Lack of change in swimming capacity (Ucrit) following acute salinity exposure in juvenile shortnose sturgeon (Acipenser brevirostrum).

The Saint John River (SJR) is home to the only Canadian population of shortnose sturgeon, Acipenser brevirostrum. Adult shortnose sturgeon routinely enter saltwater to forage, yet less is known about how juveniles cope with the associated osmoregulatory pressures. Recently, it has been shown that short-term (24 h) exposure to saltwater causes significant changes to ion and water levels in juvenile shortnose. In some species of fish, notably salmonids, it has been shown that shifts in fluid and ion levels following saltwater challenges reduce the swimming capacity. The relationship between ion concentration and swimming capacity is not well understood for sturgeon species. Our research aimed to determine whether short-term salt exposure affects swimming ability in juvenile shortnose sturgeon. Juvenile, SJR, hatchery-raised shortnose sturgeon (< 1 year old) were exposed to salinities of 0 (control), 16, or 24‰ for 24 h and then subjected to a critical swimming speed test (Ucrit) to quantify swimming ability. Following the test, the fish were weighed and blood samples were drawn to be analyzed for plasma ion and cortisol levels. While ion levels and weight loss were significantly higher in salt exposed fish, there were no significant differences in critical swimming speed or cortisol concentrations. This is in contrast to what has been observed in salmonids and Adriatic sturgeon. This suggests the hydromineral imbalance caused by moderate salt exposure is not sufficient to affect the swimming performance of shortnose sturgeon. Shortnose sturgeon are not thought to enter the saline stretches of the SJR until roughly 8 years of age, yet this research shows that much younger juveniles withstand moderate salinity for short periods, with little whole-animal ramifications.

The effect of substratum type on aspects of swimming performance and behaviour in shortnose sturgeon Acipenser brevirostrum.

The swimming performance and associated swimming behaviour (i.e. substratum-skimming, station-holding and free swimming) were assessed in shortnose sturgeon Acipenser brevirostrum during critical swimming and endurance swimming tests over a rough and a smooth substratum. It was hypothesized that the addition of a rough substratum in the swimming flume may provide a surface for the A. brevirostrum to grip and offer an energetic advantage. Substratum type did not affect the critical swimming performance, but A. brevirostrum consistently performed more bottom behaviours (i.e. substratum-skimming and station-holding) while on a smooth substratum. Acipenser brevirostrum had little contact with the rough substratum until the velocity was >1 body length s-1 . Endurance swimming time was significantly lower for A. brevirostrum over the rough bottom at the highest velocity (30 cm s-1 ) which may be attributed to the observed increase in free swimming and decrease in bottom behaviours. During endurance swimming, the rough substratum was mainly used at intermediate velocities, suggesting that there may be a stability cost associated with being in contact with the rough substratum at certain velocities.

Oxygen consumption and haematology of juvenile shortnose sturgeon Acipenser brevirostrum during an acute 24 h saltwater challenge.

This study focused on the acute physiological responses to saltwater exposure in juvenile shortnose sturgeon Acipenser brevirostrum. In two separate laboratory experiments, 2 year-old A. brevirostrum were exposed to either full (32) or half-strength (16) seawater for up to 24 h. First, oxygen consumption rates were used to estimate the metabolic costs over 24 h. Secondly, blood and muscle samples were analysed at 6, 12 and 24 h for water loss, various measures of osmoregulatory status (plasma osmolality and ions) and other standard haematological variables. Juveniles exposed to full-strength seawater showed significant decreases in oxygen consumption rates during the 24 h exposure. Furthermore, seawater-exposed fish had significantly increased plasma osmolality, ions (Na(+) and Cl(-)) and a 17% decrease in total wet mass over the 24 h exposure period. To a lesser extent, increases in osmolality, ions and mass loss were observed in fish exposed to half-strength seawater but no changes to oxygen consumption. Cortisol was also significantly increased in fish exposed to full-strength seawater. While plasma protein was elevated following 24 h in full-strength seawater, haemoglobin, haematocrit and plasma glucose levels did not change with increased salinity. These results imply an inability of juvenile A. brevirostrum to regulate water and ions in full-strength seawater within 24 h. Nonetheless, no mortality occurred in any exposure, suggesting that juvenile A. brevirostrum can tolerate short periods in saline environments.

