Quantitative characteristics of sickle cell retinopathy in optical coherence tomography angiography

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1 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 74 Quatitative characteristics of sickle cell retiopathy i optical coherece tomography agiography MINHAJ ALAM, DAMBER THAPA, JENNIFER I. LIM,2 DINGCAI CAO,2,3 AND XINCHENG YAO,2,4 Departmet of Bioegieerig, Uiversity of Illiois at Chicago, Chicago, IL 60607, USA Departmet of Ophthalmology ad Visual Scieces, Uiversity of Illiois at Chicago, Chicago, IL 6062, USA 3 dcao98@uic.edu 4 xcy@uic.edu 2 Abstract: Early detectio is a essetial step for effective itervetio of sickle cell retiopathy (SCR). Emergig optical coherece tomography agiography (OCTA) provides excellet three-dimesioal (3D) resolutio to eable label-free, oivasive visualizatio of retial vascular structures, promisig improved sesitivity i detectig SCR. However, quatitative aalysis of SCR characteristics i OCTA images is yet to be established. I this study, we coducted comprehesive aalysis of six OCTA parameters, icludig blood vessel tortuosity, vessel diameter, vessel perimeter idex (VPI), area of foveal avascular zoe (FAZ), cotour irregularity of FAZ ad parafoveal avascular desity. Compared to traditioal retial thickess aalysis, five of these six OCTA parameters show improved sesitivity for SCR detectio tha retial thickess. It is observed that the most sesitive parameters were the cotour irregularity of FAZ i the superficial layer ad avascular desity i temporal regios, while the area of FAZ, tortuosity ad mea diameter of the vessel were moderately sesitive. 207 Optical Society of America OCIS codes: ( ) Ophthalmology; ( ) Medical ad biological imagig; ( ) Optical coherece tomography; ( ) Optical diagostics for medicie; ( ) Visio system europhysiology; ( ) Physiology; ( ) Visio system - oivasive assessmet. Refereces ad liks. K. L. Hassell, Populatio estimates of sickle cell disease i the U.S, Am. J. Prev. Med. 38(4 Suppl), S52 S52 (200). 2. Sickcel Cell Guidelie Pael, Sickle Cell Disease: Screeig, Diagosis, Maagemet, ad Couselig i Newbors ad Ifats, AHCPR publicatio o (Agecy for Health Care Policy ad Research, US Public Health Service, Rockville, MD: 993). 3. M. F. Goldberg, Classificatio ad pathogeesis of proliferative sickle retiopathy, Am. J. Ophthalmol. 7(3), (97). 4. J. I. Lim, Ophthalmic maifestatios of sickle cell disease: update of the latest fidigs, Curr. Opi. Ophthalmol. 23(6), (202). 5. Q. V. Hoag, F. Y. Chau, M. Shahidi, ad J. I. Lim, Cetral macular splayig ad outer retial thiig i asymptomatic sickle cell patiets by spectral-domai optical coherece tomography, Am. J. Ophthalmol. 5(6), (20). 6. G. K. Asdouria, K. C. Nagpal, B. Busse, M. Goldbaum, D. Patriakos, M. F. Rabb, ad M. F. Goldberg, Macular ad perimacular vascular remodellig sicklig haemoglobiopathies, Br. J. Ophthalmol. 60(6), (976). 7. W. Mivielle, V. Caillaux, S. Y. Cohe, F. Chasset, O. Zambrowski, A. Miere, ad E. H. Souied, Macular microagiopathy i sickle cell disease usig optical coherece tomography agiography, Am. J. Ophthalmol. 64, (206). 8. R. K. Wag, S. L. Jacques, Z. Ma, S. Hurst, S. R. Haso, ad A. Gruber, Three dimesioal optical agiography, Opt. Express 5(7), (2007). 9. E. Moult, W. Choi, N. K. Waheed, M. Adhi, B. Lee, C. D. Lu, V. Jayarama, B. Potsaid, P. J. Rosefeld, J. S. Duker, ad J. G. Fujimoto, Ultrahigh-speed swept-source OCT agiography i exudative AMD, Ophthalmic Surg. Lasers Imagig Retia 45(6), (204). 0. T. S. Hwag, Y. Jia, S. S. Gao, S. T. Bailey, A. K. Lauer, C. J. Flaxel, D. J. Wilso, ad D. Huag, Optical coherece tomography agiography features of diabetic retiopathy, Retia 35(), (205). # Joural Received Nov 206; revised 0 Ja 207; accepted 6 Ja 207; published 23 Feb 207

