复方氨基酸螯合钙对产后奶牛PMN功能和乳腺炎发病率的影响

为明确复方氨基酸螯合钙口服液对产后奶牛外周血嗜中性粒细胞(PMN)功能和乳腺炎发病率的影响,课题组成员采用趋化试验和吞噬功能试验研究复方氨基酸螯合钙口服液对产后奶牛外周血嗜中性粒细胞功能和乳腺炎发病率进行测定。结果表明:产后每天灌服复方氨基酸螯合钙口服液1周后奶牛嗜中性粒细胞趋化功能和吞噬指数显著提高(P0.05),乳中体细胞数和隐性乳房炎发病率显著降低(P0.05)。说明复方氨基酸螯合钙口服液适合作为产后奶牛专用保健品在临床上应用。     

奶牛乳腺炎对奶牛血液指标的影响

为探讨乳腺炎奶牛血液成分的变化情况与奶牛乳腺炎发生发展的相关性,进一步研究奶牛乳腺炎的发病机理,为奶牛乳腺炎的防治提供血液学依据,采用动物血液细胞分析仪和全自动生化分析仪对临床型乳腺炎奶牛、隐性乳腺炎奶牛和健康奶牛的血液常规指标、多项血清生化指标进行检测分析和比较研究。结果表明:临床型乳腺炎奶牛、隐性乳腺炎奶牛与健康奶牛相比,相关血液常规指标的检测结果和血清生化指标的检测结果有变化。说明奶牛乳腺炎是一个全身性的病理过程,不仅乳腺组织遭到破坏,血液成分也发生了变化。     

奶牛乳腺炎的预防和治疗

<正>几年来,在国家政策强力扶持下,奶牛规模养殖场获得长足发展,但因部分奶牛场管理水平相对滞后,奶牛乳腺炎一直如影随形,很大程度上影响着牛奶的产量和品质,造成了较大经济损失,也对广大消费者的身体健康构成了威胁,控制奶牛乳腺炎意义重大。奶牛乳腺炎是乳房实质和间质发生炎症。奶牛乳腺炎的病因多由机械性刺激、感染病原微生物及化学物理性损伤所致。奶牛乳腺炎分为浆液性乳腺炎、纤维素性卡他乳腺炎、化脓性乳腺     

双城市奶牛乳腺炎的药物治疗效果分析

奶牛乳腺炎是一种较为复杂的疾病,它的危害不容小视,它不仅影响奶牛的产奶量和牛奶的品质,更危及人类的健康.本试验重点调研了双城地区奶牛乳腺炎,并对双城地区奶牛临床型乳腺炎和隐性乳腺炎进行了治疗,旨在为今后奶牛场的临床防治奶牛临床型乳腺炎和隐性乳腺炎提供一定的药物应用指导.     

奶牛乳腺炎及其防治研究进展

主要分析了诱发奶牛乳腺炎的物理性、生物性和营养性因素及其发病规律,介绍了奶牛乳腺炎的预防措施及治疗方法,以期为临床有效防治奶牛乳腺炎提供参考。     

奶牛多形核白细胞凋亡的分子调控与乳腺免疫

多形核白细胞(polymorphonuclear neutrophil leukocytes,PMN)在奶牛乳腺防御病原菌感染中充当首要防线,而PMN的凋亡在解除奶牛乳腺的炎性反应中发挥关键作用。作者综述了PMN凋亡的分子调控及其在奶牛乳腺免疫中的重要作用和意义。     

不同程度奶牛乳腺炎血清中微量元素含量变化的试验

为探讨奶牛乳腺炎血清中相关微量元素的变化是否与发病程度之间存在相关性,本试验采用原子吸收分光光度法对健康奶牛、轻度隐性乳腺炎奶牛、重度隐性乳腺炎奶牛和临床型乳腺炎奶牛血清中Zn、Cu、Fe等微量元素的含量进行测定,结果患有乳腺炎奶牛血清中Zn含量低于健康奶牛,并随炎症程度的加剧呈负相关性;Cu和Fe含量高于健康奶牛,含量随炎症程度的加剧呈正相关性.提示奶牛血清中微量元素含量的变化与乳腺炎的发病程度存在着一定的相关性.     

