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Biomedical subjects

S B Smith

Publications and source records attributed to S B Smith.

At least 19 recordsLinked to original sources

Direct mechanical measurements of the elasticity of single DNA molecules by using magnetic beads.

Single DNA molecules were chemically attached by one end to a glass surface and by their other end to a magnetic bead. Equilibrium positions of the beads were observed in an optical microscope while the beads were acted on by known magnetic and hydrodynamic forces. Extension versus force curves were obtained for individual DNA molecules at three different salt concentrations with forces between 10(-14) and 10(-11) newtons. Deviations from the force curves predicted by the freely jointed chain model suggest that DNA has significant local curvature in solution. Ethidium bromide and 4',6-diamidino-2-phenylindole had little effect on the elastic response of the molecules, but their extent of intercalation was directly measured. Conversely, the effect of bend-inducing cis-diamminedichloroplatinum (II) was large and supports the hypothesis of natural curvature in DNA.

Chemical Phenomena

Esterification of fatty acids by bovine intramuscular and subcutaneous adipose tissues.

Exogenous fatty acid esterification in intramuscular and subcutaneous adipose tissues from 72-hr fasted or ad libitum fed Angus cattle was investigated. Intramuscular (interfascicular) and subcutaneous adipose tissue snips were obtained from the longissimus dorsi muscle and were incubated with radioisotopically labeled fatty acids (palmitate, stearate, oleate, linoleate or linolenate) at three different concentrations (0.3 mM, 0.6 mM and 2.0 mM) to assess rates of fatty acid incorporation into glycerolipids. Rates of fatty acid esterification in vitro increased with fatty acid concentration in both intramuscular and subcutaneous adipose tissues. For all of the fatty acids investigated, triglycerides were the predominant products (60-85%). Subcutaneous adipose tissue had larger adipocytes and more actively (P less than 0.05) esterified fatty acids, with the exception of palmitate, than intramuscular adipose tissue. The rate of palmitate esterification was not different between tissues, although intramuscular adipose tissue esterified a greater proportion (P less than 0.10) of palmitate as triglyceride (85%) than did subcutaneous adipose tissue (75%). Relative rates of incorporation of fatty acids into lipids in intramuscular and subcutaneous adipose tissues were: palmitate greater than linolenate greater than linoleate greater than stearate. In general, 72-hr fasting did not significantly reduce the rates of fatty acid incorporation in bovine adipose tissues. Results of this study revealed that:i) rates of exogenous fatty acid incorporation into adipose tissue lipids were dependent on the medium fatty acid concentration and adipose tissue depot; and ii) the relative esterification rates of the various fatty acids in vitro did not necessarily reflect the proportion of these fatty acids in bovine adipose tissues.

Adipose Tissue

C57BL/6J-vit/vit mouse model of retinal degeneration: light microscopic analysis and evaluation of rhodopsin levels.

The C57BL/6J-vit/vit mouse is a newly described model of retinal degeneration in which photoreceptor cells die over the course of a year and the retinal pigment epithelium is unevenly pigmented. The present study utilized histological and biochemical techniques to assess the progression of the retinal degeneration in the vit/vit mouse ages 2 weeks to 8 months. Results of systematic morphometric evaluation indicated that the inner nuclear and plexiform layers of the retina are similar in thickness to age-matched C57BL/6J controls, but the outer plexiform layer is significantly thinner by 4 months. Rows of photoreceptor cells are lost at a rate of about one per month beginning at 2 months of age. By 8 months, the photoreceptor cell nuclei have diminished to only two to three rows. Inner segments of the vit/vit retina are similar in length to controls. Outer segments separate from the RPE during the first 2 months, they seem to be elongated at 2-3 months, but become severely disrupted past 4 months. Beginning at about 5 months, numerous darkly-staining cells resembling photoreceptor cell nuclei are observed in the area of the inner and outer segments and the subretinal space. Spectrophotometric analysis of rhodopsin indicated similar levels in vit/vit and controls at 6 weeks but a 50% reduction by 22 weeks. At 46 weeks, the level of rhodopsin in the mutant animal was less than 0.1 nmol per retina. The loss of rhodopsin in the vit/vit retinas correlated strongly with the decreasing number of rows of photoreceptor cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging

Synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) and developmental expression of IRBP mRNA in normal and rd mouse retinas.

