European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎

Contents lists available at ScienceDirect

European Journal of Pharmacology

journal homepage: www.elsevier.com/locate/ejphar

Full length article

Free amino acids: an innovative treatment for ocular surface disease

Dario Rusciano a,n, Anna Maria Roszkowska b, Caterina Gagliano c, Salvatore Pezzino d
a
Sooft Italia SpA, Scientific Department, Via Salvatore Quasimodo 136, 00144 Rome, Italy
b
Cornea and Refractive Surgery Section, Ophthalmology Unit, University Hospital of Messina, Italy
c
Ophthalmology Department, University of Catania, Catania, Italy
d
Sooft laboratories, Viale Andrea Doria 21, 95125 Catania, Italy

art ic l e i nf o a b s t r a c t

Article history: Amino acids are the basic constituents of living organisms, and have both a structural and an active
Received 18 December 2015 dynamic role in tissue and cell physiology. Human tears contain 23 amino acids, the relative proportion
Received in revised form of which may change with the different physiological states of the eye surface. In this review, we present
1 April 2016
a collection of data from the published literature that indicate an active role of amino acids in the
Accepted 14 April 2016
maintenance of eye surface homeostasis. Moreover, another series of published clinical data indicate that
supplementation of amino acids, either as food supplements or as a topical treatment in enriched eye
Keywords: drops, is beneficial to the eye surface, and may improve its healing in cases of eye surface disease due to
Dry eye different causes.
Aminoacids
& 2016 Elsevier B.V. All rights reserved.
Proline
Lysine
Leucine
Glycine
Taurine

1. Introduction
Dry Eye Disease (DED) was defined in 2007 by the Dry Eye
Workshop (DEWS) as ‘a multifactorial disease of the tear film and
the ocular surface that results in symptoms of discomfort, visual
disturbance, and tear film instability with potential damage to the
ocular surface’. It is usually accompanied by increased osmolarity
of the tear film and inflammation of the ocular surface, and is
considered one of the most frequent ocular disorders leading to
visual impairment (Foulks, 2007). Current therapeutic approaches
are aimed at improving signs and symptoms of DED and are re-
lated to its etiology due to either increased tear film evaporation
because of a deficiency of the lipid layer, and/or decreased pro-
duction of the aqueous phase by the main and accessory lacrimal
glands.
Currently, therapies can be systemic and/or topical, and they
are focused on tear supplementation, retention or stimulation,
coupled to the control of the underlying inflammation (Foulks,
2007; Dogru et al., 2013).
Tear supplementation is obtained with topical lubricants,
which can be hypotonic or isotonic and differ in their content of
electrolytes, surfactants and viscosity agents. Sodium hyaluronate
is frequently used as a hydrating and mucomimetic agent since it
n
Corresponding author.
E-mail address: dario.rusciano@sooft.it (D. Rusciano).
increases the stability of the precorneal tear film and improves
ocular surface wettability and smoothness thanks to its water re-
tentive and viscoelastic properties (Foulks, 2007; Aragona et al.,
2002).
Dry eye treatment by tear stimulation can be achieved by orally
administered cholinergic drugs, such as pilocarpine that is used for
severe dry eyes in Sjogren's Syndrome. The improvement of ocular
surface epithelia results from the stimulation of muscarinic re-
ceptors and of the secretory function associated with the increased
number of muciparous goblet cells (Foulks, 2007; Aragona et al.,
2006).
As regards anti-inflammatory treatment, the efficacy of topical
corticosteroids has been reported (Foulks, 2007; Aragona et al.,
2013b, 2015). The topical use of cyclosporine was also effective,
but its use is still limited to the US and is under continuing in-
vestigation (Foulks, 2007; Aragona, 2014; Stonecipher et al., 2013).
Interestingly, oral tetracyclines have demonstrated therapeutic
properties in the management of DED. They have been shown to
reduce MMP expression, lipase production and collagenase activ-
ity, and have been successfully used to treat chronic blepharitis,
rosacea and Meibomian Gland Disease (Foulks, 2007; Hessen and
Akpek, 2014; Doughty, 2016).
The use of oral supplementation with essential fatty acids was
found to be effective as an additional therapy in DED management.
Such treatment has been clinically shown to significantly reduce
conjunctival inflammatory marker expression (Foulks, 2007; Ara-
gona et al., 2005; Brignole-Baudouin et al., 2011).

http://dx.doi.org/10.1016/j.ejphar.2016.04.029
0014-2999/& 2016 Elsevier B.V. All rights reserved.

Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
2 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎

Finally, supplementation with amino acids, topical or systemic,
is a relatively recent strategy that will be the subject of this mini
review.
2. Amino acids in tears
Amino acids are essential nutrients as they constitute the main
elements of which proteins and peptides are made, and thus play
an essential part in the regulation of the whole metabolism of
living organisms. Amino acids are naturally present in human
tears. Their nature has been investigated in two different studies,
both of which reported that their relative concentration differs
from what is found in serum (ChenZhuo et al., 2002; Nakatsukasa
et al., 2011) and that this concentration changes in patients
affected by ocular dryness (Nakatsukasa et al., 2011) (Fig. 1). The
presence of free amino acids in tears and their dynamic fluctua-
tions suggest that they may play some relevant physiological role
in this specific and special environment. For instance, amino acids
may have antioxidant properties: methionine and cysteine work
as antioxidants through reversible protein oxidation (Kantorow
et al., 2004; Njie-Mbye et al., 2013). In the musculoskeletal system,
amino acids are required to increase bone and muscle mass, and to
reinforce tendons and joints (Gibala, 2000). In the epidermis,
amino acids (more specifically glycine, proline and hydroxypro-
line) are required for the synthesis of tropocollagen by stromal
fibroblasts, and for the building of collagen fibers (Wu et al., 2011).
A healthy stroma is also the key to a healthy epithelium, and ef-
ficient wound healing depends on a proper cross-talk between
fibroblasts and epithelial cells, which can be implemented by the

Fig. 1. Concentration of each amino acid in basal and reflex tears (Nakatsukasa et al., 2011).

Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
presence of amino acids (MacKay and Miller, 2003). Essential
amino acids have been successfully used in several systemic dis-
orders proving their positive effect on healing processes. In me-
tabolic pathologies, the oral supplementation with amino acids
together with conventional therapy, improve muscular protein
metabolism and cell function maintenance, promoting protein
synthesis and energy production both in peripheral skeletal
muscles and in the myocardium (Pasini et al., 2008; Aquilani,
2004). The role of the systemic use of amino acids in cardiology
was demonstrated by an improvement of muscular metabolism
and function in the elderly and in chronic heart failure. Additional
evidence in elderly patients has shown an increase of exercise
capacity by improving circulatory function, muscle oxygen con-
sumption and aerobic production of energy (Scognamiglio et al.,
2008; Aquilani et al., 2008b).
Dietary supplementation of arginine can enhance wound
healing, regulate endocrine activity and potentiate immune ac-
tivity (Efron and Barbul, 2000), significantly lowering catheter-
related sepsis rates (Caparrós et al., 2001). Arginine improves re-
productive, cardiovascular, renal, gastrointestinal, liver (Efron and
Barbul, 2000) and pulmonary functions by influencing pulmonary
hypertension (Mehta et al., 1995; G. Wu et al., 2009). Supple-
mentation of glutamine may improve protein status and im-
munocompetence, enhance nutritional management, reduce the
number of infections and augment recovery of the seriously ill
while minimizing hospital stay (Jones et al., 1999; Lacey and
Wilmore, 1990; Houdijk et al., 1998; Bollhalder et al., 2013; At-
kinson and Worthley, 2003; Platell et al., 2000; De Bandt and
Cynober, 2006). Moreover, amino acids are necessary for the
functioning of organ systems such as the nervous system, in which
they may act directly as neurotransmitters or as neurotransmitter
precursors (Fuchs et al., 2005). Amino acid imbalance can be the
cause of neurological diseases, such as Parkinson's (Luong, 2012)
or Huntington's chorea (Oepen et al., 1982), and oral supple-
mentation of amino acids may help in fighting these metabolic
deficiencies, also at the retinal neuron level (Fernstrom and
Fernstrom, 2007). In patients with severe brain injury, the sup-
plementation of branched-chain amino acids enhances the re-
trieval of disability rating scale and improves recovery from a
vegetative or minimally conscious state (Aquilani et al., 2008a,
Fig. 2. Amino acids arginine and lysine have an important role in many ocular physiological processes and can improve the clinical condition of diabetic and IOP patients.
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
3
2005).
3. Amino acids and inflammation
Some amino acids can reduce inflammation. Glutamine reduces
IL-8 production in human intestinal epithelial cells by limiting
IkBα ubiquitination and disposal, thus inhibiting the over-ex-
pression of pro-inflammatory genes and promoting the down-
regulation of the pro-inflammatory nuclear factor kappa-B (NF-kB)
pathway (Hubert-Buron et al., 2006; Fillmann et al., 2007).
Glycine, histidine and cysteine administered to human cor-
onary arterial endothelial cells can each inhibit NF-kB activation,
IL-6 production and expression of the leukocyte adhesion mole-
cule CD62E (Hasegawa et al., 2012).
CD62E (Hasegawa et al., 2012). Moreover, cysteine and histi-
dine exhibited anti-inflammatory effects in a human monocytic
leukemia cell line (Hasegawa et al., 2011). Oral supplementation of
the branched-chain amino acids isoleucine, leucine and valine
reduced inflammation and soreness in healthy subjects, likely
decreasing cytokine inflammatory chemicals released by the im-
mune system (Holecek, 2010). Orally administered L-isoleucine,
DL-isoleucine and L-leucine exhibited anti-inflammatory activity
in many test models of inflammation, except formaldehyde-in-
duced inflammation (Saxena et al., 1984). Taurine may play an
important role in inflammation associated with oxidative stress;
the taurine haloamines, TauCl and TauBr have antimicrobial and
anti-inflammatory properties (Marcinkiewicz and Kontny, 2014).
Amino acids can also be the source of relevant secondary
physiological products. In cases of inflammation, arginine is de-
graded by intracellular nitric oxide synthase (iNOS) into citrulline
and nitric oxide (NO), which is then used to sustain the in-
flammation process. NO has important physiological functions on
the ocular surface: it is involved both in the process of wound
healing after excimer laser ablation and in the protection against
allergic inflammation (Kim et al., 2002). In fact, during the in-
flammatory process there is an increased need for arginine, which
is imported into the cells by means of the cationic amino acid
transporter (hCAT), so that there is enough arginine that can be
used for the generation of NO as a defense mechanism (Fig. 2).
4 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
Accordingly, it has been shown that hCAT transporters in ocular
tissues are up-regulated during inflammation (Jäger et al., 2009).
hCAT transporters are also required for the intracellular transport
of arginine and lysine that are used for the synthesis of beta de-
fensins in tears, which are essential components of the local innate
immune system and the first line of defense against eye infections
(Jäger et al., 2010a, 2010b).
4. Amino acids and surgery
All regenerative processes require new protein synthesis for
cell replication and new extracellular matrix (ECM) production.
After mechanical or surgical traumas, wound healing will be more
efficient when all the necessary components are readily available
(MacKay and Miller, 2003). This is also true after eye surgery. In
ophthalmology, amino acids are used mainly in corneal surgery
and surface disorders. Photorefractive keratectomy (PRK) and laser
in-situ keratomileusis (LASIK) are surgical procedures that perturb
the ocular surface homeostasis (Wilson, 2001) and can result in
dry eye syndrome (De Paiva et al., 2006; Huang et al., 2012). Ne-
jima and collaborators found that indeed both procedures de-
creased the epithelial barrier function, reduced tear secretion, and
deteriorated tear film stability (Nejima et al., 2005). The efficacy of
cysteine oral supplements has been evaluated in corneal wound
healing after PRK (Fig. 3): all the eyes of patients treated with
cysteine oral supplements, in a daily dose of 200 mg, showed
shorter times to re-epithelization compared to the eyes of the
control group (Meduri et al., 2009).
Cataract surgery may also disrupt ocular surface homeostasis
and alter Meibomian gland function, thus leading to the devel-
opment of dry eye signs and symptoms (Kohlhaas, 1998; Cho and
Kim, 2009; Han et al., 2014; Li et al., 2007). Clinical data have
shown (Torres Munoz et al., 2003) that oral supplementation with
a mixture of 13 different amino acids (Fig. 4) in patients under-
going cataract surgery favors the remodeling of the stroma, de-
tected as an increase of stromal keratocytes, and induces changes
in the distribution of stromal cells and the ECM suggestive of an
active healing process (Fig. 5).
The same oral supplement with 13 amino acids has also been
used in PRK patients. In a first study (Fig. 6(a)) the corneal epi-
thelium, removed for PRK purposes, was analyzed for cytokine
(EGF and TGFβ) and polyamine expression (Roszkowska, 2006,
personal communication). Results showed that in amino acid
treated patients vs. untreated controls there was a decreased EGF
expression, suggesting decreased proliferation of epithelial cells
and keratocytes (decreased risk of hyperplasic reaction); an
Fig. 3. Oral cysteine supplementation, in a daily dose of 200 mg, reduces mean corneal wound healing time in patients after PRK (Meduri et al., 2009).
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
increased expression (35%) of TGFβ, again suggesting decreased
keratocyte proliferation and increased motility of epithelial cells
(faster re-epithelization); and a dramatic increased expression
(450%) of polyamines, suggesting regulated differentiation and
proliferation. Consistent with these results, a second study (Fig. 6
(b)) showed that oral amino acid supplementation improved ker-
atocyte growth and repopulation, thus improving the healing rate
of PRK patients who had shown re-epithelization defects (Vinci-
guerra et al., 2002).
Finally, reduced levels of amino acids in the lens may lead to
decreased synthesis of lens proteins such as the antioxidant en-
zyme glutathione (GSH) (Wu, 2009; Reddy, 1990; Berthoud and
Beyer, 2009). The effect of amino acids on mammalian eye lenses
against oxidative stress was investigated (Rathore and Gupta,
2010), and the results indicated a significant protective role of
selected amino acids in the H2O2 induced cataract model in vitro,
with a prominent role especially for arginine and proline.
5. Pre-clinical and clinical evidence
Given these results, topical eye drop formulations containing
amino acids with or without hydrating agents (such as hyaluronic
acid or hydroxy-propyl-methyl-cellulose) have been designed and
are currently available on the market. Four amino acids have been
chosen in the topical formulation: proline, lysine, leucine and
glycine. Proline and glycine are the main components of type I
collagen (Panjwani and Harding, 1978), which is the most abun-
dant protein (70% of dry weight) in the corneal stroma (Polatnick
et al., 1957). Lysine, which is present in collagen as such or as
hydroxy-lysine, although not a prominent amino acid, is critical for
the arrangement of the triple helix of collagen fibrils (Fietzek and
Kühn, 1975) and in corneal collagen is also heavily glycosylated,
thus further contributing to the spatial arrangement of the fibrils
(Panjwani and Harding, 1978) and the transparency of the corneal
stroma. Glycine and proline are also important respectively in
desmosomes (Kapprell et al., 1985) and tight junctions, in which
ZO-1 proteins contain a proline rich domain that interacts with
cytoskeletal F-actin (Fanning et al., 1998). Leucine is highly re-
presented in lumican, a proteoglycan with keratin-sulfate side
chains, which is responsible for the regulation of the collagen as-
sembly into fibrils, and necessary for corneal transparency (Amjadi
et al., 2013; Chakravarti et al., 1998). Along a similar line of
thought, however, in the field of dentistry, it has been reported
that the supplementation in vitro of these same four amino acids
(proline, glycine, lysine and leucine) may enhance collagen and
fibronectin biosynthesis by human dermal fibroblasts (Favia et al.,
 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎ 5

Fig. 4. (a–b) Chemical composition of the mixture of 13 different amino acids administered orally to patients undergoing cataract surgery or PRK (Torres Munoz et al., 2003).

