Effect of beta-adrenergic agonists on lipolysis and lipogenesis by porcine adipose tissue in vitro

The effects of the beta-adrenergic agonists isoproterenol, cimaterol, ractopamine and clenbuterol on lipolysis (release of glycerol and free fatty acids) and lipogenesis (incorporation of 14C into fatty acids from [14C]glucose) was examined in porcine adipose tissue explants in vitro. Lipolysis was stimulated by isoproterenol, cimaterol or ractopamine but not by clenbuterol. Insulin reduced the lipolytic effects of the beta-adrenergic agonists (isoproterenol, cimaterol and ractopamine). Lipogenesis was inhibited by all beta-adrenergic agonists tested (isoproterenol, cimaterol, ractopamine and clenbuterol). The antilipogenic effect of the beta-adrenergic agonists was reduced by the presence of insulin in the incubation. Although effects of the different beta-adrenergic agonists varied, all had some direct effects that could be expected to reduce adipose accretion. Effects of beta-adrenergic agonists in the pig are due in part to direct effects on adipose tissue.     

Characterization of porcine beta1- and beta2-adrenergic receptors in heart, skeletal muscle, and adipose tissue, and the identification of an atypical beta-adrenergic binding site

The objective of this study was to characterize porcine beta1- and beta2-adrenergic receptors (beta1-AR and beta2-AR) in heart, skeletal muscle, and adipose tissue by measuring the binding of a radioligand to cell membrane fragments. In skeletal muscle (LM), [3H]CGP12177 labeled a homogeneous population of beta2-AR as evidenced by the rank order of affinity of catecholamines [(-)isoproterenol > (-)epinephrine > (-)norepinephrine], a high affinity of the binding site for the beta2-AR-agonist clenbuterol (equilibrium dissociation constant, Kd = 16 nM), and a low affinity of the binding site for the beta1-AR-antagonist CGP20712A (Kd = 21 microM). The affinity of ICI118551, a ligand selective for beta2-AR in other species, was uncharacteristically low in porcine LM (Kd = 441 nM), but was consistent with a value reported for the cloned porcine beta2-AR. In heart ventricle, ligand binding revealed a predominant population of beta1-AR, judged by the rank order of affinity of catecholamines [(-)isoproterenol > (-)epinephrine > or = (-)norepinephrine] and high-affinity binding to CGP20712A (Kd = 40 nM). The Kd for ICI118551 (731 nM) was close to that observed at beta2-AR in LM, confirming that ICI118551 is not subtype-selective in the pig. Displacement studies using (-)propranolol, clenbuterol, and (-)isoproterenol revealed a second high-affinity binding site in the heart that was not a beta2-AR and could not be eliminated by guanosine 5'-triphosphate or guanylyli-midodiphosphate. In adipose tissue, an equal number of beta1- and beta2-AR was identified through the binding of clenbuterol and CGP20712A, whereas ICI118551 could not discriminate between these sites. In further experiments, we used 10 microM CGP20712A to eliminate beta1-AR binding and allow accurate Kd values to be determined at beta2-AR for nonselective ligands. Under these conditions, another binding site was observed that had a high affinity for (-)propranolol (Kd = 20 pM), which is inconsistent with beta3- or beta4-AR binding reported elsewhere. Our results indicate that porcine adipose tissue contains beta1-AR, beta2-AR, and an atypical binding site in the proportions 50, 34, and 16%, respectively, of the total binding sites labeled by [3H]CGP12177.     

