Prophylactic use of an ultra-diluted complex on polymorphonuclear leukocyte function and respiratory scores of weaned Holstein calves immediately after grouping

The aim of the study was to evaluate the effect of an ultra-diluted complex supplementation as a prophylactic strategy on immunity, performance, and respiratory scores of weaned Holstein calves immediately after grouping. Thirty-six weaned Holstein female calves (80.4±1.3 days old; 105.6±10.4 kg) were allocated to 6 pens (n=6 per pen) in a completely randomized design experiment in a double-blind, placebo-controlled trial. During a 28 days period, animals received a total mixed ration and were enrolled into two different groups (n=18 per group): 1) Control (basal diet + calcium carbonate, top-dressed at 30 g/animal/d – ultra-diluted placebo vehicle), or 2) ultra-diluted complex (basal diet + TopVita™-Real H, top-dressed at 30 g/animal/d – Sulphur:10-60 + Viola tricolor:10-14 + Caladium seguinum:10-30 + Zincum oxydatum:10-30 + Phosphorus:10-60 + Cardus marianus:10-60 + Colibacillinum:10-30 + Podophyllum:10-30 + Vehicle: calcium carbonate; q.s. 1kg). Blood samples were collected from each heifer at enrollment and 28 days later to assess polymorphonuclear leukocyte (PMNL) function and blood cell count. Body weight was assessed at enrollment and 28 days later at the end of the study. Regarding respiratory-screening process, a calf scoring system modified for calves in group pens was used. There was no effect of prophylactic ultra-diluted treatment on PMNL, nor it affected lymphocytes count and its ratio. Besides, the ultra-diluted product did not affect body weight and ADG. Further, no effect was observed in respiratory scores throughout the study period. In conclusion, the ultra-diluted complex did not improve blood cells count and PMNL function, nor it had impact on the performance of weaned Holstein calves after grouping.

Effects of ORAI calcium release-activated calcium modulator 1 (ORAI1) on neutrophil activity in dairy cows with subclinical hypocalcemia1

Hypocalcemia in dairy cows is often associated with inflammation-related disorders such as metritis and mastitis. The protein encoded by the Ca2+ release-activated calcium modulator 1 (ORAI1) gene is a membrane Ca2+ channel subunit that is activated when Ca2+ stores are depleted. Polymorphonuclear neutrophils (PMNL) have a crucial role in the defense against infection through migration, adhesion, chemotaxis, phagocytosis, and reactive oxygen species (ROS) production in response to pathogens. Whether hypocalcemia affects the activity of PMNL and if ORAI1 is involved remains unknown. To address this, PMNL were isolated at 3 d of calving from dairy cows diagnosed as clinically healthy (n = 20, CONTROL) or with plasma concentration of calcium < 2.0 mmol/L as a criterion for diagnosis of subclinical hypocalcemia (n = 20, HYPOCAL). PMNL isolated from both groups of cows were treated with or without the sarcoendoplasmic Ca2+ ATPase inhibitor thapsigargin, Ca2+ ionophore Ionomycin, and ORAI1 blocker 2APB. The intracellular Ca2+ concentration, ORAI1 abundance, ROS, phagocytosis rate, migration, and adhering capacity of treated PMNL were evaluated. Some of the in vitro assays also included use of small interfering ORAI1 RNA (siORAI1), 100 nM 1,25(OH)2D3, or 100 nM parathyroid hormone (PTH). Intracellular Ca2+ concentration was markedly lower in HYPOCAL. In addition, ORAI1 was detected in PMNL plasma membrane via FACS and was markedly lower in cows with HYPOCAL. Migration, adhesion capacity, and phagocytosis rate of PMNL were lower in response to HYPOCAL. Furthermore, plasma and PMNL concentration of nucleosome assembly protein (NAP2) and pro-platelet basic protein (CXCL7) was markedly lower with HYPOCAL. All these changes were associated with lower ROS production by PMNL. Thapsigargin and ionomycin treatment in vitro increased ORAI1 expression, migration of PMNL, adhering capacity, phagocytosis rate, and ROS production; conversely, those effects were abrogated by siORAI1 and ORAI1 inhibitor 2APB treatment. Also cytosolic Ca2+ concentration and ORAI1 abundance were increased by 1,25(OH)2D3 and PTH supplementation. Overall, the data indicate that failure of PMNL to uptake Ca2+ due to downregulation of ORAI1 during subclinical hypocalcemia is a factor contributing to impaired PMNL function. In addition, plasma PTH or 1,25(OH)2D3 could regulate ORAI1 and also participate in the regulation of PMNL activity.

Effects of inactivated Parapoxvirus ovis on polymorphonuclear leukocyte function and myeloperoxidase activity in horses

Immunomodulatory products have been used for years in veterinary medicine. Inactivated Parapoxvirus ovis (iPPVO) is currently used in equine medicine as an immunomodulator to improve the immune system and as a prophylactic treatment to prevent or treat infectious diseases. This study was designed to determine the effects of iPPVO on polymorphonuclear leukocyte (PMNL) function (phagocytosis and intracellular killing activity) and the myeloperoxidase (MPO) activity of PMNLs in horses. Twenty-four healthy English thoroughbred horses with an average age of 11 years were included in the study. Venous blood samples (10 ml) were taken before (agent-free controls) and after the administration of iPPVO (2 ml i.m. injection on Days 1, 3, and 5). PMNLs (1&nbsp;&times;&nbsp;10⁷ cells/ml) were isolated from venous blood containing EDTA (0.1 g/ml) with Ficoll-Hypaque gradient centrifugation. Cellular phagocytosis and intracellular killing activities were assayed using a modification of Alexander&rsquo;s method before and after treatment with iPPVO. MPO activity was also measured. The administration of iPPVO significantly increased the phagocytic, intracellular killing, and MPO activities of equine PMNLs (P = 0.0058, P&nbsp;= 0.0050, and P = 0.0070, respectively). This study demonstrates a strong correlation between MPO activity and PMNL function. The administration of iPPVO to horses has a supportive effect on their cellular immunity and an immunomodulatory effect against equine viral infections.

The Role of Spirochetes in Periodontal Disease

The spirochetal accumulation in subgingival plaque appears to be a function of the clinical severity of periodontal disease. It is not known how many different spirochetal species colonize the plaque, but based upon size alone, there are small, intermediate-sized, and large spirochetes. Four species of small spirochetes are cultivable, and of these, T. denticola has been shown to possess proteolytic and keratinolytic enzymes as well as factors or mechanisms which suppress lymphocyte blastogenesis and inhibit fibroblast and polymorphonuclear leukocyte (PMNL) function. All of these attributes could contribute to periodontal tissue insult. Yet independent of these potential virulence mechanisms, the overgrowth of spirochetes can be clinically useful if simply interpreted as indicating the result of tissue damage. In this case, the spirochetes would be indicators of disease and could be easily monitored by microscopic examination of plaque, or possibly by the measurement of benzoyl-DL-arginine-2-naphthylamide (BANA) hydrolytic activity in the plaque.