Whey protein plus bicarbonate supplement has little effects on structural atrophy and proteolysis marker immunopatterns in skeletal muscle disuse during 21 days of bed rest.

OBJECTIVES: To investigate the effect of whey protein plus potassium bicarbonate supplement on disused skeletal muscle structure and proteolysis after bed rest (BR). METHODS: Soleus (SOL) and vastus lateralis (VL) biopsies were sampled from ten (n=10) healthy male subjects (aged 31±6 years) who did BR once with and once without protein supplement as a dietary countermeasure (cross-over study design). The structural changes (myofibre size and type distribution) were analysed by histological sections, and muscle protein breakdown indirectly via the proteolysis markers, calpain 1 and 3, calpastatin, MuRF1 and 2, both in muscle homogenates and by immunohistochemistry. RESULTS: BR caused size-changes in myofiber cross-sectional area (FCSA, SOL, p=0,004; VL, p=0.03), and myofiber slow-to-fast type transition with increased hybrids (SOL, p=0.043; VL, p=0.037) however with campaign differences in SOL (p<0.033). No significant effect of BR and supplement was found by any of the key proteolysis markers. CONCLUSIONS: Campaign differences in structural muscle adaptation may be an issue in cross-over design BR studies. The whey protein plus potassium bicarbonate supplement did not attenuate atrophy and fibre type transition during medium term bed rest. Alkaline whey protein supplements may however be beneficial as adjuncts to exercise countermeasures in disuse.

Bone and muscle structure and quality preserved by active versus passive muscle exercise on a new stepper device in 21 days tail-suspended rats.

Human performance in microgravity is characterized by reversed skeletal muscle actions in terms of active vs. passive mode contractions of agonist/antagonist groups that may challenge principal biodynamics (biomechanical forces translated from muscle to bone) of the skeletal muscle-bone unit. We investigated active vs. passive muscle motions of the unloaded hindlimb skeletal muscle-bone unit in the 21 days tail-suspended (TS) rat using a newly designed stepper exercise device. The regimen included both active mode motions (TSA) and passive mode motions (TSP). A TS-only group and a normal cage group (CON) served as positive or negative controls. The muscle and bone decrements observed in TS-only group were not seen in the other groups except TSP. Active mode motions supported femur and tibia bone quality (5% BMD, 10% microtrabecular BV/TV, Tb.Th., Tb.N. parameters), whole soleus muscle/myofiber size and type II distribution, 20% increased sarcolemma NOS1 immunosignals vs. CON, with 25% more hybrid fiber formation (remodeling sign) for all TS groups. We propose a new custom-made stepper device to be used in the TS rat model that allows for detailed investigations of the unique biodynamic properties of the muscle-bone unit during resistive-load exercise countermeasure trials on the ground or in microgravity.

The 2nd Berlin BedRest Study: protocol and implementation.

Long-term bed-rest is used to simulate the effect of spaceflight on the human body and test different kinds of countermeasures. The 2nd Berlin BedRest Study (BBR2-2) tested the efficacy of whole-body vibration in addition to high-load resisitance exercise in preventing bone loss during bed-rest. Here we present the protocol of the study and discuss its implementation. Twenty-four male subjects underwent 60-days of six-degree head down tilt bed-rest and were randomised to an inactive control group (CTR), a high-load resistive exercise group (RE) or a high-load resistive exercise with whole-body vibration group (RVE). Subsequent to events in the course of the study (e.g. subject withdrawal), 9 subjects participated in the CTR-group, 7 in the RVE-group and 8 (7 beyond bed-rest day-30) in the RE-group. Fluid intake, urine output and axiallary temperature increased during bed-rest (p < .0001), though similarly in all groups (p > or = .17). Body weight changes differed between groups (p < .0001) with decreases in the CTR-group, marginal decreases in the RE-group and the RVE-group displaying significant decreases in body-weight beyond bed-rest day-51 only. In light of events and experiences of the current study, recommendations on various aspects of bed-rest methodology are also discussed.