2008). The results were also confirmed in vivo, after topical ad- free conditions with a mixture of these four amino acids at the
ministration to the gums of human patients (Favia et al., 2008). A same concentration (0.01%) present in artificial tears, was analyzed
conjunctival epithelial cell line challenged in vitro under serum- at 24–48 and 72 h. Results showed no effects on cell survival

Fig. 5. Clinical data show that oral supplementation favors the remodeling of the stroma, detected as an increase of stromal keratocytes, and induces changes in the
distribution of stromal cells and the ECM suggestive of an active healing process (Torres Munoz et al., 2003).

Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
6 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
Fig. 6. Effect of oral supplementation with amino acids on the corneal epithelium of PRK patients in two studies (a: Ref. Roszkowska (2006), personal communication; b: Ref.
Vinciguerra et al. (2002)).
(apoptosis monitored by Hoechst 33342) or proliferation (Ki67
proliferation marker), and a normal amount of Goblet cells, how-
ever, there was a slight increase of Muc5 mRNA and no change of
Muc1 mRNA (Micera et al., 2008, personal communication). In a
different in vitro model system, a multilayered corneal epithelium
was reconstituted on transwell filters in a serum-free medium
(Meloni et al., 2011). After mechanical scarification of the epithe-
lium, the kinetics of re-epithelization was found to be improved
after topical addition of saline plus the four amino acids, in cor-
relation with a higher expression of MMP9, a metalloprotease that
is involved in cell migration (Meloni et al., 2011; Rusciano et al.,
2009, personal communication). Similarly, re-epithelization of the
rabbit cornea after trephination showed a faster recovery after
treatment with topical eye drops containing hyaluronic acid and
amino acids (Rusciano et al., 2009, personal communication).
Dysfunctional tear syndrome is accompanied by alterations of
ocular surface epithelia and tear film, such as increased production
of inflammatory cytokines and the remodeling matrix metallo-
protease MMP9 (Chotikavanich et al., 2009; Lam et al., 2009).
Treatment of rabbits’ eyes (in which an insufficiency of tear pro-
duction had been induced by ConA intraocular injections) with
saline alone, or saline plus amino acids (Leu, Gly, Pro, Lys) showed
a significant increase of tear production, and a decrease of MMP9
expression in the amino acid treatment group after 3 days of
treatment (Marrazzo G and Platania CBM, 2012, personal
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
communication), suggesting that an improved trophism of the
ocular surface may exert a positive effect on the whole lacrimal
system. Consistent with these results, a pilot clinical study on 20
patients affected by mild to moderate dry eye (Sacchetti et al.,
2012, personal communication), showed that treatment with -
saline eye drops enriched with amino acids vs. saline eye drops
alone resulted after 1 and 3 months of treatment in a better re-
sponse for some symptoms, such as burning, itching, dryness and
tearing, in the amino acid treated patients (Fig. 7(A)). Another
clinical study compared the response of patients affected by dys-
functional tear syndrome treated with an artificial tear containing
either hyaluronic acid alone or hyaluronic acid plus amino acids
(Leu, Gly, Pro, Lys). After 1 and 3 months of treatment, all patients
showed improvements of signs and symptoms. However, those
patients treated with eye drops containing amino acids had better
corneal staining and observation at the confocal microscope
showed less hyper-reflecting cells (a sign of metabolic damage) in
the epithelial corneal layer as compared to those receiving hya-
luronic acid alone; nerve tortuosity was also improving faster and
better in the presence of topical amino acids (Aragona et al.,
2013a). Artificial tears containing hyaluronic acid plus amino acids
(Leu, Gly, Pro, Lys) can also modulate the expression pattern of
inflammatory proteins in the tears of patients affected by dys-
functional tear syndrome (Gagliano, 2013, personal communica-
tion). Starting from the first month of treatment, tear film protein
 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎ 7

Fig. 7. (A) Treatment with saline eye drops enriched with amino acids improved the signs and symptoms of patients affected my mild dry eye better than treatment with
saline eye drops alone (Sacchetti et al., 2012, personal communication), and (B) resulted in a significant decrease of inflammatory cytokines in tears (Gagliano, 2013, personal
communication).

level expression of IL-1beta, IL6, TNF-α and IFN-γ was decreased
compared to those receiving hyaluronic acid alone (Fig. 7(B)).
6. Taurine
Taurine is an amino-sulfonic acid which, differently from all the
other amino acids, contains an amino group and a sulfonic acid
group instead of the classic carboxylic group (Fig. 8).
Taurine is involved in cardiovascular function, and the devel-
opment and function of skeletal muscle, retina, and central ner-
vous system (Huxtable, 1992). In the adult's eye, it is heavily re-
presented in the retina, vitreous, lens, cornea, iris and ciliary body
(ChenZhuo et al., 2002; Heinämäki et al., 1986) with numerous
protective bioactive functions (Ripps and Shen, 2012).
Experimental studies in different animal model systems have
shown a correlation between aging, taurine availability and dis-
ease occurrence (Militante and Lombardini, 2004; Yildirim et al.,
2007). In the rat, taurine levels were found to decrease with aging
in the retina, lens tissues and cornea (Baskin et al., 1977). Taurine
induced the increase of the number of rod photoreceptors in rat
retina cultures (Altshuler et al., 1993) and it appears to act on
retinal progenitors via the glycine receptor α2 subunit (Young and
Cepko, 2004). Cats fed a taurine deficient diet developed central
retinal degeneration (Hayes et al., 1975) and exogenous taurine
administration may be helpful in preventing age related changes
in the retina (Militante and Lombardini, 2004). The effect of
taurine deficiency on photoreceptor survival was confirmed in
monkeys fed a taurine deprived diet (Imaki et al., 1987), in rats by
administering an inhibitor or competitive substrate of the taurine
transporter (Pasantes-Morales et al., 1983) and finally, achieved in
mice by knocking-out the taurine transporter (Rascher et al.,
2004). Retinal toxicity of the antiepileptic drug vigabatrin, widely
prescribed for the treatment of epilepsy (Wilby et al., 2005), is also
caused by taurine depletion (Jammoul et al., 2009). In glaucoma of
the dog, the taurine concentration seems to be markedly de-
creased in injured photoreceptors (Madl et al., 2005).
A 3-year follow-up study highlighted that treatment with
taurine, the calcium antagonist diltiazem, and vitamin E retards
the progressive visual field reduction in retinitis pigmentosa, likely
through a protective action from free radicals in affected photo-
receptors (Pasantes-Morales et al., 2002).
The beneficial effects of taurine are, in part, a result of its an-
tioxidant properties, as well as its ability to improve mitochondrial
function by stabilizing the electron transport chain and inhibiting
the generation of reactive oxygen species (Jong et al., 2012). Ex-
perimental results demonstrate that taurine attenuates hypergly-
cemia-induced apoptosis in human tubular cells via an inhibition
of oxidative stress (Verzola et al., 2002), inhibits the formation of
the Apaf-1/Caspase-9 apoptosome in cultured rat neonatal cardi-
omyocytes (Takatani et al., 2004), inhibits the m-calpain and cas-
pase-3 mediated cascades in an animal model of ischemia (Sun
and Xu, 2008) and reduces Bcl-2 in ethanol-induced apoptosis in
the developing cerebellum (J.-Y. Wu et al., 2009).

Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
8 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
Fig. 8. Chemical structure of taurine and some of its beneficial effects on the eye.
Moving from the CNS and the retina to the epithelial surface,
topical taurine application contributed to epithelial wound heal-
ing. A chitosan polymer of taurine (Tau-Gel) applied to full thick-
ness skin wounds of mice once a day for seven days, decreased
lipid peroxide and malondialdehyde formation, and increased
hydroxyproline levels (Değim et al., 2002; Ozmeri,ç et al., 2000),
thus improving wound healing. Taurine and derivatives also pos-
sess anti-inflammatory properties (Marcinkiewicz and Kontny,
2014; Marcinkiewicz et al., 2006). A clinical trial on patients af-
fected by severe epidemic keratoconjunctivitis showed that
N-chlorotaurine, a derivative of taurine, was well-tolerated, shor-
tened the duration of illness, and appeared to be a useful causative
therapeutic approach (Teuchner et al., 2005). These regenerative
properties of taurine have been exploited for the formulation of
protective polymer-based ocular surface films (Kedik et al., 2011).
Taurine has been included in contact lens cleaning solutions be-
cause of the regenerative effect on the tear film of contact lens
wearers (Grus et al., 2005). Taurine could protect corneal stroma
and epithelium from lactic acidosis caused by contact lens wearing
(Bonanno and Polse, 1987) because of its acid buffering ability
against lactic acidosis (Nakada et al., 1991). Tear proteomic studies
revealed distinct similarities between contact lens wearers and
dry eye patients that correlated with the finding that contact lens
wearers often develop dry eye symptoms. Taurine treatment
modifies the tear proteome of contact lens wearers and dry eye
patients shifting it towards the profile of healthy control subjects
(Funke et al., 2012).
Human corneal epithelial cells express Naþ -dependent os-
mosensitive taurine transport activity, thus under hypertonic
conditions there is an increase of intracellular taurine transport
that enhances cell survival, perhaps through a membrane stabili-
zation and/or an antioxidant effect (Shioda et al., 2002). In albino
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
rabbits, taurine effectively protected ocular surface tissues from
chemical damage induced by HOCl, and arrested the progression
of tissue damage (as measured by the level of lactate dehy-
drogenase activity) that had already been initiated by HOCl
(Koyama et al., 1996). A mixture of taurine, alpha-ketoglutarate,
pyruvate and pantothenate prevented corneal damage induced by
2-chloroethyl-ethyl sulfide (Varma et al., 1998).
Several studies have shown that the levels of taurine and GSH
were significantly reduced in the diabetic cataractous lens (Kilic
et al., 1999) and dietary supplementation with taurine reduced the
tissue damage and the mortality rate resulting from diabetes
(Obrosova and Stevens, 1999; Di Leo et al., 2004).
7. Conclusions
In conclusion, there are still several unanswered questions
about the occurrence and the role of amino acids in tears and the
reason why their relative concentration may fluctuate with time
and with the pathological state of the eye (they have been pro-
posed as a sensitive marker of eye conditions). However, there are
enough encouraging preclinical and clinical data suggesting that
an additional supply of amino acids, either systemic or topical,
may be useful to help the eye to recover its homeostatic equili-
brium, and a better functionality of the integrated system ocular
surface – lacrimal glands. More studies are certainly required, both
on experimental model systems and on human patients, to un-
derstand better the pharmacokinetics and the pharmacodynamics
of topical amino acids and their effects on different eye compart-
ments, such as lacrimal glands, corneal epithelial surface and
stroma. Some of these studies are already running in laboratories
and clinical settings, and will certainly be the subject of
 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
forthcoming publications on topical amino acids in the coming
years.
Declaration of interest
DR and SP are employees of Sooft Italia SpA, an Italian Phar-
maceutical Company involved in the production and commercia-
lization of food supplements and medical devices containing free
amino acids.
Acknowledgements
English proof reading of the manuscript by Dr. Antony Bridge-
wood is greatly appreciated.
References
Altshuler, D., Lo Turco, J.J., Rush, J., Cepko, C., 1993. Taurine promotes the differ-
entiation of a vertebrate retinal cell type in vitro. Development 119, 1317–1328.
Amjadi, S., Mai, K., McCluskey, P., Wakefield, D., 2013. The role of lumican in ocular
disease. ISRN Ophthalmol. 2013, 632302. http://dx.doi.org/10.1155/2013/
632302.
Aquilani, R., 2004. Oral amino acid administration in patients with diabetes mel-
litus: supplementation or metabolic therapy? Am. J. Cardiol. 93, 21A–22A. http:
//dx.doi.org/10.1016/j.amjcard.2003.11.005.
Aquilani, R., Boselli, M., Boschi, F., Viglio, S., Iadarola, P., Dossena, M., Pastoris, O.,
Verri, M., 2008a. Branched-chain amino acids may improve recovery from a
vegetative or minimally conscious state in patients with traumatic brain injury:
a pilot study. Arch. Phys. Med. Rehabil. 89, 1642–1647. http://dx.doi.org/
10.1016/j.apmr.2008.02.023.
Aquilani, R., Iadarola, P., Contardi, A., Boselli, M., Verri, M., Pastoris, O., Boschi, F.,
Arcidiaco, P., Viglio, S., 2005. Branched-chain amino acids enhance the cogni-
tive recovery of patients with severe traumatic brain injury. Arch. Phys. Med.
Rehabil. 86, 1729–1735. http://dx.doi.org/10.1016/j.apmr.2005.03.022.
Aquilani, R., Viglio, S., Iadarola, P., Opasich, C., Testa, A., Dioguardi, F.S., Pasini, E.,
2008b. Oral amino acid supplements improve exercise capacities in elderly
patients with chronic heart failure. Am. J. Cardiol. 101, 104E–110E. http://dx.doi.
org/10.1016/j.amjcard.2008.03.008.
Aragona, P., 2014. Topical cyclosporine: are all indications justified? Br. J. Oph-
thalmol. 98, 1001–1002. http://dx.doi.org/10.1136/bjophthalmol-2013-304728.
