Expression profile of genes associated with mastitis in dairy cattle

In order to characterize the expression of genes associated with immune response mechanisms to mastitis, we quantified the relative expression of the IL-2, IL-4, IL-6, IL-8, IL-10, IFN-γ and TNF- α genes in milk cells of healthy cows and cows with clinical mastitis. Total RNA was extracted from milk cells of six Black and White Holstein (BW) cows and six Gyr cows, including three animals with and three without mastitis per breed. Gene expression was analyzed by real-time PCR. IL-10 gene expression was higher in the group of BW and Gyr cows with mastitis compared to animals free of infection from both breeds (p < 0.05). It was also higher in BW Holstein animals with clinical mastitis (p < 0.001), but it was not significant when Gyr cows with and without mastitis were compared (0.05 < p < 0.10). Among healthy cows, BW Holstein animals tended to present a higher expression of all genes studied, with a significant difference for the IL-2 and IFN- γ genes (p < 0.001). For animals with mastitis no significant difference in gene expression was observed between the two breeds. These findings suggest that animals with mastitis develop a preferentially cell-mediated immune response. Further studies including larger samples are necessary to better characterize the gene expression profile in cows with mastitis.     

Activation of Plasmin in Mastitic Milk

Milk and whey samples from healthy and inflamed udder quarters of 10 Ayrshire cows were analyzed for proteolytic activity using radial caseolysis procedures, a fluorogenic coumaryl peptide substrate, and casein agarose zymography. Free lysosomal enzyme activity (N–acetyl–beta–D–glucosaminidase) was used as the criterion for inflammation. All mastitic milk samples had proteolytic activity, tentatively identified as plasmin (comigration at Mr 83 000 and characteristic fragmentation). The plasmin activities in mastitic milk were on average 2.9 μg/ml (range 0.5–12.5) as measured by radial caseolysis. Milk or whey specimens from healthy quarters were all negative except 1 in which an activity of 0.1 μg/ml was found in both specimens. The caseolytic activities were totally inhibited by 50 KITJ/ml of aprotinin, a serine proteinase inhibitor from bovine lung. No free plasminogen activator (PA) activity was found in any of the samples. Howewer, according to zymographic analyses PA molecules corresponding to urokinase were found in healthy and especially in mastitic specimens. Analysis of plasmin may provide an alternative means of screening for mastitic milk samples.     

Solexa sequencing and custom microRNA chip reveal repertoire of microRNAs in mammary gland of bovine suffering from natural infectious mastitis

Identification of microRNAs (miRNAs), target genes and regulatory networks associated with innate immune and inflammatory responses and tissue damage is essential to elucidate the molecular and genetic mechanisms for resistance to mastitis. In this study, a combination of Solexa sequencing and custom miRNA chip approaches was used to profile the expression of miRNAs in bovine mammary gland at the late stage of natural infection with Staphylococcus aureus, a widespread mastitis pathogen. We found 383 loci corresponding to 277 known and 49 putative novel miRNAs, two potential mitrons and 266 differentially expressed miRNAs in the healthy and mastitic cows’ mammary glands. Several interaction networks and regulators involved in mastitis susceptibility, such as ALCAM, COL1A1, APOP4, ITIH4, CRP and fibrinogen alpha (FGA), were highlighted. Significant down‐regulation and location of bta‐miR‐26a, which targets FGA in the mastitic mammary glands, were validated using quantitative real‐time PCR, in situ hybridization and dual‐luciferase reporter assays. We propose that the observed miRNA variations in mammary glands of mastitic cows are related to the maintenance of immune and defense responses, cell proliferation and apoptosis, and tissue injury and healing during the late stage of infection. Furthermore, the effect of bta‐miR‐26a in mastitis, mediated at least in part by enhancing FGA expression, involves host defense, inflammation and tissue damage.     

Genome-wide microRNA profiling of bovine milk-derived exosomes infected with <Emphasis Type="Italic">Staphylococcus aureus</Emphasis>

Bovine milk is rich in exosomes, which contain abundant miRNAs and play important roles in the regulation of neonatal growth and development of adaptive immunity. Here, we analyzed miRNA expression profiles of bovine milk exosomes from three healthy and three mastitic cows, and then six miRNA libraries were constructed. Interestingly, we detected no scRNAs and few snRNAs in milk exosomes; this result indicated a potential preference for RNA packaging in milk exosomes. A total of 492 known and 980 novel exosomal miRNAs were detected, and the 10 most expressed miRNAs in the six samples accounted for 80–90% of total miRNA-associated reads. Expression analyses identified 18 miRNAs with significantly different expression between healthy and infected animals; the predicted target genes of differentially expressed miRNAs were significantly enriched in immune system process, response to stimulus, growth, etc. Moreover, target genes were significantly enriched in several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways including inflammatory, immune, and cancer pathways. Our survey provided comprehensive information about milk exosomes and exosomal miRNAs involved in mastitis. Moreover, the differentially expressed miRNAs, especially miR-223 and miR-142-5p, could be considered as potential candidates for mastitis.     

Effect of in vitro zinc supplementation on HSPs expression and Interleukin 10 production in heat treated peripheral blood mononuclear cells of transition Sahiwal and Karan Fries cows

The changing climatic scenario with apprehended rise in global temperature is likely to affect the livestock adversely vis-à-vis production and reproduction. This has prompted more focus in addressing the unfavorable effects of thermal stress in livestock system. Presuming that the trace element zinc is indispensible for cellular antioxidant system and immune function, the present study was designed to investigate the effect of zinc treatment on heat stress alleviation and immune modulation in peripheral blood mononuclear cells (PBMC) of indigenous and crossbred transition cows. Twelve cows, six each of Sahiwal and Karan Fries (KF) in their second parity with confirmed pregnancy were selected for the experiment. The blood samples were collected at −21, 0 and +21 days in relation to expected date of calving. The experiment was carried out in vitro after isolating PBMC from whole blood. The 48 h cultured PBMC were subjected to assorted levels of exposures viz. 37 °C, 42 °C to impose heat stress and 42 °C+zinc to alleviate heat stress and modulate immunity. The PBMC viability was 86%, 69% and 78%, respectively. The mRNA expression of heat shock proteins (HSP 40, 70 and 90α) and Interleukin-10 (IL-10) production varied between the two breeds vis-à-vis days and levels of exposure. The mRNA expression of HSP40 and HSP70 was significantly (P<0.05) higher in Karan Fries than the Sahiwal cows. Both the breeds showed maximum expression of HSP on the day of parturition, more so in KF than Sahiwal. There was a significant (P<0.05) difference in the HSP mRNA expression at different levels of exposure. Zinc treatment to heat stressed PBMC caused a significant (P<0.05) down regulation of HSP. For immune status, anti-inflammatory cytokine, IL-10 in the culture supernatant was accessed. The IL-10 was significantly (P<0.05) higher in Karan Fries (168.18±14.09 pg/ml) than the Sahiwal cows (147.24±11.82 pg/ml). The IL-10 concentration was highest on the day of calving. Zinc treatment reduced the IL-10 concentration. From the study, it could be concluded that the zinc supplementation in heat stressed PBMC can ameliorate thermal stress and modulate immune response which can act as a model for reducing heat stress during the periparturient period in tropical livestock.     

Micro RNA-19a suppresses IL-10 in peripheral B cells from patients with atherosclerosis

Background and aimsThe interleukin (IL)-10-production B cells play an important role in the pathogenesis of atherosclerosis (Asro) with unknown mechanism. Micro RNA (miR)-17-92 cluster has strong immune regulatory activities. This study tests a hypothesis that miR-17-92 cluster suppresses IL-10 expression in B cells of Asro patients.MethodsPatients with Asro were recruited into this study. Peripheral blood samples were collected from the patients. B cells were isolated from the blood samples and analyzed to elucidate the role of miR-17-92 in the regulation of IL-10 expression.ResultsPeripheral B cells from patients with Asro show lower levels of IL-10 than that from healthy subjects. The IL-10 expression in the B cells is negatively correlated with the expression of miR-19a in the B cells. The serum levels of tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-4 in Asro patients were higher than healthy subjects. Exposure to TNF-α or IFN-γ or IL-4 suppressed IL-10 expression in B cells via increasing the expression of miR-19a in B cells, which could be abolished by Inhibition of miR-19a.ConclusionsTNF-α or IFN-γ or IL-4 suppresses IL-10 in B cells via up regulating miR-19a expression.     