Effects of adrenergic agonists and insulin on porcine adipose tissue lipid metabolism in vitro

Because this laboratory has been able to demonstrate only a small and somewhat inconsistent stimulation of glucose metabolism by insulin in porcine adipose tissue in vitro, the tissue was preincubated with insulin to attempt to enhance the hormone effect. Preincubation with or without insulin did not increase insulin stimulation. Furthermore, insulin did not stimulate triacylglycerol biosynthesis. Adrenergic hormones stimulated lipolysis in porcine adipose tissue in vitro. Several analogs of norepinephrine incubated with porcine adipose tissue in vitro did not inhibit glucose incorporation into CO2 or total lipids, in contrast to inhibition observed in adipose tissue from other species. Isoproterenol inhibited glycerol-3-phosphate incorporation into lipids; the maximal inhibition was 50% for the initial stages of the pathway. Palmitate incorporation into lipids also was inhibited 50% by isoproterenol but this may have been an artifact. Preincubation of adipose tissue, with no exogenous hormone, might decrease the concentration of endogenous adrenergic hormones and thus make the tissue more responsive to exogenous adrenergic hormones. Preincubation of porcine adipose tissue did not consistently lower the basal lipolytic rate but enhanced the stimulated lipolytic rate; the mechanism is not known. These experiments provide no evidence that preincubation is beneficial to measurement of lipolysis or glucose metabolism in porcine adipose tissue in vitro.     

Estradiol-17 beta effects on lipid metabolism of adipose tissue in nutritionally induced lean and obese ovariectomized ewes.

Nutritionally manipulated lean (68 kg) and obese (87 kg) ovariectomized ewes were administered estradiol-17 beta (E2) or sham implants. Ewes individually had ad libitum access to corn silage. Rates of de novo lipogenesis, palmitate esterification, and glycerol and fatty acid release were determined with slices of subcutaneous adipose tissue at 0, 5, and 30 d after implantation. Condition and E2 interacted over time (linear, P less than .12; quadratic, P less than .05) to affect intake. Lean ewes implanted with E2 decreased intake initially after implantation, whereas obese ewes implanted with E2 decreased intake later after implantation. The linear effect of time x condition x E2 interacted (P less than .02) for lipogenesis. Lipogenesis was inhibited in both the lean and obese ewes implanted with E2. Lean compared with obese ewes without E2 had increased lipogenesis at a faster rate over time. Esterification increased (linear, P less than .01) in the lean ewes and decreased (quadratic, P less than .01) in the obese ewes over time. A time x E2 interaction occurred for esterification (P less than .02). Glycerol and fatty acid release were variable over time within condition and E2. A 48-h adipose tissue culture determined the effect of E2 on lipid metabolism. Estradiol-17 beta decreased (P less than .05) lipogenesis, decreased (P less than .08) esterification, and maintained fatty acid and glycerol release. Data in vivo and in culture indicated that E2 acted to decrease de novo lipogenesis and palmitate esterification and had little or no effect on lipolysis.     

β‐Carotene is incorporated or mobilized along with triglycerides in bovine adipose tissue in response to insulin or epinephrine

Pasture fed cattle ingest substantial amounts of β‐carotene (β‐C). Not all of the carotenoid compound is transformed into vitamin A, but the surplus is deposited in adipose tissue (AT). The mechanisms of β‐C incorporation and mobilization are unknown. Two experiments were conducted using explants from bovine AT cultured in vitro. First, β‐C incorporation by explants from three animals was examined with different β‐C concentrations (0, 1, 5 and 20 μm ) and different times of incubation (every 5 h up to 25 h). The data showed a significant increase of β‐C concentration in explants only for 20 μm β‐C. Secondly, effects of insulin and epinephrine on β‐C and triglyceride (TG) contents of explants were studied. Explants from six animals were incubated with either hormone and 0 or 20 μm β‐C for 20 h. Both TG and β‐C contents were affected positively by insulin and negatively by epinephrine. Interestingly, changes in ratios of β‐C/TG (hormone vs. control) were similar (1.7 × 10^?3 and 1.8 × 10^?3), respectively, for insulin and epinephrine, indicating that β‐C level is directly related to TG content. We also report the presence of mRNA for β‐C 15, 15′ oxygenase in bovine AT. The in vitro culture system using explants from bovine AT is a promising model to investigate factors that might affect the accumulation and metabolism of β‐C.     

