bcl-2 protein expression in cervical intraepithelial neoplasia: no evidence of a prognostic significance in mild and moderate lesions.

BACKGROUND: The bcl-2 proto-oncogene codes for a protein which appears to block apoptosis. In our study, we examined bcl-2 protein expression in cervical squamous metaplasia, cervical intraepithelial neoplasia (CIN) and microinvasive squamous carcinoma with the aim of identifying a relationship between bcl-2 protein expression and neoplastic development and progression. MATERIALS AND METHODS: Cervical bioptic samples were obtained from 86 white women, selected consecutively from our Colposcopic Service from January 1993 to June 1994, because of abnormal pap- smear suspicious for cervical dysplasia and/or human papilloma virus (HPV) infection. Upon histologic evaluation, 41 women had CIN, 23 cervical condyloma, and 22 squamous metaplasia. Ten patients with microinvasive squamous carcinoma, matched for age and demographic characteristics, were selected from our series of invasive cervical carcinomas and immunohistochemically analyzed. The expression of primary tumor bcl-2 protein was immunohistochemically evaluated by antihuman bcl-2 monoclonal antibody (diluted 1:100, Dako, Copenhagen, Denmark) on formalin-fixed paraffin-embedded tissue. Positive staining was expressed as a percentage of positive cells per 1000 counted dysplastic cells for each case. RESULTS: Bcl-2 immunostaining was found in all the 22 squamous metaplasias, limited to the basal layer. Nineteen of the 41 CINs (46%) were bcl-2 immunoreactive, and 2 of the 10 microinvasive carcinomas (20%). By analysing CIN lesions, the bcl-2 protein showed a striking increase in the rate of positivity with increasing severity of CIN; the bcl-2 protein expression in CINs III was significantly higher than for CINs I, CINs II or microinvasive carcinomas (P = 0.03, P = 0.02, and P = 0.03 respectively). No relationship was observed between bcl-2 immunostaining and HPV infection. bcl-2 protein expression was not useful for predicting CIN I and II evolution, although the rate of persistence/progression was higher in bcl-2 positive dysplasias (7 of 9 cases, 78%) than in negative ones (13 of 21 cases, 62%) (p = 0.88). CONCLUSIONS: Based on these results, it seems possible that the increase in bcl-2 expression in higher grade of CINs implies an increasing protection against programmed cell death, but also the induction of genetic instability in dysplastic epithelial cells and a greater capacity to evolve into invasive carcinoma.

In vitro study of gingival fibroblasts from normal and inflamed tissue: age-related responsiveness.

The aim of this study was to characterize some phenotypic expressions of fibroblasts from the human oral mucosa. Gingival and lower forearm fibroblasts from young (20-30 years) and elderly (> 60 years) subjects were analyzed. Gingival fibroblasts were taken from donors with (P) and without (NP) periodontal disease, while skin biopsies were taken from healthy subjects. Cell proliferation was assessed by evaluating the cell multiplication coefficient (C.M.C.). The proliferation potential of gingival fibroblasts from elderly individuals with and without periodontopathy did not differ from that of young subjects in the same condition but differed significantly in the skin samples. Enzyme neutral endopeptidase (EC 3.4.24.11) (NEP) activity, studied as a possible marker of cell ageing, showed an age-related increase in human skin fibroblasts but not consistently in gingival fibroblasts from individuals with or without periodontal disease. Cell area and substrate adhesion were evaluated by morphometric analysis. There were no significant differences between elderly P and NP subjects, while significant differences were observed between young and elderly P subjects. In conclusion, proliferative capacity and NEP activity in gingival fibroblasts did not appear to be age-related, probably because their microenvironment is continually moistened by saliva, which continues to contain growth factors, notably EGF, even into senescence. Tissue reaction and repair are important clinical and therapeutic implications.

Immunohistochemical evaluation of type IV collagenase (72-kd metalloproteinase) in prostatic intraepithelial neoplasia.

