Effects of the administration of epidermal growth factor receptor specific inhibitor cetuximab, alone and in combination with cisplatin, on proliferation and apoptosis of Hep-2 laryngeal cancer cells.

BACKGROUND: Epidermal growth factor receptor (EGFR) overexpression and prognostic value in head and neck squamous cell cancer is the basis for targeting by anti-EGFR antibodies, which increase the efficacy of radiotherapy. In order to evaluate the best therapeutic schedule, the effects of cetuximab (C225) on Hep-2 cell proliferation, alone and in combination with cisplatin, were studied. METHODS: Hep-2 cells were treated with cetuximab alone or in combination with cisplatin. After determining cell viability with trypan blue, morphological features of apoptotic degeneration were analysed by fluorescence microscopy with Hoechst 33258 stain. RESULTS: Cetuximab alone mildly inhibited Hep-2 proliferation and showed no pro-apoptotic effects. When administered concomitantly with cisplatin, cetuximab synergistically increased inhibition of proliferation and apoptosis. CONCLUSION: The antiproliferative activity of cetuximab is consistent with its hypothesised role in inhibiting repopulation. However, the increase in the effects of pro-apoptotic agents induced by cetuximab may be even more relevant to its clinical effectiveness than the inhibition of repopulation.

[Calciphylaxis and penile necrosis: a case report and review of the literature ]. / Calcifilassi e necrosi peniena: case report e revisione della letteratura.

INTRODUCTION: Calciphylaxis is a rare clinic condition characterised by skin necrosis due to medial and intimal calcification of small and medium arteries. It's observed in patients affected by end stage renal disease associated to secondary hyperparathyroidism. Penile involvement has been documented in very few cases. We present both a case of penile calciphylaxis and a review of literature, in order to increase comprehension of pathophysiology, diagnosis and therapy of this rare disease. MATERIALS AND METHODS: A retrospective review of literature was performed after treating a case of penile calciphylaxis. We describe patient characteristics, clinical presentation, laboratory and histo-pathologic findings, therapeutic strategy and outcomes of the case. RESULTS: A 65 year-old man, affected by diabetes, chronic ischemic cardiopathy and chronic renal failure in hemodialytic treatment, was referred to our unit for the presence of increased consistency and significative pain of the distal portion of penis evolving in a complete glans necrosis. Blood levels of parathormone (PTH), calcium (Ca) and phosphorous (P) resulted pathologically elevated, promoting tissutal calcium deposition. The patient was treated with partial penectomy and the histologic findings confirm diagnosis of calciphylaxis, showing an ulcerative necrosis of glans with extensive calcium deposition and luminal narrowing of penile small arteries. CONCLUSIONS: The increase of number of patients with chronic renal failure in hemodialytic treatment could make penile calciphylaxis more prevalent in the future. Early diagnosis, lowering of pathologic blood levels of Ca and P associated to surgical treatment of necrotic lesions of the patient could be fundamental for a better prognosis of this aggressive disease.

[Carcinoma of the collecting ducts of Bellini: a case report with cutaneous metastasis as the initial clinical manifestation]. / Carcinoma dei dotti collettori del Bellini: case report con metastasi cutanea come iniziale manifestazione clinica.

INTRODUCTION: Bellini's collecting ducts carcinoma represents a rare tumor with an aggressive behaviour with a poor prognosis and often metastatic at diagnosis. We report the first case documented of Bellini tumor with an initial clinic presentation represented by a cutaneous metastasis of scalp. MATERIALS AND METHODS: All pertinent clinical information were compiled, including patient age, sex, mode of presentation, preoperative laboratory data, radiologic findings, surgery type, macro and microscopic findings, survival data. RESULTS: After reporting an histopathologic finding of cutaneous metastasis of unknown origin adenocarcinoma with poorly differentiation, a voluminous 6 cm left mesorenal mass is diagnosed through uro-CT. Consequently, it is performed a left radical transperitoneal nephrectomy with consensual exeresis of scalp cutaneous lozenge at the level of previous excision. The histopathologic diagnosis reported was Bellini tumor at stage pT3a-N2-M1. It has not reported significative responsiveness to adjuvant chemotherapy and the patient was died seven months after diagnosis of cutaneous metastasis. CONCLUSIONS: Most of Bellini's carcinoma are already metastatic at presentation. Analyzing literature, it is never documented a cutaneous metastasis as first sign at clinical presentation. In this context, radical nephrectomy, differently from others subtypes of advanced renal cell carcinoma, does not seem to improve survival of the patient but rather, it can keep a role in palliation or in the context of new chemotherapeutic protocols.