Reduced embryo yield obtained from superstimulated ewes with low circulating AMH concentration is improved by lengthening the FSH treatment.

The aim of the present study was to evaluate: 1) the association between AMH, AFC, superovulatory response and embryo yield in sheep; and 2) the effect of FSH treatment length during superstimulation of the first follicular wave on ovarian response and embryo yield, particularly in ewes with low and high AMH. The experiment was performed on 63 Polled Dorset ewes that received an ovarian superstimulatory treatment during the first follicular wave (Day 0 protocol). Ewes were administered a total dose of 240 mg of FSH distributed in six (6-dose regimen, n = 30) or eight (8-dose regimen, n = 33) decreasing doses administered 12 h apart. On Day -9 (random stage of the estrous cycle) and Day 0 (day of the first FSH dose) ovarian ultrasonography was performed and blood samples were collected for AFC and AMH determinations, respectively. A weak positive correlation between AMH and small AFC (follicles <4 mm) was observed (r = 0.23; P = 0.07), and AMH concentration was positively correlated (r = 0.29; P < 0.05) with the number of corpora lutea (CL) determined at embryo collection (i.e., 6 d after insemination). The length of FSH treatment tended (P = 0.06) to affect the ovarian response, such that the number of CL was greater in 8-dose than 6-dose treated ewes, while no differences (P > 0.10) in embryo yield outcomes were observed. For further analysis, ewes were classified into low (<7 ng/mL) and high (>10 ng/mL) serum AMH. In high AMH ewes, there were no differences (P > 0.05) in the number of CL nor embryo yield between the 6-dose and 8-dose treatment (e.g., 7.8 ± 2.4 and 8.3 ± 2.5 transferable embryos, respectively; P = 0.92). Conversely, for low AMH ewes, fertilized ova and embryo yield were greater (P ≤ 0.05) for ewes receiving the 8-dose than the 6-dose superstimulatory treatment (e.g., 8.4 ± 2.8 vs. 2.7 ± 0.9 transferable embryos, respectively, P ≤ 0.05). In conclusion, embryo production in poor responding ewes with low low circulating AMH is improved by extending the superstimulatory treatment length from 6 to 8 FSH doses.

The effects of different feeding strategies providing different levels of vitamin A on animal performance, carcass traits, and the conversion rate of subcutaneous fat color in cull-cows.

Cull cows represent a significant percentage of revenue received from the U.S. beef industry; however, cull cows are heavily price discounted at time of slaughter. This experiment's objective is to evaluate different feeding strategies and their effects on body condition score, subcutaneous fat color, and carcass yield and quality traits in cull cows. The central hypothesis is feeding a high-energy diet, with low levels of vitamin A, for 56 d will improve animal performance, carcass yield, and quality traits in addition to capturing the point (rate) of the conversion of yellow to white subcutaneous fat. In the present experiment 98 Angus crossbreed cows were utilized. Cows were fed either low vitamin A (LVA) diet consisting of whole shelled corn, soybean hulls, soybean meal, and a mineral-vitamin supplement or high vitamin A (HVA) diet, formulated using whole shelled corn, fescue hay, dry distiller grains with soluble, and a mineral-vitamin supplement for 56 d. During the 56 d feeding period, body weights and condition scores, and subcutaneous adipose samples were collected every 14 d. On day 56, cattle were slaughtered; 48 h postmortem carcass characteristics and objective color scores (subcutaneous adipose tissue) were recorded and a sample of the longissimus dorsi lumborum was collected. Subcutaneous adipose tissue samples were utilized to record subjective color scores and then ground to be analyzed for ß-carotene concentration. The longissimus dorsi lumborum samples (2.54 cm slices) were removed for Warner-Bratzler shear force (WBSF) and pH testing. Data were analyzed using the MIXED procedure of SAS. Feeding cull cows LVA resulted in differences in subcutaneous carcass fat color (P = 0.01) as well as b* values (P < 0.01) on day 56 compared with HVA. Subjective fat color scores were not different (P > 0.10) on day 0 or 14 but were different (P ≤ 0.05) on days 28, 42, and 56. Additionally, 9-cis-ß-carotene concentration on day 56 were different (P = 0.05) between treatments. A trend was noticed for all-trans-ß-carotene concentration (P = 0.10) on day 56 as well. Cull cow body weights were greater (P ≤ 0.04) when fed the LVA diet starting on days 14, 28, and 42; and a trend was noticed on day 56 (P = 0.09). Overall, cows fed the LVA treatment for 56 d exhibited decreased adipose yellowness and ß-carotene concentrations as well as increased live weights.

