Intralobular heterogeneity of perisinusoidal stellate cells in porcine liver

The aim of the present investigation was to elucidate the intralobular heterogeneity of the perisinusoidal stellate cells (fat-storing cells, lipocytes) in the porcine liver. Their three-dimensional structure, desmin immunoreactivity and vitamin-A storage were studied by use of the Golgi silver, immunocytochemical and gold chloride methods. In order to locate the stellate cells, the hepatic lobules were divided into 10 zones. The stellate cells were readily identified in Golgi preparations by their striking dendritic appearance with branching processes encompassing the sinusoids. The stellate cells in the centrolobular zones were conspicuously dendritic with longer processes in conspicuously dendritic with longer processes in comparison to those emitted by periportal elements. Such arborizations were studded with numerous thorn-like microprojections. Desmin immunoreaction in the periportal zones was stronger than that in the centrolobular zones. Vitamin-A storage in the stellate cells was well developed in zones 2–4, but reduced gradually toward the central region. The perisinusoidal etellate cells display marked heterogeneity in morphology and function based on their zonal location in the hepatic lobule.     

Managing the dominant follicle in lactating dairy cows

Reproductive efficiency is not optimal in high-producing dairy cows. Although many aspects of ovarian follicular growth in cows are similar to those observed in heifers, there are numerous specific differences in follicular development that may be linked with changes in reproductive physiology in high-producing lactating dairy cows. These include: 1) reduced circulating estradiol (E2) concentrations near estrus, 2) ovulation of follicles that are larger than the optimal size, 3) increased double ovulation and twinning, and 4) increased incidence of anovulation with a distinctive pattern of follicle growth in anovular dairy cows. The first three changes become more dramatic as milk production increases, although anovulation has not generally been associated with level of milk production. To overcome reproductive inefficiencies in dairy cows, reproductive management programs have been developed to synchronize ovulation and enable the use of timed AI in lactating dairy cows. Effective regulation of the CL, follicles, and hormonal environment during each part of the protocol is critical for optimizing these programs. This review discusses the distinct aspects of follicular development in lactating dairy cows and the methodologies that have been utilized in the past two decades in order to manage the dominant follicle during synchronization of ovulation and timed AI programs.     

Managing the dominant follicle in lactating dairy cows

Reproductive efficiency is not optimal in high-producing dairy cows. Although many aspects of ovarian follicular growth in cows are similar to those observed in heifers, there are numerous specific differences in follicular development that may be linked with changes in reproductive physiology in high-producing lactating dairy cows. These include: 1) reduced circulating estradiol (E2) concentrations near estrus, 2) ovulation of follicles that are larger than the optimal size, 3) increased double ovulation and twinning, and 4) increased incidence of anovulation with a distinctive pattern of follicle growth in anovular dairy cows. The first three changes become more dramatic as milk production increases, although anovulation has not generally been associated with level of milk production. To overcome reproductive inefficiencies in dairy cows, reproductive management programs have been developed to synchronize ovulation and enable the use of timed AI in lactating dairy cows. Effective regulation of the CL, follicles, and hormonal environment during each part of the protocol is critical for optimizing these programs. This review discusses the distinct aspects of follicular development in lactating dairy cows and the methodologies that have been utilized in the past two decades in order to manage the dominant follicle during synchronization of ovulation and timed AI programs.     

Expression of neural cell adhesion molecule (N-CAM) in perisinusoidal stellate cells of the human liver

Neural cell adhesion molecule (N-CAM) is distributed in most nerve cells and some non-neural tissues. The present immunohistochemical study has revealed, for the first time, the expression of N-CAM in perisinusoidal stellate cells of the human liver. Liver specimens were stained with monoclonal antibody against human Leu19 (N-CAM) by a streptoavidin-biotin-peroxidase-complex method. Light- and electron-microscopic analyses have shown that N-CAM-positive nerve fibers are distributed in the periportal and intermediate zones of the liver lobule. Perisinusoidal stellate cells in these zones are also positive for N-CAM. N-CAM is expressed on the surface of the cell, including cytoplasmic projections. Close contact of N-CAM-positive nerve endings with N-CAM-positive stellate cells has been observed. On the other hand, stellate cells in the centrilobular zone exhibit weak or no reaction for N-CAM. Perivascular smooth muscle cells and fibroblasts in the portal area and myofibroblasts around the central veins are negative for N-CAM. The present results indicate that the perisinusoidal stellate cells in the periportal and intermediate zones of the liver lobule characteristically express N-CAM, unlike other related mesenchymal cells, and suggest that the intralobular heterogeneity of N-CAM expression by stellate cells is related to the different maturational stages of these cells.     

