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Background: An involvement of the appendiceal orifice as a distintive skip lesion in adults with left side ulcerative colitis (UC) has been reported. The aim of our prospective study was to evaluate, by endoscopy and histology, the prevalence of periappendiceal inflammation (PAI) in children affected by UC., Methods: Fifty of 77 consecutive children undergoing total colonoscopy, who had a diagnosis of UC not extended beyond the hepatic flexure were enrolled., Results: PAI was endoscopically present in 16 of 50 patients (32%) with UC. Patients were divided in 2 groups: group A included the 16 patients with PAI, whereas group B included 34 patients without PAI. We found that among the 2 groups, PAI was more frequent in patients with new diagnosis than in those with pre-existing UC (P = 0.016). At index colonoscopy, the patients of group A had a significant major extent of disease (P = 0.013). Moreover, the histologic grade of inflammation at the ascending colon was significantly higher in group A than in group B (P = 0.014). Clinical activity, measured by pediatric ulcerative colitis activity index, and use of medication did not show significant differences among groups (P = 0.464 and P = 0.723, respectively). The use of immunosuppressant was significantly higher in group A than in group B., Conclusions: PAI is a frequent skip lesion in children with UC. It seems more frequent in patients with new diagnosis, and it is associated with a major extent of the disease and with a higher grade of histologic inflammation at the ascending colon.

Context: The urokinase plasminogen activating system is implicated in neoplastic progression, and high tissue levels of urokinase plasminogen activating system components correlate with poor prognosis in various human cancers., Objective: The objective of the study was to investigate the prognostic relevance of the urokinase plasminogen activator (uPA), its cognate receptor (uPAR), and the plasminogen activator inhibitor 1 (PAI-1) in human papillary thyroid cancer (PTC)., Design: The expression of uPA, uPAR, and PAI-1 genes was analyzed in PTC and normal matched tissues by quantitative RT-PCR. The case study consisted of 99 patients (21 males and 78 females) affected by PTC including 77 classical, 15 follicular, four tall cell, and three oncocytic variants. Forty-one patients had lymph node metastases at the time of diagnosis. All the patients underwent thyroidectomy and radioiodine therapy followed by thyroid hormone replacement therapy. Follow-up data were available for 76 patients up to 64 months., Results: The uPA, uPAR, and PAI-1 mRNA levels were significantly higher in PTC compared with normal matched tissues by 9.63 ± 1,29-, 4.82 ± 0.45-, and 5.64 ± 0.71-fold, respectively. The increased expression of uPA and uPAR correlated statistically with advanced pT and N status. The uPA was also significantly associated with advanced tumor node metastasis stages. The Kaplan-Meier analysis showed a significant association of uPA and uPAR levels with reduced patient disease-free interval (DFI), and this association was stronger in stage I patients., Conclusion: The study demonstrated that in PTC the increased gene expression of uPA and uPAR is associated with tumor invasiveness, advanced stages, and shorter DFI, suggesting their prognostic relevance. These observations warrant further investigation in larger patient populations with longer follow-up.

Background: Despite recent progress in the treatment of advanced urothelial cancer, there continues to be a need to identify new active agents and their toxicity spectra. We conducted a study using FOLFOX-4 (oxaliplatin, fluorouracil, folinic acid) in pre-treated advanced bladder cancer patients., Methods: Sixteen eligible patients with advanced disease were treated with oxaliplatin (85 mg/m(3)) on day 1 followed by fluorouracil and folinic acid (De Gramont schedule) on days 1 and 2 every 14 days until disease progression. All patients received nutritional support to increase their caloric intake. Objective responses and toxicity were evaluated. Biochemical responses (reduction of markers) and nutritional parameters (increase in body weight and albumin, and reduction in ferritin and C-reactive protein) were also considered., Results: Three patients obtained an objective response (overall response rate 19%). Hematological toxicity and stomatitis were the most commonly noted side effects, but we observed only low (3-4) grade toxicity. In four patients (25%), we observed a reduction in tumoral markers (carcinoembryonic antigen and tissutal polypeptide antigen) and modified nutritional parameters., Conclusions: Using these doses and schedules of FOLFOX-4 appears to be a promising therapy in patients pre-treated with platinum compounds. More studies are required to assess the possible role of this regimen in the treatment of advanced bladder cancer.

