Veterinarni Medicina, 53, 2008 (9): 457–468 Review Article Lactoferrin: a review L. Adlerova1, A. Bartoskova1, M. Faldyna1,2 1University of Veterinary and Pharmaceutical Sciences, Brno, Czech Republic 2Veterinary Research Institute, Brno, Czech Republic ABSTRACT: This review discusses the biological properties of the glycoprotein lactoferrin. Lactoferrin has been identified in secretions from exocrine glands and in specific granules of neutrophils. After degranulation, neu- trophils become the main source of lactoferrin in blood plasma. Lactoferrin possesses various biological func- tions, including roles in iron metabolism, cell proliferation and differentiation, and antibacterial, antiviral, and antiparasitic activity. Many of these functions do not appear to be connected with its iron binding ability. Of late, lactoferrin concentrations have been measured mostly in humans but also in some other species. However, the relationship between its concentration and physiological or pathological effects on body functions is not yet well characterised. Keywords: protein; glycoprotein; transferrin; milk; neutrophils; leukocytes; host defense; antimicrobial Contents 7.4. Antiviral activity 7.5. Antiparasitic activity 1. Introduction 7.6. Lactoferrin and host defense 2. Structure and properties of lactoferrin 7.7. Lactoferrin and tumor growth 3. Sources of lactoferrin in the organism 7.8. Lactoferrin and cell proliferation and 4. Regulation of lactoferrin synthesis differentiation 5. Lactoferrin receptors 7.9. Lactoferrin and bones 6. Lactoferrin metabolism 7.10. Enzymatic activity of ribonuclease A 7. Biological functions of lactoferrin 8. Lactoferrin in different species 7.1. Lactoferrin and iron metabolism 9. Conclusions 7.2. Antimicrobial activity 10. References 7.3. Antibacterial activity 1. Introduction was concurrently determined to be the main iron binding protein in human milk by three independ- Lactoferrin (formerly known as lactotransferrin) is ent laboratories (Groves, 1960; Johanson, 1960; a glycoprotein, and a member of a transferrin family, Montreuil et al., 1960). thus belonging to those proteins capable of binding and Subsequent research identified lactoferrin in transferring Fe3+ ions (Metz-Boutique et al., 1984). secretions from exocrine glands and in specific Lactoferrin was first isolated by Sorensen and granules of neutrophils. Neutrophils after degran- Sorensen from bovine milk in 1939. In 1960 it ulation were observed to be the main source of Supported by the Ministry of Agriculture of the Czech Republic (Grant No. MZe 0002716201) and by the Internal Grant Agency of the University of Veterinary and Pharmaceutical Sciences Brno (Grant No. 7/2006/FVL). 457 Review Article Veterinarni Medicina, 53, 2008 (9): 457–468 lactoferrin in blood plasma (Iyer and Lonnerdal, strong and can resist pH values of as low as 4, its 1993). saturation does not exceed 10% in total (Mazurier Due to the increase in its concentration during and Spik, 1980). There are three forms of lactofer- most inflammatory reactions and some viral in- rin according to its iron saturation: apolactoferrin fections, several authors classify lactoferrin as an (iron free), monoferric form (one ferric ion), and acute-phase protein (Kanyshkova et al., 2001). Its hololactoferrin (binds two Fe3+ ions). The tertiary concentration increases in all biological fluids, but structure in hololactoferrin and apolactoferrin is the highest levels have been detected in the nidus different (Jameson et al., 1998). of inflammation (Birgens, 1985). Four amino acid residues are most important for Thus, lactoferrin has a wide variety of biological iron binding (histidine, twice tyrosine, and aspartic functions, many of which do not appear to be con- acid), while an arginine chain is responsible for nected with its iron binding ability (Brock, 2002). binding the carbonate ion (Baker, 1994; Ward et al., 1996). Besides iron lactoferrin is capable of binding 2. Structure and properties of lactoferrin a large amount of other compounds and sub- stances such as lipopolysacharides, heparin, gly- Lactoferrin is a glycoprotein with a molecular cosaminoglycans, DNA, or other metal ions like weight of about 80 kDa, which shows high affinity Al3+ , Ga3+, Mn3+, Co3+, Cu2+, Zn2+ etc. However, for iron. The molecular structure and amino acid its affinity for these other ions is much lower. Apart 2– sequence of human lactoferrin were discovered in from CO3 , lactoferrin can bind a variety of other 1984. Lactoferrin was then classified as a member anions like oxalates, carboxylates, and others. In of the transferrin family, due to its 60% sequence this way it is possible for lactoferrin to affect the identity with serum transferrin (Metz-Boutique et metabolism and distribution of