Prediction of calving time in Holstein dairy cows by monitoring the ventral tail base surface temperature.

Dystocia adversely affects the health of calves and their dams. The aim of this study was to determine whether the ventral tail base surface temperature (ST) could be used to predict calving time in dairy cows. Pregnant Holstein cows were enrolled during the warm season (daily average air temperature 10-20°C; n=13) and cool season (daily average air temperature<10°C; n=22) in Hokkaido, Japan, and a wearable wireless ST sensor was attached to the surface of the ventral tail base of each cow 9-12days before the predicted calving date. The ventral tail base ST was measured every 2min until 24h after calving. Hourly maximum ventral tail base ST values were used in the analysis and changes in ventral tail base ST were expressed as residual temperatures (RTs) to exclude any circadian effects using the formula: RT=actual ST-mean ST for the same hour on the previous 3 days. In both seasons, there was a continual decrease in ventral tail base RT from approximately 24h before calving compared with the control ventral tail base RT from 120 to 97h before calving. The areas under the receiver operating characteristic curves (ROC-AUCs) for ventral tail base RT as a predictor of calving were 0.88-0.95. ROC-AUCs as a predictor of calving within 24h were higher in the warm season than in the cool season. These findings demonstrate that calving time in dairy cows can be predicted by monitoring ventral tail base ST with a wearable wireless sensor, but seasonal variability affects the accuracy of prediction of calving time.

No change in apoptosis in skeletal muscle exposed acutely or chronically to alcohol.

The pathogenic mechanisms responsible for the deleterious changes in ethanol-exposed skeletal muscle are unknown, although apoptosis may be a causal process. We therefore investigated the responses of skeletal muscle to acute or chronic ethanol exposure in male Wistar rats. In acute studies, rats were dosed with ethanol (75 mmol (3.46 g)/kg BW) and killed after either 2.5 or 6 hours. In chronic studies, rats were fed ethanol as 35% of total dietary energy for 6 weeks. Apoptosis was determined by either DNA fragmentation or TUNEL (terminal deoxynucleotidyl transferase mediated dUTP nick end labelling) assays. The results showed that apoptosis was not increased in the ethanol-exposed muscle in both acute and chronic studies compared to appropriate controls.

Alcohol increases c-myc mRNA and protein in skeletal and cardiac muscle.

The pathogenic mechanisms responsible for alcohol-induced muscle disease are unknown, although it is possible that increased proto-oncogene expression may be the causative process. Therefore, we investigated the responses of skeletal muscle c-myc protein and mRNA to a standard acute ethanol dosage regimen (75 mmol/kg/body weight [BW]) for 2.5 to 24 hours. Comparative studies were made on the heart. Acute ethanol administration in vivo led to an increase in c-myc proto-oncogene mRNA in rat skeletal and cardiac muscle. The changes in c-myc mRNA were mirrored by increases in the c-myc protein as demonstrated by immunohistochemistry. The changes in the c-myc protein were localized to the myonuclei, with no corresponding changes seen in the interstitial cell nuclei. This is the first report of altered proto-oncogene expression in muscle in response to ethanol. Increased c-myc mRNA and protein may reflect adaptive changes, a stress response, or another uncharacterized cellular adaptation.

Oestrone sulfate commences an increase around 50 days of gestation in bovine peripheral blood.

The objective of this study was to determine the peripheral plasma concentration of oestrone sulfate during early gestation in the cow with a sensitive assay system. Five Holstein heifers were inseminated on oestrus and bled from the jugular vein at regular intervals until 100 days of gestation. Oestrone sulfate, oestrone and oestradiol-17beta in blood plasma were extracted with a reverse-phase cartridge and each measured by specific radioimmunoassay. Oestrone sulfate in bovine circulation started to increase around 50 days of gestation, whereas oestrone and oestradiol-17beta remained at basal concentrations for 80 days. The plasma concentration of oestrone sulfate increased gradually and linearly from 50 days of gestation, and was drastically elevated after 80 days of gestation before the increase in unconjugated oestrogens. The rate of increase from 80 to 100 days was the greatest for oestrone sulfate concentration among all oestrogens. These results suggest that plasma oestrone sulfate concentration could be a useful indicator of pregnancy within the first trimester of gestation in the cow.

