Eosinophilia-associated muscle disorders: an immunohistological study with tissue localisation of major basic protein in distinct clinicopathological forms.

AIMS: (a) To evaluate tissue eosinophil density, location of eosinophil cytotoxic products, histopathological muscle changes and inflammatory cell types in different eosinophilia-associated myopathies that are clinicopathologically heterogeneous. (b) To determine the immunohistological range of tissue eosinophil density in non-eosinophilic inflammatory myopathies. METHODS: Muscle biopsy specimens from seven patients with blood and/or tissue eosinophilia and clinicolaboratory myopathic signs (five chronic course myopathies, one subacute onset fasciitis/myositis, one acute myositis), and from 18 non-eosinophilic inflammatory myopathies, underwent routine staining, inflammatory infiltrate immunophenotyping, immunostaining for eosinophil major basic protein (MBP) and transmission electron microscopy examination. Eosinophil and total inflammatory cell counts were statistically analysed. RESULTS: Histological examination showed occasional or no infiltrating eosinophils in all cases. MBP staining showed that tissue eosinophil density and percentages in eosinophilia-associated myopathies were significantly higher than in idiopathic myositides. Extracellular MBP diffusion, the hallmark of eosinophil cytotoxicity, was recurrent on sarcolemma and endothelium. Electron microscopy showed eosinophils close to sarcolemma, abundant mast cells, and capillary endothelial swelling. Immunostaining detected a higher mean eosinophil density in idiopathic myositides than previously assessed histologically. CONCLUSIONS: MBP immunohistology on skeletal muscle, previously performed only for acute eosinophilic polymyositis, suggests that eosinophil-mediated injury of muscle cells may occur in a wider spectrum of less aggressive eosinophilia-associated myopathies than previously thought. As conventional histology is likely to underestimate this leucocyte subset, MBP staining may be a useful tool in the analysis of tissue infiltration of eosinophils as a possible treatment target.

Lymphatic vessels in human eyelids: an immunohistological study in dermatochalasis and chalazion.

We investigated lymphatic morphology and expression of endothelin (ET-1) axis molecules in human eyelids affected by an inflammatory state (chalazion) and an age-related degenerative condition (dermatochalasis). Lymphatics were immunohistologically detected by D2-40/LYVE-1 staining. Absorbing lymphatic vessels were localized in papillary dermis and around skin appendages with distinctive morphology. In chalazion, D2-40 reactive flattened lymphatic profiles were compressed by inflammatory infiltrate; in dermatochalasis, large fully opened lymphatics were observed, with a significantly wider total area (lymphatic lumina/200x field; p < 0.05). The lymphatic density (number/200x field) in the two groups was within the same range. Lymphatic dilation is possibly dependent on reduction and fragmentation of the dermal elastic network as well as of oxytalanic fibers in the papillary dermis of dermatochalasis, as shown by Weigert's reaction. Multifunctional peptide ET-1, involved in vasomotion, inflammation and connective proliferation, was faintly and discontinuously localized on lymphatics, as was its type A receptor. In contrast, the consistent expression of type B receptor indicates that lymphatic endothelium is a physiological target for ET-1, whose effects are modulated by multiple pathophysiological conditions. Thus, vasoactive factors play a role in the physiology of richly vascularized eyelids, and therefore, morphofunctional characterization of lymphatic vessels may be useful in suggesting treatment options.

Endothelin-1 and endothelin-converting enzyme-1 in human granulomatous pathology of eyelid: an immunohistochemical and in situ hybridization study in chalazia.