Atlantic sturgeon and shortnose sturgeon exhibit highly divergent transcriptomic responses to acute heat stress.

In comparison to most modern teleost fishes, sturgeons generally display muted stress responses. While a muted stress response appears to be ubiquitous across sturgeon species, the mechanisms unpinning this muted response have not been fully described. The objective of this study was to determine the patterns of hematological and transcriptomic change in muscle tissue following an acute high temperature stress (critical thermal maxima; CTmax) in two locally co-occurring but evolutionarily distant sturgeon species (Atlantic and shortnose sturgeon). The most striking pattern found was that Atlantic sturgeon launched a vigorous transcriptomic response at CTmax, whereas shortnose sturgeon did not. In contrast, shortnose sturgeon have significantly higher cortisol than Atlantics at CTmax, reconfirming that shortnose have a less muted cortisol stress response. Atlantic sturgeon downregulated a number of processes, included RNA creation/processing, methylation and immune processes. Furthermore, a number of genes related to heat shock proteins were differentially expressed at CTmax in Atlantic sturgeon but none of these genes were significantly changed in shortnose sturgeon. We also note that the majority of differentially expressed genes of both species are undescribed and have no known orthologues. These results suggest that, while sturgeons as a whole may show muted stress responses, individual sturgeon species likely use different inducible strategies to cope with acute high temperature stress.

GnRH dose at initiation of a 5-day CO-Synch + P4 for fixed time artificial insemination in suckled beef cows.

The objective of the present study was to evaluate the effect of GnRH dose administered at initiation (GnRH-1) of a 5-day CO-Synch + P4 protocol on ovulatory response, expression of estrus, and fertility in suckled beef cows. Suckled beef cows (n = 1101) at four locations were randomized to receive either 100 or 200 µg of gonadorelin acetate at initiation (D-8) of a 5-day CO-Synch + P4 protocol concurrently with insertion of an intravaginal progesterone (P4) device. On D-3 the P4 device was removed, two doses of prostaglandin F2α were administered concurrently and a patch was applied to evaluate expression of estrus. Artificial insemination was performed 72 h after P4 device removal (D0) simultaneously with the administration of 100 µg of gonadorelin acetate (GnRH-2). Increasing GnRH dose at initiation of a 5-day CO-Synch + P4 did not enhance ovulatory response (P = 0.57) to GnRH-1, expression of estrus (P = 0.79), nor pregnancies per AI (P/AI; P = 0.91). Both follicle size (quadratic) and circulating P4 (linear) affected (P < 0.01) ovulatory response to GnRH-1 independent of dose. Cows that had ovulation to GnRH-1 had smaller (P < 0.001) follicle size on D-3 and reduced (P = 0.05) expression of estrus compared to cows that did not have ovulation to GnRH-1, however, P/AI did not differ (P = 0.75). In conclusion, increasing the dose of GnRH-1 in the 5-day CO-Synch + P4 protocol did not enhance ovulatory response, expression of estrus, or P/AI in suckled beef cows.

The influence of flume length and group size on swimming performance in shortnose sturgeon Acipenser brevirostrum.

The main objectives of this study were to determine optimal methodologies to assess the general swimming performance of juvenile shortnose sturgeon Acipenser brevirostrum. Swimming densities (group v. individual swimming) and flume length (2 v. 1 m) were altered to verify if any of those variables affected performance (i.e. time to fatigue) during critical swimming (U(crit)) and endurance tests. Results for both U(crit) and endurance swimming were not significantly different between fish swum in groups of five or fish swum individually. The U(crit) values, however, were c. 22% higher for fish swum in a longer flume. Although swimming fish in groups did not improve swimming performance, group swimming lowered the variance of the data. Results also reveal that juvenile A. brevirostrum may not possess an ability to swim at high speeds (i.e. burst phase) for long periods.