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3 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 743 tool called a sickle. Whe sickle cells travel through small blood vessels, they ca get stuck ad occlude blood flow ad thus prevet oxyge from reachig vital orgas. Cosequetly, patiets with SCD suffer from microvascular occlusios i various parts of their body, icludig the retia. Sickle cell retiopathy (SCR) is oe of the major ocular maifestatios of SCD. The uderlyig systemic ad ocular maifestatios of SCD are a result of vasoocclusive ischemia due to the blockig of blood vessels by sickle-shaped erythrocytes. SCR typically ivolves peripheral retial vessels. At this time, the cliical stagig system is of limited value i predictig progressio of SCR. Further, stadard fudus-based cliical examiatio typically shows ormal to miimal fidigs i the macular area i sickle cell patiets [3]. Although patiets are asymptomatic, may adult sickle cell patiets do harbor fidigs of SCR that precede the cliical detectio of a foveal depressio sig [4], such as macular thiig measured by optical coherece tomography (OCT) [5] or vascular abormalities i the macular regio based o fluorescei agiography (FA) [6]. However, the existig techiques are limited i detectig subcliical sigs of SCR. For istace, FA requires dye ijectio ad caot detect abormalities i ~50% of eyes i sickle cell patiets, compared with recetly emerged OCT Agiography (OCTA) [7]. New techique is desirable for detectig subcliical sig of SCR. OCTA has bee used for quatitative assessmet of retial vascular structures [8 2]. OCTA allows visualizatio of multiple retial layers with high resolutio ad therefore is more sesitive tha traditioal FA i detectig SCR [7]. Thus, OCTA may be able to further the classificatio of SCR by icludig data with progostic value. OCTA is curretly available commercially for cliical use. However, quatitative aalysis of SCR characteristics i OCTA images is yet to be established. I this study, we coducted comparative aalysis of six OCTA parameters, icludig blood vessel tortuosity, vessel diameter, vessel perimeter idex (VPI), area of foveal avascular zoe (FAZ), cotour irregularity of FAZ ad parafoveal avascular desity of cotrol ad SCR groups. The goal was to assess how the parameters correlated with retial thickess ad how sesitive the parameters could detect SCR, ad thus to establish a metric for quatitative OCTA assessmet of SCR. 2. Materials ad methods 2. Data acquisitio This study was approved by the Istitutioal Review Board of the Uiversity of Illiois at Chicago ad was i compliace with the ethical stadards stated i the Declaratio of Helsiki. The SCD patiets were recruited from Uiversity of Illiois at Chicago (UIC) Retial Cliic. The majority of patiets (N = 4, 78%) had Stage II of retiopathy with the remaiig as Stage III (N = 4, 22%). The quatitative study was based o OCTA images of 8 SCD patiets (7 males ad females; 8 Africa Americas) ad 3 cotrol subjects (2 males, female, 3 Africa Americas). The mea age was 40 years (rage 24 to 64) for the patiets ad 37 years (rage 25 to 7) for the cotrol. OCTA images of both eyes (OS ad OD) were aalyzed i this study, so the database cosisted of 36 SCR eyes ad 26 cotrol eyes. The subjects of the cotrol group were chose based o their previous ocular history, absece of ay systemic diseases, or ay visual symptoms; a ormal- appearig retia o cliical examiatio; ad a ormal reflectace OCT of the macula. SD-OCT data was acquired usig a ANGIOVUE SD-OCT agiography system (OPTOVUE, Fremot, CA, USA), with a 70-KHz A-sca rate. OCTA images were costructed usig split-spectrum amplitude-decorrelatio agiography (SSADA) algorithm with itegrated motio correctio algorithm [3]. The axial resolutio is 5 μm while lateral resolutio is 5 μm. The scaig protocol i this system provided a field of view (FOV) of 3 mm 3 mm, 6 mm 6 mm or 8 mm 8 mm. For our calculatio, we used oly 3 mm 3 mm portios from the OCTA images. OCT agiography images were exported usig the software ReVue (Optovue, Fremot, CA). The ReVue was used to segmet superficial ad