Endothelin-1 (ET-1), a potent vasoconstrictor peptide, is involved in several functions of eye pathophysiology, such as regulation of intraocular tension and retinal reactive vasoconstriction. As ET-1 pro-inflammatory and fibrosing activity is emerging in different fields of pathology, we investigated the expression of ET-1 and endothelin-converting enzyme-1 (ECE-1) in chalazia, granulomatous lesions of the eyelid. ET-1 and ECE-1 were analyzed by immunohistochemistry (IHC) in twenty surgically removed chalazia, with regard to expression in eyelid structures and inflammatory infiltrate. Phenotype of ET-1 expressing inflammatory cells was established by double immunofluorescence. The cellular localization of prepro-ET-1 (pp-ET-1) mRNA and ECE-1 mRNA was studied by nonisotopic in situ hybridization (ISH). Neutrophils (PMNs), macrophages and T-lymphocytes were scattered in stroma, around alveoli and grouped in lipogranulomas. PMNs, macrophages, basal epithelium of meibomian adenomers and central ducts immunostained for ET-1. ECE-1 protein was found in meibomian adenomers, conjunctival epithelium, tarsal mucous glands and in inflammatory cells. Hybridization signals for pp-ET-1 mRNA and ECE-1 mRNA were recognized in healthy and degenerating meibomian ducts, adenomers, inflammatory cells, as well as in vessel walls. ECE-1 mRNA was also present in conjunctival epithelium and Henle's crypts. Our findings suggest that the multifunctional peptide ET-1 may have a role in molecular genesis of tissue damage in chalazia. ET-1 cytokine activity is likely to support the migration of inflammatory cells and the setting of lipogranulomas; ET-1 stimulation might contribute to proliferation of fibroblasts and collagen synthesis. ET-1 upregulation on meibomian adenomers and ducts may further enhance granulomas formation by stimulating lipid release.

Lymphatic vessels in human sural nerve: immunohistochemical detection by D2-40.

Lymphatics were detected in the epineurium of the human sural nerve by D-240 immunostaining and confirmed by ultrastructural examination.

Influences of maternal weight on the secondary sex ratio of human offspring.

BACKGROUND: Less than optimal reproductive conditions may be associated with a secondary sex ratio biased towards females. Body weight represents a critical determinant of reproduction. Accordingly, we evaluated whether preconception body weight and weight gain during pregnancy influence the sex ratio of human offspring. METHODS: A retrospective study was performed on 10 239 births in 1997-2001. Time of conception, preconception body weight and net weight gain during pregnancy were obtained for 9284 pregnancies. RESULTS: The secondary sex ratio of mothers in the lowest quartile of pre-pregnancy body weight (<54.6 kg) was lower than that of the other three quartiles (0.497 versus 0.525; P < 0.01). In contrast, the sex ratio of children born by the women in the highest quartile of weight gain during pregnancy appeared lower than that of the first three quartiles (0.493 versus 0.516; P = 0.054). CONCLUSIONS: A low pre-pregnancy weight and a greater weight gain during pregnancy are both associated with a reduced secondary sex ratio. These data indicate that in women with non-optimal reproductive/metabolic conditions, a greater attrition is exerted on male than on female offspring.

Evaluation of biocompatibility of metallic dental materials in cell culture model.

Biocompatibility of metals for dental use was tested using a three-dimensional model consisting of oral keratinocytes cultured on de-epidermised sub-mucosa. The toxicity of orthodontic metallic wire and soldering material was assessed through parameters such as the morphology and growth rate of the keratinocytes, as well as by classical histology, scanning electron microscopy (SEM) and transmission electron microscopy (TEM). The sharp composition of metallic wires and soldering materials was assessed by Auger Electron Spectroscopy (AES). The results of our experiment showed that the new model revealed inhibition of keratinocyte growth and stratification near soldering material, whereas mucosal cells were able to grow and layer out on dental wire. It is concluded that this experimental model, which simulates the oral environment, is useful for studying the effects of materials for dental use for its sensibility and reproducibility. Moreover it can provide morpho-functional information which cannot be achieved by traditional methods.

Early outcome of myomectomy by laparotomy, minilaparotomy and laparoscopically assisted minilaparotomy. A randomized prospective study.

