Surgical treatment of sacral chordoma: survival and prognostic factors.

INTRODUCTION: Sacral chordoma is a rare low-to-intermediate grade malignant tumour. The mainstay of treatment is still surgery with en bloc and wide resection margins, which can grant the best chances of a long-term control or cure of this disease. The first aim of this paper is to collect data about survival, time to local recurrence and metastasis among patients affected by sacral chordoma and primarily treated with surgery. The second aim is to analyze the influence of level resection, tumor volume and surgical margins on local recurrence. MATERIALS AND METHODS: The study population was composed of 14 patients treated with sacral chordoma resection at the National Tumour Institute of Naples-Pascale (Italy) from January 2000 to June 2013. The median follow-up was 84 months (range 24-132 months). The follow-up was characterized by: standard radiographs, MRI, and a CT scan of the chest annually. Time to recurrence or metastasis was calculated from the date of resection to the date of diagnosis of first recurrence or metastasis. RESULTS: Out of all the patients, six died (42.86 %) during the follow-up; 6 (42.86 %) had local recurrence; 4 (28.57 %) had metastasis. At univariate analysis wide surgical margins (R0) were associated with increased survival up to a local recurrence (OR = 0.0286; 95 % CI = 0.0014-0.5739; P = 0.026); the level of resection (OR = 3.33; 95 % CI = 0.3619-30.7025; P = 0.592) and tumour volume (P = 1) did not show a statistically significant correlation. DISCUSSION: Based on our experience, we hope all patients to be treated by surgery, the only good standard treatment of this disease. The resection should result in margins as wide as possible. For these reasons, it is essential for this disease to be treated in highly specialized centres because only a complete surgery can offer a chance to care for these patients. CONCLUSIONS: Solid survival at long-term follow-up can be achieved by a surgical resection performed with wide margins.

Low FasL levels promote proliferation of human bone marrow-derived mesenchymal stem cells, higher levels inhibit their differentiation into adipocytes.

Mesenchymal stem cells (MSCs) are multipotent progenitor cells that can differentiate into several cell types. Bone marrow (BM)-MSCs mainly differentiate into osteoblasts or adipocytes. MSC interactions with their microenvironment directly affect their self-renewal/differentiation program. Here, we show for the first time that Fas ligand (FasL), a well-explored proapoptotic cytokine, can promote proliferation of BM-derived MSCs in vitro and inhibits their differentiation into adipocytes. BM-MSCs treated with a low FasL dose (0.5 ng/ml) proliferated more rapidly than untreated cells without undergoing spontaneous differentiation or apoptosis, whereas higher doses (25 ng/ml) induced significant though not massive BM-MSC death, with surviving cells maintaining a stem cell phenotype. At the molecular level, 0.5 ng/ml FasL induced ERK1/2 phosphorylation and survivin upregulation, whereas 25 ng/ml FasL induced caspase activation. Importantly, 25 ng/ml FasL reversibly prevented BM-MSC differentiation into adipocytes by modulating peroxisome proliferator-activated receptor gamma (PPARγ) and FABP4/aP2 expression induced by adipogenic medium. All such effects were inhibited by anti-Fas neutralizing antibody. The in vitro data regarding adipogenesis were confirmed using Fas(lpr) mutant mice, where higher PPARγ and FABP4/aP2 mRNA and protein levels were documented in whole tibia. These data show for the first time that the FasL/Fas system can have a role in BM-MSC biology via regulation of both proliferation and adipogenesis, and may have clinical relevance because circulating Fas/FasL levels decline with age and several age-related conditions, including osteoporosis, are characterized by adipocyte accumulation in BM.

Differences in survivin location and Bcl-2 expression in CD30+ lymphoproliferative disorders of the skin compared with systemic anaplastic large cell lymphomas: an immunohistochemical study.

