Effect of hypothyroidism on postnatal conjunctival development in rats.

AIM: To study the conjunctival development in hypothyroid genetically epilepsy-prone rats (GEPRs) with serum T3 and T4 significantly lower than in normal rats. METHODS: A structural, ultrastructural and histochemical study on the conjunctival epithelium of GEPRs and of control Sprague-Dawley (SD) rats before and after eyelid opening, with particular regard to goblet cell differentiation. RESULTS: From birth to day 12, no goblet cells were demonstrated on the conjunctival surface of both strains, so that the epithelium was formed only by a cuboidal basal layer and by a superficial layer of roundish or flattened cells. On day 16, after the eyelid opening, Alcian blue (AB)-positive goblet cells filled with homogeneous granules were demonstrated isolated, in GEPRs, or clustered, in SD rats, in both the fornices and palpebral conjunctiva. The epithelium showed a basal layer and many layers of flattened cells and was taller in SD rats (8-10 layers) than in GEPRs (6-7 layers). At 3 months, the epithelium in SD rats was higher with generally clustered goblet cells, whilst in GEPRs goblet cells were both isolated or clustered. In both strains, the goblet cells showed a marked AB/periodic acid-Schiff positivity all over the conjunctival surface and were filled with granules of different density. In both strains, goblet cells were absent at birth and their appearance, as AB-positive cells, was concomitant with eyelid opening. CONCLUSIONS: Hypothyroid rats showed a conjunctival development different than that of normothyroid rats for both epithelial and goblet cells. It appears that thyroid hormone imbalance may influence conjunctival development.

Human galectin-3 immunoexpression in thyroid follicular adenomas with cell atypia.

Human galectin-3 (hgal-3) is a beta-galactoside binding protein involved in a number of physiological and pathological processes. Increasing hgal-3 immunoexpression has been reported in several human tumors, including thyroid carcinomas, but not in benign thyroid lesions. We analyzed the immunolocalization of hgal-3 in cell compartments of benign and malignant thyroid lesions. Hgal-3 immunoperoxidase reaction was carried out on 133 thyroid tissue samples obtained from 113 patients; 20 of these were normal (NT), 85 were benign thyroid lesions [20 colloid nodules (CN), 21 nodular hyperplasias (NH), 7 focal lymphocytic thyroiditis (FLT), 15 Hashimoto's thyroiditis (HT), 22 follicular adenomas (FA)], 25 differentiated carcinomas [15 papillary carcinomas (PC), 6 follicular carcinomas (FC) and 4 Hürthle cell carcinomas (HC)] and 3 anaplastic carcinomas (AC). Among the malignant thyroid lesions, hgal-3 was detected in 12/15 (80%) PC, 3/4 (75%) HC and in 4/6 (66.6%) FC, but in none of the 3 AC. Conversely, hgal-3 immunoexpression was absent in NT and in all benign thyroid lesions, but 1/15 HT and 10/22 (45.4%) FA. In the latter, hgal-3 was mostly expressed in microfollicular areas and in five of the six atypical FA. Hgal-3 cytoplasmic-perinuclear immunolocalization was observed in the majority of thyroid carcinomas and in more than half of the FA, theoretically suggesting an involvement of this protein in thyroid tumorigenesis throughout an antiapoptotic activity. Moreover, hgal-3 expression in FA might anticipate the likelihood of evolution of these benign lesions towards malignancy.

Effect of stress on the paracellular barrier in the rat ileum.

