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Two-phase endolymphatic hydrops is a subtle experimental model for Meniere's disease. Chronic dysfunction of the endolymphatic sac, induced by dissection of the most distal part without causing damage to the intermediate part, is combined with increased endolymph production induced by administration of aldosterone which stimulates the Na/K-ATPase in the stria vascularis. A transmission electron microscopic study was performed on the endolymphatic sacs of four groups of guinea pig cochleas: controls; non-operated aldosterone-treated cochleas; operated (dissection of the endolymphatic sac) cochleas; operated and aldosterone-treated cochleas. Light and electron microscopy showed a normal morphology in the controls. Aldosterone treatment had no visible effect. Dissected ears revealed severe deviations. The epithelium of the intermediate sac was low, showed dilated lateral intercellular spaces indicating elevated fluid transport and displayed serious degenerative processes. Distally, the endolymphatic sac was completely blocked by newly formed bone. Additional aldosterone treatment had no cumulative effect on the dissected ears. [ABSTRACT FROM AUTHOR]

The foreign body reactions to collagen-immobilized polyurethane (PU-CI) films during subcutaneous implantation in rats were characterized. The underlying concept is that collagen-immobilization will improve the tissue integration. Since the method of collagen-immobilization involves the covalent coupling of collagen to an acrylic acid (AA) based surface graft, both non-modified PU and PU-AA were used as controls. Bare PU has a flat surface, whereas both PU-AA and PU-CI displayed a slightly roughened surface. Implantation showed that PU-CI induced early after implantation a far more intense foreign body reaction than PU and PU-AA. This reaction consisted of increased presence of fibrin, granulocytes and macrophages. Roughening of the surface as with PU-AA induced only a small increase in fibrin formation and cellular migration. At day 5 the reaction to PU-CI had slowed down; giant cell formation now slowly started but was decreased compared to PU and PU-AA. At day 10 capsules around each type of material looked similar, but in contrast to PU. PU-CI films could no longer be dissected from their capsules. Only at week 3 this also occurred with PU, at which time point again similar capsules with the three materials were observed. At week 6, of the three materials PU-CI showed the thinnest capsule with most immediate adherence of connective tissue. These results show that collagen-immobilization of PU increased the early tissue reaction and therefore the tissue integration. The thin capsule observed at 6 weeks may be beneficial in e.g. infectious circumstances, when easy access for immune reactions is needed. This, and the long-term performance of PU-CI will be a matter of future investigations.

The aim of this study was to evaluate short-term peripheral nerve regeneration across a 15-mm gap in the sciatic nerve of the rat, using a thin-walled biodegradable poly(DL-lactide-epsilon-caprolactone) nerve guide filled with modified denatured muscle tissue (MDMT). The evaluation was performed using transmission electron microscopy and morphometric analysis. Evaluation times ranged from 3 to 12 weeks after reconstruction. Already, 3 weeks after reconstruction, myelinated nerve fibers could be observed in the distal nerve stump. Twelve weeks after reconstruction, the number of (non)myelinated nerve fibers had significantly increased in the distal nerve stump. From this study, we can conclude that a thin-walled biodegradable poly(DL-lactide-epsilon-caprolactone) nerve guides filled with MDMT can be successfully applied in the reconstruction of severed nerves in the rat model. Furthermore, we showed fast nerve regeneration across the 15-mm nerve gap and found that the use of MDMT functioned as a mechanical support preventing a collapse of this thin-walled nerve guide.

The aim of this study was to evaluate short-term peripheral nerve regeneration across a 10-mm. gap, using a biodegradable poly(DL-lactide-epsilon-caprolacton) nerve guide, with an internal diameter of 1.5 mm and a wall thickness of 0.30 mm. To do so, we evaluated regenerating nerves using light microscopy, transmission electron microscopy and morphometric analysis after implantation of 12-mm nerve guides in the sciatic nerve of the rat. Evaluation times ranged from 3-10 weeks. Three weeks after reconstruction, myelinated nerve fibers could be observed in the distal nerve stump. Ten weeks after reconstruction, the regenerating nerves already resembled normal nerves. In conclusion, we show that poly(DL-lactide-epsilon-caprolacton) nerve guides can be successfully applied in the reconstruction of severed nerves in the rat model. Furthermore, we have observed the fastest nerve regeneration described thus far, after reconstruction using a biodegradable nerve guide.

