Abstract
Purpose
A pilot study to validate the collection of vitreous reflux (VR) after intravitreal injection using Schirmers tear strips was carried out. We assessed its efficiency for proteomics studies by estimating the differential expression of 27 cytokines using multiplexed bead array in diabetic macular oedema and proliferative diabetic retinopathy. To set, validate and assess the efficacy of Schirmer tear strips for collecting VR in patients undergoing intravitreal injections for diabetic macular oedema (DME).
Patients and methods
VR samples were collected from 11 eyes of DME patients after intravitreal injections using Schirmer tear strips. Undiluted vitrectomy samples were obtained from six eyes of non-diabetic patients with idiopathic macular hole and seven eyes of diabetic patients with high-risk proliferative diabetic retinopathy (Hr-PDR), which were also subsampled on the Schirmer tear strips. Tear sampling was done in a subset of the DME patients. Total protein concentration between VR and vitrectomy samples was compared. Levels of the set of 27 cytokines in Schirmer tear strips samples were measured. Inter-group comparison for cytokines was done using Mann–Whitney U-test.
Results
Similar protein concentration in VR samples and vitrectomy samples (P<0.05) was obtained. Tear protein contamination was not detected in VR samples. In comparison with no-DR patients, 25 and 20 of the measured 27 cytokines were significantly elevated (P<0.05) in the Hr-PDR and DME patients, respectively. As compared with no-DR patients, vascular endothelial growth factor was only moderately elevated in DME patients (P>0.05), but significantly elevated in Hr-PDR patients (P<0.05). Interleukin 1 receptor antagonist/interleukin 1b (IL1RA/IL1b) ratio was 13 times higher in DME patients as compared with Hr-PDR group.
Conclusion
We demonstrated a simple, safe method of VR sampling. This technique provides a pure, albeit small, vitreous sample for proteomics. IL1RA/IL1b ratio was found to be 13-fold higher in the DME group as compared to the Hr-PDR.
Similar content being viewed by others
Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
References
Yau JW, Rogers SL, Kawasaki R, Lamoureux EL, Kowalski JW, Bek T et al. Meta-Analysis for Eye Disease (META-EYE) Study Group. Global prevalence and major risk factors of diabetic retinopathy. Diabetes Care 2012; 35 (3): 556–564.
Amoaku WM, Saker S, Stewart EA . A review of therapies for diabetic macular oedema and rationale for combination therapy. Eye 2015; 29 (9): 1115–1130.
Capitão M, Soares R . Angiogenesis and inflammation crosstalk in diabetic retinopathy. J Cell Biochem 2016; 117 (11): 2443–2453.
Kaštelan S, Tomić M, Gverović Antunica A, Salopek Rabatić J, Ljubić S . Inflammation and pharmacological treatment in diabetic retinopathy. Mediators Inflamm 2013; 2013: 213130.
Totan Y, Güler E, Gürağaç FB . Dexamethasone intravitreal implant for chronic diabetic macular edema resistant to intravitreal bevacizumab treatment. Curr Eye Res 2016; 41 (1): 107–113.
Murthy KR, Goel R, Subbannayya Y, Jacob HK, Murthy PR, Manda SS et al. Proteomic analysis of human vitreous humor. Clin Proteomics 2014; 11 (1): 29.
Yu J, Liu F, Cui SJ, Liu Y, Song ZY, Cao H et al. Vitreous proteomic analysis of proliferative vitreoretinopathy. Proteomics 2008; 8 (17): 3667–3678.
Kim T, Kim SJ, Kim K, Kang UB, Lee C, Park KS et al. Profiling of vitreous proteomes from proliferative diabetic retinopathy and nondiabetic patients. Proteomics 2007; 7 (22): 4203–4215.
Ecker SM, Hines JC, Pfahler SM, Glaser BM . Aqueous cytokine and growth factor levels do not reliably reflect those levels found in the vitreous. Mol Vis 2011; 17: 2856–2863.
Brodie FL, Ruggiero J, Ghodasra DH, Hui JZ, VanderBeek BL, Brucker AJ . Volume and composition of reflux after intravitreal injection. Retina 2014; 34 (7): 1473–1476.
Cacciamani A, Parravano M, Scarinci F, Esposito G, Varano M, Micera A . A simple spontaneous vitreal reflux collecting procedure during intravitreal injection: set-up and validation studies. Curr Eye Res 2016; 41 (7): 971–976.
Saijyothi AV, Angayarkanni N, Syama C, Utpal T, Shweta A, Bhaskar S et al. Two dimensional electrophoretic analysis of human tears: collection method in dry eye syndrome. Electrophoresis 2010; 31 (20): 3420–3427.
Ghodasra DH, Fante R, Gardner TW, Langue M, Niziol LM, Besirli C et al. Safety and feasibility of quantitative multiplexed cytokine analysis from office-based vitreous aspiration. Invest Ophthalmol Vis Sci 2016; 57 (7): 3017–3023.
