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Biomedical subjects

C Soranzo

Publications and source records attributed to C Soranzo.

At least 19 recordsLinked to original sources
PubMed2001

Extensive traumatic soft tissue loss: reconstruction in severely injured patients using cultured hyaluronan-based three-dimensional dermal and epidermal autografts.

BACKGROUND: This report demonstrates the potential of two-stage autologous keratodermal grafting as a starting point for noninvasive reconstruction of extensive traumatic soft tissue defects. METHODS: In three severely injured patients, skin biopsies for cell cultivation were taken. Cultured "neodermis" consisting of cultured autologous fibroblasts grown on biocompatible three-dimensional scaffolds made up of benzyl ester of hyaluronan was grafted on conditioned defect areas. After ingrowth of dermal substitutes, transplantation of cultured autologous keratinocytes on hyaluronan-based laser-perforated membranes was performed. Ten days later, a 0.2-mm thin, 1:6 meshed autograft was overlaid. Clinical follow-up, histologic, and immunohistochemical findings were documented. RESULTS: Grafting with cultured autologous fibroblasts revealed a suitable dermal tissue replacement. Epithelialization was evident after transplantation of keratinocytes. Final closure of the defects with "normoelastic" tissue properties was achieved after thin mesh-grafting. CONCLUSION: Preliminary findings with the described method seem to be very promising. As in all fields of tissue engineering, long-term studies and further follow-up are required.

Adjuvants, Immunologic↗
PubMed1998

In vitro engineering of human skin-like tissue.

Coverage of large, full-thickness burns presents a challenge for the surgeon due to the lack of availability of the patient's own skin. