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Biomedical subjects

P Chieffi

Publications and source records attributed to P Chieffi.

At least 19 recordsLinked to original sources
PubMed2004

Aurora B expression in normal testis and seminomas.

Aurora/Ipl1-related kinases are a conserved family of proteins that have multiple functions during mitotic progression. High levels of Aurora kinases are characteristic of rapidly dividing cells and tumours. Aurora B encodes a protein that associates with condensing chromatin, concentrates at centromeres, and then relocates onto the central spindle at anaphase. In this study the expression and the localisation of Aurora B throughout germinal epithelial progression in normal testis and its neoplastic counterpart were analysed. Immunocytochemistry and RT-PCR analysis of mouse germinal epithelium cells showed the presence of Aurora B in spermatogonia and occasionally in spermatocytes. Western blot analysis revealed the typical Aurora B isoform ( approximately 41 kDa) in the same cellular types. A similar distribution was observed in human testis by immunohistochemistry. Moreover, the distribution and the expression of Aurora B were investigated in neoplasms derived from germ cells. Surgical samples of seminomas were analysed, and a high percentage of Aurora B positive cells (51%) was detected; the expression of Aurora B was significantly related to the MIB-1 proliferation marker (R=0.816). The data presented here demonstrate that Aurora B expression occurs in spermatogonial division. Furthermore, our results indicate that the expression of Aurora B is a consistent feature of human seminomas.

Animals↗
PubMed2003

Loss of oestrogen receptor beta, high PCNA and p53 expression and aneuploidy as markers of worse prognosis in ovarian granulosa cell tumours.

AIMS: Ovarian granulosa cell tumour (OGCT) is a sex-cord stromal tumour with a general trend toward late relapse and/or metastasis. However, mortality rate corrected for long-term follow-up shows that about 50% of patients die within 20 years of diagnosis. Classical clinicopathological parameters are unable to predict the biological behaviour of OGCT. The involvement of a recently characterized subtype of oestrogen receptor, ERbeta, in ovarian carcinogenesis has been hypothesized. METHODS AND RESULTS: We examined by immunohistochemistry the expression of ERbeta, proliferating cell nuclear antigen (PCNA) and p53 in a selected series of 30 OGCT, to evaluate their role in the prognostic evaluation of this tumour. Immunohistochemistry was performed on formalin-fixed paraffin-embedded sections. Results were compared with the DNA-ploidy of the tumours (evaluated by image analysis) and with the follow-up data of the patients. CONCLUSIONS: Loss of ERbeta expression, high PCNA expression and aneuploidy, characterized a subgroup of OGCT with a worse outcome. The identification of a high-risk subclass of OGCT may be of primary importance in addressing appropriate therapeutic strategies, offering the chance to prevent relapses and metastases by using adjunctive, specifically targetted, more aggressive therapies.

Adolescent↗
PubMed2001

Expression of the apoptosis inhibitor survivin in aggressive squamous cell carcinoma.

Deregulated expression of inhibitors of apoptosis (programmed cell death) may contribute to cancer by aberrantly extending cell viability and facilitating the insurgence of resistance to therapy. In this study, we investigated the potential expression and prognostic significance of the apoptosis inhibitor survivin in squamous cell carcinoma (SCC). A series of 135 cases of SCC including 46 oral SCC and 89 cutaneous SCC was analyzed for survivin expression by immunohistochemistry and Western blotting. Survivin was found in 57 cases (64%) of skin SCC and 26 cases (56%) of oral SCC, with weighted survivin scores ranging from 1 to 12. In contrast, normal oral epithelium, normal skin epithelium, and skin annexa did not express survivin. Survivin expression significantly (P < 0.05) segregated with high-grade and undifferentiated tumors with size >1.5 cm and invariably associated with lymph node metastasis. These data suggest that survivin expression may predictively identify cases of SCC with more aggressive and invasive clinical phenotype, potentially warranting closer follow-up protocols.

Apoptosis↗
PubMed2001

RNF4 is a growth inhibitor expressed in germ cells but not in human testicular tumors.

The RING-finger protein RNF4 modulates both steroid-receptor-dependent and basal transcription and interacts with a variety of nuclear proteins involved in cell growth control. RNF4 is expressed at very high levels in testis and at much lower levels in several other tissues. We show that in germ cells RNF4 expression is strongly modulated during progression of spermatogonia to spermatids, with a peak in spermatocytes. Analysis of human testicular germ cell tumors shows that RNF4 is not expressed in all tumors analyzed including seminomas, the highly malignant embryonal carcinomas, yolk sac, and mixed germ cell tumors. We also show that the ectopically expressed RNF4 gene inhibits cell proliferation of both somatic and germ cell tumor-derived cells. Mutation of critical cysteine residues in the RING finger domain abolished the RNF4 growth inhibition activity. Our results suggest that the lack of RNF4 expression may play a role in the progression of testicular tumors.