Elshal MF, McCoy JP . Multiplex bead array assays: performance evaluation and comparison of sensitivity to ELISA. Methods 2006; 38 (4): 317–323.
Bromberg-White JL, Glazer L, Downer R, Furge K, Boguslawski E, Duesbery NS . Identification of VEGF-independent cytokines in proliferative diabetic retinopathy vitreous. Invest Ophthalmol Vis Sci 2013; 54 (10): 6472–6480.
Zhou J, Wang S, Xia X . Role of intravitreal inflammatory cytokines and angiogenic factors in proliferative diabetic retinopathy. Curr Eye Res 2012; 37 (5): 416–420.
Murugeswari P, Shukla D, Rajendran A, Kim R, Namperumalsamy P, Muthukkaruppan V . Proinflammatory cytokines and angiogenic and anti-angiogenic factors in vitreous of patients with proliferative diabetic retinopathy and Eales' disease. Retina 2008; 28 (6): 817–824.
Maier R, Weger M, Haller-Schober EM, El-Shabrawi Y, Wedrich A, Theisl A et al. Multiplex bead analysis of vitreous and serum concentrations of inflammatory and proangiogenic factors in diabetic patients. Mol Vis 2008; 14: 637–643.
Chernykh VV, Varvarinsky EV, Smirnov EV, Chernykh DV, Trunov AN . Proliferative and inflammatory factors in the vitreous of patients with proliferative diabetic retinopathy. Indian J Ophthalmol 2015; 63 (1): 33–36.
Yoshimura T, Sonoda KH, Sugahara M, Mochizuki Y, Enaida H, Oshima Y et al. Comprehensive analysis of inflammatory immune mediators in vitreoretinal diseases. PLoS One 2009; 4 (12): e8158.
Tang J, Kern TS . Inflammation in diabetic retinopathy. Prog Retin Eye Res 2011; 30 (5): 343–358.
Adamis AP . Is diabetic retinopathy an inflammatory disease? Br J Ophthalmol 2002; 86 (4): 363–365.
Elner SG, Elner VM, Jaffe GJ, Stuart A, Kunkel SL, Strieter RM . Cytokines in proliferative diabetic retinopathy and proliferative vitreoretinopathy. Curr Eye Res 1995; 14 (11): 1045–1053.
Adamis AP, Miller JW, Bernal MT, D'Amico DJ, Folkman J, Yeo TK et al. Increased vascular endothelial growth factor levels in the vitreous of eyes with proliferative diabetic retinopathy. Am J Ophthalmol 1994; 118 (4): 445–450.
Aiello LP, Avery RL, Arrigg PG, Keyt BA, Jampel HD, Shah ST et al. Vascular endothelial growth factor in ocular fluid of patients with diabetic retinopathy and other retinal disorders. N Engl J Med 1994; 331 (22): 1480–1487.
Joussen AM, Poulaki V, Le ML, Koizumi K, Esser C, Janicki H et al. A central role for inflammation in the pathogenesis of diabetic retinopathy. FASEB J 2004; 18 (12): 1450–1452.
Klaassen I, Van Noorden CJ, Schlingemann RO . Molecular basis of the inner blood-retinal barrier and its breakdown in diabetic macular edema and other pathological conditions. Prog Retin Eye Res 2013; 34: 19–48.
Zhang X, Zeng H, Bao S, Wang N, Gillies MC . Diabetic macular edema: new concepts in patho-physiology and treatment. Cell Biosci 2014; 4: 27.
Dong N, Xu B, Chu L, Tang X . Study of 27 aqueous humor cytokines in type 2 diabetic patients with or without macular edema. PLoS One 2015; 10 (4): e0125329.
Dong N, Xu B, Wang B, Chu L, Tang X . Aqueous cytokines as predictors of macular edema in patients with diabetes following uncomplicated phacoemulsification cataract surgery. Biomed Res Int 2015; 2015: 126984.
Jonas JB, Jonas RA, Neumaier M, Findeisen P . Cytokine concentration in aqueous humor of eyes with diabetic macular edema. Retina 2012; 32 (10): 2150–2157.
Sohn HJ, Han DH, Kim IT, Oh IK, Kim KH, Lee DY et al. Changes in aqueous concentrations of various cytokines after intravitreal triamcinolone versus bevacizumab for diabetic macular edema. Am J Ophthalmol 2011; 152 (4): 686–694.
Roh MI, Kim HS, Song JH, Lim JB, Kwon OW . Effect of intravitreal bevacizumab injection on aqueous humor cytokine levels in clinically significant macular edema. Ophthalmology 2009; 116 (1): 80–86.
Funatsu H, Noma H, Mimura T, Eguchi S, Hori S . Association of vitreous inflammatory factors with diabetic macular edema. Ophthalmology 2009; 116 (1): 73–79.
Lee WJ, Kang MH, Seong M, Cho HY . Comparison of aqueous concentrations of angiogenic and inflammatory cytokines in diabetic macular oedema and macular oedema due to branch retinal vein occlusion. Br J Ophthalmol 2012; 96 (11): 1426–1430.