奶牛体细胞数与乳腺炎遗传相关性的研究进展

牛奶中的体细胞数和乳腺炎关系密切,当奶牛患乳腺炎时,牛奶中的体细胞数会显著增加,体细胞数成为奶牛乳腺炎的指示物。为了选育出抗乳腺炎的奶牛品种,首先应该研究体细胞数和乳腺炎之间的遗传力和遗传相关度。本文对国外奶牛体细胞数与遗传相关性的研究进行了综述。     

奶牛乳腺炎的诊断与防治

奶牛乳腺炎是奶牛常见的疾病之一,也是花费最大的奶牛疾病之一,分为隐性乳腺炎和临床乳腺炎。乳腺炎能够导致奶牛的产奶量大大下降。由于隐性乳腺炎不表现临床症状,难以及时发现和治疗,所以往往给奶牛生产带来很大的经济损失。在国内,许多奶牛场的奶牛乳腺炎患病率高达50％以上。     

奶牛乳腺炎主要病原菌致病性研究

奶牛乳腺炎是奶牛的常见病和多发病,是造成奶牛业经济损失的主要疾病之一。对奶牛乳腺炎主要病原菌致病性的研究,将为我国成功控制奶牛乳腺炎的流行奠定基础。应用已分离鉴定的奶牛乳腺炎主要病原菌,给小白鼠腹腔注射,通过临床症状观察和病理学检查研究其致病性。结果表明,奶牛乳腺炎主要病原菌对实验小白鼠均可产生致病性,各组织器官出现了不同程度的病理变化。     

Identification of subpopulations of bovine mammary-gland phagocytes and evaluation of sensitivity and specificity of morphologic and functional indicators of bovine mastitis

The number and function of bovine mammary-gland phagocytes were assessed in 8 lactating cows, each tested at least twice within an 8-mo period (total number of observations, 20). Macrophages and polymorphonuclear (PMN) cells were evaluated by conventional cytology, flow cytometry, fluorescent microscopy, and somatic-cell count (SCC). Phagocytosis was evaluated from the uptake of fluorescent beads and expressed as median fluorescence intensity (MFI). Two major subpopulations of phagocytes, of low or high MFI (LFI or HFI), were observed, and there were up to 4 sub-subpopulations within the HFI subpopulation of both macrophages and PMN cells. Fluorescent microscopy identified phagocytes containing up to 4 beads per cell. Cows showing ≤ 72.3% phagocytes by cytology were regarded as non-mastitic (11 observations), and those showing ≥ 80.7% phagocytes were considered to be mastitic (8 observations). Phagocyte MFI was negatively associated with mastitis; that is, the higher the MFI, the lower the SCC. The percentage of HFI PMN cells was the only indicator of mastitis with 100% sensitivity and specificity. Thus, bovine mammary-gland phagocytes consist of several subpopulations of different phagocytic ability, whose assessment more adequately predicts bovine mastitis than do morphologic indicators.     

Apoptosis of polymorphonuclear leukocytes of the juvenile bovine mammary gland during induced influx

The dynamics of apoptosis of polymorphonuclear leukocytes (PMN) during induced influx of PMN into the cavity system of the juvenile bovine mammary gland in order to investigate the role of apoptosis of PMN in the resolution of mastitis was studied. The instillation of a synthetic analogue of muramyl dipeptide into teat sinus of the sixteen mammary glands was followed by a massive influx of PMN culminating after 24 h and resolving after 96 h. Every 24 h following the influx, apoptotic PMN were microscopically detected, based on morphological characteristics. Twenty four hours after the stimulation, apoptotic PMN were already observed, and peak counts of apoptotic PMN were reached 48 h after the stimulation. The lowest differential count of apoptotic PMN, corresponding to the pre-stimulation value, was found 96 h after the stimulation. The presence of macrophages (MAC) containing phagocytized apoptotic PMN was observed by histochemical staining for myeloperoxidase (MPO) and electron microscopy. The percentage of MPO-positive macrophages increased during the resolution phase to reach peak values 48 h after the stimulation. Apoptosis of PMN and phagocytosis by macrophages may represent a removal mechanism that is important in the resolution of the induced influx of PMN in the cavity system of juvenile bovine mammary gland.     