[Endometriosis: aetiopathogenetic basis]. / Endometriosi: le basi eziopatogenetiche.

Endometriosis, defined by the presence of endometrial tissue outside the uterine cavity, is a common condition affecting 10% of women in the reproductive age. Menstrual factors reported to increase risk include dysmenorrhea, early menarche, and shorter cycle lengths. The theory of retrograde menstruation with implantation of endometrial fragments, in conjunction with peritoneal factors to stimulate cell growth is the most widely accepted. There is a growing body of evidence that immunological factors and angiogenesis play a key role in the pathogenesis of endometriosis. In women with endometriosis, there appears to be an alteration in the function of peritoneal macrophages, natural killer cells and lymphocytes, with production of growth factors and inflammatory mediators in the peritoneal fluid. Survival, adhesion, proliferation, invasion and vascularization of endometrial tissue in abdominal cavity may be the consequence of retrograde menstruation and referred to as implantation theory.

Persistent platelet activation in patients with type 2 diabetes treated with low doses of aspirin.

BACKGROUND: The percentage of diabetic patients who do not benefit from the protective effect of aspirin is larger than in other populations at cardiovascular risk. OBJECTIVE: We compared the ability of aspirin to suppress TxA2 and platelet activation in vivo, in type-2 diabetics vs. high-risk non-diabetic patients. METHODS: Urinary 11-dehydro-TXB2, plasma sCD40 L, and sP-selectin were measured, together with indices of low-grade inflammation, glycemic control, and lipid profile, in 82 patients with type-2 diabetes and 39 without diabetes, treated with low doses of aspirin. RESULTS: Urinary 11-dehydro-TxB2, plasma sCD40L and sP-selectin were significantly higher in diabetics than in controls: [38.9 (27.8-63.3) vs. 28.5 (22.5-43.9) ng mmol(-1) of creatinine, P = 0.02], [1.06 (0.42-3.06) vs. 0.35 (0.22-0.95) ng mL(-1); P = 0.0001], [37.0 (16.8-85.6) vs. 20.0 (11.2-35.6) ng mL(-1), P = 0.0001], respectively. The proportion of individuals with diabetes increased across quartiles of 11-dehydro-TxB2, sCD40L, and sP-selectin, with the highest quartiles of 11-dehydro-TxB2, sCD40L and sP-selectin, including 66%, 93.3%, and 93.3% of individuals with diabetes. Markers of platelet activation positively correlated with indices of glycemic control but not with markers of low-grade inflammation. CONCLUSIONS: Platelet dysfunction associated with insufficient glycemic control, may mediate persistent platelet activation under aspirin treatment.

Nociceptin (1-13)NH2 inhibits stimulated calcitonin-gene-related-peptide release from primary cultures of rat trigeminal ganglia neurones.

In this work we have developed and characterized primary cultures of neonatal rat trigeminal ganglia neurones; calcitonin-gene-related-peptide (CGRP) released from cells was taken as a marker of neuronal function. A significant and consistent increase in CGRP secretion was elicited by non-specific (56 mm KCl or veratridine) or specific (capsaicin) depolarizing stimuli. This paradigm was subsequently used to investigate the effects of nociceptin, an opioid-like peptide involved in central and peripheral control of nociception. We found that the nociceptin analogue nociceptin (1-13)NH2 (NOC) did not affect baseline CGRP release, but it reduced in a concentration-dependent manner CGRP release induced by all tested stimuli. NOC-induced reduction was statistically significant from 0.01 nm onward and achieved maximal effects at 10 nm. Such effects of NOC were seemingly mediated by the activation of specific ORL1 receptors, as a well-known nociceptin antagonist, N(Phe1)nociceptin (1-13)NH2, was able to completely revert NOC inhibition of capsaicin-stimulated CGRP release.

HIV-1 Gp120 protein modulates corticotropin releasing factor synthesis and release via the stimulation of its mRNA from the rat hypothalamus in vitro: involvement of inducible nitric oxide synthase.

