Publications by authors named "Filippo Biamonte"

21 Publications

  • Page 1 of 1

Immunohistochemical Analysis of DNA Repair- and Drug-Efflux-Associated Molecules in Tumor and Peritumor Areas of Glioblastoma.

Int J Mol Sci 2021 Feb 5;22(4). Epub 2021 Feb 5.

Dipartimento di Scienze della Vita e Sanità Pubblica, Sezione di Istologia ed Embriologia, Università Cattolica del Sacro Cuore-Fondazione Policlinico Universitario "Agostino Gemelli", IRCCS, 00168 Rome, Italy.

Glioblastoma (GBM), the most commonly occurring primary tumor arising within the central nervous system, is characterized by high invasiveness and poor prognosis. In spite of the improvement in surgical techniques, along with the administration of chemo- and radiation therapy and the incessant investigation in search of prospective therapeutic targets, the local recurrence that frequently occurs within the peritumoral brain tissue makes GBM the most malignant and terminal type of astrocytoma. In the current study, we investigated both GBM and peritumoral tissues obtained from 55 hospitalized patients and the expression of three molecules involved in the onset of resistance/unresponsiveness to chemotherapy: O6-methylguanine methyltransferase (MGMT), breast cancer resistance protein (BCRP1), and A2B5. We propose that the expression of these molecules in the peritumoral tissue might be crucial to promoting the development of early tumorigenic events in the tissue surrounding GBM as well as responsible for the recurrence originating in this apparently normal area and, accordingly, for the resistance to treatment with the standard chemotherapeutic regimen. Notably, the inverse correlation found between MGMT expression in peritumoral tissue and patients' survival suggests a prognostic role for this protein.
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http://dx.doi.org/10.3390/ijms22041620DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7914796PMC
February 2021

Nerve Growth Factor (NGF) modulates in vitro induced myofibroblasts by highlighting a differential protein signature.

Sci Rep 2021 Jan 18;11(1):1672. Epub 2021 Jan 18.

Research and Development Laboratory for Biochemical, Molecular and Cellular Applications in Ophthalmological Sciences, IRCCS-Fondazione Bietti, Rome, Italy.

We previously described the profibrogenic effect of NGF on conjunctival Fibroblasts (FBs) and its ability to trigger apoptosis in TGFβ1-induced myofibroblasts (myoFBs). Herein, cell apoptosis/signalling, cytokines' signature in conditioned media and inflammatory as well as angiogenic pathway were investigated. Experimental myoFBs were exposed to NGF (0.1-100 ng/mL), at defined time-point for confocal and biomolecular analysis. Cells were analysed for apoptotic and cell signalling activation in cell extracts and for some inflammatory and proinflammatory/angiogenic factors' activations. NGF triggered cJun overexpression and phospho-p65-NFkB nuclear translocation. A decreased Bcl2:Bax ratio and a significant expression of smad7 were confirmed in early AnnexinV-positive myoFBs. A specific protein signature characterised the conditioned media: a dose dependent decrease occurred for IL8, IL6 while a selective increase was observed for VEGF and cyr61 (protein/mRNA). TIMP1 levels were unaffected. Herein, NGF modulation of smad7, the specific IL8 and IL6 as well as VEGF and cyr61 modulation deserve more attention as opening to alternative approaches to counteract fibrosis.
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http://dx.doi.org/10.1038/s41598-021-81040-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7814037PMC
January 2021

Sam68 splicing regulation contributes to motor unit establishment in the postnatal skeletal muscle.

Life Sci Alliance 2020 10 4;3(10). Epub 2020 Aug 4.

Department of Movement, Human and Health Sciences, University of Rome "Foro Italico," Rome, Italy

RNA-binding proteins orchestrate the composite life of RNA molecules and impact most physiological processes, thus underlying complex phenotypes. The RNA-binding protein Sam68 regulates differentiation processes by modulating splicing, polyadenylation, and stability of select transcripts. Herein, we found that mice display altered regulation of alternative splicing in the spinal cord of key target genes involved in synaptic functions. Analysis of the motor units revealed that Sam68 ablation impairs the establishment of neuromuscular junctions and causes progressive loss of motor neurons in the spinal cord. Importantly, alterations of neuromuscular junction morphology and properties in mice correlate with defects in muscle and motor unit integrity. muscles display defects in postnatal development, with manifest signs of atrophy. Furthermore, fast-twitch muscles in mice show structural features typical of slow-twitch muscles, suggesting alterations in the metabolic and functional properties of myofibers. Collectively, our data identify a key role for Sam68 in muscle development and suggest that proper establishment of motor units requires timely expression of synaptic splice variants.
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http://dx.doi.org/10.26508/lsa.201900637DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7409371PMC
October 2020

Daily physical activity in patients on chronic haemodialysis and its relation with fatigue and depressive symptoms.