Characterization of β-adrenergic receptors in bovine intramuscular and subcutaneous adipose tissue: comparison of lubabegron fumarate with β-adrenergic receptor agonists and antagonists

Chinese hamster ovary cell constructs expressing either the β ₁-, β ₂- or β ₃-adrenergic receptor (AR) were used to determine whether a novel β-AR modulator, lubabegron fumarate (LUB; Experior, Elanco Animal Health) might exert greater potency for a specific β-AR subtype. EC₅₀ values calculated based on cAMP accumulation in dose response curves indicate that LUB is highly selective for the β ₃-AR subtype, with an EC₅₀ of 6 × 10^–9 M, with no detectible agonistic activity at the β ₂-AR. We hypothesized that the accumulation of lipolytic markers would reflect the agonist activity at each of the β-receptor subtypes of the specific ligand; additionally, there would be differences in receptor subtype expression in subcutaneous (s.c.) and intrmuscular (i.m.) adipose tissues. Total RNA was extracted from adipose tissue samples and relative mRNA levels for β ₁-, β₂-, and β ₃-AR were measured using real-time quantitative polymerase chain reaction. Fresh s.c. and i.m. adipose tissue explants were incubated with isoproterenol hydrochloride (ISO; β-AR pan-agonist), dobutamine hydrochloride (DOB; specific β ₁-AA), salbutamol sulfate (SAL; specific β ₂-AA), ractopamine hydrochloride (RAC), zilpaterol hydrochloride (ZIL), BRL-37344 (specific β ₃-agonist), or LUB for 30 min following preincubation with theophylline (inhibitor of phosphodiesterase). Relative mRNA amounts for β ₁-, β ₂-, and β ₃-AR were greater (P < 0.05) in s.c. than in i.m. adipose tissue. The most abundant β-AR mRNA in both adipose tissues was the β ₂-AR (P < 0.05), with the β ₁- and β ₃-AR subtypes being minimally expressed in i.m. adipose tissue. ISO, RH, and ZH stimulated the release of glycerol and nonesterified fatty acid (NEFA) from s.c. adipose tissue, but these β-AR ligands did not alter concentrations of these lipolytic markers in i.m. adipose tissue. LUB did not affect glycerol or NEFA concentrations in s.c. or i.m. adipose tissue, but attenuated (P < 0.05) the accumulation of cAMP mediated by the β ₁- and β ₂-AR ligands DOB and SAL in s.c. adipose tissue. Collectively, these data indicate that bovine i.m. adipose tissue is less responsive than s.c. adipose tissue to β-adrenergic ligands, especially those that are agonists at the β ₁- and β₃-receptor subtypes. The minimal mRNA expression of the β ₁- and β ₃ subtypes in i.m. adipose tissue likely limits the response potential to agonists for these β-AR subtypes.     

Effects of ovine growth hormone and other anterior pituitary hormones on lipolysis of rat and ovine adipose tissue in vitro

Ovine growth hormone ( oGH ) was tested for its effects on lipolysis of rat and ovine adipose tissue in vitro. Ovine growth hormone at 1, 5 and 25 micrograms/ml stimulated lipolysis (P less than .05) of chopped rat adipose tissue and isolated rat adipocytes incubated in the presence of 100 mU/ml adenosine deaminase and .2 micrograms/ml dexamethasone, but had no effect on lipolysis of chopped ovine adipose tissue or isolated ovine adipocytes. Isoproterenol, a beta-adrenergic agonist, stimulated lipolysis (P less than .05) of both rat and ovine adipose tissue. Contaminants of the oGH preparation used were examined for lipolytic effects. Thyroid-stimulating hormone (TSH), luteinizing hormone (LH) and adrenocorticotropic hormone (ACTH) content in oGH were measured by radioimmunoassay. When quantities of these hormones contaminating 5 and 25 micrograms oGH were tested for lipolysis in rat adipose tissue, the TSH contamination could account for some (30%) of the lipolysis observed with oGH , while the other hormones had no effect. Also, preincubation of oGH with anti-GH, but not with anti-TSH or anti-LH, removed the principle in oGH responsible for the lipolytic effect on rat adipose tissue.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of alpha2-adrenergic receptor agonists on urine production in horses deprived of food and water