Molecular biomarkers monitoring human skeletal muscle fibres and microvasculature following long-term bed rest with and without countermeasures.

The cellular mechanisms of human skeletal muscle adaptation to disuse are largely unknown. The aim of this study was to determine the morphological and biochemical changes of the lower limb soleus and vastus lateralis muscles following 60 days of head-down tilt bed rest in women with and without exercise countermeasure using molecular biomarkers monitoring functional cell compartments. Muscle biopsies were taken before (pre) and after bed rest (post) from a bed rest-only and a bed rest exercise group (n = 8, each). NOS1 and NOS3/PECAM, markers of myofibre 'activity' and capillary density, and MuRF1 (E3 ubiquitin-ligase), a marker of proteolysis, were documented by confocal immunofluorescence and immunoblot analyses. Morphometrical parameters (myofibre cross-sectional area, type I/II distribution) were largely preserved in muscles from the exercise group with a robust trend for type II hypertrophy in vastus lateralis. In the bed rest-only group, the relative NOS1 immunostaining intensity was decreased at type I and II myofibre membranes, while the bed rest plus exercise group compensated for this loss particularly in soleus. In the microvascular network, NOS3 expression and the capillary-to-fibre ratio were both increased in the exercise group. Elevated MuRF1 immunosignals found in subgroups of atrophic myofibres probably reflected accelerated proteolysis. Immunoblots revealed overexpression of the MuRF1 protein in the soleus of the bed rest-only group (> 35% vs. pre). We conclude that exercise countermeasure during bed rest affected both NOS/NO signalling and proteolysis in female skeletal muscle. Maintenance of NO signalling mechanisms and normal protein turnover by exercise countermeasure may be crucial steps to attenuate human skeletal muscle atrophy and to maintain cell function following chronic disuse.

Homer proteins and InsP(3) receptors co-localise in the longitudinal sarcoplasmic reticulum of skeletal muscle fibres.

Striated muscle represents one of the best models for studies on Ca(2+) signalling. However, although much is known on the localisation and molecular interactions of the ryanodine receptors (RyRs), far less is known on the localisation and on the molecular interactions of the inositol trisphosphate receptors (InsP(3)Rs) in striated muscle cells. Recently, members of the Homer protein family have been shown to cluster type 1 metabotropic glutamate receptors (mGluR1) in the plasma membrane and to interact with InsP(3)R in the endoplasmic reticulum of neurons. Thus, these scaffolding proteins are good candidates for organising plasma membrane receptors and intracellular effector proteins in signalosomes involved in intracellular Ca(2+) signalling. Homer proteins are also expressed in skeletal muscle, and the type 1 ryanodine receptor (RyR1) contains a specific Homer-binding motif. We report here on the relative sub-cellular localisation of InsP(3)Rs and Homer proteins in skeletal muscle cells with respect to the localisation of RyRs. Immunofluorescence analysis showed that both Homer and InsP(3)R proteins present a staining pattern indicative of a localisation at the Z-line, clearly distinct from that of RyR1. Consistent herewith, in sub-cellular fractionation experiments, Homer proteins and InsP(3)R were both found in the fractions enriched in longitudinal sarcoplasmic reticulum (LSR) but not in fractions of terminal cisternae that are enriched in RyRs. Thus, in skeletal muscle, Homer proteins may play a role in the organisation of a second Ca(2+) signalling compartment containing the InsP(3)R, but are apparently not involved in the organisation of RyRs at triads.

The olfactory adenylyl cyclase III is expressed in rat germ cells during spermiogenesis.