Int Urol Nephrol 2020 Oct 28;52(10):1959-1967. Epub 2020 Jul 28.

Università Cattolica del Sacro Cuore, Rome, Italy.

Objective: Fatigue and depressed mood are considered main impediments to physical activity in haemodialysis (HD) patients. A better understanding of their interrelationships is crucial to develop effective therapies. Moreover, measurement of daily physical activity (DPA) in HD patients is tricky, as it is usually assessed by subjective self-report questionnaires. Therefore, we aimed to objectively measure sponteanous DPA with motion sensors and to explore its relation with fatigue and depressive symptoms.

Methods: DPA was assessed for seven consecutive days in 37 HD patients based on their daily step count measured with the SenseWear™ Armband. The Fatigue Severity Scale (FSS) and Beck Depression Inventory-II (BDI-II) were administered to evaluate fatigue and depressed mood.

Results: Median DPA was 2424 steps/day, (IQR:892-4545). In 81% of subjects, DPA felt within a sedentary lifestyle classification, as they made < 5.000 steps/day. DPA did not correlate with fatigue (r = 0.04, p = 0.832), and did not significantly differ between patients categorized as clinically fatigued (n = 23, FSS ≥ 4) or not (n = 14, FSS < 4) (p = 0.654, d = 0.20). Although low-depressed subjects (n = 19, BDI-II ≤ 13) made on average 1.7 times more steps/day than high-depressed subjects (n = 18, BDI-II > 13) (p = 0.111, d = 0.60), depressive mood did also not correlate significantly with DPA (r = - 0.23, p = 0.175).

Conclusion: Objective assessment of DPA with motion sensors is feasible in HD patients and allows identifying a sedentary lifestyle. Our results suggest that spontanous DPA is determined by age rather than by fatigue or mood.
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http://dx.doi.org/10.1007/s11255-020-02578-9DOI Listing
October 2020

Ambra1 Shapes Hippocampal Inhibition/Excitation Balance: Role in Neurodevelopmental Disorders.

Mol Neurobiol 2018 Oct 27;55(10):7921-7940. Epub 2018 Feb 27.

Department of Experimental Neurosciences, IRCCS Santa Lucia Foundation, 00143, Rome, Italy.

Imbalances between excitatory and inhibitory synaptic transmission cause brain network dysfunction and are central to the pathogenesis of neurodevelopmental disorders. Parvalbumin interneurons are highly implicated in this imbalance. Here, we probed the social behavior and hippocampal function of mice carrying a haploinsufficiency for Ambra1, a pro-autophagic gene crucial for brain development. We show that heterozygous Ambra1 mice (Ambra) are characterized by loss of hippocampal parvalbumin interneurons, decreases in the inhibition/excitation ratio, and altered social behaviors that are solely restricted to the female gender. Loss of parvalbumin interneurons in Ambra1 females is further linked to reductions of the inhibitory drive onto principal neurons and alterations in network oscillatory activity, CA1 synaptic plasticity, and pyramidal neuron spine density. Parvalbumin interneuron loss is underlined by increased apoptosis during the embryonic development of progenitor neurons in the medial ganglionic eminence. Together, these findings identify an Ambra1-dependent mechanism that drives inhibition/excitation imbalance in the hippocampus, contributing to abnormal brain activity reminiscent of neurodevelopmental disorders.
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http://dx.doi.org/10.1007/s12035-018-0911-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6132777PMC
October 2018

Functional alterations of the dopaminergic and glutamatergic systems in spontaneous α-synuclein overexpressing rats.

Exp Neurol 2017 Jan 20;287(Pt 1):21-33. Epub 2016 Oct 20.

Fondazione Santa Lucia IRCCS, Via del Fosso di Fiorano 64, 00143 Rome, Italy; University of Tor Vergata, Department of Systems Medicine, Via Montpellier 1, 00133 Rome, Italy. Electronic address:

The presence of α-synuclein (α-syn) in Lewy bodies and Lewy neurites is an important characteristic of the neurodegenerative processes of substantia nigra pars compacta (SNpc) dopaminergic (DAergic) neurons in Parkinson's disease (PD) and other synucleinopathies. Here we report that Berlin-Druckrey rats carrying a spontaneous mutation in the 3' untranslated region of α-syn mRNA (m/m rats) display a marked accumulation of α-syn in the mesencephalic area, striatum and frontal cortex, accompanied to severe dysfunctions in the dorsolateral striatum. Despite a small reduction in the number of SNpc and ventral tegmental area DAergic cells, the surviving dopaminergic neurons of the m/m rats do not show clear-cut alterations of the spontaneous and evoked firing activity, DA responses and somatic amphetamine-induced firing inhibition. Interestingly, mutant DAergic neurons display diminished whole-cell Ih conductance and a reduced frequency of spontaneous excitatory synaptic currents. By contrast, m/m rats show a severe impairment of DA and glutamate release in the dorsolateral striatum, as revealed by amperometric measure of DA currents and by electrophysiological recordings of glutamatergic synaptic events in striatal medium spiny neurons. These functional impairments are paralleled by a decreased expression of the DA transporter and VGluT1 proteins in the same area. Thus, together with α-syn overload in the mesencephalic region, striatum and frontal cortex, the main functional alterations occur in the DAergic and glutamatergic terminals in the dorsal striatum of the m/m rats.
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http://dx.doi.org/10.1016/j.expneurol.2016.10.009DOI Listing
January 2017

Analysis of angiogenesis related factors in glioblastoma, peritumoral tissue and their derived cancer stem cells.