Inflammatory cell infiltration as an indicator of Staphylococcus aureus infection and therapeutic efficacy in experimental mouse mastitis

Staphylococcus aureus intramammary colonization of the mouse mammary gland induces migration of polymorphonuclear neutrophils (PMNs) similar to that observed during bovine mastitis. In the present study, a method combining acridine orange staining, fluorescence microscopy and computer-assisted image analysis has been developed to quantitate PMN infiltration in a mouse model of mastitis. This was carried out using paraffin embedded sections, and using this method, we showed that the presence of PMNs increased with the number of bacteria present in tissues. Nearly 400 and 1100 times more PMNs were counted in the mammary gland tissue after 12 and 24 h of infection, respectively, compared to mice infected for 6 h. Treatment with the antibiotic cephapirin at 10 or 25 mg/kg reduced PMN infiltration by 71 and 85%, respectively. In conclusion, this method can be used to quantitate PMN infiltration as a marker of inflammation and bacterial burden in infected tissue sections.     

Effects of bovine mammary secretion during the early nonlactating period and antibiotics on polymorphonuclear neutrophil function and morphology.

The effect of bovine mammary secretion during the early nonlactating period and of antibiotic preparations on bovine polymorphonuclear neutrophil (PMN) phagocytic function and morphology were evaluated in a series of in vitro multifactorial experiments. Benzathine cloxacillin (CL), benzathine cephapirin (CE), sodium novobiocin (NO), and a combination of dihydrostreptomycin with procaine penicillin G (DP) were prepared in the presence and absence of a peanut oil aluminum monostearate vehicle. The PMN were isolated from bovine blood, and the effect of each antibiotic preparation on PMN function and morphology was evaluated in a buffer, fat, skin, and a combination of fat with skim from bovine mammary secretion during the nonlactating period. The fat and skim were diluted with buffer to approximate their concentration in mammary secretion. Phagocytic functions of PMN were monitored by fluorescent microscopy, which made it possible to estimate both ingestion and intracellular killing of bacteria by PMN. Changes in PMN morphology were monitored by transmission electron microscopy. The ability of PMN to ingest and kill Staphylococcus aureus ATCC 25923 was significantly decreased by fat, skim, CL, CE, NO, and DP. Effects of some antibiotics on ingestion and killing of bacteria by PMN were influenced by the addition of vehicle and by interactions with mammary secretion. Neutrophil morphology was altered by fat, skim, CL, CE, NO, and DP. The detrimental effects of CL, CE, NO, and DP on PMN morphology were influenced (some significantly) by the presence of vehicle and interactions with mammary secretion. There were significant correlations among secretion- and antibiotic-induced changes in PMN ingestion of bacteria, PMN killing of bacteria, and PMN morphology.     

High milk neutrophil chemiluminescence limits the severity of bovine coliform mastitis

Polymorphonuclear neutrophil (PMN) function changes during mastitis. To investigate the contribution of milk PMN to the severity of Escherichia coli (E. coli) mastitis, chemiluminescence (CL) of blood and milk PMN and their efficiency to destroy coliform bacteria in the mammary gland were examined following the induction of E. coli mastitis in early lactating cows. To better assess and define the degree of mastitis severity, cows were classified as moderate and severe responders according to milk production loss in the non-infected quarters at post-infection hour (PIH) 48. There was an inverse relationship between pre-infection milk PMN CL and colony-forming units at PIH 6. In moderate cows, the pre-infection blood and milk PMN CL was approximately 2-fold higher than that of severe cows. The probability of severe response increased with decreasing pre-infection PMN CL. At the beginning of the infection blood and milk PMN CL was consistently higher, and milk PMN CL increased faster after infection in moderate cows. At PIH > 48 milk PMN CL in severe cows exceeded that of moderate cows. The somatic cell count (SCC) in moderate cows increased faster than colony-forming units, whereas in severe cows the results were reversed. The kinetics of CL activity for blood and milk PMN before and during the early phase of infection confirmed an impairment in PMN CL activity for severe responding cows. High pre-infection blood and milk PMN CL and the immediate increase of milk PMN CL and SCC after infection limited bacterial growth thereby facilitating the recovery of E. coli mastitis in moderate cows. Our study strengthens the idea that pre-existing milk PMN (a static part of the udder's immune defense) functions as a "cellular antibiotic" before and during infection, and low milk PMN CL is a risk factor for bovine coliform mastitis.     