BACKGROUND: To compare in the first 7 days after surgery the outcome of myomectomy performed by three laparotomic approaches: laparotomy (LT), minilaparotomy (MLT) and laparoscopically assisted minilaparotomy (LA-MLT). METHODS: Fifty-one women with 5-15 cm total myoma diameter were randomized blindly using a computer randomization list, to LT (n = 17), MLT (n = 17) or LA-MLT (n = 17). RESULTS: Mean operation length was similar in the three groups. Mean (+/- SEM) time of paralytic ileus (55.0 +/- 4.5 versus 33.4 +/- 3.4 h; P < 0.01) and discharge (141.6 +/- 5.2 versus 81.5 +/- 8.2 h; P < 0.01) was longer in LT than LA-MLT or even MLT. In comparison with LA-MLT, LT induced a greater haemoglobin decline (-3.07 +/- 0.3 versus -1.8 +/- 0.15 mg/dl; P < 0.025), and a greater post-operative stress, as documented by increased prolactin (+15.1 +/- 3.8 versus +0.16 +/- 4.5 ng/ml; P < 0.03) and decreased insulin sensitivity (fasting glucose/insulin; -7.5 +/- 2.6 versus -0.7 +/- 2.1; P < 0.02). Seven days after surgery, abdominal pain (P < 0.05) was higher after LT (3.0 +/- 0.6) than MLT (0.5 +/- 0.2) and LA-MLT (0.9 +/- 0.4). CONCLUSIONS: In selected cases, myomectomy by LA-MLT offers some advantages versus LT and, to a smaller extent, MLT.

The male disadvantage and the seasonal rhythm of sex ratio at the time of conception.

BACKGROUND: In accordance with a presumed greater fragility of male versus female pregnancies, we tested whether sex ratio (male/female ratio) of vital pregnancies is higher in seasons more favourable for reproduction. METHODS: A retrospective study was performed on 14,310 births which had occurred in our institute between 1995-2001. For each single pregnancy the time of conception was calculated by the last menstrual period recall and confirmed or redefined by ultrasound in 95.8% of cases. The sex ratio of 199,454 pregnancies which had occurred in the Modena County between 1936-1998 was also stratified according to the month of birth. RESULTS: Sex ratio of institutional deliveries was 0.511 and was identical to that obtained from the County registry. Sex ratio at birth did not show a significant seasonal variation. By contrast, sex ratio calculated at time of conception showed a seasonal rhythm, with amplitude of 2.4% and peak values in October (confidence interval: +/-43 days). The rhythm was in phase with the rhythm of conception that showed peak values in September (confidence interval: +/-37 days) and an amplitude of 7%. CONCLUSIONS: The superimposition of the phase of sex ratio and conception rhythms supports the contention that more males than females are conceived in seasons with more favourable reproductive conditions.

18q-syndrome and ectodermal dysplasia syndrome: description of a child and his family.

The 18q- syndrome [MIM #601808] is a terminal deletion of the long arm of chromosome 18. The most common deletion extends from region q21 to qter. We report here a nine-year-old boy possessing a simple 18q- deletion who had abnormalities of the brain, skull, face, tooth, hair, bone, and skin, plus joint laxity, tongue palsy, subtle sensoneural deafness, mental and speech delay, attention deficit hyperactivity disorder (ADHD), tic, and restless legs syndromes. His karyotype was 46, XY, del (18)(q21.31-qter). The size of the deletion was approximately 45 cM. Most of these abnormalities were not explained by the 18q- deletion. The family pedigree suggested the presence of a subtle involvement of ectodermal and/or mesodermal structures. Karyotypes of the other family members were normal.

Case report. Fatal rhinocerebral zygomycosis due to Rhizopus oryzae.

A case of rhinocerebral zygomycosis due to Rhizopus oryzae, arising after trauma in a 53-year-old diabetic man, is reported. Diagnosis was based on histological and mycological examination. Fragments of the colonies were observed by scanning electron microscopy. This is the first case diagnosed in Tuscany.