OBJECTIVE: To quantitate the dose- and time-related effects of IV administration of xylazine and detomidine on urine characteristics in horses deprived of feed and water. ANIMALS: 6 horses. PROCEDURE: Feed and water were withheld for 24 hours followed by i.v. administration of saline (0.9% NaCI) solution, xylazine (0.5 or 1.0 mg/kg), or detomidine (0.03 mg/kg). Horses were treated 4 times, each time with a different protocol. Following treatment, urine and blood samples were obtained at 15, 30, 60, 120, and 180 minutes. Blood samples were analyzed for PCV and serum concentrations of total plasma solids, sodium, and potassium. Urine samples were analyzed for pH and concentrations of glucose, proteins, sodium, and potassium. RESULTS: Baseline (before treatment) urine flow was 0.30 +/- 0.03 mL/kg/h and did not significantly change after treatment with saline solution and low-dose xylazine but transiently increased by 1 hour after treatment with high-dose xylazine or detomidine. Total urine output at 2 hours following treatment was 312 +/- 101 mL versus 4,845 +/- 272 mL for saline solution and detomidine, respectively. Absolute values of urine concentrations of sodium and potassium also variably increased following xylazine and detomidine administration. CONCLUSIONS AND CLINICAL RELEVANCE: Xylazine and detomidine administration in horses deprived of feed and water causes transient increases in urine volume and loss of sodium and potassium. Increase in urine flow is directly related to dose and type of alpha2-adrenergic receptor agonist. Dehydration in horses may be exacerbated by concurrent administration of alpha2-adrenergic receptor agonists.     

Histopathologic alterations induced in the lungs of sheep by use of alpha2-adrenergic receptor agonists

OBJECTIVE: To study effects of central- and peripheral-acting alpha2-adrenergic receptor agonists on lung parenchyma, platelets, and pulmonary intravascular macrophages (PIM) of sheep. ANIMALS: 12 healthy mature female sheep. PROCEDURE: Group-1 (control, n = 2) sheep received 5 ml of physiologic saline solution IV and were euthanatized 3 minutes later. Sheep of group 2 (n = 8) received xylazine (150 microg/kg of body weight, IV), then 2 sheep each were euthanatized 3, 10, or 60 minutes, or 12 hours later. Sheep (n = 2) of group 3 were given ST-91 (30 microg/kg, IV), then were euthanatized 3 minutes later. Immediately after euthanasia, the lungs were fixed intratracheally and tissue was obtained for light and electron microscopy after 1 hour. RESULTS: Pulmonary parenchymal damage or morphologic alterations in PIM and platelets were not evident in control sheep. Three minutes after xylazine administration, morphologic changes in PIM were appreciable. After 10 minutes, extensive damage to the capillary endothelium and alveolar type-I cells, intra-alveolar hemorrhage, and interstitial and alveolar edema were evident. Most PIM had complete internalization of the surface coat. Similar changes were seen 60 minutes after xylazine administration; however, by 12 hours, morphologic features of PIM and lung parenchyma were almost completely restored. Evidence of PIM activation, obvious damage to capillary endothelium, and extensive pulmonary edema also were evident 3 minutes after ST-91 administration. CONCLUSIONS: XYLAZINE induces severe pulmonary parenchymal damage when administered at clinical sedative doses in sheep; morphologic changes in PIM within 3 minutes after administration of these drugs are substantial; and platelet aggregation is not apparent.     

Response of anoestrous ewes pre-treated with a single dose of oestradiol-17 beta, or progesterone and oestradiol-17 beta, to the introduction of rams and ewes in oestrus