The synthesis and secretion of interphotoreceptor retinoid-binding protein (IRBP) was quantitatively assessed in retinas of normal and rd mutant mice using short-term organ culture with [35S]methionine. Retinas were studied at ages P9-P12, time points prior to and immediately after the onset of the degeneration of the rd retina. Soluble proteins of the retinal pellet and the incubation medium were subjected to SDS-polyacrylamide gel electrophoresis. Analysis of labeled protein bands utilized a radioactivity scanning system to quantify [35S]methionine incorporation into newly synthesized IRBP. The synthesis and secretion into the incubation medium of IRBP by rd mouse retinas was comparable to normal retinas at P9-P10 but decreased by more than 50% by P12. IRBP mRNA levels were evaluated in retinas of normal and rd mice ages P7-P14. Although IRBP mRNA expression increased in the rd mouse through P10, it decreased markedly thereafter. Previously reported immunocytochemical studies suggested that IRBP was not secreted in the rd mouse retina. The results of this study indicate, however, that rd mouse retinas, when removed from the eye, have the capacity to synthesize and secrete IRBP.

Animals

Triacylglycerol biosynthesis in bovine liver and subcutaneous adipose tissue.

1. Adipose tissue from Angus and Brahman steers incubated with [1-14C]palmitate in the absence and presence of glucose exhibited a greater rate of lipid production than liver (P < 0.05). 2. Homogenates of adipose tissue used in the glycerol-3-phosphate acyltransferase assay exhibited a greater glycerolipid specific activity (nmol lipid/mg protein/30 min) when compared to liver (P < 0.05). 3. The inverse was true for liver homogenates when calculated for tissue activity (nmol lipid/g tissue/30 min). 4. Lysophosphatidate was produced in greater (P < 0.05) amounts than all other glycerolipids in the glycerol-3-phosphate acyltransferase assay. 5. The activity of phosphatidate phosphohydrolase in liver homogenates displayed greater rates than their respective adipose tissue homogenates. 6. Diacylglycerol acyltransferase activity was greater in adipose tissue homogenates compared to liver homogenates.

Acyltransferases

Fatty acid composition and fatty acid elongase and stearoyl-CoA desaturase activities in tissues of steers fed high oleate sunflower seed.

The effects of a high oleate sunflower seed diet on tissue composition and on fatty acid elongation and desaturation enzyme activities were investigated. Three Simmental calves were fed a standard corn-based diet (2.6% fat); three others were fed the corn-based diet containing 20% high oleate sunflower seed (10.4% fat). Blood samples and perianal adipose tissue samples were obtained at 7, 90 and 180 d on trial. Samples of liver, longissimus dorsi muscle and intestinal mucosa were obtained at the termination of the experiment (195 d on trial). Plasma oleate was higher (P < 0.05) in the cattle fed the sunflower seed, and oleate, myristate (P < 0.05) and stearate (P < 0.06) were elevated in perianal adipose tissue in response to the greater lipid content of the dietary sunflower seed. Dietary sunflower seed decreased the concentration of stearate (P < 0.05) in liver. The high oleate diet significantly (P < 0.05) increased the activity of stearoyl-CoA desaturase activity in muscle, and numerical increases in desaturase activity were observed in liver, adipose and small intestine samples. Elongase activity was unaffected by diet. Because stearate is the primary fatty acid available for absorption in ruminants, elevated oleate in plasma and depressed stearate in liver of cattle fed sunflower seed may have reflected an adaptive response of stearoyl-CoA desaturase in their tissues.