4 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 744 deep ier retial vascular plexuses. It was also used to measure the retial thickess from the OCT B-scas, the thickess was measured from the outermost layer of fovea to the retial pigmet epithelium (RPE) layer. Custom-developed MATLAB (R205b, MathWorks, Ic.) procedures with graphical user iterface (GUI) were used for further image aalysis ad quatitative compariso. 2.2 Quatitative parameters for OCTA aalysis Six OCTA parameters, icludig blood vessel tortuosity, vessel diameter, VPI, area of FAZ, cotour irregularity of FAZ ad parafoveal avascular desity, were used to aalyze ad quatify the OCTA images from SCD patiets ad compare the patiet data with cotrol subjects usig Studet t-tests. Effect sizes (Cohe s d) were also calculated to quatify the differece i the outcome betwee the groups ormalized by the pooled stadard deviatios. Pearso correlatios betwee retial thickess ad the OCTA parameters were computed separately for the cotrols ad SCD patiets. Fially, caoical discrimiat aalysis was coducted to assess which measuremets (retial thickess, OCTA parameters) were most sesitive to differetiate SCD patiets vs. cotrols. For the discrimiat aalysis, the variables with small caoical loadigs (correlatios betwee the observed variables ad the discrimiat fuctio) were sequetially removed util all remaiig variables had caoical loadigs > 0.3 (a critical loadig value for a variable cosidered to be importat). The ratioale of each of these six OCTA parameters is briefly summarized i the followig sessios Blood vessel tortuosity Retial blood vessels i SCD patiets are kow to be more tortuous tha those i ormal subjects [4 6], due to sickle cell aemia. Therefore, quatitative aalysis of blood vessel tortuosity ca be valuable. I this study, quatitative aalysis of the tortuosity is coducted for large blood vessel braches i the superficial layer. As the sickle cells affects the retial vasculature, the tortuosity is visually promiet i large vessels ad they have statistically more reliable tortuosity measuremet idex [7].The first step is to recostruct e-face OCTA image of the superficial layer of retia (Fig. (a)). The secod step is to segmet the large blood vessels usig global thresholdig [2], morphological fuctios (Fig. (b)) ad fractal dimesio classificatio [8]. This biary vasculature map was the skeletoized. The skeletoizatio process removes pixels o the boudaries of vessels but does ot allow objects to break apart [9, 20]. The remaiig pixels make up the image skeleto. After this, each brach of the blood capillaries was idetified with two edpoits (poits A ad B i Fig. (c)) so that the geodesic ad Euclidia distaces for each brach could be calculated. The tortuosity of a sigle brach is defied by the distace metric [2 25] which is the ratio of geodesic distace ad Euclidea distace betwee the two edpoits. For two poits A(x, y) ad B(x2, y2) i a two dimesioal plae as show i Fig., Euclidea distace is calculated usig the followig equatio [25], Euclidea distace = ( x x ) + ( y y ). () If we defie each of the segmeted braches with [x(t),y(t)] o the iterval [t 0, t ], the geodesic distace betwee the edpoits i.e. A ad B ca be calculated with the followig equatio [25], () dy () t 2 2 t dx t Geodesic distace = + d. t (2) dt dt t0 The tortuosity of each brach of blood vessels was calculated ad the average tortuosity of the image was measured [25],

