Opposite patterns of age-associated changes in neurons and glial cells of the thalamus of human brain.

In an autopsy series of 19 individuals, age-ranged 24-94, a relatively age-spared region, the anterior-ventral thalamus, was analyzed by immunohistochemical techniques to visualize neurons (neurofilament protein), astrocytes (glial fibrillary acidic protein), microglial cells (CD68) and amyloid precursor protein. The pattern of immunoreactivity was determined by surface fractal dimension and lacunarity, the size by the field area (FA) and the spatial uniformity by the uniformity index. From the normalized FA values of immunoreactivity for the four markers studied, a global parameter was defined to give an overall characterization of the age-dependent changes in the glio-neuronal networks. A significant exponential decline of the GP was observed with increasing age. This finding suggests that early in life (age<50 years) an adaptive response might be triggered, involving the glio-neuronal networks in plastic adaptive adjustments to cope with the environmental challenges and the continuous wearing off of the neuronal structures. The slow decay of the GP observed in a later phase (age>70 years) could be due to the non-trophic reserve still available.

PDGFR expression in differential diagnosis between KIT-negative gastrointestinal stromal tumours and other primary soft-tissue tumours of the gastrointestinal tract.

AIMS: To investigate the value of platelet-derived growth factor receptors (PDGFRs) by immunohistochemistry in discriminating KIT-negative gastrointestinal stromal tumours (GISTs) from other soft-tissue neoplasms of the digestive tract. METHODS AND RESULTS: One-hundred and sixty-seven primary gastrointestinal mesenchymal tumours (125 GISTs, 15 intra-abdominal desmoids, 12 leiomyomas, eight leiomyosarcomas, three schwannomas, two solitary fibrous tumours, and one case each of inflammatory pseudotumour and fibroid polyp) were reclassified based on morphology and on the immunohistochemical panel recommended by the National Institutes of Health consensus on GIST. All cases were then tested with antibodies specific for PDGFR alpha and beta. Of 125 GISTs, 117 were KIT-positive (93.6%) and eight KIT-negative (6.4%). All the KIT-positive GISTs were negative for both PDGFRs, while all the eight KIT-negative GISTs expressed PDGFR-alpha, with two of them also coexpressing PDGFR-beta. Among the 42 non-GIST tumours, only a small percentage (26.6%) of desmoids immunostained for PDGFR-alpha, two of them coexpressing PDGFR-beta. CONCLUSIONS: Immunostaining with PDGFR-alpha is a helpful marker in discriminating between KIT-negative GISTs and other gastrointestinal mesenchymal lesions: all KIT-negative GISTs were positive for PDFGR-alpha, while none of the other gastrointestinal mesenchymal tumours analysed, except a small subset of desmoids, was reactive with anti-PDGFRs. These preliminary data demonstrate the suitability of commercially available antibodies to detect immunohistochemically the mutually exclusive expression of KIT and PDGFR-alpha previously reported in GISTs by molecular biological techniques. Since PDGFR exists in the form of a homodimer (alphaalpha, betabeta) or heterodimer (alphabeta) and two of the KIT-negative GISTs coexpressed both PDGFR isoforms, further investigations are required to elucidate the role of PDGFR-beta in GISTs.

Apolipoprotein E polymorphism and breast carcinoma: correlation with cell proliferation indices and clinical outcome.