The aim of this study was to investigate the expression of type IV collagenase (72-kd metalloproteinase, MMP-2) in prostatic intraepithelial neoplasia (PIN) in relation to normal prostate (NP) and prostatic adenocarcinoma (PAc). Twenty formalin-fixed, paraffin-embedded prostatectomy specimens, in which NP, PIN and PAc were present, were immunohistochemically examined. The NP ducts and acini not contiguous with PIN and PAc showed slight MMP-2 immunostaining in the secretory cells, with some increase in intensity at the apical border, and moderate to strong immunoreactivity of some basal cells. In NP adjacent to PIN and PAc, rare ducts and acini showed strongly stained cells either isolated or in small groups of two, located within the thickness of the epithelium, close to the basement membrane. In the majority of PIN ducts and acini, the stratified secretory cells showed moderate staining. Most of these ducts and acini also showed strongly stained cells, which were mostly isolated, and either in contact with the basement membrane or scattered among the secretory cells. Low and high grade PIN showed some difference in the frequency of dark cells, which were more numerous in the latter. A small group of neoplastic acini adjacent to high grade PIN (early invasive adenocarcinoma) was observed in one of the 20 cases. Intense immunostaining was present in the acini originating from the PIN lesion. MMP-2 immunostaining of PAc was heterogeneous in intensity and location. Cribriform and solid/trabecular PAc showed weak cytoplasmic immunostaining; both moderately and intensely stained cells were seen in the cell layer adjacent to the stroma, intense immunostaining was shown by small clusters of neoplastic cells or single neoplastic cells located in the stroma. In acinar PAc, weak cytoplasmic immunostaining for MMP-2 was seen throughout most areas of the tumours, whereas moderately and intensely stained cells were observed less frequently than in cribriform and solid/trabecular adenocarcinoma. Intense immunostaining of single or small clusters of neoplastic cells located in the stroma was occasionally observed and, as with cribriform and solid/trabecular PAc, mainly located towards the periphery of the tumour nodules. Occasional ducts and acini with PIN and foci of PAc were either completely negative or very weakly stained. In conclusion, MMP-2 immunostaining increases progressively from NP, through PIN, up to invasive PAc. These results directly support the hypothesis that increased expression of metalloproteinases is a marker of malignant conversion.

Structural features of latex gloves in dental practice.

The aim of this study was to define from a morpho-structural point of view, using scanning electron microscopy, the features of various types of disposable latex gloves commonly used in Italian dental practice (Biogel D, Trend, Pagni, J&J, Latechnics, Pehasoft, Bantex). None of the brands examined was free from morphological flaws; however, while in some of these only slight depressions were found (Biogel D, Trend), in others (Latechnics, Bantex) there was a marked lack of homogeneity in the latex structure or real holes (Pehasoft). This study emphasizes the current difficulties faced by dentists in the search for safe working conditions.

Human skeletal muscle mitochondria in aging: lack of detectable morphological and enzymic defects.

We have investigated structural and functional properties of skeletal muscle mitochondria obtained from biopsies from young and old individuals. The morphometric analysis of muscle sections revealed a tendency to an increase of total area, numerical density and volume density of mitochondria in the aged. The enzymatic activities of NADH-Coenzyme Q reductase, succinate cytochrome c reductase, ubiquinol-cytochrome c reductase exhibited a high variability of specific activities without any correlation with age. Expression of the values as enzyme turnovers reduced the variability but was unable to reveal any age-dependent modification.

The human umbilical vein in normal, hypertensive and diabetic pregnancies: immunomorphological and ultrastructural evidence.

Mother-fetus exchanges at the placental level are found to be altered in women affected by hypertensive or diabetic pregnancies following the onset of microenvironmental, circulatory, trophic or tissue disorders. Our aim was therefore to assess the alterations occurring within the umbilical cord, particularly its venous endothelial component and underlying smooth muscle layer, using transmission (TEM) and scanning electron microscopy (SEM) and immunohistochemical analyses. Immunohistochemical data appear to support the ultrastructural evidence for an activated state of these vascular structures, in both conditions (hypertension and diabetes). Furthermore, mainly during diabetic pregnancies, extracellular matrix molecules such as tenascin and fibronectin also quantitatively increase at the vein wall level. The umbilical cord seems to be a structure capable of responding actively to abnormal microenvironmental conditions which seriously threaten the health of the fetus and also the mother herself.

Fibroblast proliferation over dialysis membrane: an experimental model for "tissue" biocompatibility evaluation.

