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M Corbo

Publications and source records attributed to M Corbo.

At least 19 recordsLinked to original sources
PubMed2005

Stochastic modelling of apoptosis kinetics.

Robust quantitative estimation of average whole cell mitochondrial dysfunction is a useful tool for assessing sensitivity to apoptotic stimuli induced either by novel agents, or following manipulation of apoptotic threshold by pharmacological or functional genomics approaches. We have mathematically modelled the kinetics of whole cell mitochondrial membrane potential depolarisation within a population of cells as a Bernouli transition. An exponential distribution enables the median latency preceding mitochondrial membrane potential dissipation to be derived. The kinetic model can be fitted to in vitro single cell resolution data derived from kinetic flow cytometric studies by non-linear regression. We propose that kinetic determination of cumulative frequency distributions provides a useful approach for estimating apoptosis sensitivity across cell populations over short time-frames.

Apoptosis↗
PubMed2002

Pk11195, a mitochondrial benzodiazepine receptor antagonist, reduces apoptosis threshold in Bcl-X(L) and Mcl-1 expressing human cholangiocarcinoma cells.

BACKGROUND: Cholangiocarcinoma cells express high levels of the antiapoptotic proteins Bcl-X(L) and Mcl-1 and are markedly chemo- and radioresistant. Mitochondria have emerged as central players in apoptosis. Antiapoptotic members of the Bcl-2 protein family localise to the outer mitochondrial membrane and regulate mitochondrial release of apoptogenic proteins. Mitochondrial benzodiazepine receptor (mBzR) ligands have been shown to reverse Bcl-2 action and facilitate apoptosis. AIM: We evaluated the ability of the mBzR antagonist Pk11195 to overcome preapoptotic mitochondrial dysfunction in Egi-1 and Tfk-1, two human cholangiocarcinoma cell lines expressing high levels of Bcl-X(L) and Mcl-1. MATERIALS AND METHODS: Cells growing in culture were used to perform in vitro experiments over 48-96 hours following treatment. The cytotoxic agents used were 5 fluorouracil 10 microM and etoposide (Vp16) 10 microM, together with ultraviolet and 0.5-1 Gy x ray irradiation with or without 75 microM Pk11195. Apoptosis and mitochondrial dysfunction were measured at single cell resolution by flow cytometry using the mitochondrial fluorochrome DiOC6(3). Severe combined immunodeficient non-obese diabetic (SCID-NOD) mice with subcutaneous xenografts using the Egi-1 and Tfk-1 cell lines were treated with etoposide with or without addition of Pk11195 over a 72 hour period during which time the xenograft growth patterns were monitored. RESULTS: In vitro, the effect of Pk11195 on induction of apoptosis in cholangiocarcinoma cells following stimulation by chemotherapy or radiotherapy was found to be both time and dose dependent, with Pk11195 increasing rates of apoptosis by 50-95%. Intraperitoneal administration of Pk11195 in combination with Vp16 was found to increase the growth inhibiting effects of Vp16 on xenografts during the treatment phase. PK11195 75 microM on its own had no intrinsic cytotoxic efficacy. CONCLUSION: This is the first study to demonstrate that functional antagonism of coexpressed Bcl-X(L) and Mcl-1 proteins using the mBzR antagonist Pk11195 can facilitate apoptosis in cholangiocarcinoma following chemotherapy and radiotherapy.

Antineoplastic Agents↗
PubMed2001

Bcl-2 resistant mitochondrial toxicity mediated by the isoquinoline carboxamide PK11195 involves de novo generation of reactive oxygen species.