There is preliminary evidence that polymorphism of apolipoprotein E (apoE, protein; APOE, gene), one of the key regulatory proteins in cholesterol metabolism, influences the pathobiology of carcinoma of the colon, prostate and breast and also primary tumours of the brain. This study was designed to determine whether APOE polymorphism is related to variation in the rate of tumour cell proliferation and clinical outcome in carcinoma of the breast. One hundred and eleven infiltrating ductal carcinomas, for which follow up data were available, were included in the study. Estrogen and progesterone receptor status (ER, PR) cell proliferation index (MIB- 1) and APOE genotypes were determined from paraffin-embedded tissue by standard methods. Positive correlations were found between grade and tumour size, grade and presence of metastasis, grade and MIB-1 expression, as well as between ER and PR. Survival correlated inversely with tumour size and the presence of positive lymph nodes. Both steroid receptors correlated inversely with MIB- 1 expression. PR positive status also correlated inversely with high histological grade and presence of lymph node metastases. APOE allele frequencies resembled those of the general population. No significant associations were found between possession of either APOE epsilon2 or epsilon4 alleles and the parameters investigated. Although there is evidence to suggest that APOE epsilon4 may predispose to the development of carcinoma of the breast our data do not support the hypothesis that APOE genotype influences the rate of tumour cell proliferation or the clinical course.

Expression of p53 and bcl-2 in clinically localized prostate cancer before and after neo-adjuvant hormonal therapy.

The prognostic significance of p53 and bcl-2 expression in prostate carcinoma is currently under investigation. The aim of the present study was to analyze their expression in diagnostic biopsies and in prostatectomies performed after neo-adjuvant hormonal therapy to investigate their role in hormone resistance. One hundred and six patients with advanced prostate carcinoma were treated for 3 months with LHRH analogues before radical surgery. The expression of p53 and bcl-2 was analyzed by immunohistochemistry in all cases of prostatectomy and in available biopsies obtained before treatment, and was correlated with clinicopathologic parameters and follow-up. A significant increase in p53 expression was found following hormonal therapy, whereas no changes were observed in the expression of bcl-2. The increase in p53 did not correlate with the presence of therapy-induced morphological changes in prostate cancers, but it did correlate significantly with histologic grade and pathologic stage, biochemical progression of the disease, and short overall survival. At multivariate analysis, only grade and stage proved to be independent predictors of shorter survival. There were no correlations between bcl-2 and clinicopathologic variables whether in biopsies or in prostatectomies. The unfavorable clinical course associated with p53-positive carcinomas suggests that neo-adjuvant hormonal therapy may cause the selection of minor p53 mutated clones, rather than the induction of wild-type p53. In any case, the enhanced expression of p53 could label hormone-resistant cancers for further adjuvant therapy.

Loss of p21Waf1 expression is a strong predictor of reduced survival in primary superficial bladder cancers.

p21Waf1 is a downstream effector of p53 and belongs to the Cip1/Kip1 family of cyclin-dependent kinase inhibitors. Thus, it is a potential tumor suppressor gene and likely plays an important role in tumor development. Moreover, reduced expression of p21Waf1 has been reported to have prognostic value in several human malignancies. In this study, we evaluated the prognostic value of p21Waf1 in bladder cancer compared with other clinicopathological features and with p27Kip1 and p53 expression. A total of 96 superficial (pTa-1) human bladder carcinomas were immunohistochemically stained for p21Waf1 protein expression. Positive p21Waf1 staining (> or =5% positive nuclei) was observed in 68 of the 96 (71%) tumors. p21Waf1 expression was neither associated with tumor stage (P = 0.9) nor with tumor grade (P = 0.18) but was significantly associated with both p53 protein expression (> or =20% positive nuclei; P = 0.007) and with p53 gene mutations (P = 0.017). A significant correlation was also observed between positivity for p21Waf1 and high (>50% positive cells) p27Kip1 expression (P = 0.04). With regard to prognosis, patients whose tumors showed absence of p21Waf1 staining displayed a significantly shorter overall survival (P = 0.01 by log-rank test). However, p21Waf1 expression did not correlate with disease-free survival (P = 0.15 by log-rank test). On a multivariate analysis that also included p53 and p27Kip1 expression, negative p21Waf1 staining was an independent predictor of reduced overall survival (P = 0.004; relative risk, 5.32), stronger than age and tumor stage. These data indicate that expression of p21Waf1 protein strongly correlates with survival and might represent a useful prognostic marker in primary superficial bladder carcinomas.