Primary hypomagnesemia caused by isolated magnesium malabsorption: atypical case in adult.

Isolated magnesium malabsorption is a rare disorder, which bas been described in no more than 30 patients worldwide. Patients with this disorder typically present with convulsion and diarrhea in early infancy. Hypomagnesemia and hypocalcemia were found in a 35-year-old man with muscle cramps, who bad been diagnosed as primary hypoparathyroidism. Oral magnesium therapy corrected the low serum calcium, magnesium and parathyroid hormone levels. We report an atypical case of isolated magnesium malabsorption in an adult.

Plasma oestrone and oestradiol concentrations throughout gestation in cattle: relationship to stage of gestation and fetal number.

The objectives of the study were to characterise the peripheral plasma oestrone (E1) and oestradiol-17beta (E2) concentrations throughout gestation in the cow and to correlate this with the stage of gestation and fetal number. Cows (n = 10) were equally divided into two groups after non-surgical embryo transfer of in-vitro matured and in-vitro fertilised (IVM - IVF) embryos; Group 1 received a single embryo, Group 2 received twin embryos. Blood was collected about every third day from day 0 (day 0 was defined as first day of standing oestrus), then daily for the last 10 days of gestation and sampling was stopped one day post partum. Plasma E1 concentration exceeded that of E2 throughout gestation in both groups of cows. The time trend concentrations of plasma E1 were significantly affected by the stage of gestation (P < 0.01) and fetal number (P < 0.01) in the last two trimesters of gestation. The time trend concentrations of plasma E2 were significantly affected by the stage of gestation (P<0.01) but not foetal number (P = 0.09). In both groups there was marked preparturient increase in E1 and E2 concentrations. Plasma E2 profile between days 10 prepartum to parturition paralleled E1 in cows carrying a single foetus but was disparate during the same period in the twin-bearing cows. To conclude, our results indicate that although plasma E1 concentration was greater than E2 throughout gestation, both were related to the stage of gestation and that fetal number was correlated with circulating E1 levels in the last two trimesters of gestation.

Effects of trimebutine maleate on gastric motility in patients with gastric ulcer.

The effects of trimebutine maleate (TM), a prokinetic drug, on gastrointestinal motility in patients with gastric ulcer were investigated. Twenty patients with active gastric ulcers were allocated to two groups; 10 patients received a proton pump inhibitor alone (PPI group), given orally, and 10 patients received oral TM in combination with a PPI (PPI + TM group), each for a period of 8 weeks. Electrogastrography (EGG) and gastric emptying were measured before and after the treatment period. During the active ulcer stage, tachygastria (more than 0.06 Hz) or bradygastria (less than 0.04 Hz) in the EGG frequency were observed in 9 patients either before or after meals. During the healed ulcer stage, tachygastria or bradygastria was observed in 4 of 10 patients in the PPI group, while in the PPI + TM group, 1 patient had tachygastria and none had bradygastria. Postprandial dip (PD) was observed in 3 of the 20 patients during the active stage, while after treatment, PD was observed in 3 patients in the PPI group and in 6 patients in the PPI + TM group, respectively. Gastric emptying in the PPI group did not show any change between before and after treatment, while that in the PPI + TM group improved significantly after treatment. These results suggest that TM may have an ameliorative effect on abnormal gastric motility in patients with gastric ulcer.

Effect of stage of gestation and foetal number on plasma concentrations of a pregnancy serum protein (PSP-60) in cattle.

This study characterised the peripheral plasma concentration of PSP-60 throughout gestation, and examined the effect of stage of gestation and foetal number on this protein in Holstein cows after non-surgical embryo transfer. Cows (n=12) were divided into two groups; Group 1 contained single embryo recipient cows (n=5), Group 2 contained twin-embryo recipient cows (n=7). Blood was collected approximately every third day from day 0 (first day of standing oestrus), then daily for the last 10 days of gestation and until one day post-partum. Two of the twin-embryo recipient cows had abnormal pregnancies, consequently data from them was considered separately. In both groups PSP-60 increased progressively from about day 20 post-oestrus to 20 days pre-partum (from 0.9 +/- 0.2 to 49.7 +/- 8.7 ng ml(-1), and from 1.3 +/- 0.6 to 115 +/- 34.9 ng ml(-1) (mean +/- SEM), in singleton and twin-bearing groups, respectively). The mean concentrations between 20 and 10 days pre-partum increased dramatically by about six-fold (P<0.001) in singleton-bearing cows (from 49.7 +/- 8.7 ng ml(-1) to 283.8 +/- 73.7 ng ml(-1)) to over two-fold in twin-bearing cows (from 115 +/- 34.9 ng ml(-1) to 284 +/- 98.2 ng ml(-1)). The mean concentrations of the two groups were indistinguishable between 10 days pre-partum and parturition. Cows giving birth prematurely to stillborn calves or to a schistosomus reflexus calf exhibited abnormal PSP-60 profiles. Our findings indicate that peripheral plasma PSP-60 concentrations are correlated to the stage of gestation and foetal number, and assist in predicting foeto-placental viability.

