Effect of lactoferrin on selected immune system parameters and the gastrointestinal morphology in growing calves.

Lactoferrin (LF) is a cationic iron-binding glycoprotein that is abundantly expressed and secreted from glandular epithelial cells and a prominent component of the secondary granules of polymorphonuclear neutrophils. Various in vitro and in vivo experiments demonstrate anti-microbial, -viral, -mycotic and -inflammatory effects of LF, associated with modulations of the immune system. Effects of oral administered LF on selected immune system parameters were studied in calves. Five calves were fed LF beginning on day 3 of life with colostral milk and starting on day 6 of life milk replacer enriched with 0.16% LF was fed. The average daily intake of LF per calf was 1.5-1.6 g/day. Additional five calves served as control group with identical treatment except for the LF supplementation. At the end of the study (day 61 of life), all calves were slaughtered and various tissues were sampled for histological and gene-expression studies. LF given orally was shown to act as an immunomodulatory agent by enhancing the size of Peyer's patches in the ileum and increasing blood serum immunoglobulin G levels. In addition, the number of peripheral blood leucocytes increased and mRNA levels of various interleukins (IL) such as IL-1beta, IL-8, IL-10 and interferon gamma (IFNgamma) in those cells in response to LF treatment were enhanced. In blood, the mRNA expression of the pro-inflammatory marker genes IL-1beta and IFNgamma decreased over 10-week treatment. Additionally, LF feeding decreased villus sizes in the jejunum. Together these findings emphasize the ability of LF to stimulate prominent immune system parameters and that it has the capacity to modulate the immune responses in a positive way.

Effects of mycophenolic acid on inosine monophosphate dehydrogenase I and II mRNA expression in white blood cells and various tissues in sheep.

Mycophenolic acid (MPA) is a mycotoxin commonly found as Penicillium genus secondary metabolite in feedstuffs and silages. Feeding with MPA contaminated silages may modulate the immune system in the farm animals and can cause appetite lost, ketosis, paralysis and abortion. The aim of the present study was to characterize the long-term MPA effect on both the inosine monophosphate dehydrogenase (IMPDH) isoforms I and II mRNA expression in white blood cells (WBC) and various tissue of healthy sheep. In treated animals 300 mg MPA/day/sheep was applied. In all investigated tissues the IMPDH I and II mRNA was abundant: WBC, spleen, thymus, ileum, jejunum, kidney, liver, pharyngeal and mesenterial lymph node. An efficiency-corrected relative quantification of the IMPDH types I and II isoforms mRNA were performed by normalizing with the constant reference gene expression of beta-actin. High IMPDH I mRNA expression levels were seen in kidney > mesenterial lymph node > jejunum > spleen > pharyngeal lymph node. Medium and low abundance was found in ileum > WBC > liver > thymus. Type II mRNA was highly expressed in liver > thymus > jejunum. In pharyngeal lymph node > spleen > ileum > mesenterial lymph node > kidney > WBC medium to low IMPDH II mRNA concentrations were detected. Under MPA treatment the IMPDH I mRNA expression was not significantly regulated in WBC, only trends of down- and upregulation were observed. Surprisingly in jejunum an upregulation could be observed (P < 0.05). In pharyngeal lymph node a tendency to downregulation was shown. This may be due to frequent ruminant activities and frequent exposition of MPA to the pharyngeal lymph nodes. In contrast to type I mRNA expression, IMPDH II mRNA was significantly downregulated in ileum (3.4-fold, P < 0.01) and tendencies in downregulation could be seen in jejunum (5.1-fold, P = 0.14). In addition, significant downregulation of IMPDH II gene expression over the entire feeding experiment could be shown in WBC of MPA-treated animals compared with untreated animals (P < 0.05). In conclusion, the recent study demonstrates that feeding sheep with MPA-contaminated silage did not induce IMPDH I mRNA expression in various tissues and blood, except in jejunum, but has suppressive effects on IMPDH II mRNA expression in WBC and ileum.

Short-term effects on pro-inflammatory cytokine, lactoferrin and CD14 mRNA expression levels in bovine immunoseparated milk and blood cells treated by LPS.

Leucocytes (WBC) are recruited from peripheral blood into milk as part of the inflammatory response, mediated through cytokines or interleukins (IL) synthesized by mammary tissue and the milk somatic cells (SC). The inflammatory response is related to the concentration of SC and the cytokines produced. To investigate and to compare the kinetics of cytokine production in SC and WBC during inflammation, cell culture models were established, where SC and WBC were cultured in parallel (n = 3). In addition, macrophages or monocytes were isolated from milk and blood with antibody-coated magnetic beads and cultivated separately. Isolated cells were pure, unaltered and viable. Cultures were activated with 10 microg/ml lipopolysaccharide (LPS). After 0, 1, 2, 3, 4 and 8 h cells were harvested for RNA isolation. Cytokine [tumour necrosis factor alpha (TNFalpha), IL-1beta, IL-6] mRNA expression responses and transcriptional activity of CD14 and lactoferrin (LF) were quantified via a one-step real-time RT-PCR. Significant cytokine mRNA increases were found in all four cell culture types and genes, with peaks after 1 and 2 h (TNFalpha > IL-6 > IL-1beta). In WBC or monocytes higher LPS responses and longer persistence could be found than in corresponding milk cells (IL-1beta > IL-6 > TNFalpha). SC and macrophages are less responsive to LPS stimulation than WBC or monocytes. The strength of the immune response in the blood system is much more prominent than in the mammary gland. This may be ascribed to the role of CD14 on the cytokine production of the investigated cells, or may be caused by the blood-to-milk diapedesis. The constitutive transcription of CD14 mRNA in WBC and monocytes was found to be 6 to 15 times higher than in adequate milk cells.