To study the usefulness of calretinin as an immunohistochemistry marker in the diagnosis of cardiac myxoma (CM) and the origin of myxoma cells, we examined 24 CMs and 9 fetal hearts with immunohistochemical methods on formalin-fixed paraffin-embedded tissues. We compared 24 CMs with 10 mural thrombi, 6 jaw myxomas, and 2 papillary fibroelastomas. Calretinin expression was identified in 100% of CMs and was negative in all cases of mural thrombi, jaw myxoma, and papillary fibroelastoma. Calretinin expression by the neoplastic cells in CM was strong and diffuse and had a cytoplasmic and a nuclear pattern. Calretinin expression in fetal hearts was found in autonomic ganglia cells in the subepicardial tissue of the atria and atrial appendages, along the interatrial and atrioventricular sulci, and in the atrial septum. Results clearly indicate that calretinin can be used as a marker for the diagnosis of CM and that it is a powerful tool for the differential diagnosis, most importantly with mural myxoid thrombi. Furthermore, the positive expression of calretinin by the autonomic neurons in the fetal heart and CM supports the concept that myxoma cells may originate from endocardial sensory nerve tissue.

We studied the spatiotemporal distribution of thyroid hormone nuclear receptors (TRs) alpha1 and alpha2 and beta messenger RNA (mRNA) levels in normal human testicular tissue during development and in adulthood. Nonpathological specimens from five aborted fetuses (17 and 23 weeks of gestation, three and two cases, respectively) and from four patients undergoing orchiectomy (18 months old and 38-, 42-, and 52-yr-old, respectively) were analyzed by Northern blot, semiquantitative RT-PCR amplification using DNA sequences or specifically designed primers for the TR isoforms, and in situ hybridization. By using PCR amplification, we found that TRalpha1 and TRalpha2 are both expressed at different levels in fetal and adult testis. At all ages TRalpha2 is found at higher levels. Northern analysis showed hybridization signals corresponding to the expression of TRalpha2 and TRalpha in a ratio that increased from 2.6 at 17 weeks of gestation to 12.0 in adulthood. In fact, the expression of TRalpha1 dramatically decreased throughout development, being faintly detectable in the adult testis. Expression of TRbeta was not detected at any age studied. This finding was further confirmed by PCR, which did not amplify TRbeta either in fetal or in adult testis mRNAs. In situ hybridization studies showed the absence of TRbeta and that TRalpha1 and TRalpha2 colocalized in Sertoli cells of prepubertal testis, whereas germ and interstitial cells appeared devoid of TR mRNA signals. From these results it can be concluded that the human testis exclusively expresses TRalpha, which is localized in Sertoli cells, TRbeta being always undetectable. Fetal and prepubertal ages represent the period of maximal expression of TRalpha1 and TRalpha2. The alpha2/alpha1 ratio rises dramatically after development. These results confirm a critical window for the action of thyroid hormone in human testis, in the period of maximal expression of T3 binding isoform TRalpha1, and may account for the macroorchidism without virilization occurring when hyposecretion of thyroid hormones occurs before puberty.

In the present study we evaluated the role of T3 on the in vitro processes of mouse cumulus cell-oocyte complex expansion, oocyte meiotic maturation, and granulosa cell aromatase activity. Results obtained from cumuli oophori isolated from immature and adult mice ovaries demonstrated that T3 at all concentrations tested (0.1-100 nM) did not affect basal or FSH-induced cumulus expansion or interfere with oocyte meiotic maturation up to metaphase II stage. On the contrary, T3 inhibited in a time- and dose-dependent manner FSH-induced aromatase activity in cultured granulosa cells obtained from either adult or immature female mice. The half-maximal dose (ED50) of T3 inhibition was 0.87 +/- 0.21 nM, which is in agreement with the reported dissociation constant of T3 nuclear receptor (Kd = 0.4-5 nM) in mammalian granulosa cells. Time-course experiments demonstrated higher sensitivity to T3 of adult granulosa cells with respect to immature granulosa cells in culture. Indeed, in immature granulosa cells T3 inhibition became significantly evident only after 6 days of hormonal treatment, whereas in adult granulosa cells the inhibitory effect was present after only 2 days of treatment. (Bu)2cAMP- or 3-isobutyl-1-methyl-xanthine-stimulated aromatase activity was also significantly decreased by T3, thus suggesting that the inhibition was downstream from cAMP formation. Lastly, analysis of aromatase messenger RNA (mRNA) levels by semiquantitative RT-PCR demonstrated the ability of FSH to increase aromatase mRNA level in cultured granulosa cells by 2.4 +/- 0.5-fold. In agreement with the effect on enzyme activity, the stimulatory effect of FSH on aromatase mRNA level was greatly reduced after T3 cotreatment. In conclusion, T3 inhibition of aromatase activity may be of physiological relevance in the complex multihormonal regulation of mammalian follicle development and may contribute to explaining the alteration in female reproductive functions after thyroid hormone hypo- or hypersecretion.

