Comparison of immunoglobulin G absorption in calves fed maternal colostrum, a commercial whey-based colostrum replacer, or supplemented maternal colostrum.

Successful passive transfer of antibodies in neonatal calves can be achieved by feeding an adequate quantity and quality of maternal colostrum (MC) or colostrum replacer (CR). An alternative could be feeding low-quality maternal colostrum (LMC) with added IgG from a CR. The objective of this study was to determine if a commercial whey-based CR product containing low levels of casein (Premolac PLUS Bovine IgG; Zinpro Corporation, Eden Prairie, MN) fed to replace MC or supplement LMC could lead to adequate serum IgG levels and apparent efficiency of absorption (AEA) in neonatal dairy calves. Holstein calves (n = 20 per treatment) were separated from their dam after birth and randomly assigned to be fed 3.79 L of MC (106 g/L of IgG; 401 g of IgG fed), LMC (30 g/L IgG) supplemented with CR (41 g/L IgG; 154 g of IgG total fed; LMC-CR), or 1.3 L of 1 of 2 levels of CR (110 or 150 g of IgG fed; CR-110 or CR-150) within 1.5 h of birth. Colostrum was obtained from the first (MC) or second and third milkings (LMC) of cows from Pennsylvania State University dairy and pooled by source into large batches. Blood samples were taken from calves before colostrum feeding and 24 h after birth and were analyzed for serum total protein, total IgG, hematocrit, and Brix percentage. Calves fed MC had higher 24-h IgG values (means ± SEM) than calves fed LMC-CR (27.04 ± 1.07 vs. 22.33 ± 1.08 mg/mL, respectively). Feeding 150 g of IgG from CR led to higher 24-h serum IgG values than feeding 110 g of IgG (16.90 ± 1.09 vs. 12.79 ± 1.08 mg/mL). Serum IgG levels were different between the CR-fed calves and the calves fed LMC-CR and MC, but all had average values >10 mg/mL IgG. Calves fed LMC-CR had greater AEA than calves fed MC (54.58 ± 2.39 vs. 24.38 ± 2.36%, respectively). Among calves fed CR-110 or CR-150, AEA did not differ. Serum total protein and Brix percentage had strong correlations with actual IgG values across the entire study. We found no differences in average daily gain or health variables measured, and no differences in final hip width, withers height, or body weight for calves fed MC, LMC-CR, CR-150, or CR-110. These results indicate that CR can be fed successfully as an alternative to MC or as a supplement to colostrum with low IgG.

Short communication: Variation in serum immunoglobulin G concentrations from birth to 112 days of age in Holstein calves fed a commercial colostrum replacer or maternal colostrum.

Serum IgG concentrations in dairy calves change throughout their first weeks of life, peaking at 24 h and then steadily decreasing until calves begin to produce endogenous IgG. The objective of this study was to observe serum IgG dynamics from birth until 16 wk of life in calves fed either maternal colostrum (MC) or colostrum replacer (CR). A total of 44 Holstein calves were randomly assigned to 1 of the 4 colostrum treatments and followed throughout the study. Treatments consisted of feeding high-quality MC, low-quality MC supplemented with CR, or 1 of 2 distinct levels of IgG concentration from CR. Overall, the interaction between type of colostrum fed and sampling time was significant. Individual differences for this effect were found at d 1, 7, 14, 21, 28, and 98, while the other time points were not different.

Varying dietary protein and fat elicits differential transcriptomic expression within stress response pathways in preweaned Holstein heifers.