In the present study, we examined whether the human immunodeficiency virus type I (HIV-I) gp120 coat protein can modulate corticotropin releasing factor (CRF) secretion by using the incubation of rat hypothalamic explants as an in vitro model. Treatment of the hypothalamic fragments with recombinant gp120 resulted in a time- and concentration-dependent increase in CRF release. The maximal dose of 10 nM gp120 increased CRF release by 56.4% after 1 h, and 78.4% after 3 h, as compared with their respective controls. The intra-hypothalamic amount of CRF was also increased by 54.7% and 77.3% vs. controls after 1 and 3 h, respectively. Moreover, the action of gp120 was blocked by pretreatment with cycloheximide, suggesting that the viral protein modulates CRF secretion via an increase in its synthesis. We also investigated the effects of gp120 on CRF gene expression. RNase protection analyses of total RNA isolated from the explants indicated that 10 nM gp120 significantly increases CRF mRNA in a time-dependent manner. Furthermore, gp120 did not modify CRF mRNA stability, suggesting that the viral protein modulates CRF gene expression at the transcriptional level. Analysis of the mechanisms that mediate gp120-induced CRF synthesis was conducted. The incubation of the explants with recombinant interleukin-1 (IL-1) type I receptor antagonist (hrIL-1 ra) did not antagonize the actions of gp120 at 1 and 3 h, indicating that the effect of the latter is independent of IL-1 mediated mechanisms. The involvement of some second messenger pathways was also investigated. Specific inhibitors of cAMP-PKA, cyclo-oxygenase or heme oxygenase pathways failed to antagonize the gp120-induced increase in CRF production. By contrast, incubation with nonselective inhibitors of nitric oxide synthase (NOS), L-NAME and L-NNA, or aminoguanidine (AG), a selective inhibitor of inducible NOS (iNOS), blocked CRF release and, AG, its mRNA accumulation, stimulated by gp120, whereas selective inhibitors of endothelial and neuronal NOS had no effect. In addition, only L-NAME, L-NNA and AG were able to inhibit the gp120-stimulated production of nitrites. These results indicate that gp120 directly stimulates CRF gene expression and peptide synthesis from the rat hypothalamus in vitro via the activation of iNOS. Therefore, the actions of this viral protein on the HPA axis may, in part, reflect its ability to modulate CRF synthesis.

A functional link between heme oxygenase and cyclo-oxygenase activities in cortical rat astrocytes.

Recent evidence shows that the activation of heme oxygenase (HO) within the CNS is associated with increased prostanoid production. In this study, we investigated whether changes in HO activity induced by pharmacological manipulation are associated with parallel variations in cyclo-oxygenase (COX) activity and prostaglandin production in an in vitro paradigm of CNS cells, i.e. primary cultures of rat cortical astrocytes. Pharmacological tools commonly used to induce changes in HO activity, namely the HO enhancers hemin and CoCl(2) as well as the HO inhibitor Sn-mesoporphyrin-9 (SnMP9), were tested in our model, and the variations in COX activity associated with the above treatments were monitored by measuring a COX end product, prostaglandin E2 (PGE2), released into the incubation medium. We found that the increase in HO activity induced by hemin and/or CoCl(2) was not consistently associated with increases in prostaglandin production, whereas HO inhibition by SnMP9 was normally followed by a decrease in PGE2 release. The above effect was observed after both acute (30 min) and prolonged (24 hr) incubations, suggesting that baseline HO activity contributes to the maintenance of normal PG production in this model. Experiments with the stable HO end products biliverdin and bilirubin suggest that these products may play a role in mediating HO-induced COX activation.

Bacterial lipopolysaccharide increases prostaglandin production by rat astrocytes via inducible cyclo-oxygenase: evidence for the involvement of nuclear factor kappaB.

This study was set to investigate the mechanisms through which bacterial lipopolysaccharide (LPS) stimulates prostaglandin (PG) production in rat astrocytes. Primary cultures of rat hypothalamic astrocytes were established. Cells were treated with LPS alone or LPS plus antagonists of various pathways, and the subsequent changes in cyclo-oxygenase (COX) activity were monitored by measuring a COX end product, PGE2, released into the incubation medium. It was found that (i) LPS produced a concentration-dependent increase in PGE2 release from astrocytes. The potency of LPS was significantly increased by the addition of serum into the incubation medium; (ii) after 24 h of incubation, inducible COX (COX-2) accounts for most of the LPS-stimulated PG production, as the latter was markedly reduced by dexamethasone and the specific COX-2 inhibitor NS 398; and (iii) nuclear factor kappaB appears to play a role in the activation of COX-2 induced by LPS, since certain inhibitors of this transcription factor were able to antagonize, at least in part, the effects of LPS on PGE2 release.