Funatsu H, Yamashita H, Noma H, Mimura T, Yamashita T, Hori S . Increased levels of vascular endothelial growth factor and interleukin-6 in the aqueous humor of diabetics with macular edema. Am J Ophthalmol 2002; 133 (1): 70–77.
Usman Saeed M, Batra R, Qureshi F, Clark D . Reflux of drug during intra-vitreal anti-VEGF therapies. Semin Ophthalmol 2011; 26 (6): 357–360.
Brodie FL, Ruggiero J, Ghodasra DH, Eftekhari K, Hui JZ, Brucker AJ et al. A novel method for the measurement of reflux from intravitreal injections: data from 20 porcine eyes. Curr Eye Res 2014; 39 (7): 752–757.
Carmo A, Cunha-Vaz JG, Carvalho AP, Lopes MC . L-arginine transport in retinas from streptozotocin diabetic rats: correlation with the level of IL-1 beta and NO synthase activity. Vis Res 1999; 39 (23): 3817–3823.
Li W, Yanoff M, Jian B, He Z . Altered mRNA levels of antioxidant enzymes in pre-apoptotic pericytes from human diabetic retinas. Cell Mol Biol 1999; 45 (1): 59–66.
Demircan N, Safran BG, Soylu M, Ozcan AA, Sizmaz S . Determination of vitreous interleukin-1 (IL-1) and tumour necrosis factor (TNF) levels in proliferative diabetic retinopathy. Eye 2006; 20 (12): 1366–1369.
Abuel Asrar AM, Maimone D, Morse PH, Gregory S, Reder AT . Cytokines in the vitreous of patients with proliferative diabetic retinopathy. Am J Ophthalmol 1992; 114 (6): 731–736.
Kowluru RA, Odenbach S . Role of interleukin-1beta in the development of retinopathy in rats: effect of antioxidants. Invest Ophthalmol Vis Sci 2004; 45 (11): 4161–4166.
Gerhardinger C, Liu Y, Dagher Z . Overexpression of IL-1 receptor antagonist in the rat retina by AAV2-mediated gene transfer prevents capillary loss in experimental diabetes. Invest Ophthalmol Vis Sci 2012; 53: 5764.
Stahel M, Becker M, Graf N, Michels S . Systemic interleukin 1β inhibition in proliferative diabetic retinopathy: a prospective open-label study using Canakinumab. Retina 2016; 36 (2): 385–391.
Arend WP, Gabay C . Physiologic role of interleukin-1 receptor antagonist. Arthritis Res 2000; 2 (4): 245–248.
Arend WP . Interleukin-1 receptor antagonist. Adv Immunol 1993; 54: 167–227.
Dinarello CA . Interleukin-1 in the pathogenesis and treatment of inflammatory diseases. Blood 2011; 117 (14): 3720–3732.
Goldbach-Mansky R, Dailey NJ, Canna SW, Gelabert A, Jones J, Rubin BI et al. Neonatal-onset multisystem inflammatory disease responsive to interleukin-1β inhibition. N Engl J Med 2006; 355 (6): 581–592.
Schiff MH . Role of interleukin 1 and interleukin 1 receptor antagonist in the mediation of rheumatoid arthritis. Ann Rheum Dis 2000; 59: i103–i108.
Suffee N, Richard B, Hlawaty H, Oudar O, Charnaux N, Sutton A . Angiogenic properties of the chemokine RANTES/CCL5. Biochem Soc Trans 2011; 39 (6): 1649–1653.
Acknowledgements
We acknowledge ICER (NIH), NIRT, Chennai for the permission to use multiplex reader facility.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no conflict of interest.
Additional information
Supplementary Information accompanies this paper on Eye website
Rights and permissions
About this article
Cite this article
Srividya, G., Jain, M., Mahalakshmi, K. et al. A novel and less invasive technique to assess cytokine profile of vitreous in patients of diabetic macular oedema. Eye 32, 820–829 (2018). https://doi.org/10.1038/eye.2017.285
Received:
Accepted:
Published:
Version of record:
Issue date:
DOI: https://doi.org/10.1038/eye.2017.285
This article is cited by
-
Study of aqueous humour inflammatory mediators’ levels in a cohort of Egyptian patients with diabetic macular oedema
BMC Ophthalmology (2023)
-
Emerging applications of bioinformatics and artificial intelligence in the analysis of biofluid markers involved in retinal occlusive diseases: a systematic review
Graefe's Archive for Clinical and Experimental Ophthalmology (2023)
-
Vitreous humor proteome: unraveling the molecular mechanisms underlying proliferative and neovascular vitreoretinal diseases
Cellular and Molecular Life Sciences (2023)
-
Changes in aqueous and vitreous inflammatory cytokine levels in proliferative diabetic retinopathy: a systematic review and meta-analysis
Eye (2022)
-
The role of IL-6-174 G/C polymorphism and intraocular IL-6 levels in the pathogenesis of ocular diseases: a systematic review and meta-analysis
Scientific Reports (2020)