To identify the adenylyl cyclase (AC) genes expressed in mammalian germ cells, RT-PCR of testis and germ cell RNA was performed using degenerated primers based on the homologous region of the AC catalytic domain. This strategy yielded high-frequency amplification of a complementary DNA (cDNA) identical to type III AC (ACIII), a form previously identified as the major adenylyl cyclase expressed in the olfactory system. Ribonuclease protection studies confirmed that ACIII transcripts are present in germ cells, appear during the meiotic prophase, and accumulate during spermiogenesis. A Northern blot analysis performed on total testis RNA demonstrated the presence of a predominant transcript of 7.5 kb, suggesting that the ACIII expressed in germ cells may derive from a splicing variant different from the 4.5 kb transcripts expressed in somatic cells. To determine whether these RNAs are translated into a protein, Western blot analysis was performed using an antibody specific for the carboxyl terminus of ACIII. An immunoreactive protein of 170 kDa was detected in extracts from total testis and from germ cells. Immunofluorescence localization of this protein in the seminiferous tubules showed that ACIII was predominantly expressed in postmeiotic germ cells from round spermatids in the cap phase to maturing elongating spermatids. The ACIII antigen was located mostly on the acrosomal membrane rather than on the plasma membrane of developing spermatids. The spatial and temporal expression of ACIII in germ cells indicates a role of this AC in the acrosome formation. Together with the observation that members of the olfactory receptor family and an olfactory phosphodiesterase are expressed in spermatids, these findings suggest that a signal transduction system used in olfaction is also used during gamete development.

DEFT, a novel death effector domain-containing molecule predominantly expressed in testicular germ cells.

Apoptosis is a physiological process by which multicellular organisms eliminate unwanted cells. Death factors such as Fas ligand induce apoptosis by triggering a series of intracellular protein-protein interactions mediated by defined motifs found in the signaling molecules. One of these motifs is the death effector domain (DED), a stretch of about 80 amino acids that is shared by adaptors, regulators, and executors of the death factor pathway. We have identified the human and rat complementary DNAs encoding a novel protein termed DEFT (Death EFfector domain-containing Testicular molecule). The N-terminus of DEFT shows a high degree of homology to the DEDs found in FADD (an adaptor molecule) as well as procaspase-8/FLICE and procaspase-10/Mch4 (executors of the death program). Northern blot hybridization experiments have shown that the DEFT messenger RNA (mRNA) is expressed in a variety of human and rat tissues, with particularly abundant expression in the testis. In situ hybridization analysis further indicated the expression of DEFT mRNA in meiotic male germ cells. In a model of germ cell apoptosis induction, an increase in testis DEFT mRNA was found in immature rats after 2 days of treatment with a GnRH antagonist. Unlike FADD and procaspase-8/FLICE, overexpression of DEFT did not induce apoptosis in Chinese hamster ovary cells. Although cotransfection studies indicated that DEFT is incapable of modulating apoptosis effected by FADD and procaspase-8/FLICE, interactions between DEFT and uncharacterized DED-containing molecules in the testis remain to be studied in the future. In conclusion, we have identified a novel DED-containing protein with high expression in testis germ cells. This protein may be important in the regulation of death factor-induced apoptosis in the testis and other tissues.

Type 4 cyclic adenosine monophosphate-specific phosphodiesterases are expressed in discrete subcellular compartments during rat spermiogenesis.

The type 4 cAMP-specific phosphodiesterases (PDE4) are a family of closely related enzymes with similar catalytic domains and divergent amino- and carboxyl-terminus domains. Multiple PDE proteins with heterogeneous amino termini are derived from each gene. To understand the significance of this heterogeneity, the expression and localization of variants derived from PDE4A and PDE4D genes was investigated during spermatogenesis in the rat. RNase protection analysis with mRNA for testes at different ages of development showed that two transcripts (PDE4D1 and PDE4D2) are expressed at day 10 and 15 of age and become undetectable thereafter. An additional PDE4D transcript appears at day 30 and increased during testid maturation. This latter transcript codes for a long variant of the PDE4D gene and is expressed in germ cells as demonstrated by RNase protection with RNA from isolated pachytene spermatocytes and round spermatids. The presence of a corresponding PDE4D protein with a molecular mass of 98 kDa was established by immunoprecipitation and Western blot analysis with antibodies specific for PDE4D and by immunoaffinity chromatography purification of the 98 kDa variant from isolated germ cells. PDE4A transcripts were also expressed in pachytene spermatocytes and round spermatids. Two polypeptides encoded by these PDE4A transcripts were expressed in pachytene spermatocytes, reached a maximum in round spermatids, and declined thereafter. Immunofluorescence analysis demonstrated a localization of the PDE4D protein in the manchette and in a periacrosomal region of the developing spermatid, a localization confirmed by immunogold electron microscopy. Conversely, the PDE4A was mostly soluble in the cytoplasm of round spermatids. These data demonstrate that PDE4D and PDE4A variants are expressed at different stages and localized in distinct subcellular structures of developing spermatids. Different properties of the mRNAs derived from the two genes and localization signals are responsible for the temporal and spatial expression of the different PDE4 isoenzymes.

