Immunological attributes of blood and milk neutrophils isolated from crossbred cows during different physiological conditions (original) (raw)
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Frontiers in Veterinary Science, 2021
Milk somatic cell counts (SCCs) have been used as a gold standard to monitor mammary health as well as an indicator of raw milk quality. The present work was undertaken to compare the changes in the milk SCC, milk differential leukocyte counts (DLCs), phagocytic activity (PA) of milk neutrophils and macrophages (by nitroblue tetrazolium assay), extracellular trap formation (PicoGreen assay) and mRNA expression of various genes in milk neutrophils and macrophages (reverse transcription-polymerase chain reaction), and milk plasma cortisol concentration (enzyme-linked immunosorbent assay) in healthy, subclinical mastitis (SCM), and clinical mastitis (CM) cows. Milk was collected from healthy, SCM, and CM cows grouped based on their SCCs and California mastitis test with eight cows in each group. Milk SCC was estimated by SCC counter, and DLC was done after staining the milk slide under a microscope at 100×. Total SCCs in healthy, SCM, and CM cows were on an average of 128.30, 300.3, and 694.40 × 10 3 cells/mL, respectively. Milk DLCs indicated a lower percentage of macrophage and lymphocytes and a higher (p < 0.05) percentage of neutrophils in SCM and CM compared to healthy milk. The percentage of mature segmented neutrophils was lower, whereas immature band neutrophils were higher (p < 0.05) in the SCM and CM groups as compared to healthy cows. The viability, in vitro PA, and extracellular trap formation of neutrophils were lower (p < 0.05) in SCM and CM milk samples as compared to healthy samples. However, the PA of macrophage remained unchanged in all the studied groups. The relative mRNA expression of Toll-like receptors (TLR2, TLR4), myeloperoxidase, and interleukin 2α (IL-2α) receptor (CD25) were minimum in healthy samples and increased (p < 0.05) with the progress of mammary inflammation. However, CD44 decreased (p < 0.05), and CD62L remained unchanged in mastitis as compared to healthy cows. Plasma cortisol concentrations were higher (p < 0.05) in mastitis as compared to healthy cows and were negatively correlated with the number of milk macrophages and the functions of milk phagocytes. Estimation of total SCC, milk DLC, and activity of milk phagocytes is essential for effective control and prevention of incidence of mastitis in dairy cows.
Immunologic Research, 2019
Neutrophils represent the first line of innate immunity and are the most prominent line of cellular defence against invading microorganisms. On stimulation, they can quickly move through the walls of veins and into the tissues of the body to immediately attack or monitor the foreign antigens. Neutrophils are highly versatile and sophisticated cells which are endowed with highly sensitive receptor-based perception systems. They were traditionally classified as short-lived phagocytes actively involved during infection and inflammation, but recently, it has been seen that neutrophils are capable of detecting the presence of sperms during insemination as well as an implanting embryo in the female reproductive tract. These specialised phagocytes play a major role in tissue remodelling and wound healing, and maintain homeostasis during parturition, expulsion of placenta, folliculogenesis, corpus luteum formation and luteolysis. Here, we review the role played by neutrophils in maintaining homeostasis during normal and inflammatory conditions of dairy cattle. We have summarised the alteration in the expression of some cell adhesion molecules and cytokines on bovine neutrophils during different physiological and physiopathological conditions. Some emerging issues in the field of neutrophil biology and the possible strategies to strengthen their activity during the period of immunosuppression have also been discussed.
American Journal of Veterinary Research, 2004
Objective—To determine whether proinflammatory mediators and glucocorticoids affect CD62L(L-selectin) expression on peripheral blood neutrophils from cows in various stages of lactation.Animals—100 healthy dairy cows during early (13.1 ± 0.79 days after parturition; n = 31), peak (58.7 ± 1.64 days after parturition; 31), and mid (137.2 ± 2.59 days after parturition; 38) lactation.Procedure—In vitro effects of relevant proinflammatory mediators that are released in response to mastitis caused by gram-negative bacteria such as lipopolysaccharide (endotoxin), tumor necrosis factor-α, and platelet-activating factor (PAF) on CD62L expression on bovine neutrophils were assessed by flow cytometry. Influences of cortisol and dexamethasone on CD62L expression on bovine neutrophils were also investigated.Results—Basal CD62L expression on neutrophils from cows during early, peak, and mid lactation were similar. Lipopolysaccharide and tumor necrosis factor-α had no effect on CD62L expression on...
Journal of dairy science, 2006
The objectives of this experiment were to compare the circulating concentrations of cortisol and determine whether these correlated with the expression profiles of a set of candidate apoptosis genes in neutrophils of Holstein-Friesian cows milked once vs. 3 times daily for 28 d postpartum. Cows on the once-daily milking regimen had significantly higher plasma cortisol concentrations on d 3, 14, and 28 postpartum than did those milked 3 times daily. On d 3 postpartum, when differences in cortisol and neutrophil counts were highest between the groups, mean mRNA abundance of nuclear factor kappaB p65 subunit, IkappaBalpha, X-linked inhibitor of apoptosis; and heat shock protein 70 were higher in neutrophils of the cows milked once daily than in cows milked 3 times daily. However, no correlations were detected among plasma cortisol concentration, neutrophil count, or neutrophil gene expression in this study. Results suggest that the modest neutrophilia associated with once daily milking...