Aragona, P., Aguennouz, M., Rania, L., Postorino, E., Sommario, M.S., Roszkowska, A.
M., De Pasquale, M.G., Pisani, A., Puzzolo, D., 2015. Matrix metalloproteinase
9 and transglutaminase 2 expression at the ocular surface in patients with
different forms of dry eye disease. Ophthalmology 122, 62–71. http://dx.doi.
org/10.1016/j.ophtha.2014.07.048.
Aragona, P., Bucolo, C., Spinella, R., Giuffrida, S., Ferreri, G., 2005. Systemic omega-6
essential fatty acid treatment and pge1 tear content in Sjögren's syndrome
patients. Invest. Ophthalmol. Vis. Sci. 46, 4474–4479. http://dx.doi.org/10.1167/
iovs.04-1394.
Aragona, P., Di Pietro, R., Spinella, R., Mobrici, M., 2006. Conjunctival epithelium
improvement after systemic pilocarpine in patients with Sjogren's syndrome.
Br. J. Ophthalmol. 90, 166–170. http://dx.doi.org/10.1136/bjo.2005.078865.
Aragona, P., Papa, V., Micali, A., Santocono, M., Milazzo, G., 2002. Long term treat-
ment with sodium hyaluronate-containing artificial tears reduces ocular sur-
face damage in patients with dry eye. Br. J. Ophthalmol. 86, 181–184.
Aragona, P., Rania, L., Roszkowska, A.M., Spinella, R., Postorino, E., Puzzolo, D., Mi-
cali, A., 2013a. Effects of amino acids enriched tears substitutes on the cornea of
patients with dysfunctional tear syndrome. Acta Ophthalmol. 91, e437–e444.
http://dx.doi.org/10.1111/aos.12134.
Aragona, P., Spinella, R., Rania, L., Postorino, E., Sommario, M.S., Roszkowska, A.M.,
Puzzolo, D., 2013b. Safety and efficacy of 0.1% clobetasone butyrate eyedrops in
the treatment of dry eye in Sjögren syndrome. Eur. J. Ophthalmol. 23, 368–376.
http://dx.doi.org/10.5301/ejo.5000229.
Atkinson, M., Worthley, L.I.G., 2003. Nutrition in the critically ill patient: part I.
Essential physiology and pathophysiology. Crit. Care Resusc. J. Australas. Acad.
Crit. Care Med. 5, 109–120.
Baskin, S.I., Cohn, E.M., Kocsis, J.J., 1977. The effect of age on taurine levels in eye
tissues. Exp. Eye Res. 24, 315–319.
Berthoud, V.M., Beyer, E.C., 2009. Oxidative stress, lens gap junctions, and cataracts.
Antioxid. Redox Signal. 11, 339–353. http://dx.doi.org/10.1089/ars.2008.2119.
Bollhalder, L., Pfeil, A.M., Tomonaga, Y., Schwenkglenks, M., 2013. A systematic
literature review and meta-analysis of randomized clinical trials of parenteral
glutamine supplementation. Clin. Nutr. Edinb. Scotl. 32, 213–223. http://dx.doi.
org/10.1016/j.clnu.2012.11.003.
Bonanno, J.A., Polse, K.A., 1987. Corneal acidosis during contact lens wear: effects of
hypoxia and CO2. Invest. Ophthalmol. Vis. Sci. 28, 1514–1520.
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
9
Brignole-Baudouin, F., Baudouin, C., Aragona, P., Rolando, M., Labetoulle, M., Pisella,
P.J., Barabino, S., Siou-Mermet, R., Creuzot-Garcher, C., 2011. A multicentre,
double-masked, randomized, controlled trial assessing the effect of oral sup-
plementation of omega-3 and omega-6 fatty acids on a conjunctival in-
flammatory marker in dry eye patients. Acta Ophthalmol. 89, e591–e597. http:
//dx.doi.org/10.1111/j.1755-3768.2011.02196.x.
Caparrós, T., Lopez, J., Grau, T., 2001. Early enteral nutrition in critically ill patients
with a high-protein diet enriched with arginine, fiber, and antioxidants com-
pared with a standard high-protein diet. The effect on nosocomial infections
and outcome. J. Parenter. Enter. Nutr. 25, 299–308 (discussion 308–309).
Chakravarti, S., Magnuson, T., Lass, J.H., Jepsen, K.J., LaMantia, C., Carroll, H., 1998.
Lumican regulates collagen fibril assembly: skin fragility and corneal opacity in
the absence of lumican. J. Cell Biol. 141, 1277–1286.
ChenZhuo, L., Murube, J., Latorre, A., del Río, R.M., 2002. Different concentrations of
amino acids in tears of normal and human dry eyes. Adv. Exp. Med. Biol. 506,
617–621.
Chotikavanich, S., de Paiva, C.S., Li, D.Q., Chen, J.J., Bian, F., Farley, W.J., Pflugfelder, S.
C., 2009. Production and activity of matrix metalloproteinase-9 on the ocular
surface increase in dysfunctional tear syndrome. Invest. Ophthalmol. Vis. Sci.
50, 3203–3209. http://dx.doi.org/10.1167/iovs.08-2476.
Cho, Y.K., Kim, M.S., 2009. Dry eye after cataract surgery and associated in-
traoperative risk factors. Korean J. Ophthalmol. 23, 65–73. http://dx.doi.org/
10.3341/kjo.2009.23.2.65.
De Bandt, J.-P., Cynober, L., 2006. Therapeutic use of branched-chain amino acids in
burn, trauma, and sepsis. J. Nutr. 136, 308S–313SS.
Değim, Z., Celebi, N., Sayan, H., Babül, A., Erdoğan, D., Take, G., 2002. An in-
vestigation on skin wound healing in mice with a taurine-chitosan gel for-
mulation. Amino Acids 22, 187–198. http://dx.doi.org/10.1007/s007260200007.
De Paiva, C.S., Chen, Z., Koch, D.D., Hamill, M.B., Manuel, F.K., Hassan, S.S., Wilhel-
mus, K.R., Pflugfelder, S.C., 2006. The incidence and risk factors for developing
dry eye after myopic LASIK. Am. J. Ophthalmol. 141, 438–445. http://dx.doi.org/
10.1016/j.ajo.2005.10.006.
Di Leo, M.A.S., Santini, S.A., Silveri, N.G., Giardina, B., Franconi, F., Ghirlanda, G.,
2004. Long-term taurine supplementation reduces mortality rate in strepto-
zotocin-induced diabetic rats. Amino Acids 27, 187–191. http://dx.doi.org/
10.1007/s00726-004-0108-2.
Dogru, M., Nakamura, M., Shimazaki, J., Tsubota, K., 2013. Changing trends in the
treatment of dry-eye disease. Expert Opin. Invest. Drugs 22, 1581–1601. http:
//dx.doi.org/10.1517/13543784.2013.838557.
Doughty, M.J., 2016. On the prescribing of oral doxycycline or minocycline by UK
optometrists as part of management of chronic Meibomian Gland Dysfunction
(MGD). Contact Lens Anterior Eye 39, 2–8. http://dx.doi.org/10.1016/j.
clae.2015.08.002.
Efron, D., Barbul, A., 2000. Role of arginine in immunonutrition. J. Gastroenterol. 35
(Suppl 12), S20–S23.
Fanning, A.S., Jameson, B.J., Jesaitis, L.A., Anderson, J.M., 1998. The tight junction
protein ZO-1 establishes a link between the transmembrane protein occludin
and the actin cytoskeleton. J. Biol. Chem. 273, 29745–29753.
Favia, G., Mariggio, M.A., Maiorano, F., Cassano, A., Capodiferro, S., Ribatti, D., 2008.
Accelerated wound healing of oral soft tissues and angiogenic effect induced by
a pool of aminoacids combined to sodium hyaluronate (AMINOGAM). J. Biol.