Efficacy of dietary and injected vitamin E for poults.

An experiment was conducted to compare the efficacy of two dietary sources and an injectable form of vitamin E (VE) to improve the VE status of poults. Six of the treatments consisted of a factorial arrangement of three concentrations and two sources of dietary VE. Turkeys in these treatments received 12, 80, or 150 IU of either dl-alpha-tocopheryl acetate or d-alpha-tocopherol (d-alpha-TOC)/kg of diet. The seventh treatment consisted of a single subcutaneous injection of d-alpha-TOC at 1 d of age. Poults in this treatment were subcutaneously injected in the dorsal area of the neck with 25 IU of d-alpha-TOC, this amount being approximately equivalent to the amount poults would consume if their diet was supplemented with 150 IU of VE/kg during their 1st wk of life. Concentration, source, or route of VE administration did not affect growth parameters, plasma creatine kinase, plasma triglycerides, or liver lipid peroxidation as measured by the thiobarbituric acid reactive substances assay (TBARS). Plasma, red blood cells (RBC), and liver alpha-TOC decreased from hatching to 14 d of age in poults fed either source of VE. The use of 80 or 150 IU of dietary VE (either source) reduced (P < 0.05) the extent of depletion of alpha-TOC at all ages and also reduced the susceptibility of RBC to hemolysis. There was no effect of source of dietary VE on concentration of alpha-TOC in plasma, RBC, or liver, or on RBC hemolysis. Subcutaneous injection of 25 IU of d-alpha-TOC at Day 1 increased (P < 0.05) alpha-TOC concentration until 7 d of age. Also, d-alpha-TOC injection reduced (P < 0.05) RBC susceptibility to hemolysis through 21 d of age. Data showed that one single subcutaneous injection of 25 IU of d-alpha-TOC at 1 d of age was as effective as 80 IU or more of dietary VE through 21 d to improve the alpha-TOC status of poults.

Influence of supplemental dietary fat on changes in vitamin E concentration in livers of poults.

An experiment was conducted to determine the influence of supplemental dietary fat on alpha-tocopherol (TOC) stored in the livers of young turkeys during the first 21 d after hatching. The four dietary treatments were obtained by supplementing a corn-soybean meal diet with 8% sucrose (SUC), 8% animal-vegetable fat (AVF), 8% tallow (TAL), or 8% coconut oil (COC). All diets were supplemented with 12 IU of dl-alpha-tocopheryl acetate (vitamin E)/kg of diet. Body weight at 21 d of age was not affected by dietary fat, whereas feed efficiency was improved (P < .05) by added fat, irrespective of source. Liver TOC (micrograms per gram of liver and micrograms per total liver weight) decreased markedly between 1 to 14 d of age, irrespective of fat source. Average TOC concentration in liver was 78.9 micrograms/g at 1 d, but was only .5 microgram/g at 14 d. Between 14 and 21 d of age, total liver TOC increased slightly in all treatment groups. No diet effect was observed on the liver TOC concentration until 21 d of age. At this time, poults fed TAL had less (P < .05) TOC in liver than those fed COC and AVF. The data show that neither the presence of supplemental dietary fat nor fat source changed the pattern of marked decrease in liver TOC during the first 14 d after hatching.

Influence of supplementing corn-soybean meal diets with vitamin E on performance and selected physiological traits of male turkeys.