Oncotarget 2016 Nov;7(48):78541-78556

Institute of Histology and Embryology, "A. Gemelli" Faculty of Medicine, Catholic University of the Sacred Heart, Rome, Italy.

The formation of new blood vessels represents a crucial event under both physiological and pathological circumstances. In this study, we evaluated by immunohistochemistry, and/or Western blotting and/or quantitative real time-PCR the expression of HIF1α, HIF2α, VEGF, VEGFR1 and VEGFR2 in surgical glioblastoma multiforme (GBM) and peritumoral tissue samples obtained from 50 patients as well as in cancer stem cells (CSCs) isolated from GBM (GCSCs) and peritumoral tissue (PCSCs) of 5 patients. We also investigated the contribution of both GCSCs and PCSCs on the behavior of endothelial cells (ECs) in vitro. Immunohistochemistry demonstrated the expression of angiogenesis markers in both GBM and peritumoral tissue. In addition, in vitro tube formation assay indicated that both GCSCs and PCSCs stimulate EC proliferation as well as tube-like vessel formation. An increased migration aptitude was mainly observed when ECs were cultured in the presence of GCSCs rather than in the presence of PCSCs. These findings suggest that relevant neoangiogenetic events may occur in GBM. In particular, VEGF/VEGFR co-expression in PCSCs leads to hypothesize the involvement of an autocrine signaling. Moreover, our results suggest that both GCSCs and PCSCs own the skill of activating the "angiogenic switch" and the capability of modulating EC behavior, indicating that both cell types are either responsive to angiogenic stimuli or able to trigger angiogenic response. Together with our previous findings, this study adds a further piece to the challenging puzzle of the characterization of peritumoral tissue and of the definition of its real role in GBM pathophysiology.
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http://dx.doi.org/10.18632/oncotarget.12398DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5346658PMC
November 2016

Progenitor/Stem Cell Markers in Brain Adjacent to Glioblastoma: GD3 Ganglioside and NG2 Proteoglycan Expression.

J Neuropathol Exp Neurol 2016 Feb;75(2):134-47

Characterization of tissue surrounding glioblastoma (GBM) is a focus for translational research because tumor recurrence invariably occurs in this area. We investigated the expression of the progenitor/stem cell markers GD3 ganglioside and NG2 proteoglycan in GBM, peritumor tissue (brain adjacent to tumor, BAT) and cancer stem-like cells (CSCs) isolated from GBM (GCSCs) and BAT (PCSCs). GD3 and NG2 immunohistochemistry was performed in paired GBM and BAT specimens from 40 patients. Double-immunofluorescence was carried out to characterize NG2-positive cells of vessel walls. GD3 and NG2 expression was investigated in GCSCs and PCSCs whose tumorigenicity was also evaluated in Scid/bg mice. GD3 and NG2 expression was higher in tumor tissue than in BAT. NG2 decreased as the distance from tumor margin increased, regardless of the tumor cell presence, whereas GD3 correlated with neoplastic infiltration. In BAT, NG2 was coexpressed with a-smooth muscle actin (a-SMA) in pericytes and with nestin in the endothelium. Higher levels of NG2 mRNA and protein were found in GCSCs while GD3 synthase was expressed at similar levels in the 2 CSC populations. PCSCs had lower tumorigenicity than GCSCs. These data suggest the possible involvement of GD3 and NG2 in pre/pro-tumorigenic events occurring in the complex microenvironment of the tissue surrounding GBM.
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http://dx.doi.org/10.1093/jnen/nlv012DOI Listing
February 2016

Estrogen administration modulates hippocampal GABAergic subpopulations in the hippocampus of trimethyltin-treated rats.

Front Cell Neurosci 2015 5;9:433. Epub 2015 Nov 5.

Institute of Anatomy and Cell Biology, Università Cattolica del Sacro Cuore Rome, Italy.