Increases in milk replacer dietary energy subsequently increase growth and weight in preweaned dairy heifers. However, the underlying effects of dietary component increases on key functional pathways have yet to be fully investigated. Elucidating these relationships may provide insights into the mechanisms through which protein and fat are partitioned for tissue growth and metabolism. We hypothesized that genes within key growth and metabolic pathways would be differentially expressed between calves fed a protein- and fat-restricted diet and calves fed a protein- and fat-enhanced diet. The objectives of this study were to (1) identify genes differentially expressed between dietary restricted calves and enhanced calves and (2) determine the key regulatory pathways influenced by these genes. Preweaned Holstein heifers (n = 12; 6 ± 0.02 d of age) were randomly assigned to 1 of 2 milk replacer diets: enhanced (28.9% crude protein, 26.2% fat; n = 6) or restricted (20.9% crude protein, 19.8% fat; n = 6). Growth measures included average daily gain and gain-to-feed ratio. After 56 d, calves were killed for tissue collection. Samples from longissimus dorsi, adipose, and liver tissues were collected and RNA was isolated for RNA sequencing analysis. The MIXED procedure of SAS (SAS Institute Inc., Cary, NC) was used to evaluate relationships of growth with dietary energy. Fixed effects included date of collection and time (day). Random effects included sire and birth weight. The RNA sequencing analysis was performed using CLC Genomics Workbench (Qiagen, Germantown, MD), and the Robinson and Smith exact test was used to identify differentially expressed genes between diets. The Protein Analysis Through Evolutionary Relationships (PANTHER) database was then used to identify functional categories of differentially expressed genes. Enhanced calves had increased growth rates and feed efficiency compared with restricted calves (average daily gain = 0.76 and 0.22, respectively; gain-to-feed ratio = 0.10 and 0.06, respectively). There were 238 differentially expressed genes in adipose, 227 in longissimus dorsi, and 40 in liver. We identified 10 genes concordant among tissues. As expected, functional analyses suggested that the majority of genes were associated with metabolic or cellular processes, predominantly cell communication and cell cycle. Overall, it appears that varying levels of dietary protein and fat influence calf growth and development through metabolic processes, including oxidative phosphorylation and glyceroneogenesis. However, protein- and fat-restricted calves appeared to experience metabolic stress at a cellular level, as evidenced by an upregulation in stress response pathways, including genes in the p53 pathway. Calves could be fed at a higher level of protein and fat to decrease the prevalence of metabolic stress at the cellular level, but evidence indicating the presence of inflammatory stress and adipose fibrosis in enhanced calves prompts further investigation of the effects of milk replacer component levels.

Higher plane of nutrition pre-weaning enhances Holstein calf mammary gland development through alterations in the parenchyma and fat pad transcriptome.

BACKGROUND: To reduce costs of rearing replacement heifers, researchers have focused on decreasing age at breeding and first calving. To increase returns upon initiation of lactation the focus has been on increasing mammary development prior to onset of first lactation. Enhanced plane of nutrition pre-weaning may benefit the entire replacement heifer operation by promoting mammary gland development and greater future production. METHODS: Twelve Holstein heifer calves (< 1 week old) were reared on 1 of 2 dietary treatments (n = 6/group) for 8 weeks: a control group fed a restricted milk replacer at 0.45 kg/d (R, 20% crude protein, 20% fat), or an accelerated group fed an enhanced milk replacer at 1.13 kg/d (EH, 28% crude protein, 25% fat). At weaning (8 weeks), calves were euthanized and sub-samples of mammary parenchyma (PAR) and mammary fat pad (MFP) were harvested upon removal from the body. Total RNA from both tissues was extracted and sequenced using the Illumina HiSeq2500 platform. The Dynamic Impact Approach (DIA) and Ingenuity Pathway Analysis (IPA) were used for pathway analysis and functions, gene networks, and cross-talk analyses of the two tissues. RESULTS: When comparing EH vs R 1561 genes (895 upregulated, 666 downregulated) and 970 genes (506 upregulated, 464 downregulated) were differentially expressed in PAR and MFP, respectively. DIA and IPA results highlight a greater proliferation and differentiation activity in both PAR and MFP, supported by an increased metabolic activity. When calves were fed EH, the PAR displayed transcriptional signs of greater overall organ development, with higher ductal growth and branching, together with a supportive blood vessel and nerve network. These activities were mediated by intracellular cascades, such as AKT, SHH, MAPK, and Wnt, probably activated by hormones, growth factors, and endogenous molecules. The analysis also revealed strong communication between MFP and PAR. CONCLUSION: The transcriptomics and bioinformatics approach highlighted key mechanisms that mediate the mammary gland response to a higher plane of nutrition in the pre-weaning period.

Efficacy of colostrum replacer versus maternal colostrum on immunological status, health, and growth of preweaned dairy calves.