Epithelioid sarcoma of the penis: a case report and review of the literature.

A rare case of epithelioid sarcoma (ES) of the penis is presented. The patient, a 35-year-old man, was initially treated as having Peyronie's disease, but the subsequent appearance of a subcutaneous nodule displayed a 'distal-type' ES. At immunohistochemical phenotypification, the tumor was positive for vimentin, cytokeratins and epithelial membrane antigen (EMA), as well as for some other multidirectional antibodies, including a membranous reaction for CD99. The review of 11 similar cases so far reported in the literature led to the conclusion that the clinicopathological characteristics of penile ES are basically the same as those of tumors in more classical locations: the age at diagnosis ranged from 23 to 43 years, the interval between first manifestations and diagnosis averaged 37 months (delayed diagnosis being common because of the slow growth rate and the harmless appearance of the lesion), the typical sign was a superficial nodule or mass, usually accompanied (better preceded) by urethral stenosis, dysuria and erectile disturbances. Total or partial penectomy was the treatment of choice in most patients, but lack of adequate follow up did not permit any definitive conclusion to be reached regarding its efficacy.

Apolipoprotein E polymorphism and central nervous system tumors: correlation with cell proliferation indices and clinical outcome.

AIMS: This study was designed to determine whether the polymorphism of apolipoprotein E (apoE), one of the key regulatory proteins in cholesterol metabolism, is related to varying susceptibility to central nervous system (CNS) neoplasms, and to evaluate any possible interaction between this polymorphism and tumor cell proliferation or clinical outcome. METHODS AND RESULTS: 53 CNS tumors were selected. Follow-up and survival data were available for 36 patients. ApoE genotypes and cell proliferation indices (nucleolar organizer regions, MIB-1, PCNA, p53) were determined from paraffin-embedded tissue by standard methods. Each of the indices of cell proliferation correlated positively with tumor grade and negatively with duration of clinical follow-up and survival. There was a non-significant trend for apoE epsilon2 allele carriers to have high-grade tumors and apoE epsilon4 allele carriers to have low-grade tumors. Possession of apoE epsilon4 was associated with a more advanced age of disease presentation (p < 0.01) and a longer duration of follow-up (p < 0.04). No significant correlations were found between possession of either apoE epsilon2 or apoE epsilon4 alleles and indices of cell proliferation. CONCLUSIONS: These preliminary findings suggest that possession of apoE epsilon4 allele may correspond to a more favorable clinical course in terms of more advanced age of disease presentation, and longer duration of follow-up and survival in patients with CNS neoplasms.

Communication: expression of the novel inhibitor of apoptosis survivin in normal and neoplastic skin.

Apoptosis plays a fundamental part in epidermal homeostasis, and apoptotic cells have been detected in normal and diseased skin. Little is known, however, on the inhibitory mechanisms of apoptosis at the skin level. In addition to bcl-2, a novel inhibitor of apoptosis designated survivin and structurally analogous to IAP apoptosis inhibitors has been recently identified. The expression of survivin in normal and pathologic skin was investigated. Immunohistochemical studies revealed that survivin is expressed in basal keratinocytes, but not in suprabasal epidermal layers, with a pattern similar to bcl-2. In western blots, the anti-survivin antibody recognized a single band of 16.5 kDa in protein extracts from normal human keratinocytes in culture, in agreement with the predicted size of survivin. In addition, survivin immunoreactivity was detected in benign and malignant melanocytic lesions, with strong expression in invasive lesions of melanomas. Whereas survivin staining was undetectable in benign epithelial tumors, such as seborrheic keratoses, it was observed in all epidermal layers in Bowen's disease. Interestingly, at variance with bcl-2, survivin was markedly expressed in squamous cell carcinoma, but virtually lacking in basal cell carcinoma, suggesting that these two apoptosis inhibitors may act through different anti-apoptotic pathways. Deregulation of survivin may influence both epidermal homeostasis and the development of melanoma and nonmelanoma skin cancer.