Given the well-documented involvement of estrogens in the modulation of hippocampal functions in both physiological and pathological conditions, the present study investigates the effects of 17-beta estradiol (E2) administration in the rat model of hippocampal neurodegeneration induced by trimethyltin (TMT) administration (8 mg/kg), characterized by loss of pyramidal neurons in CA1, CA3/hilus hippocampal subfields, associated with astroglial and microglial activation, seizures and cognitive impairment. After TMT/saline treatment, ovariectomized animals received two doses of E2 (0.2 mg/kg intra-peritoneal) or vehicle, and were sacrificed 48 h or 7 days after TMT-treatment. Our results indicate that in TMT-treated animals E2 administration induces the early (48 h) upregulation of genes involved in neuroprotection and synaptogenesis, namely Bcl2, trkB, cadherin 2 and cyclin-dependent-kinase-5. Increased expression levels of glutamic acid decarboxylase (gad) 67, neuropeptide Y (Npy), parvalbumin, Pgc-1α and Sirtuin 1 genes, the latter involved in parvalbumin (PV) synthesis, were also evident. Unbiased stereology performed on rats sacrificed 7 days after TMT treatment showed that although E2 does not significantly influence the extent of TMT-induced neuronal death, significantly enhances the TMT-induced modulation of GABAergic interneuron population size in selected hippocampal subfields. In particular, E2 administration causes, in TMT-treated rats, a significant increase in the number of GAD67-expressing interneurons in CA1 stratum oriens, CA3 pyramidal layer, hilus and dentate gyrus, accompanied by a parallel increase in NPY-expressing cells, essentially in the same regions, and of PV-positive cells in CA1 pyramidal layer. The present results add information concerning the role of in vivo E2 administration on mechanisms involved in cellular plasticity in the adult brain.
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http://dx.doi.org/10.3389/fncel.2015.00433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4633568PMC
November 2015

Toll-Like Receptors and Tissue Remodeling: The Pro/Cons Recent Findings.

J Cell Physiol 2016 Mar 1;231(3):531-44. Epub 2015 Sep 1.

Department of Ophthalmology, University Campus Bio-Medico, Rome, Italy.

The Toll-like Receptor (TLR) family ensures prompt response towards pathogens, protecting the host against infections, and guarantees a realistic balance between protective and detrimental activities. Multiple regulating mechanisms characterize TLR activity that is not limited to innate and adaptive antimicrobial immune responses, as observed in the inflammatory (either infective, allergic, or autoimmune) responses associated with tissue remodeling. Following the insult and the arise of inflammatory response, tissue remodeling takes place and might develop in fibrosis, depending on microenvironment as a result of imbalanced fibroblasts (FBs) and myofibroblasts (myoFBs) activation/survival. The process is driven by an epithelial-fibroblast-immune cell cross-talk. While the main FB function is the matrix metabolism for tissue homeostasis or repair, the myoFB differentiation represents a crucial step in attempting repair of injury. FBs/myoFBs provide more than structural support at site of injury, synthesizing and/or reacting to different cytokines, growth factors, neuromediators and soluble/lipid mediators. TLR-bearing FBs/myoFBs might contribute at the innate immune level, providing a second line of protection/defense as well as being a target/effector cell of tissue remodeling. TLRs might also interfere with acute inflammation as well as with established fibrosis, triggering structural/functional changes in agreement with the genetic background, the site of lesion, the entity of associated infection, the poor blood circulation or the pharmacological treatments, all together strictly influencing tissue repair/remodeling process. This review will focus on the recent findings on TLRs at launch and long-lasting tissue remodeling process, that strongly suggest TLRs as optional targets for future therapies.
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http://dx.doi.org/10.1002/jcp.25124DOI Listing
March 2016

Quantitative neuroanatomy of all Purkinje cells with light sheet microscopy and high-throughput image analysis.

Front Neuroanat 2015 27;9:68. Epub 2015 May 27.

National Institute of Optics, National Research Council Sesto Fiorentino, Italy ; European Laboratory for Non-Linear Spectroscopy Sesto Fiorentino, Italy ; Department of Physics and Astronomy, University of Florence Sesto Fiorentino, Italy ; International Center for Computational Neurophotonics Sesto Fiorentino, Italy.

Characterizing the cytoarchitecture of mammalian central nervous system on a brain-wide scale is becoming a compelling need in neuroscience. For example, realistic modeling of brain activity requires the definition of quantitative features of large neuronal populations in the whole brain. Quantitative anatomical maps will also be crucial to classify the cytoarchtitectonic abnormalities associated with neuronal pathologies in a high reproducible and reliable manner. In this paper, we apply recent advances in optical microscopy and image analysis to characterize the spatial distribution of Purkinje cells (PCs) across the whole cerebellum. Light sheet microscopy was used to image with micron-scale resolution a fixed and cleared cerebellum of an L7-GFP transgenic mouse, in which all PCs are fluorescently labeled. A fast and scalable algorithm for fully automated cell identification was applied on the image to extract the position of all the fluorescent PCs. This vectorized representation of the cell population allows a thorough characterization of the complex three-dimensional distribution of the neurons, highlighting the presence of gaps inside the lamellar organization of PCs, whose density is believed to play a significant role in autism spectrum disorders. Furthermore, clustering analysis of the localized somata permits dividing the whole cerebellum in groups of PCs with high spatial correlation, suggesting new possibilities of anatomical partition. The quantitative approach presented here can be extended to study the distribution of different types of cell in many brain regions and across the whole encephalon, providing a robust base for building realistic computational models of the brain, and for unbiased morphological tissue screening in presence of pathologies and/or drug treatments.
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http://dx.doi.org/10.3389/fnana.2015.00068DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4445386PMC
June 2015