Plasma bovine pregnancy-associated glycoprotein concentrations throughout gestation in relationship to fetal number in the cow.

This study characterized the peripheral plasma bovine pregnancy-associated glycoprotein (bPAG) profile throughout gestation and examined the effect of stage of gestation and fetal number on this profile in Holstein cows after non-surgical embryo transfer. Cows (n = 12) were divided into three groups: group 1 = normal singleton pregnancies (n = 5); group 2 = normal twin pregnancies (n = 5); group 3 = abnormal twin pregnancies (n = 2). Blood was collected about every third day from day 0 (defined as the first day of standing estrus), then daily for the last 10 days of gestation, and sampling was stopped one day postpartum. The time-related changes in plasma bPAG concentrations were significantly (P < 0.01) affected by the stage of gestation and fetal number (P < 0.01), except during the last 10 days of gestation. In both normal pregnancy groups, bPAG concentration increased rapidly during the first trimester (0.5 +/- 0.1 to 14.6 +/- 1.7 ng/ml and 1.0 +/- 0.6 to 21.8 +/- 4.8 ng/ml, in singleton and twin-bearing groups respectively), then progressively between days 160 and 20 prepartum (31.6 +/- 6.2 to 114.3 +/- 31.3 ng/ml and 41.6 +/- 7.4 to 155.8 +/- 36.6 ng/ml in singleton and twin-bearing cows respectively). The mean concentration between days 20 and 10 prepartum approximately tripled (P < 0.001) in both these groups of cows (114.3 +/- 31.1 to 493.0 +/- 75.3 ng/ml and 155.8 +/- 36.6 to 409.3 +/- 114.7 ng/ml in singleton and twin-bearing cows respectively), but between days 10 prepartum and parturition the values increased about threefold (P < 0.01) in the singleton group (493.0 +/- 75.3 to 1352.8 +/- 286.5 ng/ml) and fivefold (P < 0.001) in the twin-bearing group (409.3 +/- 114.7 to 2154.0 +/- 505.7 ng/ml). The two cows in group 3 that gave birth prematurely to a stillborn calf or to a schistosomus reflexus calf exhibited an aberrant bPAG profile. Our results indicate that peripheral bPAG concentrations are correlated to the stage of gestation and fetal number, and that the profile of the peripheral plasma concentrations provides a useful indication of the feto-placental status.

Rat hepatoma Reuber H-35 cells produce factors that promote the hatching of mouse embryos cultured in vitro.