Commercially available colostrum replacers (CR) are commonly used when maternal colostrum (MC) is unavailable, for managerial convenience, to ensure quality consistency at first feeding, or in disease control and eradication programs. The objective of this study was to determine the efficacy of feeding First Day Formula (Accelerated Genetics, Baraboo, WI) CR versus pooled MC on immunological status, growth, and health of preweaned dairy calves. A total of 1,220 Jersey and Jersey × Holstein calves born on a California Central Valley dairy farm were assigned after birth to receive either CR or MC following a systematic allocation procedure. Calves assigned to MC were tube fed 2.8 L of MC, and calves assigned to CR were tube fed a total of 500 g of CR (150 g of immunoglobulin G; IgG) mixed into 1.9 L of water at 1 h ± 5 min after the calf was born. A subset of calves was selected for passive transfer (n = 592) and growth (n = 268) analyses. Although both coliform count and total bacteria count were low for MC and CR fed to calves during the study, the predicted probability of calves receiving contaminated liquid feed (coliform count >10,000 cfu/mL) at first feeding was reduced for calves fed CR (1.5%) compared with calves fed MC (6.1%). The mean blood concentration of IgG was lower for calves fed CR than for calves fed MC (19.6 vs. 23.4 mg/mL). However, the apparent efficiency of absorption of IgG did not differ between treatments (34.4 and 35.9% for CR and MC, respectively). Total proteins were lower in calves fed CR compared with MC at 24 h (5.16 vs. 5.84 g/dL, respectively). Calves fed CR were 1.5 kg lighter at weaning and gained 0.03 kg less per day (0.30 vs. 0.33 kg/d, respectively) than calves fed MC before weaning. Height at weaning did not differ between the 2 treatment groups. Calves fed CR tended to have a higher predicted probability of not being treated for diarrhea than calves fed MC (0.142 vs. 0.110, respectively). However, when the disease was present, CR had a higher number of treatment days compared with MC (11.6 vs. 10.8 d, respectively). The hazard ratio of dying did not differ between MC and CR; however, CR calves had a numerically higher risk (hazard ratio = 1.347) of dying compared with calves that received MC. In conclusion, IgG absorption and serum concentration of calves were adequate when calves were fed either CR or MC. The CR-fed calves had a lower probability of receiving contaminated liquid feed and performed similar in terms of health compared with calves receiving high-quality MC, although they were slightly lighter at weaning. Therefore, the CR evaluated in this study is a valid alternative to high-quality (>50 mg of IgG/mL) MC.

Feeding an enhanced diet to Holstein heifers during the preweaning period alters steroid receptor expression and increases cellular proliferation.

Preweaning diet and estradiol treatment alters mammary development. Our objectives were to study the effects of diet and estradiol on proliferation of mammary epithelial cells and expression of estrogen receptor α (ESR1) and progesterone receptors (PGR) in these cells. Thirty-six Holstein heifer calves were raised on (1) a control milk replacer fed at 0.44 kg of powder/head per day, dry matter (DM) basis (restricted, R; 20.9% crude protein, 19.8% fat, DM basis), or (2) an enhanced milk replacer fed at 1.08 kg of powder/head per day, DM basis (Enhanced, EH; 28.9% crude protein, 26.2% fat, DM basis). Milk replacer was fed for 8 wk. At weaning, a subset (n = 6/diet) of calves were euthanized and had tissue harvested. Remaining calves received estradiol implants (E2) or placebo and were euthanized at wk 10 to harvest tissue. Treatments were (1) R, (2) R + E2 (R-E2), (3) EH, and (4) EH + E2 (EH-E2). One day before euthanasia calves were given bromo-2'-deoxyuridine (BrdU; 5 mg/kg of body weight). At euthanization, mammary parenchyma was removed and fixed. Tissue sections from zone 1 (cisternal), 2 (medial), and 3 (distal) within the mammary gland were stained with hematoxylin and eosin and antibodies to measure expression of ESR1, PGR, and incorporation of BrdU. At wk 8, R-fed calves had more PGR-expressing cells in distal parenchyma; however, PGR expression intensity was greater in EH-fed calves. The proportion of cells expressing ESR1 was not affected by diet, but expression intensity (receptors per positive cell) was greater in EH-fed calves across all zones (62-81%). Overall, the percent BrdU-positive epithelial cells was 2 and 0.5 fold greater for EH-fed calves in zone 2 and 3. The proportion of labeled cells was greater in terminal ductal units than in subtending ducts, and treatment effects were more evident in terminal ductal units. At wk 10, calves treated with estradiol had 3.9-fold greater PGR expression intensity. The intensity and percent of cells expressing ESR1 was lowest in estradiol-treated calves. Overall, estradiol-treated calves had the greatest number of proliferating epithelial cells. Moreover, in zone 3, EH-E2 calves had a higher percentage of proliferating cells than in all other treatments. Results indicate both diet and estradiol administration alter proliferation rates of the mammary epithelium and that changes in expression of ESR1 and PGR are involved in enhanced mammary development. The data support our hypothesis that enhanced preweaning feeding increases the mammary tissue responsiveness to mammogenic stimulation.