Acetyltransferases

Nitrogen and lipid metabolism in heifers fed at increasing levels of intake.

The relationship between N and lipid metabolism was investigated in heifers fed five different levels of feed intake (five heifers per treatment group). Targeted ME intakes were 84, 123, 157, 191, and 225 kcal per kt.75 per day, which were .76x, 1.12x, 1.43x, 1.74x, and 2.05x (times) the estimated ME requirement for maintenance. After 120 d on trial, the heifers were moved to a confinement building for 7 d, and feces and urine were collected over a 3-d period. On the 1st d of confinement, blood samples were collected every 15 min for a total of 15 samples. Because the group fed at 1.43x maintenance was fed improperly during the period in confinement, this group was omitted from the study. Adipose tissue samples were obtained at slaughter to obtain in vitro measures of lipid metabolism. As feed intake increased, N retention increased (P less than .05) from 1.7 to 24.3 g/d. Daily urinary N tau-methylhistidine excretion was significantly different between the .76x and 1.74x treatments (769 vs 1,575 mumol/d, respectively). The fractional breakdown rate of myofibrillar proteins also was significantly different between these two groups of heifers (1.49 vs 2.44%/d, respectively). Plasma glucose and insulin were lowest (P less than .05) at the lowest level of feed intake. Conversely, plasma nonesterified fatty acids were lowest (P less than .05) in those animals receiving the highest level of feed intake. Subcutaneous adipocytes were smallest (93 microns) in the heifers fed at 33% ad libitum intake and largest in heifers fed at 76 or 90% ad libitum intake (115 and 110 microns, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue

In vitro metabolism of glucose by bovine reproductive tissues obtained during the estrous cycle and after calving.

Two experiments were conducted to determine the effect of physiological state (stage of the estrous cycle or time after calving) on the in vitro metabolism of glucose by reproductive tissues. In Exp. 1, the corpus luteum and ipsilateral uterine horn were collected at surgery from 15 cows at early (d 6 to 7), middle (d 10 to 13), or late (d 17 to 19) stages of the estrous cycle. Luteal or endometrial tissues (45 mg) were incubated (4 h, 37 degrees C) in metabolic flasks containing Nutrient Mixture F-10 (3 mL), increasing concentrations of glucose (1, 2, 5, 10, or 15 mM), and 1 microCi of [U-14C]glucose. Luteal tissue collected at the middle stage of the estrous cycle had greater (P less than .04) rates of glucose uptake (17.8 vs 12.1 ng/mg of wet tissue per min) and oxidation (138.6 vs 67.7 pg/mg of wet tissue per min) and a lower (P less than .02) rate of metabolism of glucose to lactate (10.5 vs 13.3 ng/mg of wet tissue per min) than tissue collected at the late stage. Compared to tissue collected at early and late stages, endometrial tissue collected at the middle stage of the estrous cycle had a similar (P greater than .5) rate of glucose uptake (29.6 vs 27.8 and 26.1 ng/mg of wet tissue per min), a lower (P less than .02) rate of metabolism of glucose to lactate (13.8 vs 16.6 and 16.4 ng/mg of wet tissue per min), and a greater (P less than .1) rate of oxidation of glucose (84.1 vs 65.8 and 72.3 pg/mg of wet tissue per min). In Exp. 2, the previously gravid uterine horn was collected at surgery on d 20 (n = 8) or 30 (n = 7) after calving, or the uterine horn ipsilateral to the preovulatory follicle was collected at first observed estrus after calving (n = 8). Uterine tissue obtained at first estrus after calving had a greater (P less than .003) rate of glucose uptake and oxidation (88.8 vs 77.3 and 75.5 pg/mg of tissue per min) than d-20 or d-30 endometrium.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

Ruminal biohydrogenation of fatty acids from high-oleate sunflower seeds.

