Hepatocyte apoptosis in dairy cows with fatty infiltration of the liver

The objective of the present study was to analyse the apoptotic process of liver cells in dairy cows with fatty infiltration of the liver using indicators of DNA damage and immunohistochemistry. For this purpose, sixteen dairy cows with fatty liver were examined. On clinical examination, the physical condition of the animals was fair in nine and poor in seven cows. The most dominant clinical signs were reduced ruminal motility, inappetance and/or anorexia and recumbency. Postmortem examination, in seven cases, revealed enlarged liver (18-33kg), icteric carcasses and distended gallbladder. Laboratory results included neutrophilia, hypochloraemia, decreased concentrations of total bilirubin and increased concentrations of β-hydroxy butyric acid, non-esterified fatty acids and insulin. The activities of aspartate aminotransferase, γ-glutamyl transpeptidase, creatine kinase and lactate dehydrogenase were high. Histopathological examination of hepatic specimen showed lipid drops in cytosol with indistinct cellular membranes. In control hepatic cells, the DNA was tightly compressed and maintained the circular disposition of the normal nucleus. However, in the diseased cows, the damaged DNA migrated from the core toward the anode, forming a tail of a comet. Compared to controls, numerous ssDNA and caspase-3-positive cells were detected in the liver. To the authors' knowledge, this is the first study to document accelerated apoptosis of hepatocytes in dairy cows with fatty infiltration of the liver.     

Hepatocyte apoptosis in dairy cows with fatty infiltration of the liver

The objective of the present study was to analyse the apoptotic process of liver cells in dairy cows with fatty infiltration of the liver using indicators of DNA damage and immunohistochemistry. For this purpose, sixteen dairy cows with fatty liver were examined. On clinical examination, the physical condition of the animals was fair in nine and poor in seven cows. The most dominant clinical signs were reduced ruminal motility, inappetance and/or anorexia and recumbency. Postmortem examination, in seven cases, revealed enlarged liver (18–33 kg), icteric carcasses and distended gallbladder. Laboratory results included neutrophilia, hypochloraemia, decreased concentrations of total bilirubin and increased concentrations of β-hydroxy butyric acid, non-esterified fatty acids and insulin. The activities of aspartate aminotransferase, γ-glutamyl transpeptidase, creatine kinase and lactate dehydrogenase were high. Histopathological examination of hepatic specimen showed lipid drops in cytosol with indistinct cellular membranes. In control hepatic cells, the DNA was tightly compressed and maintained the circular disposition of the normal nucleus. However, in the diseased cows, the damaged DNA migrated from the core toward the anode, forming a tail of a comet. Compared to controls, numerous ssDNA and caspase-3-positive cells were detected in the liver. To the authors’ knowledge, this is the first study to document accelerated apoptosis of hepatocytes in dairy cows with fatty infiltration of the liver.     

Identification of plasma and hepatic parameters related to metabolic robustness in dairy cows

Blood plasma and hepatic parameters were identified that describe the differences between metabolically robust or vulnerable dairy cows grouped according to their past health status. Data from a field study on dairy cows were used from which metabolically challenged dairy cows were selected that had a milk fat percentage of >4.5 mg/g and a fat to protein ratio of >1.5 in their previous early lactation. The selected cows were either classified as metabolically robust or vulnerable based on the occurrence of various metabolic and (re)production disorders in their previous lactations. Blood and liver tissue samples were collected in week 3 ante partum (a.p.) (-3 wk), in week 4 (+4 wk) and in week 13 (+13 wk) post-partum (p.p.). Plasma concentrations of metabolites and hormones and mRNA expression of genes involved in metabolic pathways in the liver were used as variables for a two-group discriminant analysis (DA). Average discriminant scores (centroids) were different (p < 0.05) in -3 wk, +4 wk and in +13 wk. In -3 wk, significant variables that best explained the differences between metabolically robust and vulnerable cows were parity, plasma triglycerides, glucose and mRNA abundance of carnitine palmitoyltransferase 2 (CPT2). In addition, based on the classification matrix, 69% of the dairy cows were correctly classified. In +4 wk, identified significant parameters were parity, plasma glucose and urea, and 67% of the cows were correctly classified. In +13 wk, significant variables that explained the differences between the groups were parity, mRNA abundance of acyl-CoA synthetase long-chain 1 and CPT1, and 66% of the cows were correctly classified. In conclusion, the identified variables may distinguish from metabolically challenged cows, those cows that had a poorer health performance in their previous lactations.     