Application of the ELISPOT method for comparative analysis of interleukin (IL)-6 and IL-10 secretion in peripheral blood of patients with astroglial tumors

Glioblastoma, (grade IV astrocytoma), is characterized by rapid growth and resistance to treatment. Identification of markers of aggressiveness in this tumor could represent new therapeutic targets. Interleukins (IL)-6 and IL-10 may be considered as possible candidates, regulating cell growth, resistance to chemotherapy and angiogenesis. ELISPOT method provides a useful tool for the determination of the exact cell number of peripheral lymphocytes secreting a specific cytokine. IL-6 and IL-10 secretion levels were determined using ELISPOT methodology in peripheral blood mononuclear cells of 18 patients with astrocytic neoplasms (3 grade II and 15 grade IV), in parallel with 18 healthy controls. Additionally, immunohistochemical expression of these two cytokines was performed in paraffin-embedded neoplastic tissue in 12 of these patients. The secretion of IL-6 from peripheral monocytes was significantly higher in glioma patients compared to controls (P = 0.0003). In addition, IL-10 secretion from peripheral mononuclear and tumor cells of glioma patients was also higher as compared to healthy controls (P = 0.0002). Based on immunohistochemical staining, IL-6 expression was localized in tumor cells and macrophages as well as in areas of large ischemic necrosis, while the major source of IL-10 expression in glioblastomas was the microglia/macrophage cells. It is suggested that IL-10 contributes to the progression of astrocytomas by suppressing the patient’s immune response, whereas IL-6 provides an additional growth advantage. This study demonstrates for the first time the usefulness of ELISPOT in estimating the secretion of IL-6 and IL-10 from peripheral blood and the correlation of their expression in neoplastic cells. Christina Piperi and Penelope Korkolopoulou have equally contributed to this work.     

Microbial Diversity of Bovine Mastitic Milk as Described by Pyrosequencing of Metagenomic 16s rDNA

Dairy cow mastitis is an important disease in the dairy industry. Different microbial species have been identified as causative agents in mastitis, and are traditionally diagnosed by bacterial culture. The objective of this study was to use metagenomic pyrosequencing of bacterial 16S rRNA genes to investigate bacterial DNA diversity in milk samples of mastitic and healthy dairy cows and compare the results with those obtained by classical bacterial culture. One hundred and thirty-six milk samples were collected from cows showing signs of mastitis and used for microbiological culture. Additionally, 20 milk samples were collected from healthy quarters. Bacterial DNA was isolated from the same milk samples and the 16S rRNA genes were individually amplified and pyrosequenced. Discriminant analysis showed that the groups of samples that were most clearly different from the rest and thus easily discriminated were the normal milk samples from healthy cows and those characterised by culture as Trueperella pyogenes and Streptococcus spp. The mastitis pathogens identified by culture were generally among the most frequent organisms detected by pyrosequencing, and in some cases (Escherichia coli, Klebsiella spp. and Streptococcus uberis mastitis) the single most prevalent microorganism. Trueperella pyogenes sequences were the second most prevalent sequences in mastitis cases diagnosed as Trueperella pyogenes by culture, Streptococcus dysgalactiae sequences were the second most prevalent sequences in mastitis cases diagnosed as Streptococcus dysgalactiae by culture, and Staphyloccocus aureus sequences were the third most prevalent in mastitis cases diagnosed as Staphylococcus aureus by culture. In samples that were aerobic culture negative, pyrosequencing identified DNA of bacteria that are known to cause mastitis, DNA of bacteria that are known pathogens but have so far not been associated with mastitis, and DNA of bacteria that are currently not known to be pathogens. A possible role of anaerobic pathogens in bovine mastitis is also suggested.     

