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HomemieyecareLactoferrin

Lactoferrin

Lactoferrin is a ubiquitous mucous membrane protein that forms part of the innate immune system. It has reported ocular activity as an anti-inflammatory, antimicrobial and wound healing promoter.

Lactoferrin is a natural component of milk and is a safe nutraceutical that, in the U.S., is already in phase 2 clinical trials for treatment of severe sepsis and diabetic skin ulcers. Currently oral lactoferrin is available in Australia as an unscheduled medicine. Topical ocular formulations have been patented but are yet to be manufactured. Lactoferrin presents as a future novel treatment for a range of conditions commonly encountered by optometrists.

When Dr. Stephen DeFelice coined the term “nutraceutical” over 20 years ago, he defined it as “any substance that is in a food or a part of a food and provides medical or health benefits, including the prevention and treatment of disease”. Lactoferrin is a key nutraceutical commonly purified from milk where it is at an average concentration of 2 mg/mL. High levels of this protein are present in mucous membrane secretions and the secondary granules of neutrophils. In the tear film lactoferrin is the second most abundant protein, after lysozyme, and has functions ranging from antimicrobial activity and immune modulation to wound healing.

The Australian market for nutraceutical products is estimated to be worth AUD$950 million per annum and includes minerals, vitamins, proteins, herbs and foods that are of assistance, beyond their nutritional value, in the management of pathologies. These are regulated by the Therapeutics Goods Act 1989 under ‘Complementary Medicines’. The Therapeutic Goods Administration designates access to these compounds as either unscheduled, over the counter (OTC) or prescription medicines, although generally they are the former and available to consumers for self-medication from retail outlets.

In the tear film lactoferrin is the second most abundant protein, after lysozyme, and has functions ranging from antimicrobial activity and immune modulation to wound healing

In the tear film lactoferrin accounts for a quarter of the total protein and is mostly secreted by the acinar cells of the lacrimal gland to a concentration of approximately 2mg/mL. Lower tear film lactoferrin levels are reported in dry eye, herpes simplex keratitis, ocular allergic conditions and systemic infections such as HIV and Hepatitis C. The possibility of a causal link has led researchers to explore lactoferrin’s potential as an ocular therapeutic agent.

Reduced lactoferrin

The two major groups of aqueous deficient dry eye, Sjögren and non-Sjögren, are associated with a reduction in lactoferrin levels. Sjögren’s syndrome dry eye is an autoimmune disorder that severely reduces the lacrimal gland’s capacity to secrete tears by destroying its exocrine cells. As these cells are the primary source of lactoferrin in the tear film, a reduction in their population lowers the lactoferrin concentration.

Non-Sjögren dry eye is attributed to reduced neural stimulation of the lacrimal gland. In this group, the capacity to produce lactoferrin and tears remains unchanged but the innervation is diminished.1 Two small clinical trials found that by attempting to correct this deficiency in severe dry eye patients, with 270 mg/day of lactoferrin, there was a lessening of symptoms, an increase in tear film stability and reduced staining.2

Contact Lenses and Lactoferrin

Interactions between tear film lactoferrin and contact lenses have also been explored. No change in tear film lactoferrin levels are reported across contact lens materials or wear modalities, although lactoferrin is deposited in trace levels on all common contact lens materials. A possible benefit of these deposits has been uncovered by investigators at the Brien Holden Vision Institute, previously the Institute for Eye Research (IER)), who found lactoferrin-coated lenses host fewer viable bacteria.3

The most notable contact lens related condition associated with a change in lactoferrin levels is contact lens papillary conjunctivitis (CLPC). Current theory holds that mechanical and antigenic stimuli are required for the development of CLPC and its progression to the more severe giant capillary conjunctivitis (GPC). The mechanical interaction is between the lens and the conjunctiva while deposits on the lens contain antigens that raise production of immunoglobulins and activate the complement system. As a decrease in tear film lactoferrin is found in both CLPC and GPC, it has been proposed that the loss of lactoferrin’s inhibitory effect on the pro-inflammatory mediator C3a within the complement cascade may be a factor in the pathogenesis of these conditions.4 This indicates supplementation with lactoferrin could be of assistance, if administered prophylactically, to those at risk of these adverse events.

The Immune Response

In addition to suppression of the complement pathway, lactoferrin also regulates other aspects of the immune response. Receptors for lactoferrin are found on myeloblasts, monocytes, macrophages, lymphocytes and epithelial cells. These interactions are complex and not fully elucidated. Generally speaking, lactoferrin is released in inflammation and acts to curtail excessive inflammatory responses both directly, through antigen binding and iron uptake, and indirectly, by stimulating negative feedback signalling pathways within other cells. The capacity for exogenous lactoferrin to act as a local anti-inflammatory agent has been demonstrated in-vivo for gastrointestinal epithelia5 and cutaneous tissue6.

Lactoferrin’s modes of action include iron sequestration, membrane disruption and digestion of bacterial proteins. The first of these to be identified was iron scavenging. Because iron is an essential co-factor, required for the growth of many invasive bacteria, when lactoferrin binds free iron in tears, it interrupts pathogenic bacterial proliferation. Lactoferrin is also able to disrupt bacterial cell membranes. The interaction of the positively charged amino acids at the terminus of lactoferrin and the negatively charged bacterial membrane increases their permeability. This may kill the bacteria or at least increase their susceptibility to other antimicrobials in the tear film.