Regul. Homeost. Agents 22, 109–116.
Fernstrom, J.D., Fernstrom, M.H., 2007. Tyrosine, phenylalanine, and catecholamine
synthesis and function in the brain. J. Nutr. 137, 1539S–1547S (discussion
1548S).
Fietzek, P.P., Kühn, K., 1975. Information contained in the amino acid sequence of
the alpha1(I)-chain of collagen and its consequences upon the formation of the
triple helix, of fibrils and crosslinks. Mol. Cell. Biochem. 8, 141–157.
Fillmann, H., Kretzmann, N.A., San-Miguel, B., Llesuy, S., Marroni, N., González-
Gallego, J., Tuñón, M.J., 2007. Glutamine inhibits over-expression of pro-in-
flammatory genes and down-regulates the nuclear factor kappaB pathway in an
experimental model of colitis in the rat. Toxicology 236, 217–226. http://dx.doi.
org/10.1016/j.tox.2007.04.012.
Foulks, G.N., 2007. Research in dry eye: report of the Research Subcommittee of the
International Dry Eye Workshop. Ocul. Surf. 5, 179–193.
Fuchs, S.A., Berger, R., Klomp, L.W.J., de Koning, T.J., 2005. D-amino acids in the
central nervous system in health and disease. Mol. Genet. Metab. 85, 168–180.
http://dx.doi.org/10.1016/j.ymgme.2005.03.003.
Funke, S., Azimi, D., Wolters, D., Grus, F.H., Pfeiffer, N., 2012. Longitudinal analysis of
taurine induced effects on the tear proteome of contact lens wearers and dry
eye patients using a RP-RP-Capillary-HPLC-MALDI TOF/TOF MS approach. J.
Proteom. 75, 3177–3190. http://dx.doi.org/10.1016/j.jprot.2012.03.018.
Gibala, M.J., 2000. Nutritional supplementation and resistance exercise: what is the
evidence for enhanced skeletal muscle hypertrophy? Can. J. Appl. Physiol. 25,
524–535.
Grus, F.H., Kramann, C., Bozkurt, N., Wiegel, N., Bruns, K., Lackner, N., Pfeiffer, N.,
2005. Effects of multipurpose contact lens solutions on the protein composition
of the tear film. Contact Lens Anterior Eye 28, 103–112. http://dx.doi.org/
10.1016/j.clae.2005.06.004.
Han, K.E., Yoon, S.C., Ahn, J.M., Nam, S.M., Stulting, R.D., Kim, E.K., Seo, K.Y., 2014.
Evaluation of dry eye and meibomian gland dysfunction after cataract surgery.
Am. J. Ophthalmol. 157, 1144–1150. http://dx.doi.org/10.1016/j.ajo.2014.02.036.
Hasegawa, S., Ichiyama, T., Sonaka, I., Ohsaki, A., Hirano, R., Haneda, Y., Fukano, R.,
Hara, M., Furukawa, S., 2011. Amino acids exhibit anti-inflammatory effects in
human monocytic leukemia cell line, THP-1 cells. Inflamm. Res. 60, 1013–1019.
http://dx.doi.org/10.1007/s00011-011-0362-1.
10 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
Hasegawa, S., Ichiyama, T., Sonaka, I., Ohsaki, A., Okada, S., Wakiguchi, H., Kudo, K.,
Kittaka, S., Hara, M., Furukawa, S., 2012. Cysteine, histidine and glycine exhibit
anti-inflammatory effects in human coronary arterial endothelial cells. Clin.
Exp. Immunol. 167, 269–274. http://dx.doi.org/10.1111/j.1365-2249.2011.04519.
x.
Hayes, K.C., Carey, R.E., Schmidt, S.Y., 1975. Retinal degeneration associated with
taurine deficiency in the cat. Science 188, 949–951.
Heinämäki, A.A., Muhonen, A.S., Piha, R.S., 1986. Taurine and other free amino acids
in the retina, vitreous, lens, iris-ciliary body, and cornea of the rat eye. Neu-
rochem. Res. 11, 535–542.
Hessen, M., Akpek, E.K., 2014. Dry eye: an inflammatory ocular disease. J. Oph-
thalmic Vis. Res. 9, 240–250.
Holecek, M., 2010. Three targets of branched-chain amino acid supplementation in
the treatment of liver disease. Nutrition 26, 482–490. http://dx.doi.org/10.1016/
j.nut.2009.06.027.
Houdijk, A.P., Rijnsburger, E.R., Jansen, J., Wesdorp, R.I., Weiss, J.K., McCamish, M.A.,
Teerlink, T., Meuwissen, S.G., Haarman, H.J., Thijs, L.G., van Leeuwen, P.A., 1998.
Randomised trial of glutamine-enriched enteral nutrition on infectious mor-
bidity in patients with multiple trauma. Lancet 352, 772–776. http://dx.doi.org/
10.1016/S0140-6736(98)02007-8.
Huang, J.C.-C., Sun, C.-C., Chang, C.-K., Ma, D.H.-K., Lin, Y.-F., 2012. Effect of hinge
position on corneal sensation and dry eye parameters after femtosecond laser-
assisted LASIK. J. Refract. Surg. 1995 (28), 625–631. http://dx.doi.org/10.3928/
1081597X-20120815-07.
Hubert-Buron, A., Leblond, J., Jacquot, A., Ducrotté, P., Déchelotte, P., Coëffier, M.,
2006. Glutamine pretreatment reduces IL-8 production in human intestinal
epithelial cells by limiting IkappaBalpha ubiquitination. J. Nutr. 136, 1461–1465.
Huxtable, R.J., 1992. Physiological actions of taurine. Physiol. Rev. 72, 101–163.
Imaki, H., Moretz, R., Wisniewski, H., Neuringer, M., Sturman, J., 1987. Retinal de-
generation in 3-month-old rhesus monkey infants fed a taurine-free human
infant formula. J. Neurosci. Res. 18, 602–614. http://dx.doi.org/10.1002/
jnr.490180414.
Jäger, K., Bönisch, U., Risch, M., Worlitzsch, D., Paulsen, F., 2009. Detection and
regulation of cationic amino acid transporters in healthy and diseased ocular
surface. Invest. Ophthalmol. Vis. Sci. 50, 1112–1121. http://dx.doi.org/10.1167/
iovs.08-2368.
Jäger, K., Garreis, F., Dunse, M., Paulsen, F.P., 2010a. Cationic amino acid transporters
and beta-defensins in dry eye syndrome. Dev. Ophthalmol. 45, 12–15. http://dx.
doi.org/10.1159/000315015.
Jäger, K., Garreis, F., Posa, A., Dunse, M., Paulsen, F.P., 2010b. Functional relationship
between cationic amino acid transporters and beta-defensins: implications for
dry skin diseases and the dry eye. Ann. Anat. Anat. Anz. Off Organ Anat. Ges.
192, 65–69. http://dx.doi.org/10.1016/j.aanat.2010.01.006.
Jammoul, F., Wang, Q., Nabbout, R., Coriat, C., Duboc, A., Simonutti, M., Dubus, E.,
Craft, C.M., Ye, W., Collins, S.D., Dulac, O., Chiron, C., Sahel, J.A., Picaud, S., 2009.
Taurine deficiency is a cause of vigabatrin-induced retinal phototoxicity. Ann.
Neurol. 65, 98–107. http://dx.doi.org/10.1002/ana.21526.
Jones, C., Palmer, T.E., Griffiths, R.D., 1999. Randomized clinical outcome study of
critically ill patients given glutamine-supplemented enteral nutrition. Nutrition
15, 108–115.
Jong, C.J., Azuma, J., Schaffer, S., 2012. Mechanism underlying the antioxidant ac-
tivity of taurine: prevention of mitochondrial oxidant production. Amino Acids
42, 2223–2232. http://dx.doi.org/10.1007/s00726-011-0962-7.
Kantorow, M., Hawse, J.R., Cowell, T.L., Benhamed, S., Pizarro, G.O., Reddy, V.N.,
