-
ILAR journal / National Research Council, Institute of Laboratory Animal Resources 09/2012; ILAR Journal(54). · 2.33 Impact Factor
-
ILAR journal / National Research Council, Institute of Laboratory Animal Resources 09/2012; · 2.33 Impact Factor
-
Alba Rocco,
Eleonora Liguori,
Giuseppe Pirozzi,
Virginia Tirino,
Debora Compare,
Renato Franco,
Fabiana Tatangelo,
Raffaele Palaia,
Francesco Paolo D'Armiento,
Giorgia Pollastrone, Andrea Affuso,
Enrico Coppola Bottazzi,
Stefania Masone,
Giovanni Persico,
Gerardo Nardone
[show abstract]
[hide abstract]
ABSTRACT: Emerging evidence suggests that tumors contain and are driven by a cellular component that displays stem cell properties, the so-called cancer stem cells (CSCs). CSCs have been identified in several solid human cancers; however, there are no data about CSCs in primary human gastric cancer (GC). By using CD133 and CD44 cell surface markers we investigated whether primary human GCs contain a cell subset expressing stem-like properties and whether this subpopulation has tumor-initiating properties in xenograft transplantation experiments. We examined tissues from 44 patients who underwent gastrectomy for primary GC. The tumorigenicity of the cells separated by flow cytometry using CD133 and CD44 surface markers was tested by subcutaneous or intraperitoneum injection in NOD/SCID and nude mice. GCs included in the study were intestinal in 34 cases and diffuse in 10 cases. All samples contained surface marker-positive cells: CD133+ mean percentage 10.6% and CD133+/CD44+ mean percentage 27.7%, irrespective of cancer phenotype or grade of differentiation. Purified CD133+ and CD133+/CD44+ cells, obtained in sufficient number only in 12 intestinal type GC cases, failed to reproduce cancer in two mice models. However, the unseparated cells produced glandular-like structures in 70% of the mice inoculated. In conclusion, although CD133+ and CD133+/CD44+ were detectable in human primary GCs, they neither expressed stem-like properties nor exhibited tumor-initiating properties in xenograft transplantation experiments. J. Cell. Physiol. 227: 2686–2693, 2012. © 2011 Wiley Periodicals, Inc.
Journal of Cellular Physiology 02/2012; 227(6):2686 - 2693. · 3.87 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Animal experiments are necessary for a better understanding of diseases and for developing new therapeutic strategies. The mouse (Mus musculus) is currently the most popular laboratory animal in biomedical research. Experimental procedures on animals often require anesthesia and/or analgesia to obtain adequate immobilization and to reduce stress or pain. Mice anesthesia is challenging for several reasons including the animals' size, metabolic rate, and the high risk of hypothermia and hypoglycemia. Moreover, anesthetic agents influence physiological parameters, further interfering with experimental results. Small animal imaging procedures are increasingly used in biomedical research both because the animals allow in vivo monitoring and because they are readily available for longitudinal and noninvasive studies as well as investigations into the evolution of diseases and the effects of new therapies. Anesthesia must adapt to the imaging technique, the procedure length, and the aim of the study. The purpose of this article is to review the existing literature on anesthetic protocols adopted in mice for molecular imaging studies and to report our experience.
ILAR journal / National Research Council, Institute of Laboratory Animal Resources 01/2012; 53(1):55-69. · 2.33 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Animal experiments are necessary for a better understanding of diseases and for developing new therapeutic strategies. The mouse (Mus musculus) is currently the most popular laboratory animal in biomedical research. Mice imaging procedures are increasingly used in preclinical research because they allow in vivo monitoring and they are readily available for longitudinal and noninvasive studies as well as investigations into the evolution of diseases and the effects of new therapies. New imaging techniques and sophisticated laboratory animal imaging tools are currently producing a large body of evidence about the possible interference of anesthesia with different imaging methods that have the potential to compromise the results of in vivo studies. The purpose of this article is to review the existing literature on molecular imaging studies in mice, to describe the effects of different anesthetic protocols on their outcome, and to report our own experience with such studies.