Differential expression of interferon-gamma,IL-4 and IL-10 in peripheral blood mononuclear cells during early pregnancy of the bovine

Maternal systemic immune response is regulated by conceptus-derived signals through peripheral blood mononuclear cells (PBMCs) via blood circulation during early pregnancy in cattle. In this study, the PBMCs from day 18 in non-pregnant cows and days 14, 18 and 30 in pregnant cows were used to explore the expression of interferon-gamma (IFN-γ), interleukin 4 (IL-4) and IL-10, and the plasma progesterone (P4) concentration was also determined. The results showed that the expression levels of mRNA and the protein of IFN-γ were lower and that IL-4 and IL-10 were higher in the PBMCs from the pregnant cows than in those of non-pregnant cows. From this study, early pregnancy induced a lower Th1 immunity (IFN-γ) and a higher Th2 immunity (IL-4 and IL-10) in the PBMCs, which may be related to interferon-tau and P4, thereby contributing to successful pregnancy in cattle.     

Innate immunity and metabolomic responses in dairy cows challenged intramammarily with lipopolysaccharide after subacute ruminal acidosis

Subacute ruminal acidosis (SARA) is a prevalent metabolic disorder in dairy cows known to elicit local and systemic immune responses. We recently showed that cows experiencing SARA and challenged intramammarily with lipopolysaccharide (LPS) experienced stronger metabolic disturbances compared with cows without SARA. Therefore, we hypothesized that cows experiencing SARA have a modulated innate immune response and impaired plasma metabolome compared with healthy cows when experiencing an acute mastitis challenge. A total of 18 Simmental cows were subjected either to a Control (CON, n=6) or SARA (n=12) feeding regimen, receiving either 40% or 60% concentrates for 30 days. Thereafter, six SARA (SARA-LPS) and the CON (CON-LPS) cows were intramammarily challenged with 50 µg LPS from Escherichia coli (O26 : B6), while the remaining six SARA cows (SARA-PLA) received a placebo. Blood and milk samples were analyzed for acute phase proteins and a targeted ESI-LC-MS/MS-based metabolomics approach was performed in blood samples 24 h after the LPS challenge. The LPS infusion caused a strong increase in immune response variables, with a higher concentration of milk amyloid A 48 h after the LPS challenge in SARA-LPS compared with CON-LPS cows. Cows receiving the LPS infusion had a lower plasma concentration of several amino acids and lysophosphatidylcholines but without differences in SARA cows and healthy cows. In conclusion, our results revealed that an intramammary LPS infusion increased acute phase proteins and modulated the blood metabolome. While no systemic differences between SARA and healthy cows were observed, cows experiencing SARA showed a higher concentration of an acute phase protein at the local level of the mammary gland. Further research is required to elucidate the underlying mechanisms and to evaluate its clinical significance for udder health.     

Effect of vitamin D treatments on plasma metabolism and immune parameters of healthy dairy cows

The objective of this study was to investigate the possible beneficial effect of vitamin D repletion on certain immune parameters of vitamin D insufficient dairy cows. Twenty dairy cows in late lactation were treated daily with vitamin D in five different ways: sunlight exposure (SUN), D₂ supplementation combined with sunlight exposure (D2SUN), D₂ supplementation (D2), D₃ supplementation (D3), and D₂ and D₃ supplementation combined (D2D3). The cows had very low vitamin D levels at d 0 because of the vitamin D deprivation before the study. After 1 month of vitamin D repletion, all cows had plasma 25(OH)D levels within the normal range. Total 25(OH)D concentration was significantly higher in SUN, D2SUN and D2D3 than D2 or D3 at the end of the study. However, milk yield, as well as protein and fat content of the milk, was not influenced by vitamin D treatments. There was no difference obtained in the measured immune parameters: Leucocyte populations, somatic cell count, immunoglobulin concentrations in plasma and milk, and antigen-stimulated cytokine productions did not change in response to vitamin D repletion or difference in vitamin D sources, and no relations to plasma 25(OH)D levels were identified. Despite the fact that plasma 25(OH)D increased from a very low level to normal range, the present study did not show any effect of vitamin D repletion on the tested immune parameters of healthy dairy cows. Therefore, in this study, it was concluded that repletion to physiologically normal plasma 25-hydroxyvitamin D levels of vitamin D-depleted healthy dairy cows had no influence on immune parameters.     

