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Aqueous humor cytology of pseudophakic bullous keratopathy
Aqueous humor cytology of pseudophakic bullous keratopathy Audrey W. Tuberville, M.D. William M. Poston, M.D. Thomas O. Wood, M.D. Memphis, Tennessee
ABSTRACT The long-term effects of the interaction between an IOL and the immune system of the implanted patient are not yet clear. This study presents the results of aqueous humor (AH) cytological analysis of 51 patients undergoing intraocular surgery for cataract, and aphakic (ABK) or pseudophakic bullous keratopathy (PBK). Cataract AH was acellular (N 8). The mean ABK AH cell count was 8.5 ± 6.4 cells per high power field (hpi) of cytologically filtered specimens; mean PBK cell count was 30.2 ± 13.0. If inflamed eyes are defined as those with more than one cell/hpf, a statistically significant greater number of PBK eyes than ABK eyes were inflamed (93% versus 44%, P < 0.003 chi square). More than 99% of the nucleated cells were mononuclear histiocytes. One PBK patient had many multinucleated giant cells in the AH.
=
Key Words: aqueous humor cytology, implant associated uveitis, mononuclear histiocyte, multinucleated foreign-body giant cells, pseudophakic bullous keratopathy
Many modifications in intraocular lens (IOL) design have been made in the last decade to achieve a style that combines good visual rehabilitation, simple insertion techniques, and few postoperative complications. Most technical problems during the operative and immediate postoperative period have been addressed. The long-term effects of the interaction between the implant and the patient's immune system are not yet entirely clear. Implant surgeons have previously reported that a few individuals undergoing IOL surgery may develop a long-term uveitis with the attendant serious sequelae of intraocular inflammation. I -3 Experimental evidence has shown that certain IOL components, namely the nylon and polypropylene loops, are capable of activating the serum complement system in vitro, thereby generating substances that could stimulate inflammation within the eye. 4 The
clinical importance of this is presently under investigation. Although a few histopathological reports of autopsy or enucleated eyes that contain implants are in the literature,5,6 little has been published concerning the ocular histopathology or the pathophysiology of implant-associated uveitis. Few opportunities to reenter the implanted human eye to collect specimens for inspection exist. This study presents the results of aqueous humor cytological analysis of 51 patients undergoing intraocular surgery for cataract and aphakic or pseudophakic bullous keratopathy. MATERIALS AND METHODS Aqueous humor (AH) of 51 eyes undergoing intraocular surgery was examined to determine cell numbers and morphology. Specimens from eight eyes
From the Department of Ophthalmology, University of Tennessee Center for the Health Sciences, Memphis, Tennessee, and the Department of Pathology, Baptist Memorial Hospital, Memphis, Tennessee. Supported in part by an Earl Wilson Award of Fight for Sight, Inc., New York, and by NEI grant EY04086, National Institutes of Health, Bethesda, Maryland. Presented in part at the U.S. Intraocular Lens Symposium, New Orleans, Louisiana, March 1983. Reprint requests to Audrey W. Tuberville, M.D., 956 Court Avenue, Room 2D-29, Memphis, Tennessee 38163. AM INTRA-OCULAR IMPLANT SOC
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with cataract, 16 eyes with aphakic bullous keratopathy (ABK), and 27 eyes with pseudophakic bullous keratopathy (PBK) were examined. Lens styles ofPBK patients were iris or anterior chamber supported. All operative procedures were performed by one surgeon. Aqueous humor from each patient was collected in the following manner: After retrobulbar anesthesia and digital massage, entrance into the anterior chamber was made with a 27-gauge needle on a plastic tuberculin syringe. About 100 Jotl of AH was aspirated; air (0.5 cc) was drawn into the syringe and the specimen was immediately placed on crushed ice. Within one hour, the specimen was centrifuged at 800 RPMs for three minutes in a microfilter apparatus (Schleicher and Schuell, Inc., Keene, New Hampshire) containing a 9-mm, 5.0 Jot pore size nitrocellulose filter. The filter was immediately fixed in 95% ethanol to avoid drying artifacts. Following staining with Papanicolaou stain, the filters were processed in the usual manner and permanently mounted for microscopic examination. The filters were examined with the x40 objective for cell counts. Forty random fields were counted by scanning the filter twice and the mean was calculated. The results are expressed as cells per high power field (cells/hpf). The slides were examined with the oil immersion objective for differential morphology.