Feeding a higher plane of nutrition and providing exogenous estrogen increases mammary gland development in Holstein heifer calves.

Feeding heifers a higher plane of nutrition postweaning but before puberty can negatively affect mammary gland development and future milk yield. However, enhanced nutrition preweaning may promote development and future production. Our objectives were to determine the effects of enhanced feeding preweaning and exogenous estrogen immediately postweaning on mammary gland development and the composition of the mammary parenchyma (PAR) and mammary fat pad (MFP). Thirty-six Holstein heifer calves (<1 wk old) were reared on 1 of 2 dietary treatments for 8 wk: (1) a restricted milk replacer fed at 0.45 kg/d (R; 20% crude protein, 20% fat), or (2) an enhanced milk replacer fed at 1.13 kg/d (EH; 28% crude protein, 25% fat). Upon weaning, calves from each diet (n=6) were given either a placebo or estrogen implant for 2 wk, creating 4 treatments: R, R + estrogen (R-E2), EH, and EH + estrogen (EH-E2). Calves were housed individually with ad libitum access to water. Starter feeding began at wk 5 and was balanced between treatments. Udders were evaluated by palpation and physical measurements weekly. Subsets of calves were killed at weaning (n=6 per diet) and at the conclusion of the trial (n=6 per treatment). Udders were removed, dissected, and weighed. At wk 8, EH calves had longer front and rear teats. Providing estrogen to EH calves increased the length of rear teats during wk 9 and 10. Enhanced-fed calves had 5.2-fold more trimmed mammary gland mass than R calves. Providing estrogen to EH calves further increased mammary gland weight. Masses of PAR and MFP were markedly greater for EH calves than for R calves (e.g., 7.3-fold greater PAR tissue). Estrogen increased the mass of both PAR and MFP in EH calves. Feeding a higher plane of nutrition increased total protein, DNA, and fat in the MFP and total protein and DNA in the PAR. Dual-energy x-ray absorptiometry estimates of mammary fat mass were highly correlated with biochemical analyses of fat content. From histological study, we observed that the degree of expansion of epithelium into the adjacent stromal tissue and the complexity of ductal development were minimal in R, increased in EH, and increased by estrogen in both dietary treatments. Results provide compelling evidence that preweaning nutrition and estrogen administration immediately postweaning markedly increase mammary gland development in dairy calves. Cellular and molecular mechanisms responsible for these differences are currently under study.

Growth, intake, and health of Holstein heifer calves fed an enhanced preweaning diet with or without postweaning exogenous estrogen.

Research has shown that changes in nutrition both before and after weaning can affect mammary development. Additionally, estrogen is known to be a potent mammogenic stimulant. Our objectives were to determine effects of altered preweaning feeding and exogenous estradiol postweaning on growth, intake, and health. Thirty-six Holstein heifer calves were reared on (1) a restricted milk replacer (MR) diet fed at 0.44kg powder dry matter (DM)/day [R; 20.9% crude protein (CP), 19.8% fat, DM basis], or (2) an enhanced MR fed at 1.08kg powder DM/d (EH; 28.9% CP, 26.2% fat, DM basis). The MR feeding was reduced 50% during wk 8 to prepare for weaning. Starter was offered after wk 4 but balanced between treatments. Body weight and frame were measured weekly with intakes and health monitored daily. At weaning, a subset of calves were slaughtered (n=6/diet). Enhanced-fed calves had greater carcass, thymus, liver, spleen, and mammary gland (parenchyma and mammary fat pad) weights. The EH calves also had greater average daily gain (ADG) starting during wk 1 (0.36 vs. -0.06kg/d) and lasting through wk 7 (1.00 vs. 0.41kg/d). Remaining calves received estrogen implants or placebo and were slaughtered at the end of wk 10, creating 4 treatments: (1) R, (2) R + estrogen (R-E2), (3) EH, and (4) EH + estrogen (EH-E2). Postweaning ADG was similar between R, EH, and EH-E2 calves, but greater in R-E2 calves than E calves. The EH-E2 calves had the heaviest mammary glands, and R-E2 calves had heavier mammary glands than R calves. The EH calves consumed more MR DM, CP, and fat preweaning. The R-fed calves consumed more starter DM preweaning. Fecal score was greater for EH calves (1.74 vs. 1.50) preweaning, but days medicated did not differ. Fecal scores were lower for R-E2 calves postweaning. Improved preweaning feeding of calves increased body weights and frame measures. Differences in body weights remained postweaning. Enhanced-fed calves showed greater ADG during the preweaning period but not postweaning. Exogenous estrogen may elicit diet-dependent growth responses. Analysis of collected samples will allow determination of cellular and molecular processes responsible for the marked differences in mammary development observed.