Hejtmancik, J.F., 2004. Methionine sulfoxide reductase A is important for lens
cell viability and resistance to oxidative stress. Proc. Natl. Acad. Sci. USA 101,
9654–9659. http://dx.doi.org/10.1073/pnas.0403532101.
Kapprell, H.P., Cowin, P., Franke, W.W., Ponstingl, H., Opferkuch, H.J., 1985. Bio-
chemical characterization of desmosomal proteins isolated from bovine muzzle
epidermis: amino acid and carbohydrate composition. Eur. J. Cell Biol. 36,
217–229.
Kedik, S.A., Levachev, S.M., Panov, A.V., Sakaeva, I.V., Kharlov, A.E., Grigor’eva, O.A.,
Zhavoronok, E.S., Cherta, Y.V., Zaitsev, M.A., An, H.K., 2011. Formation of ul-
trathin protective films for ophthalmological use from aqueous solutions of
polymers and taurine. Pharm. Chem. J. 45, 245–247. http://dx.doi.org/10.1007/
s11094-011-0606-y.
Kilic, F., Bhardwaj, R., Caulfeild, J., Trevithick, J.R., 1999. Modelling cortical catar-
actogenesis 22: is in vitro reduction of damage in model diabetic rat cataract by
taurine due to its antioxidant activity? Exp. Eye Res. 69, 291–300. http://dx.doi.
org/10.1006/exer.1999.0697.
Kim, J.C., Cheong, T.B., Park, G.S., Park, M.H., Kwon, N.S., Yoon, H.Y., 2002. The role of
nitric oxide in ocular surface diseases. Adv. Exp. Med. Biol. 506, 687–695.
Kohlhaas, M., 1998. Corneal sensation after cataract and refractive surgery. J. Cat-
aract Refract. Surg. 24, 1399–1409.
Koyama, I., Nakamori, K., Nagahama, T., Ogasawara, M., Nemoto, M., 1996. The re-
activity of taurine with hypochlorous acid and its application for eye drops.
Adv. Exp. Med. Biol. 403, 9–18.
Lacey, J.M., Wilmore, D.W., 1990. Is glutamine a conditionally essential amino acid?
Nutr. Rev. 48, 297–309.
Lam, H., Bleiden, L., de Paiva, C.S., Farley, W., Stern, M.E., Pflugfelder, S.C., 2009. Tear
cytokine profiles in dysfunctional tear syndrome. Am. J. Ophthalmol. 147,
198–205. http://dx.doi.org/10.1016/j.ajo.2008.08.032.
Li, X.-M., Hu, L., Hu, J., Wang, W., 2007. Investigation of dry eye disease and analysis
of the pathogenic factors in patients after cataract surgery. Cornea 26, S16–S20.
http://dx.doi.org/10.1097/ICO.0b013e31812f67ca.
Luong, 2012. The beneficial role of thiamine in parkinson's disease: preliminary
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
report. J. Neurol. Res. . http://dx.doi.org/10.4021/jnr145e
MacKay, D., Miller, A.L., 2003. Nutritional support for wound healing. Altern. Med.
Rev. J. Clin. Ther. 8, 359–377.
Madl, J.E., McIlnay, T.R., Powell, C.C., Gionfriddo, J.R., 2005. Depletion of taurine and
glutamate from damaged photoreceptors in the retinas of dogs with primary
glaucoma. Am. J. Vet. Res. 66, 791–799.
Marcinkiewicz, J., Kontny, E., 2014. Taurine and inflammatory diseases. Amino Acids
46, 7–20. http://dx.doi.org/10.1007/s00726-012-1361-4.
Marcinkiewicz, J., Kurnyta, M., Biedroń, R., Bobek, M., Kontny, E., Maśliński, W.,
2006. Anti-inflammatory effects of taurine derivatives (taurine chloramine,
taurine bromamine, and taurolidine) are mediated by different mechanisms.
Adv. Exp. Med. Biol. 583, 481–492.
Meduri, A., Grenga, P.L., Scorolli, L., Ceruti, P., Ferreri, G., 2009. Role of cysteine in
corneal wound healing after photorefractive keratectomy. Ophthalmic Res. 41,
76–82. http://dx.doi.org/10.1159/000187623.
Mehta, S., Stewart, D.J., Langleben, D., Levy, R.D., 1995. Short-term pulmonary va-
sodilation with l-arginine in pulmonary hypertension. Circulation 92,
1539–1545. http://dx.doi.org/10.1161/01.CIR.92.6.1539.
Meloni, M., De Servi, B., Marasco, D., Del Prete, S., 2011. Molecular mechanism of
ocular surface damage: application to an in vitro dry eye model on human
corneal epithelium. Mol. Vis. 17, 113–126.
Militante, J., Lombardini, J.B., 2004. Age-related retinal degeneration in animal
models of aging: possible involvement of taurine deficiency and oxidative
stress. Neurochem. Res. 29, 151–160.
Nakada, T., Hida, K., Kwee, I.L., 1991. pH-lactate dissociation during anoxic insult:
taurine effect. Neuroreport 2, 325–328.
Nakatsukasa, M., Sotozono, C., Shimbo, K., Ono, N., Miyano, H., Okano, A., Hamuro,
J., Kinoshita, S., 2011. Amino Acid profiles in human tear fluids analyzed by
high-performance liquid chromatography and electrospray ionization tandem
mass spectrometry. Am. J. Ophthalmol. 151. http://dx.doi.org/10.1016/j.
ajo.2010.11.003, pp. 799–808.e1.
Nejima, R., Miyata, K., Tanabe, T., Okamoto, F., Hiraoka, T., Kiuchi, T., Oshika, T., 2005.
Corneal barrier function, tear film stability, and corneal sensation after pho-
torefractive keratectomy and laser in situ keratomileusis. Am. J. Ophthalmol.
139, 64–71. http://dx.doi.org/10.1016/j.ajo.2004.08.039.
Njie-Mbye, Y.F., Kulkarni-Chitnis, M., Opere, C.A., Barrett, A., Ohia, S.E., 2013. Lipid
peroxidation: pathophysiological and pharmacological implications in the eye.
Front. Physiol. 4, 366. http://dx.doi.org/10.3389/fphys.2013.00366.
Obrosova, I.G., Stevens, M.J., 1999. Effect of dietary taurine supplementation on GSH
and NAD(P)-redox status, lipid peroxidation, and energy metabolism in diabetic
precataractous lens. Invest. Ophthalmol. Vis. Sci. 40, 680–688.
Oepen, G., Cramer, H., Bernasconi, R., Martin, P., 1982. Huntington’s disease - im-
balance of free amino acids in the cerebrospinal fluid of patients and offspring
at-risk. Arch. Psychiatr. Nervenkrankh. 231, 131–140.
Ozmeriç, N., Ozcan, G., Haytaç, C.M., Alaaddinoğlu, E.E., Sargon, M.F., Senel, S., 2000.
Chitosan film enriched with an antioxidant agent, taurine, in fenestration de-
fects. J. Biomed. Mater. Res. 51, 500–503.
Panjwani, N.A., Harding, J.J., 1978. Isolation and hydroxylysine glycoside content of
some cyanogen bromide-cleaved fragments of collagen from bovine corneal
stroma. Biochem. J. 171, 687–695.
Pasantes-Morales, H., Quesada, O., Cárabez, A., Huxtable, R.J., 1983. Effects of the
taurine transport antagonist, guanidinoethane sulfonate, and beta-alanine on
the morphology of rat retina. J. Neurosci. Res. 9, 135–143. http://dx.doi.org/
10.1002/jnr.490090205.
Pasantes-Morales, H., Quiroz, H., Quesada, O., 2002. Treatment with taurine, dil-
tiazem, and vitamin E retards the progressive visual field reduction in retinitis
pigmentosa: a 3-year follow-up study. Metab. Brain Dis. 17, 183–197.