Effects of dihydroheptaprenol on the neutrophil function of postpartum dairy cows

Neutrophil responses were evaluated in dairy cows at parturition and on 1, 2, 3, 5 and 7 days after they were given dihydroheptaprenol (DHP) (0.5 mg/kg body weight) at parturition. The chemiluminescent index increased significantly (P less than 0.05) from parturition to 1 day after parturition in cows injected with DHP. No significant differences were observed in the number of neutrophils between untreated and DHP-treated groups of dairy cows during the immediate postparturient period.     

头胎与经产奶牛围产期生产性能和血清生化指标的比较

为比较头胎奶牛与经产奶牛在围产阶段生产性能和血清生化指标的差异,本研究选取临床健康的头胎和经产奶牛各10头,在同等条件下饲喂全混合日粮,于预产期前7、4、2、1 d及产后0、1、2、4、7、14 d晨喂前尾静脉采血并分离血清,测定血糖、甘油三酯和血钙浓度,产后11~40 d记录产奶量,于产后第30天采集乳样,测定乳成分和体细胞数。结果显示,头胎奶牛与经产奶牛血清甘油三酯、乳脂、乳蛋白、乳糖和乳干物质等指标均无显著差异（P>0.05）;经产牛产后日均产奶量和乳体细胞数均显著高于头胎牛（P<0.05）;在分娩前第4天和分娩后第2、4天时,头胎牛血糖浓度显著高于经产牛（P<0.05）,但在分娩时,头胎牛血糖浓度显著低于经产牛（P<0.05）;头胎牛在产前第1、2天、分娩当天及产后第2天的血钙水平显著高于经产牛（P<0.05）。     

Effects of diet energy density and milking frequency in early lactation on tumor necrosis factor-alpha responsiveness in dairy cows

A whole blood stimulation assay (WBA) with Escherichia coli lipopolysaccharide (LPS) and an enzyme-linked immunosorbent assay (ELISA) were established to measure the production of tumor necrosis factor-alpha (TNF-alpha) in bovine plasma. The assays were used to study the effect of time around parturition, and diet energy density, and milking frequency on TNF-alpha responsiveness of dairy cows in early lactation. Forty cows were included in a 2 x 2 factorial block design. One factor was high (H) versus low (L) diet energy density and the other factor was two versus three daily milkings. Blood samples were collected in weeks -3, -1, 2, 3, 5, 9, and 13 around parturition, and investigated for the TNF-alpha production ex vivo and CD14+ monocytes. The TNF-alpha response, CD14+ monocyte number, and CD14 expression level on monocytes were significantly increased in the weeks close to parturition. However, dips of varying sizes were observed for the measured parameters in week 3 after calving. Diet and milking frequency had no effect on the TNF-alpha response ex vivo or CD14 expression level on monocytes, but cows fed diet H had significantly higher numbers of CD14+ monocytes than cows fed diet L. The WBA with LPS was a fast reliable method for repeated measurements of TNF-alpha responsiveness in cattle. Previous findings of increased TNF-alpha responses in periparturient cows were confirmed, whereas diet energy concentration and milking frequency had no effect on the TNF-alpha responsiveness in early lactation.     