BACKGROUND: Cutaneous CD30+ lymphoproliferative disorders (LPDs) are a spectrum of disease associated with a favourable prognosis. Systemic anaplastic large cell lymphoma (ALCL), although morphologically and phenotypically similar, differs in clinical presentation and has a less favourable biological behaviour. Dysregulation of apoptosis, the process regulating cell population by programmed death, can explain the differences among these disorders. OBJECTIVES: We investigated the expression of two inhibitors of apoptosis, survivin and Bcl-2 protein, in serial skin lesion samples from CD30+ LPDs compared with systemic ALCL. METHODS: Immunohistochemical analysis with antibodies against anaplastic lymphoma kinase (ALK)-1 protein, survivin and Bcl-2 protein was performed in 10 cutaneous CD30+ LPDs (five lymphomatoid papulosis, five ALCL) and 18 systemic ALCLs. Reverse transcription-polymerase chain reaction studies for ALK and ALK/nucleophosmin were also performed. RESULTS: Cutaneous CD30+ LPDs shared a heterogeneous expression of cytoplasmic survivin with all systemic ALCLs, and of Bcl-2 with systemic ALK- ALCLs; however, they differ from systemic ALK- ALCLs because they lack nuclear survivin (P = 0.045), and from systemic ALK+ ALCLs by a higher expression of Bcl-2 (P = 0.045) and a lack of ALK-1. Overall, coexpression of Bcl-2 and nuclear survivin in CD30+ LPDs was associated with a less favourable disease survival. CONCLUSIONS: The different patterns of expression of Bcl-2 and survivin in CD30+ LPDs might have an impact on their different biological and clinical behaviour. Moreover, nuclear localization of survivin, similarly to ALK, may be a useful marker for predicting a systemic form of ALCL with cutaneous presentation.

Assessment of cyclin D1 overexpression as a prognostic factor in soft tissue sarcomas: role of laparoscopy and core needle biopsy.

BACKGROUND: This study was designed to test cyclin D1 as a prognostic marker in patients with soft tissue sarcomas (STS), and to evaluate the usefulness of laparoscopy for determining cyclin D1 overexpression. METHODS: The records of 62 patients with STS were collected: 28 with retroperitoneal STS (RSTS) and 34 with extremity STS (ESTS). A total of 51 patients underwent surgical resection, whereas 11 did not undergo surgery because of advanced tumor stage. Preoperative-intraoperative laparoscopic staging was performed for patients judged to be resectable at preoperative imaging. RESULTS: Cyclin D1 was overexpressed in 30 (58.8%) of 51 resected patients and in 10 (90.9%) of 11 nonresected patients. Laparoscopy avoided unnecessary laparotomy in 9 (32.1%) of 28 RSTS patients. CONCLUSIONS: High tumor grade, positive surgical margins, local recurrence, distant metastases, and cyclin D1 overexpression were related to poor survival. Multivariate analysis demonstrated cyclin D1 to be the only independent factor. Laparoscopy was shown to be useful for avoiding useless laparotomies.

Combined multistep approach in a locally advanced rectal cancer with sacral invasion: case report.

Composite pelvic resection with sacrectomy may provide good local control in case of locally advanced rectal cancer infiltrating the sacral bone. A combined multidisciplinary approach including chemotherapy and radiotherapy is here presented for a case of rectal tumor invading the sacrum.

In vitro H2O2 stress and patterns of mitochondrial damage in the NCTC 2544 continuous cell line--a morphologic and morphometric study.

The morpho-functional and energy condition of NCTC 2544 cells exposed for 1 hr to a high concentration of H2O2 (500 microM) was studied at 4 and 24 hr to investigate the short- and medium-term biomolecular mechanisms affecting energetic mitochondrial capability. Morphometric data obtained from ultrastructural investigations clearly showed significant modifications of the different mitochondrial parameters--numerical density (Nv), volume density (Vv) and Vv/Nv ratio, in interkinetic, apoptotic and mitotic cells after H2O2 exposure (from 4 to 24 hr). These results were confirmed by the detection at 24 hr of mitochondrial cytochrome c release in the cytosol, indicating impairment in mitochondrial membrane permeability. Data supporting these observations were obtained from the MTT test which showed reduced cell viability in H2O2 treated cultures at 4 hr and an even greater decrement at 24 hr. In conclusion our data imply that significant cause-effect relationships exist between the toxicity of reactive oxygen species (i.e. 500 microM H2O2) and morpho-structural mitochondrial damage in interkinetic, apoptotic and mitotic cells, respectively. They support previous results present both in the literature and also in one of our earlier papers which show that early nuclear DNA damage could initiate mitochondrial or intrinsic apoptotic pathway after H2O2 exposure.

Absence of somatic mutations in natriuretic peptide receptor type A gene in human aldosterone-secreting adenomas.