Three experiments were conducted to determine the effects of supplementing practical diets of male turkeys with dl-alpha-tocopheryl acetate (TA). In Experiment 1, a factorial arrangement of dietary treatments [0, 12, 50, 150, and 300 IU TA/kg with 0 or 300 mg ascorbic acid (AA)/kg] was used. These 10 treatments were fed to poults from 1 to 41 d of age. From 41 to 118 d of age, the AA treatments were discontinued, and the 300 IU TA treatment groups were changed to 12 IU TA/kg. Neither TA nor AA treatments affected 41-d BW, feed to gain ratio (FE), or livability. No effects of dietary TA concentrations on turkey performance were observed through 118 d of age alpha-Tocopherol (TOC) concentrations of plasmas and livers were increased by increments of dietary TA, with substantial liver storage when toms were fed 150 IU TA/kg from 1 to 118 d. Supplementing diets with 0, 25, 50, 75, or 100 IU TA/ kg in Experiments 2 and 3 had no effect on performance of toms through 119 and 105 d, respectively. alpha-Tocopherol concentrations of plasma and red blood cells (RBC) increased linearly with increments of dietary TA. The same was true for livers in Experiment 2. Susceptibility of RBC to hemolysis induced by 400 microM t-butyl hydroperoxide (TBH) in Experiment 2 decreased with increasing dietary TA, and these decreases corresponded to increases in TOC concentration of RBC. However, the relationships between hemolysis and dietary TA or RBC TOC were inconsistent in Experiment 3 and varied according to concentration of TBH (200, 300, or 400 microM) and age of the toms. At 105 d of age, RBC of toms fed no supplemental TA were resistant to hemolysis, irrespective of dietary TA and TBH concentration. In Experiment 3, there were no indications of dietary TA effects on plasma peroxide concentration or activity of plasma creatine kinase. A positive relationship between dietary TA and blastogenic responses of blood lymphocytes was observed with concanavalin A when toms were at 44 d but not at 23 or 86 d of age. The overall data indicate that corn-soybean meal diets containing from 6 to 20 IU TOC/kg, but no supplemental TA supported satisfactory performance and well-being of male turkeys from 1 d of age to market ages when the turkeys were free of disease, as was true in the research reported here.

Effect of early nutrient restriction on broiler chickens. 2. Performance and digestive enzyme activities.

An experiment was conducted to determine the effect of two early nutrient restriction programs on performance, selected characteristics of the gastrointestinal tract (GIT), and activities of digestive enzymes of broiler chickens. Three hundred and sixty male broiler (Ross x Ross) chicks kept in floor pens were assigned to three groups. The control group (C) was given ad libitum access to feed from 1 to 48 d of age. Another group was restricted from 11 to 14 d (R4) of age to an energy intake of .74 x BW.67 kcal ME/d, and a third group was restricted from 7 to 14 d (R7) of age to an energy intake of 1.5 x BW.67 kcal ME/d. Then, both restricted groups were given ad libitum access to feed through 48 d. Body weight and feed intake were determined weekly and selected carcass characteristics were measured at 48 d of age. Broilers also were sampled at 7, 14, 21, and 42 d of age to obtain data on components of the GIT (proventriculus, gizzard, pancreas, and small intestine) and activities of selected digestive enzymes. Feed-restricted groups were lighter in body weight (P < .01) at 14 and 48 d of age than the C group but were superior in overall feed efficiency. No treatment effects were observed for percentage yields of breast meat and abdominal fat pad. Absolute weights of GIT components were significantly reduced at 14 d of age by feed restriction. However, GIT components increased in weight more quickly after refeeding than did the whole body. Restricted groups had reduced (P < .01) specific activities of jejunal alkaline phosphatase and pancreatic trypsin, amylase, and lipase as compared with the C group at 14 d of age but not at 21 and 42 d of age. Relative activities for jejunal maltase and sucrase were greater (P < .01) at 21 d of age in the R4 and R7 groups than in the C group. The present data show that feed restriction results in transient changes in organs and activities of digestive enzymes, suggesting a functional adaptation to feed restriction.