Comparison of Productive Performance and Serum Biochemical Indices of Primiparous and Multiparous Dairy Cows During the Perinatal Period

In order to study the differences of productive performances and serum biochemical indices between primiparous and mulitiparous dairy cows during the perinatal period, ten primiparous and ten multiparous healthy dairy cows were selected and were fed the same total mixed ration.Blood samples were collected from the caudal vein before the morning feeding on -7, -4, -2, -1, 0, 1, 2, 4, 7 and 14 d after parturition, and serum glucose, triglyceride and calcium concentrations were determined. Milk yields were recorded every day from 11 to 40 days after parturition. Milk samples were collected from each experimental cow on 30th day after parturition and milk compositions and somatic cell counts were determined. The results indicated that no significant difference was observed in serum triglyceride, milk fat, milk protein, milk lactose and milk dry matter between primiparous and multiparous dairy cows (P>0.05). Mutiparous cows had greater daily milk yield and higher milk somatic cell count than primiparous cows (P<0.05). Primiparous cows had significant higher serum glucose level on -4, 2 and 4 d after parturition (P<0.05), but significant lower serum glucose level on calving date (P<0.05) when compared with multiparous cows. Higher serum calcium concentrations were found on -2, -1 and 2 d after parturition in primiparous cows than multiparous cows (P<0.05).     

Peripartum changes in antibody producing capability of lymphocytes from dairy cows.

B-lymphocyte populations and the in vitro antibody producing activity of lymphocytes were measured to evaluate the lymphocyte function of periparturient dairy cows. Changes in B-lymphocyte populations were not significantly different in cows during this period. Significantly decreased numbers of plaque-forming cells were observed in lymphocytes from cows at parturition and 3 days after parturition compared to those in lymphocytes obtained 14 days after parturition.     

Effect of suckling on the reproductive performance and metabolic status of obese Japanese black cattle during the early postpartum period

The aim of our study was to investigate the effects of suckling on reproductive performance and metabolic status of obese (mean body condition score of more than 4.0 on a scale of 1-5) maternal Japanese Black cows during early postpartum period. We used 7 postpartum Japanese Black cattle. Four cows were suckled ad libitum (suckled) until completion of their first artificial insemination (AI), while 3 cows were not suckled at all because they were separated from their calves immediately after parturition (non-suckled). Body weight and plasma concentrations of metabolites and hormones were measured from wk 1 to 9 postpartum. Ovarian activity was detected using plasma progesterone concentration, and all cows received their first AI after application of the Ovsynch protocol at approximately 4 months postpartum. Although body weights of non-suckled cows increased during experimental period (P<0.05), those of suckled cows remained unchanged. Plasma concentrations of glucose of non-suckled cows were higher at wk 2 postpartum (P<0.05) and their levels of non-esterified fatty acid tended to be lower at wk 1 and 2 postpartum compared with suckled cows (P<0.1); however, these differences between groups were not observed with progression of postpartum period. In addition, plasma insulin concentrations of non-suckled cows were higher than those of suckled cows during experimental period (P<0.05). During sampling period (wk 0 to 9 postpartum), onset of normal ovarian cycle was observed in all non-suckled and 2 of 4 suckled cows, and it was delayed in other 2 suckled cows compared with non-suckled cows; however, 3 suckled cows conceived at the first AI after application of the Ovsynch protocol; none of non-suckled cows conceived at this time. Overall, we suggest that suckling seems to reduce increase of body weight after parturition, although it does not improve obesity, and influences conception despite delay in resumption of normal ovarian cyclicity in obese Japanese Black cows.     

Effect of hoof trimming before the dry period on productive performance in perinatal dairy cows

To investigate the effect of hoof trimming before the dry period, the hooves of 10 cows (trimmed group) were trimmed at 79.6 +/- 8.6 days before parturition and the hooves of 52 cows were left untrimmed (control group). Blood biochemistry and hormone concentrations were investigated for 6 cows from each group. The daily milk yield after parturition in the trimmed group tended to be higher than that of the control group. Between 0 and 30 days after parturition, the levels of non-esterified fatty acids were significantly lower stet, and the plasma glucose and glucose disposal rates were significantly higher in the trimmed group. The plasma cortisol concentration was stable before and after parturition in the trimmed group. Hoof trimming before the dry period appears to reduce stress and maintain the nutritional conditions of perinatal dairy cows.     

过瘤胃烟酸和胆碱对围产期奶牛泌乳性能和肝脂质代谢的影响

旨在研究日粮中添加过瘤胃保护烟酸(rumen-protected niacin,RPN)和过瘤胃保护胆碱(rumen-protected choline,RPC)对围产期奶牛泌乳性能和肝脂质代谢的影响,为缓解奶牛脂肪肝提供理论依据.本试验选用24头健康、胎次相近的中国荷斯坦围产期奶牛,根据2×2试验设计分为4组,每组6...     

