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Lenscapsule material of human cataractous lenses studied by LM, SEM and TEM
0739-6260/90 $3.00 +0.00
Micron and Microscopica Ada, Vol. 21. No. 4, Pp. 261—262. 1990. Printed in Great Britain.
Pergamon Press plc
LENSCAPSIJLE MATERIAL OF HUMAN CATARACTOUS LENSES STUDIED BY LM, SEM and TEM D.Kalicharan’, W .L.Jongebloed’ and J G.F.Worst2 .
Lab. Histology & Ceilbiology RUG, Oostersingel 69/1, Groningen 2 Eye Physician and Surgeon, Haren, The Netherlands
INTRODUCTION: Earlier SEM-studies of capsules of human cataractous lenses sometimes gave conificting results at interpretation, due to inferior fixation procedures, resulting in the existance of artificial structures difficult to discriminate from pathological findings (Los et aL, 1989; Jongebloed et al., 1990). The purpose of this study was a reinvestigation of the cellular elements in anterior capsular cataracts, performed at lightmicroscopical (LM) and ultrastructural (TEM/SEM) level at a standardized fixing procedure. MATERIALS & METHODS: Capsules of cataract lenses of patients of advanced age, extracted during extracapsular cataract surgery, were prepared for LM, SEM and TEM. Capsules were mounted in cold sodiumcacodylate buffer (pH 7.4, 400 mOsm, 4°C.)on silicon-rubber and subsequently fixed in buffered G.A. (2%, pH 7.4, 440 mOsm., 24 h.). For LM/TEM the specimens were post-fixed in a buffered mixture of 1% 0s0 4 and 1.5% K4Fe(CN)6 (pH 7.4, 4°C,2h), rinsed in buffer, dehydrated and flat embedded after proper orientation. For LM 1 ~&m sections were stained with basic fuchsine/toluidine blue. Ultrathin sections were stained using 10% uranyl acetate in methanol and lead citrate, and examined with a Akashi 002A electronmicroscope. For SEM the G.A.-flxed specimens were washed with buffer, immersed in a mixture of 2% Tannicacid /2% Arginine-HCI /2% Guanidine-HC1 (16h ,20°C),washed carefully with aquabidest (15 mm.) and post-fixed with 2% 0s04 (8h)(Malick et aL, 1975). After dehydration with graded alcohol and critical point drying via C02, the specimens were observed without Au-sputtering in a JEOL SEM at 15-25 kV. RESULTS: At light microscopical level the lenscapsule demonstrated proliferative changes with multilaniellar arrangements of the capsular epitheial cells and an increase in the pleomorphic appearance of cell and nucleus. At the ultrastructural level the subcapsular epithelium showed atrophy and blebbing of cells at the overlying anterior capsule. Processes in the cytoplasm like the existance of picnotic nuclei, ballooning of the mitochondria, detachment of epitheial cells from the basal membrane, the appearance of vacuoles of various sizes and the presence of thin filaments are indicative for progressive celldegradation. CONCLUSION: It is possible that cataractogenesis is associated with fysiological disbalance and ultrastructural metaplastic and degenerative alterations as shown. REFERENCES: M. Hogan, J. Alvarado and J. Wedell (1971). Histology of the human eye. W.B. Saunders Comp. W.L. Jongebloed, L.I. Los, F. Dijk and J.G.F. Worst (1990). Morphology of donor lens-capsule material. Doc. Ophthalmol., in press. L.I. Los, W.L. Jongebloed and JGF Worst (1989). Lens-capsule material of human and animal origin, studied by SEM. Doc. Ophthalmol.72: 357-365. L.E. Malick and R.B. Wilson (1975). Evaluation of a modified technique for SEM examination of verterbrate specimens without evaporated layers. Scanning Electron Microscopy. 0. Johari (ed.) lIT Research Institute, Chicago, IL, 259-266. 261
262
I). Kaileliardit et a!.
41 Fig. I A stilized drawing of the lens, cortex, epithelium (A. B. C), capsule (D) and zonular atta ments (G) (Hogan et al., 1971). Fig. 2 LM image of the pleomorphic capsular epithelial cells (CEC). x930 Fig. 3 Low power TEM showing several atrophic capsular epithelial cells (CEC) near the capsule of an area similar in Fig. 2. x3600 Fig. 4 Part of the capsular epithelial cell (CEC) cytoplasm showing ultrastructure of higher swelli with disintegration of cristae in mitochondria. x45000 Fig. 5 SEM showing characteristic multiple capsular epithelial cell (CEC) morphology with blcbs a ruffles. x2100
Micron and Microscopica Ada, Vol. 21. No. 4, Pp. 261—262. 1990. Printed in Great Britain.