ILAR journal / National Research Council, Institute of Laboratory Animal Resources 01/2012; 53(1):70-81. · 2.33 Impact Factor
-
Alba Rocco,
Eleonora Liguori,
Giuseppe Pirozzi,
Virginia Tirino,
Debora Compare,
Renato Franco,
Fabiana Tatangelo,
Raffaele Palaia,
Francesco Paolo D'Armiento,
Giorgia Pollastrone, Andrea Affuso,
Enrico Coppola Bottazzi,
Stefania Masone,
Giovanni Persico,
Gerardo Nardone
[show abstract]
[hide abstract]
ABSTRACT: Emerging evidence suggests that tumors contain and are driven by a cellular component that displays stem cell properties, the so-called cancer stem cells (CSCs). CSCs have been identified in several solid human cancers; however, there are no data about CSCs in primary human gastric cancer (GC). By using CD133 and CD44 cell surface markers we investigated whether primary human GCs contain a cell subset expressing stem-like properties and whether this subpopulation has tumor-initiating properties in xenograft transplantation experiments. We examined tissues from 44 patients who underwent gastrectomy for primary GC. The tumorigenicity of the cells separated by flow cytometry using CD133 and CD44 surface markers was tested by subcutaneous or intraperitoneum injection in NOD/SCID and nude mice. GCs included in the study were intestinal in 34 cases and diffuse in 10 cases. All samples contained surface marker-positive cells: CD133(+) mean percentage 10.6% and CD133(+)/CD44(+) mean percentage 27.7%, irrespective of cancer phenotype or grade of differentiation. Purified CD133(+) and CD133(+)/CD44(+) cells, obtained in sufficient number only in 12 intestinal type GC cases, failed to reproduce cancer in two mice models. However, the unseparated cells produced glandular-like structures in 70% of the mice inoculated. In conclusion, although CD133(+) and CD133(+)/CD44(+) were detectable in human primary GCs, they neither expressed stem-like properties nor exhibited tumor-initiating properties in xenograft transplantation experiments.
Journal of Cellular Physiology 09/2011; 227(6):2686-93. · 3.87 Impact Factor
-
Rita Mancini,
Enrico Giarnieri,
Claudia De Vitis,
Donatella Malanga,
Giuseppe Roscilli,
Alessia Noto,
Emanuele Marra,
Carmelo Laudanna,
Pietro Zoppoli,
Pasquale De Luca, [......],
Giuseppe Mesiti,
Luigi Aurisicchio,
Alberto Ricci,
Salvatore Mariotta,
Lara Pisani,
Claudio Andreetti,
Giuseppe Viglietto,
Erino A Rendina,
Maria Rosaria Giovagnoli,
Gennaro Ciliberto
[show abstract]
[hide abstract]
ABSTRACT: Malignant pleural effusions (MPEs) could represent an excellent source to culture a wide variety of cancer cells from different donors. In this study, we set up culture conditions for cancer cells deriving from MPEs of several patients affected by the most frequent form of lung cancer, namely the subset of non small cell lung cancers (NSCLC) classified as Lung Adenocarcinomas (AdenoCa) which account for approximately 40% of lung cancer cases. AdenoCa malignant pleural effusions gave rise to in vitro cultures both in adherent and/or in spheroid conditions in almost all cases analyzed. We characterized in greater detail two samples which showed the most efficient propagation in vitro. In these samples we also compared gene profiles of spheroid vs adherent cultures and identified a set of differentially expressed genes. Finally we achieved efficient tumor engraftment in recipient NOD/SCID mice, also upon inoculation of small number of cells, thus suggesting indirectly the presence of tumor initiating cells.