various substances al., 1984). (Baker, 1994). Three different isoforms of lactoferrin have been The ability to keep iron bound even at low pH isolated. Lactoferrin-α is the iron binding form, but is important, especially at sites of infection and has no ribonuclease activity. On the other hand inflammation where, due to the metabolic activ- lactoferrin-β and lactoferrin-γ demonstrate ribo- ity of bacteria, the pH may fall under 4.5. In such nuclease activity but they are not able to bind iron a situation lactoferrin also binds iron released (Furmanski et al., 1989). from transferrin, which prevents its further usage Lactoferrin is comprised of a single polypep- for bacterial proliferation (Valenti and Antonini, tide chain containing 703 amino acids folded into 2005). two globular lobes. These lobes, also called C – Lactoferrin has demonstrated remarkable re- (carboxy) and N – (amino) terminal regions, are sistance to proteolytic degradation by trypsin connected with a α-helix. Each lobe consists of and trypsin-like enzymes. The level of resistance two domains known as C1, C2, N1, and N2. The do- is proportional to the degree of iron saturation mains create one iron binding site on each lobe. (Brock et al., 1976; Brines and Brock, 1983; Iyer Lactoferrin molecules contain (according to the and Lonnerdal, 1993). species and protein) varying numbers of sites for potential glycosylation, mostly on the surface of the molecule. The most common sacharide is mannose; 3. Sources of lactoferrin in the organism around 3% are hexoses, and 1% hexosamines. The degree of glycosylation varies and determines the Lactoferrin expression can first be detected in rate of resistance to proteases or to very low pH. two- and four-cell embryos during embryonic de- Lactoferrin’s capability of binding iron is two velopment, then throughout the blastocyst stage times higher than that of transferrin, which can up to implantation. Lactoferrin cannot be detected serve in some cases as donor of Fe3+ ions for from the time of implantation until halfway through lactoferrin. Two ferric ions can be bound by one gestation. Later, it is found in neutrophils and epi- lactoferrin molecule. One carbonate ion is always thelial cells of forming reproductive and digestive bound by lactoferrin concurrently with each ferric systems (Ward et al., 1999). ion (Aisen and Liebman, 1972; Metz-Boutique et The predominant cell types involved in lactofer- al., 1984; Baker, 1994). Although this bond is very rin synthesis are of the myeloid series and secretory 458 Veterinarni Medicina, 53, 2008 (9): 457–468 Review Article epithelia (Baynes and Bezwoda, 1994). In adults, the blood increases during infection, inflammation higher levels of lactoferrin are present in milk and (Birgens, 1985), excessive intake of iron, or tumor colostrum (Masson and Heremans, 1971; Brock, growth (Levay and Viljoen, 1995). 1980). It is also found in most mucosal secretions such as uterine fluid, vaginal secretion, seminal fluid, saliva, bile, pancreatic juice, small intestine 4. Regulation of lactoferrin synthesis secretions, nasal secretion, and tears (Masson et al., 1966; Baker, 1994; Levay and Viljoen, 1995; The regulation of lactoferrin synthesis depends on Lonnerdal and Iyer, 1995; Kikuchi et al., 2003; Baker the type of cells producing this protein. The amount and Baker, 2005). of lactoferrin synthesized in the mammary gland The production of lactoferrin by human kidneys is controlled by prolactin (Green and Pastewka, was described by Abrink et al. (2000). Lactoferrin 1978), whereas its production in reproductive tis- is expressed and secreted throughout the collecting sues is determined by estrogens (Pentecost and tubules, and in the distal part of the tubules it may Teng, 1987; Walmer et al., 1992; Teng et al., 2002). be reabsorbed. These results show that the kidney The synthesis of lactoferrin in endometrium is in- produces lactoferrin in a highly ordered manner fluenced by not only estrogens but also epidermal and that only a minor fraction of this protein is growth factor (Nelson et al., 1991). Exocrine glands secreted into the urine. Therefore, lactoferrin is produce and secrete lactoferrin in a continuous thought to have important functions in both the manner. In neutrophils, lactoferrin is synthesized immune defense of the urinary tract and in general during their differentiation (when promyelocytes iron metabolism. develop into myelocytes) and is afterwards stored Neutrophils are an important source of lacto- in specific granules. Mature neutrophils cease to ferrin in adults. Indeed, most plasma lactoferrin produce lactoferrin (Masson et al., 1969). originates from neutrophils (Iyer and Lonnerdal, Lactoferrin levels might vary with gender and 1993). Lactoferrin is predominantly