[Magnetic resonance with endorectal coil in the local staging of prostatic carcinoma. Comparison with histologic macrosections in 40 cases]. / La Risonanza Magnetica con bobina endorettale nella stadiazione locale del carcinoma prostatico. Confronto con macrosezioni istologiche in 40 casi.

PURPOSE: Endorectal coil MRI is widely used in the diagnostic workup of prostate cancer, but diagnostic accuracy rates reported in the literature are quite variable. We report our personal experience with endorectal coil MRI in the local staging of prostate carcinoma. MATERIAL AND METHODS: Forty consecutive patients with histologically proved prostate carcinoma were examined with endorectal coil MRI at high field strength (1.5 T). All patients underwent a sagittal T1-weighted SE location sequence (TR 400, TE 20), an axial T1-weighted SE (TR 400, TE 20), two axial T2-weighted FSE sequences (TR 3000, TE 102, ETL 8) with and without fat suppression, and a coronal T2-weighted FSE sequence (TR 3000, TE 102, ETL 8); an axial Fast Multiplanar Spoiled Gradient Recalled (FMSPGR) dynamic sequence after Gd-DTPA injection was also performed in 18 patients. MR staging of local tumor spread was done according to the current literature criteria. All patients were submitted to radical prostatectomy, and histologic macrosections on the same plane as MR images were obtained from surgical specimens. MR and histologic staging were compared to assess MR accuracy in detecting capsular infiltration, seminal vesicles and apex involvement. The diagnostic yield of Gd-DTPA was also investigated. RESULTS: MRI correctly staged 31 of 40 cases (77.5%). MR accuracy was 80% in detecting capsular infiltration (85.7% sensitivity and 73.6% specificity), 90% in seminal vesicle involvement (91.6% sensitivity, 89.2% specificity) and 72.5% in apex involvement (79.1% sensitivity, 62.5% specificity). Dynamic studies with Gd-DTPA did not improve staging accuracy in any case. DISCUSSION AND CONCLUSIONS: In agreement with most of the current literature, MRI showed moderate overall accuracy in the local staging of prostate carcinoma. Particularly, MRI had good accuracy in detecting seminal vesicle involvement but moderate sensitivity and specificity in demonstrating capsular infiltration and apex involvement. Due to its high cost, MRI should not be routinely used in prostate cancer staging but should be reserved to the patients whose clinical and serological data suggest extraprostatic tumor spread, whose preoperative demonstration could avoid noncurative surgery.

Loss of P27Kip1 expression correlates with tumor grade and with reduced disease-free survival in primary superficial bladder cancers.

p27Kip1 is a member of the Cip1/Kip1 family of cyclin-dependent kinase inhibitors and is a potential tumor suppressor gene. We previously reported a deregulated expression of p27Kip1 in a series of human cancer cell lines and in primary breast and colon cancers. Moreover, p27Kip1 has been reported as an important prognostic factor in primary lung, breast, colon, and prostate cancers. In this study, we evaluated the prognostic value of p27Kip1 in a series of 96 superficial (pTa-1) human bladder carcinomas. High (>50% positive cells), moderate (25-50%), and low (<25%) p27Kip1 staining was observed in 39 (41%), 19 (20%), and 38 (39%) of the 96 primary superficial bladder cancers, respectively. No significant association was found between the expression level of p27Kip1 and tumor stage. Decreased p27Kip1 staining correlated with higher tumor grade (P = 0.001). Interestingly, a significant association was observed between increased expression of p27Kip1 and positivity for p53 (>20% positive cells; P < 0.001). A significant correlation was also observed between low expression of p27Kip1 and decreased disease-free survival (P = 0.0003 by log-rank test) and overall survival (P = 0.01 by log-rank test). Furthermore, on multivariate analysis, low p27Kip1 protein expression was an independent predictor of reduced disease-free survival (P = 0.018; relative risk = 1.95) second only to tumor stage. These data indicate that p27Kip1 protein is frequently expressed at low level in poorly differentiated tumors and suggest that this protein might represent a useful prognostic marker for disease recurrence and overall survival in superficial bladder carcinomas.