Associations among exposure to methylmercury, reduced Reelin expression, and gender in the cerebellum of developing mice.

Neurotoxicology 2014 Dec 14;45:67-80. Epub 2014 Oct 14.

Laboratory of Developmental Neuroscience and Neural Plasticity, University Campus Biomedico, Via A. del Portillo 21, 00198 Rome, Italy. Electronic address:

Genetic risk factors acting during pregnancy or early after birth have been proposed to account for the exponential increase of autism diagnoses in the past 20 years. In particular, a potential link with exposure to environmental mercury has been suggested. Male sex constitutes a second risk factor for autism. A third potential genetic risk factor is decreased Reelin expression. Male heterozygous reeler (rl(+/-)) mice show an autism-like phenotype, including Purkinje cells (PCs) loss and behavioral rigidity. We evaluated the complex interactions between 3 risk factors, i.e. genetic status, sex, and exposure to methylmercury (MeHg), in rl(+/-) mice. Mice were exposed to MeHg during the prenatal and early postnatal period, either at a subtoxic dose (2 ppm in Dams' drinking water), or at a toxic dose (6 ppm Dams' drinking water), based on observations in other rodent species and mice strains. We show that: (a) 2 ppm MeHg does not cause PCs loss in the different animal groups, and does not enhance PCs loss in rl(+/-) males; consistent with a lack of overt neurotoxicity, 2 ppm MeHg per se does not cause behavioral alterations (separation-induced ultrasonic calls in newborns, or sociability and social preference in adults); (b) in stark contrast, 6 ppm MeHg causes a dramatic reduction of PCs number in all groups, irrespective of genotype and sex. Cytochrome C release from mitochondria of PCs is enhanced in 6 ppm MeHg-exposed groups, with a concomitant increase of μ-calpain active subunit. At the behavioral level, 6 ppm MeHg exposure strongly increases ultrasonic vocalizations in all animal groups. Notably, 6 ppm MeHg significantly decreases sociability in rl(+/-) male mice, while the 2 ppm group does not show such as decrease. At a subtoxic dose, MeHg does not enhance the autism-like phenotype of male rl(+/-) mice. At the higher MeHg dose, the scenario is more complex, with some "autism-like" features (loss of sociability, preference for sameness) being evidently affected only in rl(+/-) males, while other neuropathological and behavioral parameters being altered in all groups, independently from genotype and sex. Mitochondrial abnormalities appear to play a crucial role in the observed effects.
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http://dx.doi.org/10.1016/j.neuro.2014.09.006DOI Listing
December 2014

Isolation of cancer stem cells from three human glioblastoma cell lines: characterization of two selected clones.

PLoS One 2014 14;9(8):e105166. Epub 2014 Aug 14.

Institute of Histology and Embryology, Medical School, Catholic University of the Sacred Heart, Rome, Italy.

Cancer stem cells (CSC) were isolated via a non-adherent neurosphere assay from three glioma cell lines: LI, U87, and U373. Using a clonal assay, two clones (D2 and F11) were selected from spheres derived from LI cells and were characterized for the: expression of stem cell markers (CD133, Nestin, Musashi-1 and Sox2); proliferation; differentiation capability (determined by the expression of GalC, βIII-Tubulin and GFAP); Ca(2+) signaling and tumorigenicity in nude mice. Both D2 and F11 clones expressed higher levels of all stem cell markers with respect to the parental cell line. Clones grew more slowly than LI cells with a two-fold increase in duplication time. Markers of differentiation (βIII-Tubulin and GFAP) were expressed at high levels in both LI cells and in neurospheres. The expression of Nestin, Sox2, and βIII-Tubulin was down-regulated in D2 and F11 when cultured in serum-containing medium, whereas Musashi-1 was increased. In this condition, duplication time of D2 and F11 increased without reaching that of LI cells. D2, F11 and parental cells did not express voltage-dependent Ca(2+)-channels but they exhibited increased intracellular Ca(2+) levels in response to ATP. These Ca(2+) signals were larger in LI cells and in spheres cultured in serum-containing medium, while they were smaller in serum-free medium. The ATP treatment did not affect cell proliferation. Both D2 and F11 induced the appearance of tumors when ortotopically injected in athymic nude mice at a density 50-fold lower than that of LI cells. All these data indicate that both clones have characteristics of CSC and share the same stemness properties. The findings regarding the expression of differentiation markers and Ca(2+)-channels show that both clones are unable to reach the terminal differentiation. Both D2 and F11 might represent a good model to improve the knowledge on CSC in glioblastoma and to identify new therapeutic approaches.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0105166PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4133365PMC
March 2016

Characterization of NGF, trkA (NGFR) , and p75 (NTR) in Retina of Mice Lacking Reelin Glycoprotein.