Resistance to apoptosis is a major obstacle preventing effective therapy for malignancy. Mitochondria localized anti-death proteins of the Bcl-2 family play a central role in inhibiting apoptosis and therefore present valid targets for novel therapy. The peripheral benzodiazepine receptor (PBR) shares a close physical association with the permeability transition pore complex (PTPC), a pivotal regulator of cell death located at mitochondrial contact sites. In this study we investigated the cytotoxicity of the PBR ligand, PK11195, in the micromolar concentration range. PK11195 induced antioxidant inhibitable collapse of the inner mitochondrial membrane potential (DeltaPsi(m)) and mitochondrial swelling in HL60 human leukaemia cells, but not in SUDHL4 lymphoma cells (which exhibited a higher level of reduced glutathione and relative tolerance to chemotherapy or pro-oxidant induced DeltaPsi(m)dissipation). PK11195 induced the production of hydrogen peroxide that was not inhibited by Bcl-2 transfection, nor depletion of mitochondrial DNA. ROS production was however blocked by protonophore, implicating a requirement for DeltaPsi(m). Our findings suggest that PK11195-induced cytotoxicity relies upon Bcl-2 resistant generation of oxidative stress; a process only observed at concentrations several orders of magnitude higher that required to saturate its receptor.

Antineoplastic Agents↗
PubMed2001

Human IgM anti-GM1 autoantibodies modulate intracellular calcium homeostasis in neuroblastoma cells.

Increased titers of IgM anti-GM1 antibodies are present in some patients with Lower Motor Neuron Disease (LMND) or Motor Neuropathy (MN), but their pathogenic role and the mechanism of action are unclear. Previous studies have shown that the B subunit of Cholera Toxin (CT), which binds and crosslinks ganglioside GM1, modulate intracellular calcium in murine neuroblastoma cells via the activation of L-type voltage-dependent calcium channels (VGCC). Therefore, using a fluorimetric approach, we have examined the hypothesis that the pentameric IgM anti-GM1 antibodies, could similarly alter calcium concentration in N18 neuroblastoma cells. Sera with human IgM anti-GM1 antibodies were obtained from 5 patients with LMND and 2 patients with MN. Human IgG anti-GM1, IgM anti-Myelin Associated Glycoprotein (MAG), IgM anti-sulfatide antibodies and lectin peanut agglutinin (PNA), that recognizes specifically the Gal(betal-3)GalNAc epitope, were used as control sera. Direct application of either human IgM anti-GM1 antibodies or the B subunit of CT to N18 neuroblastoma cells induced a sustained influx of manganese ions, as indicated by a quench of the intracellular fura-2 fluorescence. Furthermore, the dihydropyridine L-type channel antagonists completely inhibited the manganese influx, suggesting that it is due to activation of an L-type VGCC. The magnitude of the influx was correlated with antibody titers. None of human IgG anti-GM1, IgM anti-MAG, IgM anti-sulfatide antibodies or PNA induce an ion influx, pointing to the selective participation of the pentameric IgM isotype of anti-GM1 in the modulation of L-type calcium channels opening. Given that L-type calcium channels are present on motor neurons, the modulation of L-type calcium channels by IgM GM1 antisera may have important implications in diseases such as LMND and MN.

Adult↗
PubMed2000

Normal and chronic phase CML hematopoietic cells repopulate NOD/SCID bone marrow with different kinetics and cell lineage representation.

INTRODUCTION: Chronic myelogenous leukemia is characterized by a clonal expansion of abnormal hematopoietic cells, which eventually replaces normal hematopoiesis. We wanted to test the hypothesis that the growth kinetics of CML and normal hematopoietic cells are different. MATERIALS AND METHODS: We compared the growth kinetics and the phenotype of engraftment of chronic phase CML and normal human CD34(+) precursor cells in the bone marrow of immune deficient mice. RESULTS: High levels of engraftment of normal precursors occurred early and consisted of myeloid, erythroid, megakaryocytic, and lymphoid elements. This level and pattern of engraftment were maintained at later assessments. The level of CML cell engraftment was initially much lower, but it increased progressively at late time-points with no indication of a plateau in growth. Early engraftment of CML cells consisted almost entirely of myeloid and mast cells but soon after only mast cells were detectable. Conversely mast cells were infrequent in mice engrafted with normal progenitors. CONCLUSION: We conclude that in contrast to normal cell engraftment, engraftment of CML cells in NOD/SCID mice is characterized by a slow but progressive myeloid infiltration, which eventually consists almost entirely of mast cells.

