Non-specific immunity and ketone bodies. I: In vitro studies on chemotaxis and phagocytosis in ovine neutrophils

The in vitro effects of the ketone bodies beta-OH-butyrate (2.4 or 4.8 mmol/l) and acetoacetate (2.4 or 4.8 mmol/l) on the uptake of latex particles (1.09 microns) and chemotaxis were investigated in ovine neutrophils. Because the acetoacetate used was a lithium salt, the effect of 2.4 or 4.8 mmol/l lithium chloride was also tested. Neutrophils from eight non-lactating, non-pregnant ewes were studied. The uptake of latex particles, as measured by a spectrophotometric method, showed wide individual variation. The phagocytotic activity was unaffected by 2.4 mmol/l ketone bodies and LiCl, but it was significantly inhibited by 4.8 mmol/l beta-OH-butyrate and activated by 4.8 mmol/l LiCl. The latter result could be masking an inhibitory effect of acetoacetate. Chemotactic movements of neutrophils, as evaluated in a modified Boyden chamber using homologous zymosan-activated serum (ZAS) as a chemoattractant, were slightly but significantly reduced by a 2.4 mmolar concentration of the ketone bodies, administered singly or simultaneously, and by LiCl. We conclude therefore that the inhibitory effect of lithium-acetoacetate could be due to its lithium component. The 4.8 mmol/l dose of acetoacetate and beta-OH butyrate significantly decreased chemotaxis only when both compounds were added simultaneously. No effect of 4.8 mmol/l LiCl was observed. These results suggest that ketone bodies, in particular beta-OH butyrate, could directly influence particle uptake and chemotaxis in neutrophils. Although other factors could decrease the efficiency of the immune system in ketotic ruminants, the effects of the ketone bodies on neutrophils functions may explain the high frequency of infectious disease during 'ketotic syndrome'. The immunomodulatory effect of lithium needs to be evaluated further and it should be considered when testing lithium compounds.     

The influence of ketone bodies and glucose on interferon, tumor necrosis factor production and NO release in bovine aorta endothelial cells

Bovine aorta endothelial cells (BAECs) were used to determine the effect of ketone bodies and glucose on in vitro interferon (IFN), tumor necrosis factor (TNF) and nitric oxide (NO) production. BAECs were incubated for 4 and 24h with the ketone bodies: 3.8mmol/l beta-hydroxybutyrate (BHB), 1mmol/l acetoacetate (AcAc) and 5. 2mmol/l acetone (Ac), used separately or in a mixture together with cytokine inducers: Newcastle disease virus (NDV) and lipopolysaccharide (LPS). BHB alone (but not AcAc or Ac) and a mixture of ketone bodies caused a significant decrease in IFN titers induced by NDV and LPS and in TNF titers induced by LPS. Glucose used at concentrations of 5.55, 3.33 and 1.66mmol/l did not influence cytokine production.NO measured by the nitrite content in culture medium was released spontaneously from BAECs. A slight enhancement of NO release was observed after infection of BAECs with NDV; however, treatment with LPS caused inhibition of the release. The mixture of ketone bodies used with NDV or LPS enhanced NO release. However, when cells were incubated in the medium with 1. 66mmol/l glucose (mimicking low plasma glucose level in ketotic cows) a significant decrease in NO release was observed. This enhancing effect of ketone bodies and inhibition by low glucose in the final effect balanced each other, and the amounts of NO released in the medium with 1.66mmol/l glucose and with the mixture of ketone bodies resembled those produced at 3.33mmol/l glucose without ketone bodies. The significance of these effects of ketone bodies and glucose concentrations on cytokine and NO production in the immunity of ketotic cows has been discussed.     