Chronic treatment with the antidepressant amitriptyline decreases CRF-R1 receptor mRNA levels in the rat amygdala.

Using semi-quantitative in situ hybridization, corticotropin-releasing factor (CRF) and CRF receptor 1 (CRF-R1) mRNA levels were determined in the rat hypothalamus and amygdala after short-term (10 days) and chronic (4 weeks) treatment with the antidepressant amitriptyline. We found that chronic treatment with amitriptyline produced a significant decrease in CRF mRNA (to 33% of control) in the hypothalamic paraventricular nucleus (PVN). Short-term or chronic amitriptyline treatment had no effect on CRF-R1 mRNA levels in the PVN. However, after chronic treatment, there was a significant decrease of CRF-R1 mRNA levels in the lateral + basolateral (to 60% of control), and in the medial (to 70% of control) amygdala nuclei. These results suggest that the tricyclic antidepressant amitriptyline may exert part of its effects through modulation of hypothalamic CRF and of CRF-R1 gene expression in the amygdala.

[Sarcomatoid carcinoma of the bladder]. / Carcinoma sarcomatoide della vescica.

Two cases of bladder sarcomatoid carcinoma, a rare tumor (0.3% of all bladder histotypes) are described and the difficult histological diagnosis and the utility of immunoassay markers analysed. Moreover, clinical observations are shortly discussed.

Bacterial lipopolysaccharide increases interleukin-6 and prostaglandin release in rat cortical type I astrocytes by different mechanisms: role of anti-inflammatory agents.

LPS stimulated IL-6 release in a concentration-dependent manner from rat cortical type I astrocytes. This stimulatory action was completely abolished by Dexamethasone (DEX), but was not affected by indomethacin (IND), a 5-cyclooxigenase inhibitor. LPS-induced IL-6 release was partially inhibited by BW 4AC, a 5-lipoxygenase inhibitor. LPS concentration-dependently increased the release of PGE2 from type I astrocytes, an effect completely inhibited by IND. To rule out the possibility that DEX was inhibiting LPS-induced IL-6 release by blocking IL-6 gene expression, we tested the effect of DEX on interleukin 1beta(IL-1)-induced IL-6 release. DEX slightly inhibited IL-1-induced IL-6 release, while IL-1 releasing action on IL-6 was significantly reduced by IND. The involvement of nitric oxide (NO) generation on LPS-induced IL-6 release was also studied. We found that L-NO-arginine, an inhibitor of nitric oxide synthase, concentration-dependently reduced LPS-induced IL-6 release in astrocytes. In conclusion, we provide evidence that LPS action on IL-6 and PGE2 release can be ascribed to the activation of different transduction mechanisms, which can be pharmacologically dissected with the aid of anti-inflammatory drugs.

[Correlation between the leak point pressure and the clinical grade of incontinence]. / Correlazione tra leak point pressure e grado clinico d'incontinenza.

We evaluated one-hundred and forty-six women with stress urinary incontinence (SUI), mean age 61.5 years, with clinical examination, urodynamics and patient history, grading the subjective degree of SUI according to SEAPI QMM classification. SUI was grade 1 in 73 pts (mean LPP 107, 7 cmH2O, mean maximal urethral closure pressure 59, 13 cmH2O), grade 2 in 36 (mLPP 55, 4 cmH2O, mMUCP 50, 3 cmH2O), grade 3 in 37 (mLPP 32, 29 cmH2O, mMUCP 33, 76 cmH2O). There is statistically significant difference in mLPP (p = 0.001) and mMUCP (p = 0.02) among three groups. The grade of SUI increases as the likelihood that LPP will be < or = 90 cmH2O or < or = 60 cmH2O (72.2% of pts with grade 2 has a LPP < or = 60 cmH2O, 100% of pts with grade 3 has a LPP < or = 60 cmH2O). Women with severe leakage and/or predisposing factor (PF) to intrinsic sphincter deficiency are likely to have a low LPP: all patients with SUI grade 3 and PF have a LPP < or = 60 cmH2O, 77% of pts with SUI grade 3 or PF has a LPP < or = 60 cmH2O. Women with higher grades of leakage and PF are significantly more likely to have a very low LPP and intrinsic sphincter deficiency.