Int J Cell Biol 2014 30;2014:725928. Epub 2014 Jan 30.

IRCCS-G.B. Bietti Foundation, Rome, Italy ; Laboratory of Ophthalmology, IRCCS-G.B. Bietti Foundation, Via Alvaro del Portillo 21, 00128 Rome, Italy.

Both Reelin and Nerve Growth Factor (NGF) exert crucial roles in retinal development. Retinogenesis is severely impaired in E-reeler mice, a model of Reelin deficiency showing specific Green Fluorescent Protein expression in Rod Bipolar Cells (RBCs). Since no data are available on Reelin and NGF cross-talk, NGF and trkA(NGFR)/ p75(NTR) expression was investigated in retinas from E-reeler versus control mice, by confocal microscopy, Western blotting, and real time PCR analysis. A scattered increase of NGF protein was observed in the Ganglion Cell Layer and more pronounced in the Inner Nuclear Layer (INL). A selective increase of p75(NTR) was detected in most of RBCs and in other cell subtypes of INL. On the contrary, a slight trend towards a decrease was detected for trkA(NGFR), albeit not significant. Confocal data were validated by Western blot and real time PCR. Finally, the decreased trkA(NGFR)/ p75(NTR) ratio, representative of p75(NTR) increase, significantly correlated with E-reeler versus E-control. These data indicate that NGF-trkA(NGFR)/ p75(NTR) is affected in E-reeler retina and that p75(NTR) might represent the main NGF receptor involved in the process. This first NGF-trkA(NGFR)/ p75(NTR) characterization suggests that E-reeler might be suitable for exploring Reelin-NGF cross-talk, representing an additional information source in those pathologies characterized by retinal degeneration.
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http://dx.doi.org/10.1155/2014/725928DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3928862PMC
March 2014

Knock down of caveolin-1 affects morphological and functional hallmarks of human endothelial cells.

J Cell Biochem 2013 Aug;114(8):1843-51

Department of Anatomy, Histology, Forensic Medicine and Orthopedics, Section of Histology and Medical Embryology, Sapienza University of Rome, Rome, Italy.

Caveolin-1 (CAV1) is the principal structural component of caveolae which functions as scaffolding protein for the integration of a variety of signaling pathways. In this study, we investigated the involvement of CAV1 in endothelial cell (EC) functions and show that siRNA-induced CAV1 silencing in the human EC line EA.hy926 induces distinctive morphological changes, such as a marked increase in cell size and formation of stress fibers. Design-based stereology was employed in this work to make unbiased quantification of morphometric properties such as volume, length, and surface of CAV1 silenced versus control cells. In addition, we showed that downregulation of CAV1 affects cell cycle progression at G1/S phase transition most likely by perturbation of AKT signaling. With the aim to assess the contribution of CAV1 to typical biological processes of EC, we report here that CAV1 targeting affects cell migration and matrix metalloproteinases (MMPs) activity, and reduces angiogenesis in response to VEGF, in vitro. Taken together our data suggest that the proper expression of CAV1 is important not only for maintaining the appropriate morphology and size of ECs but it might represent a prospective molecular target for studying key biological mechanisms such as senescence and tumorigenesis.
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http://dx.doi.org/10.1002/jcb.24526DOI Listing
August 2013

The neuroprotective and neurogenic effects of neuropeptide Y administration in an animal model of hippocampal neurodegeneration and temporal lobe epilepsy induced by trimethyltin.

J Neurochem 2012 Jul 21;122(2):415-26. Epub 2012 May 21.

Institute of Anatomy and Cell Biology, Università Cattolica del Sacro Cuore, Rome, Italy.

The effects of intracerebroventricular administration of neuropeptide Y (NPY), which is believed to play an important role in neuroprotection against excitotoxicity and in the modulation of adult neurogenesis, were evaluated in an animal model of hippocampal neurodegeneration and temporal lobe epilepsy represented by trimethyltin (TMT) intoxication. A single TMT injection (8 mg/kg) causes, in the rat brain, massive neuronal death, selectively involving pyramidal neurons, accompanied by glial activation and enhanced hippocampal neurogenesis. Our data indicate that intracerebroventricular administration of exogenous NPY (at the dose of 2 μg/2 μL, 4 days after TMT-administration), in adult rats, exerts a protective role in regard to TMT-induced hippocampal damage and a proliferative effect on the hippocampal neurogenic niche through the up-regulation of Bcl-2, Bcl2l1, Bdnf, Sox-2, NeuroD1, Noggin and Doublecortin genes, contributing to delineate more clearly the role of NPY in in vivo neurodegenerative processes.
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http://dx.doi.org/10.1111/j.1471-4159.2012.07770.xDOI Listing
July 2012

Perseverative responding and neuroanatomical alterations in adult heterozygous reeler mice are mitigated by neonatal estrogen administration.