RESULTS Cell Counts Aqueous humor of eight cataract patients was acellular. Cell counts of patients with bullous keratopathy were extremely variable. The mean cells/hpf ± S.E.M. of ABK patients was 8.5 ± 6.4 (range 0.0-105.0 cell/hpf); ofPBK patients, 30.2, ± 13.0 (range 0.4-285.8) (Tables 1 and 2). Although the mean AH cell count of PBK patients was more than three times greater than that of ABK patients, the Students' t test cannot be used to compare the results because of the high variability of Table 2. Pseudophakic bullous keratopathy. Patient Number
Cataract Technique
Lens Style
Cellslhpf
1
IC*
ACt
0.40
Table 1. Aphakic bullous keratopathy.
AC
0.70
IC
IF+-4Ioop
1.10
4
IC
Metal loop
1.50
5
IC
IF-4loop
2.20
6
EC§
IF
2.75
7
IC
IF-4loop
5.20
8
IC
IF
5.40 5.70
9
IC
IF
10
IC
IF
6.20
11
IC
AC-flexible loop
6.40
12
IC
AC-flexible loop
6.50
Cataract Technique
1
IC*
0.0
13
IC
IF-4loop
6.50
2
IC
0.0
14
IC
AC Choyce
7.10
0.0
15
IC
IF -Copeland
7.50
IC
Iris 4 loop
7.80
IC
4
IC
0.1
16
5
IC
0.0
17
EC
IF-4loop
7.80
6
ECt
0.5
18
IC
AC Choyce
9.80
7
IC
0.5
19
IC
AC
11.30
IC
IF -Copeland
11.80
8
IC
0.5
20
9
IC
0.9
21
IC
IF-4loop
23.00
10
IC
2.6
22
EC
IF -metal 2 loop
25.10
11
IC
3.5
23
EC
IF-4 loop
36.20
12
EC
3.6
24
IC
IF-4loop
41.40
13
IC
3.9
25
IC
IF-Medallion
60.80
14
EC
5.5
26
IC
AC Choyce
230.00
27
EC
IF-4loop
285.80
15
IC
9.9
16
IC
105.0
Mean ± S.E.M. = 8.5 ± 6.4
* IC t EC 330
IC
Patient Number
3
Cells/hpf
2 3
= intracapsular = extracapsular
Mean ± S.E.M.
* IC t AC + IF § EC
AM INTRA-OCULAR IMPLANT SOC
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= intracapsular = anterior chamber = iris fixated = extracapsular
VOL 10, SUMMER 1984
= 30.2 ±
13.0
counts. The data, however, can be considered in another fashion. We can assume that there is a positive correlation between degree of inflammation and increasing cell counts in each group. If we define inflamed eyes as those with more than one cell/hpf, we find seven of 16 (43.8%) ABK patients and 25 of 27 (92.6%) PBK patients in this category (Table 3). This difference is statistically significant at the P < 0.003 level, chi square test. Table 3. Aqueous humor cell counts. ABK Cellslhpf
PBK
Number
Percent
Number
Percent
9 7
56.3 43.7*
2 25
7.4 92.6*
0.0-0.9
> 1.0
* P < 0.003, chi square
Cell Morphology
More than 99% of the nucleated cells in the bullous keratopathy specimens were monocytic histiocytes. The monocytic cells, appearing singly or in clumps were large with a moderate amount of finely granulated cytoplasm (Figure 1). Typically, the cells had a beanshaped, indented, or folded nucleus; many contained phagocytized pigment granules. An occasional polymorphonuclear leukocyte was present as well as rare pigmented epithelial cells, apparently desquamated from the iris. Many free pigment granules were seen on the filters. One specimen from a pseudophakic bullous keratopathy patient contained many multinucleated, foreign body giant cells (Figure 2).
Fig. 1.
(Tuberville) Mononuclear histiocytes from aqueous humor of a patient with pseudophakic bullous keratopathy. Cells typically have bean-shaped or folded nuclei with granular cytoplasm. Many cells have pigment-granule phagosomes in the cytoplasm.