类风湿性关节炎患者外周血单个核细胞 IL-10 基因启动子甲基化状态研究

白细胞介素-10（interleukin-10,IL-10）是在类风湿性关节炎中发挥重要免疫调节作用的细胞因子,其基因失活与已分化的Th1和Th2细胞染色质结构重塑有关。为了探讨IL-10基因启动子甲基化及基因失活在类风湿性关节炎(Rheumatoid Arthritis,RA)发病和进展中的作用,采用逆转录聚合酶链反应(RT-PCR)、酶联免疫吸附实验(enzyme linked immunosorbent assay,ELISA)及甲基化特异性聚合酶链反应(MSP), 分别检测34例类风湿性关节炎患者和30例健康人外周血单个核细胞 IL-10 mRNA、蛋白表达水平及基因启动子甲基化状态。结果显示,病例组IL-10 mRNA及蛋白表达均低于健康对照组,无统计学差异（P>0.05）。病例组甲基化率(85.29%)高于健康对照组(43.33%), 具有统计学差异(x² =12.439,P=0.000)。IL-10基因启动子甲基化状态与其mRNA表达呈显著负相关(r=-0.579, P=0.001), 与所累关节数显著相关,但与血沉（ESR）、C反应蛋白（CRP）、类风湿因子(RF)、年龄均无相关性(P>0.05)。IL-10 mRNA表达与年龄、所累关节数、ESR、CRP及RF均无相关性(P>0.05)。上述结果提示,启动子甲基化可能是IL-10基因失活的重要机制,IL-10基因异常高甲基化状态可能参与了RA的发生发展。     

High level expression of the bioactive human interleukin-10 in milk of transgenic mice

Human interleukin-10 (hIL-10) has wide spectrum of anti-inflammatory activities and has shown a potential to be used for treatment of inflammatory or immune illness. In this study, transgenic mice that over-express human interleukin-10 (IL-10) in their milk were generated using a bovine beta-casein/human IL-10 hybrid gene. After cloning of the IL-10 gene, a 22 kb hybrid gene was constructed by linking a 10 kb promoter sequence of the bovine beta-casein gene to the cloned 12 kb IL-10 gene. In six of the eight transgenic mice, the transgene RNA was expressed only in the mammary gland and in the other two mice, it was also slightly expressed in the lung. The highest human IL-10 level in milk was 1620 microg x ml(-1). Notably, transgenes in all the eight transgenic mice were expressed regardless of the integration site even though no correlation was shown between the copy numbers of the transgene and expression level. These results suggest that the genomic sequence of the human IL-10 gene can induce the IL-10 expression at high levels under the control of the bovine beta-casein promoter.     

Prevalence of Prototheca spp. on dairy farms in Poland – a cross-country study

The Prototheca algae have recently emerged as an important cause of bovine mastitis globally. Here, we present results of a first large-scale, cross-country survey on the prevalence of Prototheca spp. in dairy cows, and their environment in Poland. A total of 1211 samples were collected and microbiologically analysed. Included within this number were milk (n = 638), body swabs (n = 374) and environmental samples (n = 199), originating from 400 dairy cows and their surroundings, on 16 dairy farms, based in all major provinces of the country. Prototheca spp. were the third, after Streptococcus and Staphylococcus spp., most common mastitis pathogens. The overall prevalence of protothecal mastitis was 8.3% (33/400), with the majority (75.8%) of cases having a subclinical course, and all but one attributable to P. zopfii genotype 2. Prototheca spp. were cultured from body swabs of both healthy and mastitic cows, yet the isolation rate among the latter was conspicuously lower (12.3% vs. 17.8%). Forty-two (21.2%) environmental samples yielded growth of Prototheca spp. However, no clear association between Prototheca mastitis in dairy cows and the algal isolation from the herd environment was found. Nor was there any association between the environmental recovery of the algae and farm management practices.     

