histochemical differentiation of lean and obese fetal adipocytes indicates that fetal adipocytes become sensitive to external stimuli between 70 and 90 d of gestation.     

Mitotic activity in fetal and early postnatal porcine adipose tissue

Tritiated thymidine autoradiography and histochemistry were used to study the development of subcutaneous adipose tissue of lean and obese fetuses and postnatal pigs. A pattern of tritiated thymidine uptake by pre-adipocytes and adipocyte lipid accumulation was demonstrated during the growth of the fetal pig. In the youngest fetuses there was a period of intense stromal cell mitotic activity before any adipocyte lipid accumulation. During subsequent fetal development, clusters of tightly arranged stromal cells were formed. Lipid accumulation occurred only in these cell clusters. During this time of cell cluster formation and lipid accumulation, mitotic activity was minimal. In obese fetuses, stromal cell mitotic activity overlapped temporarily with the cell cluster formation and lipid accumulation period. In early postnatal pigs, fat cell clusters increased in size until they "physically filled" the adipose tissue. In pigs 3 d and older, there was extensive mitotic activity of cells within the fat cell clusters. The synthesis of this second bed of pre-adipocytes and the altered developmental pattern in the obese fetuses is suggested to be due to the influence of a high fat diet. The significance of these findings in terms of plausible links between pre-adipocyte mitosis and lipid accumulation is discussed.     

Glucocorticoids and the differentiation of porcine preadipocytes

The function of glucocorticoids in the differentiation of porcine preadipocytes was examined. Stromal-vascular cell cultures (containing preadipocytes) derived from adipose tissue of the perirenal, ham and shoulder regions of neonatal pigs were incubated in the presence of hydrocortisone at 0 to 100 ng/ml medium. Perirenal cells did not respond to hydrocortisone with an increase in enzyme expression, nor did they demonstrate growth characteristics similar to those of cultures derived from the ham or shoulder. Cultures from the shoulder and ham regions demonstrated dose-responsive increases in enzymatic expression to hydrocortisone. Enzymatic responses by cultures derived from the ham region were lower than responses by cultures from the shoulder region as measured by changes in the activities of sn-glycerol-3-phosphate dehydrogenase and lipoprotein lipase. Addition of insulin to the medium did not produce a synergistic effect with glucocorticoid on differentiation as determined by these enzymatic parameters. However, [14C]glucose metabolism by the cells in culture was synergistically increased by insulin and glucocorticoid supplementation of the medium. The ability of hydrocortisone to induce differentiation of porcine preadipocytes in vitro suggests that the changes that occur in plasma glucocorticoid concentrations during late gestation may play an important role in the rapid development of s.c. adipose tissue in the fetal pig. Secondly, the differences in culture characteristics and hormone responses of cells derived from different locations of adipose tissue formation indicate that differences may exist in the regulation of the growth and development of preadipocytes from different anatomical locations.     

Ontogenic development of brown adipose tissue in Angus and Brahman fetal calves

Brahman calves experience greater neonatal mortality than Angus calves if cold-stressed. To establish a developmental basis for this, three fetuses of each breed type were taken at 96, 48, 24, 14, and 6 d before expected parturition, and at parturition. Overall fetal BW tended (P = 0.08) to be greater for Angus than for Brahman fetuses. There was no difference between breed types in total brown adipose tissue (BAT) mass or grams of BAT/kg BW. Brown adipocyte density decreased 56%, whereas lipogenesis from acetate and glucose in vitro decreased 97% during the last 96 d of gestation in both breed types. Glycerolipid synthesis from palmitate declined by 85% during the last trimester but still contributed 98% to total lipid synthesis at birth. The fetal age x breed interaction was significant for lipogenesis from glucose (P = 0.05) and palmitate (P = 0.005); rates were higher at 96 d before birth in Brahman BAT but declined to similar rates by birth. Uncoupling protein-1 (UCP1) mRNA tripled during gestation in both breed types (P = 0.002), whereas mitochondrial cross-sectional area did not change (P = 0.14) during gestation. Neither the breed nor the age x breed effect was significant (P > or = 0.24) for UCP1 mRNA concentration or mitochondrial cross-sectional area. In both breed types, a marked decrease in BAT UCP1 mRNA between 24 and 14 d prepartum was associated with a similar reduction in lipogenesis from palmitate and a noticeable change in BAT mitochondrial morphology, as the mitochondria became more elongated and the cristae became more elaborate. Uncoupling protein-1 mRNA initially was elevated in Angus tailhead s.c. adipose tissue, but was barely detectable by birth, and tended to be greater overall (P = 0.09) in Angus than in Brahman BAT. If uncoupling protein activity in s.c. adipose tissue persists after birth, then s.c. adipose tissue may contribute more to thermogenesis in Angus newborn calves than in Brahman calves. In contrast, we did not observe differences in ontogenic development of perirenal BAT that could explain the documented differences in thermogenic capacity between Angus and Brahman newborn calves.     