Net nutrient absorption and liver metabolism in lactating dairy cows fed supplemental dietary biotin

The effect of feeding supplemental biotin on net absorption and metabolism of nutrients by the portal-drained viscera (PDV; the gut, pancreas, spleen and associated fat) and liver of lactating dairy cows was measured. Three cows in early to mid-lactation catheterised for measurements of net nutrient absorption and metabolism by the PDV and liver were fed a total-mixed ration with or without supplemental biotin at 20 mg/day using a switch-back design (ABA v. BAB) with three 2-week periods. There were no effects of feeding biotin on dry matter intake (22.2 kg/day), milk yield (29.5 kg/day) or milk composition. There was also no effect of feeding biotin on net release of glucose by the liver, net liver removal of glucose precursors (propionate, alanine, lactate) or net liver release of β-hydroxybutyrate. Feeding biotin increased net PDV release of ammonia. Reasons for the response are not certain, but a numerical increase in net PDV release of acetate suggests that rumen or hindgut fermentation was altered. Results of the present study do not support the hypothesis that supplemental biotin increases liver glucose production in lactating dairy cows.     

Embryonic mortality and the uterine environment in first-service lactating dairy cows

Embryos were collected non-surgically from the tip of one uterine horn of 23 lactating dairy cows on Day 7 of pregnancy. Embryos were classified on the basis of morphological criteria as normal (n = 12) or abnormal (n = 13). Abnormal embryos were further classified as cleavage stage (n = 9) or morula/blastocyst (n = 4). Cows producing an abnormal embryo did not differ in days post partum at oestrus, age or parity from cows producing a normal embryo. Cows with an abnormal morula/blastocyst also did not differ with respect to days post partum at oestrus from cows with abnormal cleavage-stage embryos but cows with an abnormal morula/blastocyst were significantly older and of greater parity than cows with an abnormal cleavage-stage embryo. Hepes-saline-PVP solution (30 ml) was initially infused into the uterine tip, mixed and then withdrawn with a syringe. Analysis of this fluid revealed that the concentrations of glucose, total protein, calcium, magnesium and potassium were significantly higher in the flushings from the uterus of cows with abnormal embryos than from cows with normal embryos and zinc and phosphorus tended to be higher in the uterine flushings of cows with abnormal embryos. Phosphorus, total protein, calcium and magnesium tended to be higher in the flushings from cows with abnormal morulae/blastocysts than from cows with abnormal cleavage-stage embryos. Plasma progesterone did not differ between cows with normal or abnormal embryos or in cows with abnormal morulae/blastocysts or abnormal cleavage-stage embryos. Most embryonic mortality therefore occurred before Day 5 (during cleavage) in these cows.(ABSTRACT TRUNCATED AT 250 WORDS)     

Proteomic analysis of physiological function response to hot summer in liver from lactating dairy cows

Lactation performance of dairy cattle is susceptible to heat stress. The liver is one of the most crucial organs affected by high temperature in dairy cows. However, the physiological adaption by the liver to hot summer conditions has not been well elucidated in lactating dairy cows. In the present study, proteomic analysis of the liver in dairy cows in spring and hot summer was performed using a label-free method. In total, 127 differentially expressed proteins were identified; most of the upregulated proteins were involved in protein metabolic processes and responses to stimuli, whereas most of the downregulated proteins were related to oxidation–reduction. Pathway analysis indicated that 3 upregulated heat stress proteins (HSP90α, HSP90β, and endoplasmin) were enriched in the NOD-like receptor signaling pathway, whereas several downregulated NADH dehydrogenase proteins were involved in the oxidative phosphorylation pathway. The protein–protein interaction network indicated that several upregulated HSPs (HSP90α, HSP90β, and GRP78) were involved in more interactions than other proteins and were thus considered as central hub nodes. Our findings provide novel insights into the physiological adaption of liver function in lactating dairy cows to natural high temperature.     