Interaction of dendritic cells with skin endothelium: A new perspective on immunosurveillance.

The goal of this study was to determine the mechanisms by which dendritic cells (DCs) in blood could interact with endothelium, a prerequisite to extravasation into tissues. Our results indicate that DCs express both HECA-452-reactive and nonreactive isoforms of P-selectin glycoprotein ligand 1 (PSGL-1) and can tether and roll efficiently on E- and P-selectin under flow conditions in vitro. Freshly isolated blood DCs were further observed to roll continuously along noninflamed murine dermal endothelium in vivo. This interaction is strictly dependent on endothelial selectins, as shown by experiments with blocking antibodies and with E- and P-selectin-deficient mice. We hypothesize that DCs in blood are constitutively poised at the interface of blood and skin, ready to extravasate upon induction of inflammation, and we showed that cutaneous inflammation results in a rapid recruitment of DCs from the blood to tissues. We propose that this is an important and previously unappreciated element of immunosurveillance.

Intranasal oxytocin treatment does not attenuate the hypothalamo-pituitary-adrenal axis in beef heifers subjected to isolation stress or restraint and isolation stress.

Changes in the physiological, psychological, and behavioral manifestations of stress have been observed in association with increases in circulating oxytocin (OXT). Providing OXT intranasally has been shown to attenuate stressor-induced hypothalamo-pituitary-adrenal (HPA) axis activation in humans and rodents; however, anxiolytic effects may be context and species specific. The present study aimed to investigate the effect of intranasal OXT supplementation on stressor-induced activation of the HPA axis in beef cattle. We hypothesized that OXT would attenuate activation of the HPA axis, ultimately decreasing plasma cortisol and adrenocorticotropic hormone (ACTH). Twenty-eight Bos taurus heifers were blocked by bodyweight and randomly allocated to one of four treatment groups, in a 2 × 2 factorial arrangement: (1) saline, isolated, standing, and unrestrained (S-isolation stress [IS], 0.015 mL/kg BW 0.9% isotonic saline, n = 7); (2) saline, isolated, and restrained (S-restraint and isolation stress [RIS]; 0.015 mL/kg BW 0.9% isotonic saline; n = 7); (3) OXT, IS (OXT-IS, 0.3 IU/kg BW oxytocin; n = 7); and (4) OXT and RIS (OXT-RIS, 0.3 IU/kg BW oxytocin; n = 7). Oxytocin and saline were administered intranasally. Intranasal treatments were given followed by a waiting time of 30 min when each of the stress treatments was applied for 2 h. Blood samples were collected via jugular catheters directly after stressor application and every 10 min thereafter, for 2 h. Cortisol concentrations increased over time in animals exposed to RIS (P < 0.01) and decreased over time in animals exposed to IS (P < 0.01). Concentrations of ACTH decreased over time for the IS-treated heifers but remained elevated for the RIS-treated heifers (P < 0.01). Under the conditions of the present study, OXT treatment did not affect measured indicators of HPA axis activation. A treatment × time interaction (P < 0.01) was detected for OXT, such that OXT heifers exhibited greater initial OXT concentrations followed by a decline; saline-treated heifers had consistently stable oxytocin concentrations. The RIS-treated heifers increased their glucose (P < 0.01) and lactate (P < 0.01) concentrations throughout the application of the stressors compared with the IS-treated heifers. Overall, restraint stress increased cortisol and oxytocin in B taurus heifers compared with heifers subjected only to isolation. Finding a more intermediate stress model may better allow for detection of the effects of oxytocin on the stress response.

Short communication: pharmacokinetics of oxytocin administered intranasally to beef cattle.