不同能量摄入对围产期奶牛脂肪组织胰岛素受体mRNA丰度的影响

将年龄、胎次相同,年产奶量大于5000kg的围产期健康奶牛30头随机分为3组,每组10头。按照中国奶牛饲养标准(2000),低能组饲喂减少20%日粮(能量摄入80%组),对照组饲喂标准日粮(能量摄入100%组),高能组饲喂增加20%日粮(能量摄入120%组),试验从产前14天开始至产后28天结束,产后各组奶牛均饲喂标准日粮。采用半定量RTPCR方法检测了不同能量摄入对围产期奶牛脂肪组织胰岛素受体(InsR)mRNA丰度的影响。结果表明:各组脂肪InsRmRNA相对表达量从产前14天至产后28天均呈现先升高后降低的趋势,产后14天达到最大值,且产后高于产前;低能组产前InsRmRNA相对表达量低于高能组和正常组,产后1天至产后28天高于高能组和正常组,产后14天显著高于高能组(P<0.01)。可见,干乳期低能饲喂奶牛,产后脂肪InsRmRNA表达增加,提示产前适当降低能量摄入水平,有利于胰岛素抑制泌乳初期脂肪动员作用的发挥。     

Fecal shedding of coliform bacteria during the periparturient period in dairy cows

OBJECTIVE: To determine whether numbers of coliform bacteria in feces of dairy cattle changed during the periparturient period and whether fluctuations were associated with changes in dry-matter intake. ANIMALS: 12 healthy Holstein cows. PROCEDURE: Fecal samples were collected on a semi-regular basis (i.e., 3 to 7 times/wk) beginning 4 to 6 weeks before the anticipated parturition date and continuing through the third day (5 cows) or second week (7 cows) after parturition, and total numbers of fecal coliform bacteria were determined. Daily feed intake of 7 cows was monitored. RESULTS: For 11 cows, fecal coliform bacterial counts between 34 and 25 days prior to parturition were low and relatively constant (< 102 change in number of bacteria). Coliform bacteria were not detected in 4 to 8% of fecal samples from 10 cows. All cows had a 10(4) to 10(7) increase in number of colony forming units/g of feces near the time of parturition. Number of fecal coliform bacteria peaked within 7 days of parturition in 9 cows and within 12 days of parturition in 3. Number of fecal coliform bacteria was not correlated with feed intake. CONCLUSIONS AND CLINICAL RELEVANCE: Cows may have large increases in fecal coliform bacteria count during the periparturient period; however, periparturient cows do not continually shed high numbers of coliform bacteria, and coliform bacteria may not always be detectable by conventional culture methods. Changes in fecal coliform bacteria count did not correlate with changes in dry-matter intake.     

Prolactin, estradiol, and progesterone changes in paretic and nonparetic cows during the periparturient period

We studied the relationship of serum prolactin, estradiol-17 beta, and progesterone concentrations to plasma calcium, phosphorus, and free hydroxyproline concentrations, as well as to dry matter intake, in 14 aged dairy cows (mean of 4.5 parities), 7 of which became paretic, from 28 days before to 4 days after calving. Plasma calcium and phosphorus concentrations and dry matter intake decreased more at parturition in paretic cows than in nonparetic cows. Prolactin concentrations were not different between paretic and nonparetic cows, but were variable. Concentrations of estradiol were higher in paretic cows from 15 to 5 days before parturition, whereas hydroxyproline concentration was lower in paretic cows on days 10 through 3 before parturition. Progesterone concentration was lower in paretic cows and decreased earlier at parturition, compared with that in nonparetic cows. The findings suggested that high estradiol concentrations in late pregnancy inhibit bone resorption and predispose aged cows to parturient paresis. The earlier decrease in progesterone concentration at parturition and lower concentrations throughout late pregnancy might have contributed to the greater inappetence in paretic cows at parturition. The importance of prolactin in the pathogenesis of parturient paresis is not clear.     