PLoS ONE 01/2011; 6(7):e21320. · 4.09 Impact Factor
-
Marcello Mancini,
Emilia Vergara,
Giuliana Salvatore,
Adelaide Greco,
Giancarlo Troncone, Andrea Affuso,
Raffaele Liuzzi,
Paolo Salerno,
Maria Scotto di Santolo,
Massimo Santoro,
Arturo Brunetti,
Marco Salvatore
[show abstract]
[hide abstract]
ABSTRACT: The objective of the study was to explore high-frequency ultrasound (HFUS) for noninvasive microimaging of thyroid in living mice. Thyroid examination was performed by HFUS in 10 normal C57BL/6 mice, eight mice treated by propylthiouracil, and 22 Tg-TRK-T1 transgenic mice. The dimension of the gland and the presence of nodules were evaluated. Nodules were classified as malignant (hypoechogenicity, poorly defined margins, internal microcalcification, irregular shapes, and extra glandular extension) or not, and the findings were compared with histological data. Thyroid images were successfully obtained in all the animals analyzed. Normal thyroid reached a volume of 4.92 microl (range 2.11-4.92 microl). Mice with propylthiouracil-induced goiter showed diffuse thyroid enlargement (median volume 6.67 microl, range 4.09-8.82 microl). In 19 of 22 Tg-TRK-T1 mice (86%), HFUS identified a nodular process (the smallest detected nodule had a diameter of 0.46 mm). Eleven nodules were classified as malignant and eight as benign. Compared with histological analysis, HFUS showed a sensitivity of 100% in the detection of thyroid nodules and a specificity of 60% (two of the nodules identified by HFUS were not confirmed at the histology). The specificity and sensitivity of HFUS in predicting the malignancy of the thyroid nodules were 83 and 91%, respectively. Thus, HFUS is an accurate imaging modality that can potentially replace more invasive techniques, and, therefore, it represents a significant advancement in phenotypic assessment of mouse models of thyroid cancer.
Endocrinology 08/2009; 150(10):4810-5. · 4.46 Impact Factor
-
Francesco Errico,
Robert Nisticò,
Giuseppe Palma,
Mauro Federici, Andrea Affuso,
Elisa Brilli,
Enza Topo,
Diego Centonze,
Giorgio Bernardi,
Yuri Bozzi,
Antimo D'Aniello,
Roberto Di Lauro,
Nicola B Mercuri,
Alessandro Usiello
[show abstract]
[hide abstract]
ABSTRACT: In the present study, we demonstrate a direct role for d-aspartate in regulating hippocampal synaptic plasticity. These evidences were obtained using two different experimental strategies which enabled a non-physiological increase of endogenous d-aspartate levels in the mouse hippocampus: a genetic approach based on the targeted deletion of d-aspartate oxidase gene and another based on the oral administration of d-aspartate. Overall, our results indicate that increased d-aspartate content does not affect basal properties of synaptic transmission but enhances long-term potentiation in hippocampal slices from both genetic and pharmacological animal models. Besides electrophysiological data, behavioral analysis suggests that altered levels of d-aspartate in the hippocampus do not perturb basal spatial learning and memory abilities, but may selectively interfere with the dynamic NMDAR-dependent processes underlying cognitive flexibility.