Interleukin-10 and interleukin-18 promoter polymorphisms in an Italian cohort of patients with undifferentiated carcinoma of nasopharyngeal type

Purpose: Cytokines such as IL-10 and IL-18 seem to be involved in the inflammatory response of undifferentiated carcinoma of nasopharyngeal type (UCNT). The aim of this study was to evaluate the correlation between functional single nucleotide polymorphisms (SNPs) in the promoter region of IL-10 and IL-18 genes and the virological and clinical characteristics in a large case series of Caucasian patients suffering from UCNT, a tumor regularly associated with the Epstein Barr Virus (EBV). Methods: Eighty-nine patients with histologically confirmed UCNT and 130 healthy donors were included in our study. DNA was examined for the polymorphisms of IL-10 gene at positions –1082, −819, −592 by direct sequencing and IL-18 gene at position −607 and −137 by allele –specific PCR. EBV DNA serum viremia was evaluated by QC-PCR. Results: The distributions of the IL-10 and IL-18 genetic variants were not different between UCNT patients and healthy controls. The frequency of IL-10 –1082G allele, which is associated with high IL-10 expression, showed a nearly statistically significant increase in UCNT patients EBV DNA-negative as compared to healthy controls (OR=3.3 95% CI: 1.2–9.8). Subjects with C/C or C/G combined IL-18 genotypes showed an increased risk of being with Stages III-IV (OR=2.1 95% CI: 1.2–6.6). Conclusion: This study was performed to improve the definition of the pathogenetic factors implicated in UCNT by addressing the correlation between cytokine polymorphisms and clinical parameters. This is the first study investigating the possible role of the IL-18 and IL-10 polymorphisms in the development and outcome of UCNT. In our genetic analysis there is no evidence for involvement of IL-10 promoter polymorphisms alone in the genetic predisposition to this tumor. On the other hand, IL18 genetic variants may represent a genetic risk factor for tumor aggressiveness.     

Antimicrobial resistance and toxin gene profiles of Staphylococcus aureus strains from Holstein milk

Isolation of Staphylococcus aureus (Staph. aureus) from Holstein milk samples with mastitis and nonmastitis was conducted to estimate its prevalence, antimicrobial resistance and toxin genes. A total of 353 milk samples were collected from three Chinese Holstein herds. Fifty‐three Staph. aureus isolates collected from 29 Staph. aureus‐positive samples were characterized via antimicrobial susceptibility, toxin genes and Pulsed‐field Gel Electrophoresis (PFGE) profiles. The prevalence of Staph. aureus was 4·0–9·5% in mastitic and 7·3–11·5% in nonmastitic samples in the analysed herds. Approximately 61·0% of Staph. aureus strains isolated from mastitis cows were resistant to ≥10 antimicrobials compared with 0% of isolates with nonmastitis. The most frequently observed super antigenic toxin gene was pvl (41·5%) followed by seh + pvl (13·2%). We did not find mecA‐positive methicillin‐resistant Staph. aureus (MRSA) strains, while mecA‐negative MRSA strains were identified in the three herds. PFGE results suggested potential transmission of Staph. aureus strains in different farms. These results open new insights into Staph. aureus transmission and antimicrobial resistance of Holstein dairy cows and into developing strategies for udder health improvement of dairy cattle.     