Pasini, E., Aquilani, R., Dioguardi, F.S., D’Antona, G., Gheorghiade, M., Taegtmeyer,
H., 2008. Hypercatabolic syndrome: molecular basis and effects of nutritional
supplements with amino acids. Am. J. Cardiol. 101, 11E–15E. http://dx.doi.org/
10.1016/j.amjcard.2008.02.074.
Platell, C., Kong, S.E., McCauley, R., Hall, J.C., 2000. Branched-chain amino acids. J.
Gastroenterol. Hepatol. 15, 706–717.
Polatnick, J., La Tessa, A.J., Katzin, H.M., 1957. Comparison of collagen preparations
from beef cornea and sclera. Biochim. Biophys. Acta 26, 365–369.
Rascher, K., Servos, G., Berthold, G., Hartwig, H.-G., Warskulat, U., Heller-Stilb, B.,
Häussinger, D., 2004. Light deprivation slows but does not prevent the loss of
photoreceptors in taurine transporter knockout mice. Vis. Res. 44, 2091–2100.
http://dx.doi.org/10.1016/j.visres.2004.03.027.
Rathore, M.S., Gupta, V.B., 2010. Protective effect of amino acids on eye lenses
against oxidative stress induced by hydrogen peroxide. Asian J. Pharm. Clin. Res.
3, 166–169.
Reddy, V.N., 1990. Glutathione and its function in the lens–an overview. Exp. Eye
Res. 50, 771–778.
Ripps, H., Shen, W., 2012. Review: taurine: a “very essential” amino acid. Mol. Vis.
18, 2673–2686.
Saxena, R.N., Pendse, V.K., Khanna, N.K., 1984. Anti-inflammatory and analgesic
properties of four amino-acids. Indian J. Physiol. Pharmacol. 28, 299–305.
Scognamiglio, R., Testa, A., Aquilani, R., Dioguardi, F.S., Pasini, E., 2008. Impairment
in walking capacity and myocardial function in the elderly: is there a role for
nonpharmacologic therapy with nutritional amino acid supplements? Am. J.
Cardiol. 101, 78E–81E. http://dx.doi.org/10.1016/j.amjcard.2008.03.005.
Shioda, R., Reinach, P.S., Hisatsune, T., Miyamoto, Y., 2002. Osmosensitive taurine
transporter expression and activity in human corneal epithelial cells. Invest.
Ophthalmol. Vis. Sci. 43, 2916–2922.
Stonecipher, K.G., Chia, J., Onyenwenyi, A., Villanueva, L., Hollander, D.A., 2013.
 D. Rusciano et al. / European Journal of Pharmacology ∎ (∎∎∎∎) ∎∎∎–∎∎∎
Health claims database study of cyclosporine ophthalmic emulsion treatment
patterns in dry eye patients. Ther. Clin. Risk Manag. 9, 409–415. http://dx.doi.
org/10.2147/TCRM.S49754.
Sun, M., Xu, C., 2008. Neuroprotective mechanism of taurine due to up-regulating
calpastatin and down-regulating calpain and caspase-3 during focal cerebral
ischemia. Cell. Mol. Neurobiol. 28, 593–611. http://dx.doi.org/10.1007/
s10571-007-9183-8.
Takatani, T., Takahashi, K., Uozumi, Y., Shikata, E., Yamamoto, Y., Ito, T., Matsuda, T.,
Schaffer, S.W., Fujio, Y., Azuma, J., 2004. Taurine inhibits apoptosis by pre-
venting formation of the Apaf-1/caspase-9 apoptosome. Am. J. Physiol. Cell
Physiol. 287, C949–C953. http://dx.doi.org/10.1152/ajpcell.00042.2004.
Teuchner, B., Nagl, M., Schidlbauer, A., Ishiko, H., Dragosits, E., Ulmer, H., Aoki, K.,
Ohno, S., Mizuki, N., Gottardi, W., Larcher, C., 2005. Tolerability and efficacy of
N-chlorotaurine in epidemic keratoconjunctivitis–a double-blind, randomized,
phase-2 clinical trial. J. Ocul. Pharmacol. Ther. J. Assoc. Ocul. Pharmacol. Ther.
21, 157–165. http://dx.doi.org/10.1089/jop.2005.21.157.
Torres Munoz, I., Grizzi, F., Russo, C., Camesasca, F.I., Dioguardi, N., Vinciguerra, P.,
2003. The role of amino acids in corneal stromal healing: a method for eval-
uating cellular density and extracellular matrix distribution. J. Refract. Surg.
1995 (19), S227–S230.
Varma, S.D., Devamanoharan, P.S., Ali, A.H., Brozetti, J., Petrali, J., Lehnert, E., Weir,
A., 1998. Half mustard (CEES) induced damage to rabbit cornea: attenuating
effect of taurine-pyruvate-alpha-ketoglutarate-pantothenate mixture. J. Ocul.
Pharmacol. Ther. J. Assoc. Ocul. Pharmacol. Ther. 14, 423–428.
Verzola, D., Bertolotto, M.B., Villaggio, B., Ottonello, L., Dallegri, F., Frumento, G.,
Berruti, V., Gandolfo, M.T., Garibotto, G., Deferran, G., 2002. Taurine prevents
apoptosis induced by high ambient glucose in human tubule renal cells. J. In-
vestig. Med. 50, 443–451.
Please cite this article as: Rusciano, D., et al., Free amino acids: an innovative treatment for ocular surface disease. Eur J Pharmacol
(2016), http://dx.doi.org/10.1016/j.ejphar.2016.04.029i
11
Vinciguerra, P., Camesasca, F.I., Ponzin, D., 2002. Use of amino acids in refractive
surgery. J. Refract. Surg. 1995 (18), S374–S377.
Wilby, J., Kainth, A., Hawkins, N., Epstein, D., McIntosh, H., McDaid, C., Mason, A.,
Golder, S., O’Meara, S., Sculpher, M., Drummond, M., Forbes, C., 2005. Clinical
effectiveness, tolerability and cost-effectiveness of newer drugs for epilepsy in
adults: a systematic review and economic evaluation. Health Technol. Assess. 9,
1–157 (iii–iv).
Wilson, S.E., 2001. Laser in situ keratomileusis-induced (presumed) neurotrophic
epitheliopathy. Ophthalmology 108, 1082–1087.
Wu, G., 2009. Amino acids: metabolism, functions, and nutrition. Amino Acids 37,
1–17. http://dx.doi.org/10.1007/s00726-009-0269-0.
Wu, G., Bazer, F.W., Burghardt, R.C., Johnson, G.A., Kim, S.W., Knabe, D.A., Li, P., Li, X.,
McKnight, J.R., Satterfield, M.C., Spencer, T.E., 2011. Proline and hydroxyproline
metabolism: implications for animal and human nutrition. Amino Acids 40,
1053–1063. http://dx.doi.org/10.1007/s00726-010-0715-z.
Wu, G., Bazer, F.W., Davis, T.A., Kim, S.W., Li, P., Marc Rhoads, J., Carey Satterfield, M.,
Smith, S.B., Spencer, T.E., Yin, Y., 2009a. Arginine metabolism and nutrition in
growth, health and disease. Amino Acids 37, 153–168. http://dx.doi.org/
10.1007/s00726-008-0210-y.
Wu, J.-Y., Wu, H., Jin, Y., Wei, J., Sha, D., Prentice, H., Lee, H.-H., Lin, C.-H., Lee, Y.-H.,
Yang, L.-L., 2009b. Mechanism of neuroprotective function of taurine. Adv. Exp.
Med. Biol. 643, 169–179. http://dx.doi.org/10.1007/978-0-387-75681-3_17.
Yildirim, Z., Kiliç, N., Ozer, C., Babul, A., Take, G., Erdogan, D., 2007. Effects of taurine
in cellular responses to oxidative stress in young and middle-aged rat liver.
Ann. N. Y. Acad. Sci. 1100, 553–561. http://dx.doi.org/10.1196/annals.1395.061.
Young, T.L., Cepko, C.L., 2004. A role for ligand-gated ion channels in rod photo-
receptor development. Neuron 41, 867–879.