The objective of these experiments was to examine methods of modifying the fatty acid composition of bovine tissues. In the first experiment, four steers were fitted with duodenal fistulas and were assigned to four diets in a Latin square design. The steers were fed a control diet or the same diet containing 10% high-oleate partially crushed sunflower seeds, serum-coated sunflower seeds, and heat-treated, serum-coated sunflower seeds for 5 d. Samples of digesta and feces were collected on d 5. The inclusion of sunflower seeds (plain or serum-coated) in the diet increased (P less than .05) the digesta concentration of stearate. The percentage of stearate in the digesta and feces was increased (P less than .05) from 51 to 67% and from 64 to 74%, respectively, when steers were fed the untreated sunflower seed. The fecal concentration of oleate was increased (P less than .05) by dietary sunflower seeds in steers that were fed the serum-coated, unheated sunflower seeds. In a second experiment, heifers (four per group) were fed a corn-based control diet or diets containing 10% of high-oleate sunflower oil encapsulated with calcium alginate, either plain, coated with blood meal, or with blood meal integrated into the pellet. After 50 d on treatment, samples of perianal adipose tissue were obtained by biopsy. The fatty acid composition of the adipose tissue was not modified by the inclusion of the encapsulated oleate in the diet. In summary, limited ruminal bypass of sunflower seed oleate was accomplished with sunflower seed but not with encapsulated oleate.

Adipose Tissue

Model and computer simulations of the motion of DNA molecules during pulse field gel electrophoresis.

A model is presented for the motion of individual molecules of DNA undergoing pulse field gel electrophoresis (PFGE). The molecule is represented by a chain of charged beads connected by entropic springs, and the gel is represented by a segmented tube surrounding the beads. This model differs from earlier reptation/tube models in that the tube is allowed to leak in certain places and the chain can double over and flow out of the side of the tube in kinks. It is found that these kinks often lead to the formation of U shapes, which are a major source of retardation in PFGE. The results of computer simulations using this model are compared with real DNA experimental results for the following cases: steady field motion as seen in fluorescence microscopy, mobility in steady fields, mobility in transverse field alternation gel electrophoresis (TFAGE), mobility in field inversion gel electrophoresis (FIGE), and linear dichroism (LD) of DNA in agarose gels during PFGE. Good agreement between the simulations and the experimental results is obtained.

Chemical Phenomena

Acylation and glycosylation of rhodopsin in the rd mouse.

Retinas of 9-10-day-old rd and control mice were incubated for 2 hr with [14C]leucine along with either tritiated palmitic acid or galactose to investigate the acylation or glycosylation, respectively, of rhodopsin. Although other laboratories have reported that phosphorylation of rhodopsin is not detectable in rd retinas, the two post-translational modifications of rhodopsin investigated in the present work are detectable. The rod outer segments (ROS) were separated from the retinal debris containing the rough endoplasmic reticulum (RER) of photoreceptor cells by vortexing and then by linear sucrose gradients. The rhodopsin from the RER was purified by affinity chromatography and gel electrophoresis. In the acylation studies, the mean ratio of palmitate to leucine in the rd mouse was nearly twice that of controls (11.73 +/- 2.84 v. 6.81 +/- 1.04). Possible explanations for the disparity between the two groups could include: (1) a diminished internal pool size of the fatty acid; or (2) acylation of amino acids such as serine or threonine which normally are not acylated in rhodopsin. Treatment of purified rhodopsin with 1 M hydroxylamine released similar amounts of palmitate from the rd mice and controls. Hence, the higher ratio of palmitate to leucine in rd mice is apparently due to a diminished internal pool size. In the glycosylation studies, the ratio of galactose to leucine was very similar between rd mice and controls, 1.7 +/- 0.43 v. 2.47 +/- 0.74. Protein content and specific activity were determined for the crude ROS preparations and for the remaining retinal debris. Although the amount of ROS protein differed significantly between the two groups, the specific activities did not.(ABSTRACT TRUNCATED AT 250 WORDS)

Acylation

Transient hyperglycosylation of rhodopsin with galactose.