The present study reports on a biological model based on fibroblast proliferation applied to 3 different types of flat-plate dialysis membrane, in order to ascertain whether the artificial materials currently used in hemodialysis cause in vitro cellular proliferation. The study plan we followed involved plate membrane isolation from non-used dialyzers and used dialyzers, observed through scanning electron microscopy (SEM) both before and after testing with human fibroblasts by means of cell culture. Fibroblast growth was assessed by phase contrast light microscopy examination and cytometric DNA content evaluation. Our investigations proved that the artificial materials we considered interact with fibroblast cultures. Noticeable proliferative response was observed both after contact with unused material and on mediation by the protein layer absorbed on the membrane surface at the end of dialysis sessions. In this last case fibroblast proliferative activity appeared higher than that observed with unused membranes, showing that the soluble molecules entrapped in the protein layer appeared able to exert a biological activity even in vitro tests.

Osteoconductive properties of methylpyrrolidinone chitosan in an animal model.

Bone defects were surgically produced in the tibiae of rabbits and medicated with freeze-dried methylpyrrolidinone chitosan. Histological observations 60 d after surgery showed a considerable presence of neoformed bone tissue, as opposed to controls, originating from the pre-existing bone as well as from the periosteum. The cationic nature and the chelating ability of the methylpyrrolidinone chitosan apparently favoured mineralization. Endosteal-periosteal and bone marrow osteoblast-like precursors, stimulated by growth factors entrapped in the coagulum-polysaccharide mixture, gave rise to intramembranous bone formation. The ultrastructural examination evidenced that bone osteoid was followed by mineralization of the tissue.

Osteoconduction exerted by methylpyrrolidinone chitosan used in dental surgery.

Surgical wounds from wisdom tooth avulsions were medicated with freeze-dried methylpyrrolidinone chitosan, a gel-forming resorbable biopolymer obtained from crab chitosan by chemical modification. Methylpyrrolidinone chitosan promoted osteoconduction and the space left after avulsion was filled with newly formed bone tissue, which conferred desirable mechanical and physiological characteristics to the healed would site. Morphological evidence obtained from biopsies confirmed the radiographic data. Methylpyrrolidinone chitosan was progressively depolymerized by lysozyme and was no longer detected 6 months after surgery. None of the 10 patients reported adverse effects over one year of observation.

Placental villi-decidua interactions in normal and hypertensive pregnancies: a morphological quantitative study.

The decidual response to the implantation of the embryo is characterized by physical modifications to the uterine wall, with proliferation of the stromal cells which later change into decidual cells. We performed associated morphological and morphometrical studies to assess how the placental villi and decidua intersect, both in normal terminal pregnancy and in hypertensive patients in whom microenvironmental modifications induced by hypertension may cause significant alterations in mother-fetus relationships. In placentas of hypertensive women our morphometric analyses showed a higher number of chorionic villi-decidua interactions (p less than 0.05) with a more clumped distribution (p less than 0.05) and a smaller surface area of single interaction (p less than 0.001), in association with a higher number (p less than 0.005), and greater areas (p less than 0.01) of decidual cells. These data demonstrate how the placenta can enhance mother-fetus contacts impaired as the result of a hypertensive condition.

[Our experience in vascular trauma of the limbs]. / La nostra esperienza in tema di traumatologia vascolare degli arti.

Progress in the management of complex vascular injuries of the limbs has not eliminated the necessity for amputations. Physicians' failure to suspect vascular injury on admission in traumatized patients causes delay in the diagnosis and management of arterial injury that ultimately leads to limb loss. High index of suspicion, early recognition by clinical evaluation, routine use of Doppler flowmeter and broad indication of preoperative angiography result in prompt repair of these lesions in the majority of cases. In managing the lower popliteal or trifurcation vessels, distal radial and ulnar artery as well as nerve injuries, the Authors believe microsurgical techniques should always be employed.

Effects of chemical pretreatments on dentin bonding.

The aim of this study was to evaluate several chemical pretreatments on the shear bond strength of three dentin bonding agents (DBAs) and to try to correlate dentin morphology with bond strength using scanning electron microscopy (SEM). The DBAs tested were: Gluma Primer in association with Scotchbond DC, Scotchbond DC alone and Clearfil New Bond. Bond strengths of Scotchbond DC and Clearfil New Bond were not significantly modified or reduced by acidic dentin pretreatments. Gluma/Scotchbond adhesion was increased by several treatments but only maleic acid treatment produced shear bond strengths significantly higher than EDTA treatment. SEM evaluation of dentin treatments revealed a wide variety of morphological changes in the dentin surface with partial and complete removal of smear layer by acidic solutions, and only minor modification by amino acid solutions. Only maleic acid was capable of the complete removal of the smear layer and smear plugs coupled with extensive exposure of dentin collagen fibrils.