Effect of early nutrient restriction on broiler chickens. 1. Performance and development of the gastrointestinal tract.

An experiment was conducted to determine the effects of early nutrient restriction on performance and development of the gastrointestinal tract of broiler chickens. Four hundred male broiler (Ross x Ross) chicks raised in floor pens were assigned to two treatment groups. One group was given ad libitum access to feed from 1 to 48 d of age. The second group was feed restricted from 7 to 14 d of age to an energy intake of 1.5 x BW.67 kcal ME/d and then given ad libitum access to feed from 14 to 48 d. Body weight and feed intake were determined weekly. At 49 d of age, birds were processed for carcass yield, abdominal fat pad measurement, and body composition analysis. Broilers were also sampled at 7, 14, 21, and 41 d of age for proventriculus, gizzard, small intestine (duodenum, jejunum, and ileum), pancreas, and liver weights and for intestinal length measurements. Total DNA, protein:DNA, and RNA:DNA ratios of livers and jejuna were determined as indexes of changes in cell size and number. Feed-restricted broilers failed to catch up to the Control birds in BW at 48 d of age but were superior (P < .01) in overall feed efficiency. No treatment effects were observed on breast meat yields or abdominal fat. Moreover, percentage carcass fat, crude protein, ash, and dry matter were not affected by restricted feeding. Body weight and weights of gastrointestinal organs were reduced (P < .01) by feed restriction at 14 d of age. Restricted feeding, however, did not decrease the relative weights of organs, except for liver. Feed restriction also resulted in a reduction (P < .01) of liver cell number and size and a decrease in jejunum cell number. All organs recovered normal weight on refeeding, and all cellular constituent ratios (e.g., RNA:DNA, RNA:protein, and protein:DNA) returned to normal by 41 d of age. Absolute and relative weights of supply organs (e.g., proventriculus, gizzard, small intestine, liver, and pancreas) were less affected by feed restriction and responded more quickly to refeeding than the whole body.

Posthatching changes in the immunoglobulin A concentration in the jejunum and bile of turkeys.

An experiment was conducted to document the age-related changes in IgA concentration in the small intestine of newly hatched turkey poults reared in floor pens and to determine whether infection with stunting syndrome (SS) affects age-related changes. Day-old turkey poults were dose per os with .5 mL of saline carrier (control) or with .5 mL of one of two dilutions (250- or 2.5 x 10(6)-fold) of a "crude" SS-causing inoculum. Inoculation with the 250-fold dilution depressed body weight gain (P less than .01) throughout the experiment and impaired feed efficiency (P less than .05) at 5 and 9 days of age as compared with the control group. After 9 days of age, all inoculated poults utilized feed more efficiently than did control poults (P less than .01). Stunting syndrome did not affect IgA concentrations in either bile or jejunum at any specific age. Age-related changes in IgA concentrations, however, were observed. Bile IgA decreased from 1 to 9 days of age, and then increased until 29 days of age. The IgA concentration in jejunal tissue increased linearly from 1 to 29 days of age (P less than .01), whether expressed as IgA concentration per gram of wet tissue or as percentage of total protein in jejunum. Age-related changes in IgA concentration in both bile and jejunum suggest that the secretory immune system associated with the digestive mucosa is not fully developed at the time of hatch.

Research note: vitamin E status of turkey poults as influenced by different dietary vitamin E sources, a bile salt, and an antioxidant.

