Supplementary table 1. Correlation of endothelial cell density among graft, 3, 6, and 12 months after Descemet s automated stripping endothelial keratoplasty. Graft 3 months 6 months 12 months Graft <0.001 0.091 0.050 (0.999) (0.483) (0.698) 3 months <0.001 0.859 0.723 (0.999) (<0.001) (<0.001) 6 months 0.091 0.859 0.898 (0.483) (<0.001) (<0.001) 12 months 0.050 0.723 0.898 (0.698) (<0.001) (<0.001) Correlation coefficient* (P value) *Spearman correlation analysis
Supplementary table 2. Preoperative aqueous cytokine levels stratified by endothelial cell density of 1,200 cells/mm 2 at 12 months after Descemet s automated stripping endothelial keratoplasty. Control (N=33) ECD 1200 (N=35) ECD < 1200 (N=29) P value P value P value P value* Protein 0.31 ± 0.23 0.94± 0.84 1.31 ± 0.79 0.003 <0.001 0.18 <0.001 IL-1α 49.3 ± 25.5 68.7 ± 48.1 99.5 ± 99.7 0.20 0.17 >0.99 0.032 IL-1β 3.1 ± 10.1 2.5 ± 3.0 7.8 ± 12.9 >0.99 0.17 0.72 0.303 IL-4 20.5 ± 5.8 31.2 ± 17.4 52.9 ± 50.6 0.008 <0.001 0.52 0.005 IL-6 85 ± 399 956 ± 1371 1915 ± 3536 <0.001 <0.001 >0.99 <0.001 IL-8 46.4 ± 146 80.7 ± 115.6 83.4 ± 96.5 0.005 0.012 >0.99 0.002 IL-10 1.9 ± 0.7 3.6 ±2.2 14.7 ± 33.5 <0.001 0.004 >0.99 <0.001 IL-12p70 6.4 ± 0.9 10.9 ± 6.6 15.3 ± 12.9 0.003 <0.001 >0.99 0.001 IL-13 7.1 ± 1.4 9.7 ± 4.7 9.8 ± 5.1 0.045 0.018 >0.99 0.007 IL-17A 4.0 ±2.1 8.3 ± 10.7 11.8 ± 10.3 0.014 <0.001 0.39 0.001 MIP-1α 9.4 ± 2.5 14.1 ± 8.1 16.8 ± 21.7 0.54 >0.99 0.63 0.047 MIP-1β 345 ± 116 298.2 ±283.7 389 ± 626 0.55 0.44 >0.99 0.237 MCP-1 592 ± 485 705.1 ± 668.3 1283 ± 830 0.39 <0.001 0.01 <0.001 TNF-α 76.4 ± 31.2 162 ± 178 114 ± 99 0.006 0.51 0.39 0.001 IFN-α 4.1 ± 0.7 5.3 ± 3.2 5.3 ± 4.2 0.51 0.91 >0.99 0.141 IFN -γ 55.6 ± 11.9 74.8 ± 47.7 132 ± 122 0.47 <0.001 0.02 <0.001 E-Selectin 2149 ± 264 3737 ± 2287 4836 ± 5070 <0.001 <0.001 >0.99 <0.001 P-Selectin 3724 ± 753 8679 ± 7737 13556 ± 17279 <0.001 <0.001 0.28 <0.001 sicam-1 2027 ± 2585 3375 ± 2659 6920 ± 6596 0.046 <0.001 0.13 <0.001 IP10 257 ± 514 174 ± 142 380 ± 458 0.85 0.25 >0.99 0.192 Mean ± SD Protein:(mg/ml), Cytokines:(pg/ml) Mann-Whitney U test, compared ECD 1200 group to control group. Mann-Whitney U test, compared ECD < 1200 group to control group. Mann-Whitney U test, compared ECD < 1200 group to ECD 1200 group. *Trends test ECD: endothelial cell density, IL: interleukin, MIP: macrophage inflammatory protein, MCP: monocyte chemotactic protein, TNF: tumor necrosis factor, GM-CSF: granulocyte-macrophage colony-stimulating factor, IFN: interferon, sicam: soluble intracellular adhesion molecule, IP10: interferon gamma-induced protein 10