Rhodopsin's oligosaccharide chains contain predominantly two types of sugar residues: mannose and N-acetylglucosamine. In the present work, bovine and rat rhodopsin were analysed biochemically for the presence of a third sugar, galactose. Treatment of bovine rod outer segments (ROS) with galactose oxidase followed by reduction with tritium-labeled sodium borohydride revealed the presence of existing molecules of galactose on rhodopsin. Rats injected intravitreally with [3H]galactose and [14C]leucine and maintained in darkness were killed 1 hr, 6 hr, 1, 3 or 5 days following the injection. Retinas were collected for subcellular fractionation and rhodopsin from each of the fractions was purified by ConA sepharose chromatography and SDS-PAGE. During the first 6 hr, galactose selectively labeled rhodopsin in the Golgi-enriched fraction resulting in increased [3H]/[14C] ratios in both Golgi and ROS. The data suggested that trimming was occurring at the transition from Golgi to ROS. Furthermore, a decrease in isotope ratio in the ROS between 6 hr and 1 day suggested further trimming of rhodopsin after membrane assembly in the ROS. Additional in vivo experiments demonstrated existing molecules of galactose on rhodopsin's oligosaccharide chain using lectin affinity chromatography. Rats injected intravitreally with [35S]methionine were dark-adapted for 2 hr. Following subcellular fractionation of retinas, ConA purified rhodopsin from ROS was applied to one of two additional lectin columns: Ricinus communis agglutinin (RCA) or Griffonia simplicifolia I (GSA). Eight to nine percent of the labeled rhodopsin was bound to and eluted from RCA, whereas none bound to GSA, indicating the presence of a beta-galactoside. The RCA agarose eluted protein co-electrophoresed with a rhodopsin standard and was light sensitive. Galactose was shown to be the terminal sugar on this subset of rhodopsin and was not capped by neuraminic acid. Binding of rhodopsin's oligosaccharide to RCA was abolished by pre-treatment with beta-galactosidase. Decreased binding of rhodopsin to RCA was observed following intravitreal injection of castanospermine but not swainsonine. Of those two inhibitors of glycoprotein trimming, only castanospermine would be expected to prevent the addition of galactose to the oligosaccharide. The association of galactose with rat rhodopsin appeared to be a transient one. At 2 hr, 8-9% of rhodopsin contained galactose, at 6 hr only 2.2% had galactose and by 24 hr less than 1% did. The galactose was trimmed from rhodopsin's oligosaccharide presumably after its role was complete. Separation of rhodopsin of the plasma membranes from rhodopsin of discs indicated that 75% of the galactose-containing rhodopsin was in the plasma membrane and only 25% was in the discs. These findings suggested a possible role for galactose in new disc formation with subsequent removal after the discs are sealed.

Animals

Lipogenesis in acute and 48-hour cultures of bovine intramuscular and subcutaneous adipose tissue explants.

In this study, the interactions among breed of cattle, adipose tissue site and specific incubation conditions were investigated. Subcutaneous and i.m. adipose tissues were obtained from 10 Angus and 9 Santa Gertrudis steers immediately postmortem. Adipose tissue explants were incubated acutely for 2 h immediately at slaughter or after being cultured 48 h with or without 1 mU/ml insulin and 30 mg/ml bovine serum albumin; the incorporation of 14C-labeled acetate and glucose (5 mM, plus 5 mM unlabeled lactate) into lipid fractions was measured. AT the same chronological age, Angus steers had a more youthful lean maturity score, higher USDA marbling score and higher USDA quality grade (P less than .05) than did carcasses from Santa Gertrudis steers. The lower marbling score of the Santa Gertrudis steers was paralleled by smaller i.m. adipocytes (P less than .05) relative to Angus steers. Pentose cycle reductase and NADP-malate dehydrogenase activities were greater in Angus i.m. adipose tissue than in Santa Gertrudis i.m. adipose tissue, which would provide more reducing equivalents (NADPH) and glycerol for fatty acid biosynthesis and triacylglycerol esterification. Correspondingly, Angus i.m. adipose tissue exhibited a greater rate of lipogenesis from acetate and glucose (P less than .05) than did Santa Gertrudis i.m. adipose tissue in acute incubations. The presence of insulin resulted in higher rates of lipogenesis from acetate in Angus s.c. adipose tissue than in Santa Gertrudis s.c. adipose tissue after 48 h of explant culture. These data indicate that i.m. and s.c. adipose tissues exhibit aspects of lipid metabolism unique to each tissue and suggest that breed-related differences in adipose tissues may explain the divergent responses to insulin observed in different laboratories.