BACKGROUND: Restraint stress induces permeability changes in the rat small intestine but little is known of the ultrastructural events leading to defects of the paracellular sealing or of the short term evolution of these alterations. METHODS: In the present study, we performed transmission electron microscopy in the terminal ileum perfused with lanthanum after two hours of immobilisation stress and in non-stressed control rats. Moreover, immunohistochemistry of the tight junction (TJ) associated proteins, occludin and zonula occludens 1 (ZO-1), was carried out together with western blot analysis of the transmembrane protein occludin. TJ morphology was also assessed after a 22 hour recovery period. RESULTS: Immobilisation stress induced a significant increase in epithelial permeability to the lanthanum tracer (p<0.005) which recovered completely after 22 hours. Compared with unstressed controls, in stressed rats no differences were found on freeze fracture analysis. The TJ related immunofluorescence signals of occludin and of ZO-1 were irregularly distributed in stressed rats after two hours but returned to a normal pattern at 24 hours although with minor intensity. No quantitative alterations in occludin were detectable in stressed rats by immunoblot whereas a perinuclear concentration of occludin was observed by immunolocalisation. CONCLUSIONS: Immobilisation stress induced an increase in TJ permeability in the rat terminal ileum. These changes were mainly due to modifications and redistribution of the TJ transmembrane protein occludin and of the plaque protein ZO-1 whereas protein synthesis, at least that of occludin, was not affected by stress.

Myofibroblasts in adolescent varicocele: an ultrastructural and immunohistochemical study.

Myofibroblasts of the testes play an important role in the morphofunctional integrity of the seminiferous tubule. Previous studies in adults with varicocele have demonstrated an involvement of this cell population that tends to transform into fibroblasts. The aim of the present study was to try and verify the morphological features of myofibroblasts in the adolescent with left idiophatic varicocele. Twenty-two testicular biopsies were obtained from adolescents (aged 13-18 years, mean 15.8) and operated for left idiophatic varicocele. Biopsies were processed for electron microscopy (TEM) and immunofluorescence studies. The latter determined the level of myofibroblasts by using alpha-smooth muscle isoactin, a specific marker of myofibroblasts and, hence, excluding fibroblasts. TEM observations revealed a normal ultrastructure of myofibroblasts that was similar to that for the controls but an increased presence of extracellular matrix. The immunofluorescence study always demonstrated strong cell positivity to anti alpha-smooth muscle isoactin as also seen in the controls. This study demonstrates that adolescents with varicocele have well preserved myofibroblasts and do not show any evidence of transformation into fibroblasts, this has already been demonstrated in adult varicocele. These observations could represent an important factor for the understanding of the reversal of growth failure of the testes observed after early treatment.

Morphological changes induced by prolonged darkness in the retinal pigment epithelium of the turtle (Pseudemys scripta elegans).

The retinal pigment epithelium of Vertebrates was shown to be sensitive to cyclic oscillations of light and darkness. The morphological changes induced by prolonged darkness on the retinal epithelial cells of the freshwater turtle were studied, with particular regard to their localization and to their reversibility if animals are recovered under cyclic light. The eyes were processed for light and electron microscopy and a morphological and morphometric analysis was performed on the specimens. After 7 days of prolonged darkness, the vitreal extremity of some epithelial cells was partially detached; on the basal zone the infoldings were missing and vesicles and tubules, often arranged in rows, were observed. After 30 days of prolonged darkness, partial or complete double layers of epithelial cells were present: the superficial layer was connected, by means of the apical fringes, to the photoreceptors, whilst the deepest layer showed vesicles and tubules on its basal zone. After 7 days of recovery to L:D = 12:12, no cyclic activity was demonstrated and only occasional double layers of cells were present; on the basal surface isolated basal infoldings were present where two adjacent cells were joined together. It could be concluded that the detachment of the apical part of some cells, rapidly covered by the lateral sliding of the adjacent cells, and the substitution of the basal infoldings with vesicles and tubules could represent the morphological response of the retinal epithelium to the functional changes induced by prolonged darkness.

Morphological changes of the conjunctival epithelium in contact lens wearers evaluated by impression cytology.