Detection of human papillomavirus DNA in urinary bladder carcinoma by in situ hybridisation.

AIMS: To investigate the sensitivity of an in situ hybridisation system to detect human papillomavirus (HPV) infection in transitional cell bladder cancer and to evaluate the advantages of analysing multiple biopsies; to examine the correlation between HPV tumour infection detected by in situ hybridisation and the presence of serum anti-HPV antibodies detected by enzyme linked immunosorbent assay (ELISA); and to relate the presence of viral infection to grade, stage, and follow up in cases of bladder cancer. METHODS: The in situ hybridisation technique was used with broad spectrum and type specific (6/11, 16/18, 31/33/35) probes against HPV DNA in formalin fixed, paraffin embedded tissues from 43 cases of bladder cancer. The results were analysed for the presence and type of papillomavirus and correlated with clinicopathological variables. RESULTS: The presence of HPV DNA was identified by the in situ hybridisation technique in 17 of 43 cases of bladder cancer; 12 of these were serum antibody positive and 10 had had multiple biopsies. Fifteen of the cases that were negative for HPV DNA by in situ hybridisation had positive serum serology when tested by ELISA. In 14 cases, the HPV was either types 16/18 or types 31/33/35, both of which carry high oncogenic risk. The stage (p < 0.05) and grade (NS) of the tumour and the outcome on follow up (p < 0.05) were correlated with the presence of HPV infection. CONCLUSIONS: ELISA is not useful in identifying patients with HPV positive bladder cancer, but the use of several probes and multiple biopsies increases the detection rate of HPV in neoplastic tissues. The association between tumour virus infection and high grade/high stage tumours and worse outcome suggests that HPV infection of neoplastic tissue has a negative effect on the behaviour and evolution of transitional cell bladder carcinoma.

Analysis of 4-ABP-DNA adducts and p53 alterations in urinary bladder carcinoma.

BACKGROUND: Activated intermediates of 4-aminobiphenyl (4-ABP) are able to covalently interact with DNA to form adducts. There is a large body of evidence indicating that carcinogen-DNA adduct formation can be one of the cancer initiating mechanisms. MATERIALS AND METHODS: (4-ABP)-induced DNA damage in association with p53 overexpression and mutations were evaluated in specimens of urothelial bladder cancers from 106 patients. RESULTS: 4-ABP-DNA adduct levels resulted higher in smokers compared to non smokers, with a borderline statistical value. p53 nuclear overexpression was related to tumor grading, while no significant correlation with stage, 4-ABP-DNA adducts, smoking habit, and disease recurrence could be observed. Concerning molecular analysis, p53 point mutations were found in 17 of 106 cases (16%) and mutational pattern was significantly associated both with higher grade and stage, but no correlation was found with disease recurrence. CONCLUSIONS: These results suggest that other sources, in addition to tobacco smoke, may contribute to 4-ABP-DNA adducts formation in bladder tissue and that p53 expression/mutation cannot be considered a prognostic factor in bladder cancer.

Does p53 immunostaining improve diagnostic accuracy in urine cytology?