5 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 745 Geodesic distacebetweetwo edpoits of a vessel brachi Tortuosity =. i= Euclidea distacebetweetwo edpoits of a vessel brachi (3) where i is the ith brach ad is the umber of brach Mea diameter of blood vessel As the sickle cells affect the blood flow iside the vessels ad thus cause structural chage i the vasculature, the mea diameter of blood vessels is a importat parameter to aalyze i OCTA images. Large blood vessels from superficial e-face OCTA image were measured to quatify this parameter. The large vessel area was calculated from the vessel map (Fig. (b)) ad legth was calculated from the skeleto map (Fig. (c)). Mea vessel diameter was the calculated as the ratio of vessel area ad legth usig the followig equatio [2], Meavessel width i=, j= i, j = = = ( ) (, ) B i, j. S i j where B (i,j) represets the pixels occupied by the vessels ad S (i,j) represets the pixels occupied by the vessel skeleto (i, j represet the row ad colum positios of each pixel of the image). With this parameter, localized vascular dilatio would be easily idetified ad it could serve as a marker for vascular abormalities Vessel perimeter idex (VPI) Large blood vessels i the e-face OCTA image were used to measure this parameter. From the biary vasculature map, a vessel perimeter map (Fig. 3(a)) was obtaied by detectig the edge of vessels ad deletig the pixels that were ot close to the edge of the vessels. The VPI was calculated as the ratio of perimeter pixel area ad total image area usig the followig equatio [2], Vessel perimeter idex i=, j= = i =, j= ( ) P i, j. I i j (, ) where P (i,j) represets the pixels withi the vessel perimeters (white pixels o Fig. 3(a)) ad I (i,j) represets all the pixels i the vessel perimeter map. This parameter is a good marker of overall chage of vessel legth i OCTA images Area of FAZ Sice the SC disease directly affects the vessel texture with tortuosity ad dilatio, it is also iterestig to compare the chage i foveal avascular area for cotrol subjects ad SCD patiets. The FAZ cotour was semi-automatically demarcated ad the FAZ was segmeted from the OCTA images (Fig. 4(a), 4(d)) usig a active cotour model [26, 27], where the seed poit was maually placed at the ceter of the fovea. The pixel size of the OCTA image is a kow parameter ad from this iformatio, the area of the avascular regio was calculated usig the followig equatio [2], 2 ( ) (, ) FAZ = ( Area of sigle pixel i µm A i j. (6) i=, j= where A (i,j) represets the pixels occupied by the segmeted avascular regio. (4) (5)

6 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS Cotour irregularity of FAZ A cotour irregularity parameter ca be used to express border irregularity ad is also a excellet descriptor of spiculatio, i.e, spiked ature of the cotour [28]. We illustrate the effects of icreasig border roughess usig cotours of avascular area (Fig. 4(c), 4(f)). The bigger the value of cotour irregularity, the more irregular ad spiked the cotour of the avascular regio is. Cotour irregularity parameter was calculated with the followig equatio [28], Cotour irregularity i=, j= = i =, j= ( ) O i, j. R i j (, ) where O (i,j) represets the pixels occupied by the FAZ cotour ad R (i,j) represets the pixels occupied by the perimeter of the referece circle. The cotour of the origial avascular regio was measured; a circle of the same area as the avascular regio was used as a referece Desity of parafoveal avascular regio I order to quatify the desity of parafoveal avascular regio, fractal dimesio (FD) aalysis of both the superficial ad deep layers were coducted [8]. FD has bee cosidered as a potetial biomarker for retia-based disease detectio [29 3]. Fractal is o-euclidea