Relationship between concentration of citrate and ketone bodies in cow's milk

The authors' hypothesis is that the members of the tricarboxylic acid cycle (TCA cycle) such as citrate decrease in association with increased ketone body formation. To prove this hypothesis the connection between ketone bodies and citrate formation of milk was studied. A fluorimetric method was used to determine citrate and a headspace sampling gas chromatographic (GC) method was developed for determination of ketone bodies. Under real conditions of milk sampling, transport and storage, preserved milk samples of 119 clinically healthy dairy cows obtained in the 48 hours after milking were investigated. A low level of acetoacetate (ACAC) was found in all samples. This fact can be explained by the spontaneous decarboxylation of acetoacetate during sample storage (previously decarboxylised acetoacetate = pdACAC) and, consequently, the majority of the amount of acetoacetate in the samples (AC + pdACAC) appeared in the measured acetone concentrations. Based on the measured acetone concentration of milk samples two groups were formed retrospectively: HA (high-acetone) group (n = 41) with an AC + pdACAC concentration of > 0.4 mmol/l and a LA (low-acetone) group (n = 78) with an AC + pdACAC level of < or = 0.4 mmol/l. In the milk of cows of Group HA a positive correlation (r = +0.623) and linear connection between acetone (AC + pdACAC) and beta-hydroxybutyrate (BOHB) levels was found [BOHB = 2.491 + 0.586 x (pdAC + ACAC)]. Furthermore, in this group a negative correlation between citrate and BOHB and AC + pdACAC was also established (r = -0.579). Focusing on the results of this group the authors found a significant drop of AC + pdACAC and citrate during the metabolically critical first 1-4 weeks of lactation. For this reason they suggest that simple, easy, automated methods (i.e. flow injection analysis, Fourier transformation infrared analysis) should be introduced for the simultaneous determination of acetone and citrate concentration in milk to make the evaluation of the energy status of high-producing dairy cows easier and more certain.     

Pharmacokinetics and toxic effects of lithium chloride after intravenous adminstration in conscious horses

OBJECTIVE: To determine the pharmacokinetics and toxic effects associated with IV administration of lithium chloride (LiCl) to conscious healthy horses. ANIMALS: 6 healthy Standardbred horses. PROCEDURE: Twenty 3-mmol boluses of LiCl (0.15 mmol/L) were injected IV at 3-minute intervals (total dose, 60 mmol) during a 1-hour period. Blood samples for measurement of serum lithium concentrations were collected before injection and up to 24 hours after injection. Behavioral and systemic toxic effects of LiCl were also assessed. RESULTS: Lithium elimination could best be described by a 3-compartment model for 5 of the 6 horses. Mean peak serum concentration was 0.561 mmol/L (range, 0.529 to 0.613 mmol/L), with actual measured mean serum value of 0.575 mmol/L (range, 0.52 to 0.67 mmol/L) at 2.5 minutes after administration of the last bolus. Half-life was 43.5 hours (range, 32 to 84 hours), and after 24 hours, mean serum lithium concentration was 0.13+/-0.05 mmol/L (range, 0.07 to 0.21 mmol/L). The 60-mmol dose of LiCl did not produce significant differences in any measured hematologic or biochemical variables, gastrointestinal motility, or ECG variables evaluated during the study period. CONCLUSIONS AND CLINICAL RELEVANCE: Distribution of lithium best fit a 3-compartment model, and clearance of the electrolyte was slow. Healthy horses remained unaffected by LiCl at doses that exceeded those required for determination of cardiac output. Peak serum concentrations were less than steady-state serum concentrations that reportedly cause toxic effects in other species.     

Ketone measurements using dipstick methodology in cats with diabetes mellitus

O bjectives : To compare the results of urine and plasma ketone dip test in a group of diabetic cats with possible ketosis or ketoacidosis, using laboratory plasma β-hydroxybutyrate measurements as the gold standard.
M ethods : According to clinical examinations, plasma β-hydroxybutyrate measurements and venous blood gas analysis, 54 cats with diabetes mellitus were classified as non-ketotic (n=3), ketotic (n=40) or ketoacidotic (n=11). Plasma and urine acetoacetate concentrations were determined using urine reagent strips.
R esults : Although there was a significant positive correlation between blood and urine ketone measurements (r=0·695, P<0·001), the results differed significantly (Z=−3·494, P<0·001). Using the differential positive rates, the best cut-off value to detect cats with ketoacidosis was 1·5 mmol/l for urine and 4 mmol/l for plasma. The sensitivity/specificity was 82/95 per cent for urine and 100/88 per cent for plasma, respectively.
C linical S ignificance : The urine and plasma ketone dip tests have a different diagnostic accuracy, and results have to be interpreted differently. Because of its high sensitivity, the plasma ketone dip test performs better than the urine ketone dip test to identify cats with impending or established ketoacidosis.     

Beta-hydroxybutyrate levels in peripheral blood and ketone bodies supplemented in culture media affect the in vitro chemotaxis of bovine leukocytes.