Short communication: Initial evidence supporting existence of potential rumen epidermal stem and progenitor cells.

The bovine rumen epidermis is a keratinized multilayered tissue that experiences persistent cell turnover. Because of this constant cell turnover, epidermal stem cells and their slightly more differentiated daughter cells, epidermal progenitor cells, must exist in the stratum basale of rumen epidermis. To date, these 2 epidermal cell populations and any unique cellular markers they may possess remain completely uncharacterized in the bovine rumen. An important first step in this new research area is the demonstration of the relative abundance and existence of markers for these cells in rumen tissue. A related second step is to document rumen epidermal proliferative responses to an extrinsic signal such as nutrient concentration within the rumen. The objectives of this experiment were to evaluate the extrinsic effect of diet on (1) gene expression of 6 potential rumen epidermal stem or progenitor cell markers and (2) rumen epidermal cell proliferation within the stratum basale. Twelve preweaned Holstein heifers were fed either a restricted diet (R) or an enhanced diet (EH). Animals on R received a milk replacer (MR) diet fed at 0.44kg of powder dry matter (DM)/d (20.9% crude protein, 29.8% fat, DM basis) and EH received MR at 1.08kg of powder dry matter/d (28.9% crude protein, 26.2% fat, DM basis). All calves had access to a 20% crude protein starter and were weaned during wk 7 of the experiment. Lifetime DM intake was 0.73kg of DM/calf per day for R (5.88 Mcal of net energy/calf per day) and 1.26kg of DM/calf per day for EH (10.68 Mcal of net energy/calf per day). Twenty-four hours before slaughter heifers received an intravenous dose of 5-bromo-2'-deoxyuridine to label proliferating cells. Heifers were slaughtered at 8 wk of age, and rumen samples from the ventral sac region were obtained and stored in RNA preservative and processed for routine histology. Quantitative real-time reverse transcriptase PCR was used to analyze relative abundance of genes. Candidate genes for markers of epidermal stem and progenitor cells were ß1-integrin (ITGB1), tumor protein p63 (TP63), keratin-14 (KRT14), Notch-1 (NOTCH1), Leu-rich repeat-containing G protein-coupled receptor 5-expressing (LGR5), and musashi-1 (MSI1). All genes were detected in the rumen tissue; ITGB1 was increased in EH compared with R. 5-Bromo-2'-deoxyuridine immunohistochemistry revealed that both R and EH rumen tissue had proliferating cells within the stratum basale of the rumen epidermis at the time of analysis. The EH diet resulted in an additive effect on cell proliferation. The percentage of cells in the stratum basale synthesizing DNA in preparation for mitosis nearly doubled (23.8±2.4% for EH vs. 14.7±2.0% for R) compared with calves fed R. This work represents the first attempt at characterizing rumen epidermal stem and progenitor cells. We demonstrated the relative abundance and existence of potential markers in rumen tissue and showed a rumen epidermal proliferative response to the extrinsic stimulus of nutrient concentration in the form of diet.

Short communication: Effects of increasing protein and energy in the milk replacer with or without direct-fed microbial supplementation on growth and performance of preweaned Holstein calves.

Forty-four Holstein calves were fed a direct-fed microbial (DFM) and 1 of 2 milk replacers to evaluate calf performance and growth. Treatments were (1) a control milk replacer [22:20; 22% crude protein (CP) and 20% fat], (2) an accelerated milk replacer (27:10; 27% CP and 10% fat), (3) the control milk replacer with added DFM (22:20+D), and (4) the accelerated milk replacer with added DFM (27:10+D). Dry matter intake, rectal temperatures, respiration scores and rates, and fecal scores were collected daily. Body weight, hip and withers height, heart girth, blood, and rumen fluid samples were collected weekly. Effects of treatment, sex, week, and their interactions were analyzed. Calves fed an accelerated milk replacer, regardless of DFM supplementation, consumed more CP and metabolizable energy in the milk replacer. No treatment differences were found for starter intake or intake of neutral detergent fiber or acid detergent fiber in the starter. Calves fed the accelerated milk replacer had greater preweaning and weaning body weight compared with calves fed the control milk replacer. Average daily gain was greater during the preweaning period for calves fed the accelerated milk replacer, but the same pattern did not hold true during the postweaning period. Feed efficiency did not differ among treatments. Hip height tended to be and withers height and heart girth were greater at weaning for calves fed the accelerated milk replacer compared with calves fed the control milk replacer. Fecal scores were greatest in calves fed DFM. Overall acetate, propionate, butyrate, and n-valerate concentrations were lower in calves fed the accelerated milk replacer, but DFM did not have an effect. Rumen pH was not different. Blood metabolites were unaffected by DFM supplementation, but calves fed the accelerated milk replacer had increased partial pressure of CO2, bicarbonate, and total bicarbonate in the blood. Direct-fed microbial supplementation did not appear to benefit the calf in this trial.