Animals↗
PubMed1999

Assessment of value and applications of in vitro testing of topical dermatological drug products.

The FDA recently issued a guidance covering practices of scaleup and post approval changes with semisolids (SUPAC-SS). This guidance outlines the steps that must be taken by a company to maintain certification of its semisolid dermatological products after quantitative changes have been made in their compositions and/or after changes have been made in the sourcing of their key ingredients, in their processing, in their batch sizes, and/or after their site of manufacture has been relocated. A key element within the guidance is a release test to be used to determine if the diffusional release of a drug found in a formulation is the same after changes have been made to the formulation as it was prior to implementing the changes. The AAPS-FDA sponsored workshop was set up to explore this qualifying test. The stated aims of the workshop were: a) to illustrate the methodology and techniques of in vitro release testing, b) to show the sensitivity of in vitro release with respect to manufacturing variables and to variations in components and composition (of specific formulations), c) to recognize in vitro release testing as a useful procedure for SUPAC documentation, d) to highlight and evaluate other applications of in vitro release testing, e) to explore the degree to which in vitro release testing and bioavailability may be related, and f) to evaluate the role of in vitro release testing of topical dosage forms as a tool to improve product quality.

Administration, Topical↗
PubMed1998

The kinetics and extent of engraftment of chronic myelogenous leukemia cells in non-obese diabetic/severe combined immunodeficiency mice reflect the phase of the donor's disease: an in vivo model of chronic myelogenous leukemia biology.

In vitro studies have provided little consensus on the kinetic abnormality underlying the myeloid expansion of chronic myelogenous leukemia (CML). Transplantation of human CML cells into non-obese diabetic mice with severe immunodeficiency disease (NOD/SCID mice) may therefore be a useful model. A CML cell line (BV173) and peripheral blood cells collected from CML patients in chronic phase (CP), accelerated phase (AP), or blastic phase (BP) were injected into preirradiated NOD/SCID mice. Animals were killed at serial intervals; cell suspensions and/or tissue sections from different organs were studied by immunohistochemistry and/or flow cytometry using antihuman CD45 monoclonal antibodies (MoAbs), and by fluorescence in situ hybridization (FISH) for the BCR-ABL fusion gene. One hour after injection, cells were sequestered in the lungs and liver, but 2 weeks later they were no longer detectable in either site. Similar short-term kinetics were observed using 51Cr-labeled cells. The first signs of engraftment for BV173, AP, and BP cells were detected in the bone marrow (BM) at 4 weeks. At 8 weeks the median percentages of human cells in murine marrow were 4% (range, 1 to 9) for CP, 11% (range, 5 to 36) for AP, 38.5% (range, 18 to 79) for BP, and 54% (range, 31 to 69) for BV173. CP cells progressively infiltrated BM (21%) and spleen (6%) by 18 to 20 weeks; no animals injected with the cell line or BP cells survived beyond 12 weeks. The rate of increase in human cell numbers was higher for BP (7.3%/week) as compared with CP (0.9%/week) and AP (0. 5%/week). FISH analysis with BCR and ABL probes showed that some of the human cells engrafting after injection of CP cells lacked a BCR-ABL gene and were presumably normal. We conclude that CML cells proliferate in NOD/SCID mice with kinetics that recapitulate the phase of the donor's disease, thus providing an in vivo model of CML biology.

Animals↗
PubMed1998

Metabolic neuropathies.