The effects of coculture and conditioned medium of rat hepatoma Reuber H-35 cells on the subsequent in vitro development and hatching of mouse 2-cell embryos were examined. The hatching of embryos obtained from CD-1 mice was accelerated by coculture with Reuber H-35 cells in the presence of 3 mg/ml BSA. The promoting effect on complete hatching from zona pellucida was evident even in cell-conditioned medium containing 60 micrograms/ml BSA. In the presence of 60 micrograms/ml BSA, more than 20% of embryos completely hatched, whereas none hatched in the control culture. The promoting activity was also found in both the M(r) < 10,000 and the M(r) > 10,000 subfractions of the conditioned medium separated by ultrafiltration. The cell number per blastocyst was increased to 1.1- to 1.3 times the control by culturing embryos from the 2-cell stage with the conditioned medium or its subfractions. The effective target of promoting factors for complete hatching was after the morula stage, and blastocysts hatched completely even when incubated in conditioned medium for 6 h. Inhibitors of DNA polymerase alpha, protein synthesis, and protein kinase partially reduced (40-90% inhibition) the promoting effect of the conditioned medium. On the other hand, protease inhibitors showed no effect. In a caseinolytic assay, protease activity was undetectable in the conditioned medium. Incubating the 125I-labeled proteins derived from the M(r) > 10,000 fraction with blastocysts revealed that at least 9 proteins with apparent molecular masses of 76, 60, 49, 38, 34, 31, 24, 22, and 18 kDa specifically bound to or accumulated in the embryos. Moreover, reverse-transcriptase polymerase chain reaction showed that Reuber H-35 cells expressed mRNAs for epidermal growth factor, transforming growth factors alpha and beta 1, and stem cell factor. These results indicated that embryonic development and the process of zona hatching was accelerated by factors synthesized by Reuber H-35 cells. This and other studies demonstrated that Reuber H-35 cells exert positive (later than 2-cell stage) and negative (at 2-cell stage) effects upon the development of mouse embryos at different embryonic stages. These factors will serve as valuable tools to clarify the proliferating and differentiating mechanisms of the preimplantation embryo.

Maternal plasma estrone sulfate profile during pregnancy in the cow; comparison between singleton and twin pregnancies.

Peripheral plasma estrone sulfate (E(1)-S) concentrations were characterized throughout gestation in singleton and twin bearing cows by a direct radioimmunoassay method. Maternal plasma E1-S was detectable from around day 100 and after that its concentration increased progressively to term in both singleton (n = 5) and twin bearing (n = 5) cows. Twin bearing cows had a significantly higher E1-S concentrations in some time from mid-gestation to term when compared to singleton cows. E1-S concentration in the twin bearing cows increased rapidly in the third trimester and peaked (16.7 ng/ml) on the day of calving. In the singleton cows the concentration increased gradually and peaked (7.1 ng/ml) about 10 days before calving and then subsequently decreased. Our results indicate that singleton and twin bearing cows show a disparate E1-S profile from mid-gestation to term.

Peripheral cortisol levels throughout gestation in the cow: effect of stage of gestation and foetal number.

This study characterized the peripheral plasma cortisol profile throughout gestation and examined the effect of stage of gestation and foetal number in Holstein cows after non-surgical embryo transfer. Cows (n = 10) were divided into two groups: Group 1 = single embryo recipient cows (n = 5); and group 2 = twin-embryo recipient cows (n = 5). Mean plasma cortisol concentrations remained basal (2-4 ng ml-1) in both groups up to 2 days prepartum increased significantly (P < 0.05) to peak at parturition day, and then declined rapidly 1 day post-partum. Twin-bearing cows had significantly (P < 0.01) higher mean plasma cortisol concentration on the day of parturition than in the singleton cows. There was no effect of the stage of gestation on cortisol levels in either group (P > 0.1), except in the last 48 h prior to parturition. A single cow giving birth prematurely had 100% higher plasma cortisol levels on the day of parturition and 1 day post-partum than cows giving birth at term.

Plasma bovine placental lactogen concentration throughout pregnancy in the cow; relationship to stage of pregnancy, fetal mass, number and postpartum milk yield.

This study characterized the peripheral plasma placental lactogen (bPL) profile throughout gestation and examined the relationship between the stage of gestation, fetal mass, number, and postpartum lactation with circulating levels of bPL in Holstein cows after nonsurgical embryo transfer. Cows (n = 12) were divided into two groups: Group 1 = single embryo recipient cows (n = 5); Group 2 = twin embryo recipient cows (n = 7). Blood was collected about every third day from Day 0 (Day 0 was defined as the first day of standing estrus), then daily for the last 10 d of gestation, and sampling was stopped 1 d postpartum. The cows were milked twice daily at 0800 and 1800 hr. Two twin-embryo recipient cows had abnormal pregnancies; therefore, their data were excluded from that of the group and reported separately. The time trend concentrations of plasma bPL were significantly affected by the stage of gestation (P < 0.01) but not fetal number (P < 0.21). In both groups bPL levels remained low during the first two trimesters, then increased rapidly (P < 0.01) to peak concentrations between Days 200 and 220, and stabilized at this elevated level until parturition. Postpartum milk yields were indistinguishable between the singleton and twin bearing cows. Calf birth weight and postpartum lactation were both correlated (P < 0.01) to peripheral bPL concentration in singleton cows, however, this relationship decreased with a subsequent increase in fetal number. Cows giving birth prematurely to stillborn calves or to a schistosomus reflexus calf exhibited a deviating pBL profile. These results indicate that peripheral bPL levels are positively associated with the stage of gestation but not with fetal number. Otherwise, the peripheral pattern of bPL is a valuable index for predicting feto-placental viability.

