This study is a retrospective single-center observational study. This study followed the tenets of the Declaration of Helsinki and was approved by the Ethics Committee of Shanghai General Hospital (No. 2016KY003). Data were obtained from the database of ophthalmology in Shanghai General Hospital. Written informed consent was obtained from all subjects.
Patients with untreated clinically significant DME in the study eye without previous anti-VEGF drug or steroid treatment and without clinically significant macular edema in fellow eye were included in the study. A central subfield thickness of 320 μm for males and 305 μm for females was followed for clinically significant macular edema (for Heidelberg device)(16). Intravitreal conbercept (0.5 mg/0.05 mL) injections were performed by one retinal specialist. Patients were followed-up for 12 months between August 2010 and December 2013. All patients received three injections in the first three months, and then a pro re nata (PRN) strategy was then followed. Each patient underwent a thorough ophthalmic examination, including BCVA, slit lamp observation, and SD-OCT measurements, before the first injection and at each follow-up visit. The exclusion criteria were the following: (1) any non–diabetes-related macular edema; (2) any ophthalmological or neurological disease that could affect or could have affected the visual acuity (uncontrolled glaucoma, uveitis, retinal macular traction or macular epiretinal membrane); and (3) an eye with a history that could potentially affect the retinal layer thickness, e.g., vitrectomy, laser photocoagulation and cataract extraction.
The BCVA was tested at baseline and monthly thereafter using Early Treatment Diabetic Retinopathy Study (ETDRS) charts at 4 meters. The serum glycated hemoglobin level was assessed at baseline.
Optical Coherence Tomography
A 5.4×5.4-mm area of the macular region centered on the fovea was examined by the Spectralis SD-OCT (Heidelberg Engineering, Heidelberg, Germany) with display mode; volume scans of 97 sections were centered on the fovea, and 5 B-scan images at each section were averaged. For standardization, all examinations were performed by one well-trained technician who was masked to the identity of the subjects and was not involved in the data analysis. A build-in automatic recognition system enabled scanning of precisely the same location during the follow-up examinations.
To identify eyes/patients with increased thickness in the central subfield, the protocol proposed by DRCR.net was followed. Foveal thickness was manually segmented by one retinal specialist (Y.S.) using the SD-OCT software (Spectralis version 18.104.22.168, Heidelberg Engineering, Heidelberg, Germany). All scans of a particular eye were graded consistently by the same reader to counteract a potential annotation bias. The whole retina was divided into three circles as follows: central subfield (diameter of 1 1000μm) and inner (diameter of 3000μm) and outer (diameter of 6000μm) rings. The inner and outer rings were divided into the superior, inferior, nasal and temporal regions, and the average retinal thickness in these total 9 regions was measured automatically. The Heidelberg Software recognizes retinal tissue interfaces and, using these landmarks, allows the software to handle the following retinal layers: retinal nerve fiber layer (RNFL), ganglion cell layer (GCL), inner plexiform layer (IPL), inner nuclear layer (INL), outer plexiform layer (OPL), outer nuclear layer (ONL), and the photoreceptor–retinal pigment epithelium complex (PR). All layers were first segmented using the built-in automated software, and manual correction of artifacts was then performed to obtain precise results.
Microperimetry was performed on all subjects using the MP1 Microperimeter. (version 1.7.0., Nidek Technologies, Gamagori, Japan) in the first visit and the 3-month follow-up after injection. To avoid bias resulting from instrument variability, all examinations were calibrated accurately by the same experienced technician. The following parameters were used: a fixation target consisting of a red ring 2° in diameter; white, monochromatic background at 4asb, stimulus size Goldman III with 200ms projection time; Stimulus light attenuation was set at 10dB, with a threshold 4-2-1 strategy (14, 15).
A macular 10° program is applied, in which 40 stimuli was used: the central 2° area (diameter of 600μm) covered with 8 stimuli; superior, inferior, nasal and temporal areas were each targeted with 4 stimuli (totally 32 stimuli) located 6° (diameter of 1800μm) and 10° (diameter of 3000μm) from the center of the fovea. Mean sensitivity were calculated in central area (diameter=1000μm) and the four quadrants in the inner ring (diameter=3000μm).
All statistical analyses were performed using SPSS version 18.0 for Windows software (SPSS, Chicago, IL, USA). All continuous results are expressed as the mean ± standard deviation. The changes of retina thickness at baseline and after one year follow-up on each region of each layer were compared by paired t-test. Correlations between the OCT parameters changes and injection times were evaluated using Spearman correlation coefficients because the injection times were not normally distributed while all the other parameters were evaluated using Pearson correlation coefficients. We consider r>0.5 as well positively correlated and r= 0.3 to 0.5 as moderate positively correlated. Univariate linear regression and stepwise backward multivariate linear regression analyses were performed to evaluate the contribution of each characteristic to BCVA gain at month 12 compared to baseline, and p< 0.05 was considered significant.