Brown adipose tissue development and metabolism in ruminants

We conducted several experiments to better understand the relationship between brown adipose tissue (BAT) metabolism and thermogenesis. In Exp. 1, we examined perirenal (brown) and sternum s.c. adipose tissue in 14 Wagyu x Angus neonates infused with norepinephrine (NE). Perirenal adipocytes contained numerous large mitochondria with well-differentiated cristae; sternum s.c. adipocytes contained a few, small mitochondria, with poorly developed cristae. Lipogenesis from acetate was high in BAT but barely detectable in sternum s.c. adipose tissue. In Exp. 2, we compared perirenal and tailhead adipose tissues between NE-infused Angus (n = 6) and Brahman (n = 7) newborn calves. Brahman BAT contained two-to-three times as many total beta-receptors as Angus BAT. The mitochondrial UCP1:28S rRNA ratio was greater in Brahman BAT than in BAT from Angus calves. Lipogenesis from acetate and glucose again was high, but lipogenesis from palmitate was barely detectable. Tail-head s.c. adipose tissue from both breed types contained adipocytes with distinct brown adipocyte morphology. In Exp. 3, three fetuses of each breed type were taken at 96, 48, 24, 14, and 6 d before expected parturition, and at parturition. Lipogenesis from acetate and glucose in vitro decreased 97% during the last 96 d of gestation in both breed types, whereas the UCP1 gene expression tripled during gestation in both breed types. At birth, palmitate esterification was twice as high in Angus than in Brahman BAT and was at least 100-fold higher than in BAT from NE-infused calves from Exp. 2. Uncoupling protein-1 mRNA was readily detectable in tailhead s.c. adipose tissue in all fetal samples. In Exp. 4, male Brahman and Angus calves (n = 5 to 7 per group) were assigned to 1) newborn treatment (15 h of age), 2) 48 h of warm exposure (22 degrees C) starting at 15 h of age, or 3) 48 h of cold exposure (4 degrees C) starting at 15 h of age. Brahman BAT adipocytes shrank with cold exposure, whereas Angus BAT adipocytes did not. Similarly, BAT from neonatal lambs (Exp. 5; n = 6 per group) was depleted of lipid in response to cold exposure, although UCP1 gene expression persisted. In Exp. 4, NE stimulated lipogenesis from palmitate in BAT incubated in vitro. Lipogenesis from palmitate was higher in Angus than in Brahman BAT, and increased with both warm and cold exposure. These studies suggest that BAT from Brahman calves may be exhausted of lipid shortly after birth during times of cold exposure.     

Effects of pre-natal glucocorticoids on testicular development in sheep

We tested the hypothesis that acute pre-natal exposure to high levels of synthetic glucocorticoid (betamethasone) would alter fetal testicular development through actions on gonadal glucocorticoid receptors (GRs). Pregnant Merino ewes bearing singleton male fetuses (n = 24) were allocated randomly among four equal groups to be injected intramuscularly with saline or betamethasone (0.5 mg/kg) either on day 109 of gestation or on both day 109 and day 116 of gestation. Fetal testes were collected at post-mortem, 5 days after each treatment. The volume of interstitial tissue and the volume, length and diameter of the sex cords were measured, and Sertoli cells and gonocytes were counted. For cord volume and interstitial tissue volume, control testes demonstrated maturational changes as fetal age advanced from 109 to 116 days of gestation. For that period, the single injection of betamethasone significantly reduced Leydig cell proliferation (P < 0.05), but had no effect on Sertoli cell numbers. Immunohistochemistry was used to localize GR and proliferating cell nuclear antigen in testicular cells. GR immunoexpression in Leydig cells was higher in fetuses exposed to betamethasone at 109 days of gestation than in control fetuses. Sertoli cells showed low levels of GR. It was concluded that, during mid-gestation, a brief period of glucocorticoid treatment could affect testicular development in male sheep fetuses. The mechanism probably involves direct effects on Leydig cells, as these cells express extra-GR in response to the treatment. Sertoli cells seem to produce less GR than Leydig cells, perhaps explaining their lack of response to betamethasone. These outcomes may have important implications for future fertility in male offspring.     