structures that show self-similarity at differet scales. The FD of a structure provides a measure of its texture complexity [32]. The retial blood vessels ad capillaries are complex ad rarely have a exact Euclidea shape. Therefore, they ca be precisely described by fractal aalysis. May retial diseases such as SC disease ivolve vessel abormalities, icludig drop out zoes i betwee vascular structures. This requires detailed aalysis of the retial vasculature to uderstad their role i disease pathophysiology. I our case, local fractal dimesio (LFD) aalysis which idetifies local variatios i the vascular etwork was calculated from the OCTA images by usig a movig widow of size ( 2w+ ) ( 2w+ ) usig the followig equatio [33], ( ) ( ) q i, j = LFD[p i+ k,j+ k ; w < k < w]. (8) where p( i, j ) is the itesity OCTA image, (, ) (7) q i j is the fractal dimesio of the itesity image ad (i,j) correspods to the locatio of each pixel i the image. Local FD was calculated usig widow sizes (i pixels) of 3 3, 5 5, 7 7, 9 9 ad (w =,2,3,4.5). The FD varies across the image; it is higher i larger vessels compared to that i smaller vessel or o-vascular regio [8]. Therefore, the LFD was ormalized ad plotted as a cotour plot (Fig. 5(a), 5(b)). The ormalized LFD with a value close to idicates large vessels ad a value close to 0 idicates o-vessel regios, so the regios were classified as follows: the blood vessels ad capillaries lie betwee a value of 0.7 ad ; the ovascular regios lie betwee a value of 0.0 ad 0.3 ad the smaller gaps betwee vessels lie betwee a value of 0.3 ad 0.7, we defie this area as grey zoe. The desity calculatio is expressed i percetage by takig the ratio of the total pixels with correspodig FD values (0 to 0.3 for o-vascular regio, 0.7 to for vessel ad 0.3 to 0.7 for grey zoe) to the total pixels i the aalyzed widow [8]. Here, vessel desities were calculated i three circular parafoveal regios of diameter mm, 2 mm ad 3 mm as show i Fig. 5(a) ad four parafoveal sectors, amely, asal (N), superior (S), temporal (T), ad iferior (I) of a circular zoe of diameter 3 mm as show i Fig. 5(b).

7 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS Results I Sectio 3. we report the OCTA measuremets ad their respective variatio with SCD. I Sectio 3.2, we provide comparative iformatio of retial thickess ad discrimiat aalysis. Amog 36 SCR OCTA images i total, OD ad 3 OS OCTA images were excluded due to severe image distortios. 3.. OCTA parameters 3.. Blood vessel tortuosity Figure illustrates represetative OCTA image (Fig. (a)), segmeted blood vessel map (Fig. (b)) ad skeletoized blood vessel map (Fig. (c)). The retial vasculature i SCD patiets becomes more complex as the vessels become more tortuous ad twisted. As show i Fig. (d), SCD eyes had much higher tortuosity tha the cotrol eyes (48.07% vs. 3.52%, p < 0.00, Cohe s d = 3.69). O average, a tortuosity icrease of 6.56% was observed i SCD patiets compared to cotrol subjects. Fig.. Measurig tortuosity of OCTA images. (a) OCTA raw image, (b) Segmeted large blood vessel map, (c) Skeletoized blood vessels braches with idetified edpoits (for a radom vessel brach, A ad B edpoits are show with red dots), (d) Compariso of tortuosity i cotrol ad SC patiets (error bars are stadard deviatios) Mea diameter of blood vessel Figure 2 illustrates the mea diameters of large blood vessels of superficial OCTA images. It was observed that the averaged diameter of blood vessels of cotrol subjects ad SCD patiets were μm ad μm, respectively. The mea diameter icreased about 29.4% i SCD patiets, reflectig a sigificat dilatio i the large blood vessels of SCD patiets (p<0.0, Cohe s d = 3.8). Fig. 2. Compariso of mea diameter of blood vessels i cotrol ad SC patiets (superficial layer).