AIMS: To determine if the frequency distribution of time to oestrus observed after the introduction of rams, and additional ewes in oestrus, is affected by the administration of oestradiol- 17beta. To determine if the fertility following mating at the first induced oestrus in progesterone-primed ewes may be increased with administration of oestradiol-17beta. METHODS: In Experiment 1, ewes received 40 mug oestradiol- 17beta 3 (E3; n=61) or 5 (E5; n=56) days before the introduction of rams; 61 ewes were controls (C1). In Experiment 2, a controlled internal drug-releasing (CIDR) device was inserted in 95 ewes on Day -5 (Day 0 = introduction of rams). In addition, 47 ewes received oestradiol-17beta on Day 0 (CE), and the remaining 48 were controls (C2). In both experiments, additional ewes, brought into oestrus between Days 0 and 2 by hormonal treatments, were introduced with the rams. Onset of oestrus was estimated by visual observation of ewes marked by rams. Pregnancy status of ewes in Experiment 2 was determined using ultrasound, 30 days after oestrus. RESULTS: The total number of ewes detected in oestrus in Experiment 1 was similar between the three groups. The frequency distribution of ewes in oestrus in the E5 group differed from that of ewes in both C1 (p=0.05) and E3 (p<0.001). A similar number of ewes were detected in oestrus between Days 16-20, but the proportion of ewes in oestrus between Days 21-26 was greater for ewes in C1 than E5 or E3. Mean interval to the onset of oestrus was shorter in C1 than E3 which was shorter than E5 ewes (p<0.001). In Experiment 2, the total number of ewes in oestrus were similar between groups. Ewes in oestrus between Days 1-4 (65% and 64% for C2 and CE, respectively) and conception rates over the same period (81% and 80% for C2 and CE, respectively) were also similar between groups. CONCLUSIONS: Administration of oestradiol-17beta to anoestrous ewes altered the pattern of onset of oestrus, but it did not affect submission or conception rates, regardless of whether or not a short period of progesterone priming with a new CIDR device was included prior to the introduction of rams.     

Influence of the beta 2-adrenergic agonist clenbuterol on insulin-stimulated lipogenesis in mouse adipocytes

Growing mice fed the beta 2-adrenergic agonist clenbuterol (CB; 20ppm) had increased rate of growth and altered composition of gain (greater protein and less fat). Adipocytes prepared from the epididymal fat pads of treated and untreated mice were used to examine the influence of CB on lipid metabolism. Using cells from untreated mice, CB stimulated lipolysis to an equivalent maximum rate as epinephrine (EPI), but CB was far less potent (EC50 (microM); CB = 5, EPI = 0.2). Both CB and EPI inhibited insulin-stimulated lipogenesis over the physiological range of insulin concentrations. This inhibition was expressed as a dose-dependent decrease in tissue sensitivity to insulin and a decrease in maximal lipogenic capacity. Inhibition of maximal rate, but not of insulin sensitivity, could be stimulated by the addition of palmitate without EPI or CB. Adipocytes isolated from CB-treated mice did not differ from controls in sensitivity to insulin or in activity of fatty acid synthetase. Increased lipolysis and reduced lipogenesis as observed in vitro with CB are consistent with reduced fat accretion in CB-treated mice. However, the absence of detectable changes in adipocyte lipogenesis from CB-fed mice leaves open the question of the relevance of altered lipid metabolism to the observed changes in body composition.     

Effects of suckling, progestogen-impregnated pessaries or hysterectomy on ovarian function in autumn-lambing postpartum ewes

In three experiments, we examined the effects of suckling, progestogen treatment, hysterectomy or exogenous gonadotropin releasing hormone (GnRH) on ovarian function in autumn-lambing, postpartum ewes. In each experiment, GnRH was injected on approximately d 25 postpartum. Suckling reduced (P less than .01) GnRH-induced release of luteinizing hormone (LH) but not of follicle stimulating hormone (FSH), and reduced (P less than .05) the proportion of ewes that developed corpora lutea in response to GnRH. Suckling had no effect on duration (8.8 d) of GnRH-induced luteal phases. Progestogen prior to GnRH increased (P less than .01) the duration of the first luteal phase (10.1 vs 7.6 d; progestogen-treated ewes vs control ewes), but progestogen did not affect the release of LH or FSH. Progestogen treatment did not alter the interval from parturition to the first detected estrus (42.6 d). The concentration of 13,14-dihydro-15-keto-PGF2 alpha (PGFM) just after lambing was greater than 400 pg/ml of jugular plasma, but concentrations of PGFM declined thereafter. Hysterectomy the day after lambing hastened (P less than .001) the decline in concentrations of PGFM, indicating that prostaglandins from the postpartum uterus probably caused the high concentrations of PGFM in jugular plasma. Hysterectomy reduced (P less than .05) the interval from parturition to detectable luteal function (19.6 vs 25.3 d) and enhanced (P less than .001) luteal production of progesterone. This study of autumn-lambing ewes indicates that the uterus has a negative effect on ovarian function and that suckling and progestogen affect ovarian response to GnRH.     