Pergamon Press plc
LENSCAPSIJLE MATERIAL OF HUMAN CATARACTOUS LENSES STUDIED BY LM, SEM and TEM D.Kalicharan’, W .L.Jongebloed’ and J G.F.Worst2 .
Lab. Histology & Ceilbiology RUG, Oostersingel 69/1, Groningen 2 Eye Physician and Surgeon, Haren, The Netherlands
INTRODUCTION: Earlier SEM-studies of capsules of human cataractous lenses sometimes gave conificting results at interpretation, due to inferior fixation procedures, resulting in the existance of artificial structures difficult to discriminate from pathological findings (Los et aL, 1989; Jongebloed et al., 1990). The purpose of this study was a reinvestigation of the cellular elements in anterior capsular cataracts, performed at lightmicroscopical (LM) and ultrastructural (TEM/SEM) level at a standardized fixing procedure. MATERIALS & METHODS: Capsules of cataract lenses of patients of advanced age, extracted during extracapsular cataract surgery, were prepared for LM, SEM and TEM. Capsules were mounted in cold sodiumcacodylate buffer (pH 7.4, 400 mOsm, 4°C.)on silicon-rubber and subsequently fixed in buffered G.A. (2%, pH 7.4, 440 mOsm., 24 h.). For LM/TEM the specimens were post-fixed in a buffered mixture of 1% 0s0 4 and 1.5% K4Fe(CN)6 (pH 7.4, 4°C,2h), rinsed in buffer, dehydrated and flat embedded after proper orientation. For LM 1 ~&m sections were stained with basic fuchsine/toluidine blue. Ultrathin sections were stained using 10% uranyl acetate in methanol and lead citrate, and examined with a Akashi 002A electronmicroscope. For SEM the G.A.-flxed specimens were washed with buffer, immersed in a mixture of 2% Tannicacid /2% Arginine-HCI /2% Guanidine-HC1 (16h ,20°C),washed carefully with aquabidest (15 mm.) and post-fixed with 2% 0s04 (8h)(Malick et aL, 1975). After dehydration with graded alcohol and critical point drying via C02, the specimens were observed without Au-sputtering in a JEOL SEM at 15-25 kV. RESULTS: At light microscopical level the lenscapsule demonstrated proliferative changes with multilaniellar arrangements of the capsular epitheial cells and an increase in the pleomorphic appearance of cell and nucleus. At the ultrastructural level the subcapsular epithelium showed atrophy and blebbing of cells at the overlying anterior capsule. Processes in the cytoplasm like the existance of picnotic nuclei, ballooning of the mitochondria, detachment of epitheial cells from the basal membrane, the appearance of vacuoles of various sizes and the presence of thin filaments are indicative for progressive celldegradation. CONCLUSION: It is possible that cataractogenesis is associated with fysiological disbalance and ultrastructural metaplastic and degenerative alterations as shown. REFERENCES: M. Hogan, J. Alvarado and J. Wedell (1971). Histology of the human eye. W.B. Saunders Comp. W.L. Jongebloed, L.I. Los, F. Dijk and J.G.F. Worst (1990). Morphology of donor lens-capsule material. Doc. Ophthalmol., in press. L.I. Los, W.L. Jongebloed and JGF Worst (1989). Lens-capsule material of human and animal origin, studied by SEM. Doc. Ophthalmol.72: 357-365. L.E. Malick and R.B. Wilson (1975). Evaluation of a modified technique for SEM examination of verterbrate specimens without evaporated layers. Scanning Electron Microscopy. 0. Johari (ed.) lIT Research Institute, Chicago, IL, 259-266. 261
262
I). Kaileliardit et a!.
41 Fig. I A stilized drawing of the lens, cortex, epithelium (A. B. C), capsule (D) and zonular atta ments (G) (Hogan et al., 1971). Fig. 2 LM image of the pleomorphic capsular epithelial cells (CEC). x930 Fig. 3 Low power TEM showing several atrophic capsular epithelial cells (CEC) near the capsule of an area similar in Fig. 2. x3600 Fig. 4 Part of the capsular epithelial cell (CEC) cytoplasm showing ultrastructure of higher swelli with disintegration of cristae in mitochondria. x45000 Fig. 5 SEM showing characteristic multiple capsular epithelial cell (CEC) morphology with blcbs a ruffles. x2100