Adipose Tissue

Fatty acid elongation and desaturation enzyme activities of bovine liver and subcutaneous adipose tissue microsomes.

Assay conditions were established for the fatty acid elongation and the delta 9 desaturase enzyme systems of bovine liver and adipose tissue microsomes; rat liver microsomes were used as a reference. Overall fatty acid elongation was determined by measuring the incorporation of [2-14C]malonyl-coenzyme A (CoA) to 14C-labeled stearate. Rat liver elongation activity was .50 +/- .02 nmol.min-1.mg protein-1; bovine liver microsomal elongation activity was substantially lower (P less than .05), with a mean value of .15 +/- .02 nmol.min-1.mg protein-1. The elongation activity of bovine s.c. adipose tissue microsomes (.42 +/- .10 nmol.min-1.mg protein-1) was not different (P greater than .05) from the activity observed in rat liver microsomes. To determine the fatty acid delta 9 desaturase activity, microsomes were incubated in the presence of [1-14C]stearoyl-CoA and nicotinamide adenine dinucleotide (reduced form) (NADH), and the production of radioactively labeled oleate was quantified. Microsomal delta 9 desaturase activity was similar in rat liver and bovine s.c. adipose tissue microsomes with rates of .15 +/- .04 and .21 +/- .05 nmol.min-1.mg protein-1, respectively. However, no desaturase activity was detected in bovine liver microsomes, indicating that the liver is not a major site of oleate synthesis in this species. To investigate differences in fatty acid metabolism relative to breed type, eight Angus and seven Braford heifers were slaughtered at approximately 12 mo of age. Subcutaneous fat thickness over the 12th-13th thoracic vertebrae was greater in the Angus heifers than in the Braford heifers. However, no differences (P greater than .05) were observed in mean adipocyte size or number of cells per gram of adipose tissue between the Angus and Braford heifers. Similarly, there were no significant differences between the Angus and Braford s.c. adipose tissues for microsomal fatty acid elongation or delta 9 desaturation, or for nicotinamide adenine dinucleotide phosphate (NADP)-malate dehydrogenase, fatty acid synthetase, or the pentose cycle reductases. The inability of bovine liver to convert stearate to oleate was in agreement with the fatty acid composition of the liver lipid, which had a smaller percentage of oleate and a higher percentage of stearate than s.c. adipose tissue.

Acetyltransferases

Relationship between fatty acid-binding protein activity and marbling scores in bovine longissimus muscle.

Studies were conducted in an attempt to establish a relationship between fatty acid-binding protein (FABP) activity and marbling score in bovine longissimus muscle. Longissimus muscle was obtained from four 20-mo-old Charolais-Hereford crossbred heifers, three 16-mo-old Angus steers, and four 18-mo-old Angus steers. Immediately after slaughter, longissimus muscles were removed for the extraction of FABP. Supernatant (S104) fractions containing 41.3 to 144 mg of protein (depending on animal group) were eluted over Sephadex columns, and elution fractions were analyzed for the binding of radiolabeled palmitoyl-coenzyme A (CoA). Specific activities of FABP were 23, 32, and 101 nmol palmitoyl-CoA bound/mg protein for the Charolais-Hereford, 16-mo-old Angus, and 18-mo-old Angus cattle, respectively. These preliminary results suggested that longissimus muscle FABP activity was positively correlated with marbling score. To test specifically for this possibility, longissimus muscle was obtained at slaughter from each of four Wagyu steers, Angus heifers and Braford heifers. Marbling scores taken at the 12th-13th rib junction were Sm45, Sm43, and SI50 for the Wagyu, Angus, and Braford cattle, respectively. Interfascicular adipose tissue was exhaustively removed from sections of the 5th to 8th thoracic region of the longissimus muscle to eliminate any contribution of adipose tissue to FABP activity. For each animal, 300 mg of the S104 were eluted over Sephadex columns. Specific activities for the Wagyu, Angus, and Braford longissimus muscle FABP were 3.1, 3.8, and 3.9 pmol palmitoyl-CoA bound/mg protein, respectively, and were not different (P greater than .05) among the three animal groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue

Growth, carcass traits, and fatty acid profiles of adipose tissues from steers fed whole cottonseed.

To investigate the impact of dietary whole cottonseed (WCS) level on fatty acid composition, growth, and carcass traits, 45 Hereford steers were assigned to diets containing 0, 15, or 30% dietary WCS. The 15 and 30% WCS contributed an estimated 3.3 and 6.6% additional lipid, respectively, to the diets. After being fed for 54 d, all animals were weighed and slaughtered, and carcass measurements were obtained. There were no differences (P greater than .05) among dietary treatment groups in live weight or ADG for the 54-d feeding period. Control steers had larger (P less than .05) longissimus muscle areas than steers fed 30% WCS, which accounted for the advantage in yield grade (P less than .05) exhibited by the control group. Feeding of 30% WCS resulted in minor increases in linoleic and total polyunsaturated fatty acid content of perinephric fat expressed in both normalized (area percentage) and gravimetric (g/100 g of fresh tissue) formats. There were no significant differences in the monounsaturated or saturated fatty acid content of adipose tissues from animals fed the different diets. Subcutaneous adipose tissue samples were higher (P less than .01) in total unsaturates but had lower (P less than .05) proportions of C18:0 and C18:1 than perinephric samples. Feeding WCS at the levels reported herein only had minor effects on fatty acid composition of beef adipose tissues.

Adipose Tissue

The localization and timing of post-translational modifications of rat rhodopsin.

Rat retinas were labeled by incubation with, or intravitreal injection of, [14C]leucine along with tritiated palmitic acid, glucosamine or galactose. At selected intervals, subcellular fractions were prepared on linear sucrose gradients and rhodopsin was extracted and purified by affinity chromatography and gel electrophoresis. Time courses revealed that leucine rapidly and transiently labeled the rhodopsin in the rough endoplasmic reticulum (RER), with a maximum at 1.5 hr post-injection. Subsequently, the rod outer segments (ROS) contained the labeled rhodopsin, with the ROS labeling maximally at 6-12 hr. Palmitate labeling followed the same pattern but was subject to a delay, presumably because of a large intracellular pool of the fatty acid. With palmitate the RER rhodopsin was not maximally labeled until 12 hr. The acylation of rhodopsin takes place in the RER sometime after the polypeptide has been translated but before transport to the Golgi. Glucosamine labeling was also delayed because of intracellular pools of the sugar or its metabolic derivatives. But because of secondary glycosylation in the Golgi, the rhodopsin in the ROS also labeled maximally with glucosamine at about 6 hr. Administration of [3H]galactose resulted in the labeling of rhodopsin both in vivo and in vitro, in part possibly because of its conversion to mannose and subsequent insertion into the core oligosaccharide on the RER. However, in the ROS the ratio of tritium, derived from [3H]galactose, to [14C]leucine decreased by a factor of 2 between 6 and 24 hr post-injection. Moreover, between 6 and 12 hr post-injection, labeled rhodopsin molecules in the ROS underwent a shift in mobility on gels indicative of trimming to a lower molecular weight. Thus some sugar residues may be added to the rhodopsin in the inner segment and removed in the ROS.

Animals