The role of ketone bodies on chemotactic capacities of leukocytes was characterized in two experiments. Experiment I was performed to investigate the association between serum beta-hydroxybutyrate concentrations (BHB) and in vitro chemotaxis of leukocytes. Cows were divided into low-BHB, medium-BHB, and high-BHB ones and classified according to their BHB. Leukocytes from high-BHB cows had a significantly lower chemotactic differential than leukocytes from low-BHB cows (p < 0.01). The effect of adding ketone bodies into in vitro chemotaxis cultures on leukocytes chemotaxis was studied in Experiment II. Either individual or a combination of commercial ketone bodies - sodium salts of BHB (BHBA), lithium salt of acetoacetate (ACAC), and acetone (Acetone) - were diluted in culture media and divided into eight concentrations corresponding to concentrations of bovine subclinical and clinical ketosis. For leukocytes from medium- and high-BHB cow, the chemotactic indexes of leukocytes were reduced by ACAC and Acetone. Chemotactic differentials of cultures with ACAC and acetone supplementation from both sources of leukocytes were significantly lower than that of the control culture (p < 0.05). For leukocytes from high-BHB cows, chemotactic indexes were suppressed in a ketone-body environment. In conclusion, leukocytes from naturally-occurring ketotic cows have lower chemotactic differentials than those from non-ketotic cows, and a chemotactic capacity indicated by a chemotactic differential is impaired when leukocytes migrate in an environment with ketone bodies in vitro.     

Plasma glucose, ketone bodies, insulin, glucagon and enteroglucagon in cows: diurnal variations related to ketone levels before feeding and to the ketogenic effects of feeds

Ingestions of a moderately ketogenic silage twice daily were followed by transient increments in plasma insulin and ketone bodies and decreases in plasma glucose. Ketone bodies and glucose were negatively correlated throughout the day, but the insulin elevations culminated before the maximal effects on ketone bodies and glucose were established. Cows with varying glucose levels before morning feeding reacted to a highly ketogenic silage by decreasing their glucose level uniformly to about 3 mmol/l, in spite of a widely varying feeding-induced insulin increment. Hay-feeding caused insulin increments of the same magnitude as silage-feeding, but the glucose decrease and the ketone increment was much smaller. The results indicate some direct action of ketone bodies on blood sugar regulation, in addition to effects mediated by insulin. The role of ketone bodies as the insulinotropic factor was not confirmed. The insulin level after feeding seems to be determined by the carbohydrate status of the animal before feeding. No significant changes in plasma glucagon were observed after feeding, and no consistent differences in plasma levels of this hormone were found when non-ketonemic, ketonemic, and clinically ketotic cows were compared. The plasma level of enteroglucagon (GLI) was positively correlated to the relative amount of concentrates consumed, but no relation to plasma glucose was found.     

Agonist-induced adherence of equine neutrophils to fibronectin- and serum-coated plastic is CD18 dependent.

Adherence to vascular endothelium and extracellular matrix proteins is a pre-requisite for neutrophil accumulation at sites of inflammation. In this study, equine neutrophil adherence to fibronectin and autologous serum-coated plastic in response to PAF, hrIL-8, hrC5a and PMA has been measured. In addition, the mechanisms involved have been investigated using monoclonal antibodies (MoAbs) against the beta2 integrin CD18. PAF and hrC5a caused similar, concentration dependent, increases in adherence to fibronectin- and serum-coated plastic (maximum responses 19 +/- 4% and 19 +/- 3% for PAF and 15 +/- 4% and 16 +/- 2% for hrC5a on fibronectin- and serum-coated plastic, respectively). Adherence in response to PMA, although not reaching a maximum over the time course studied, was of a similar magnitude on the two surfaces (41 +/- 1% and 38 +/- 2% with 10(-7) M PMA on fibronectin- and serum-coated plastic, respectively). In contrast, the maximum adherence caused by hrIL-8 was significantly lower on fibronectin- than on serum-coated plastic (9 +/- 3% vs. 17 +/- 2%; 10(8) x M hrIL-8). Pre-incubation with MoAbs against CD18 (H20A and 6.5E) caused concentration related inhibition of stimulus-induced adherence to both fibronectin- and serum-coated plastic. Equine neutrophil adherence in response to PAF, hrIL-8, hrC5a and PMA therefore appears to be mediated by a CD18 dependent mechanism.     