Leading on from this, it has been demonstrated that lactoferrin can be applied as a synergistic agent, increasing the efficacy of other antibiotics. Lactoferrin’s antimicrobial action also extends to interactions with bacterial virulence factors. For example, lactoferrin can digest the adhesion proteins secreted by enteropathogenic E. coli, preventing colonisation. Despite these multiple actions of lactoferrin against bacteria, there are still species able to express protective mechanisms. Fortunately, many of the bacteria associated with adverse ocular events including Escherichia coli, Haemophilus influenzae, Bacillus subtilis, Streptococcus spp. and Pseudomonas spp. are affected by lactoferrin.

Bacterial biofilms are a particular problem in contact lens wear, as these colonies are more resistant to cleaning systems. Biofilms are tight clusters of bacteria that adhere to a surface and produce their own protective matrix. Lactoferrin is able to inhibit biofilm formation in Pseudomonas aeruginosa by depriving it of iron. When P. aeruginosa is unable to bind iron it remains in a motile state that is unable to attach and form colonies. However, once a biofilm is formed, the bacteria become 100 fold more resistant to lactoferrin.7 In the case of Staphylococcus epidermis, lactoferrin is able to increase the susceptibility of the biofilm colony to other antimicrobials.8 Thus, lactoferrin in the tear film may reduce biofilm formation and increase the biofilm’s permeability to other agents.

Protection Against Viruses

The protective role of lactoferrin is not limited to an anti-bacterial role but also extends to viruses. Those species with demonstrated lowered invasiveness in the presence of lactoferrin include rotavirus, enterovirus and adenovirus. This is largely attributed to blocking of the viral binding sites on potential host cells. To a lesser extent, lactoferrin also augments the immune response against viral infection.

A recently uncovered property of lactoferrin at the Brien Holden Vision Institute is the promotion of corneal wound repair. Increased epithelial healing rates have been observed for alkali burns treated with lactoferrin. This is associated with an up-regulation of growth factor, PDGF, and cytokine, IL-6, that stimulate cells to migrate more rapidly over the site of an injury. In the hope of developing an agent that reduces the ocular morbidity associated with corneal lesions, this continues to be an exciting area of research at the institute.9

Ben Ashby, BOptom (Hons), is a PhD candidate at the Brien Holden Vision Institute and School of Optometry and Vision Science, University of NSW.
Dr Qian Garrett, BSc PhD, is Manager of Biological Sciences and Senior Research Scientist at the Brien Holden Vision Institute and a Visiting Fellow at the School of Optometry and Vision Science, University of NSW.
Professor Mark Willcox, BSc PhD, is Chief Scientific Officer at the Brien Holden Vision Institute, Professor of the School of Optometry and Vision Science, University of NSW and Executive Director of Science and Core Capabilities at the Vision Cooperative Research Centre.

References

1. Ohashi, Y., Ishida, R., Kojima, T., Goto, E., Matsumoto, Y., Watanabe, K., Ishida, N., Nakata, K., Takeuchi and T., Tsubota, K., Abnormal protein profiles in tears with dry eye syndrome, American Journal of Ophthalmology, 2003. 136(2): p. 291-299.
2. Yamamoto, Y., Dogru, M., Matsumoto, Y., Saeki, M., Goto, E., Ishioka, M., and Tsubota, K., Tear function and ocular surface alterations with lactoferrin treatment in severe dry eyes, Invest. Ophthalmol. Vis. Sci., 2005. 46(5): p. 2046-.
3. Williams, T.J., Schneider, R.P. and Willcox, M.D.P., The effect of protein-coated contact lenses on the adhesion and viability of gram negative bacteria, Current Eye Research, 2003. 27(4): p. 227-235.
4. Ballow, M., Donshik, P.C., Rapacz, P., Samartino, L, Tear lactoferrin levels in patients with external inflammatory ocular disease, Invest Ophthalmol Vis Sci, 1987. 28(3): p. 543-545.
5. Togawa J., Nagase H., Tanaka K., Inamori M., Nakajima A., Ueno, N., Saito, T., and Sekihara, H.,) Oral administration of lactoferrin reduces colitis in rats via modulation of the immune system and correction of cytokine imbalance, J. Gastroenterol. Hepatol, 2002 17(12): p. 1291-1298
6. Kimber, I., Cumberbatch, M., Dearman, R. J., Headon, D. R., Bhushan, M., and Griffiths, C. E., Lactoferrin: influences on Langerhans cells, epidermal cytokines, and cutaneous inflammation, Biochem. Cell Biol., 2002 80(1):p. 103-107
7. Singh, P.K., Parsek, M.R., Greenberg, E.P. and Welsh, M.J., A component of innate immunity prevents bacterial biofilm development, Nature, 2002. 417(6888): p. 552-555.
8. Leitch, E.C. and Willcox, M.D.P., Lactoferrin increases the susceptibility of S. epidermidis biofilms to lysozyme and vancomycin, Current Eye Research, 1999. 19(1): p. 12-19.
9. Pattamatta, U., Willcox, M.D.P., Stapleton, F., Cole, N. and Garrett, Q., Bovine lactoferrin stimulates human corneal epithelial alkali wound healing in vitro, Invest Ophthalmol Vis Sci, 2008. 5: p. 1636-1643.

“The protective role of lactoferrin is not limited to an anti-bacterial role but also extends to viruses”