Animals↗
PubMed2001

Expression of PCNA in the testis of the lizard, Podarcis s. sicula: an endogenous molecular marker of mitotic germinal epithelium proliferation.

Proliferating cell nuclear antigen (PCNA) plays an essential role in nucleic acid metabolism as a component of the replication and repair machinery. This protein encircles DNA and can slide bidirectionally along the duplex binding to DNA polymerase delta and epsilon. It is well known that PCNA interacts with proteins involved in the cell cycle. The PCNA interactions with different cellular proteins and the importance of these interactions are discussed. To examine mitotic germinal epithelium proliferation during annual discontinuous spermatogenesis in the lizard Podarcis s. sicula, temporal and the spatial PCNA expression were investigated, and provide a useful endogenous molecular marker.

Animals↗
PubMed2001

Variations of proline-rich kinase Pyk2 expression correlate with prostate cancer progression.

Proline-rich kinase 2 (Pyk2), also known as CAKbeta (cell adhesion kinase beta), is a cytoplasmic tyrosine kinase that is structurally related to focal adhesion kinase. Pyk2 is expressed in different cell types including brain cells, fibroblasts, platelets, and other hemopoietic cells. Pyk2 is rapidly tyrosine phosphorylated in response to diverse extracellular signals acting via different post receptor pathways. We have investigated whether this protein kinase is functionally expressed in normal and neoplastic prostate tissues. In this study, we demonstrate that Pyk2 is expressed only in normal epithelial prostate tissue and in benign prostatic hyperplasia, whereas its expression progressively declines with an increasing grade of malignancy of prostate cancer.

Enzyme Activation↗
PubMed2000

PCNA in the testis of the frog, Rana esculenta: a molecular marker of the mitotic testicular epithelium proliferation.

Proliferating cell nuclear antigen (PCNA) plays an essential role in nucleic acid metabolism as a component of the replication and repair machinery. This toroidal-shaped protein encircles DNA and can slide bidirectionally along the duplex. One of the well-established functions for PCNA is its role as the processing factor for DNA polymerase delta and epsilon. It has become apparent that PCNA interacts with proteins involved in the cell cycle. The PCNA interactions with different cellular proteins and the importance of these interactions are also discussed. To examine the different mitotic testicular epithelium proliferation during the annual discontinuous frog (Rana esculenta) spermatogenesis, the temporal and the spatial PCNA expression are described and give a useful endogenous molecular marker.

Animals↗
PubMed2000

Association between the expression of E1A oncogene and increased sensitivity to growth inhibition induced by sustained levels of cAMP in rat thyroid cells.

OBJECTIVE: The aim of this study was to investigate: (i) whether a persistent increase of cAMP interferes with the proliferation of transformed thyroid cells, and (ii) whether the degree of malignancy is correlated with the sensitivity to a transient and/or sustained increase in intracellular cAMP levels. DESIGN AND METHODS: To address these questions we used thyroid cell lines transformed with E1A oncogene from adenoviruses 5 (PC E1A cell line) or 2 (PC HE4 cell line), or infected with the polyoma murine leukemia virus (PC PyMLV cell line) carrying the middle T gene of the polyoma virus, or, finally, expressing both E1A and PyMLV. These cell lines present various degrees of malignancy: PC EIA and PC HE4 cells are not tumorigenic; PC PyMLV cells induce non-invasive tumors after a long latency period; and PC EIA+PyMLV cells are highly tumorigenic. RESULTS AND CONCLUSIONS: Thyroid cell proliferation required the transient increase of intracellular cAMP levels, while persistent elevation of cAMP blocked the proliferation of normal thyroid PC Cl 3 cells and of PC Cl 3 cells transformed by a variety of different oncogenes. In addition, sustained levels of cAMP induced apoptosis in cells carrying the adenovirus EIA oncogene, but not in cells transformed with other oncogenes or in the wild-type PC Cl 3 cells. Furthermore, middle T gene of the polyoma virus seemed to afford protection only from apoptosis induced by cAMP when middle T is present in thyroid cells along with the E1A gene.

Adenovirus E1A Proteins↗
PubMed2000

Estradiol-induced mitogen-activated protein kinase (extracellular signal-regulated kinase 1 and 2) activity in the frog (Rana esculenta) testis.