Effect of bovine somatotropin and food deprivation on β-adrenergic and A1 adenosine receptor binding in adipose tissue of lactating cows

Lactating Holstein cows were used to assess the effect of bovine somatotropin (bST; n = 8) and fasting (FAST; n = 4) on ligand binding to β-adrenergic (BAR) and TYP e-1 adenosine (A₁R) receptors in adipose tissue. Cows received exogenous bST (sometribove; 40 mg/d) or no hormone (control) for 4 d in a single-reversal design with a 7-d interval between treatment periods. Subcutaneous adipose tissue biopsies were taken on day 4 of each treatment. Eight d after the bST regimen, 4 cows were fasted for 3 d and adipose biopsies were taken. Ligand binding was quantified with a postnuclear, total adipose tissue membrane preparation (100,000 × g pellet). Binding to BAR and A₁R was assessed with the antagonists [¹²⁵I]iodocyanopindolol (ICP) and [³H]8-cyclopentyl-1,3-dipropylxanthine (DCPCX), respectively. The binding affinity (K_d) of BAR for ICP was not affected by bST but was enhanced by FAST; maximal binding (B_max) was increased with bST treatment (P < 0.06) and reduced by FAST (61%, P < 0.01). K_d values for DCPCX binding to A₁R were not changed by bST or FAST. bST did not affect B_max for A₁R; however, FAST reduced the B_max by 38%. Data highlight the differential regulation of BAR and A₁R by bST and FAST.     

Effects of estradiol-17 beta, naloxone and gonadotropin releasing hormone on postpartum secretion of luteinizing hormone in fall-lambing ewes

The interaction among exogenous estradiol-17 beta, naloxone and gonadotropin releasing hormone (GnRH) in the control of luteinizing hormone (LH) secretion was studied in intact postpartum ewes nursing their offspring. One-half of 30 fall-lambing ewes were implanted subcutaneously with an estradiol-17 beta containing Silastic capsule between postpartum d 1 and 12 which doubled their serum concentrations of estradiol (16.0 +/- .1 vs 8.4 +/- .1 pg/ml). Blood samples were collected from implanted and non-implanted ewes at 15-min intervals for 5 h on d 3, 8, 13, 20 and 28 postpartum. Pre-injection samples were collected for 1 h, and ewes were injected with saline, naloxone (NAL;1 mg/kg) or GnRH (100 micrograms/ewe). When averaged across all days and implant groups, serum LH in the three post-NAL samples was higher (P less than .05) than in the three pre-NAL samples (3.6 +/- 1.2 vs .6 +/- .2 ng/ml). Post-GnRH concentrations of serum LH were lower (P less than .05) in estradiol-implanted ewes than in non-implanted ewes on d 8 and 13, but there were no differences in any LH characteristics on d 20 and 28 after implant removal on d 12. In non-implanted ewes, serum LH responses to GnRH increased (P less than .05) eightfold from d 3 (3.8 +/- 1.4 ng/ml) to d 8 (31.6 +/- 1.4 ng/ml), remained elevated through d 20, but declined by d 28 (10.8 +/- 1.4 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of exogenous estradiol-17 beta and progesterone on naloxone-reversible inhibition of the release of luteinizing hormone in ewes