The Stability and Automatic Determination of Ketone Bodies in Blood Samples Taken in Field Conditions

An automated, spectro-photometric determination of blood acetoacetate and β-hydroxybutyrate was developed with a Gilford 3500 autoanalyzer. The stability of ketone bodies was studied in different conditions. An immediate precipitation with 0.6 M perchloric acid and cooling the sample effectively prevent the loss of acetoacetate from samples during transport to the laboratory (at 4°C a 6 % loss of acetoacetate was noted during 24 h). Freezing the sample makes it practically stable (less than 2 % loss of acetoacetate per week during a study lasting 2 months). At room temperature (20°C) the sample’s acetoacetate was instable and disappeared with a rate of 6 % per h. β-hydroxybutyrate was stable in precipitated samples. Because the precipitation also retains the sample’s glucose, 3 main parameters for the indication of ketosis could be analyzed automatically from the same sample with a total capacity of 40 samples in 2½ h.     

Growth hormone release induced by an amino acid mixture from primary cultured anterior pituitary cells of goats

The effects of amino acids on growth hormone (GH) release and cytosolic calcium concentration ([Ca²⁺]_i) were investigated in caprine anterior pituitary cells cultured for 3 d in Dulbecco modified Eagle medium. The addition of an amino acid mixture consisting of seven nonessential amino acids (NEAA: l-Asp, Gly, l-Ala, l-Ser, l-Pro, l-Asn, and l-Glu; concentration of each 12.5–200 μmol/l) in the medium significantly raised GH release from the cultured cells in a concentration-dependent manner with the maximum release at 200 μmol/l NEAA. Although an addition of l-Asp (0.1–100 μmol/l) caused a significant rise in GH release in a concentration-dependent manner, neither the individual amino acids contained in NEAA except l-Asp nor others (l-Leu, l-Phe, l-Gln, l-Met, and l-Arg) caused a rise in GH release when added alone to the medium. The rise in GH release induced by NEAA (200 μmol/l) and GH-releasing hormone (GHRH, 10 nmol/l) was significantly reduced by the addition of EGTA (l.8 mmol/l) and nifedipine (1 μmol/l) to the medium, respectively. The addition of NEAA (200 μmol/l) caused a rapid and transient [Ca²⁺]_i increase, followed thereafter by a steady increase. The prior addition of nifedipine (1 μmol/l), which itself significantly reduced the basal [Ca²⁺]_i, completely abolished the response induced by NEAA or GHRH. From these findings, we conclude that: 1) NEAA raises GH release and [Ca²⁺]_i in cultured caprine anterior pituitary cells, and 2) Ca²⁺ influx from the medium may be responsible for the cellular action of NEAA.     

Cell surface hydrophobicity of Actinomyces pyogenes determined by hexadecane adherence- and salt aggregation studies.

Cell surface hydrophobicities of Actinomyces pyogenes were determined by measuring the adherence of the bacteria to hexadecane droplets and by salt aggregation tests. Among 42 A. pyogenes cultures tested 25 (60%) adhered strongly (adherence greater than or equal to 75%) and 17 (40%) less pronounced (adherence between 25-75%) to the hexadecane droplets. Pre-treatment of the bacteria with proteolytic enzymes completely eliminated the adherence properties whereas heat treatment had no effect. The salt aggregation studies revealed that 4 (10%) cultures aggregated in ammonium sulfate solutions of a molarity of 0.05 mol/l, 5 (12%), 14 (33%) and 3 (7%) cultures in ammonium sulfate solutions with molarities of greater than or equal to 1.5 mol/l, greater than or equal to 3 mol/l and greater than or equal to 4.5 mol/l, respectively. No aggregation at all could be observed with 16 (38%) of the cultures. Pronase treatment completely eliminated the salt aggregation reactions, trypsin- and heat treatment had no effect. The results from hexadecane adherence and salt aggregation did not correspond. The differences in surface hydrophobicities, possibly related to adherence properties of A. pyogenes, could be used for epidemiological typing of individual cultures of this bacterial species.     

Suppressing effects of short-chain fatty acids on growth hormone (GH)-releasing hormone-induced GH release in isolated anterior pituitary cells of goats.