A large epidemiological study has documented that one-third of diabetic patients have peripheral neuropathy. Diabetes duration, poor glycaemic control, smoking and hypertension are all independent predictors of the incidence of diabetic polyneuropathy. High prevalence of autonomic dysfunctions, both sympathetic and parasympathetic, has been found in patients with nonalcoholic chronic liver disease. The pathogenesis of metabolic neuropathy is unclear; even immunologic factors might play a role in the development of diabetic autonomic neuropathy. No specific treatments are available for these neuropathies. Correction of metabolic derangement is fundamental, as shown by the amelioration of peripheral nerve function obtained after successful simultaneous pancreas-kidney transplantation. The therapeuthic potentials of neurotrophins for the prevention and treatment of diabetic neuropathy have to be confirmed in future studies.

Diabetes Mellitus, Type 1↗
PubMed1997

BCL-2 antisense therapy in patients with non-Hodgkin lymphoma.

BACKGROUND: Overexpression of BCL-2 is common in non-Hodgkin lymphoma and leads to resistance to programmed cell death (apoptosis) and promotes tumorigenesis. Antisense oligonucleotides targeted at the open reading frame of the BCL-2 mRNA cause a specific down-regulation of BCL-2 expression which leads to increased apoptosis. Lymphoma grown in laboratory animals responds to BCL-2 antisense oligonucleotides with few toxic effects. We report the first study of BCL-2 antisense therapy in human beings. METHODS: A daily subcutaneous infusion of 18-base, fully phosporothioated antisense oligonucleotide was administered for 2 weeks to nine patients who had BCL-2-positive relapsed non-Hodgkin lymphoma. Toxicity was scored by the common toxicity criteria, and tumour response was assessed by computed tomography scan. Efficacy was also assessed by quantification of BCL-2 expression; BCL-2 protein levels were measured by flow cytometry in samples from patients. FINDINGS: During the course of the study, the daily dose of BCL-2 antisense was increased incrementally from 4.6 mg/m2 to 73.6 mg/m2. No treatment-related toxic effects occurred, apart from local inflammation at the infusion site. In two patients, computed tomography scans showed a reduction in tumour size (one minor, one complete response). In two patients, the number of circulating lymphoma cells decreased during treatment. In four patients, serum concentrations of lactate dehydrogenase fell, and in two of these patients symptoms improved. We were able to measure BCL-2 levels by flow cytometry in the samples of five patients, two of whom had reduced levels of BCL-2 protein. INTERPRETATION: In patients with relapsing non-Hodgkin lymphoma, BCL-2 antisense therapy led to an improvement in symptoms, objective biochemical and radiological evidence of tumour response, and down-regulation of the BCL-2 protein in some patients. Our findings are encouraging and warrant further investigations of BCL-2 antisense therapy in cancer treatment.

Adult↗
PubMed1997

Motor nerve biopsy studies in motor neuropathy and motor neuron disease.

The clinical presentation of motor neuropathy often resembles that of motor neuron disease, sometimes leading to an erroneous diagnosis. Moreover, the underlying pathological process in motor neuropathy has been rarely investigated and there are no systematic studies of the affected motor nerves. We describe a new motor nerve biopsy procedure, performed in 15 patients: 6 with motor neuropathy and 9 with motor neuron disease. The motor branch from the anterior division of the obturator nerve to the gracilis muscle in the thigh was biopsied. In both groups of patients the motor nerves exhibited depletion of myelinated nerve fibers. In motor neuropathy there was a significantly higher density of regenerative clusters of small myelinated fibers in comparison to motor nerves from patients with motor neuron disease. In addition, in 3 patients with motor neuropathy there was evidence for demyelination with thinly myelinated axons and small onion bulb formations. These pathological studies of motor nerve biopsies can help to differentiate motor neuropathy from motor neuron disease.

Adult↗
PubMed1997

Diagnostic biopsy of the motor nerve to the gracilis muscle. Technical note.

A biopsy specimen of the motor nerve to the gracilis muscle was obtained in 151 patients to aid in the diagnosis of neuromuscular disorders. The procedure is simple, safe, and has low morbidity and complication rates. The surgical technique is described. Indications for the use of this technique are motor neuropathy, motor neuron disease, Guillain-Barre syndrome, and mixed neuropathy with prominent motor symptoms.