Immunolocalization of proliferating perisinusoidal cells in rat liver

Summary There is now substantial evidence that perisinusoidal (Ito or fat-storing) cells are the principal source of extracellular matrix proteins during hepatic fibrogenesis. In rat liver these cells express the intermediate fiament protein desmin; this is now widely used as an immunohistochemical marker for these cells. It has been shown that in experimental models of acute and chronic liver injury there is an increase in the number of desmin-positive perisinusoidal cells prior to the deposition of matrix proteins; however, these studies have failed to establish whether local proliferation is involved in this expansion of the desmin-positive perisinusoidal cell population.In order to investigate the kinetics of the perisinusoidal cell response, we have developed a novel double-labelling immunohistochemical technique for the simultaneous demonstration of desmin and incorporated bromodeoxyuridine in proliferating perisinusoidal cells in sections of fixed paraffiin-embedded rat liver. Application of this technique to a model of acute liver injury (single dose carbon tetrachloride by gavage) has shown that expansion of the perisinusoidal cell population is contributed to by local proliferation, with a labelling index of 18.7% 2 days following injury.     

Exogenous glucagon effects on health and reproductive performance of lactating dairy cows with mild fatty liver

Severe fatty liver, a metabolic disease of dairy cows in early lactation, results in decreased health and reproductive performance, but can be alleviated by treatment with i.v. injections of glucagon. Mild fatty liver in cows effects on health and reproductive performance were determined by treatment with 14-day s.c. injections of glucagon at 7.5 or 15 mg/day. Multiparous Holstein cows (n=32) were grouped into Normal and Susceptible based on liver triacylglycerol concentrations (>1% liver tissue biopsy wet weight) at day 8 postpartum (day 0=day of parturition). Susceptible cows (n=24) were assigned randomly to three groups and s.c. injected with 0mg glucagon [60 ml 0.15M NaCl] [n=8] (same for Normal cows), 2.5 mg glucagon, or 5 mg glucagon every 8 h for 14 days, beginning day 8 postpartum. Mild fatty liver resulted in an increased number of days with elevated body temperature during the injection period, an increased incidence of mastitis after glucagon treatment, increased days to first estrus and insemination, increased days before conception occurred, and decreased conception rate. In cows with mild fatty liver, glucagon (15 mg/day) decreased the number of days with elevated body temperature and the incidence of mastitis after hormone treatment. From these results, we suggest that mild fatty liver is detrimental to health and reproduction of dairy cows and, furthermore, that exogenous glucagon decreases some of these detrimental effects.     

Direct mobilization of retinol from hepatic perisinusoidal stellate cells to plasma.

We have studied the mechanism for mobilization of retinol from stellate cells. Our data show that perisinusoidal stellate cells isolated from liver contained retinol-binding protein (RBP) mRNA. By Western blot analysis we found that cultivated liver stellate cells secreted RBP into the medium. Cultivated stellate cells loaded in vitro with [3H]retinyl ester mobilized radioactive retinol as a complex with RBP. Furthermore, exogenous RBP added to the medium of cultured stellate cells increased the secretion of retinol to the medium. These data suggest that liver stellate cells in vivo mobilize retinol directly to the blood and that a transfer to parenchymal cells for secretion as holo-RBP is not required. The direct mobilization of retinol from liver stellate cells as retinol-RBP to blood is indirectly supported by the demonstration of RBP mRNA production and RBP secretion by lung stellate cells. The data suggest that the same mechanism for retinol mobilization may exist in hepatic and extrahepatic stellate cells. This is, vitamin A-storing stellate cells in liver, lungs, and probably also in other organs may synthesize their own RBP (or alternatively use exogenous RBP) and mobilize holo-RBP directly to the blood.     