Supplementary table 3. Preoperative aqueous protein and cytokine levels stratified by endothelial cell density of 1,000 and 2,000 cells/mm 2 at 12 months after Descemet s automated stripping endothelial keratoplasty. Aqueous Control ECD 2000 1000 ECD < 2000 ECD < 1000 P value P value P value P value P value* P value# P value* cytokine N=33 N=9 N=34 N=21 Protein 0.31 (0.23) 0.91 (0.72) 1.08 (0.88) 1.34 (0.78) - - - - - - <0.001 IL-1α 49.3 (25.5) 80.4 (30.7) 72.3 (51.5) 108.4 (118.2) 0.13 0.25 >0.99 >0.99 0.98 >0.99 0.243 IL-1β 3.1 (10.1) 3.7 (4.5) 3.3 (6.1) 8.91 (13.9) >0.99 >0.99 0.132 >0.99 >0.99 0.55 0.091 IL-4 20.5 (5.8) 38.7 (18.7) 31.2 (17.3) 61.7 (57.4) 0.019 0.011 <0.001 >0.99 >0.99 0.97 <0.001 IL-6 6.4 (0.9) 673 (942) 1170 (1676) 2318 (3996) 0.001 <0.001 <0.001 >0.99 >0.99 >0.99 <0.001 IL-8 19.3 (1.2) 61.4 (82.5) 93.0 (127.9) 85.9 (87.8) 0.63 0.001 0.008 >0.99 >0.99 >0.99 <0.001 IL-10 1.9 (0.7) 3.7 (1.6) 4.5 (4.2) 18.2 (39.1) 0.02 <0.001 0.02 >0.99 >0.99 >0.99 <0.001 IL-12p70 6.4 (0.9) 12.1 (7.3) 10.0 (6.12) 18.2 (14.1) 0.09 0.04 <0.001 >0.99 >0.99 0.13 <0.001 IL-13 7.1 (1.4) 10.1 (2.6) 9.6 (4.8) 10.3 (5.9) 0.17 0.099 0.067 >0.99 >0.99 >0.99 0.004 IL-17A 4.0 (2.1) 6.9 (5.3) 8.4 (10.5) 14.3 (11.4) 0.82 0.009 <0.001 >0.99 0.57 0.64 <0.001 MIP-1α 9.4 (2.5) 15.5 (10.2) 13.1 (7.3) 19.1 (25.2) >0.99 0.627 >0.99 >0.99 >0.99 >0.99 0.366 MIP-1β 345 (116) 353 (297) 323 (286) 400 (721) >0.99 >0.99 0.35 >0.99 >0.99 >0.99 0.058 MCP-1 449 (24) 950 (1182) 734 (581) 1387 (777) >0.99 0.79 <0.001 >0.99 0.26 0.004 <0.001 TNF-α 76.4 (31.2) 163 (122) 153 (169) 120 (122) 0.69 0.006 >0.99 >0.99 >0.99 0.36 0.050 IFN-α 4.1 (0.7) 5.2 (3.3) 5.1 (3.0) 5.7 (4.8) 0.63 >0.99 >0.99 >0.99 >0.99 >0.99 0.318 IFN -γ 55.6 (11.9) 91.0 (48.4) 69.0 (42.7) 156 (136) 0.14 >0.99 <0.001 0.51 >0.99 <0.001 <0.001 E-Selectin 2149 (264) 4964 (2858) 3353 (1988) 5456 (5588) <0.001 <0.001 <0.001 0.67 >0.99 0.27 <0.001 P-Selectin 3724 (752) 11368 (10917) 10328 (14918) 11469 (11088) 0.047 <0.001 <0.001 >0.99 >0.99 0.61 <0.001 sicam-1 2027(2585) 4276 (3248) 3729 (4134) 7761 (6427) 0.13 0.15 <0.001 >0.99 >0.99 0.06 <0.001 IP10 257 (514) 158 (152) 252 (323) 341 (410) >0.99 0.91 0.77 >0.99 >0.99 >0.99 0.168 Mean (SD) Protein:(mg/ml), Cytokines:(pg/ml) Mann-Whitney U test, compared ECD > 2000 group to control group. Mann-Whitney U test, compared 1000 < ECD < 2000 group to control group. Mann-Whitney U test, compared ECD < 1000 group to control group. Mann-Whitney U test, compared ECD > 2000 group to 1000 < ECD < 2000 group *Mann-Whitney U test, compared ECD > 2000 group to ECD < 1000 group #Mann-Whitney U test, compared 1000 < ECD < 2000 group to ECD < 1000 group, *trends test ECD: endothelial cell density, IL: interleukin, MIP: macrophage inflammatory protein, MCP: monocyte chemotactic protein, TNF: tumor necrosis factor, IFN: interferon, sicam: soluble intracellular adhesion molecule, IP10: interferon gamma-induced protein 10