Several lines of evidence support a key role of estradiol-17beta (E(2)) in male fertility. We have used a non-mammalian vertebrate model, the frog Rana esculenta, to investigate the regulation of extracellular signal-regulated kinase 1 and 2 (ERK1/2) activity in the testis during the annual sexual cycle and to study whether E(2 )exerts a role in spermatogenesis through the regulation of ERK1/2 activity. ERK1/2 proteins are present in the cytoplasm and nucleus of the primary and secondary spermatogonia (SPG), and in the nucleus of primary spermatocytes. The annual E(2) profile shows a progressive increase during active spermatogenesis with a peak in the month of June. In parallel, ERK1/2 are highly phosphorylated during the period of active spermatogenesis (from April to July) compared with the regressive period (September/October) and winter stasis (from November to March). E(2) treatment induces the proliferation of primary SPG, possibly via the activation of ERK1/2, and this effect is counteracted by the anti-estrogen ICI 182-780.

Animals↗
PubMed1999

Endogenous insulin-like growth factors regulate the proliferation of TSH-independent mutants derived from FRTL5 cells.

TSH-independent mutant clones (M cells) derived from FRTL5 cells, proliferate vigorously in the absence of TSH. The growth of M cells is stimulated by IGF-I in a dose-dependent fashion, but it is not influenced by TSH. Sm1.2, an antibody against IGF-I cross-reacting with IGF-II, significantly decreases basal DNA synthesis in the M cells. Binding of 125I-IGF-I to M cells is significantly lower than that to FRTL5 cells. M cells produce in their culture medium IGF-like peptides which appear to influence their basal DNA synthesis and the availability of type I receptors to bind exogenous IGF-I.

Animals↗
PubMed1999

Fos localization in cytosolic and nuclear compartments in neurones of the frog, Rana esculenta, brain: an analysis carried out in parallel with GnRH molecular forms.

C-fos activity was determined in the brain of the frog, Rana esculenta, during the annual sexual cycle. The localization of GnRH molecular forms (mammalian- and chicken-GnRHII) was also carried out to determine whether or not the proto-oncogene and the peptides showed a functional relationship. Northern blot analysis of total RNA revealed the presence of a single strong signal of c-fos like mRNA of 1.9 Kb during February and April. This was followed by expression of c-Fos protein (Fos) in several brain areas during March and July shown by immunocytochemistry. In particular, the olfactory region, the lateral and medial pallium, the nucleus lateralis septi, the ventral striatum, the caudal region of the anterior preoptic area, the suprachiasmatic nucleus, the ventral thalamus, tori semicircularis and ependymal layers of the tectum were immunostained. There was no overlap between Fos immunoreactive perikarya and GnRH immunoreactive perikarya (e.g. gonadotrophin-releasing hormone (GnRH) in the rostral part and Fos in the caudal region of the anterior preoptic area). Interestingly, a cytoplasmic localization of Fos was also observed by immunocytochemistry and gel retardation experiments supported this observation. Cytoplasmic extracts from September-October animals bound the AP1 oligonucleotide. The complex was not available in the nuclear extracts from the same preparation, suggesting that, besides Fos, Jun products were also present. Conversely, nuclear but not cytosolic binding was detected in the brain of animals collected in July. In conclusion, we show that Fos and GnRH activity does not correlate in the frog brain and, for the first time in a vertebrate species, we give evidence of a cytoplasmic AP1 complex in neuronal cells.

Animals↗
PubMed1999

C-Jun phosphorylation (Ser-63) in the testis of the lizard, Podarcis s. sicula.

Proto-oncogenes play an important role in the regulation of cellular growth and differentiation. C-Jun activity has been studied in the testis of a non mammalian vertebrate, the lizard Podarcis s. sicula, during two different periods: winter stasis and the breeding season. C-Jun protein was localized by immunocytochemistry in the cytoplasm of the spermatogonia (SPG) and stage I and II spermatocytes (SPC) during the winter stasis (from December until March), while the protein was present in the nuclei of the same cells during the active spermatogenic period (April/May). The different localization of c-Jun has been confirmed by Western blot and immunoprecipitation analysis. In addition, when Jun is present in the nuclear compartment, it is phosphorylated on Ser-63 and is complexed with Fos protein. These data suggest that the nuclear localization of the Jun protein in the SPG and stage I and II SPC, with strong phosphorylation on Ser-63 during the breeding period, could be the signal of increasing transcriptional activity in the lizard testis.

Animals↗
PubMed1997

17 beta-estradiol effects on mast cell number and spermatogonial mitotic index in the testis of the frog, Rana esculenta.