The role of endogenous opioids in controlling luteinizing hormone (LH) secretion was studied by injecting the opioid antagonist naloxone into intact and ovariectomized ewes that were treated with estradiol-17 beta (E2) and progesterone (P4). The existence of a naloxone-reversible inhibition of LH release was examined in five experiments using a total of 52 mature ewes. Naloxone at a dosage of 1 mg/kg disinhibited release of LH and abruptly increased serum concentrations of LH in a variety of experimental models. This naloxone-reversible inhibition of LH secretion was apparent in all experimental models that involved P4-induced inhibition of basal LH secretion but not in one model in which P4 inhibited the LH surge. Specific effects of E2 on naloxone-reversible inhibition of LH varied among experimental models. When prolonged administration of P4 alone appeared to lose its LH-inhibitory potency, E2 restored inhibition of LH as well as the naloxone-reversible state. Whenever E2 acted synergistically to suppress basal LH secretion in models involving brief (5 d) exposure to P4, E2 appeared to antagonize the naloxone-reversible state. In summary, P4-induced suppression of LH secretion appeared to be mediated by endogenous opioids, but the apparent interaction of E2 and opioids in LH suppression varied among experiments.     

PCR detection and prevalence of alpha-, beta-, beta 2-, epsilon-, iota- and enterotoxin genes in Clostridium perfringens isolated from lambs with clostridial dysentery.

Clostridium perfringens isolated from lambs with dysentery (n=117) were analysed by a DNA amplification technique, the polymerase chain reaction (PCR), in order to determine the prevalence of the alpha-, beta-, beta 2-, epsilon-, iota- and enterotoxin genes. The most prevalent toxin type of C. perfringens found was type B, containing the alpha-, beta-, and epsilon-toxin genes, representing 46% of the cases with clostridial dysentery. C. perfringens type C containing the alpha-, and beta-toxin genes was isolated in 20% and type D, which is characterized by the alpha- and epsilon-toxin genes, was isolated in 28% of all isolates. The recently discovered, not yet assigned beta 2-toxigenic type of C. perfringens was represented in 6% of all isolates. No C. perfringens type A containing the alpha-toxin alone and no type E, which harbours the ADP-ribosylating iota-toxin, were found in the diseased animals. None of the samples contained the enterotoxin gene. Only one type of C. perfringens was found in a given herd, revealing the epidemiological use of PCR toxin gene typing of C. perfringens. The animals originated from 79 different herds with sizes ranging from 30 to 250 animals, bred in the area of northern Greece.     

Preliminary observations on expression of transforming growth factors beta1 and beta3 in equine full-thickness skin wounds healing normally or with exuberant granulation tissue

OBJECTIVE: To determine whether transforming growth factor (TGF)-beta1 and -beta3 expression differs between equine limb wounds healing normally and those healing with experimentally induced exuberant granulation tissue (EGT). STUDY DESIGN: Six wounds were created on the lateral aspect of both metacarpi of each horse; one forelimb was untreated, and the other was bandaged to stimulate the development of EGT. Sequential wound biopsies allowed comparison of growth factor expression between the two types of wound. ANIMALS: Four horses (2 to 4 years of age; 350 to 420 kg). METHODS: Wounds were assessed grossly, histologically, and by enzyme-linked immunosorbent assay (ELISA) for TGF-beta1 and -beta3 expression at 12 and 24 hours and 2, 5, 10, and 14 days postoperatively. RESULTS: Bandaged wounds developed EGT. In all wounds, TGF-beta1 peaked early and remained elevated at 14 days. Peak TGF-beta1 concentration was higher in wounds with EGT, but not significantly so. Expression of TGF-beta3 differed from TGF-beta1, with peak TGF-beta3 concentrations being delayed. Concentrations of TGF-beta3 were higher in wounds healing normally, but this difference was not significant. CONCLUSIONS: During both normal and exuberant wound repair, the expression of TGF-beta1 occurred earlier than TGF-beta3 expression. Wounds healing with EGT tended to have higher concentrations of fibrogenic TGF-beta1 and lower concentrations of antifibrotic TGF-beta3 than wounds healing normally, although these differences were not statistically significant. CLINICAL RELEVANCE: This study suggests that the production of EGT in bandaged wounds may be related to increased expression of fibrogenic TGF-beta1 and decreased expression of antifibrotic TGF-beta3. Further investigation of the roles of TGF-beta1 and -beta3 may be important in understanding the molecular control of EGT in horses.