The involvement of tetrodotoxin-sensitive Na+ channels and receptor-operated nonspecific Ca2+ channels, and the effects of short-chain fatty acids, on growth hormone (GH) release induced by GH-releasing hormone (GHRH) were investigated in cultured and freshly isolated caprine anterior pituitary cells. In 3-d cultured cells in Dulbecco's modified Eagle's medium, an increase in GH release induced by GHRH (10 nmol/l) was moderately, but significantly, reduced by a voltage-sensitive Na+ channel antagonist tetrodotoxin (1 micromol). The GHRH-induced GH increase, which was not affected by a simultaneous addition of a receptor-operated nonspecific Ca2+ channel antagonist tetramethrine (0.1 mmol/l), was significantly reduced by a voltage-sensitive L-type Ca2+ channel antagonist nifedipine (1 micromol/l). Propionate and butyrate at 10 mmol/l, however, not only suppressed basal GH release but also significantly reduced the GH increase induced by 10 nmol/l of GHRH. The inhibitory action of these acids was also reproduced by an addition of beta-hydroxy butyrate (10 mmol/l) and octanoate (10 mmol/l). In freshly isolated and perifused cells, butyrate (10 mmol/l) as well as somatostatin (100 nmol/l) significantly reduced the GH increase induced by GHRH. From these findings we conclude that tetrodotoxin-sensitive Na+ channels and voltage-dependent L-type Ca2+ channels are involved in the cellular mechanism for GHRH-induced GH release, and that short-chain fatty acids such as propionate and butyrate have a direct action on somatotrophs to reduce basal and GHRH-induced GH release, in caprine somatotrophs.     

Pharmacokinetics of lithium in sheep

Lithium was administered to adult sheep to estimate its pharmacokinetic parameters and a suitable dosage for chronic psychopharmacological experiments. The triexponential decline of plasma lithium levels was tentatively interpreted with a three-compartment, open-model of distribution. Sheep differed from other species by the following features: high faecal excretion of lithium, recycling of lithium through the salivary secretion and low absorption rate of the orally-administered drug. A chronic oral administration of 1 mmol/kg once daily provided minimal plasma levels of about 1 mmol/l, without toxic side-effects. The results are discussed with respect to comparative pharmacokinetics of lithium and its potential use in the treatment of behavioral disorders of farm animals.     

Detection of ketosis in dairy cows by analysis of exhaled breath

Summary

In four healthy cows an elevation of ketone bodies was induced by reduction of feed intake. Two cows became clearly ketotic while the other two cows showed only slight increases in ketone body concentrations in serum and milk. Acetone concentrations in exhaled breath were measured by gas chromatography combined with mass spectrometry. These values were correlated with concentrations of serum β‐hydroxybutyric acid (r=0.81) and milk acetoacetate+acetone (r=0.70). It is concluded that the ketotic state of dairy cows can be detected by analysis of exhaled breath. This offers a potential non‐invasive method of determining the metabolic state of dairy cows.     

The role of insulin, glucagon, dexamethasone, and leptin in the regulation of ketogenesis and glycogen storage in primary cultures of porcine hepatocytes prepared from 60 kg pigs

A study was conducted to elucidate hormonal control of ketogenesis and glycogen deposition in primary cultures of porcine hepatocytes. Hepatocytes were isolated from pigs (54-68 kg) by collagenase perfusion and seeded into collagen-coated T-25 flasks. Monolayers were established in medium containing fetal bovine serum for 1 day and switched to a serum-free medium for the remainder of the culture period. Hepatocytes were maintained in DMEM/M199 containing 1% DMSO, dexamethasone (10(-6) or 10(-7) M), linoleic acid (3.4 x 10(-5) M), and carnitine (10(-3) M) for 3 days. On the first day of serum-free culture, insulin was added at 1 or 100 ng/ml and glucagon was added at 0, 1, or 100 ng/ml. Recombinant human leptin (200 ng/ml) was added during the final 24 h; medium and all cells were harvested on the third day. Concentrations of acetoacetate and beta-hydroxybutyrate (ketone bodies) in media and glycogen deposition in the cellular compartment were determined. Ketogenesis was highly stimulated by glucagon (1 and 100 ng/ml) and inhibited by insulin. In contrast, glycogen deposition was stimulated by insulin and attenuated by glucagon; high insulin was also associated with a reduction in the ketone body ratio (acetoacetate:beta-hydroxybutyrate). High levels of dexamethasone stimulated ketogenesis, but inhibited glycogen deposition at low insulin. Culture of cells with leptin for 24 h, over the range of insulin, glucagon, and dexamethasone concentrations had no effect on either glycogen deposition or ketogenesis. These data suggest that while adult porcine hepatocytes are indeed sensitive to hormonal manipulation, leptin has no direct influence on hepatic energy metabolism in swine.     