8 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS Vessel perimeter idex (VPI) Figure 3(a) illustrates the biary vessel perimeter map obtaied from the superficial raw OCTA image (Fig. (a)). VPI provides a good estimatio of the chage i vessel legth for SCD patiets. As show i Fig. 3(b), the VPI i cotrol subjects is 0.8% ad it is 8.3% i SCD patiets, so there is a decrease of 2.49% i VPI for SCD patiets (p<0.05, Cohe s d = 2.4). Fig. 3. (a) Vessel perimeter map, (b) Compariso of VPI i cotrol ad SCD patiets (superficial layer) Area of FAZ Figure 4(a) ad 4(d) illustrates the demarcatio of FAZ i raw OCTA images, i cotrol ad SCD patiets respectively with the biary FAZ segmetatio (Fig. 4(b) ad 4(e)) ad cotour map (Fig. 4(c) ad 4(f)). It was observed that the average area of avascular regio also icreases i SCD patiets (52% i deep ad 53% i superficial layer). This shows that, for SCD patiets FAZ icreases, which is possibly a result of rapid drop out of retial vessels ear foveal area (p<0.00, Cohe s d = 4.5). The compariso is illustrated i Fig. 4(g) Cotour irregularity of FAZ From the segmeted FAZ (Fig. 4(b), 4(e)) we also measured the cotour of the regio (Fig. 4(c), 4(f)). As there are complex vascular structure ad vessel abormalities, the cotour becomes more irregular i SCD patiets rather tha a smooth shape i cotrol subjects. The cotour irregularity idex for cotrol subjects are.0 ad. for deep ad superficial layers respectively.

9 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 749 Fig. 4. (a) OCTA image with demarcatio for ormal eye, (b) Segmeted Avascular regio for ormal eye (c) FAZ cotour for ormal eye, (d) OCTA image with demarcatio for diseased eye, (e) Segmeted avascular regio for diseased eye, (f) FAZ cotour for diseased eye, (g) Compariso of area of FAZ i cotrol ad SCD patiets for deep ad superficial layers, (h) Compariso of area of FAZ i cotrol ad SCD patiets for deep ad superficial layers. Our results show that the cotour of FAZ from cotrol subjects has 0-% deviatio from a ideal circular cotour whereas the cotour from the SCD patiets has aroud 46-47% deviatio (compariso show i Fig. 4(h)). The irregularity ad spiculatio icrease by almost 36% i case of SCD patiets for both deep ad superficial layers. This clearly idicates the feasibility of this parameter (cotour irregularity) as a biomarker of SC disease (p<0.00, Cohe s d = 4.52) Desity of parafoveal avascular regio Figure 5 illustrates the cotour maps with ormalized values of local fractal dimesio correspodig to OCTA raw images; it also shows the differet regios or zoes of the image where FD aalysis was coducted (Fig. 5(a), 5(b)). A detailed vasculature desity aalysis was coducted with the OCTA database. SC disease ca lead to vessel drop out zoes i the vascular structure of retia, so we measured the desity of o-vascular regio, vessels, itermediate gaps (grey zoes) ad compared the chages i desity i defiite zoes. The desity compariso was doe i three circular parafoveal regios of diameter mm, 2 mm

10 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 750 ad 3 mm (Fig. 5(a)) ad four parafoveal sectors, amely, asal (N), superior (S), temporal (T), ad iferior (I) of a circular zoe of diameter 3 mm (Fig. 5(b)). The avascular regio icreased sigificatly i SCD patiets (p<0.0, Cohe s d = 3.24), the vessel desity ad grey zoe desity decreased to compesate for the icrease of o-vascular regios. Fig. 5. Cotour maps created with ormalized values of local fractal dimesio. (a) Circular zoes of diameter, 2 ad 3mm, (b) Nasal, Superior, Temporal ad Iferior regios. FD aalysis was coducted o the differet regios specified i a ad b. Table. Vascular desity chages i differet zoes Vascular desity chages i differet zoes Superficial Layer Deep Layer Avascular regio Grey Zoe Vessel desity Avascular regio Grey Zoe Vessel desity Circle 8.37% * 5.72% * 2.65% * 5.82% * 2.64% * 3.8% * Circle % ** 4.43% ** 8.27% ** 9.66% ** 3.53% ** 6.2% ** Circle % ** 4.83% ** 8.42% ** 3.85% ** 4.2% ** 9.64% ** N 2.60% ** 4.25% ** 8.35% ** 4.40% ** 4.53% ** 9.87% ** S 2.55% * 2.98% * 9.57% * 4.45% * 5.57% * 8.88% * T 4.34% *** 3.45% *** 0.92% *** 7.26% *** 6.8% *** 0.46% *** I 2.35% * 2.57% * 9.78% * 3.44% * 4.77% * 8.67% * A summary of the vasculature desity aalysis of FD cotour map is show i the Table ad illustrated i Fig. 6 where we ca observe the icrease of avascular regio desity for each specific zoe ad the decrease of grey zoe ad large vessel desities. For each sectio i Table the sigificace of the t-test is marked as: * for p < 0.05; ** p < 0.0; *** p < 0.00.