PURPOSE: Conjunctival changes induced by rigid, gas-permeable and soft contact lenses (CL) were investigated using impression cytology. A scoring system based on seven parameters (specimen cellularity, cell-to-cell contacts, nucleus/cytoplasm ratio, chromatin, goblet cell distribution, keratinisation, inflammatory cells) was used to evaluate the morphological results. METHODS: One hundred and seventy-two CL-wearing eyes and 40 control eyes were examined. The population was divided into three groups: group 1, control eyes; group 2, asymptomatic CL-wearing eyes; group 3, CL-wearing eyes with intolerance problems. Impression cytology was carried out and all specimens were evaluated according to a scoring system. RESULTS: Group 1 specimens always showed normal features in the conjunctival epithelium. In group 2, rigid and gaspermeable lenses produced greater changes in conjunctival morphology than soft lenses, especially as regards cellularity, nucleus/cytoplasm ratio, goblet cell distribution and total score. On the contrary, in group 3 wearers of soft lenses showed higher partial and total scores than wearers of gas-permeable and rigid lenses. CONCLUSIONS: In asymptomatic CL wearers conjunctival morphology was better preserved in wearers of soft lenses. In patients with intolerance problems, wearers of soft lenses showed the worst cytological features. Impression cytology, evaluated with this scoring system, can be used to demonstrate epithelial damage occurring in CL-wearing patients.

Ultrastructural study of the conjunctival epithelium in the Mongolian gerbil (Meriones unguiculatus).

The conjunctival epithelium of the Mongolian gerbil (Meriones unguiculatus) was studied using the transmission and scanning electron microscopy. The superficial cells of the conjunctival epithelium were isoprismatic in shape and were covered, on their luminal surface, with microplicae. They were connected with the adjacent cells by junctional complexes and desmosomes. Some of the superficial cells were partially or completely detached from the underlying intermediate cells: this morphological pattern was interpreted as an expression of cellular renewal of the conjunctival surface. Goblet cells were either isolated or gathered in clusters: in any case, they were firmly connected with the adjacent epithelial cells. The apical part of the goblet cells was covered with a great number of long microvilli: they showed a variety of morphological aspects, which were interpreted as occurring in a sequential pattern during the secretory process. The Mongolian gerbil can be considered among the laboratory animals used for ophthalmic research.

Morphological and morphometric analysis on the rabbit conjunctival goblet cells in different hormonal conditions.

An altered conjunctival mucous secretion was reported in pregnancy or oral contraceptive use. Four groups of rabbits (males, dioestrous females, oestrous females, pregnant) were studied to determine whether sex and/or different physiological conditions could influence conjunctival goblet cells structure and ultrastructure. Light microscopy, transmission electron microscopy, and morphometry were performed. In males and in oestrous females the intracytoplasmic secretory granules were filled with granular material, whilst in pregnant and dioestrous females the granules were formed by a more homogenous and dense secretory material. The number of goblet cells was not statistically different in the groups studied, whilst pregnant animals showed the largest mean diameter. As to the secretory granules, their mean area was larger in dioestrous females, whilst their optical density was highest in pregnant animals. These observations indicate that the morphology of conjunctival goblet cell may vary according to sex and to different physiological conditions: this may account for the peculiar mucous secretion demonstrated during pregnancy.

Morphological differentiation of the conjunctival goblet cells in the chick (Gallus domesticus).

BACKGROUND: These is no consensus in the literature regarding the differentiation of conjunctival goblet cells in vertebrates. METHOD: The conjunctival epithelium of the chick was studied before and after hatching in order to demonstrate the morphological evolution of the goblet cells. The entire conjunctiva was processed for light microscopy either on semithin sections stained with toluidine blue-pironine or on traditional sections stained with Alcian blue pH 2.5-PAS. RESULTS: It was possible to demonstrate that goblet cells underwent remarkable changes in their secretory activity. At 12 h after hatching, isolated Alcian blue-positive cells were present in the fornix. At 24 h after hatching, cells positive for both Alcian blue and PAS were scattered among epithelial cells. Two days after hatching, cells which reacted positively only to PAS were also present. CONCLUSION: It is suggested that the differentiation of conjunctival goblet cells occurs first in the fornix, probably due to the particular vascular environment of this region, and then spreads all over the conjunctiva.