Concentrations of metabolites and hormones in the blood plasma of lactating dairy cows during treatment with recombinant bovine somatotropin (BST)]

Sixteen lactating dairy cows were submitted to six injections in a four-week interval of either 640 mg prolonged release bST or of 640 mg saline (control, n = 8). The bST-treated animals were pair fed to the control cows, which were fed according to requirements. Fourteen days after each injection, blood samples were derived and analysed for metabolites and hormones. Plasma contents averaged 0.16 mmol/l non esterified fatty acids (NEFA), 2.69 mmol/l glucose, 0.43 mmol/l beta-hydroxybutyrate, 0.024 mmol/l acetoacetate, 78.8 g/l protein, 4.6 mmol/l urea, 0.81 micrograms/l insulin, 44.4 ng/l thyroxine (T4) and 1.41 ng/l triiodothyronine (T3). The bST-application was without effect on the analysed plasma contents.     

Effect of ketone bodies on crop emptying in the chicken

1. The effect of ketone bodies on crop emptying was studied in chickens in 2 experiments. In the first, the effect of β‐hydroxybutyrate, acetoacetate or acetone on relative crop content was measured. The effects of dietary medium and long chain triacylglycerols upon serum β‐hydroxybutyrate were investigated in the second.

2. β‐Hydroxybutyrate, but not acetoacetate and acetone, delayed crop emptying in a dose dependent fashion. Serum β‐hydroxybutyrate concentration was high in chicks given medium chain triacylglycerol, when compared with long chain triacylglycerol.

3. The results suggest that delayed crop emptying induced by medium chain triacylglycerol could partly be explained by an enhanced concentration of serum β‐hydroxybutyrate, which is the result of the rapid oxidation of medium chain fatty acids.     

Biochemical blood profile in 20 female veiled chameleons (Chamaeleo calyptratus) Aged 7, 9 and 11 Months

BackgroundVeiled chameleons (Chamaeleo calyptratus) are more frequently presented to veterinary clinics due to reproductive diseases that lead to high morbidity, especially in captive-bred females. The objective of this study was to evaluate blood biochemical profiles of healthy, young captive females veiled chameleons aged seven, nine and eleven months.MethodsBlood samples were taken from 20 female veiled chameleons at the age of 7, 9 and 11 months. The biochemical profile was analyzed using the VetScan VS2 analyzer with an Avian/Reptile Profile rotor. The ionized calcium (iCa) concentration was measured by i-STAT analyzer with a CHEM+8 cartridge.ResultsMean concentration of glucose (15.8 ± 1.3 mmol/l) and uric acid (244.3 ± 143.2 µmol/l) at the age of seven months were significantly higher than mean concentrations of glucose (12.8 ± 16.5 mmol/l) and uric acid (134.9 ± 113.2 µmol/l) at nine months of age and mean concentration of glucose (13.2 ± 0.9 mmol/l) and uric acid (129.4 ± 109.2 µmol/l) at 11 months of age, respectively. Total calcium (4.2 ± 0.8 mmol/l) and ionized calcium (1.51 ± 0.16 mmol/l) were significantly higher at seven months of age compare to nin months. Mean activity of aspartate aminotransferase at nine months (6.2 ± 2.3 µkat/l) was significantly higher (P=0.004) than the mean activity of aspartate aminotransferase at seven months of age (4.7 ± 1.0 µkat/l). Concentrations of potassium at eleven months (8.3 ± 2.1 mmol/l) were significantly higher (P ˂ 0.001) than the mean concentration of potassium at seven months (5.4 ± 1.3 mmol/l) and also at ninth months (6.1 ± 1.6 mmol/l). No significant differences were found among the mean concentrations of total protein (57.5 ± 14.6, 61.9 ± 9.2, 59.6 ± 12.2 g/l), albumin (32.8 ± 5.4, 33.6 ± 4.1, 33.4 ± 7.1 g/l), globulin (27.4 ± 5.4, 26.6 ± 4.3, 26.3 ± 5.4 g/l) and mean activity of creatine kinase (25.4 ± 10.3, 30.1 ± 21.0, 25.6 ±13.2 µkat/l) at any of the ages.Conclusions and clinical relevanceIn all females, ovarian activity was already evident in the first part of monitoring, and the measured values could be interpreted as values for young female veiled chameleons during active ovarian activity.     