Providing the neuropeptides oxytocin and vasopressin intranasally increased concentrations in plasma and cerebral spinal fluid in humans and primates, respectively. This is of interest because of the documented anxiolytic effects of oxytocin observed in humans and rodents. To date, a transnasal approach of hormone administration has not been investigated in beef cattle. Defining the pharmacokinetics of intranasal oxytocin in cattle is necessary for determining optimum sampling and dosing timelines for future investigations. Five, weaned Bos taurus steers were used in a 3 × 3 Latin square design. Treatments included 1) 0.33 IU oxytocin/kg BW (A, n = 5), 2) 0.66 IU oxytocin/kg BW (B, n = 5), and 3) 1.32 IU oxytocin/kg BW (C, n = 5). Steers were acclimated to handling and restraint procedures for 4 wk leading up to the start of the experiment. Frequent blood collection occurred every 2 min for the first 30 min and every 5 min for the second 30 min, relative to administration of intranasal treatment. No treatment by time interaction was detected; however, there was an effect of time (P < 0.001) and treatment (P = 0.002) on oxytocin concentrations over time. Pharmacokinetic parameters, determined by PKSolver excel add-in, demonstrated an average maximum concentration (CMAX) of 63.3 pg/mL at 3.5 min after intranasal dose administration. An average half-life (T1/2) of 12.1 min after intranasal administration was determined. Pharmacokinetic parameters to a single bolus were not dose-dependent.

Thyrotropin releasing hormone: development of inactivation system during maturation of the rat.

Whereas thyrotropin releasing hormone is rapidly and extensively degraded by plasma of adult rats, no appreciable loss of biological or immunological activity is caused by plasma from rats 4 or 16 days old. The plasma of neonatal rats does not appear to contain an inhibitor of thyrotropin releasing hormone peptidase or a peptidase with altered substrate affinity. The development of an active peptidase in rat plasma suggests a physiological role for inactivation of thyrotropin releasing hormone.

Oxygen consumption, ammonia excretion and protein use in response to thermal changes in juvenile Atlantic salmon Salmo salar.

Experiments were designed to examine the effects of various temperature challenges on oxygen consumption and ammonia excretion rates and protein utilization in juvenile Atlantic salmon Salmo salar. Fish acclimated to 15 degrees C were acutely and abruptly exposed to either 20 or 25 degrees C for a period of 3 h. To simulate a more environmentally relevant temperature challenge, a third group of fish was exposed to a gradual increase in temperature from 15 to 20 degrees C over a period of 3 h (c. 1.7 degrees C h(-1)). Oxygen consumption and ammonia excretion rates were monitored before, during and after the temperature shift. From the ammonia excretion and oxygen consumption rates, protein utilization rates were calculated. Acute temperature changes (15-20 degrees C or 15-25 degrees C) caused large and immediate increases in the oxygen consumption rates. When the temperature was gradually changed (i.e. 1.7 degrees C h(-1)), however, the rates of oxygen consumption and ammonia excretion were only marginally altered. When fish were exposed to warmer temperatures (i.e. 15-20 degrees C or 15-25 degrees C) protein use generally remained at pre-exposure (15 degrees C) levels. A rapid transfer back to 15 degrees C (20-15 degrees C or 25-15 degrees C) generally increased protein use in S. salar. These results indicate that both the magnitude and the rate of temperature change are important in describing the physiological response in juvenile salmonids.

Behaviour and performance of juvenile shortnose sturgeon Acipenser brevirostrum at different water velocities.

Critical swimming speeds (mean +/-s.e.) for juvenile shortnose sturgeon Acipenser brevirostrum were 34.4 cm s(-1)+/- 1.7 (2.18 +/- 0.09 body lengths, BL s(-1)). Swimming challenges at 10, 20 and 30 cm s(-1) revealed that juvenile A. brevirostrum are relatively poor swimmers, and that the fish did not significantly modify their swimming behaviour, although they spent more time substratum skimming (i.e. contact with flume floor) at 30 cm s(-1) relative to 10 cm s(-1). When present, these behavioural responses are probably related to morphological features, such as flattened rostrum, large pectoral fins, flattened body shape and heterocercal tail, and may be important to reduce the costs of swimming.

Physiological and genetic analyses of inbred mouse strains with a type I iodothyronine 5' deiodinase deficiency.