Fig. 2.
(Tuberville) Multinuclear foreign-body giant cell, one of many found in the aqueous humor of one patient with pseudophakic bullous keratopathy.
DISCUSSION At least three factors may be involved in the development of bullous keratopathy following cataract extraction and IOL implantation: the presence of preoperative corneal endothelial dystrophy; intraoperative endothelial cell trauma; and the presence of some measure of chronic postoperative intraocular inflammation. Since we found no cells present in aqueous humor from cataract patients, we can assume that aqueous humor cells are an indication of intraocular inflammation. If we define inflamed eyes as those with more than one cell/hpf in cytologically filtered specimens, we found that a statistically significant greater number of bullous keratopathy patients with an IOL (PBK) than those without an IOL (ABK) have inflamed eyes (93% versus 44%, P < 0.003, chi square). Although it might be expected that eyes with bullous keratopathy would have some cells in the aqueous humor as a result of corneal inflammation, we have demonstrated a significant difference between implanted and nonimplanted patients, all of whom had edematous corneas. Seven percent of PBK patients with less than one cell/hpf (Table 2) may indicate that some patients in this category had significant preoperative corneal dystrophy or intraoperative endothelial trauma without a significant degree of postoperative inflammation. Conversely, 44% of ABK patients with more than one cell/hpf may indicate that these patients had intraoperative complications such as vitreous loss resulting in chronic postoperative inflammation. Although there have been no previous reports of human aqueous humor cytology, Wolter has published several reports regarding cells found clinging to
AM INTRA-OCULAR IMPLANT SOC J-VOL 10, SUMMER 1984
331
IOLs removed from human patients. 7-10 Several cell types were found on the implant including mononuclear histiocytes both quiescent and in the reactive epithelioid stage. 7 Pigment granules, small fibroblast-like cells, and many multinucleated giant cells were also seen. 8-10 Wolter believes that soon after implantation all IOLs are covered with a proteinaceous membrane that supports a rich cell life. 9 It must be kept in mind, however, that the rigid, Choyce-style anterior chamber lenses and the Medallion-type pupillary lenses in that study are considered the most traumatic to uveal structures of the implant styles still in use. It must also be noted that all the patients included in our study had anterior chamber or pupillary lenses. Our very preliminary studies on aqueous humor of eyes with posterior chamber lenses indicate much lower aqueous humor cell counts. Examination of the aqueous humor and IOLs from PBK eyes developing after extracapsular cataract extraction and posterior chamber implantation should yield information on the long-term intraocular inflammatory effects associated with this lens style, which is becoming increasingly popular.
332
REFERENCES 1. Obstbaum SA, Galin MA: Cystoid macular oedema and ocular inflammation: The corneo-retinal inflammatory syndrome. Trans Ophthalmol Soc UK 99:187-191, 1979 2. Shepard DD: Indications for intraocular lens removal. Ophthalmic Surg 8(3):144-148, 1977 3. Drews RC: Inflammatory response, endophthalmitis, corneal dystrophy, glaucoma, retinal detachment, dislocation, refractive error, lens removal, and enucleation. Ophthalmology 85:164-175, 1978 4. Tuberville AW, Galin MA, Perez HD, Banda D, et al: Complement activation by nylon- and polypropylene-looped prosthetic intraocular lenses. Invest Ophthalmol Vis Sci 22:727-733,1982 5. Smith R: Histopathological studies of eyes enucleated after failure of intra-ocular acrylic lens operations. Br J Ophthalmol 40:473-479, 1956 6. Kincaid MC, Green WR, LliffWJ: Granulomatous reaction to Choyce style intraocular lens. Ophthalmic Surg 13:292-299, 1982 7. Wolter JR: Cell life on the surface of lens implants. Graefe's Arch Clin Exp Ophthalmol 218:244-249, 1982 8. Wolter JR: Pigment in cellular membranes on intraocular lens implants. Ophthalmic Surg 13:726-732, 1982 9. Wolter JR, Felt DP: Proliferation of fibroblast-like cells on failing intraocular lenses. Ophthalmic Surg 14:57-64, 1983 10. Wolter JR: Foreign body giant cells on intraocular lens implants. Graefe's Arch Glin Exp Ophthalmol 219:103-111, 1982
AM INTRA-OCULAR IMPLANT SOC
J-
VOL 10, SUMMER 1984
ABSTRACT The long-term effects of the interaction between an IOL and the immune system of the implanted patient are not yet clear. This study presents the results of aqueous humor (AH) cytological analysis of 51 patients undergoing intraocular surgery for cataract, and aphakic (ABK) or pseudophakic bullous keratopathy (PBK). Cataract AH was acellular (N 8). The mean ABK AH cell count was 8.5 ± 6.4 cells per high power field (hpi) of cytologically filtered specimens; mean PBK cell count was 30.2 ± 13.0. If inflamed eyes are defined as those with more than one cell/hpf, a statistically significant greater number of PBK eyes than ABK eyes were inflamed (93% versus 44%, P < 0.003 chi square). More than 99% of the nucleated cells were mononuclear histiocytes. One PBK patient had many multinucleated giant cells in the AH.