Reconstruction of parodontal tissue with chitosan.

Chitosan ascorbate, obtained by mixing chitosan with ascorbic acid and sodium ascorbate, was produced in a gel form suitable for the treatment of periodontitis according to current dental surgery. While chitosan ascorbate underwent degradation in vitro, especially in the presence of atmospheric oxygen and at pH 6.0, the protection from oxygen offered by the surgical cements and the physiological pH value permitted chitosan ascorbate to play an important biological role in vivo, where it kept a honeycomb structure, as indicated by SEM on biopsies taken on 10 patients. The proliferation and organization of the cells were thus favoured with a subsequent enhanced capability of reconstructing a histoarchitectural tissue. Chitosan was progressively reabsorbed by the host, with very satisfactory clinical recoveries of the 52 defects treated, for which tooth mobility and pocket depths were significantly reduced.

[Hypertension and syncytiotrophoblast: morpho-structural aspects]. / Ipertensione e sinciziotrofoblasto: aspetti morfo-strutturali.

Hypertension is a pathological condition that involves maternal fetal relationship. In hypertension placenta displays a syncytiotrophoblast plasmalemma with aspects of anomalous behaviour concerning Intramembranous Particles (IMP) and actin content of microvilli cytoskeleton. Decrease of syncytiotrophoblast microvilli IMP and microvilli actin further sustain the tendency of hypertensive placenta to show some features of immaturity that might deeply influence fetal-maternal exchanges during pregnancy associated with pathological status.

[Intrinsic potential of the gingival interdental epithelium and its "therapeutic" induction using brushing. Clinico-morphologic aspects]. / Potenzialità intrinseche dell'epitelio interdentale gengivale e loro induzione "terapeutica" mediante spazzolamento. Aspetti clinico-morfologici.

Toothbrushing technique may represent an important tool to improve gingival keratinization. Our experience evidenced a close relationship between this endoral therapy and interdental epithelial recovery of gingiva, after two months of treatment. Mechanical or microenvironmental stimuli and genetically determined potentialities are the main factors involved in this clinical-therapeutical recovery to modulate structural epithelial behaviour.

Root curettage and gingival repair in periodontitis.

Twelve patients, aged 35 to 60, having adult periodontitis were treated with scaling and root planning performed with ultrasonic and hand scalers. Biopsies were taken, gingival crevicular fluid and clinical parameters were analyzed at different times, and the healing process was monitored. Findings showed a clear correlation between the amount of crevicular fluid and the histological inflammatory pattern, whereas no relationship between crevicular fluid and gingival pocket depth was apparent. In this study, no differences were found in the histological features of the repair process triggered by either type of scaler.

Cytoskeleton and extracellular matrix of cutaneous vessels in inflammatory processes: immunomorphological study.

In pathological conditions, vascular modifications occur in various stages involving both vessel structure and adjacent extracellular matrix. The relationships between vascular cells and surrounding microenvironmental stroma are mediated by cytoskeleton. Our investigation showed a high number of vimentin- and actin-positive cells in the vascular cutaneous bed, mainly related to reactive vascularization phenomena, whereas vessel cells with a desmin-positive reaction were barely detectable. Furthermore, in newly formed vessels ultrastructure showed that basement membrane synthesis strictly depends on close contact between the endothelium and extracellular matrix. Our data give structural evidence of the close morphofunctional interactions existing between vascular cells and extracellular matrix.

Fibronectin distribution during lymph node development in guinea pig: an immunohistochemical study.

We carried out an immunohistochemical study on mesenteric guinea pig lymph nodes, from the 10th day prepartum till the 26th day postpartum, to assess the role of fibronectin in their organization during development. This glycoprotein is diffusely distributed in embryonic lymph nodes, suggesting a primer function during organogenesis. After birth, in fact, it is less widespread and is mainly localized around sinuses and vessels. Our data, supporting the important role of this glycoprotein during lymph node organization, are in agreement with the results obtained in other tissues and organs.