LACTOFERRIN AND BACTERIAL GROWTH

Iron is an essential growth factor for virtually all bacteria. Lactoferrin chelates iron and impairs bacterial multiplication and has a direct bactericidal action on some microorganisms.     

Resistance to infection of the external eye: The role of tears

The external eye is continuously exposed to an environment containing potentially pathogenic microorganisms. One of the mechanisms which protects the eye from infection is the tear layer. We review the current knowledge of those antimicrobial substances known to be present in tears and the role they might play in preventing infection. These substances include lysozyme, lactoferrin, beta-lysin, and the antibody-complement system of proteins.     

Serum and plasma measurements of neutrophil granule proteins during cardiopulmonary bypass: a model to estimate human turnover of lactoferrin and myeloperoxidase

During extracorporeal circulation (ECC), granule proteins such as lactoferrin and myeloperoxidase escape to the plasma in large amounts. ECC was used to study the turnover of these proteins and compare their variations in plasma and serum. During ECC both proteins rose to very high levels and the variations were similar in both serum and plasma (r = 0.93 for lactoferrin and 0.80 for myeloperoxidase). The initial elimination after termination of operation (t 1/2) was, for lactoferrin 0.75 and for myeloperoxidase 0.5 h. Both proteins followed a second order kinetics of elimination with a t 1/2 for lactoferrin of about 9 and for myeloperoxidase of about 25 h. 48 and 72 h postoperatively serum levels, but not plasma levels, of myeloperoxidase rose again 10-fold. We conclude that serum and plasma measurements of neutrophil granule proteins are complementary. The period after disconnection of the patient from the extracorporeal device may be used to estimate the turnover kinetics of neutrophil granule proteins.     

Donor neutrophil function after plateletpheresis

BACKGROUND: Neutrophils are important mediators of inflammation and may be activated by foreign surfaces in apheresis systems. Because most of the WBCs are returned to the donor, it was investigated whether artificial activation leads to altered donor neutrophil function. STUDY DESIGN AND METHODS: Three apheresis systems (Amicus, Autopheresis-C, and CS-3000; all: Baxter Fenwal) were investigated. Preapheresis and postapheresis blood samples were drawn from 10 volunteer donors, with all three apheresis systems used in random order for each donor. Changes in neutrophil phagocytic ability, oxidative burst, and expression of L-selectin and CD11b were measured by flow cytometry, and plasma concentrations of myeloperoxidase and lactoferrin were measured by EIA. Complement activation was evaluated by quantification of C3bc and the terminal complement complex by EIA. RESULTS: Neutrophil expression of L-selectin increased after apheresis (p = 0.02), and the production of oxygen radicals was reduced (p = 0.01). This effect was possibly a result of priming. Complement was not activated. There were no significant differences in neutrophil function after apheresis with any of the three apheresis systems. CONCLUSIONS: Neutrophil function was altered after apheresis, although to a very small extent, and contact between neutrophils and the foreign surface in the apheresis systems is found to be a biotolerant procedure.     

转基因山羊羊乳中重组人乳铁蛋白性质的检定与研究

目的:检定与研究转基因山羊 CLF123-1羊乳中重组人乳铁蛋白(rhLF)及其分子特性。方法:利用 SDS～PAGE,West～ern-blotting 及 Edman 降解法测定分析转基因山羊羊乳中 rhLF 的相对分子质量、免疫学特性和 N-末端15个氨基酸残基;利用紫外分光光度法比较 rhLF、天然人乳铁蛋白(hLF)与 Fe~(3 )离子结合的动力学过程,测定 Fe~(3 )与 LF 发生结合反应达到平衡状态后在279 nm 和465 nm 的吸收度值,Fe~(3 )与 LF 的物质的量比值分别为:0:1,0.5:1,1:1,2:1,4:1。结果:rhLF 的相对分子质量为(8.06±0.15)万(2次实验,6条电泳谱带计算结果);Western-blotting 结果显示 rhLF 可特异地与兔抗人 LF 抗体发生特异性结合;rhLF 的 N-末端1～15个氨基酸残基的序列为 G R R R R S V Q W X T V S Q P;rhLF 和 hLF 与 Fe~(3 )结合特性与趋势几乎完全一致。结论:本文所研究的转基因山羊羊乳中的乳铁蛋白是与人乳铁蛋白分子特性一致的 rhLF。     

牛乳铁蛋白的质量控制

目的:研究牛乳铁蛋白(BLF)的质量控制.方法:采用高效毛细管电泳法(HPCE)对牛乳铁蛋白的含量进行测定.结果:牛乳铁蛋白的浓度在100.0～600.0 μg·ml-1与其峰面积成良好的线形关系,r=0.999 5.结论:该法操作简便,精密度好,结果准确可靠,可作为牛乳铁蛋白质量标准的重要指标.     