Molecular and Cellular Neuroscience 03/2008; 37(2):236-46. · 3.66 Impact Factor
-
Gaetano Calì,
Mariastella Zannini,
Patrizia Rubini,
Carlo Tacchetti,
Barbara D'Andrea, Andrea Affuso,
Tim Wintermantel,
Oreda Boussadia,
Daniela Terracciano,
Daniel Silberschmidt,
Elena Amendola,
Mario De Felice,
Günther Schütz,
Rolf Kemler,
Roberto Di Lauro,
Lucio Nitsch
[show abstract]
[hide abstract]
ABSTRACT: We have conditionally inactivated the E-cadherin gene in the thyroid follicular cells of mouse embryo to unravel its role in thyroid development. We used the Cre-loxP system in which the Cre-recombinase was expressed under the control of the tissue-specific thyroglobulin promoter that becomes active at embryonic d 15. At postnatal d 7, thyroid follicle lumens in the knockout mice were about 30% smaller with respect to control mice and had an irregular shape. E-cadherin was almost completely absent in thyrocytes, beta-catenin was significantly reduced, whereas no change in gamma-catenin was detected. alpha-Catenin was also reduced on the cell plasma membrane. Despite the dramatic loss of E-cadherin and beta-catenin, cell-cell junctions were not affected, the distribution of tight junction proteins was unaltered, and no increase of thyroglobulin circulating in the blood was observed. In addition, we found that other members of the cadherin family, the R-cadherin and the Ksp-cadherin, were expressed in thyrocytes and that their membrane distribution was not altered in the E-cadherin conditional knockout mouse. Our results indicate that E-cadherin has a role in the development of the thyroid gland and in the expression of beta-catenin, but it is not essential for the maintenance of follicular cell adhesion.
Endocrinology 07/2007; 148(6):2737-46. · 4.46 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Free D-aspartic acid and NMDA are present in the mammalian central nervous system and endocrine glands at significant concentrations, but their physiological role is still matter of debate. The only enzyme known to metabolize in vitro selectively these D-amino acids is D-aspartate oxidase (DDO). To clarify the role in vivo of the enzyme, we generated mice with targeted deletion of Ddo gene by homologous recombination. Mutated animals showed increased amounts of both D-aspartic acid and NMDA in all tissues examined demonstrating a physiological role of DDO in the regulation of their endogenous levels.
Gene 07/2006; 374:50-7. · 2.34 Impact Factor
-
Elena Amendola,
Pasquale De Luca,
Paolo Emidio Macchia,
Daniela Terracciano,
Annamaria Rosica,
Gennaro Chiappetta,
Shioko Kimura,
Ahmed Mansouri, Andrea Affuso,
Claudio Arra,
Vincenzo Macchia,
Roberto Di Lauro,
Mario De Felice
[show abstract]
[hide abstract]
ABSTRACT: Congenital hypothyroidism with thyroid dysgenesis (TD) is a frequent human condition characterized by elevated levels of TSH in response to reduced thyroid hormone levels. Congenital hypothyroidism is a genetically heterogeneous disease. In the majority of cases studied, no causative mutations have been identified and very often the disease does not show a Mendelian transmission. However, in approximately 5% of cases, it can be a consequence of mutations in genes encoding the TSH receptor or the transcription factors TITF1, FOXE1, or PAX8. We report here that in mouse models, the combination of partial deficiencies in the Titf1 and Pax8 genes results in an overt TD phenotype that is absent in either of the singly deficient, heterozygous mice. The disease is characterized by a small thyroid gland, elevated levels of TSH, reduced thyroglobulin biosynthesis, and high occurrence of hemiagenesis. The observed phenotype is strain specific, and the pattern of transmission indicates that at least two other genes, in addition to Titf1 and Pax8, are necessary to generate the condition. These results show that TD can be of multigenic origin in mice and strongly suggest that a similar pathogenic mechanism may be observed in humans.