Adult↗
PubMed1996

Anti-MAG and anti-SGPG antibodies in neuropathy.

We compared the binding of human antibodies from patients with neuropathy to the myelin-associated glycoprotein (MAG), to its cross-reactive glycolipid sulfoglucuronyl paragloboside (SGPG), and to sections of peripheral nerve. Titers were correlated with the clinical presentation and results of electrophysiological and pathological studies. Most patients had a predominantly sensory or sensorimotor demyelinating neuropathy and highly elevated antibodies to both MAG and SGPG, but 2 had highly elevated antibodies to MAG alone, and 1 to SGPG alone. Two patients had predominantly motor neuropathy and highly elevated antibodies to SGPG which reacted with MAG by Western blot but not by enzyme-linked immunosorbent assay. One patient had amyotrophic lateral sclerosis and antibodies to SGPG but not to MAG. These studies indicate that the neuropathic syndrome associated with anti-MAG or -SGPG antibodies are more heterogeneous than previously suspected, and that although most of the antibodies react with both MAG and SGPG, some may react with MAG or SGPG alone.

Aged↗
PubMed1996

[Colpocytology in patients on hormonal contraceptives].

In this paper, the authors analyse retrospectively 737 colpocytologic examinations done on patients on hormonal contraception, on IUD contraception and in a control group. A careful examination of data revealed no statistically relevant differences among the various groups.

Adolescent↗
PubMed1994

Identification of Gal(beta 1-3)GalNAc bearing glycoproteins at the nodes of Ranvier in peripheral nerve.

A subset of human anti-GM1 ganglioside antibodies cross-reacts with Gal(beta 1-3)GalNAc bearing glycoproteins in peripheral nerve and spinal cord. The same oligosaccharide determinant is recognized by the lectin peanut agglutinin (PNA) which binds at the nodes of Ranvier in intact peripheral nerve. The Gal(beta 1-3)GalNAc bearing glycoproteins were isolated using PNA lectin affinity chromatography followed by separation on Western blot, and the proteins were subjected to partial amino acid sequence analysis. Two major PNA binding glycoproteins were identified in peripheral nerve and spinal cord; one had an approximate molecular weight of 120 kD and had sequence homology to the oligodendrocyte-myelin glycoprotein (OMgp). The other migrated between 70 and 80 kD and had sequence homology to the hyaluronate binding domain of versican, which has been reported to share sequence homology with the 70 kD proteins hyaluronectin and the glial hyaluronic acid binding protein (GHAP). By immunocytochemistry, OMgp was localized to the paranodal region of myelin, and the protein homologous to the hyaluronate binding domain of versican was localized to the nodal gap in peripheral nerve. These PNA binding glycoproteins might be target antigens for autoantibodies in peripheral nerve.

Amino Acid Sequence↗
PubMed1994

A pleiotropic response is induced in F9 embryonal carcinoma cells and rhino mouse skin by All-trans-retinoic acid, a RAR agonist but not by SR11237, a RXR-selective agonist.

We evaluated SR11237, a retinoid X receptor (RXR)-specific compound, for its pharmacologic effects on cell differentiation in F9 embryonal carcinoma cells and rhino mouse epidermis. SR11237 can cause RXR/RXR homodimers to form and transactivate a reporter gene containing a RXR-response element. We confirmed, using nuclear receptor co-transfection assays in COS-1 cells, that SR11237 is effective at transactivating a chloramphenicol acetyltransferase reporter gene through RXRs but not retinoic acid receptors. When SR11237 was tested for its ability to modulate cell differentiation, it was inactive on F9 embryonal carcinoma cells and rhino mouse skin. Because differentiation in these systems is known to be regulated by RAR-specific compounds, such as all-trans-retinoic acid and (E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1-prope nyl benzoic acid], our results with SR11237 are compatible with the concept that classical retinoid pleiotropic responses are mediated by RXR/RAR heterodimeric nuclear receptors rather than through RXR/RXR homodimers.

Animals↗