The frequent change of the transitional cell carcinoma of the urinary tract accounts for the fact that cytological abnormalities in urinary specimens are often not sufficient to enable a definitive diagnosis of malignancy. The purpose of this work was to evaluate the possible use of p53 protein in increasing the diagnostic accuracy of urinary cytology. The expression of p53 was investigated by immunocytochemistry in two groups of urinary specimens, one cytologically positive and the other cytologically negative for cancer. Immunostaining was carried out using a monoclonal antibody to p53. In the positive group, in which bladder cancer was confirmed by cystoscopy and biopsy (31 cases), positive reaction for p53 was found in 55% of the cases (17 cases). In the negative group (92 cases), presence of cancer was histologically ascertained in 64 cases and in this group 15 cases (23.4%) showed positive p53 staining. In the remaining 28 cases of this group, where TCC was not present, 7 cases showed p53 positivity in non-neoplastic urothelial cells. This result shows that, while immunocytochemical detection of p53 in urinary specimens may be used for prognostic evaluation of patients with bladder cancer, it does not contribute to the diagnostic accuracy in cases with morphologically inconclusive or negative cytology. The sensitivity and specificity of the method in detecting bladder carcinoma were 23.5 and 75%, respectively.

[Molecular biology in bladder carcinoma: contributions of immunohistochemistry]. / Biologia molecolare nel carcinoma vescicale: contributi di immunoistochimica.

Despite the insights genetics and molecular biology have given to a better understanding of the mechanisms which lead to the onset and development of bladder carcinoma, the factors that influence its unpredictable and, at times, particularly aggressive outcome are still largely unknown. Also in bladder carcinoma the study of cellular differentiation markers has been replaced by that of genotypic alterations, and, mainly with the help of immunohistochemistry, of the expression of genes involved in cell proliferation and death, such as MTS1, TP53, Rb, c-myc, Bcl-2, c-erb-B2. So far, anyway, no independent and reliable indicator able to predict the outcome of the single tumour has been identified, and this issue seems to be best addressed by studies of the altered expression of more than one oncoprotein simultaneously. Fairly identical is the question arised by TP53 mutations, which, while worsening the evolution of advanced muscle-infiltrating tumours, hold a still unclear and debated meaning in superficial tumours. It is anyway clear that molecular analysis only may enable to reliably detect the presence of any TP53 mutations. As a matter of fact, the multiplicity of genetic mutations, the frequent transcript variations and the intrinsic limits of immunohistochemistry may explain the discrepancy between immunohistochemical and molecular analysis results, with specificity and sensitivity levels clinically not acceptable. To date, anyway, the biological and clinical meaning of this discrepancy has still to be clarified, as well as the clinical meaning, if any, of p53 overexpression in the absence of gene mutations.

[Bulletin of breast disease: results of the 1990-1994 quinquennium]. / Bollettino della patologia mammaria: risultati del quinquennio 1990-1994.

All mammary lesions diagnosed at the Institute of Pathological Anatomy of the University of Modena have been systematically filed since 1990 and reported in a bulletin, which is issued twice a year and delivered to health operators. So far, 5.188 cases of breast lesions, comprising 1.999 non-neoplastic pathologies, 1.040 benign tumors, 1.943 primary malignant neoplasms and 206 recurrences, have been filed. During the quinquennium 1990-1994, a progressive numerical reduction in diagnoses of non-neoplastic lesions coupled to an increase of benign tumors has been observed, whereas the number of primary malignant tumors has not changed. In particular, a statistically significant increase in diagnoses of carcinoma-in-situ and of fibrocystic disease associated with moderate-risk lesions (atypical hyperplasias) has been detected, whereas the number of cases of single fibrocystic disease has decreased. This reduction, however, is not significant. A slight increase of breast carcinomas smaller than 1 cm and 2 cm, coupled to a decrease of those exhibiting dimensions between 2 and 5 cm, has been found. The collection and systematic analysis of cases of mammary lesions appears to represent a useful tool to study the incidence of different breast pathologies in the general populations. It can also be viewed as a simple way to test the reliability of diagnostic methods used for selection of surgical cases.

Adrenergic and serotoninergic receptors mediate the immunological activation of corticosterone secretion in male rats.