Impression cytology of the conjunctival epithelium in patients with vernal conjunctivitis.

The alterations in the conjunctival epithelium during the course of vernal conjunctivitis were examined by conjunctival impression cytology. The study was carried out on 53 patients with vernal conjunctivitis and 20 normal subjects as control. The results of impression cytology demonstrated that all cytological parameters were significantly modified in vernal conjunctivitis patients; the earliest alterations were found in the distribution of goblet cells, in the intercellular junctions, in the chromatin morphology and in the degree of keratinisation. The morphometric comparison showed that in vernal conjunctivitis patients the mean number of goblet cells per field was significantly higher than in controls. Moreover the mean diameter of goblet cells was smaller in patients than in controls. Impression cytology can, therefore, be a simple, non-invasive and cheap method for the study of the ocular surface in vernal conjunctivitis.

Embryological considerations on a multiple vascular anomaly in a child.

Coiling and kinking of the internal carotid artery are considered of remarkable importance in the pathogenesis of cerebral ischaemic episodes, especially in adults. The authors describe the case of a seven months old infant who underwent aortic and bilateral cerebral angiography because of severe neurological symptomatology. Besides serious malformations of the aorta and major vessels, both coiling of the left internal carotid artery and kinking of the right internal carotid artery were found. The arterious anomalies of the aortic branches are probably due to the deranged development of the two subclavian arteries and, in particular, of the right one. Owing to the patient's age, the carotid alterations are also consequent to an abnormal development of the arterious district deriving from the primitive aortic archs, especially the third arch.

[The synaptic ribbons in the retina of the turtle (Pseudemys scripta elegans): ultrastructural and chronobiological study]. / I nastri sinaptici nella retina della tartaruga (Pseudemys scripta elegans): indagini ultrastrutturali e cronobiologiche.

The Authors investigate the morphology and the behaviour of the synaptic ribbons (NS) in the outer plexiform layer of the retina of the turtle Pseudemys scripta elegans during the 24 hrs cycle; the animals were subjected to a light-dark cycle = 12:12 hours. New ultrastructural features are demonstrated in the rod arciform density and in the fibers connecting the central axis of the ribbon with the presynaptic membrane (Figs. 1 A-B). As to chronobiological data, the existence of different kinds of ribbons, each typical of a single period of the 24 hrs cycle is excluded: identical NS can likewise be observed at different hours of the day and under various environmental light stimulations (Figs. 2-3-4). Furthermore, it is also demonstrated that during the light period, every rod synapse in the outer plexiform layer has a typical ultrastructural topography of its own (Figs. 5 A-B-C). During the dark period, the ribbon with its neighbouring vesicles moves towards the nucleus, thus leaving the synaptic region (Figs. 6 A-B-C). From the analysis of our results, we are of the opinion that all previously demonstrated changes in number are only in part dependent on cyclic disruption and reconstruction. On the contrary, they can be the result of a different topographical arrangement of the organelles. When NS modify their relationship with the presynaptic membrane of the rods, they may behave as switch on the nervous circuits of the retina and play a facilitatory or inhibitory role on the discharge of synaptic vesicles during the different periods of the 24 hrs cycle.

[Post-natal maturation of the retina in the albino rat. I. The pigment epithelium]. / Maturazione post-natale della retina nel ratto albino: I. L'epitelio pigmentato.

The Authors studied the postnatal development of the retinal pigment epithelium in the albino rat, in order to elucidate its morphological and functional evolution, correlated to the numerous functional roles played in Vertebrates (Scheme 1). At birth, epithelial cells show few cytoplasmic organules and the apical surface provided of small depressions. From the third to the fifth postnatal day the first apical microfolds surround the depressions. From the seventh to the ninth day inner segments develop, whilst the apical surface of the epithelial cells is covered by many finger-like microfolds. During the eleventh postnatal day the buds of the outer segments and many lamellar microfolds can be demonstrated. During the sixteenth day the retina reaches its adult morphology. It is therefore well-evident that birth, similarly to many other Vertebrates, is not the last step, but only a moment, in the development of the retina: this process is completed only during postnatal life, when environmental light is able to stimulate every ocular structure.