Endocrinology 01/2006; 146(12):5038-47. · 4.46 Impact Factor
-
Nicoletta Potenza,
Carmine Vecchione,
Antonella Notte,
Assunta De Rienzo,
Annamaria Rosica,
Lisa Bauer, Andrea Affuso,
Mario De Felice,
Tommaso Russo,
Roberta Poulet,
Giuseppe Cifelli,
Gabriella De Vita,
Giuseppe Lembo,
Roberto Di Lauro
[show abstract]
[hide abstract]
ABSTRACT: Ras proteins are highly related GTPases that have key roles in regulating growth, differentiation and tumorigenesis. Gene-targeting experiments have shown that, out of the three mammalian ras genes, only K-ras is essential for normal mouse embryogenesis, and that mice deprived of H-ras and/or N-ras show no major phenotype. We generated mice (HrasKI) in which the K-ras gene had been modified to encode H-Ras protein. HrasKI mice produce undetectable amounts of K-Ras but-in contrast to mice homozygous for a null K-ras allele-they are born at the expected mendelian frequency, indicating that H-Ras can be substituted for K-Ras in embryonic development. However, adult HrasKI mice show dilated cardiomyopathy associated with arterial hypertension. Our results show that K-Ras can be replaced by H-Ras in its essential function in embryogenesis, and indicate that K-Ras has a unique role in cardiovascular homeostasis.
EMBO Reports 06/2005; 6(5):432-7. · 7.36 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The thyroid gland originates from the ventral floor of the foregut as a thickening of the endodermal cell layer. The molecular mechanisms underlying the early steps of thyroid morphogenesis are not known. Gene targeting experiments have contributed to the identification of several transcription factors, in general playing a role in the proliferation, survival, and migration of the thyroid cell precursors. The experiments reported here analyze the expression of the transcription factors Titf1, Hhex, Pax8, and Foxe1 in the thyroid primordium of null mutants of each of them. We found that most of these transcription factors are linked in an integrated regulatory network, each of them controlling the presence of other members of the network. The expression of Foxe1 is regulated in an intriguing fashion as it is strongly dependent on the presence of Pax8 in thyroid precursor cells, while the expression of the same gene in the pharyngeal endoderm surrounding the primordium is dependent on Sonic hedgehog (Shh)-derived signaling. Moreover, by the generation of mouse mutants expressing Foxe1 exclusively in the thyroid primordium, we provide a better understanding of the role of Foxe1 in these cells in order to acquire the competence to migrate into the underlying mesenchyme. In conclusion, we provide the first evidence of gene expression programs, controlled by a hierarchy of transcription factors expressed in the thyroid presumptive gut domain and directing the progression of thyroid morphogenesis.
Developmental Biology 01/2005; 276(2):464-75. · 4.07 Impact Factor
-
Daniela Spano,
Igor Branchi,
Annamaria Rosica,
Maria Teresa Pirro,
Antonio Riccio,
Pratibha Mithbaokar, Andrea Affuso,
Claudio Arra,
Patrizia Campolongo,
Daniela Terracciano,
Vincenzo Macchia,
Juan Bernal,
Enrico Alleva,
Roberto Di Lauro
[show abstract]
[hide abstract]
ABSTRACT: The development and the function of central nervous system depend on thyroid hormones. In humans, the lack of thyroid hormones causes cretinism, a syndrome of severe mental deficiency. It is assumed that thyroid hormones affect the normal development and function of the brain by activating or suppressing target gene expression because several genes expressed in the brain have been shown to be under thyroid hormone control. Among these, the Rhes gene, encoding a small GTP-binding protein, is predominantly expressed in the striatal region of the brain. To clarify the role of Rhes in vivo, we disrupted the Rhes gene by homologous recombination in embryonic stem cells and generated mice homozygous for the Rhes null mutation (Rhes(-/-)). Rhes(-/-) mice were viable but weighed less than wild-type mice. Furthermore, they showed behavioral abnormalities, displaying a gender-dependent increase in anxiety levels and a clear motor coordination deficit but no learning or memory impairment. These results suggest that Rhes disruption affects selected behavioral competencies.
Molecular and Cellular Biology 08/2004; 24(13):5788-96. · 5.53 Impact Factor
-
Gaetano Calì,
Mariastella Zannini,
Patrizia Rubini,
Carlo Tacchetti,
Barbara D'Andrea, Andrea Affuso,
Tim Wintermantel,
Oreda Boussadia,
Daniela Terracciano,
Daniel Silberschmidt,
Elena Amendola,
Mario De Felice