Psychoneuroendocrinology 2010 Oct 7;35(9):1374-87. Epub 2010 May 7.

Department of Cell Biology & Neuroscience, Istituto Superiore di Sanità, Roma, Italy.

According to the "extreme-male brain" theory, elevated fetal testosterone levels may partly explain the skewed sex ratio found in Autism Spectrum Disorders (ASD). Correcting this testosterone imbalance by increasing estrogen levels may mitigate the abnormal phenotype. Accordingly, while control heterozygous reeler (rl/+) male mice - a putative model of neuroanatomical and behavioral endophenotypes in ASD - show a decreased number of Purkinje cells (PC) compared to control wild-type (+/+) littermates, neonatal estradiol administration has been shown to correct this deficit in the short-term (i.e. on postnatal day 15). Here, we further investigated the neuroanatomical and behavioral abnormalities of rl/+ male mice and the potential compensatory effects of neonatal treatment with estradiol. In a longitudinal study, we observed that: i) infant rl/+ mice showed reduced motivation for social stimuli; ii) adult rl/+ male mice showed reduced cognitive flexibility; iii) the number of amygdalar parvalbumin-positive GABAergic interneurons were remarkably reduced in rl/+ mice; iv) neonatal estradiol administration into the cisterna magna reverted the abnormal profile both at the behavioral and at the neuroanatomical level in the amygdala but did not compensate for the cerebellar abnormalities in adulthood. This study supports the view that an increased excitation-to-inhibition ratio in the cerebellum and in the amygdala during a critical window of development could be crucial to the social and cognitive phenotype of male rl/+ mice, and that acute estradiol treatment during this critical window may mitigate symptoms' severity.
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http://dx.doi.org/10.1016/j.psyneuen.2010.03.012DOI Listing
October 2010

Interactions between neuroactive steroids and reelin haploinsufficiency in Purkinje cell survival.

Neurobiol Dis 2009 Oct 10;36(1):103-15. Epub 2009 Jul 10.

Laboratory of Developmental Neuroscience and Neural Plasticity, University Campus Bio-Medico, Via Alvaro del Portillo 21, 00128 Rome, Italy.

We determined total Purkinje cell (PC) numbers in cerebella of wild-type (+/+) and heterozygous (rl/+) reeler mice of either sex during early postnatal development; in parallel, we quantified levels of neuroactive steroids in the cerebellum with mass spectrometry. We also quantified reelin mRNA and protein expression with RT-PCR and Western blotting. PC numbers are selectively reduced at postnatal day 15 (P15) in rl/+ males in comparison to +/+ males, +/+ females, and rl/+ females. Administration of 17beta-estradiol (17beta-E) into the cisterna magna at P5 increases PC numbers in rl/+ males, but not in the other groups; conversely, estrogen antagonists 4-OH-tamoxifen or ICI 182,780 reduce PC numbers in +/+ and rl/+ females, but have no effect in males. Testosterone (T) levels at P5 are much higher in males than in females, reflecting the perinatal testosterone surge in males. In addition, rl/+ male cerebella at P5 show a peculiar hormonal profile in comparison with the other groups, consisting of increased levels of T and 17beta-E, and decreased levels of dihydrotestosterone. RT-PCR analysis indicated that heterozygosity leads to a 50% reduction of reelin mRNA in the cerebellum in both sexes, as expected, and that 17beta-E upregulates reelin mRNA, particularly in rl/+ males; reelin mRNA upregulation is associated with an increase of all major reelin isoforms. These effects may represent a novel model of how reelin deficiency interacts with variable perinatal levels of neuroactive steroids, leading to gender-dependent differences in genetic vulnerability.
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http://dx.doi.org/10.1016/j.nbd.2009.07.001DOI Listing
October 2009

Reelin haploinsufficiency reduces the density of PV+ neurons in circumscribed regions of the striatum and selectively alters striatal-based behaviors.

Psychopharmacology (Berl) 2009 Jun 10;204(3):511-21. Epub 2009 Mar 10.

Department of Experimental Neurology, IRCCS Santa Lucia Foundation, Rome, Italy.

Rationale: Reelin, a large extracellular matrix glycoprotein, is down-regulated in the brain of schizophrenic patients and of heterozygous reeler mice (rl/+). The behavioral phenotype of rl/- mice, however, matches only partially the schizophrenia hallmarks.