Estrogen affects mast cell activity and cellular proliferation in several vertebrate tissues. Due to the presence of mast cells in the interstitial tissue of the testis and due to the annual changes of spermatogonial proliferation and estradiol level in the frog, Rana esculenta, we have studied the possible regulation of mast cell number (MCN) and primary spermatogonial mitosis exerted by 17 beta-estradiol (E2). MCN changed in the testis during the annual reproductive cycle, showing peaks in December and in May. Administration of E2 elicited an increase of MCN both in intact and hypophysectomized frogs, and this effect was counteracted by tamoxifen. In vitro experiments indicated that E2 induced an increase of both MCN and the primary spermatogonial mitotic index. Moreover, a significant decrease of intratesticular androgen content was measured in E2-treated testes. All the in vitro effects induced by E2 were counteracted by tamoxifen. In conclusion, our data indicate that in Rana esculenta testis E2 increase MCN and primary spermatogonial mitotic index via intratesticular mechanisms. A possible involvement of the decrease of androgen levels exerted by E2 in the mechanism underlying the increase of MCN is also considered.

Animals↗
PubMed1997

Number of mast cells in the Harderian gland of the lizard Podarcis sicula sicula (Raf): the annual cycle and its relation to environmental factors and estradiol administration.

The Harderian gland of the lizard Podarcis sicula sicula (Raf) contains connective tissue type mast cells whose numbers vary during the year showing two peaks, one in spring the other in winter. No sex differences are found throughout the year. Thermal and photoperiodic manipulations indicate that only temperature influences mast cell number (MCN) both in winter and in summer but not in spring. In animals exposed to high temperatures in February (but not in May) MCN declined, while exposure to low temperature in July had the opposite effect. Estradiol treatment of the February and April lizards increased MCN, an effect counteracted by the synthetic antiestrogen tamoxifen; in July lizards, this did not occur. In animals exposed to a high temperature in February, estradiol had no effect, as in animals exposed to low temperatures in July. These data suggest that in spring MCN seems to be more responsive to hormonal stimuli rather than external cues (temperature), while in summer MCN is more sensitive to temperature than to hormonal stimuli (estradiol). Both humoral and external factors are concluded to influence mast cell numbers in the Harderian gland of the lizard P. sicula sicula.

Animals↗
PubMed1997

Proto-oncogene activity in the testis of the lizard, Podarcis s. sicula, during the annual reproductive cycle.

Since proto-oncogenes play a central role in the regulation of cellular growth and differentiation, localization of MYC, FOS, and JUN proteins has been studied in the testis of the lizard, Podarcis s. sicula, during the annual reproductive cycle by immunocytochemistry using antisera against c-myc, c-fos, and c-jun products. MYC was localized in the nuclei of spermatogonia (SPG), I and II spermatocytes (SPC), and spermatids (SPT). Strong immunoreactivity was detected in Sertoli cells just prior to the onset of the early spring spermatogenic wave coinciding with the androgen peak. FOS protein was present in the nuclei of SPG and SPC. In SPG an exclusive nuclear localization was seen during the active spermatogenic period (February-March and September). A perinuclear localization was observed during other months. Immunoreactivity in Sertoli cells was also observed during the periods of active spermatogenesis. JUN protein was localized in the cytoplasm of SPG as well as in I and II SPC and was detected in the nuclei of I and II SPC during April and October when spermatogenic waves occur. These data suggest that proto-oncogene activities have regulatory roles in the spermatogenesis of the lizard.

Amino Acid Sequence↗
PubMed1997

Oestrogen control of the sexual dimorphism in the Harderian gland of Xenopus laevis.

Xenopus laevis shows a sexual dimorphism of the electrophoretic pattern of Harderian gland (HG) proteins. The male pattern displays three protein fractions whose molecular sizes are approx. 205, 180 and 78 kDa, respectively, and which are absent in the female pattern. Conversely, the female pattern displays two protein fractions of approx. 190 and 76 kDa, respectively. This sexual dimorphism led us to hypothesize a sex steroid control of the HG. Administration of 17beta-oestradiol to male Xenopus converts the male protein pattern into the female one, while the administration of testosterone to the female has no effect. In this respect neither Northern analysis nor the RNase-protection assay performed using a 213 bp encoding for the androgen-binding domain reveals the presence of an androgen receptor mRNA in Xenopus HG. Conversely, Northern analysis has shown an oestrogen receptor mRNA whose size is approx. 6.5 kb and the RNase-protection assay performed by using a 197 bp encoding for the oestrogen-binding domain has also displayed the presence of an oestrogen receptor mRNA in the female HG but not in the male one. In addition, the oestrogen administration to male Xenopus induces the appearance of an oestrogen receptor mRNA. Androgen administration to female toad is ineffective. Taken together, all these findings suggest that in Xenopus laevis oestrogens are involved into the HG physiology. The appearance of an oestrogen receptor mRNA in the oestradiol treated males supports the hypothesis of the occurrence of autoinduction of oestrogen receptor mRNA expression in the HG.

Animals↗