11 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 75 Fig. 6. Compariso of vascular desity i differet sectios of the OCTA image i cotrol ad patiet eyes. Cotrol eye: (a) Avascular regio desity, (b) Vessel desity, (c) Grey zoe desity; Patiet eye: (d) Avascular regio desity, (e) Vessel desity, (f) Grey zoe desity. We have also summarized the results for the first five parameters i Table 2. It illustrates the effect of SCD for the parameters ad their sesitivity to it. By observig the P values from the t-test results we get the idicatio whether the chage due to SCD is sigificat or ot. Table 2. Quatitative compariso of parameters Parameters Average Tortuosity Average Diameter of Blood Vessels (μm) Vessel Perimeter Idex Area of FAZ (mm 2 ) FAZ Cotour Irregularity Retial Layers Superficial Superficial Superficial Deep Superficial Deep Superficial Cotrol Patiets Chage i parameters 6.56% 29.40% 2.49% 52% 53% 36% 36% P value <0.00 <0.0 <0.05 <0.00 <0.00 Cohe s d Retial thickess ad discrimiat aalysis Retial thickess was sigificatly lower i SCD patiets tha cotrols (93.6 ± 5.3 um vs ± 6.44 um, p < 0.00, Cohe s d = 3.68). Noe of the OCTA parameters were sigificatly correlated with the retial thickess either i cotrols or SCD patiets except for the area of FAZ i Circle of the superficial layer i SCD patiets (r = 0.73, p < 0.00). This correlatio aalysis suggested that OCTA parameters provided additioal iformatio of retial health tha retial thickess.