Veterinary Immunology and Immunopathology, 2015
The transition period is known to be the most critical phase in the life of high yielding dairy cow. Changes in the immune functions have been observed during the transition period which may account for the onset of clinical and subclinical (e.g. inflammatory response) problems at calving or at the beginning of lactation however this relationship has not yet been adequately investigated. Thus, to establish the potential of the periparturient dairy cow's immune system to respond to stimuli, two challenges [an ex vivo whole blood stimulation assay (WBA) with lipopolysaccharides and a carrageenan skin test (CST)] were performed in addition to characterizing the metabolic and inflammatory profile. The WBA was performed using 0, 0.01 and 5 g LPS/mL on whole blood and CST was administered by subcutaneous injection of 0.7 mL solution containing 4.2 mg of carrageenan to the shoulder region of the cows. These tests were performed on 10 Holstein-Friesian cows at −45 ± 2, −20 ± 2, −3, 3, 7, 28 ± 2 days from parturition (DFP). Cows were also monitored for health status, body condition score, milk yield. The results demonstrate a higher production of IL-1 and IL-6 from leukocytes after LPS stimulation around calving (from −3 to 3 DFP) compared to −45 DFP (P < 0.05). Moreover, IL-6 (but not IL-1) was able to reach close to the maximum response at the lower stimulus intensity (0.01 g LPS/mL), maintaining a higher response over a longer time in early lactation. The release of higher levels of IL-6 in the transition period, with low LPS dose, suggests its crucial role in the regulation of inflammatory response around calving. The response of cows to CST decreased a few days before calving (−3 DFP) compared with response at −45 and 28 DFP (P < 0.05), and remained low in the first week of lactation. This result suggests the reduction of the functionality of some vascular factors, which decreases diapedesis. Overall, the WBA and CST tests confirm changes in immunocompetence around calving. These tests are able to better describe the changes of the innate immune response at a local and systemic level, mainly when combined with conventional metabolic and inflammatory indices.
2001
Waller K: Changes in some blood micronutrients, leukocytes and neutrophil expression of adhesion molecules in periparturient dairy cows. Acta vet. scand. 2001, 42, 139-150.-Dairy cows are highly susceptible to infectious diseases, like mastitis, during the period around calving. Although factors contributing to increased susceptibility to infection have not been fully elucidated, impaired neutrophil recruitment to the site of infection and changes in the concentrations of some micronutrients related with the function of the immune defence has been implicated. Most of the current information is based on studies outside the Nordic countries where the conditions for dairy cows are different. Therefore, the aim of the study was to evaluate changes in blood concentrations of the vitamins A and E, the minerals calcium (Ca), phosphorous (P), and magnesium (Mg), the electrolytes potassium (K) and sodium (Na) and the trace elements selenium (Se), copper (Cu) and zinc (Zn), as well as changes in total and differential white blood cell counts (WBC) and expression of the adhesion molecules CD62L and CD18 on blood neutrophils in Swedish dairy cows during the period around calving. Blood samples were taken from 10 cows one month before expected calving, at calving and one month after calving. The results were mainly in line with reports from other countries. The concentrations of vitamins A and E, and of Zn, Ca and P decreased significantly at calving, while Se, Cu, and Na increased. Leukocytosis was detected at calving, mainly explained by neutrophilia, but also by monocytosis. The numbers of lymphocytes tended to decrease at the same time. The mean fluorescent intensity (MFI) of CD62L and CD18 molecules on blood neutrophils remained constant over time. The proportion of CD62L + neutrophils decreased significantly at calving. The animals were fed according to, or above, their requirements. Therefore, changes in blood levels of vitamins, minerals and trace elements were mainly in response to colostrum formation, changes in dry matter intake, and ruminal metabolism around calving. Decreased levels of vitamins A and E, and of Zn at calving might have negative implications for the functions of the immune defence. The lower proportion of CD62L+ neutrophils at calving may result in less migration of blood neutrophils into the tissues, and might contribute to the increased susceptibility to infections at this time.