Establishment of a reliable extraction method of estrone sulfate from bovine plasma for detection of the peripheral level during the regular estrous cycle by radioimmunoassay.

A simple extraction and assay technique of estrone sulfate in bovine blood was developed with the object of detecting the peripheral level of estrone sulfate in a normal estrous cycle or in early pregnancy. Estrone sulfate in bovine plasma was extracted with a small reversed phase cartridge. The steroid conjugate retained in the cartridge was eluted with 40% (v/v) methanol. Estrone sulfate was separately recovered from other steroids by the stepwise increase in methanol concentration in the elution solvent. The recoveries of estrone sulfate eluted with 40% methanol were more than 90%, irrespective of the applied plasma volume. The concentration measured by radioimmunoassay with the eluent of 40% methanol was consistent for plasma extraction volumes of 0.5-2.0 ml. The change of estrone sulfate in bovine peripheral plasma during the regular estrous cycle was determined with a small reversed phase cartridge for extraction and 40% methanol for elution. The change in estrone sulfate was found to be similar to the change of estrone and estradiol-17 beta. The concentration of estrone sulfate was not higher than that of both estrogens in cattle.

Rat hepatoma reuber H-35 cells produce a 2-cell stage-specific inhibitor of the cleavage of mouse embryos.

We purified an embryonic stage-specific inhibitor produced by rat hepatoma Reuber H-35 cells against cleaving mouse 2-cell embryos and defined its biological properties. Zygotes obtained from CD-1 mice (a strain that shows a 2-cell block in vitro) or C57BL/6 and B6C3F1 mice (strains that do not) were cultured in media with and without 50 microM EDTA, respectively. The development of the zygotes from all strains was arrested at the 2-cell stage when zygotes were cocultured with Reuber H-35 cells. However, the embryos from C57BL/6 and B6C3F1 were less sensitive than those from CD-1 against the inhibitory effects of development. This inhibitory effect was also evident in medium conditioned with the Reuber H-35 cells. The factor from the conditioned medium was separated into its < 10 000 M(r) fraction by ultrafiltration and was further purified in fraction B-25 as a single peak by reverse-phase column chromatography. An incubation as short as 3-h during the late 2-cell stage (G2 phase) with fraction B-25 suppressed cleavage in 61.5% of the CD-1 embryos (30.3% in control culture). Although the inhibitory effect was reversible, embryos that cleaved again either degenerated or were retarded at various stages in their subsequent development. Additionally, a long-term incubation of developing zygotes with the inhibitory factor caused a significant reduction in [3H]thymidine (TdR) incorporation into the DNA of CD-1 2-cell embryos as well as developmental arrest at the interphase of the 2-cell stage. These results indicated that this factor will serve as a valuable tool with which to clarify the proliferating mechanism of the preimplantation embryo.

Effect of fetal mass, number and stage of gestation on pregnancy-specific protein B concentrations in the bovine.