Purpose: In oral and maxillofacial surgery palatal mucosal grafts are routinely used to cover mucosal defects caused by vestibuloplasty. However, the quantity of palatal mucosa is a limiting factor in more extensive operations. This study investigated whether autologous cultured sheets of mucosa can serve as a dressing for these wounds., Materials and Methods: Punch biopsies (diameter, 4 mm) were taken from the hard palate of eight patients (five men, three women; mean age 43 years). Epithelial cells were enzymatically dissociated from these tissue specimens and grown in vitro in the presence of a fibroblast feeder layer. Within 3 weeks, a transplantable epithelial sheet of about 20 cm2 was obtained. The sheet was detached from the culture flask by enzyme treatment and fixed to a carrier of Vaseline (Cheeseborough Ponds Inc, Greenwich, CT) gauze. Using a split-mouth technique, the sheet was placed on half of a mucosal defect created by vestibuloplasty, while the other half of the defect was covered by a conventional split-thickness palatal graft. Both the cultured and conventional graft were held in place by the patient's relined denture fixed with perimandibular sutures. One week postsurgery, the denture and Vaseline gauze were removed. Three months after vestibuloplasty, biopsy specimens of each grafted site were taken and processed for light and transmission electron microscopy (LM, TEM)., Results: Three months postsurgery, the grafted mucosa of both sites bore close resemblance to palatal mucosa. Both the cultured and split-thickness grafts were vascularized, did not evoke a homograft reaction, and showed a smooth graft/lip mucosal junction and minimal wound contraction. LM and TEM revealed that both types of grafts formed a fully differentiated keratinizing mucosa with a well-developed basement membrane and rete ridges, comparable with the histology and ultrastructure of palatal mucosa in situ., Conclusion: It was concluded from this study that cultured mucosa can serve as a proper dressing for mucosal defects after vestibuloplasty.

In this morphological study the (ultra)structural changes that lead to contraction of detached cultured epithelium were investigated. Keratinocytes, isolated from human skin and oral mucosa, were grown to form stratified cell sheets. The multilayers were examined with light and electron microscopy before, during and after detachment from the culture vessel. Attached epithelium had a stretched morphology with flattened cells and nuclei. Evidence is provided that after enzymatical detachment with dispase (1) basal cells became columnar by contraction of actin bundles in the basal cortex, which was accompanied by blebbing of the basal cell membrane; (2) in all cell layers cytokeratin bundles contracted resulting in displacement of desmosomes and a spherical shape of the cells and nuclei. By slow dispase-detachment at 4 degrees C or by quick mechanical detachment, shrinkage of the sheet was partly suppressed but contraction of cytokeratin and related events occurred indicating that these were the result of the spontaneous reassembly of the intermediate filament system. The results suggested that the shape and ultrastructure of all cells in an epithelial multilayer are dependent on the interaction of the basal cells with the underlying extracellular matrix.

A classification of the existing multitude of cystic lesions of the brain is proposed, which allows an understanding of their genesis and consequent therapeutic implications, as well as their diagnostic characteristics. Essentially, cerebral cystic lesions may be classified into the following categories: Cysts containing CSF-like fluid, which include ex vacuo type cysts, such as leptomeningeal cysts, and cysts following surgical resection; cysts with fluid secreting walls and CSF-like content, such as arachnoid cysts; cysts associated with dysgenesis, for example Dandy-Walker cysts. The ex vacuo cysts increase craniospinal compliance, whereas the other cysts with CSF-like content do not; they are not per se expansive, however, although their occasional location along CSF pathways may cause obstruction and hydrocephalus. Another category includes cysts with a lining of non-neural epithelium like colloid cysts, epidermoid cysts, or craniopharyngiomas. They may increase in size and cause symptoms by compression, although not at the rate of tumour-associated cysts. The cysts associated with gliomas and other tumours have a pathogenesis bearing upon blood-brain barrier impairment and formation of vasogenic oedema. Finally, one may distinguish a category of cysts with infectious origin, such as brain abscesses and hydatid cysts. The cysts with CSF-like contents may be recognised by their magnetic resonance characteristics resembling those of CSF, whereas cysts containing proteinaceous fluid are associated with blood-brain barrier impairment and consequent contrast enhancement. The cysts with a lining of non-neural epithelium exhibit diverse properties of attenuation on computed tomography (CT) and magnetic resonance imaging (MRI), depending on the nature of their cyst contents.