Apoptosis and necrosis of blood and milk polymorphonuclear leukocytes in early and midlactating healthy cows.

Increased milk somatic cell counts (SCC) are used as an indicator for bovine mastitis. During mastitis, polymorphonuclear leukocytes (PMN) become the predominant cell type. Shortly after parturition, the severity of mastitis is increased and several PMN functions are downregulated. Apoptotic and necrotic processes of PMN could influence SCC and PMN functions. In this study, the percentages of apoptotic and necrotic PMN in blood and milk from early and midlactating healthy cows were compared. Apoptosis and necrosis of PMN were quantified using a dual-color flow cytometric procedure with fluorescein labeled annexin-V (green) and propidium iodide (red). Using this technique three different subpopulations of bovine PMN could be detected: apoptotic cells (high intensive green fluorescence), necrotic cells (high intensive green and high intensive red fluorescence) and viable cells (low intensive green and low intensive red fluorescence). Following a 4 h incubation of blood from both groups of cows at 37 degrees C to induce apoptosis, the mean percentage of apoptotic blood PMN was significantly higher (P < 0.01) in early lactating cows (15.1%, n = 9) compared with midlactating cows (5.3%, n = 10). The mean percentage of necrotic PMN remained lower than 5% in all cows. In contrast to blood, no significant difference was found between the percentage of apoptotic PMN in milk from early (41.2%, n = 7) and midlactating cows (34.0%, n = 8). The percentage of necrotic PMN in milk from early lactating cows (25.9%, n = 7) was significantly higher than that in midlactating cows (14.2%, n = 8) (P < 0.05). Higher percentages of apoptotic as well as necrotic PMN were consistently found in milk compared to blood in all cows. From these results, it can be concluded that spontaneously induced apoptosis was higher in blood PMN from early lactating cows than in blood PMN from midlactating cows. The higher percentage of necrotic milk PMN in early lactating cows than in midlactating cows could be explained by the induction of secondary necrosis.     

Effect of apoptosis on phagocytosis, respiratory burst and CD18 adhesion receptor expression of bovine neutrophils

Polymorphonuclear neutrophil leukocytes (PMN) play an important role in intramammary defense against infections by Escherichia coli. During mastitis, PMN are confronted with various inflammatory mediators that can modulate their function. In severely diseased cows, increased concentrations of lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-alpha (TNF-alpha) are detected in plasma. Binding of LPS to membrane bound CD14 molecules on monocytes cause release of inflammatory mediators such as TNF-alpha. Because apoptosis of PMN promotes resolution of inflammation and because the LPS and TNF-alpha response in milk and blood is related to the severity of E. coli mastitis, the effect on apoptosis of bovine PMN of increased concentrations LPS and TNF-alpha was studied together with the functionality of apoptotic PMN.Bovine PMN apoptosis, as determined with annexin-V, was induced with high concentrations of either LPS (1000 and 10,000ng/mL) or TNF-alpha (10,000ng/mL) in whole blood following a 6h incubation at 37 degrees C. The apoptosis inducing effect of LPS on PMN was not inhibited following coculture with either anti-bovine TNF-alpha or anti-ovine CD14 monoclonal antibodies. When compared to controls, apoptotic PMN had a similar level of CD18 expression but lacked phagocytic and respiratory burst activity. This is the first study reporting the effects of apoptosis on bovine PMN function. These functional impairments in apoptotic PMN could be important in contributing to the establishment of intramammary infection. Well functioning PMN could finally determine the severity of mastitis following an invasion of bacteria in the mammary gland.     