Endotoxin decreases corticotropin-releasing factor receptor 1 messenger ribonucleic acid levels in the rat pituitary.

Bacterial endotoxins produce profound activation of the hypothalamo-pituitary-adrenal axis, mediated by stimulation of hypothalamic CRF neurons. Although a number of studies have described direct pituitary actions of inflammatory mediators, the effects of inflammatory stimuli on the sensitivity of corticotropes to CRF remain to be elucidated. The aim of this study was to determine the effects of inflammatory stress on the CRF receptor 1 (CRF-R1) messenger RNA (mRNA) levels in the rat pituitary. The systemic injection of endotoxin [lipopolysaccharide (LPS); 50 microg/kg, i.v.] increased plasma concentrations of ACTH and corticosterone. Ribonuclease protection analysis of total RNA isolated from individual whole pituitaries indicated that LPS produced a significant decrease in CRF-R1 mRNA that was evident by 2 h after injection (to 57% of control) and more marked by 6 h (to 38% of control). To evaluate whether the decrease in CRF-R1 mRNA was dependent upon increased exposure to CRF and/or vasopressin (AVP), LPS was injected with an anti-CRF antiserum, a CRF receptor antagonist (Astressin), or anti-AVP antiserum. A strong inhibition of the ACTH response to LPS was produced by pretreatment with anti-CRF antiserum, Astressin, or anti-AVP antiserum. However, these treatments had no effect on the decrease in CRF-R1 mRNA produced by LPS, indicating that neither CRF nor AVP are obligatory mediators of this pituitary response. The hypothesis that LPS might have direct pituitary effects on CRF-R1 mRNA levels was tested in vitro. Indeed, decreases in CRF-R1 mRNA to 43% and 53% of the control level were observed in rat anterior pituitary cell cultures that were treated with either LPS itself or the inflammatory mediator interleukin-1beta, respectively. Collectively, these results show that CRF receptor mRNA levels in the pituitary of the rat are markedly reduced by systemic LPS treatment and that this decrease is not dependent upon increased exposure of the pituitary to CRF or AVP, but may involve direct effects within the pituitary of either LPS itself or ensuing cytokine production.

[The conservative surgery of renal carcinoma]. / Chirurgia conservativa del carcinoma renale.

In the period 1986-1997, 387 cases of renal carcinoma were operated upon, at the Department of Urology, Parma General Hospital (Italy). Among these, thirty patients (all together 31 operations, 26 men and 5 women, mean age 58 +/- 11.3 years) have had conservative, nephron-sparing surgery; in 8 patients, conservative procedure was mandatory, due to previous contralateral nephrectomy or renal unreliability (4 RCC, 1 TCC, 1 severe injury, 1 pyonephrosis, 1 end stage insufficiency); in 23 patients, with normal contralateral kidney, the tumor was less than 4 cm in diameter and unique. Preoperatively, all cases had been staged by abdominal TC, chest X-ray, bone scan, renal angiography. 23 of 30 cases showed pathological stages I-II (pT1-T2), while 8 patients had stage III (pT3) tumors. After dismissal we recommended: abdominal echography after three months; again US and TC, chest X-ray after further three months. Then US and/or TC every six months, should the former results suggest a relapse, either locally and/or at a distance. Mean follow-up was 40 months. 6/30 patients (19.3%) died of metastatic disease (mean survival time: 27 months). 25 patients are alive and tumor free after a mean follow-up of 43.1 months. Immediate postoperative complications were 2 cases of urinary fistula treated by ureteral stenting.

[Valsalva leak point-pressure (LPP) and maximal urethral closure pressure (MUCP) in women with stress urinary incontinence (SUI)]. / Valsalva leak point-pressure (LPP) e massima pressione di chiusura uretrale (MUCP) in donne con incontinenza urinaria da stress (IUS).