A model of ischemic isolated acute liver failure in pigs: standardizing monitoring and treatment.

BACKGROUND: Acute liver failure (ALF) models in pigs have been widely used for evaluating newly developed liver support systems. But hardly any guidelines are available for the surgical methods and the clinical management. METHODS: The study validated several standard operating procedures describing in detail the surgical method and intensive care monitoring and treatment (control of potassium, glucose and bicarbonate levels, cardiovascular and intracranial pressure monitoring, etc.). ALF was induced in animals with a mean of 56 kg. Two surgical methods were compared: ligation of hepatic arteries with either end-to-side portacaval shunt (ESPS) and bile duct ligation or side-to-side portacaval shunt (SSPS) without bile duct ligation. RESULTS: During total portal vein clamping, the animals in the ESPS group developed severe hypotension, splanchnic congestion and metabolic acidosis. One animal died after approximately 1.5 h. This model therefore represents a multiorgan failure model rather than an isolated ALF model. In the SSPS group, none of these side effects were observed, while clinical, laboratory and histopathological signs of ALF were evident. CONCLUSIONS: A reproducible model in pigs representing ALF can be established with the help of the standardized monitoring and treatment procedures presented.

Review: The pre-pubertal bovine mammary gland: unlocking the potential of the future herd.

Historically, pre-pubertal development of the bovine mammary gland (MG) has received little attention compared to later development. Recent evidence suggests not only that this period represents a very active time in the development of the MG but also that the first 90 days of life can partially dictate future productivity of the lactating cow. The MG, often considered quiescent during early life (first 3 months), is now known to increase in size by over 60-fold in the same period. The importance of sex steroids in MG development is well classified, but a complex signaling network exists among estrogen, progesterone and other growth factors and hormones. Complicating our understanding of this developmental period further is the discovery that pre-weaning nutrition of the calf not only influences the growth of the mammary parenchyma but may also alter the way in which it responds to mammogenic stimuli. Recent data suggest that feeding calves a higher plane of nutrition improves the ability of the mammary epithelium to respond to estradiol and also alters the way in which the mammary parenchyma and fat pad communicate. It is clear that early life nutrition, although able to influence the MG, is still poorly understood mechanistically. For example, additional evidence suggests that increased feeding rates in early life alter the morphology of myoepithelial cells in the mammary epithelium. Further data have also suggested a role for other cell types, such as immune cells, in the penetration of the mammary parenchyma into the fat pad during the early life development of the MG suggesting that mammary development is not only controlled by the local tissue population (parenchyma and fat pad) but perhaps systemically by other tissue types (i.e., immune system). Understanding the roles of these various stimuli and signaling pathways as they relate to the development of the MG in early life may hold the key to unlocking the potential for the optimal development of this crucial organ and, in turn, may lead to improvements in other phases of mammary development and milk yield potential.

Long-term selection of chickens for body weight alters muscle satellite cell behaviors.