Somatic mutations of genes codifying for key regulatory proteins are the cause of different types of hormone-secreting adenomas. Natriuretic peptides (NP) are the strongest inhibitors of aldosterone secretion but aldosterone-secreting adenomas (aldosteronomas) are resistant to this inhibition and have reduced binding sites for NPs. The objective of this study was to sequence the entire coding region of the NP receptor type A (NPRA, codified by the Npr1 gene) to find loss-of-function somatic mutations. Total RNA was extracted from eight aldosteronomas and cDNA was synthesized. NPRA mRNA expression was evaluated by Northern blot analysis and compared with beta-actin mRNA as the housekeeping gene. Twelve primer couples were designed on the basis of the Npr1 gene organization to amplify, by PCR, all 22 coding exons of the gene. The two strands of amplified DNAs were purified and directly sequenced by automated capillary sequencer. NPRA mRNA expression did not differ among aldosteronomas. Npr1 open reading frame sequences obtained from eight aldosteronomas did not contain any mutation. The coding sequences of all 22 exons were identical in all samples and identical to published sequences. In the 3'-untranslated region (3'-UTR) a new length difference 3C/4C polymorphism was found at position 15 129 (three adenomas were 3C/4C and two were 3C/3C). Such a 3C/4C polymorphism was present in genomic DNA from 80 control subjects (25, 4C/4C; 40, 3C/4C; 15, 3C/3C). Mutations in the coding exons of the Npr1 gene do not appear to be a common cause of aldosteronomas. Moreover, the exons of Npr1 encoding for the translated portion of mRNA do not appear to be prone to polymorphisms. The polymorphism identified in the 3'-UTR might affect mRNA stability resulting in lower receptor synthesis, but it is not likely to confer a predisposition to the development of aldosteronomas.

PPARgamma expression in normal human placenta, hydatidiform mole and choriocarcinoma.

Peroxisome proliferator-activated receptor (PPAR) gamma belongs to a subclass of nuclear hormone receptors that execute their transcriptional functions as heterodimers with the retinoid X receptors (RXR). PPARgamma plays a pivotal role in cellular differentiation. This study investigated PPARgamma protein expression in normal human placentas, hydatidiform moles and choriocarcinoma, using immunohistochemical and Western blot analyses. In first trimester normal placenta, PPARgamma was mainly localized in the nuclei of the villous cytotrophoblastic cells, whereas at term it was mainly localized in the nuclei of the syncytiotrophoblast. Extravillous cytotrophoblast of cell islands and cell columns also showed nuclear PPARgamma immunostaining. A striking result was the altered expression patterns of PPARgamma in pathological tissues; PPARgamma showed a reduced immunostaining in the trophoblastic diseases. In hydatidiform moles, PPARgamma was mainly localized in the nuclei of the trophoblastic collections of the pathological villi and in the extravillous trophoblastic cells, whereas in the choriocarcinoma, only a few trophoblastic cells showed weak PPARgamma nuclear immunostaining. These findings suggest an involvement of PPARgamma in trophoblast differentiation during normal placental development. The down-regulation of PPARgamma expression in the gestational trophoblastic diseases analysed in this study provides a new insight into the progression of these pathologies.

Coenzyme Q10 levels in idiopathic and varicocele-associated asthenozoospermia.

Levels of coenzyme Q10 (CoQ10) and of its reduced and oxidized forms (ubiquinol, QH2, and ubiquinone, Qox) have been determined in sperm cells and seminal plasma of idiopathic (IDA) and varicocele-associated (VARA) asthenozoospermic patients and of controls. The results have shown significantly lower levels of coenzyme Q10 and of its reduced form, QH2, in semen samples from patients with asthenospermia; furthermore, the coenzyme Q10 content was mainly associated with spermatozoa. Interestingly, sperm cells from IDA patients exhibited significantly lower levels of CoQ10 and QH2 when compared to VARA ones. The QH2/Qox ratio was significantly lower in sperm cells from IDA patients and in seminal plasma from IDA and VARA patients when compared with the control group. The present data suggest that the QH2/Qox ratio may be an index of oxidative stress and its reduction, a risk factor for semen quality. Therefore, the present data could suggest that sperm cells, characterized by low motility and abnormal morphology, have low levels of coenzyme Q10. As a consequence, they could be less capable in dealing with oxidative stress which could lead to a reduced QH2/Qox ratio. Furthermore, the significantly lower levels of CoQ10 and QH2 levels in sperm cells from IDA patients, when compared to VARA ones, enable us to hypothesize a pathogenetic role of antioxidant impairment, at least as a cofactor, in idiopathic forms of asthenozoospermia.