Supplementary table 4. Correlation between preoperative cytokine levels and percentage of endothelial cell density loss. %ECD loss at 6 months %ECD loss at 12months r P value 95% CI r P value 95% CI Protein 0.293 0.053 0.087 to 0.636 0.320 0.041 0.005 to 0.578 IL-1α -0.270 0.066-0.428 to 0.187-0.148 0.321-0.202 to 0.407 IL-1β 0.058 0.706-0.240 to 0.389-0.027 0.858-0.162 to 0.440 IL-4 0.093 0.481-0.259 to 0.287 0.289 0.031 0.021 to 0.519 IL-6-0.007 0.961-0.350 to 0.192 0.047 0.727-0.131 to 0.404 IL-8 0.021 0.874-0.312 to 0.233 0.085 0.528-0.216 to 0.328 IL-10-0.069 0.602-0.312 to 0.233 0.039 0.775 0.028 to 0.529 IL-12p70 0.103 0.446-0.229 to 0.331 0.272 0.047-0.003 to 0.444 IL-13 0.060 0.720-0.270 to 0.411 0.070 0.680-0.257 to 0.424 IL-17A 0.355 0.021 0.049 to 0.601 0.380 0.013 0.087 to 0.619 MIP-1α -0.289 0.067-0.507 to 0.132-0.198 0.215-0.245 to 0.407 MIP-1β -0.250 0.063-0.446 to 0.103-0.141 0.309-0.249 to 0.318 MCP-1 0.303 0.022 0.038 to 0.528 0.470 0.0002 0.232 to 0.656 TNF-α -0.178 0.260-0.411 to 0.241-0.155 0.328-0.490 to 0.136 IFN-α 0.358 0.158-0.440 to 0.227 0.404 0.109-0.308 to 0.358 IFN -γ 0.218 0.049-0.007 to 0.495 0.392 0.0026 0.139 to 0.597 E-Selectin 0.286 0.039 0.006 to 0.524 0.293 0.0352 0.0133 to 0.530 P-Selectin 0.093 0.482-0.201 to 0.333 0.050 0.710-0.249 to 0.297 sicam-1 0.291 0.028 0.06 to 0.519 0.355 0.0068 0.096 to 0.569 IP10-0.075 0.570-0.302 to 0.243-0.023 0.862-0.059 to 0.462 *Spearman s correlation analysis IL: interleukin, MIP: macrophage inflammatory protein, MCP: monocyte chemotactic protein, TNF: tumor necrosis factor, IFN: interferon, sicam: soluble intracellular adhesion molecule, IP10: interferon gamma-induced protein 10
Supplementary table 5. Correlation between preoperative cytokine levels and endothelial cell density in eyes after cataract surgery. %ECD loss at 6 months Cataract surgery ECD at 6 months r P value* r P value* Protein 0.231 0.519 0.171 0.589 IL-1α 0.200 0.557 0.100 0.766 IL-1β 0.073 0.833-0.137 0.677 IL-4 0.018 0.972-0.077 0.811 IL-6 0.330 0.322-0.220 0.498 IL-8 0.169 0.619-0.228 0.489 IL-10 0.136 0.693-0.018 0.967 IL-12p70 0.245 0.468 0.109 0.755 IL-13 0.164 0.634 0.055 0.881 IL-17A 0.005 0.995-0.091 0.781 MIP-1α 0.273-0.221 0.064 0.854 MIP-1β 0.091 0.796 0.273 0.418 MCP-1 0.136 0.694 0.193 0.572 TNF-α 0.264 0.435-0.055 0.881 IFN-α 0.255 0.451-0.164 0.634 IFN -γ 0.110 0.587 0.055 0.881 E-Selectin 0.227 0.503 0.209 0.539 P-Selectin 0.046 0.902 0.193 0.572 sicam-1 0.255 0.451 0.046 0.903 IP10 0.355 0.286 0.073 0.839 *Spearman s correlation analysis IL: interleukin, MIP: macrophage inflammatory protein, MCP: monocyte chemotactic protein, TNF: tumor necrosis factor, IFN: interferon, sicam: soluble intracellular adhesion molecule, IP10: interferon gamma-induced protein 10
Supplementary table 6. Multivariate analysis: history of trabeculectomy was associated with postoperative endothelial cell density. ECD at 3 months ECD at 6 months ECD at 12 months β P value β P value β P value Trabeculectomy -0.352 0.011-0.287 0.028-0.278 0.026 Laser iridotomy 0.038 0.776-0.230 0.076-0.300 0.018 Lens status -0.098 0.478-0.100 0.445-0.216 0.085 Graft size 0.085 0.512 0.141 0.258 0.192 0.105 Preop steroid use -0.062 0.650 0.046 0.731-0.058 0.646 Adjusted R 2 0.114 0.169 0.278 VIF = 1.14 * Lens status, laser iridotomy and history of trabeculectomy were dichotomized as categorical variables for multivariate regression analysis as follows: history of trabeculectomy: absence = 0, presence = 1, laser iridotomy: absence = 0, presence = 1, lens status: phakia = 0, pseudophakia = 1 ECD: endothelial cell density, VIF: variance inflation factor