Influence of Body Condition on Serum Metabolic Indicators of Lipid Mobilization and Oxidative Stress in Dairy Cows During the Transition Period

The objectives of this study were to examine the influence of body condition of cows on metabolic and antioxidative status, as well as to investigate the relationship between metabolic indicators of lipid mobilization and oxidative stress during transition period. The study was conducted on 24 Holstein‐Friesian dairy cows divided into 2 groups according to their body condition score (BCS) as optimal (n = 12; BCS from 3.25 to 3.75) or adipose (n = 12; BCS ≥4). Metabolic status (glucose, triglycerides, total cholesterol, HDL cholesterol, NEFA and BHB), paraoxonase‐1 (PON1) and apolipoprotein A‐I (ApoA‐I) were analysed in sera taken on days ?30, ?10, ?2, 0, 5, 12, 19, 26 and 60 relative to parturition. Adipose cows had significantly higher glucose concentration at parturition being significantly decreased after parturition on days 12 and 19. Total cholesterol and HDL‐C concentrations were the lowest at parturition and significantly higher on days 26 and 60 after parturition in both groups of cows. Both investigated groups had significantly higher NEFA concentration from parturition until day 19 after parturition, indicating energy deficit and an increased lipid mobilization after calving. There were no significant differences in BHB concentration during transition period in both groups. No significant differences were found in PON1 activity and ApoA‐I concentration during transition period in both groups of cows. However, in adipose cows, although not significantly different, PON1 was decreased from calving until day 19 after parturition indicating a disturbance in antioxidative status in adipose cows. PON1 significantly positively correlated with total cholesterol and HDL‐C concentrations and negatively with NEFA indicating a strong relationship of PON1 with lipid metabolism. Significant positive correlation between NEFA and BHB in both groups of cows points out on energy deficit during transition period that cows tend to overcome by lipid mobilization providing alternative source of energy needed for parturition and lactation.     

干乳期能量摄入水平对围产期奶牛肝载脂蛋白B100mRNA丰度的影响

将年龄、胎次相同，年产奶量大于5000kg的围产期健康奶牛30头随机分为3组，每组10头。Ⅰ组饲喂中国奶牛饲养标准（2000）减少20％日粮（能量摄入80％组）。Ⅱ组饲喂中国奶牛饲养标准（2000）日粮（能量摄入100％组），Ⅲ组饲喂中国奶牛饲养标准（2000）增加20％日粮（能量摄入120％组）。试验从产前28d开始至产后56d结束，产后各组奶牛均饲喂标准泌乳日粮。采用内对照RT—PCR方法，检测了干乳期摄入不同能量的围产期奶牛肝脏活体组织载脂蛋白B100（apoB100）mRNA丰度。结果表明。apoB100mRNA相对表达量，产前至产后Ⅰ、Ⅲ组均呈先升高后降低的趋势，Ⅲ组产后apoB100mRNA丰度降低了61．45％，Ⅰ组呈逐渐降低的趋势，各组均在产后28d达最低值（Ⅰ组，P〈0．05；Ⅱ、Ⅲ组，P〈0．01）。产前14d至产后1d，apoB100mRNA相对表达量Ⅲ组高于Ⅰ组，Ⅰ组高于Ⅱ组，组间差异极显著（P〈0．01）；产后28d。Ⅰ组高于Ⅱ、Ⅲ组（P〈0．05）。由此可见，围产期奶牛能量摄入水平对肝脏载脂蛋白B100mRNA丰度有显著影响。     

Effects of feeding intensity during the dry period on leukocyte and lymphocyte sub-populations, neutrophil function and health in periparturient dairy cows