Smooth muscle cells of the media in the dilatative pathology of ascending thoracic aorta: morphology, immunoreactivity for osteopontin, matrix metalloproteinases, and their inhibitors.

The etiopathogenesis of thoracic aortic aneurysms is currently an issue of debate. The present study investigated ultrastructural, morphometric, and immunohistochemical aspects of smooth muscle cells (SMCs) in chronic aneurysm of the thoracic aorta (aneurysm group), aortic dilatation associated with valvular disease (valvular group), and dissection of the thoracic aorta (dissection group). Fragments of the ascending aorta that had been taken from the patients during coronary bypass surgery were used as controls. No significant difference was observed in the density of SMCs between the 3 pathologic groups put together and the controls. Only separate analysis of SMC density in each of the pathologic groups showed that the valvular group samples had significantly smaller amounts of SMCs in the internal layer of the media than the dissection group samples and controls. Ultrastructural analysis, in situ end labeling, propidium iodide assay, and DNA laddering did not show apoptosis of SMCs in the samples investigated. Ultrastructure of SMCs characteristic of the synthetic phenotype, together with increased expression of osteopontin in the media of pathologic thoracic aortas indicated the transition of SMCs from the contractile to the synthetic phenotype. Immunohistochemical investigation showed that medial SMCs in the samples taken from aortas of all 3 pathologic groups expressed stronger immunoreactivity for matrix metalloproteinase 1, 2, and 9 and tissue inhibitor of metalloproteinase 1 and 2 than the controls. The present study shows that during the formation of aneurysms, dissection of the thoracic aorta, or aortic dilatation associated with valvular disease, loss of SMCs was not of great importance with respect to their transition from the contractile to the synthetic type in leading to increased production of matrix metalloproteinases.

Carbon dioxide therapy in the treatment of localized adiposities: clinical study and histopathological correlations.

The authors report their experience using carbon dioxide (CO2) therapy for the treatment of 48 female patients presenting adipose accumulations, located on the thighs, knees, and/or abdomen; a Carbomed Programmable Automatic Carbon Dioxide Therapy apparatus was used. In light of the effects of CO2 on the microcirculation recently described in the literature, we expected this gas, which we administered subcutaneously, to positively affect the physiological oxidative lipolytic process. The aim of our study was to evaluate the effect of this therapy on localized adiposities. As such, we describe the method we used and report the results observed in the areas treated (in terms of reduction in maximum circumference) as well as side effects. Furthermore, we assessed the effect of subcutaneous administration of CO2 on the microcirculation by showing changes in the Laser Doppler signal and in the concentration of transcutaneous oxygen tension (tcPO2). Pre- and posttreatment biopsies of tissues were performed in seven patients in order to study the changes induced by the use of CO2 on both adipose and connective tissues. All data obtained were statistically analyzed; values of P < 0.05 were considered significant.

Hemolytic drugs aniline and dapsone induce iron release in erythrocytes and increase the free iron pool in spleen and liver.

Incubation of rat erythrocytes with the hydroxylated metabolites of aniline and dapsone (4-4'-diaminodiphenylsulfone), phenylhydroxylamine and dapsone hydroxylamine, respectively, induced marked release of iron and methemoglobin formation. On the contrary, no release of iron nor methemoglobin formation was seen when the erythrocytes were incubated with the parent compounds (aniline and dapsone). The acute intoxication of rats with aniline or dapsone induced a marked increase in the erythrocyte content of free iron and methemoglobin, indicating that the xenobiotics are effective only after biotransformation to toxic metabolites in vivo. Prolonged administration of aniline or dapsone to rats produced continuous release of iron from erythrocytes. Marked iron overload was seen in the spleen and in the liver Kupffer cells, as detected histochemically. The spleen weight in these subchronically treated animals was significantly increased. The free iron pool was markedly increased in the spleen and to a lower extent in the liver. The possible relationships between iron release in erythrocytes, oxidative damage seen in senescent cells, hemolysis, overwhelmed capacity of spleen and liver to keep iron in storage forms and subsequent increase in low molecular weight, catalitically active iron is discussed.