Supplementary table 7. Correlation between preoperative cytokine levels and percentage of endothelial cell density loss in eyes excluding Fuchs endothelial corneal dystrophy. %ECD loss at 6 months %ECD loss at 12months r P value 95% CI r P value 95% CI Protein 0.227 0.203-0.136 to 0.537 0.258 0.135-0.093 to 0.552 IL-1α -0.303 0.092-0.596 to 0.062-0.207 0.274-0.522 to 0.157 IL-1β 0.034 0.861-0.347 to 0.405 0.062 0.746-0.315 to 0.422 IL-4 0.251 0.123-0.080 to 0.532 0.417 0.010 0.097 to 0.658 IL-6-0.053 0.735-0.369 to 0.270 0.040 0.801-0.276 to 0.348 IL-8-0.028 0.865-0.345 to 0.295 0.081 0.612-0.238 to 0.383 IL-10-0.064 0.696-0.376 to 0.262 0.085 0.594-0.234 to 0.387 IL-12p70 0.265 0.108-0.069 to 0.546 0.375 0.0266 0.037 to 0.636 IL-13 0.255 0.108-0.159 to 0.592 0.309 0.127-0.101 to 0.629 IL-17A 0.606 0.0008 0.282 to 0.805 0.582 0.0014 0.249 to 0.792 MIP-1α -0.350 0.068-0.646 to 0.038-0.311 0.109-0.620 to 0.082 MIP-1β -0.288 0.075-0.560 to 0.039-0.108-0.503-0.410 to 0.216 MCP-1 0.424 0.0079 0.122 to 0.661 0.616 <0.0001 0.360 to 0.785 TNF-α -0.109 0.567-0.460 to 0.272-0.198 0.295-0.529 to 0.196 IFN-α -0.289 0.143-0.611 to 0.113-0.120 0.552-0.487 to 0.284 IFN -γ 0.479 0.0023 0.179 to 0.698 0.623 <0.0001 0.370 to 0.790 E-Selectin 0.564 0.0003 0.280 to 0.758 0.585 0.0002 0.309 to 0.771 P-Selectin 0.122 0.452-0.206 to 0.426 0.178 0.259-0.142 to 0.465 sicam-1 0.356 0.028 0.032 to 0.613 0.506 0.0012 0.213 to 0.716 IP10-0.270 0.093-0.543 to 0.053-0.127 0.423-0.423 to 0.193 *Spearman s correlation analysis IL: interleukin, MIP: macrophage inflammatory protein, MCP: monocyte chemotactic protein, TNF: tumor necrosis factor, GM-CSF: granulocyte-macrophage colony-stimulating factor, IFN: interferon, sicam: soluble intracellular adhesion molecule, IP10: interferon gamma-induced protein 10
Supplementary table 7. Preoperative demographics of DSAEK patients. Solitary Combined P value DSAEK DSAEK # Eyes (n) 43 21 - Sex, n (%) Male 14 (32.6) 7 (33.3) Female 29 (67.4) 14 (66.7) 0.95* Age (y) 74.8 ± 9.5 73.1 ± 9.1 0.38 BCVA (logmar) 1.37 ± 0.60 1.13 ± 0.96 0.08 IOP (mmhg) 13.1 ± 4.2 10.8 ± 3.4 0.01 Axial length (mm) 23.75 ± 2.33 23.40 ± 1.52 0.76 ECD (cells/mm 2 ) NA NA - CCT (μm) 754 ± 122 716 ± 84 0.26 DM, n (%) 6 (14) 4 (19) 0.59* Presence of glaucoma, n (%) 14 (33) 3 (14) 0.12* Number of Previous intraocular surgeries (n) 2.0 ± 1.5 0.5 ± 0.8 <0.001 Mean ± SD DSAEK: Descemet s automated stripping endothelial keratoplasty, SD: standard deviation, BCVA: best-corrected visual acuity, logmar: logarithm of minimal angle resolution, IOP: intraocular pressure, AL: axial length, ECD: endothelial cell density, CCT: central corneal thickness, DM: Diabetes mellitus, IOL: intraocular lens, NA: not available *: Chi-square test # : DSAEK combined with cataract surgery