=
Key Words: aqueous humor cytology, implant associated uveitis, mononuclear histiocyte, multinucleated foreign-body giant cells, pseudophakic bullous keratopathy
Many modifications in intraocular lens (IOL) design have been made in the last decade to achieve a style that combines good visual rehabilitation, simple insertion techniques, and few postoperative complications. Most technical problems during the operative and immediate postoperative period have been addressed. The long-term effects of the interaction between the implant and the patient's immune system are not yet entirely clear. Implant surgeons have previously reported that a few individuals undergoing IOL surgery may develop a long-term uveitis with the attendant serious sequelae of intraocular inflammation. I -3 Experimental evidence has shown that certain IOL components, namely the nylon and polypropylene loops, are capable of activating the serum complement system in vitro, thereby generating substances that could stimulate inflammation within the eye. 4 The
clinical importance of this is presently under investigation. Although a few histopathological reports of autopsy or enucleated eyes that contain implants are in the literature,5,6 little has been published concerning the ocular histopathology or the pathophysiology of implant-associated uveitis. Few opportunities to reenter the implanted human eye to collect specimens for inspection exist. This study presents the results of aqueous humor cytological analysis of 51 patients undergoing intraocular surgery for cataract and aphakic or pseudophakic bullous keratopathy. MATERIALS AND METHODS Aqueous humor (AH) of 51 eyes undergoing intraocular surgery was examined to determine cell numbers and morphology. Specimens from eight eyes
From the Department of Ophthalmology, University of Tennessee Center for the Health Sciences, Memphis, Tennessee, and the Department of Pathology, Baptist Memorial Hospital, Memphis, Tennessee. Supported in part by an Earl Wilson Award of Fight for Sight, Inc., New York, and by NEI grant EY04086, National Institutes of Health, Bethesda, Maryland. Presented in part at the U.S. Intraocular Lens Symposium, New Orleans, Louisiana, March 1983. Reprint requests to Audrey W. Tuberville, M.D., 956 Court Avenue, Room 2D-29, Memphis, Tennessee 38163. AM INTRA-OCULAR IMPLANT SOC
J-
VOL 10, SUMMER 1984
329
with cataract, 16 eyes with aphakic bullous keratopathy (ABK), and 27 eyes with pseudophakic bullous keratopathy (PBK) were examined. Lens styles ofPBK patients were iris or anterior chamber supported. All operative procedures were performed by one surgeon. Aqueous humor from each patient was collected in the following manner: After retrobulbar anesthesia and digital massage, entrance into the anterior chamber was made with a 27-gauge needle on a plastic tuberculin syringe. About 100 Jotl of AH was aspirated; air (0.5 cc) was drawn into the syringe and the specimen was immediately placed on crushed ice. Within one hour, the specimen was centrifuged at 800 RPMs for three minutes in a microfilter apparatus (Schleicher and Schuell, Inc., Keene, New Hampshire) containing a 9-mm, 5.0 Jot pore size nitrocellulose filter. The filter was immediately fixed in 95% ethanol to avoid drying artifacts. Following staining with Papanicolaou stain, the filters were processed in the usual manner and permanently mounted for microscopic examination. The filters were examined with the x40 objective for cell counts. Forty random fields were counted by scanning the filter twice and the mean was calculated. The results are expressed as cells per high power field (cells/hpf). The slides were examined with the oil immersion objective for differential morphology.