Journal of Thermal Biology, 2018
The present study was undertaken to compare the effect of different seasons on the mammary immunity of three Indian native breeds of cows (Tharparkar, Gir and Sahiwal) well adapted to the tropical region. For this milk samples were collected from cows in winter (THI= 57, comfortable zone), hot-dry (HD; THI= 76, heat stressful zone) and hot-humid (HH; THI= 82, severe heat stress) and estimated for milk somatic cell counts (SCC), phagocytic activity (PA) of milk neutrophils, milk cortisol and heat shock proteins and function associated genes in milk neutrophils. Milk SCC was evaluated using a cell counter and differential cell counts measured microscopically. Cortisol was quantified in skimmed milk by competitive ELISA. Milk PA was estimated using nitro blue tetrazolium assay, and for gene expression studies, milk neutrophils were isolated and studied for heat shock proteins (HSP40, HSP70, HSP90α) and cell adhesion molecules (CD11b, CD25, CD44) using real-time polymerase chain reaction. All the studied parameters increased in HD and HH seasons with highest values observed in Sahiwal cows. However, PA of neutrophil was highest in Tharparkar cows in winter and decreased gradually at higher THI values during hot seasons. Milk cortisol was positively correlated with expression of various CD molecules and HSPs (p<0.05) in milk neutrophils but negatively correlated (p<0.05) with PA during HH season in all breeds. The study revealed that Indian native cows were at considerable risk in HH season and Sahiwal cows were more heat stressed followed by Gir and Tharparkar cows, respectively, and thus may require managemental interventions. Also, the higher expression of HSP70 and CD25 with increasing THI levels in hot seasons makes them suitable biological markers for quantifying heat stress.
Journal of dairy science, 2009
Polymorphonuclear neutrophilic leukocytes (PMNL) play an important role in the first line cell-mediated immune defense of the body in general and of the mammary gland against mastitis pathogens in particular. Reduced viability of PMNL close to parturition may explain the high incidence of infectious diseases and the high prevalence of intramammary infections (IMI) in periparturient dairy heifers. Apoptosis of blood PMNL 1 wk before the expected calving date and of blood and milk PMNL at 1 to 4 d in milk was determined using flow cytometry. Information on heifer and gland characteristics was collected before calving and in early lactation. Data were analyzed using multivariable, multilevel regression analysis. Supplementation of a commercial mineral/vitamin mix before calving was associated with less blood (14.4 +/- 2.9 vs. 22.4 +/- 2.1%) and milk PMNL apoptosis (19.0 +/- 1.1 vs. 26.4 +/- 0.9%) near calving, presumably related to higher blood selenium concentrations. Both blood and m...
Biological Rhythm Research, 2017
Chances of mammary infections are comparatively higher in high producing cows during harsh environmental conditions and are usually characterised by changes occurring in various somatic cells secreted in the milk and their activities. The present study was conducted to record diurnal rhythmicity in milk somatic cell counts (SCC), neutrophil: macrophage (N: M) ratio, phagocytic activity (PA) of milk neutrophils and plasma cortisol concentrations during different seasons and parity in high producing Karan Fries cows. Values of milk SCC, N: M ratio and plasma cortisol levels were lowest during thermoneutral (TN), intermediate in winter and highest during the summer season. Diurnal rhythm in the milk SCC and N: M ratio was noticed in the summer while plasma cortisol exhibited diurnal rhythm in both winter and summer seasons. Milk SCC, N: M ratio and plasma cortisol increased in multiparous cows, but diurnal variation was noticed only in the N: M ratio and plasma cortisol in cows having more than four parity. Phagocytic activity of milk neutrophils was highest during TN, intermediate in winter and lowest during the summer season. Phagocytic activity was higher and similar in cows up to fourth parity but decreased in subsequent lactation cycles. Diurnal rhythm in the PA was noticed in winter and summer seasons and in cows having more than four parity where morning samples showed higher phagocytosis as compared to the evening samples. These results can be used for immunomodulatory interventions and therapeutic approaches in treating mastitis of crossbred cows reared under tropical conditions.
Veterinary World, 2015
The present study was aimed to study the use of cell counts as an early indicator of mammary health. Materials and Methods: Milk and blood cell counts were estimated from 8 healthy, 8 subclinical (SCM), and 8 clinically mastitis (CM) groups of Karan Fries (KF) cows. Results: Total leucocyte counts and neutrophil percent in blood and milk somatic cells and milk neutrophil percent of healthy cows increased significantly (p<0.05) in SCM cows and CM cows. Viability of blood and milk neutrophils was more in healthy cows, but decreased significantly (p<0.05) in SCM and CM cows. Significant (p<0.05) decrease were also observed in both the blood and milk lymphocytes and monocytes of SCM and CM cows. Phagocytic activity (PA) of blood neutrophils also decreased significantly (p<0.05) in SCM cows. There was no difference between the PA of SCM and CM cows. Milk neutrophil percent was more in the SCM and clinically infected milk than in the blood of these cows. About 96-97% of the neutrophils had segmented nucleus in both healthy and subclinical milk, whereas, 2-3% were having band shaped or immature nuclei. There was a significant decrease in the segmented neutrophils, whereas, band neutrophils increase significantly to about 5% in the infected milk of mastitic cows. Viability of the milk neutrophils decreased more in case of subclinical and clinical milk as compared to that of blood. PA was found to be highest in the milk of healthy group of cows, but decreased significantly (p<0.05) in subclinically infected cows. However, there was no difference between the PA of milk neutrophils of SCM and CM cows. PA of milk was also found to be significantly lower in the milk of healthy cows when compared to that of blood neutrophils. Conclusion: This study indicated that percent neutrophils and their type in conjunction with milk somatic cell counts can be used as a more reliable indicator of mammary health in cows.