This review presents various aspects of the technological development, and their assessment in the design of a contrast agent for MRI, tailored to visualise tumours in the brain. First, it was demonstrated that magnetite as a contrast agent exhibited a much stronger relaxivity than gadolinium. The prepared magnetite particles bound to dextran, were also shown to be of appropriate size by electron microscopy. After their intravenous injection into rats with blood-brain barrier disruption, the lesion was strongly enhanced by T2-shortening. Furthermore, monoclonal antibodies directed against small cell lung carcinoma, proved to be able to penetrate into tumours, which had been raised by implantation of the small cell lung carcinoma cells into the brains of nude rats. As to the essential step, it was demonstrated in vitro that magnetite particles coupled to monoclonal antibodies by the biotin-streptavidin binding, could be bound to the target cells of the antibody, changing the relaxation rates of the latter. Finally it could be shown in vitro that an alternative approach, using lymphocytes to be targeted to tumour cells, also proved feasible, in that these lymphocytes could be labelled with magnetite that had been incorporated into liposomes. Further developments will be the in vivo assessment of the acquired progress in experimental animals, before clinical application is warranted.

Nerve regeneration using artificial biodegradable conduits is of increasing interest. The aim of this study is to evaluate the regeneration and maturation of a nerve after long-term implantation (2 years) of a biodegradable poly-L-lactide/poly-epsilon-caprolactone (PLLA/PCL) copolymeric nerve guide in the sciatic nerve of the rat. After harvesting, we evaluated both the regenerated nerves and the controls, using light microscopy, transmission electron microscopy, and morphometric techniques. Remnants of biomaterial were still present after 2 years of implantation, but the foreign body reaction was very mild at this stage, due to the rounded shapes of the polymer debris. Morphometric analysis showed significant differences between the regenerated nerve and the normal sciatic nerve: the number of myelinated fibers is higher, and the mean fiber diameter of the myelinated fibers in the regenerated nerve is smaller. In conclusion, the results demonstrate that the new PLLA/PCL nerve guide can provide optimal conditions for regeneration and maturation of damaged nerves.

In this study, the authors have examined the mechanism of the formation of tumor cysts. Cyst fluid samples were obtained during surgery and by percutaneous aspiration from 22 patients with cystic cerebral gliomas. The concentration of protein was measured in the cyst fluid and blood plasma. Analysis of brain tumor cyst fluids revealed that plasma proteins constituted a major fraction (92%) of cyst fluid proteins; moreover, the protein fractions occurred in concentrations (relative to the plasma concentrations) that were around 50-fold of those in cerebrospinal fluid. This strongly indicates blood-brain barrier disruption. Evidence from computed tomographic and magnetic resonance imaging scans as well as from electron microscopy of tumor cyst walls suggests the transition of spongy edematous tissue in or around tumors into the contents of associated cysts. Pathophysiologically, blood-brain barrier breakdown is inherent to the occurrence of vasogenic brain edema. It is therefore plausible that the development of cysts is related to peritumoral vasogenic edema.

A new method of thymus transplantation is introduced, in which the graft is directly connected with the recipient's vascular system. This procedure was used both in euthymic rats and congenitally athymic nude rats. At all tested intervals after transplantation thymus grafts hardly differed from the recipient's own thymus in immunohistology and lymphocyte yield. In athymic nude rats, T cell-dependent immunity, tested by mitogen- and alloantigen-induced T cell responses, as well as by antibody production and delayed-type hypersensitivity after ovalbumin administration, showed that vascular thymus grafts could generate T cell functions to euthymic control levels. We conclude that the technique of vascular thymus transplantation represents a valuable tool, either in fundamental research on thymus function, or for the purpose of immune (re)constitution.