The utilization of canola and its constituents by lactating dairy cows

Three feeding trials and one nylon bag trial were conducted to determine the effect of supplementing a barley-based control diet with 3.5% canola oil (CO), 22% presscake (CPC) or 9% whole seed (WCS) on feed intake, digestibility, milk yield and composition of lactating dairy cows. Ruminal utilization of canola meal (CM), CPC and WCS was also determined. Increasing the level of fat in the diet had no significant effect on intake of concentrate or digestible energy, or on total tract digestibility of dry matter (DM), crude protein (CP) and acid detergent fibre. Addition of canola in the form of CPC and WCS gave greater energy and ether extract digestibility than C and CO (P < 0.05). Diet had no significant effect on milk production, yield of milk CP, milk lactose + ash, gross energetic efficiency of milk production, milk urea or minerals. Milk fat and 4% fat corrected milk (FCM) yield were similar with the C and CPC diets, and with the CO and WCS diets. But the CO and WCS diets gave less milk fat and FCM than the C diet (P < 0.05). Milk crude protein was higher (P < 0.05) on the WCS diet than on the C, CO and CPC diets, which were similar. Diets WCS, C and CO promoted similar levels of blood urea (BU) but BU levels with CPC and CO were lower than with the C diet (P < 0.05). Ruminal DM and CP disappearance of CM was lower than for WCS and CPC at all incubation times (P < 0.05).     

Comparison of two estrus synchronization and resynchronization treatments in lactating dairy cows

Reproductive performance in cows following synchronization of estrus with intravaginal progesterone releasing devices (IVD) has varied with the length of treatment, cyclic status and prolonged return to estrus intervals in some cows following first AI. The objective of this study was to compare two methods of synchronizing and resynchronizing estrus on the reproductive performance of lactating dairy cows. Cows were treated with an IVD (Day 0) for 7 days (n = 350) or 8 days (n = 350), cloprostenol (0.5 mg i.m.) at the time of device removal and estradiol benzoate (EB) at the time of device insertion (1.5mg i.m.), and again 9 days later (1.0 mg i.m.). Cows were also resynchronized starting on Days 23 and 46 by reinsertion of IVDs for either 7 or 8 days and treatment with EB (1mg i.m.) at the time of device insertion and again 9 days later. Cows were inseminated on detection of estrus for 4 days after removal of devices at each of the synchronized estrous cycles. No significant differences in reproductive performance were detected between each treatment throughout the study period. Synchrony of estrus was more precise at the first and second estrus after treatment with an IVD for 8 days compared to 7 days. Cows classified as anestrous had lower reproductive performance than cows classified as cycling and had longer intervals to estrus at the second (P < 0.001) and third estrus (P < 0.06), but not at the first estrus (P = 0.09). Mean time to onset of estrus after IVD removal was less in cows treated with an IVD for 8 days compared to 7 days at each synchronized estrus (P < 0.01). More Holstein-Friesian cows were classified as non-pregnant and not detected in estrus than crossbreed cows (15.7%, 54/343 versus 9.0%, 24/266; [P < 0.05). The results of the study suggested that the main effects of the treatments that were used to synchronize and resynchronize estrus were to alter the timing and synchrony of estrus without affecting fertility.     

Effect of bovine somatotropin on reproductive function in lactating dairy cows

Several recent experiments have reported that chronic treatment with bovine somatotropin (bST) increased the number of days open without affecting the services per conception. The physiological basis for these effects was examined. Eleven lactating Holstein cows received daily injections of bST (40 mg) and 10 received daily injections of vehicle. Treatment was initiated between 32 and 85 d post partum and continued for up to 180 d. Eight of 11 bST-treated cows experienced at least one period of extended ovarian acyclicity during treatment. Only 3 of 10 control cows did so (P = 0.05). Concentrations of progesterone during luteal phases were lower in bST-treated cows than in controls (P = 0.06). Baseline concentrations of LH were suppressed in bST-treated cows compared with those of controls (P < 0.04). Neither the pulse frequency of LH nor the expression of estrous behavior was affected by bST (P > 0.30). These results indicate that chronic administration of a high dose of bST can reduce reproduction performance by promoting ovarian acyclicity.     