Correlated changes in reproductive components accompanying 10 generations of selection for improved sow productivity index.

Reproductive components were compared between a line of sows selected (S) for improved sow productivity index (SPI = 6.5 x number born alive + adjusted 21-d litter weight) and sows from an unselected control (C) line. Generation 9 and 10, second-parity, Landrace sows were chosen from both the S (n = 35) and C (n = 33) line. Sows were slaughtered at a commercial slaughter plant at approximately 75 d of gestation and their reproductive tracts were recovered. Reproductive tracts were evaluated for uterine weight (UTWT), uterine horn length (UTLN), ovulation rate (OR), number of fully formed fetuses (NF), number of mummified fetuses (NM), percentage of fetal survival (FS = NF/OR), fetal space (FSPACE = UTLN/[NF + NM]), and fetal position, sex, and weight. Select-line sows had greater NF (P less than .10) and higher FS (P less than .10) than C-line sows. Select-line sows had longer (P less than .05), and heavier (P less than .01) uteri than C-line sows. However, uterine length adjusted for NF was not different between the two lines. Uterine weight adjusted for NF was greater in S-line sows (P less than .05). Select-line sows had greater total fetal weight (TFWT) (P less than .05) than did C-line sows. Female fetuses positioned between two male fetuses were lighter in weight than all other female fetuses (P less than .01). Male fetuses positioned between two female fetuses did not differ in weight from all other male fetuses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Porcine preadipocyte proliferation and differentiation: a role for leptin?

The present study was designed to determine whether porcine leptin can alter the proliferation and differentiation of the porcine preadipocyte. The stromal vascular cell fraction of neonatal pig s.c. adipose tissue was isolated by collagenase digestion, filtration, and subsequent centrifugation. For differentiation studies, cells were seeded on six-well tissue culture plates and proliferated to confluency in 10% (vol/vol) fetal bovine serum (FBS) in Dulbecco's modified Eagle medium/F12 (DMEM/F12; 50:50). Cultures were differentiated using 2.5% pig serum (vol/vol) and recombinant porcine leptin at concentrations of 0 to 1,000 ng/mL alone or in combination with porcine insulin (100 nM), dexamethasone (1 microM), or IGF-1 (250 ng/mL). After 7 d of lipid filling, cultures were harvested for analysis of sn-glycerol 3 phosphate dehydrogenase (GPDH) and lipoprotein lipase (LPL). The GPDH and LPL activities are measures of preadipocyte differentiation. Data were corrected for protein content of the cultures. For proliferation experiments, 24 h after seeding cells with 10% FBS in DMEM/F12 in 25-cm2 tissue culture flasks, cells were switched to 5% FBS and supplemented with 0 to 1,000 ng of porcine leptin or 1,000 ng of murine leptin. Cell proliferation was measured by 3H-thymidine incorporation in preconfluent cultures over 24 h on d 4 of culture. At confluency, cells were switched to a medium to promote differentiation and lipid filling (2.5% pig serum, 100 nM insulin, 1 microM dexamethasone) for 7 d. Cells were harvested from the flasks and adipocytes were separated from stromal cells by Percoll gradient centrifugation. In a series of experiments, leptin alone or in combination with insulin, dexamethasone, or IGF-I did not affect differentiation as measured by the activity of GPDH and LPL. Leptin at any concentration did not inhibit differentiation induced by insulin, dexamethasone, or IGF-I; however, leptin at 1,000 ng/mL stimulated a 30% increase in preadipocyte proliferation (P = 0.007; n = 6) and a 27% increase in stromal cell proliferation (P < 0.001; n = 6). These results indicate that, at most, porcine leptin may contribute to the recruitment of new adipocytes within the adipose tissue.