Muscle satellite cells (SC) are resident stem-like cells that play an integral role in skeletal muscle growth and repair. Understanding how SC maintain their identities and dynamic properties is critical to animal growth. However, the genetic and environmental factors governing SC behaviors and the underpinning mechanisms remain unknown. To explore whether genetic selection influences SC behaviors, we used 2 lines of chickens selected for over 50 generations with over a 10-fold difference in body weight at 56 d of age-the Virginia high weight selection (HWS) and low weight selection (LWS) lines. To study these 2 lines, we performed both in vivo and in vitro experiments. In vivo, we studied the abundance of SC in normal physiological settings and tested their functional roles in muscle regeneration using a muscle injury model. In vitro, we isolated SC from chicken skeletal muscle and assayed their ability to proliferate and differentiate under cultured conditions. Immunohistochemical staining of breast muscle (pectoralis major) revealed that muscle fibers from HWS chickens possessed more SC than those from LWS. Further analysis showed that the SC pool from HWS muscles contained a higher percentage of activated SC compared to that of LWS. When isolated SC from HWS and LWS muscles were cultured, HWS SC exhibited greater abilities to proliferate and differentiate than those SC from LWS. To test whether the observed in vitro differences in SC properties could be confirmed in vivo, we subjected chicken breast muscle to barium chloride to induce muscle injury and regeneration. Consistent with in vitro data, breast muscle in HWS chicken experienced a faster and more robust recovery than that of LWS, as evidenced by quicker regeneration and larger muscle fiber size. Taken together, these findings suggest divergent selection for body weight not only results in correlated responses in SC number, but also changes SC growth kinetics. Further dissection of the molecular mechanism will aid the identification of the target molecules for growth intervention in chickens.

Satellite cell-mediated breast muscle regeneration decreases with broiler size.

Satellite cells (SCs) reside between the sarcolemma and basal lamina of muscle fibers and are the primary contributor of DNA for post-hatch muscle growth and repair. Alterations in SC content or properties by intrinsic and extrinsic factors can have detrimental effects on muscle health and function, and ultimately meat quality. We hypothesized that disrupted SC homeostasis may account in part for the increased breast myopathies observed in growing broilers. To test this hypothesis, we selected broilers with different body weights at comparable ages and studied SC characteristics in vitro and in vivo. Data shows that SC numbers in the breast muscles decrease (P < 0.001) and their inherent abilities to proliferate and differentiate diminish (P < 0.001) with age and size. Further, when breast muscle is presented with an insult, muscle of larger broilers regenerates more slowly than their smaller, age-matched counterparts arguing that SC quality changes with size and age. Together, our studies show that birds with greater muscle hypertrophy have less SCs with diminished ability to function, and suggest that aggressive selection for breast growth in broilers may exhaust SC pools when birds are grown to heavier processing weights. These findings provide new insights into a possible mechanism leading to breast myopathies in the poultry industry and provide targets for mitigating adverse fresh breast quality.

Characterization of an enhancer element in the proximal promoter of the mouse glucagon receptor gene.

The 5'-flanking region of the mouse glucagon receptor has been previously cloned and two promoter regions were characterized. Functional analysis of the proximal promoter was now performed to characterize cis-acting element(s) regulating basal gene expression. Promoter analysis using deletion constructs in a rat cell line (CA-77) expressing the glucagon receptor, showed that the region from -64 to +127 relative to the proximal transcription start site was sufficient for maximal proximal promoter activity. A DNA sequence spanning the -28 to -16 region organized as an imperfect palindrome was demonstrated to be functional as a cis-acting enhancer. Constructs including several copies of this motif strongly increased activity of the heterologous thymidine kinase promoter. Gel mobility shift assays performed with different DNA fragments spanning this region confirmed that it specifically bound nuclear protein(s) from CA-77 cells, mouse MIN-6 cells or mouse liver. Mutations in the core sequence of this site impaired both reporter gene activity and nuclear protein binding. The palindrome is a novel DNA sequence with no homology to existing transcription factor binding site database. This is the first characterization of a functional cis-acting sequence into the proximal promoter of the mouse glucagon receptor that may support constitutive expression of the gene.

An examination of the working case definition of chronic fatigue syndrome.

PURPOSE: Chronic fatigue syndrome (CFS) currently is defined by a working case definition developed under the leadership of the United States Centers for Disease Control and Prevention (CDC) based on a consensus among experienced clinicians. We analyzed the experience from one large center to examine the adequacy of the case definition. PATIENTS AND METHODS: Predefined clinical and laboratory data were collected prospectively from 369 patients with debilitating fatigue, of whom 281 (76%) met the major criteria of the original CDC case definition for CFS: (1) fatigue of at least 6 months' duration, seriously interfering with the patient's life; and (2) without evidence of various organic or psychiatric illnesses that can produce chronic fatigue. The same clinical data were obtained from 311 healthy control subjects and two comparison groups with diseases that can present in a similar fashion; relapsing-remitting multiple sclerosis (n = 25) and major depression (n = 19). RESULTS: All of the minor criteria symptoms from the original CDC case definition distinguished patients with debilitating chronic fatigue from healthy control subjects, and many distinguished the patients with chronic fatigue from the comparison groups with multiple sclerosis and depression: myalgias, postexertional malaise, headaches, and a group of infectious-type symptoms (ie, chronic fever and chills, sore throat, swollen glands in the neck or underarm areas). In addition, two other symptoms not currently part of the case definition discriminated the chronic fatigue patients from the control/comparison groups: anorexia and nausea. Physical examination criteria only infrequently contributed to the diagnosis. Patients meeting the CDC major criteria for CFS also met the minor criteria in 91% of cases. CONCLUSION: Patients meeting the major criteria of the current CDC working case definition of CFS reported symptoms that were clearly distinguishable from the experience of healthy control subjects and from disease comparison groups with multiple sclerosis and depression. Eliminating three symptoms (ie, muscle weakness, arthralgias, and sleep disturbance) and adding two others (ie, anorexia and nausea) would appear to strengthen the CDC case definition of CFS.

