Analysis of human neutrophil granule protein composition in chronic myeloid leukaemia by immuno-electron microscopy

Summary In this study immuno-electron microscopy was used to assay, semi-quantitatively, the granule contents of elastase, lactoferrin, lysozyme and myeloperoxidase in human peripheral blood neutrophils from 13 chronic myeloid leukaemia patients in the chronic phase of the disease and from normal non-smoking donors. The fixation conditions that adequately preserved the antibody binding capacities of these antigens and reasonably preserved the ultrastructure of the neutrophils were selected by light-microscopic immunoperoxidase cytochemistry on cytospin smears. Immunogold cytochemistry on LR White resin sections localised elastase and myeloperoxidase to the primary granules, lactoferrin to the secondary granules and lysozyme to both types of granule. When applicable, peroxidase cytochemistry was combined with immunogold staining making it easier to distinguish the primary from the secondary granules. A comparison of the immunolabelling density values obtained for the leukaemic and normal states revealed no significant abnormalities in the immunoreactivity patterns for any of these neutrophil granule antigens in the leukaemic patients. All 13 patients gave normal immunostaining reactivities for these neutrophil granule proteins. Consequently the distribution patterns of these proteins, as shown in this study, cannot be used as indices in distinguishing chronic myeloid leukaemic neutrophils from normal neutrophils.     

人乳铁蛋白在转基因马铃薯块茎中的表达

人乳铁蛋白(human lactoferrin,hLF)是人体非特异性免疫系统的重要成员之一,具有抗细菌、真菌和抗病毒活性及其他多种功能.报道将hLF基因的cDNA与马铃薯(Solarium tuberosum L.)块茎专一性表达patain基因启动子融合后通过农杆菌介导导入马铃薯,PCR检测证实获得了多个转基因株系,RT-PCR阳性结果说明hLF mRNA在马铃薯植株中得到了表达.同时,经过ELISA及Western blot检测证实,转基因马铃薯表达了hLF并具有人乳铁蛋白的活性.     

Emulsions Based on the Interactions Between Lactoferrin and Chitosans

In this work, purification of lactoferrin from whey was performed with high recovery rate. Lactoferrin was then exploited in the preparation of food emulsions. Two tertiary emulsions, formed by olive oil, lecithin, chitosan, and lactoferrin, were compared: both the emulsions showed similar turbidity and stability. In the secondary emulsion formed by oil/lecithin/chitosan, the pH was increased to 9 before addition of lactoferrin. Then, lactoferrin was added, and the pH was stabilized above pH 9. Lactoferrin was found in amounts of 1 to 2.5 mg/ml in the multiple experiments. A fraction of the added lactoferrin was also present in a milky layer above the emulsion layer. This was, to our knowledge, the first study of emulsions made exploiting the interactions between lactoferrin and chitosan. It was noted that chitosan droplets remained soluble, although the hydrocolloid solubility occurs at pH lower than 5.9. These results showed the feasibility of manufacturing lactoferrin-based emulsions as functional foods.     

粪乳铁蛋白和C 反应蛋白对溃疡性结肠炎的评价

目的:研究粪乳铁蛋白(1actoferrin,LF)和C反应蛋白(C-reactive protein,CRP)对溃疡性结肠炎的严重程度、活动性和对治疗反应的评价作用。方法:选取我院2009-2010年43例特发性溃疡性结肠炎患者,同时进行对照平行研究。对患者进行临床评估、内镜和病理检查,了解疾病活动性、严重程度,并检测LF、CRP在治疗前后的水平。结果:与对照组相比较,病例组生物学标志都升高(P=0.000),特发性溃疡性结肠炎严重组粪LF和CRP更高,治疗后,生物学指标下降到与Mayo分数齐平,ROC曲线下面积分别为0.953和0.721,其灵敏度和特异性分别为:LF(90.7%,100%)和CRP(44.2%,100%)。结论:在对特发性溃疡性结肠炎的活动性、严重程度和对治疗反应的评价中,粪LF和CRP是非常有用的生物学指标。     

pH-induced structural changes of apo-lactoferrin and implications for its function: a molecular dynamics simulation study

The minor protein in milk, lactoferrin (Lf), is known for a variety of biological functions, and has been investigated as a protective encapsulant for probiotic bacteria in health-promoting food products. Lf is likely to be exposed to extreme pH conditions which are known to have disruptive influences on its functionality. The molecular mechanisms underlying these pH-dependent changes are not well-understood. To explore the potential of Lf as an encapsulant, molecular dynamics (MD) simulations were applied to study its conformational changes under extreme acidic (pH 1.0) or basic (pH 14.0) conditions, relative to neutral pH. Simulations indicate that the structure of apo-Lf is relatively stable at neutral pH, while acidic and basic pH result in substantially greater flexibility, partly induced by the loss of contacts between the N- and C-terminal lobes, causing them to undergo extensive relative bending and twisting motions. Basic pH causes greater structural disruption compared to acidic exposure. The latter has greater influence on the N-terminus, with increased fluctuations and disruptions of inter-residue contacts compared to those at neutral pH; while basic pH was found to more prominently disrupt contacts at the C-terminus. These results help elucidate possible functional consequences on Lf of exposure to extreme pH conditions.     