Primary monotypic epithelioid angiomyolipoma of bone.

AIMS: Monotypic epithelioid angiomyolipoma is a distinct and definable variant of angiomyolipoma, composed of monomorphous epithelioid cells that show HMB45 immunoreactivity. Angiomyolipoma, including its morphological variants, belongs to the family of perivascular epithelioid cell tumour. METHODS AND RESULTS: The tumour was examined using immunohistochemical staining and by transmission electron microscopy. Neoplastic cells showed a cytoplasmic granular positivity for HMB45. CONCLUSIONS: Extrarenal angiomyolipomas are rare and, to the best of our knowledge, this is the first reported case of a primary monotypic epithelioid angiomyolipoma of bone in a patient without evidence of tuberous sclerosis.

Neoadjuvant chemotherapy for intermediate/high-grade soft tissue sarcomas: five-year results with epirubicin and ifosfamide.

BACKGROUND: Neoadjuvant chemotherapy for intermediate/high-grade soft tissue sarcomas (STS) may provide some advantages for facilitating the surgical resection of the tumor and for disease control. However its role as induction therapy before surgery should still be proved. PATIENTS AND METHODS: Twenty-one patients with intermediate/high-grade STS and tumor size > or = 5 cm were consecutively treated from 1997 to 2001 with neoadjuvant chemotherapy based on epirubicin 60 mg/m2/day on days 1 and 2 and ifosfamide 1.8 gr/m2/day on days 1 through 5 every three weeks. Evaluation of objective tumor response and toxicity were carried out according to WHO criteria. RESULTS: Nine partial responses were documented; stable disease in 11 patients, progressive disease in one patient. Apart from nine cases of grade 4 neutropenia, the treatment was generally well-tolerated. Twelve patients underwent conservative and limb salvage surgery. CONCLUSION: This therapeutic approach seems to be effective in facilitating surgery. Neutropenia was the most significant toxicity but it was preventable or medically treatable with G-CSF support.

Src-dependence and pertussis-toxin sensitivity of urokinase receptor-dependent chemotaxis and cytoskeleton reorganization in rat smooth muscle cells.

The catalytically inactive precursor of urokinase-type plasminogen activator (pro-u-PA) induced a chemotactic response in rat smooth muscle cells (RSMC) through binding to the membrane receptor of urokinase (u-PA receptor [u-PAR]). A soluble form of u-PAR activated by chymotrypsin cleavage as well as a peptide located between domain 1 and 2 of u-PAR reproduced the effect of pro-u-PA on cell migration. The chemotactic pro-u-PA effect correlates with a dramatic reorganization of actin cytoskeleton, of adhesion plaques, and with major cell shape changes in RSMC. Pro-u-PA induced a decrease in stress fiber content, membrane ruffling, actin ring formation, and disruption leading to the characteristic elongated cell shape of motile cells with an actin semi-ring located close to the leading edge of cells. u-PAR effects on both chemotaxis and cytoskeleton were sensitive to pertussis toxin and, hence, possibly require G proteins. u-PAR effects are accompanied by a relocation of u-PAR, vitronectin receptor (VNR) alphavbeta3, beta1 integrin subunit, and Src tyrosine kinase to the leading membrane of migrating cells. In conclusion, our data show that pro-u-PA, via binding to u-PAR, controls a signaling pathway, regulated by tyrosine kinases and possibly G proteins, leading to cell cytoskeleton reorganization and cell migration.

Multicentric giant cell tumor with viral-like inclusions associated with Paget's disease of bone: a case treated by steroid therapy.

We report a case of giant cell tumor of bone (GCTB) associated with Paget's disease unsuccefully treated with radiotherapy for some years but dramatically reduced in size with high dose of dexamethasone within few days. An ultrastructural study showed intranuclear virus-like inclusions in the multinucleated giant cells. The patient was then switched to prednisone plus diclofenac and he is still in almost complete remission after two years. The patient was from Avellino area, a small town in Southern Italy.