[Morphological adaptations of the eyes of vertebrates: retinal trophism and the response to environmental stimuli]. / Adattamenti morfologici nell'occhio dei Vertebrati: trofismo retinico e risposte agli stimoli ambientali.

The results of many investigations on the morphology of the eyes of Vertebrates are proposed, comparing our personal data with the literature available on this argument. It is firstly taken into account the retinal nourishment: it takes place according to direct and indirect mechanisms (Scheme 1). The former (Scheme 2) consist of intraretinal blood vessels and are particularly rare among the Vertebrates; the latter are more numerous and can be classified as constant (choriocapillaris) (Schemes 3A, B), if they are present in all the species up today investigated, or unconstant [Müller cells increased in their number and size (Scheme 5A), papillary cone (Schemes 5B, 8B), membrana vascularis retinae (Schemes 6A, C), falciform process (Schemes 6B, 7A, B, 8B), pecten oculi (Schemes 8A, B)], if they can be demonstrated only in some species, even if belonging to different classes. Their structural and ultrastructural organization, likewise their embryological processes, are examined in detail. The effects of the cyclic changes of light and darkness during a 24 hrs period on the eyeball are then examined; quantitative and qualitative modifications of the mitochondria (Schemes 9A, B) and of the synaptic ribbons (Schemes 10A, B) in the outer plexiform layer, of the photosensitive disks of rods and cones, and of the secretory cells and of the excretory ducts of the Meibomian tarsal glands (Schemes 11A, B, 12A, B) are described. If the animals are exposed to prolonged darkness, no structural changes can be demonstrated; nevertheless, a dark environment can induce the differentiation of peculiar structural specializations, such as the retinal or choroidal tapetum lucidum (Schemes 13, 14, 15). On the contrary, the light, if prolonged or of high intensity, is able to induce irreversible lesions on the photoreceptors and on the pigment epithelium, according to similar mechanisms in all the animals up to now investigated (Scheme 16). In conclusion, it is confirmed the peculiar structural complexity of the eyeball in all the classes of Vertebrates, due to adaptation to the various environmental requests and developed either during the evolutionary processes or during the different steps of the life of a single animal.

The biological evolution of the Leeds-Keio ligament in the human knee. A histological and ultrastructural study.

The authors report their experience in the reconstruction of the anterior cruciate ligament using the Leeds-Keio ligament in 12 patients, 7 by open arthrotomy and 5 by an arthroscopic technique. The biological evolution of the new ligament was studied macroscopically and histologically in the first 6 cases treated by open arthrotomy. Arthroscopic monitoring and biopsy was carried out 4 to 10 months after operation. In 5 cases the new ligament appeared to be intact and histologically vital, with proliferation of new tissue along the new ligament. There were broken filaments of dacron hanging from the intercondylar cavity in only one case. The first macroscopic, histological and ultrastructural results show that despite a few limitations the Leeds-Keio ligament has the features required for replacement of the anterior cruciate ligament, although it will be necessary to wait 4 or 5 years before fully evaluating the durability and function of the ligament and the best operative method.

Effects of a stable analogue of PGE2 (11-deoxy-13,14-didehydro-16 (S)-methylester methyl PGE2: FCE 20700) on the secretory processes of conjunctival goblet cells of rabbit.

The effects of the instillation of a PGE2-analogue (11-deoxy-13,14-didehydro-16 (S)-methyl PGE2 methylester: FCE 20700) in the conjunctival sac of the rabbit were studied by means of two methods. The former is a clinical study (Dohlman test), the latter is a morphological investigation (semithin sections) on specimens of the conjunctival mucosa. From both methods it was possible to demonstrate that the FCE 20700 instillation enhanced the mucous lacrimal secretion in rabbits.