RESULTS Cell Counts Aqueous humor of eight cataract patients was acellular. Cell counts of patients with bullous keratopathy were extremely variable. The mean cells/hpf ± S.E.M. of ABK patients was 8.5 ± 6.4 (range 0.0-105.0 cell/hpf); ofPBK patients, 30.2, ± 13.0 (range 0.4-285.8) (Tables 1 and 2). Although the mean AH cell count of PBK patients was more than three times greater than that of ABK patients, the Students' t test cannot be used to compare the results because of the high variability of Table 2. Pseudophakic bullous keratopathy. Patient Number
Cataract Technique
Lens Style
Cellslhpf
1
IC*
ACt
0.40
Table 1. Aphakic bullous keratopathy.
AC
0.70
IC
IF+-4Ioop
1.10
4
IC
Metal loop
1.50
5
IC
IF-4loop
2.20
6
EC§
IF
2.75
7
IC
IF-4loop
5.20
8
IC
IF
5.40 5.70
9
IC
IF
10
IC
IF
6.20
11
IC
AC-flexible loop
6.40
12
IC
AC-flexible loop
6.50
Cataract Technique
1
IC*
0.0
13
IC
IF-4loop
6.50
2
IC
0.0
14
IC
AC Choyce
7.10
0.0
15
IC
IF -Copeland
7.50
IC
Iris 4 loop
7.80
IC
4
IC
0.1
16
5
IC
0.0
17
EC
IF-4loop
7.80
6
ECt
0.5
18
IC
AC Choyce
9.80
7
IC
0.5
19
IC
AC
11.30
IC
IF -Copeland
11.80
8
IC
0.5
20
9
IC
0.9
21
IC
IF-4loop
23.00
10
IC
2.6
22
EC
IF -metal 2 loop
25.10
11
IC
3.5
23
EC
IF-4 loop
36.20
12
EC
3.6
24
IC
IF-4loop
41.40
13
IC
3.9
25
IC
IF-Medallion
60.80
14
EC
5.5
26
IC
AC Choyce
230.00
27
EC
IF-4loop
285.80
15
IC
9.9
16
IC
105.0
Mean ± S.E.M. = 8.5 ± 6.4
* IC t EC 330
IC
Patient Number
3
Cells/hpf
2 3
= intracapsular = extracapsular
Mean ± S.E.M.
* IC t AC + IF § EC
AM INTRA-OCULAR IMPLANT SOC
J-
= intracapsular = anterior chamber = iris fixated = extracapsular
VOL 10, SUMMER 1984
= 30.2 ±
13.0
counts. The data, however, can be considered in another fashion. We can assume that there is a positive correlation between degree of inflammation and increasing cell counts in each group. If we define inflamed eyes as those with more than one cell/hpf, we find seven of 16 (43.8%) ABK patients and 25 of 27 (92.6%) PBK patients in this category (Table 3). This difference is statistically significant at the P < 0.003 level, chi square test. Table 3. Aqueous humor cell counts. ABK Cellslhpf
PBK
Number
Percent
Number
Percent
9 7
56.3 43.7*
2 25
7.4 92.6*
0.0-0.9
> 1.0
* P < 0.003, chi square
Cell Morphology
More than 99% of the nucleated cells in the bullous keratopathy specimens were monocytic histiocytes. The monocytic cells, appearing singly or in clumps were large with a moderate amount of finely granulated cytoplasm (Figure 1). Typically, the cells had a beanshaped, indented, or folded nucleus; many contained phagocytized pigment granules. An occasional polymorphonuclear leukocyte was present as well as rare pigmented epithelial cells, apparently desquamated from the iris. Many free pigment granules were seen on the filters. One specimen from a pseudophakic bullous keratopathy patient contained many multinucleated, foreign body giant cells (Figure 2).
Fig. 1.
(Tuberville) Mononuclear histiocytes from aqueous humor of a patient with pseudophakic bullous keratopathy. Cells typically have bean-shaped or folded nuclei with granular cytoplasm. Many cells have pigment-granule phagosomes in the cytoplasm.
Fig. 2.
(Tuberville) Multinuclear foreign-body giant cell, one of many found in the aqueous humor of one patient with pseudophakic bullous keratopathy.
