Transcript Anti-Inflammatory Activity of Topical Azithromycin on Corneal Innate

Anti-Inflammatory Activity of Topical Azithromycin
on Corneal Innate Immune Responses
Zahra Sadrai MD, Amir Reza Hajrasouliha MD, Sunil Chauhan PhD, Daniel
Saban PhD, Reza Dana MD, MPH, MSc.
Schepens Eye Research Institute,
Massachusetts Eye & Ear Infirmary,
Department of Ophthalmology,
Harvard Medical School, Boston, MA
Financial Disclosure: The authors have no financial interest in the product mentioned in this poster.
Funding Sources: Inspire Pharmaceuticals provided research support for this study.
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
INTRODUCTION
Corneal inflammation is a critical facet of many ocular pathologies, including corneal angiogenesis and
corneal allograft rejection and represents a leading cause of blindness worldwide (1). BM-derived antigen
presenting cells (APCs) in the cornea and ocular surface comprise diverse subsets of CD45+ cells including
macrophages (CD11b+) that normally reside in the stroma, and CD11c+ dendritic cells in the epithelium (2,
3). Innate immunity, the major mechanism for acute inflammatory response, involves cellular trafficking into
the cornea in response to traumatic, noxious, or microbial stimuli (2, 4-6). Cytokines, the molecular
components of innate immune responses, coordinate leukocyte migration in immunity and inflammation (3).
Macrolides are broad-spectrum antibiotics that are widely used to treat bacterial infections (7). There is also
in vitro evidence that macrolides have anti-inflammatory and immunomodulatory activities. Studies have
demonstrated that macrolides suppress the activation of NF-kB and the release of pro-inflammatory
cytokines in vivo (8, 9).
We hypothesized here that Azithromycin (AZM), a broad-spectrum macrolide antibiotic recently approved for
ocular infection, has potential immunomodulatory effects on corneal inflammation.
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
PURPOSE
To investigate the potential anti-inflammatory effects of AZM on corneal innate
immune responses.
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
METHODS
Cornea of 6-8 week old BALB/c mice underwent thermal cautery to induce inflammation and leukocyte influx.
Fifteen corneas were randomly divided into three groups treated topically either with AZM ophthalmic solution
1% (AzaSite®; Inspire Pharmaceuticals, Inc, NC, USA), the relevant vehicle (DuraSite®; Inspire
Pharmaceuticals, Inc, NC, USA), or prednisolone acetate 1% twice per day. Corneas were harvested at
various time-points to characterize the inflammatory infiltrate via FACS analysis, and to quantitate expression
of cytokines via real time PCR. All experiments were approved by the IACUC of the Schepens Eye Research
Institute and adhered to the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.
Days 1, 3, 7, 10, 14
Naïve
Flow cytometry CD45,
CD11c
Cauterized Cornea
Vehicle (Bid)
Cauterized Cornea
AZM (Bid)
Cauterized Cornea
Prednisolone (Bid)
Real Time PCR IL-10,
IL-6
Anti-Inflammatory
ActivityInflammation
of Topical Azithromycin
on
Amelioration of Corneal
by Topical
CornealAzithromycin
Innate Immune
Responses
(AZM)
RESULTS
B.
*
Cell numbers per cornea
Cell numbers per cornea
A.
*
Figure 1. Relative to the vehicle control, the AZM-treated group showed a significant reduction in
total infiltration of CD45+ (pan-leukocyte marker) cells at day 1 (30%) and day 7 (39%), which was
similarly observed in the prednisolone-treated group (A). Dendritic cells (CD11c+) demonstrated a
reduced infiltration at day 7 in the AZM-treated (35% reduction) and prednisolone-treated (40%
reduction) groups.
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
IL-10
IL-6
Vehicle
40
Vehicle
50
AZM
45
35
40
30
Fold increase compared to naive
Fold increase compared to naive
AZM
25
20
15
10
5
0
35
30
25
20
15
10
5
0
Day 1
Day 3
Day 7
Day 10
Day 14
Day 1
Day 3
Day 7
Day 10
Day 14
Figure 2. Corneas treated with AZM increased the expression level of IL-10 in all time-points
(A). The relative expression of IL-6 was variable at different time-points (B).
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
CONCLUSIONS
Following an inflammatory insult, topical AZM reduced infiltration of
leukocytes considerably to levels comparable to the prednisolone-treated
group. This was further supported by an associated increase in antiinflammatory cytokine IL-10. Thus, these data suggest that topical AZM
holds endogenous anti-inflammatory properties.
Anti-Inflammatory Activity of Topical Azithromycin on
Corneal Innate Immune Responses
References
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Streilein JW. Immunology and immunopathology of corneal transplantation. Chem Immunol 1999:73:186-206.
2.
Dana MR. Corneal antigen-presenting cells: diversity, plasticity, and disguise. The Cogan lecture. Invest Ophthalmol Vis
Sci. 2004;45:722-727.
Delves PJ, Roitt IM. The immune system. First of two parts. N Engl J Med. 2000 6;343(1):37-49.
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Hamrah P, Huq SO, Liu Y, Zhang Q, Dana MR. Corneal immunity is mediated by heterogeneous population of
antigenpresenting cells. J Leukoc Biol 2003;74:172-178.
Hamrah P, Zhang Q, Liu Y, Dana MR. Novel Characterization of MHC Class II–Negative Population of Resident Corneal
Langerhans Cell–Type Dendritic Cells. Invest Ophthalmol Vis Sci. 2003;43:639-646.
Hamrah P, Zhang Q, Liu Y, Dana MR. The Corneal Stroma Is Endowed with a Significant Number of Resident Dendritic
Cells. Invest Ophthalmol Vis Sci. 2003;44:581-589.
G.G. Zhanel, M. Dueck, D.J. Hoban, L.M. Vercaigne, J.M. Embil, A.S. Gin and J.A. Karlowsky, Review of macrolides
and ketolides: focus on respiratory tract infections, Drugs 61 2001; 61:443–498.
O. Culic, V. Erakovic and M.J. Parnham, Anti-inflammatory effects of macrolide antibiotics, Eur. J. Pharmacol. 2001;
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