No reliable connective tissue stains for GMA sections were available until recently. However, the use of toluidine blue in combination with basic fuchsin appeared to be a rapid and reliable connective tissue stain for glycol methacrylate (GMA) embedded tissue.

Microporous, compliant, biodegradable vascular grafts prepared from a mixture of polyurethane (95% weight) and poly-L-lactic acid (5% weight) can function as a temporary scaffold for the regeneration of the arterial wall in small-caliber arteries. This study was undertaken to document the sequential events leading to this regeneration. Therefore, polyurethane/poly-L-lactic acid vascular grafts were implanted into the abdominal aorta of rats (N = 28) and were harvested at regular intervals from 1 hour up to 12 weeks after implantation. The implants were evaluated by means of light and electron microscopy. At each time of harvesting, the implants were patent and showed arterial pulsations. No stenosis or dilatation was observed. Endothelial cells grew from the adjacent aortic intima across the anastomoses, from day 6 onward, to form an almost complete neointima after 6 weeks of implantation. Smooth muscle cells also grew from the adjacent aortic media over the graft lattice through the platelet-fibrin coagulum from day 6 onward. The smooth muscle cells, predominantly longitudinally arranged at week 6, but also circularly arranged in some areas at week 12, formed a neomedia in which elastic laminae regenerated. Polymorphonuclear leukocytes and monocytes initially invaded the graft lattices. Fibroblasts, histiocytes, and capillaries grew from the perigraft tissue into the polyurethane/poly-L-lactic acid lattices from day 6 onward, which resulted in the formation of a neoadventitia. The polyurethane/poly-L-lactic acid lattices started to disintegrate from day 12 onward. The regenerative processes in the disintegrating polyurethane/poly-L-lactic acid grafts resulted in the formation of neoarteries, which were of sufficient strength, compliance, and thromboresistance to function as small-caliber arterial substitutes.

Morphological and enzyme ultracytochemical evidence is presented to support the contention that the walls of arachnoid cysts secrete fluid. Clinical evidence has already suggested this phenomenon, including intracranial pressure elevation and expansion in some cases, and the observation that arachnoid cysts constitute closed compartments with a fluid content that cannot be derived from other cerebrospinal fluid-containing spaces. Ultrastructurally, the cyst lining showed a similarity to subdural neurothelium and the neurothelial lining of arachnoid granulations in such morphological features as intercellular clefts with sinusoid dilatations, desmosomal intercellular junctions (upon which tonofilaments may be abutting), pinocytotic vesicles, multivesicular bodies, lysosomal structures, and the presence of a basal lamina. Some of these features, together with the presence of microvilli on the luminal surface, are consistent with fluid secretion. Moreover, enzyme cytochemistry demonstrated (Na+ + K+)-ATPase in the plasma membranes lining the cavity, either directly (the apical membranes), or via the intercellular clefts (the basolateral membranes), and, with alkaline phosphatase occupying the opposite plasma membranes, this structural organization indicates fluid transport toward the lumen. It may be surmised that arachnoid cysts derive from subdural neurothelium differentiating towards arachnoid villus mesothelium.

The morphological similarity of the lining of arachnoid cysts to subdural neurothelium and the mesothelium of arachnoid granulations suggested that the latter tissues might be the origin of arachnoid cysts. Transport Na+-K+-adenosine triphosphatase was shown by enzyme ultracytochemistry to be an indication of secretory activity in the lining of arachnoid cysts and in the endothelial lining of arachnoid granulations. This secretory activity suggests the existence of a biochemical mechanism for cerebrospinal fluid absorption at these granulations separate from the mechanisms already demonstrated.

