D Kalicharan

A COMBINED SCANNING AND TRANSMISSION ELECTRON-MICROSCOPIC INVESTIGATION OF HUMAN (SECONDARY) CATARACT MATERIAL

Cataract lenses from patients of advanced age were processed for SEM by standard pre-fixation followed by treatment by the Tannine-Arginine-Osmiumtetroxide (TAO) method and critical point drying, and for TEM by standard pre-fixation followed by vibratomation, standard post-fixation, ultramicrotome sectioning and staining with uranyl acetate/lead citrate. Secondary cataract material was brought onto a Millipore filter, fixed by standard methods, dried in air and sputter-coated with Au. Both SEM and TEM images revealed degeneration processes in lensfibre material, such as swelling of the lensfibre, protrusion of the cytoplasm, fibrillation of the cell membrane, loss of the nucleus, spherical bodies of various sizes between 0.5–1.5 μm, sometimes surrounded by a (double) membrane with different contrast but without cellular evidence, and small and large vacuoles partly filled with granular material both in and at the periphery of the lensfibre-body. The secondary cataract material on the Millipore filter revealed erythrocytes and more or less spherical bodies with high contrast, measuring between 0.5–1.5 μm, often referred to as Elschnigs pearls, besides non-definable organic material. The SEM and TEM micrographs of the cataract lens material strongly suggest that the spherical bodies with sizes of approximately 0.5–1.5 micrometer and high contrast without cellular evidence, are similar to the more or less spherical bodies found in the secondary cataract material on the filter, referred to as Elschnigs pearls.

HUMAN CAPSULE EPITHELIAL-CELL DEGENERATION A LM, SEM AND TEM INVESTIGATION

The degeneration of the capsule epithelium of cataractous lenses has been studied with LM, SEM en TEM with emphases on TEM. The observed degeneration of the epithelial cells can be described as follows: The cell nucleus becomes picnotic and desintegrates as result of change of the chromatin. Degeneration of the cytoplasm starts with swelling of the mitochondria, coming into existence of filamentous networks and balloon-like bulges of the nucleus. Repelling of the cell nucleus due to a porosity of the plasm membrane. Collapse of the cell due to degeneration of the cell and loss of the cytoplasmic contents, leaving finally only a swollen framework of cell walls.

(An)aerobic bacteria found in secondary-cataract material

Twentyfour patients, who had marked reduction of vision due to secondary-cataract developed after an ECCE, were treated by surgical cleaning of the posterior lens capsule. During this procedure globular secondary-cataract material was removed and collected for morphological examination by SEM and TEM. Fragments of various sizes and shapes, including some with a ‘golfball’ structure, were seen; these closely resembled particles frequently found in cataractous lenses. In addition, in 18 patients micro-organisms were found: rod-shaped bacteria, cocci, and in 2 cases yeasts. These findings were the more remarkable because these were clinically quiet eyes with no signs of intra-ocular inflammation and cultures have been persistently negative. We imagine that these bacteria must have entered the eye during the cataract extraction and have settled there without causing an infection.

Study of the substructure of the Morgagni and Brunescens cataract with the TAO non-coating technique

Lens tissue from a Morgagni cataract was examined by SEM and TEM. For SEM, after prefixation with glutaraldehyde and postfixation with the tannic acid/arginine/OsO4 non-coating (TAO) technique, and for TEM, after prefixation with glutaraldehyde, postfixation with OsO4/K4Fe(CN)6 and poststaining with uranyl acetate/lead citrate. The TAO technique seems to be a particularly suitable postfixation method for the SEM investigation of cararact tissue because of the presence of the protein structures present. The cortical region showed areas of radially, instead of concentrically, arranged lens fibres, degenerated lens fibres with holes (vacuoles), broken ball and socket connections between the lens fibres, and oval or spherical structures varying in size from 0.5–20 μm, the largest resembling a golfball, arising from the cytoplasm of degenerating lens fibres. The smallest, 0.2–0.5 μm, appear to have been expelled from the furrowed lens epithelium.

Cell-ingrowth in a silicone plombe

A male patient (42 years) who had been treated for retinal detachment by the implantation of a silicone plombe into the sclera, returned to the clinic 8 years after implantation because of inflammation of the tissue and partial protrusion from the sclera. After removal of the plombe this was processed for TEM and SEM and examination of the plombe material after 8 years in situ could be carried out in order to get information about the cell-biomaterial interface. A large proportion of the silicone pores was filled with cellular material, including macrophages, giant cells and erythrocytes thus indicating a foreign body granuloma. The external surface of the pores showed a granular osmiophilic dense amorphous layer including extracellular debris. Engulfing of silicone particles by macrophages and the evidence of long-term tissue response suggest partial biodegradation of the silicone and certainly not complete inertness as was formerly claimed.

Reaction of the rabbit corneal endothelium to nylon sutures

Nylon and stainless steel sutures separately placed deeply into rabbit corneas by splitting the stroma for a few millimeters, without closing sutures, remained in the cornea for two, four and six weeks respectively. In contrast to the stainless steel sutures an extensive tissue reaction could be observed clinically around the nylon sutures within a few days and was still present after 4–6 weeks. On the endothelial side, covering of the nylon with fibroblast cells took place very slowly, this in contrast with stainless steel. Irritation of the tissue and an oedematous appearance of the endothelium around the nylon suture was the result. At first a kind of collagen network, often mixed with inflammatory cells, was deposited on the nylon material before fibroblast cells could grow in. Covering of the stainless steel started quite soon after implantation, without preliminary deposition of collagen material. The reason for this phenomenon must be sought in the high free surface energy of the stainless steel, which attracts cells, in contrast to nylon which has a very low free surface energy.

LENSFIBRE DEGENERATION AT CATARACT LENSES - A LM, SEM AND TEM INVESTIGATION

The degeneration process of lensfibres in a cataractous lens, described as the biochemical changes of a part of the lensproteins, can be characterised morphologically as follows: Emulsification of a part of the lensfibre-mass and the development of open spaces between the lensfibres with the formation of vacuoles (spherical structures) with a granular or a compact contents with high contrast, which is squeezed into the intercellular space. Degeneration of the ball & socket system interdigitation and microvilli domain system and the formation of almost empty rectangular structures in somewhat radial array. Balloon-like bulging of the cytoplasm of the lenscell, degeneration of the cytoplasm membrane and visualisation of a micro-filamentous network with enclosed cell organelles.

Morphological aspects of human lens capsules

Lens capsules of patients of advanced age, obtained after extracapsular cataract surgery, were carefully prepared for a combined LM, TEM and SEM investigation, after preliminary washing and mounting onto a holder in a buffer solution. After pre-fixation with GA, samples were postfixed for LM/TEM and OsO4/K4Fe(CN)6 and stained with toluidineblue/basic fuchsine for LM and with uranyl acetate/lead citrate for TEM; for SEM the GA-pre-fixed samples were post-fixed by the Tannine Arginine-OsO4 non-coating technique. At LM-level discrimination between healthy and degenerating cells was possible after toluidine staining. At SEM-level protrusion of the cell nucleus and fibrillation and blebbing of the cell membrane as the result of capsular degeneration could be observed with the TAO-method. At TEM-level protrusion of the cell nucleus, degeneration of the cytoplasm, ballooning of the mitochondria, the presence of microfilaments and the occurrence of vacuoles were visible as the result of capsular degeneration on cataract formation.