One-hundred and twenty-two women with USI have been evaluated with clinical examination and urodynamics and divided in two groups: only in 74 patients with urinary loss during the Valsalva manoeuvre, LPP was compared to MUCP by linear regression analysis and its ability (cut-off = 60 cmH2O) to predict a MUCP < or = 20 cmH2O was tested. Weak correlations were observed between MUCP and LPP (r = 0.56). Fifty-two patients presented a LPP < or = 60 cmH2O, in 6 of them MUCP was < or = 20 cmH2O; none with LPP > 60 cmH2O showed a MUCP < or = 20 cmH2O. Median MUCP and intravesical pressures at the instant of leakage of patients with LPP < or = 60 cmH2O were significantly different from those of patients with LPP > 60 cmH2O (p < 0.01). The specificity and positive predictive value of LPP < or = 60 cmH2O for the detection of a "low pressure urethra" were respectively 32% and 11.5%, while sensibility and negative predictive value were 100%. LPP can not be regarded as a specific test for urethral sphincteric deficits. For its sensibility, it can be an useful screening tool for patients at high risk of type III urinary incontinence.

Evidence for the neuronal origin of immunoreactive interleukin-1 beta released by rat hypothalamic explants.

In this study, we have investigated the release of immunoreactive interleukin-1 beta (irIL-1 beta) from the rat hypothalamus in vitro. It was found that (1) tissue explants release sizable amounts of irIL-1 beta (ranging from 0.43 to 0.52 pg/mg of wet tissue) in 20 min incubations; (2) basal release in significantly increased by depolarization induced with 56 mM KCl; (3) K(+)-induced irIL-1 beta release is inhibited by the specific blocker of N-type calcium channels, omega-conotoxin, and by verapamil, but not by nifedipine; (4) K(+)-induced release is also inhibited by the Na+ channel blockers tetrodotoxin and lidocaine; (5) irIL-1 beta release is significantly increased by noradrenalin; such increase is antagonized by verapamil and the beta-blocker propranolol, but not by the alpha-blocker phentolamine. The present evidence suggests that irIL-1 beta released by rat hypothalamic explants following KCl depolarization is neuronal in origin.

[Conservative treatment of high-risk (T1G3) transitional carcinoma]. / Il trattamento conservativo nel carcinoma transizionale ad alto rischio (T1G3).

OBJECTIVE: This study evaluates the outcome of patients (pts) with primary T1G3 bladder cancer treated by transurethral resection (TUR) alone or followed by intravesical prophylaxis (BCG/Doxorubicin). Cistectomy was considered at disease progression. METHODS: Between 1/89 and 5/95 thirty-one pts with primary T1G3 bladder cancer were treated by TUR, in 24 followed by intravesical prophylaxis (13 with BCG, 11 with Doxorubicin). 7 pts had only TUR. RESULTS: At 42 months median follow up 45.2% pts (14/31) are disease free. The recurrence rate was 25.8% (8/31) and progression of disease was seen in 29.0% (9/31); mortality rate was 22.6% (7/31). In 13/31 pts treated by TUR + BCG 53.8% pts (7/13) are disease free. The recurrance rate was 23.1% (3/13) and progression of disease was seen in 23.1% (3/13) of cases; mortality rate was 23.1% (3/13). In 11/31 pts treated by TUR+Doxorubicin 54.5% pts (6/11) are disease free. The recurrance rate was 18.2% (2/11), progression of disease was seen in 27.3% (3/11) of cases of mortality rate of 9.1% (1/11). In 7/31 pts treated by TUR alone 14.3% pts (1/7) are disease free. The recurrance rate was 42.9% (3/7) and progression of disease was seen in 42.9% (3/7) of cases and mortality rate of 42.9% (3/7). Cistectomy was considered in 4 pts (3 for disease progression and 1 because of no disease free interval). The other pts with progression were not treated surgically because of their poor performance status. CONCLUSION: At a 42 months median follow up 77.4% pts (24/31) are alive (83.3% pts treated by TUR+intravesical prophylaxis). 64.5% pts (20/31) still have their bladder (66.6% pts treated by TUR+intravesical prophylaxis (16/24). We did not find a significative difference between prophylaxis with immunotherapy or chemotherapy. In conclusion we believe that the conservative management of high risk bladder transitional cell carcinoma T1G3 is feasible and allow us to plan cistectomy only in pts with progression or recurrance with no free interval without losing survival.