An experiment was conducted to determine the effects of level and chemical form of dietary vitamin E on alpha-tocopherol status of poults. The effects of a dietary bile salt and an antioxidant on concentrations of alpha-tocopherol in serum and liver were also tested. Six dietary treatments were obtained by supplementing a corn-soybean meal diet with 12 IU of DL-alpha-tocopheryl acetate (TA)/kg (LE), 12 IU of TA plus 800 mg of sodium taurocholate/kg (LB), 12 IU of TA plus 500 mg of ethoxyquin/kg (LS), 12 IU of D-alpha-tocopheryl polyethylene glycol 1,000 succinate (TPGS)/kg (LT), 100 IU of TA/kg (HE), and 100 IU of TPGS/kg (HT). Growth rate and feed efficiency of poults were unaffected (P > .05) by dietary treatments. The HE diet increased alpha-tocopherol in liver (P < .01) at 14 and 21 days of age. Liver and serum alpha-tocopherol concentrations were unaffected by dietary TPGS (LT and HT diets) at any age. Serum alpha-tocopherol concentration was unaffected by dietary treatments at 5 days of age. The HE diet, however, increased (P < .01) serum alpha-tocopherol at 9, 14, and 21 days of age. Age-related changes in alpha-tocopherol concentration were observed. Both liver and serum alpha-tocopherol decreased markedly from 1 to 14 days of age. The HE diet only partly alleviated the reduction of alpha-tocopherol in liver and serum. The water-soluble form of vitamin E, TPGS, dietary sodium taurocholate, or dietary ethoxyquin, did not prevent the marked decline in alpha-tocopherol concentration of liver and serum during the 21-day experiment.

Dietary effects on stunting syndrome in poults.

Experiments were done to determine the effect of feeding diets of different ingredient composition to poults experimentally infected with stunting syndrome (SS) at 1 day of age. In Experiment 1, feeding a complex diet (CPX) containing fish meal and sunflower meal as the main protein sources eliminated the adverse effects of SS inoculation on performance traits as compared with SS effects on poults fed a corn and soybean meal (CS) diet. In Experiments 2 and 3, the effects of SS were more severe than in Experiment 1. In these experiments, the CPX diet only partly overcame the adverse effects of SS on performance (i.e., in Experiment 2, growth depressions from 2 to 5 days of age were 90.3 and 59.6% in SS-inoculated poults fed the CS and CPX diets, respectively, as compared with uninoculated, control poults fed the same diets). Properties of the CPX diet that made it effective in reducing the severity of SS were not evident from the results of Experiment 3. Replacing soybean meal with soy protein or canola meal was ineffective as compared with the use of a mixture of sunflower meal, fish meal, meat and bone meal, and corn gluten meal.

Responses of turkey poults to virginiamycin as influenced by litter condition and experimentally induced stunting syndrome.

Two experiments were conducted to evaluate the effect of virginiamycin (VM, 22 mg/kg of diet) on performance of uninfected (CON) turkey poults and those infected (INO) with stunting syndrome and reared on used woodshavings (Experiment 1) or on clean or used woodshavings (Experiment 2). Virginiamycin improved BW (P less than .001) and feed efficiency (FE) (P less than .05) from 1 to 29 days of age, irrespective of type of litter or disease condition. The increase in BW induced by VM, however, was greatest when poults were kept on used litter, resulting in significant (P less than .05) VM by litter interaction. Induced stunting syndrome depressed BW (P less than .01) to 29 days of age and impaired FE from 1 to 9 days of age (P less than .05) and from 5 to 9 days of age (P less than .01) in Experiments 1 and 2, respectively. Virginiamycin did not prevent early adverse effects of INO on BW and FE, but facilitated notable recovery of INO poults relative to INO poults not fed VM. Virginiamycin increased specific activities of maltase and sucrase of the jejunum of CON poults in Experiments 1 and 2; in Experiment 2, this VM effect was evident irrespective of type of litter. Maltase-specific activity and sucrase were reduced by INO (P less than or equal to .05 and P less than or equal to .01 in Experiments 1 and 2, respectively) and VM did not modify this effect. The maltase and sucrase data suggest that VM improved BW and FE of CON poults, in part, by helping to maintain digestive and absorptive functions of the small intestine during the early growth period, but, in the instance of INO poults, VM was not effective in this regard.

Effects of early immune stress and changes in dietary metabolizable energy on the development of newly hatched turkeys. 1. Growth and nutrient utilization.