12 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS 752 Caoical discrimiat aalysis usig retial thickess ad the OCTA parameters idicated most of the OCTA parameters had higher caoical loadigs tha retial thickess. The most sesitive OCTA parameters with caoical loadigs > 0.3 icluded: ) cotour irregularity of FAZ i superficial layer, 2) avascular area of Circle 2 i superficial layer, 3) avascular area of the temporal regio i superficial layer, ad 4) avascular area of the iferior regio i deep layer. These four variables could idividually or joitly differetiate the SCD patiets vs. cotrols with 00% correct rate. 4. Discussios Six OCTA parameters, i.e., blood vessel tortuosity, diameter, VPI, area of FAZ, cotour irregularity of FAZ ad parafoveal avascular desity were developed for quatitative assessmet of OCTA images. 36 SCR ad 26 ormal OCTA images were used for comparative aalysis. Amog the 36 SCR OCTA images, OD ad 3 OS OCTA images were excluded due to severe image distortios. Pathological chage of the retia, eye motio ad eface OCTA projectio artifacts were the mai reasos behid the distortio. ReVue software utilizes SSADA algorithm for OCTA costructio, with itegrated motio correctio algorithm. Potetial icorporatio of removal algorithm of projectio artifacts [34, 35] may further improve the OCTA image quality. A Pearso correlatio aalysis was coducted to test the relatioship betwee traditioal retial thickess measuremet ad each of these six OCTA parameters. The aalysis revealed that most of OCTA parameters are ot sigificatly correlated with the retial thickess except for the area of FAZ i Circle ( mm diameter or 0.5 mm radius from the ceter) of the superficial layer. This suggests that the OCTA may provide additioal iformatio o disease associated vasculature chage tha retial thickess oly iformatio from traditioal OCT. Morphological distortio of retial blood vessels i SCD patiets occurs due to sickle cell aemia [4 6]. Quatitative aalysis of blood vessel tortuosity revealed 9.07% tortuosity icremet i SCD group, compared to that i cotrol group. A 29.40% icremet of blood vessel diameter was observed i SCD group. A 2.49% decrease was observed for the VPI of the SCD group, which is cosistet to the observed icremet of blood vessel diameter. FAZ was cosistetly elarged ad cotour irregularity was icreased i SCD patiets. The FAZ cotour irregularity is closely related to tortuosity icrease which makes the retial vessels more irregular ad spiked i shape. I cotrary, retial blood vessel desities i both superficial ad deep layers were decreased i SCD retias, compared to ormal oes. It is kow that proliferative SCD affects the peripheral retial vasculature, ad its maifestatios iclude capillary dropout, arteriolar veular aastomoses, developmet of retial eovascularizatio ad pigmetary chages. Our quatitative aalysis of retial vasculature with FD aalysis cofirms this effect of SCD o retial vessels. Aother importat aspect of this study was to test the sesitivity of each OCTA parameter for detectig SCR. The caoical discrimiat aalysis showed that OCTA parameters were more sesitive tha retial thickess ad the most sesitive OCTA parameter was cotour irregularity of FAZ i superficial layer, avascular desity i circle 2 of superficial layer, avascular desity i temporal regio i superficial layer ad avascular desity i the iferior regio i deep layer. These variables could correctly differetiate the SCD patiets from the cotrol subjects. Accordig to the mea calculatio of the parameter (supported by t-test ad Cohe s d idex), it was cosistetly observed that the most sesitive parameter was the area of FAZ (about 52-53% chage i SCD patiets) ad cotour irregularity (both superficial ad deep layers) (about 36% chage) (Table 2). Tortuosity, mea diameter of the vessels ad avascular desity i temporal regios are moderately sesitive i SCD patiets (about 4-29% chage). It ca be see that the cotour irregularity i superficial layer ad avascular desity i temporal regios are the most sesitive parameters accordig to both discrimiat aalysis ad t-test results obtaied.

13 Vol. 8, No. 3 Mar 207 BIOMEDICAL OPTICS EXPRESS Coclusios Six parameters have bee used for quatitative assessmet of OCTA images. Comparative aalysis of cotrol ad SCD groups reveals statistically sigificat differeces for all of these six parameters. It is observed that the most sesitive parameters were the cotour irregularity i superficial layer ad avascular desity i temporal regios while the area of FAZ, tortuosity ad mea diameter of the vessel were moderately sesitive. It is cofirmed that the parafoveal o-vascular regio desity icreases as there are vessel drop outs due to SCD. As the ovascular regio icreases, the grey zoe ad vessel desity decreases. The study establishes use of the parameters as bio-marker for potetial SCR diagoses ad provides a metric for quatifyig chages i retial vasculature i SCD patiets. Fudig This research was supported i part by NIH grats R0 EY023522, R0 EY024628, P30 EY00792; by NSF grat CBET ; by Richard ad Loa Hill edowmet; by urestricted grat from Research to Prevet Blidess; by Mario H. Scheck Chair edowmet. Ackowledgmet The authors thak Mr. Mark Jaowicz for his help o OCTA data acquisitio.