Preparation,optimization and cellular uptake study of tanshinone I nanoemulsion modified with lactoferrin for brain drug delivery

Tanshinone I (TSI) is one of the bioactive compound obtained from the root of Salvia miltiorrhiza which is a well-known traditional Chinese medicine (TCM) used for the treatment of various diseases. Although TSI possesses several pharmacological effects, it has poor water solubility, blood–brain barrier (BBB) permeability and brain bioavailability. Therefore, in the present study, we developed TSI nanoemulsion (TSI-NE) modified with a brain targeting ligand (Lactoferrin (Lf)) to improve the BBB permeability. Pseudo-ternary phase diagrams were used to optimize the formulation. The optimal TSI-NE and TSI-Lf-NE were prepared and characterized. Finally, the uptake of TSI-Lf-NE by mouse brain microvascular endothelial cell line (bEnd.3 cells) was assessed using Coumarin-6 as a fluorescent probe. The results of the study showed that the stable optimal formulation of O/W nanoemulsion was successfully developed and modified with Lf. The cellular uptake study has shown that the fluorescence intensity (FI) increased with time over the incubation period. The FI at all time intervals increased in the following order: Coumarin-6-Solution＜Coumarin-6-NE＜Coumarin-6-Lf-NE. The results suggest that the BBB permeability of Coumarin-6-Lf-NE was better than those of Coumarin-6-NE and Coumarin-6 solution. Lf modified nanoemulsion has great potential for improving the brain delivery of TSI.     

Deficiency of the specific granule proteins,R‐binder/transcobalamin I and lactoferrin,in plasma and saliva: A new disorder

The mechanisms of hereditary deficiency of R binder, which originates in neutrophils and exocrine gland epithelium, are unknown and may be multiple. This led us to examine if defective R binder synthesis also involves proteins that colocalize with it in neutrophil‐specific granules and exocrine epithelial cells and may be under common regulatory control. Stored plasma and saliva samples from five unrelated R binder–deficient patients and control subjects were assayed for R binder, lactoferrin, cationic antimicrobial protein‐18, neutrophil gelatinase–associated lipocalin, gelatinase, lysozyme, and myeloperoxidase. One patient, patient A, had lactoferrin levels below the limits of detection in both plasma and saliva in addition to his R binder deficiency. Although his deficiency involved lactoferrin as well, he had no history of predisposition to infection. PCR amplification of his R binder gene promoter region and the beginning of the first exon revealed no DNA abnormalities. His son and the son of his equally deficient brother, both presumptive heterozygotes, had mild deficiency of both R binder and lactoferrin. The results show that R binder deficiency exists in at least two forms. One, presumably the less common of the two forms, is the new hereditary entity described here, which is characterized by deficiency of more than one specific granule protein in both plasma and saliva. Despite this more widely distributed absence of the proteins than is found in congenital specific granule deficiency, infection posed no clinical problem in the affected patient. © 2001 Wiley‐Liss, Inc.     

Expression of lactoferrin on human granulocytes: analysis with polyclonal and monoclonal antibodies

Lactoferrin (LF), an iron-binding protein present in specific granules of neutrophils, is expressed on membrane after granulocyte activation. It may represent a target for anti-neutrophil cytoplasmic antibodies (ANCA) in patients affected by some immunomediated diseases. We recently produced two MoAbs, AGM 2.29 and AGM 10.14, that recognize two spatially distant epitopes of human LF. In this study we perform a cytometric analysis in order to evaluate the expression of LF on the surface of granulocytes obtained from freshly drawn blood or after purification, in both the presence and absence of stimuli. Our results demonstrate that LF is not constitutively expressed on membrane of circulating neutrophils. After priming with phorbol myristate acetate (PMA) or tumour necrosis factor-alpha (TNF-α), an increased mean fluorescence intensity (MFI) was obtained on neutrophils stained with polyclonal anti-LF antibodies and with AGM 2.29. The kinetics of LF expression during activation demonstrated a progressive increase in MFI within 45 min. No increase in MFI was documented when primed granulocytes were stained with MoAb AGM 10.14, thus indicating that the epitope recognized by AGM 10.14 is not exposed at the cell surface. Following membrane permeabilization, performed in order to analyse the binding of anti-LF MoAbs to cytoplasmic LF, a marked increase in MFI was obtained by staining granulocytes with both anti-LF MoAbs. Indirect immunofluorescence (IIF) analysis confirmed that AGM 2.29 and AGM 10.14 reacted with human granulocytes, showing a cytoplasmic pattern on formalin–acetone-fixed neutrophils and a perinuclear one on ethanol-fixed cells.     

Lactoferrin prevents invasion and inflammatory response following E. coli strain LF82 infection in experimental model of Crohn's disease

BackgroundCrohn's disease is a multifactorial disease in which an aberrant immune response to commensal intestinal microbiota leads to chronic inflammation. The small intestine of patients with Crohn's disease is colonized by a group of adherent-invasive Escherichia coli strongly able to adhere and invade intestinal epithelial cells lactoferrin is an iron-binding glycoprotein known to have anti-bacterial and anti-inflammatory activities.AimsWe explore the ability of bovine lactoferrin to modulate the interactions between the adherent-invasive E. coli strain LF82 and intestinal epithelial cells as well as the inflammatory response.MethodsBacterial adhesion and invasion assays were used to assess the antimicrobial activity of lactoferrin. Electron microscopy was used to characterize bacteria–cell interactions. The mRNA expression of pro-inflammatory cytokines was measured both in cultured cells and in biopsies taken from intestine of patients affected by Crohn's disease.ResultsLactoferrin inhibited bacterial invasion through minimally affecting adhesion. This divergence was due to a mannose-dependent lactoferrin binding to the bacterial type 1 pili and consequent bacterial aggregation on the intestinal epithelial cell surface. Expression of pro-inflammatory cytokines, such as TNF-alpha, IL-8, and IL-6, was markedly inhibited by lactoferrin both in cultured and Crohn-derived intestinal cells.ConclusionsBovine lactoferrin might function via an antibacterial and/or anti-inflammatory mechanism in the treatment of Crohn's disease.