Comparison of artificial insemination versus embryo transfer in lactating dairy cows

Conception rates (CR) are low in dairy cows and previous research suggests that this could be due to impaired early embryonic development. Therefore, we hypothesized that CR could be improved by embryo transfer (ET) compared with AI. During 365 days, 550 potential breedings were used from 243 lactating Holstein cows (average milk production, 35 kg/day). Cows had their ovulation synchronized (GnRH-7d-PGF(2alpha)-3d-GnRH) and they were randomly assigned for AI immediately after the second GnRH injection (Day 0) or for transfer of one embryo 7 days later. Circulating progesterone concentrations and follicular and luteal size were determined on Days 0 and 7. Pregnancy diagnosis was performed on Days 25 or 32 and pregnant cows were reevaluated on Days 60-66. Single-ovulating cows with synchronized ovarian status had similar CR on Days 25-32 with ET (n = 176; 40.3%) and AI (n = 160; 35.6%). Pregnancy loss between Days 25-32 and 60-66 also did not differ (P = 0.38) between ET (26.2%) and AI (18.6%). When single (n = 334) and multiple (n = 57) ovulators were compared, independent of treatment, multiple ovulators had greater (P < 0.001) circulating progesterone concentrations on Day 7 (2.7 ng/ml versus 1.9 ng/ml) and there was a tendency (P = 0.10) for a greater CR in multiple ovulators (50.9% versus 38.1%). However, there was no difference in CR between AI and ET cows with multiple ovulations (50.0% versus 51.7%). In single-ovulating cows, CR tended to be lower for AI than ET in cows ovulating smaller follicles (diameter < or = 15 mm; 23.7% versus 42.3%; P = 0.06) but not average-diameter follicles (16-19 mm; 41.2% versus 37.3%; P = 0.81) or larger (> or =20 mm; 34.3 versus 51.0%; P = 0.36) follicles. Thus, although ET did not improve overall CR in lactating cows, follicle diameter and number of ovulating follicles may determine success with these procedures.     

Comparison of oestrous synchronization regimens for lactating dairy cows.

The objective of this experiment was to evaluate various programmes for synchronization of oestrus. The focus of the study was to evaluate rates of detection of oestrus, synchrony of oestrus, pregnancy rate, and effect of ovarian status at initiation of the programmes on rates of detection of oestrus and pregnancy rate. Spring-calving, lactating dairy cows (n = 2009) were allocated at random to one of six treatments: (1) A (n = 335), progestogen (controlled intravaginal drug release; CIDR) inserted per vaginum 10 d before breeding season for 8 d, 10 microg of buserelin at CIDR insertion, PGF2alpha treatment on the day prior to CIDR removal, and AI of cows detected in oestrus within 6 d after CIDR withdrawal; (2) B (n = 330), as in A, plus 1 mg of oestradiol benzoate i.m. 10 h post CIDR withdrawal; (3) C (n = 347), as in A, except buserelin was replaced by 10 mg of oestradiol benzoate; (4) D (n = 335), as in A, plus PGF2alpha and oestradiol benzoate at CIDR insertion; (5) E (n = 332), CIDR containing a 10 mg oestradiol benzoate capsule inserted per vaginum for 12 d; or (6) F (n = 330), as in E, plus PGF2alpha on the day prior to CIDR withdrawal. The oestrous detection rate (number of cows detected in oestrus within 6 days of CIDR withdrawal as a proportion of the number of cows submitted for synchronization of oestrus) and oestrous synchrony (oestrous detection rate within 2 d of CIDR withdrawal), respectively, were greater (P<0.05) following B (95.7% of 330, 98.7% of 316) compared with any of the other programmes for synchronization of oestrus (A: 87.5 of 335, 79.4% of 293; C: 86.7% of 347, 80.0% of 301; D: 90.1% of 335, 89.8% of 302; E: 74.4% of 332, 70.4% of 247; F: 76.4% of 330, 78.5% of 252). The oestrous detection rate was reduced (P<0.05) among cows in metoestrus administered E (64.0% of 50) relative to similar cows administered F (82.8% of 64). Pregnancy rate was greater (P<0.05) following B (57.9% of 330) than A (48.9% of 335, P = 0.06), C (43.2% of 347), E (40.0% of 332), and F (35.1% of 330) but not D (59.3% of 302), when based on those cows presented for oestrous synchronization programmes. In conclusion, 1 mg of oestradiol benzoate administered 10 h post CIDR withdrawal (B) resulted in the best overall oestrous detection, oestrous synchrony, and pregnancy rates, which would be beneficial to a fixed-time AI program.     