In this study we characterized the peripheral plasma pregnancy-specific protein-B (PSPB) profile throughout gestation and examined the effect of stage of gestation, fetal mass and number on this profile in Holstein cows after non surgical embryo transfer. Cows (n = 12) were divided into 2 groups: Group 1 = single embryo recipient cows (n = 5), Group 2 = twin-embryo recipient cows (n = 7). Blood was collected approximately every third day from Day 0 (Day 0 = first day of standing estrus), then daily for the last 10 d of gestation, and sampling was stopped 1 d post partum. Two twin-embryo recipient cows had abnormal pregnancies; therefore, their data were excluded from the group. The time trend concentrations of plasma PSPB were significantly affected by the stage of gestation (P < 0.001) and fetal number (P < 0.001). In both groups PSPB increased gradually, with the mean levels being significantly higher (P < 0.01) in the twin-bearing group from Day 50 onwards (0.7 +/- 0.2 vs 9.2 +/- 4.5 ng/ml, singleton and twin-bearing cows, respectively) except for Day 10 pre-partum. By mid-gestation (Day 140), mean PSPB levels increased in the singleton (P < 0.001) cows by thirty-fold (21.2 +/- 3.2 ng/ml) as opposed to a ten-fold (98.4 +/- 13.2 ng/ml) increase in the twin-bearing (P < 0.001) group. The mean PSPB concentrations between Days 30 to 20 prepartum dramatically increased by about 700 to 200% in singleton (128.8 +/- 46.3 to 745.6 +/- 66.7 ng/ml) and twin-bearing cows (375.6 +/- 130.4 to 861.5 +/- 127.9 ng/ml), respectively. The PSPB levels between Day 10 prepartum to parturition were significantly higher (P < 0.001) in the twin-bearing group than in the singleton group (745.6 +/- 66.7 to 1627.4 +/- 238.9 ng/ml vs 861.5 +/- 127.9 to 3103.0 +/- 643.0 ng/ml in singleton and twin-bearing groups, respectively). Calf birthweight was correlated (P < 0.01) to peripheral PSPB concentration in singleton cows; however, this relationship decreased with the subsequent increase in fetal number. Cows giving birth prematurely to stillborn calves or to a schistosomus reflexus calf exhibited abnormal PSPB profiles. These results indicate that peripheral PSPB levels are correlated to the stage of gestation and fetal number. In addition, the peripheral pattern of PSPB is a valuable guage for predicting fetoplacental viability.

Sex steroid levels throughout gestation in cows carrying normal and malformed fetuses.

Birth of malformed/stillborn calves is a liability to farmers and diagnosis of the condition early in gestation would be of immense economic benefit. We report on peripheral plasma progesterone (P4), estrone (E1) and estradiol (E2) concentrations quantified by radioimmunoassay throughout gestation in twin embryo recipient cows carrying normal (cow N), freemartin(cow F) and schistosomus reflexus fetuses (cow S). The undulating plasma P4 profiles were identical in all three cows throughout gestation apart from that the concentration in cow F dramatically declined on day 254 and it subsequently gave birth to stillborn calves. The plasma E1 concentration progressively increased in cow N to peak at parturition and then rapidly declined a day after parturition. E1 levels were lower in cow F than in cow N and exhibited a sudden increase in concentration at day 254 of gestation followed by a dramatic decline. Cow S had lower E1 levels throughout gestation than cow N and showed an undulating profile. The plasma E2 profile paralleled the plasma E1 profile in all the cows but the E2 concentration throughout gestation was lower than the E1 levels. Plasma E1 and E2 levels declined to < 20 pg/ml in cow N a day after parturition as opposed to > 150 pg/ml E1 and > 20 pg/ml E2 levels, respectively, in cows F and S. Our results indicate that E1 and E2 are better than P4 as prognostic indicators of fetal in-utero status as well as the number of fetuses a cow is gestating.

Luteotropic effect of pregnant mare serum gonadotropin in cattle.

Pregnant mare serum gonadotropin (PMSG) is a long acting luteotropin with follitropin activity. Cattle were used to examine whether a smaller dose of PMSG has substantial luteotropic effect without excessive follitropic effects. Eleven Japanese Black heifers were randomly assigned to two groups. Animals were administered 500 IU PMSG on the day of ovulation (Day 0) to promote the formation of corpus luteum (group A; N = 5) or Day 7 to stimulate the luteal function (group B; N = 6). Four of them were given injections of saline on Days 0 and 7 of the preceding estrous cycle for control. All animals were examined by palpation per rectum every other morning and bled every day for steroids analyses. The length of estrous cycle was shortened by the treatment in group A compared with the previous cycle, whereas it was extended in group B (P < 0.05). Progesterone secretion was not enhanced in group A, but it was significantly elevated and sustained on higher levels in group B (P < 0.05) as compared with the control. Although estradiol-17 beta concentrations were significantly increased in both PMSG-treated groups (P < 0.05), no excessive follicular development was observed. It is concluded that 500 IU PMSG administered on Day 7 enhances luteal function without excessive follicular development, whereas the administration on Day 0 has an adverse effect on luteal function.