The present study reports the modulation of basement membrane (BM) components, laminin, entactin, and type IV collagen, expression in prepubertal rat Sertoli cell by the thyroid hormone T3. Immunocytochemical studies of permeabilized Sertoli cells in culture showed that T3 treatment (10[-7] M for 24 h) increased the number of cells staining positive for laminin and/or entactin (from 58 +/- 5.3% to 86.4 +/- 6.5%, P < 0.01). In contrast, a strong inhibition of type IV collagen immunopositivity was observed. Western blot analysis of Sertoli cell-conditioned media indicated that T3 treatment significantly (P < 0.01) increased the level of secreted entactin by 60-65% without affecting the levels of laminin A and B1/B2 chains. Moreover, thyroid hormone treatment of Sertoli cells significantly reduced type IV collagen secretion by 62% (P < 0.05). Slot blot analysis of poly-A RNA demonstrated a significant (P < 0.01) increase in the level of entactin messenger RNA (mRNA) by 140% (P < 0.01) and a 50% reduction of type IV collagen alpha1 chain mRNA after thyroid hormone treatment. No effect of the hormone was observed on the accumulation of the laminin B1 and B2 chain mRNAs in Sertoli cell cultures. These effects cannot be ascribed to changes in the degradation of BM components, because no effect of thyroid hormone was observed on plasminogen activators or metalloproteinase secretion by Sertoli cells. These observations indicate the Sertoli cell as a source of entactin within the testis, demonstrate the ability of T3 to differentially regulate the expression of BM components, and can be regarded as a part of the integrated mechanism by which thyroid hormone affects testicular development and differentiation.

Primary cultures of prepubertal rat Sertoli cells secrete two major tissue inhibitors of metalloproteinases: TIMP-1 (M(r) 28K) and TIMP-2 (M(r) 21 K). FSH stimulated Sertoli cell TIMP-1 and TIMP-2 activity in a time- and dose-dependent manner and also stimulated TIMP-1 and TIMP-2 protein and messenger RNA levels. These effects were mimicked by the cAMP analog, 8-bromo-cAMP, and the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine. The protein kinase C activating phorbol ester phorbol myristate acetate (TPA) stimulated TIMP-1 but not TIMP-2 activity and messenger RNA levels. Cycloheximide and actinomycin-D inhibited basal TIMP-1 and TIMP-2 activity and inhibited the ability of FSH, 8-bromo-cAMP, and TPA to stimulate TIMP activity. The protein kinase A (PKA) inhibitor AMP Rp isomer did not affect basal TIMP-1 and TIMP-2 activity or TPA-stimulated TIMP-1 activity. However, the PKA inhibitor markedly reduced FSH and 3-isobutyl-1-methylxanthine stimulation of TIMP-1 and TIMP-2 activity. FSH, 8-bromo-cAMP, and TPA stimuli induced DNA binding complexes capable of binding to a TIMP-1 AP-1 site consensus sequence oligonucleotide. The AP-1 site binding complex(es) induced by all three treatments reacted with antibodies directed broadly against fos and jun protooncogene families and against the specific family members c-fos, junB, and junD but not c-jun proteins. Constitutive cAMP response element binding activity capable of binding an artificial cAMP response element binding site oligonucleotide was demonstrated in Sertoli cell nuclear extracts. This activity was minimally modulated by FSH, 8-bromo-cAMP, or TPA treatment. In summary, Sertoli cells secrete TIMP-1 and TIMP-2 that can be coordinately up-regulated by FSH through a cAMP, PKA-dependent pathway. a convergence of TPA, FSH, and cAMP mediated signals in prepubertal Sertoli cells may occur with the induction of specific AP-1 site binding complex(es) containing jun and fos proteins. Our data suggest that FSH stimulation of TIMP-2 expression may be regulated independently to that of TIMP-1. We propose that the ability of FSH to stimulate Sertoli cell TIMP activity suggests a central role for this hormone in the control of extracellular matrix turnover during testicular development at the level of metalloproteinase inhibition.