Expression of Met protein and urokinase-type plasminogen activator receptor (uPA-R) in papillary carcinoma of the thyroid.

Met protein encoded by MET oncogene is the high affinity receptor for hepatocyte growth factor (HGF)/scatter factor (SF). HGF/SF has to be cleaved in its heterodimeric form by the urokinase-type plasminogen activator (uPA) to become active as a ligand for Met receptor. The expression of Met protein and of the high affinity receptor for uPA (uPA-R) was investigated in 39 samples of papillary carcinoma using immunohistochemistry. Reactivity for Met protein was present in 33 of 34 tumours, mostly with a diffuse pattern of staining. Reactivity for uPA-R was present in 78 per cent of papillary tumours and exhibited a pattern of staining similar to that of Met protein. Staining for uPA-R was present in 23 of 25 cases (92 per cent) of papillary carcinoma with prominent sclerosis, and in only 1 of 7 cases (14 per cent) without sclerosis. Peritumoural normal thyroid, follicular adenomas, and follicular carcinomas were negative for Met protein and for uPA-R. Hyperfunctioning tall thyroid cells showed weak membrane reactivity for uPA-R and for Met protein. The findings of immunohistochemistry were confirmed at the mRNA level using in situ hybridization, since the signal for uPA-R and Met RNAs was detected in most tumour cells of five cases of papillary carcinoma.

A urokinase-sensitive region of the human urokinase receptor is responsible for its chemotactic activity.

The role of urokinase-type plasminogen activator (uPA) and its receptor (uPAR/CD87) in cell migration and invasion is well substantiated. Recently, uPA has been shown to be essential in cell migration, since uPA-/- mice are greatly impaired in inflammatory cell recruitment. We have shown previously that the uPA-induced chemotaxis requires interaction with and modification of uPAR/CD87, which is the true chemoattracting molecule acting through an unidentified cell surface component which mediates this cell surface chemokine activity. By expressing and testing several uPAR/CD87 variants, we have located and functionally characterized a potent uPAR/CD87 epitope that mimics the effects of the uPA-uPAR interaction. The chemotactic activity lies in the region linking domains 1 and 2, the only protease-sensitive region of uPAR/CD87, efficiently cleaved by uPA at physiological concentrations. Synthetic peptides carrying this epitope promote chemotaxis and activate p56/p59(hck) tyrosine kinase. Both chemotaxis and kinase activation are pertussis toxin sensitive, involving a Gi/o protein in the pathway.

Osteomyelitis by Paracoccidioides brasiliensis (South American blastomycosis): cytologic diagnosis on fine-needle aspiration biopsy smears: a case report.

A case of osteomyelitis caused by Paracoccidioides brasiliensis primarily diagnosed by means of fine-needle aspiration biopsy is reported here in a 60-yr-old Italian patient who had lived in Venezuela for 40 yr. The cytologic and electron microscopic features of the exudate aspirated from a left femoral osteolytic area are described, and the differential diagnosis of this mycotic infection is discussed briefly.

Integrin-dependent induction of functional urokinase receptors in primary T lymphocytes.

In order to reach the sites of inflammation, lymphocytes leave the bloodstream and migrate into peripheral tissues, in a process involving integrin-mediated adhesion to the vascular endothelium, followed by transmigration across the endothelial barrier and through the underlying interstitial matrix. We have investigated the role of the plasminogen activator/plasmin system in normal T cell migration. Receptors for urokinase plasminogen activator (uPAR) were not expressed in resting T lymphocytes, but could be efficiently induced at the mRNA and protein level by coclustering of the antigen receptor complex and beta1 or beta2 integrins, through a signalling pathway involving both protein kinase C activation and an increase in intracellular cyclic AMP. Catalytic activation of plasminogen by uPAR-expressing T cells promoted their migration through an extracellular matrix in vitro. Plasmin-induced invasion was inhibited by plasmin-and urokinase inhibitors and by anti-uPAR antibodies. Finally, cytofluorimetric and immunohistochemical analysis of primary human tumor specimens showed the presence of uPAR positive infiltrating T cells in vivo. Collectively, these findings suggest that plasminogen activation may play a role in lymphocyte migration in vivo, and that integrin-dependent expression of membrane-associated endopeptidases could represent an additional step in the regulated process of leukocyte transmigration.