Soil intake of lactating dairy cows in intensive strip grazing systems

Involuntary soil intake by cows on pasture can be a potential route of entry for pollutants into the food chain. Therefore, it appears necessary to know and quantify factors affecting soil intake in order to ensure the food safety in outside rearing systems. Thus, soil intake was determined in two Latin square trials with 24 and 12 lactating dairy cows. In Trial 1, the effect of pasture allowance (20 v. 35 kg dry matter (DM) above ground level/cow daily) was studied for two sward types (pure perennial ryegrass v. mixed perennial ryegrass–white clover) in spring. In Trial 2, the effect of pasture allowance (40 v. 65 kg DM above ground level/cow daily) was studied at two supplementation levels (0 or 8 kg DM of a maize silage-based supplement) in autumn. Soil intake was determined by the method based on acid-insoluble ash used as an internal marker. The daily dry soil intake ranged, between treatments, from 0.17 to 0.83 kg per cow in Trial 1 and from 0.15 to 0.85 kg per cow in Trial 2, reaching up to 1.3 kg during some periods. In both trials, soil intake increased with decreasing pasture allowance, by 0.46 and 0.15 kg in Trials 1 and 2, respectively. In Trial 1, this pasture allowance effect was greater on mixed swards than on pure ryegrass swards (0.66 v. 0.26 kg reduction of daily soil intake between medium and low pasture allowance, respectively). In Trial 2, the pasture allowance effect was similar at both supplementation levels. In Trial 2, supplemented cows ate much less soil than unsupplemented cows (0.20 v. 0.75 kg/day, respectively). Differences in soil intake between trials and treatments can be related to grazing conditions, particularly pre-grazing and post-grazing sward height, determining at least in part the time spent grazing close to the ground. A post-grazing sward height lower than 50 mm can be considered as a critical threshold. Finally, a dietary supplement and a low grazing pressure, that is, high pasture allowance increasing post-grazing sward height, would efficiently limit the risk for high level of soil intake, especially when grazing conditions are difficult. Pre-grazing and post-grazing sward heights, as well as faecal crude ash concentration appear to be simple and practical tools for evaluating the risk for critical soil intake in grazing dairy cows.     

Early postabortion recovery of Neospora-infected lactating dairy cows

This study was designed to assess the reproductive performance of Neospora caninum–infected dairy cows (Bos taurus) after abortion. The population examined was composed of 92 aborting cows subsequently subjected to at least one artificial insemination (AI) attempt. Of these animals, 68 were N. caninum seropositive and 24 were seronegative. Only animals producing at least 25 kg milk at the estrus after abortion were inseminated. Normal uterine involution was verified at first AI after abortion in cows showing estrous signs within 30 d of abortion. Of the 92 aborting cows, 73 (79.3%) became pregnant after abortion: 57 (83.8%) and 16 (66.7%) of the N. caninum seropositive and seronegative cows, respectively. The factors days of gestation and lactation number at abortion failed to differ between the two Neospora serostatus groups, based on Student's t-test or Kruskal-Wallis tests, respectively. The interval from abortion to first AI, the number of AIs necessary for an animal to become pregnant, and the interval from abortion to next pregnancy in cows becoming pregnant were lower for the N. caninum–seropositive than for the N. caninum–seronegative cows (P < 0.0001). Logistic regression analysis indicated no significant effects of herd, lactation number, days of gestation at abortion, and the interval from abortion to AI on the likelihood of pregnancy at first AI after abortion. Based on the odds ratio, Neospora-seropositive cows were 6.22 times more likely to become pregnant at first AI after abortion compared with their seronegative partners. Fifty-three of the Neospora-seropositive cows (78%) were inseminated within 30 d after abortion and 26 (49%) became pregnant, whereas only one seronegative cow (4.2%) was ready for insemination within this 30-d period and remained open after first AI. These results indicate that Neospora-infected cows suffering abortion can be inseminated with a likelihood of high fertility within the first month of abortion. We recommend checking that uterine involution is normal early after abortion in N. caninum–seropositive cows.