The possibility that FSH stimulates the phospholipase A2 (PLA2) pathway was studied in cultured immature Sertoli cells. FSH induced [3H]-arachidonic acid (AA) release from prelabeled cells in a time- and concentration-dependent fashion (ED50 = 21.8 +/- 1.9 ng/ml). This response could be fully prevented by pretreatment of cells with the PLA2 inhibitor, mepacrine. That PLA2 was the main enzyme responsible for cleavage of AA from membrane phospholipids was directly shown by PLA2 activity assay using vesicles of radiolabeled phosphatidylcholine (PC) as substrate. Furthermore, FSH stimulated eicosanoid generation in a time-dependent manner through the cyclooxygenase but not the lipoxygenase pathway. In fact, higher levels of prostaglandin (PG) E2, F2 alpha, and the stable products of PGI2 and thromboxane A2 (6-keto PGF1 alpha and thromboxane B2, respectively) were generated by the gonadotropin-treated cells as compared to control cells. The effect was inhibited by mepacrine, further supporting the pivotal role of PLA2 in the release of the eicosanoid precursor, AA. Finally, the effect of the main product of FSH-induced AA metabolism, i.e., PGE2, was studied. Intracellular cAMP accumulation in Sertoli cells was stimulated by the prostanoid in a dose-dependent manner (ED50 = 2.3 +/- 0.37 nM). PGE2 also significantly stimulated aromatase activity, a specific marker of Sertoli cell functions, measured as 17 beta-estradiol production (ED50 = 4.7 +/- 0.8 nM). Similar results were obtained with PGF2 alpha. Our findings show that FSH, through the activation of PLA2, leads to AA release with consequent metabolism by the cyclooxygenase pathway.(ABSTRACT TRUNCATED AT 250 WORDS)

By means of in vivo and in vitro approaches, we studied the effect of thyroid hormone on postnatal development of rat testis. T3 treatment in neonatal rats is associated with an increase of testis size of about 60%, compared to coeval controls. Increased number of both Sertoli and germ cells and enlarged diameter of seminiferous cords were found in hyperplastic testes. In the T3-treated group, TSH serum levels were low and a slight increase of FSH was found. In vitro treatment of neonatal testis fragments by 10(-7) M T3 for 3 days increased the number of gonocytes (P < 0.001 vs control) and decreased the percentage of degenerating germ cells (P < 0.001 vs control). In the adult testis, both in vivo and in vitro treatments with thyroid hormone did not induce morphological modifications, thus demonstrating that the critical window of thyroid hormone effectiveness coincides with the prepuberal period. Since thyroid hormone stimulates Sertoli cells to secrete growth factors and nutrients for germ cell development, we suggest that the increased testicular size and germ cell number following the T3 treatment is mediated by a direct hormonal effect on the somatic cell of the seminiferous epithelium.

The aim of this study was to investigate the ontogenesis of the nuclear T3 receptor among the different cell types in the rat testis from fetuses at the 19th day of gestation and animals 1, 5, 15, 20, and 60 days after birth. Whole testis, tubular fraction, nontubular fractions, and Sertoli cells cultured in vitro or enriched in vivo by irradiation were used. The results demonstrate that high affinity, low capacity T3-binding sites are localized only in Sertoli cells; the binding specificity and affinity (Kd ranges from 0.8 +/- 0.2 to 2.6 +/- 0.4 nM) do not change significantly with the age of the animals and are comparable to those observed for T3 receptors in other mammalian tissues. In the whole testis, the concentration of receptors changes during gonadal development, being maximally expressed in the fetus (154.3 +/- 8.1 fg T3 bound/10(6) nuclei) and from 1 (203.4 +/- 10.9 fg T3 bound/10(6) nuclei) to 5 (185.3 +/- 15.1 fg T3 bound/10(6) nuclei) days of postnatal life, decreasing significantly at 15 and 20 days (65.4 +/- 2.0 and 57.9 +/- 1.9 fg T3 bound/10(6) nuclei, respectively) and being virtually absent in the adult. The same change in receptor concentration was found in Sertoli cells obtained by different techniques. This ontogenetic profile coincides with the pattern of Sertoli cell proliferation and differentiation, thus suggesting a role of thyroid hormones in the regulation of growth and maturation of the somatic cells of the seminiferous epithelium.

The plasma concentrations of mineralocorticoid hormones, basal and after stimulation and suppression with ACTH, can identify the heterozygotes in a family with two siblings with 17 alpha-hydroxylase deficiency. Both parents and one sibling had elevated levels of plasma deoxycorticosterone, corticosterone, 18-hydroxydeoxycorticosterone, and 18-hydroxycorticosterone, but normal cortisol and aldosterone concentrations. Stimulation with ACTH effected additional increases in the elevated steroid and cortisol levels, but not in aldosterone, further increasing the discrepancy and the ratio between 18-hydroxycorticosterone and aldosterone. One sibling had normal steroid patterns and an 18-hydroxycorticosterone to aldosterone ratio. Suppression of ACTH restored the steroids to low normal levels. In addition, the ratio of the gas chromatographic analysis of the total major urinary metabolites of corticosterone to total metabolites of cortisol was greater, and the sum of urinary androsterone and etiocholanolone to total corticosterone and cortisol metabolites was less in the heterozygotes than in normal subjects. This identifies deficient 17-hydroxylation, which is required for the production of cortisol and C-19 steroids. These criteria appear unique for the 17 alpha-hydroxylase defect in the heterozygote.