Morphological Quality of Oocytes and Blood Plasma Metabolites in Repeat Breeding and Early Lactation Dairy Cows

The objectives of the study were to evaluate the morphological quality of oocytes in repeat breeder and early lactation cows and to determine the possible associations between the quality of oocytes and a range of blood metabolites. Oocyte quality and a range of metabolites were compared between 29 repeat breeder and 13 early lactation cows. The yield of oocytes from the repeat breeders was lower than that from the early lactation cows (4.4 ± 0.2 vs 5.4 ± 0.6, p < 0.05). Percentages of abnormal oocytes for the repeat breeders and the early lactation cows were 52.5% and 37.9%, respectively (p < 0.001). An excess of abnormal oocytes to normal was found in 55.2% of the studied repeat breeders (65.8% vs 34.2%, p < 0.05). Total protein, glucose and aspartate aminotransferase did not differ (p > 0.05) between the repeat breeders with an excess of abnormal oocytes (81 ± 1.0 g/l, 3.5 ± 1.0 mmol/l and 68.5 ± 3.7 U/l), those with the prevalence of normal oocytes (84 ± 1.0 g/l, 3.6 ± 0.1 mmol/l and 73.2 ± 3.5 U/l) and the early lactation cows (83 ± 2.0 g/l, 3.7 ± 0.1 mmol/l and 74.5 ± 3.6 U/I). The repeat breeders with an excess of abnormal oocytes had higher (p < 0.05) urea (5.2 ± 0.2 mmol/l) level than in those with the prevalence of normal oocytes (4.8 ± 0.2 mmol/l) and the early lactation cows (4.7 ± 0.2 mmol/l). A trend for higher total cholesterol and lactate dehydrogenase activity was found in the repeat breeders with an excess of abnormal oocytes. In conclusion, it is suggested that possible causes of repeat breeding in dairy cows may include impaired oocytes. An excess of abnormal oocytes in the repeat breeder cows was associated with elevated blood plasma levels of urea.     

Rumen fermentation with special reference to volatile fatty acid production during reproduction. 1. Methods and results of volatile fatty acid levels and production rates in vitro

The target of the investigations was to register part of the synthesis performance in the rumen of ewes during the complete reproduction cycle. With the help of in-vitro experiments with artificial rumen the concentration and production rates of volatile fatty acids in the rumen fluid were measured. From the experiments the measured data of fertility-accentuated crossbreeding (experiment 1) and crossbreeding (experiment 2) ewes were contrasted. The average concentrations of volatile fatty acids in ewes are 95.4 m mol/l in experiment 1 and 109.2 m mol/l in experiment 2 during the early stage gestation, 121.4 m mol/l in experiment 1 and 99.8 m mol/l in experiment 2 in the last stage stage of gestation, 129.6 m mol/l in experiment 1 and 112.8 m mol/l in experiment 2 during lactation and 106.6 m mol/l in experiment 1 and 112.9 m mol/l in experiment 2 during the dry period. The production rates of volatile fatty acids calculated form their concentration amount to 4.8 m mol/l X h in experiment 1 and 2.6 m mol/l X h in experiment 2 in the early stage of gestation, 3.5 m mol/l X h in experiment 1 and 3.1 m mol/l X h in experiment 2 in the last stage of gestation, 3.2 m mol/l X h in experiment 1 and 2.7 m mol/l X h in experiment 2 during lactation and 3.7 m mol/l X h in experiment 1 and 2.9 m mol/l X h in experiment 2 in the dry period. The correlation between the concentration and the production rate of volatile fatty acids is not significantly negative in either of the ewe experiments. The scattering of the individual values is wide so that the individual influence of the test animals as well as the influence of the in-vitro method used permit the conclusion that a significant statement on the influence of the genotype and the stage of reproduction on the production rates cannot be made.