Two experiments were conducted to document the effects of an early immunologic stress and changes in dietary ME(n) on growth and nutrient utilization of newly hatched turkeys. Treatments in both experiments consisted of a complete factorial arrangement of two types of injection and four isonitrogenous diets. Turkeys were injected i.p. with saline (SAL) or a solution of lipopolysaccharide (LPS) (100 micrograms LPS/mL SAL) at 1, 3, and 5 d of age. In Experiment 1, two diets were formulated to contain 2,800 kcal ME(n)/kg. One was a corn-soybean meal-based diet (CSBM) and the other contained 8% Solkafloc (SKF). A third diet (3,100 kcal ME(n)/kg) was formulated by substituting 8% sucrose (SUC) for the 8% SKF. The fourth diet included in Experiment 1 was formulated to contain 3,700 kcal ME(n)/kg. The CSBM and SUC diets were also included in Experiment 2. Two additional diets tested in Experiment 2 were the CSBM diet containing 74.5 mg ibuprofen/kg (IBU) and a corn-soybean meal-based diet with a ME(n) value of 3,100 kcal/kg (CS31). Injection with LPS reduced (P < .05) BW of turkeys throughout Experiment 1 and until 9 d of age in Experiment 2, as compared with injection with SAL, irrespective of dietary treatment. The reduction in BW was mainly due to a decrease in feed intake (FI) (P < .05). Turkeys fed diets with 3,100 kcal ME(n)/kg were heavier (P < .05) than those fed diets with 2,800 kcal ME(n)/kg, irrespective of injection. Inclusion of ibuprofen to the CSBM diet from 1 to 14 d improved (P < .05) BW and feed efficiency (P < .01) of turkeys at 14 d of age, compared with turkeys fed the CSBM diet. Determined ME(n) was not affected by LPS injection. Adverse effects of LPS injection on growth of turkey poults were mainly the consequence of a reduced FI and not of altered nutrient utilization. These effects were not fully alleviated by feeding a diet with 3,100 kcal ME(n)/kg.

Integrin receptor alpha 6 beta 1 is localized at specific sites of cell-to-cell contact in rat seminiferous epithelium.

With the aim of investigating the presence and role of integrin receptors in cell-to-cell interactions during spermatogenesis, we have immunolocalized alpha 6A integrin chain in the rat seminiferous epithelium. In both prepubertal and adult seminiferous epithelium, the antigen was found to be restricted to definite sites of intercellular contact, following a stage-specific distribution almost invariably identical to that known for beta 1 chain. In the adult seminiferous epithelium, positivity for both antigens was found exclusively around the profiles of elongating and maturing spermatids and, in most but not all stages, at a characteristic suprabasal position. In the prepubertal rat, the antigen is localized along a very regular suprabasal line of intercellular contacts. In immunoprecipitation experiments on both seminiferous epithelium explants and Sertoli cell cultures from 3-wk-old rats, anti-alpha 6A antibody coprecipitates a polypeptide of 118 kDa, presumably corresponding to beta 1 chain. These data strongly suggest that the integrin heterodimer alpha 6A beta 1 is expressed at sites of intercellular contact in the rat seminiferous epithelium. The stage-specific and restricted pattern observed by immunofluorescence suggests that this integrin is involved in regulatory interactions between Sertoli cells and germ cells during spermatogenesis.

Heterologous expression and purification of recombinant rolipram-sensitive cyclic AMP-specific phosphodiesterases.

With the cloning of cDNAs coding for the different phosphodiesterase 4 (PDE4) isoenzymes present in mammals, homogeneous preparations of these forms have become readily available. This strategy has greatly facilitated the understanding of the properties of the myriad of isoforms derived from the four PDE4 genes found in mammals, and has opened a new avenue to develop inhibitors with a different degree of selectivity for each isoform. Here we describe the strategies and methods used to express PDE4 in bacterial, yeast, insect, and mammalian cell heterologous systems, and review the advantages and disadvantages of each of these expression strategies. In addition, procedures to purify the recombinant proteins are described. The recently developed purification of a PDE4 by immunoaffinity chromatography provides a rapid and efficient method to prepare large quantities of PDE4. This method should be very useful for structural and kinetic studies on the PDE4D isoforms.