CD44 is highly expressed on milk neutrophils in bovine mastitis and plays a role in their adhesion to matrix and mammary epithelium

Mastitis, inflammation of the mammary gland, is a common and economically important disease in dairy animals. Mammary pathogenic organisms, such as Escherichia coli, invade the teat canal,milk ducts, and mammary alveolar space, replicate in mammary secretions, and elicit a local inflammatory response characterized by massive recruitment of blood polymorphonuclear neutrophil leukocytes (PMN) into the alveoli and milk ducts. CD44 is a trans-membrane glycoprotein previously shown to play a role in mediation and control of blood PMN recruitment in response to inflammatory signals. Here we show, for the first time, increased expression of CD44 on recruited milk PMN in bovine mastitis and the expression of a CD44 variant, CD44v10, on these PMN. Furthermore, we demonstrate that CD44 mediates specific adhesion of bovine blood PMN to hyaluronic acid and mammary epithelial cells. Our results suggest that in mastitis CD44 plays a role in recruiting blood PMN into the mammary glands, the exact nature of this role needs to be elucidated.     

The bovine neutrophil: Structure and function in blood and milk

Migration of polymorphonuclear neutrophil leukocytes (PMN) into the mammary gland provide the first line of defense against invading mastitis pathogens. Bacteria release potent toxins that activate white blood cells and epithelial cells in the mammary gland to secrete cytokines that recruit PMN that function as phagocytes at the site of infection. While freshly migrated PMN are active phagocytes, continued exposure of PMN to inhibitory factors in milk such as fat globules and casein, leads to altered PMN morphology and reduced phagocytosis. In the course of phagocytosing and destroying invading pathogens, PMN release chemicals that not only kill the pathogens but that also cause injury to the delicate lining of the mammary gland. This will result in permanent scarring and reduced numbers of milk secretory cells. The life span of PMN is limited by the onset of apoptosis. To minimize damage to mammary tissue, PMN undergo a specialized process of programmed cell death known as apoptosis. Macrophages quickly engulf and phagocytose apoptotic PMN, thereby minimizing the release of PMN granular contents that are damaging to tissue. The PMN possess an array of cell surface receptors that allow them to adhere and migrate through endothelium and to recognize and phagocytose bacteria. One receptor found on phagocytes that is receiving considerable attention in the control of infections by Gram-negative bacteria is CD14. Binding of lipopolysaccharide (LPS) to membrane bound CD14 causes release of tumor necrosis factor-alpha and sepsis. Binding of LPS to soluble CD14 shed from CD14-bearing cells results in neutralization of LPS and rapid recruitment of PMN to the site of infection. Recent advances in the fields of genomics and proteomics should greatly enhance our understanding of the PMN role in controlling intramammary infections in ruminants. Further, manipulation of PMN, through either recombinant proteins such as soluble CD14 that enhance PMN response or agents that mediate PMN apoptosis, may serve as novel therapeutics for the treatment of mastitis.     

Expression of β2-integrin on monocytes and blood polymorphonuclear leukocytes in the periparturient period in dairy cows

The hypothesis that an altered expression of CD11/CD18 on bovine circulating monocytes, polymorphonuclear leukocytes (PMN), or both, contributes to an increased mastitis susceptibility in periparturient cows was tested. Expression of CD18 and CD11a, -b, -c on bovine monocytes and PMN were assessed in 8 Friesian-Holstein cows by flow cytometry from 2 wk before calving to 5 wk after calving. Minor changes in adhesion molecule expression levels were detected throughout the experimental period. Compared with PMN, monocytes exhibited an expression level that was similar for CD18, higher for CD11a and CD11c, but lower for CD11b. Differences in density may reflect the relative importance of these adhesion molecules on both leukocyte types. In this study, the decreased number of milk resident macrophages and PMN observed during the periparturient period could not be attributed to changes of CD11/CD18 levels on circulating leukocytes.