In order to elucidate the mechanism of action of immune agents on corticosterone secretion, the present study evaluated the possible involvement of some neuronal pathways (serotoninergic, noradrenergic/adrenergic) in the lipopolysaccharide (LPS)-induced corticosterone release in male rats. Serotoninergic antagonists, mianserin (5-HT2C receptor blocker) or pindolol (5HT1A receptor blocker) or noradrenergic/adrenergic antagonists, prazosin (alpha 1-adrenoceptor blocker) or propranolol (beta-adrenoceptor blocker), were intraperitoneally (i.p.) injected before (5 min) the administration of LPS. In each experiment a group of rats i.p. injected with vehicle served as controls. Animals were sacrificed by decapitation 90 min after administration of LPS and trunk blood was collected for corticosterone radioimmunoassay. Results showed that pretreatment with mianserin, but not with pindolol, significantly reduced plasma corticosterone levels following administration of LPS (p < 0.05); prazosin attenuated the plasma corticosterone response to LPS (p < 0.05), while propranolol did not induce significant change. The present study indicated that serotoninergic and noradrenergic/adrenergic pathways are involved in the immunoneuroendocrine modulation of hypothalamus-pituitary-adrenal function in rats. In particular, it is probably mediated by the activation of 5-HT2C receptors and of alpha 1-adrenoceptors, while type 1A serotonin receptors or beta-adrenoceptors do not seem to be involved in such a phenomenon.

Estrous cycle- and acute stress-related changes of rat ovarian immunoreactive corticotropin-releasing factor.

Corticotropin-releasing factor (CRF), the major regulator of the stress response within the central nervous system, is also present at peripheral sites, including the gonads, and the gene encoding its own receptor can be finely induced in selective ovarian compartments in both control and stressful conditions during the gonadal life cycle. The present study, therefore, investigated the influence of both gonadal function and estrous cycle on the immunoreactive CRF (irCRF) contents in the immature and adult rat ovary. In addition, the effect of an acute (5 min) or chronic intermittent (twice a day for 4 days) cold swimming stress on ovarian irCRF contents was evaluated. High-performance liquid chromatography (HPLC), gel-chromatography (Sephadex G-75, 45 x 1 cm) and a direct radioimmunoassay were performed to measure irCRF ovarian contents. The HPLC elution profile of irCRF in ovarian tissues of adult rats was superimposable on that of synthetic rat/human CRF and gel-chromatograms performed according to the phase of the estrous cycle revealed higher irCRF contents at proestrus. Total irCRF ovarian content was undetectable both in control and acute stressed immature rats, while adult rats showed the highest values at proestrus (p < 0.0001). The acute stress exposure induced a significant increase (p < 0.0001) of irCRF ovarian contents only at proestrus, without affecting irCRF at the other phases of the estrous cycle. Finally, no significant changes were found in ovarian irCRF after chronic intermittent stress. The proestrus-related changes of ovarian irCRF, confirming the adult ovary as an extrahypothalamic source of CRF, may constitute a neuropeptidergic signal involved in the gonadal reproductive cycle. Furthermore, the stress-related changes of ovarian irCRF indicated that the gonad may be locally sensitive to acute stressful stimuli.

Bicuculline enhances the corticosterone secretion induced by lipopolysaccharide and interleukin-1 alpha in male rats.