Effects of venlafaxine on extracellular concentrations of 5-HT and noradrenaline in the rat frontal cortex: augmentation via 5-HT1A receptor antagonism.

Venlafaxine is a novel serotonin/noradrenaline reuptake inhibitor (SNRI) which has been shown clinically to be an effective antidepressant (AD) with a faster onset of action than serotonin specific reuptake inhibitors (SSRI). Preclinically, venlafaxine has been shown to potently inhibit dorsal raphe neuronal (DRN) firing through a 5-HT1A mediated mechanism, in a similar manner to SSRIs. Here we demonstrate the acute neurochemical effects of venlafaxine on extracellular concentrations of 5-HT and noradrenaline (NA) from the rat frontal cortex using in vivo microdialysis. Administration of venlafaxine (3-50 mg/kg s.c.) resulted in a significant dose-dependent increase in extracellular NA, but produced no significant increase in 5-HT concentrations. Combination treatment with the selective 5-HT1A antagonist WAY100635 produced a dose-dependent augmentation of venlafaxine-induced (3-30 mg/kg s.c) extracellular 5-HT concentrations, but had no further effect on NA above that produced by venlafaxine alone. WAY100635, at doses as low as 0.03 mg/kg s.c., maintained this potentiation effect. The beta-adrenergic/5-HT1A receptor antagonist (+/-)pindolol and the selective 5-HT1B/D antagonist GR127935 produced no significant augmentation of venlafaxine-induced changes in either 5-HT or NA. Using the alpha1 and alpha2-adrenoceptor antagonists, prazosin and idazoxane, we also demonstrate the role of the alpha-adrenoceptors in the augmentation of venlafaxine-induced changes. The possible mechanisms underlying venlafaxines improved clinical AD action and the potential for further enhancement of this SNRIs clinical effects are discussed.

Inhibition of protein synthesis by didemnins: cell potency and SAR.

Synthetic and naturally occurring didemnins are potent and specific inhibitors of protein synthesis in vitro. Structure-activity analysis indicates a requirement for the intact macrocycle; however, the smaller ring size represented by the didemnin analogue, tamandarin A, is equipotent to didemnin B. Replacement of the N,O-dimethyltyrosine by a N-methylphenylalanine or N-methylleucine residue is also well-tolerated. The rank order for inhibition of protein synthesis in vitro appears to be retained in MCF-7 cells, albeit at much higher potency. This increase in potency is explained for the first time by data indicating that MCF-7 cells can accumulate didemnin B up to 2-3 orders of magnitude compared to the growth medium.

Vaginal birth after cesarean delivery: an admission scoring system.

OBJECTIVE: To develop a scoring system to predict the likelihood of vaginal birth in patients undergoing a trial of labor after previous cesarean delivery using factors known at the time of hospital admission. METHODS: Trial of labor was attempted in 5022 patients who were assigned randomly to score derivation and score testing groups. Multivariate logistic regression modeling was used in the score derivation group to develop a predictive scoring system for vaginal birth. The scoring system was then applied to the testing group to evaluate its predictive ability. RESULTS: Five variables significantly affected the mode of birth and were incorporated into a weighted scoring system. Rates of successful vaginal birth after cesarean ranged from 49% in patients scoring 0-2 to 95% in patients scoring 8-10. Increasing score was associated linearly with increasing probability of vaginal birth after cesarean. CONCLUSION: Increasing scores correlate with increasing probability of vaginal birth after cesarean. The admission vaginal birth after cesarean scoring system may be useful in counseling patients regarding the option of vaginal birth or repeat cesarean delivery. This information could be particularly valuable for the patient who opts for trial of labor but has second thoughts about her mode of birth when labor begins.