Inbred mouse strains differ in their capacity to deiodinate iododioxin and iodothyronines, with strains segregating into high or low activity groups. Metabolism of iododioxin occurs via the type I iodothyronine 5'deiodinase (5'DI), one of two enzymes that metabolize thyroxine (T4) to 3,5,3'-triiodothyronine (T3). Recombinant inbred strains derived from crosses between high and low activity strains exhibit segregation characteristic of a single allele difference. Hepatic and renal 5'DI mRNA in a high (C57BL/6J) and low (C3H/HeJ) strain paralleled enzyme activity and concentration, in agreement with a recent report. 5'DI-deficient mice had twofold higher serum free T4 but normal free T3 and thyrotropin. Brown adipose tissue 5'DII was invariant between the two strains. Southern analyses using a 5'DI probe identified a restriction fragment length variant that segregated with 5'DI activity in 33 of 35 recombinant inbred strains derived from four different pairs of high and low activity parental strains. Recombination frequencies using previously mapped loci allowed assignment of the 5'DI gene to mouse chromosome 4 and identified its approximate chromosomal position. We propose the symbol Dio1 to denote the mouse 5'DI gene. Conserved linkage between this segment of mouse chromosome 4 and human HSA1p predicts this location for human Dio1.

Effects of metal mining effluent on Atlantic salmon (Salmo salar) and slimy sculpin (Cottus cognatus): using artificial streams to assess existing effects and predict future consequences.

In the summer of 2000, the effects of metal mine discharge on fish growth and exercise performance were assessed at a Zn-Pb-Cu mine in New Brunswick, Canada. Juvenile Atlantic salmon (Salmo salar) were exposed to 0%, 20%, and 80% treated metal mine effluent in a mobile, fish-only artificial stream system. Fish were fed commercial salmon pellets throughout the study. Young-of-the-year slimy sculpins (Cottus cognatus) were exposed to the same treatments in a multitrophic level, modular artificial stream system or mesocosm, in which the fish were dependent on seeded algae and invertebrates for nutrition. Treatment concentrations were chosen to represent existing discharge dilutions (80%) and a scenario of reduced effluent discharge (20%) as predicted upon mine closure (scheduled for 2008). Al, Ba, B, Fe, Mn, Sr, Tl, Ti, and Zn increased in a concentration-dependent fashion across the three treatments. Salmon body burdens of Ba, Cd, Li, Cu, Mn, Se, Sr, and Zn were increased in the 80% treatment, while Tl increased across all treatment levels. Mortalities and depressions in growth in both fish species paralleled treatment concentrations (80%>20%>0%). Salmon liver weight was significantly greater in fish exposed to 20% and 80% effluent in a concentration-dependent fashion. Exercise performance in fish, as assessed by the ability to recover from forced exercise, showed little effect of treatment. The contamination of the receiving environment by mine discharges has led to loss of fish, making it impossible to study the system in situ. However, the use of the artificial stream systems enabled us to assess effects of present conditions on fish, as well as the potential impacts of mine reclamation. The 20% discharge predicted following mine reclamation is potentially favourable for the reinstitution of native fishes into the system.

Juvenile atlantic and shortnose sturgeons (family: Acipenseridae) have different hematological responses to acute environmental hypoxia.

Experiments were conducted to determine the behavioral and physiological responses to acute hypoxic challenges in Atlantic (Acipenser oxyrinchus) and shortnose (Acipenser brevirostrum) sturgeons. We measured the ventilatory rate following a 45-mmHg hypoxic challenge, as well as a variety of hematological parameters, including O2 transport and hormonal, ionic, and metabolic variables, following a 1-h exposure to either 75- or 30-mmHg hypoxic challenges. Compared to fish in normoxic conditions (Pwo2 150 mmHg), juveniles of both species increased their ventilatory rate by approximately 40% when exposed to a 1-h challenge at 45 mmHg Pwo2. Hematological variables (e.g., hematocrit, hemoglobin, and Na+ and Cl- levels) did not change substantially following a 1-h challenge at 75 mmHg Pwo2. Conversely, a severe hypoxic challenge of 30 mmHg caused changes in several hematological variables (e.g., whole blood glucose and plasma cortisol and lactate levels). Most of these hematological parameters returned to prehypoxic levels within 2 h. Severe environmental hypoxia elicited the same basic pattern of response in both species; however, maximal plasma lactate levels were higher in Atlantic sturgeons, and maximal cortisol levels were higher in shortnose sturgeons. Whether these species differences are related to dissimilar hypoxia-tolerance, ecological, and/or endocrinological characteristics between these two species is not entirely clear.