The objective of this investigation was to study (1) the numbers of leukocytes, (2) the proportions of lymphocytes expressing CD4, CD8, WC1, B or IL2R and (3) neutrophil phagocytosis and oxidative burst activity in blood around parturition in three groups of dairy cows fed different levels of a total mixed ration during the last eight weeks before calving. All cows were fed ad libitum during the first eight weeks of lactation. Serum concentration of the acute phase protein serum amyloid A (SAA), the milk somatic cell count (SCC) and disease incidence were also recorded. Special emphasis was given to the weeks just before and just after calving as dairy cows are known to be immune suppressed during this period. Dry period diet had only minor effects on leukocyte numbers, and did not influence neutrophil phagocytosis and oxidative burst. In addition, no effect was observed on disease incidence or SAA concentrations. However, an increase in the proportion of B-lymphocytes and a decrease in the proportion of WC1+ T lymphocytes were observed after calving in cows fed high or low energy rations during the dry period, but not in cows fed a medium energy ration. The weeks just before and after parturition were characterised by neutrophilia, eosinopenia, lymphopenia and monocytosis, but time had no effect on neutrophil phagocytosis and oxidative burst. The proportions of CD4+, CD8+, B+ and IL-2R+ lymphocytes increased in early lactation relative to the mid dry period. In addition, the concentration of SAA increased dramatically at calving. The results emphasise the need for more studies to clarify the complex interactions between nutrition and immunity during the peripartum period in dairy cattle.     

The impact of the chromium supplementation on insulin signalling pathway in different tissues and milk yield in dairy cows

Thirty days before expected time of parturition, 20 Holstein cows were divided into ?Cr and +Cr groups. From day 25 before parturition (BP ) up to day 30 after parturition (AP ), +Cr cows received 10 mg of Cr (chromium‐enriched yeast) daily. Muscle and adipose tissue samples were taken at days ?30, ?10, +7 and +10 related to parturition, when body condition score (BCS ) was also determined. Hepatic tissue samples were taken at days ?10 and +7. Tissue samples were used for determination of the insulin signalling pathway protein expressions. Intravenous glucose tolerance test (IVGTT ) was performed at days ?28, ?7, +10 and +30. Milk yield was recorded during first 14 weeks AP . Milk composition was obtained at days 7 and 28 AP . At day 10 BP , protein content of β ‐subunit of insulin receptor (IR β ) was significantly higher (p ? 0.05) in muscle, and phosphorylation of insulin receptor substrate 1 at serine 307 (pIRS ‐1 Ser³⁰⁷) was significantly lower (p ? 0.05) in hepatic tissue of +Cr group. After parturition, pIRS ‐1 Ser³⁰⁷ was significantly lower in muscle tissue at days 7 and 28 (p ? 0.05 and p ? 0.001, respectively), while phosphorylation of Akt at serine 473 (pA kt Ser⁴⁷³) was significantly higher (p ? 0.01) in hepatic tissue at day 7 AP in +Cr group. Chromium had opposite effect on insulin kinetics during IVGTT s obtained BP and AP . Insulin secretion was significantly reduced at day 7 BP and significantly enhanced at day 10 AP , when NEFA concentration was also significantly increased. Milk yield and ECM value were depressed in +Cr group. DMI and BCS were significantly enhanced in +Cr group at day 7 BP . In conclusion, chromium modulates insulin signalling pathway in dairy cows, but targeted signalling molecules are different in antepartal then post‐partal period, probably due to duration of exposure to chromium and different energy status between those periods.     

Effects of active egg white product/ Clostridium butyricum Miyairi 588 additive on peripheral leukocyte populations in periparturient dairy cows

The leukocyte populations of periparturient dairy cows were analyzed after administration of active egg white/Clostridium butyricum Miyairi additive. Sixty-eight Holstein milking cows were divided into 3 groups. Group A was administered active egg white product (AEWP)/Clostridium butyricum Miyairi 588 (Miyairi 588) additive (n=23). Group B was administered Miyairi 588 only (n=23), and Group C was the control group (n=22). The challenged groups were administered 100 g of AEWP + Miyairi 588, or Miyairi 588 alone, daily for 60 days from 1 month before until 1 month after paturition. Blood samples were collected from all groups three times (1 month before, 1 week after and 1 month after parturition) for analysis of the peripheral leukocyte population. The results showed significantly higher numbers of CD4+ cells in Group A compared with Group C 1 week after paturition. AEWP/Miyairi 588 additive may enhance the number of CD4+ T cells in periparturient dairy cows.