Lipopolysaccharide (LPS)-induced inflammatory stress activates the hypothalamus-pituitary-adrenal (HPA) function. Interleukin-I (IL-1) is one of the key factors during this event; however, the mechanisms mediating IL-1 stimulation of HPA axis are still unclear. The present study evaluated the possible involvement of gamma-aminobutyric acid (GABA) in LPS-induced activation of HPA axis in adult male rats. In addition, the possible existence of diurnal changes of LPS-induced HPA axis activity was also investigated. Bicuculline (0.8 mg/kg BW), a GABA-A receptor antagonist and GABA (1 g/kg BW) were intraperitoneally (ip) injected 15 min before LPS (2 mg/kg BW, ip) or recombinant human IL-1 alpha (microgram/rat) administration in intact rats. Control animals received an equivalent volume of 0.9% saline. Rats were sacrificed at 60 min or 90 min after LPS, or IL-1 alpha or saline injection. Plasma corticosterone levels were measured by radioimmunoassay. Results showed that pretreatment with bicuculline enhanced both LPS- and IL-1-induced corticosterone secretion; while pretreatment with GABA significantly reduced the LPS-stimulated corticosterone release (p < 0.05, vs LPS alone). The effect is dependent on the time of sampling and such effect of bicuculline or GABA was not observed when rats were stimulated in the evening. In addition, the maximal changes of plasma corticosterone following LPS administration in the evening were significantly lower than in the morning (p < 0.01). The present study provides evidence that GABA is involved, at least in part, in the neuroendocrine regulation of LPS/interleukin-1a-induced corticosterone secretion via GABA-A receptor in rats. In addition, the response of plasma corticosterone to LPS has a diurnal variation, which corresponds to a diurnal change of GABAergic modulation of the immunoneuroendocrine response.

Argyrophilic nucleolar organizer regions and bromodeoxyuridine and 3[H]-thymidine labelling indices in colorectal cancer.

The count of argyrophilic nucleolar organizer regions (AgNORs) has been proposed as a useful method for evaluating cell replication in human tumours. The current study was undertaken to compare AgNOR values in colorectal cancers with two better established methods for investigating cell proliferation such as bromodeoxyuridine (BrdUrd) and 3[H]-thymidine (3[H]dT) labelling indices (LIs). Because some concern still exists regarding accuracy and reproducibility of AgNOR quantifying methods, we carried out a control study by independently repeating the same measurements (number, area and area per silver-stained NOR particle) in two centres with different operators and computer-assisted image analysers on 40 colorectal carcinomas. AgNOR values recorded in the two centres were strictly correlated (r = 0.75; P < 0.001 for number; r = 0.62, P < 0.01 for area; r = 0.63, P < 0.001 for area per silver-stained NOR particle) and the range of values were almost identical. Then, AgNOR values were compared with BrdUrd and 3[H]dT LIs, respectively obtained by in vivo incorporation and in vitro incubation in the same series of colorectal carcinomas. No correlation was found between AgNOR values and BrdUrd or 3[H]dT LIs. BrdUrd and 3[H]dT LIs were instead reciprocally significantly correlated. No evident correlation was seen between LIs or AgNOR values and clinico-pathological parameters of the tumour. In conclusion, in colorectal neoplasms, AgNOR values did not appear to relate with more direct parameters of cell proliferation. It follows that AgNOR reliability as a biomarker of cell proliferation remains questionable.

Detection of human papillomavirus in extragenital Bowen's disease using in situ hybridization and polymerase chain reaction.

Extragenital Bowen's disease (EBD) has rarely been studied for the presence of human papillomaviruses (HPVs). Twenty consecutive patients with EBD were investigated for the presence of HPVs using in situ hybridization with a generic probe that can detect HPV DNA types 6, 11, 16, 18, 30, 31, 33, 35, 45, 51, and 52 and specific probes for HPV DNA types 6/11, 16/18, and 31/33/35. All cases were tested with the polymerase chain reaction (PCR) technique employing the L1 consensus primer pair, MY11 (primer for the positive strand) and MY9 (primer for the negative strand) complementary to genital and dermal HPV types. Seven caucasian patients, five males and two females, with an average age of 70.4 years, showed positive in situ hybridization (ISH) for HPV DNA. The positivity varied from 5 to 40% of neoplastic cells. Three of seven of the ISH DNA-positive cases showed a positive PCR for DNA HPVs. The role of HPVs in human tumors is not fully understood since oncogenic types of HPVs have been found in normal tissue and the actions of cofactors have been postulated. Bowen's disease usually occurs in elderly people in whom the efficiency of the immune systems may be compromised. The association between HPV infection and low efficiency of the immune response may be responsible for HPV-related Bowen's disease in elderly people.