A new model for studying differentiation and growth of epidermal cultures on hyaluronan-based carrier.

Here, a three-dimensional model based on fragments of human de-epidermized dermis (DED) is prepared in order to study the performance of a microperforated, hyaluronan-based membrane as a carrier of cultured epidermal cells. Hyaluronic acid is, in fact, considered to be an optimal biomaterial allowing proliferation of both keratinocytes and melanocytes, and it is already used for clinical aims. The carrier with subconfluent human epidermal cultures is positioned onto the DED and kept in culture until a new epidermis is formed. This model system allowed to study the migration and growth of human epidermal cells from the carrier, resembling 'in vivo' re-epithelization.

Case report. Cutaneous phaeohyphomycosis caused by Cladosporium oxysporum.

The case of a 66-year-old woman with Cushing syndrome and a 1-year history of papulo-nodular lesions on the right leg is reported. Biopsy revealed septate hyphae and yeast-like cells in granulomatous dermo-hypodermal lesions. Culture of biopsy fragments on Sabouraud glucose agar without cycloheximide produced colonies that were olive green on top and greenish black underneath. On the basis of microscope findings and scanning electron microscopy observation of fragments of colonies, a diagnosis of cutaneous phaeohyphomycosis due to Cladosporium oxysporum was made. The patient was initially treated with itraconazole, which led to clinical improvement, but mycological recovery was obtained after a course of ketoconazole, made necessary by the presence of pituitary adenoma. To our knowledge, this is the first reported case of subcutaneous phaeohyphomycosis due to Cl. oxysporum.

A case of acral Darier's disease.

A sporadic case of acral Darier's disease in a 20-year-old woman is reported. The disease was diagnosed on the basis of clinical, histological and ultrastructural data. Only few cases of exclusively acral clinical manifestations of Darier's disease have been described in the literature.

Two cases of cutaneous phaeohyphomycosis by Alternaria alternata and Alternaria tenuissima.

Two cases of cutaneous phaeohyphomycosis, one with a nodular appearance and the other with an erythematous infiltrating patch, are reported in immunocompromised patients. Diagnosis was based on histological examination, which revealed hyphae and round-shaped fungal cells in a granulomatous dermal infiltrate, and on identification of the moulds when biopsy fragments were cultured on Sabouraud-dextrose agar without cycloheximide. The pathogens were Alternaria tenuissima in the first case and A. alternata in the second. The fungi were examined by scanning electron microscopy. The patients were checked for bone and lung involvement and were then treated with surgical excision and itraconazole, and itraconazole only, respectively, with clinical and mycological resolution.

The behaviour of Sarcoptes scabiei var. hominis in human skin: an ultrastructural study.

The biology of Sarcoptes scabiei var. hominis is poorly understood because of the lack of an in vivo or in vitro propagation system. To obtain more information on the mite behaviour in its natural habitat we conducted an ultrastructural study of burrows in a number of patients with common scabies. Scanning electron microscopy furnished attractive images of the tunnel, parasite body and eggs architecture and demonstrated the presence of holes in the tunnel roof probably representing aeration structures. Transmission electron microscopy showed a marked keratinocyte damage around burrowing mites, well evident ahead of the mite capitulum also. Faecal pellets containing keratinocyte micro-organelles (melanosomes and mitochondria) were documented in posterior midgut. For the first time we disclosed the adhesion mechanism of eggs to the burrow floor. We showed that the typical finger-like projections of the outer layer of the egg shell gradually disappear where the eggs are in contact with the tunnel floor. This allows the inner layer of the egg shell to fuse and stick with the damaged keratinocytes lining the tunnel floor. Our observation substantiates that Sarcoptes scabiei produces a proteolytic substance (salivary secretions?) that has a key role in its life cycle allowing burrowing, feeding and eggs-burrow adhesion.