Supplementation of antioxidant micronutrients reduces stress and improves immune function/response in periparturient dairy cows and their calves

BackgroundPeriparturient period induces stress in cows which fluctuates hormonal and metabolic function and causes immune suppression. Apart from impairing the health, production, and reproduction of cows, it also influences the well-being of newborn calves by decreasing the colostrum quality. Micronutrients are known for optimal health and production and their effects on parturition stress, immune response in both cow and its calf need to be explored.AimThe aim of this study was to see the effect of oral supplementation of micronutrients during the prepartum period on the health status of crossbred dairy cows and subsequently on their newborn calves.MethodsA total of 42 healthy multiparous cows were selected and randomly divided into five groups with seven cows in each group, i.e. control (Basal Diet, BD), VA group (BD + vitamin A, 10⁵ IU), Zn group (BD + zinc sulphate, 60 ppm), VE group (BD + vitamin E, 2500 IU), and combined supplementation (CS) group (BD + combination of VA, Zn, and VE). The supplements were offered in compounded concentrate DM (100 g) to individual cows once daily before the morning feeding and the remaining portion was incorporated in the TMR. Feeding was started one month before the expected days of calving till calving. Blood samples were collected from cows at days -15, -7, -3, 0, +3, +7, and +15 relative to the day of calving. Blood samples from newborn calves and milk samples of cows were collected at days 0, +3, +7, and +15. Milk somatic cell counts (SCC) were estimated using a cell counter. Cortisol was estimated by ELISA kit in blood and milk plasma of cows and in the blood plasma of their calves. Total immunoglobulins (Ig) were estimated in milk of cows and serum of calves using zinc sulphate turbidity method. Blood neutrophils from cows and calves were studied for phagocytic activity (PA) using nitro blue tetrazolium (NBT) assay.Data were analysed by repeated-measures two-way ANOVA using the mixed procedure of SAS, and the pairwise comparison was performed using a multiple comparison test (Tukey).ResultsCombined supplementation of micronutrients decreased (P < 0.05) maternal blood plasma (control vs. CS group, 5.98 ± 0.20 vs. 3.86 ± 0.23 ng/mL) and milk plasma (3.96 ± 0.13 vs. 2.71 ± 0.10 ng/mL) cortisol, milk SCC (3.05 ± 0.11 vs. 2.12 ± 0.10 × 10⁵ cells/mL) and increased (P < 0.05) total milk Ig concentration (18.80 ± 0.11 vs. 23.04 ± 0.57 mg/mL) and the PA of blood neutrophils (0.84 ± 0.03 vs. 1.07 ± 0.03). Similarly, lower blood cortisol concentration (9.69 ± 0.35 vs. 6.02 ± 0.18 ng/mL) and higher (P < 0.05) total Ig (23.26 ± 0.11 vs. 30.34 ± 0.70 mg/mL) and PA of blood neutrophils (0.37 ± 0.02 vs. 0.52 ± 0.02) were observed in the calves born to CS group of cows as compared to the control. Highest (P < 0.05) positive effects (lower stress levels and higher immune response) of treatment were noticed in CS group followed by VE group and then Zn group. However, VA group didn’t differ from the control group.ConclusionOur results indicate that micronutrient interventions during the prepartum period can improve the health status of dairy calves and subsequently the well-being of their calves.     

Involvement of IL-10 and Bcl-2 in resistance against an asbestos-induced apoptosis of T cells

To analyze the possibility that immunological alteration in asbestos-related diseases (ARDs) such as asbestosis (ASB) and malignant mesothelioma (MM) may affect the progression of cancers, a human adult T cell leukemia virus-immortalized T cell line (MT-2Org) was continuously exposed to 10 μg/ml of chrysotile-B (CB), an asbestos. After at least 8 months of exposure, the rate of apoptosis in the cells became very low and the resultant subline was designated MT-2Rst. The MT-2Rst cells were characterized by (i) enhanced expression of bcl-2, with regain of apoptosis-sensitivity by reduction of bcl-2 by siRNA, (ii) excess IL-10 secretion and expression, and (iii) activation of STAT3 that was inhibited by PP2, a specific inhibitor of Src family kinases. These results suggested that the contact between cells and asbestos may affect the human immune system and trigger a cascade of biological events such as activation of Src family kinases, enhancement of IL-10 expression, STAT3 activation and Bcl-2 overexpression. This speculation was partially confirmed by the detection of elevated bcl-2 expression levels in CD4 + peripheral blood T cells from patients with MM compared with those from patients with ASB or healthy donors. Further studies will be required to verify the role of T cells with enhanced bcl-2 expression in tumor progression induced by asbestos exposure.