Inflammatory skin disease in K14/p40 transgenic mice: evidence for interleukin-12-like activities of p40.

The proinflammatory cytokine interleukin-12, a p35/p40 heterodimer, is produced by resident cells in skin and has been implicated as a pathogenetic factor in T-cell-mediated skin diseases. Secretion of heterodimeric interleukin-12 is always accompanied by production of p40 monomer and p40/p40 homodimer. To investigate the possible in vivo role of p40 per se, we generated mice that constitutively express monomeric and homodimeric p40 in basal keratinocytes. These mice spontaneously developed an eczematous skin disease that was characterized by hyperkeratosis, focal epidermal spongiosis, and a mixed inflammatory infiltrate composed of T cells (CD4+), macrophages, eosinophils, mast cells, and few neutrophils. Fluorescence-activated cell sorter analysis of transgenic epidermal cell suspensions revealed induction of major histocompatibility complex class II molecules on keratinocytes and a 2-3-fold increase in the content of Langerhans cells. Cytokines produced by these activated epidermal cells include interleukin-1alpha and tumor necrosis factor alpha. The skin disease in K14/p40 mice was similar to that of littermate mice that received injections of interleukin-12, suggesting overlapping in vivo functional properties. As induction of interferon-gamma is a major function of interleukin-12, we tested the in vitro ability of transgenic p40 to induce interferon-gamma. In contrast to interleukin-12, transgenic p40 did not stimulate interferon-gamma secretion by cultured splenocytes. We conclude that transgenic p40 and interleukin-12 are equally capable of initiating cutaneous inflammation. Despite these in vivo similarities, there is a clear functional difference between interleukin-12 and transgenic p40 in vitro, suggesting that interferon-gamma is not a major factor contributing to interleukin-12-like activities of transgenic p40.

Photoaffinity labeling of rat type I iodothyronine deiodinase.

The photoreactive compound p-nitrophenyl-2-diazo-3,3,3-trifluoropropionate (PAL) was coupled to [125I]rT3, T4, or T3 and incubated with liver and kidney microsomes of hypo-, hyper-, or euthyroid rats to identify the type I iodothyronine deiodinase. Various substrates or inhibitors of the enzyme, including rT3, T4, T3, 6-n-propylthiouracil (PTU), and iopanoic acid, were used as competitors to establish the specificity of protein labeling. The PAL derivatization enhanced the behavior of T4 and T3 as substrates for the type I enzyme. No specific labeling of microsomal proteins was observed with either rT3 or T4-PAL, presumably due to deiodination of the labeled compound. In contrast, T3-PAL labeled a 27-kDa band, the presence of which paralleled thyroid status. The labeling of only this protein was blocked by either substrates or enzyme inhibitors in a dose-dependent fashion, with a rank order of potency predicted by the activity of such compounds in type I enzyme assays. The specific nature of these competitions provides further evidence that this 27-kDa protein, identified in previous studies using N-bromoacetyl [125I]T3 or -T4, contains the active site of the rat type I deiodinase. This is in agreement with the mol wt of the rat type I deiodinase deduced from the recently identified cDNA coding for this protein.

Plasma TSH levels, by radioimmunoassay, during the estrous cycle of the rat.

Between 1000 and 1200 hr on any given morning, female rats displaying regular 4-day estrous cycles were bled by decapitation. Plasma TSH concentrations were determined by homologous radioimmunoassay (RIA), in terms of NIAMDD-Rat-TSH-I-1 standards. Mean plasma TSH concentrations plus or minus SE were 1.2 plus or minus 0.1, 1.5 plus or minus 0.2, 2.3 plus or minus 0.6, and 1.1 plus or minus 0.1 ng/ml for rats killed at proestrus, estrus, diestrus-1, and diestrus-2, respectively. Statistical analysis revealed no significant differences among the TSH levels, except for the elevation at diestrus-1, which is of borderline significance at the 0.05 probability level. Our findings, based on RIA, are in contrast to previous reports, based on bioassays, of an elevation of plasma TSH at estrus.