Objectives: We recently reported (Marrone et al., Eur J Neurosci 24:20062-22070, 2006) that homozygous reeler mutants (rl/rl) exhibit reduced density of parvalbumin-positive (PV+) GABAergic interneurons in anatomically circumscribed regions of the neostriatum. Assuming that in rl/+ mice may also show regional reduction of striatal GABAergic interneurons, behavioral impairments should selectively emerge in tasks depending on specifically altered striatal circuits.

Materials And Methods: We mapped the density of striatal PV+ interneurons in rl/+ and wild-type (+/+) mice and measured their performance in tasks depending on distinct striatal subregions.

Results: Our findings show that, contrary to what would be expected on the basis of gene dosage criteria, the striatal regions in which rl/rl mice exhibited decreased density of PV+ interneurons were either unaltered (rostral striatum) or equally altered (dorsomedial and ventromedial intermediate striatum, caudal striatum) in rl/+ mice. The anatomical findings were paralleled by behavioral deficits in fear extinction and latent inhibition, respectively, requiring the dorsomedial and ventromedial striatal regions. Conversely, active avoidance performance, which requires the dorsolateral region, was unaffected.

Conclusions: Reelin haploinsufficiency alters the density of PV+ neurons in circumscribed regions of the striatum and selectively disrupts behaviors sensitive to dysfunction of these targeted regions. This aspect should be considered when designing experiments aimed at evaluating the impact of reelin haploinsufficiency in schizophrenia-associated cognitive disturbances in rl/+ mutants.
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http://dx.doi.org/10.1007/s00213-009-1483-xDOI Listing
June 2009

Combined treatment with atorvastatin and minocycline suppresses severity of EAE.

Exp Neurol 2008 May 14;211(1):214-26. Epub 2008 Feb 14.

Department of Pharmacology, University of Florence, Viale Pieraccini n. 6, Florence, Italy.

Multiple sclerosis (MS) is the most common inflammatory demyelinating disorder of the central nervous system (CNS). An approach to improve MS treatment is to identify a rational combination of new medications or existing therapies that impact different aspects of the disease process. Statins are effective in the treatment of MS animal models and are promising candidates for future treatment. Minocycline ameliorates clinical severity of experimental autoimmune encephalomyelitis (EAE) and exhibits several anti-inflammatory and neuroprotective activities. In this study, we tested whether the combination of these two drugs could produce beneficial effects in EAE mice immunized with myelin oligodendrocyte protein (MOG). Our findings show that combined treatment, compared to using the medications alone, resulted in a significant reduction in disease severity, in both the acute and chronic phases of the disease, along with attenuation of inflammation, demyelination and axonal loss. Stereological analysis revealed that the combined treatment significantly guarded against neuroinflammation and neurodegeneration. Moreover, a significant suppression of anti-MOG antibody production in animals treated with the two medications was found. In conclusion, our findings prove that this combination of drugs is neuroprotective and suppresses the severity of EAE. Furthermore, this pharmacological approach appears to be promising as a future therapeutic strategy to control MS.
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http://dx.doi.org/10.1016/j.expneurol.2008.01.022DOI Listing
May 2008

Altered cortico-striatal synaptic plasticity and related behavioural impairments in reeler mice.

Eur J Neurosci 2006 Oct;24(7):2061-70

Department of Experimental Neurology, IRCCS Santa Lucia Foundation, Rome, Italy.

Reelin-deficient mice have been used to investigate the role of this extracellular protein in cortico-striatal plasticity and striatum-related behaviours. Here we show that a repetitive electrical stimulation of the cortico-striatal pathway elicited long-term potentiation (LTP) in homozygous reeler (rl/rl) mice, while causing long-term depression in their wild-type (+/+) littermates. The N-methyl-D-aspartic acid (NMDA) receptor antagonist D-(-)-2 amino-5-phosphonopentanoic acid prevented the induction of LTP in (rl/rl) mice, thus confirming that this form of synaptic plasticity was NMDA receptor-dependent. Interestingly, in the presence of tiagabine, a blocker of gamma-aminobutyric acid (GABA) re-uptake system, the probability that (rl/rl) mice showed LTP decreased significantly, thus suggesting an impaired GABAergic transmission in reeler mutants. Consistent with this view, a decreased density of parvalbumin-positive GABAergic striatal interneurons was found in (rl/rl) mice in comparison to (+/+) mice. Finally, compatible with their abnormal striatal function (rl/rl) mice exhibited procedural learning deficits. Our data, showing alterations in cortico-striatal plasticity largely depending on a depressed GABAergic tone, delineate a mechanism whereby the lack of reelin may affect cognitive functions.
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http://dx.doi.org/10.1111/j.1460-9568.2006.05083.xDOI Listing
October 2006