DISCUSSION At least three factors may be involved in the development of bullous keratopathy following cataract extraction and IOL implantation: the presence of preoperative corneal endothelial dystrophy; intraoperative endothelial cell trauma; and the presence of some measure of chronic postoperative intraocular inflammation. Since we found no cells present in aqueous humor from cataract patients, we can assume that aqueous humor cells are an indication of intraocular inflammation. If we define inflamed eyes as those with more than one cell/hpf in cytologically filtered specimens, we found that a statistically significant greater number of bullous keratopathy patients with an IOL (PBK) than those without an IOL (ABK) have inflamed eyes (93% versus 44%, P < 0.003, chi square). Although it might be expected that eyes with bullous keratopathy would have some cells in the aqueous humor as a result of corneal inflammation, we have demonstrated a significant difference between implanted and nonimplanted patients, all of whom had edematous corneas. Seven percent of PBK patients with less than one cell/hpf (Table 2) may indicate that some patients in this category had significant preoperative corneal dystrophy or intraoperative endothelial trauma without a significant degree of postoperative inflammation. Conversely, 44% of ABK patients with more than one cell/hpf may indicate that these patients had intraoperative complications such as vitreous loss resulting in chronic postoperative inflammation. Although there have been no previous reports of human aqueous humor cytology, Wolter has published several reports regarding cells found clinging to
AM INTRA-OCULAR IMPLANT SOC J-VOL 10, SUMMER 1984
331
IOLs removed from human patients. 7-10 Several cell types were found on the implant including mononuclear histiocytes both quiescent and in the reactive epithelioid stage. 7 Pigment granules, small fibroblast-like cells, and many multinucleated giant cells were also seen. 8-10 Wolter believes that soon after implantation all IOLs are covered with a proteinaceous membrane that supports a rich cell life. 9 It must be kept in mind, however, that the rigid, Choyce-style anterior chamber lenses and the Medallion-type pupillary lenses in that study are considered the most traumatic to uveal structures of the implant styles still in use. It must also be noted that all the patients included in our study had anterior chamber or pupillary lenses. Our very preliminary studies on aqueous humor of eyes with posterior chamber lenses indicate much lower aqueous humor cell counts. Examination of the aqueous humor and IOLs from PBK eyes developing after extracapsular cataract extraction and posterior chamber implantation should yield information on the long-term intraocular inflammatory effects associated with this lens style, which is becoming increasingly popular.
332
REFERENCES 1. Obstbaum SA, Galin MA: Cystoid macular oedema and ocular inflammation: The corneo-retinal inflammatory syndrome. Trans Ophthalmol Soc UK 99:187-191, 1979 2. Shepard DD: Indications for intraocular lens removal. Ophthalmic Surg 8(3):144-148, 1977 3. Drews RC: Inflammatory response, endophthalmitis, corneal dystrophy, glaucoma, retinal detachment, dislocation, refractive error, lens removal, and enucleation. Ophthalmology 85:164-175, 1978 4. Tuberville AW, Galin MA, Perez HD, Banda D, et al: Complement activation by nylon- and polypropylene-looped prosthetic intraocular lenses. Invest Ophthalmol Vis Sci 22:727-733,1982 5. Smith R: Histopathological studies of eyes enucleated after failure of intra-ocular acrylic lens operations. Br J Ophthalmol 40:473-479, 1956 6. Kincaid MC, Green WR, LliffWJ: Granulomatous reaction to Choyce style intraocular lens. Ophthalmic Surg 13:292-299, 1982 7. Wolter JR: Cell life on the surface of lens implants. Graefe's Arch Clin Exp Ophthalmol 218:244-249, 1982 8. Wolter JR: Pigment in cellular membranes on intraocular lens implants. Ophthalmic Surg 13:726-732, 1982 9. Wolter JR, Felt DP: Proliferation of fibroblast-like cells on failing intraocular lenses. Ophthalmic Surg 14:57-64, 1983 10. Wolter JR: Foreign body giant cells on intraocular lens implants. Graefe's Arch Glin Exp Ophthalmol 219:103-111, 1982
AM INTRA-OCULAR IMPLANT SOC
J-
VOL 10, SUMMER 1984