Lactoferrin Interacts with Nucleotides

Lactoferrin (LF) is an iron‐binding glycoprotein found predominantly in milk and in granulocytes. LF is extremely polyfunctional protein some biological functions of which are determined by its capacity to bind iron, but many other functions are iron‐independent. In this article we show for the first time that LF interacts with a number of various mononucleotides.     

A structural comparison of human serum transferrin and human lactoferrin

The transferrins are a family of proteins that bind free iron in the blood and bodily fluids. Serum transferrins function to deliver iron to cells via a receptor-mediated endocytotic process as well as to remove toxic free iron from the blood and to provide an anti-bacterial, low-iron environment. Lactoferrins (found in bodily secretions such as milk) are only known to have an anti-bacterial function, via their ability to tightly bind free iron even at low pH, and have no known transport function. Though these proteins keep the level of free iron low, pathogenic bacteria are able to thrive by obtaining iron from their host via expression of outer membrane proteins that can bind to and remove iron from host proteins, including both serum transferrin and lactoferrin. Furthermore, even though human serum transferrin and lactoferrin are quite similar in sequence and structure, and coordinate iron in the same manner, they differ in their affinities for iron as well as their receptor binding properties: the human transferrin receptor only binds serum transferrin, and two distinct bacterial transport systems are used to capture iron from serum transferrin and lactoferrin. Comparison of the recently solved crystal structure of iron-free human serum transferrin to that of human lactoferrin provides insight into these differences.     

Proadipogenic effects of lactoferrin in human subcutaneous and visceral preadipocytes

Lactoferrin has been associated with insulin sensitivity in vivo and in vitro studies. We aimed to test the effects of lactoferrin on human subcutaneous and visceral preadipocytes. Human subcutaneous and visceral preadipocytes were cultured with increasing lactoferrin (hLf, 0.1, 1, 10 μM) under differentiation conditions. The effects of lactoferrin on adipogenesis were studied through the expression of different adipogenic and inflammatory markers, AMPK activation and Retinoblastoma 1 (RB1) activity. The response to insulin was evaluated through ^Ser473AKT phosphorylation. In both subcutaneous and visceral preadipocytes, lactoferrin (1 and 10 μM) increased adipogenic gene expressions and protein levels (fatty acid synthase, PPARγ, FABP4, ADIPOQ, ACC and STAMP2) and decreased inflammatory markers (IL8, IL6 and MCP1) dose-dependently in parallel to increased insulin-induced ^Ser473AKT phosphorylation. In addition to these adipogenic effects, lactoferrin decreased significantly AMPK activity (reducing ^pThr172AMPK and ^pSer79ACC) and RB1 activity (increasing the ^pser807/811RB1/RB1 ratio). In conclusion, these results suggest that lactoferrin promotes adipogenesis in human adipocytes by enhancing insulin signaling and inhibiting RB1 and AMPK activities.     

Twenty-five years of research on bovine lactoferrin applications

Lactoferrin (LF) was identified as a milk protein in 1960. Large-scale manufacturing of bovine LF (bLF) was established more than 20 years ago. Using this commercially available material, research for bLF applications has advanced from basic studies to clinical studies, and bLF has been applied to commercial food products for the last 25 years. During this period, it was found that LF is digested by gastric pepsin to generate a multi-potent peptide, lactoferricin. It was also demonstrated that oral administration of bLF augments host protection against infections via antimicrobial action and immunomodulation of the host. In addition, researchers have demonstrated that oral administration of bLF prevents cancer development. In this review, we look back on 25 years of bLF research and development.     

A structural framework for understanding the multifunctional character of lactoferrin

Lactoferrin (Lf) is widely distributed, in mammalian milks, other secretory fluids and white blood cells, and its biology is complex. The three-dimensional structure of this important protein was determined in 1987, giving the first atomic view of any member of the transferrin family. This review examines how structural knowledge has contributed to our understanding of Lf function, and what we have yet to understand. The internal structure of Lf is highly conserved, and is dedicated to binding iron, which is sequestered in two almost identical sites, one in each lobe of the molecule. The processes of iron binding and release, and the accompanying conformational changes, are well understood. Some functional properties of Lf derive from this property, both through iron scavenging, and because the structure and dynamics of Lf are altered by its iron status. On the other hand, the external structure (its molecular surface) is much more variable between different Lfs, making it more difficult to identify functionally important sites. One key feature is clear - the cationic N-terminus and associated lactoferricin domain on the N-lobe of Lf. Recent work shows that this region, in addition to its role in antibacterial activity and probable role in DNA binding, is also involved in complex formation with other proteins. Other parts of the surface are more variable and may result in functional differences between the Lfs of different species. Finally, it may be time to re-examine the importance of glycosylation, given the growing evidence that many pathogens depend on binding to glycans for pathogenesis.