The ultrastructure of a new type of vascular graft, prepared from a mixture of polyurethane (95 weight %) and poly-L-lactic acid (5 weight %), was examined six weeks after implantation into the abdominal aorta of rats. These microporous, compliant, biodegradable, vascular grafts function as temporary scaffolds for the regeneration of the arterial wall. Smooth muscle cells, covering the grafts, regenerated a neo-media underneath an almost completely regenerated endothelial layer (neo-intima). These smooth muscle cells varied in morphology from normal smooth muscle cells to myofibroblasts. They were surrounded by elastic laminae and collagen fibers. Macrophages, epithelioid cells, multinucleated giant cells, fibroblasts and capillaries were present in the disintegrating graft lattices. The epithelioid cells and multinucleated giant cells engulfed polymer particles of the disintegrating grafts. The regeneration of the endothelial and smooth muscle cells is similar to the natural response of arterial tissue upon injury. The presence of macrophages, epithelioid cells, multinucleated giant cells, fibroblasts and capillaries in the graft lattices resembles the natural response of tissue against foreign body implants. Both of these responses result in the formation of a neo-artery that possesses sufficient strength, compliance and thromboresistance to function as a small caliber arterial substitute.

The in vitro adhesion and spreading of human endothelial cells (HEC) on hydrophobic poly(ethylene terephthalate) (PETP) and moderately wettable tissue culture poly(ethylene terephthalate) (TCPETP) were studied with light microscopy and electron microscopy. Numbers of HEC adhering on TCPETP were always higher than those found on PETP. When cells were seeded in the presence of serum, extensive cell spreading on both PETP and TCPETP was observed after the first 30 min. Thereafter, spread cells appeared to withdraw from the PETP surface, resulting in irregularly shaped cells. Complete cell spreading occurred on TCPETP. Complete cell spreading also occurred on PETP and TCPETP when HEC had first been seeded from phosphate buffer solution and serum was supplied after 30 min. Furthermore, HEC spread on both PETP and TCPETP when the surfaces were precoated with protein(s), which promotes cell adhesion. However, when plasma was used for the coating, spread cells did not proliferate in a monolayer pattern. This study shows that TCPETP is, in general, a better surface for adhesion and proliferation of HEC than is PETP, suggesting that vascular prostheses with a TCPETP-like surface will perform better in vivo than prostheses made of PETP.

To obtain improved transmission electron microscopy sections for cell biological and interface evaluation of implanted biomaterials we present an improved embedding procedure. Standard problems in preparation and sectioning, like dissolution of the biomaterial, or holes and chatter in the sections can be prevented by introducing butyl-2,3-epoxypropylether as an intermedium between the dehydration series and the Epon resin. Most biomaterials were not affected by this chemical agent. The introduction of butyl-2,3-epoxypropylether resulted in completely homogeneous Epon blocks which enabled us to cut 50 nm sections, free of holes and chatter. The biomaterials did not dislodge during the process of sectioning and the cell-polymer interface remained intact for electron microscopical evaluation.

The tetracyclines specifically inhibit mitochondrial protein synthesis when present at the same low concentrations as used for their antibacterial action. Inhibition of mitochondrial protein synthesis leads to decrease in the oxidative energy-generating capacity of cells. Therefore, the presence of tetracyclines may result in proliferation arrest. In the present study we show that continuous intravenous administration of polyvinylpyrrolidone (PVP) induces the formation of granulomas in the normal rat liver; the rats usually die within 2 weeks of continuous PVP treatment. Athymic (nude) rats appear to be more resistent to the deleterious effects of PVP as they survive the treatment for at least 5 weeks. Although the livers of the PVP-treated nude rats are heavily infiltrated with phagocytic cells, they seldom show granulomas. Reconstitution of nude rats with syngenic thymocytes leads, on the other hand, to extensive granuloma formation. Normal rats treated continuously with PVP plus doxycycline, however, all survive, their livers showing only a few very small granulomas and the normal low number of phagocytic cells. We conclude that the formation of granulomas induced by PVP is a process which is mediated by T-lymphocytes. Because doxycycline prevents this kind of granuloma formation it seems likely that doxycycline not only impairs the proliferation and differentiation of T-lymphocytes but also of monocytes and macrophages.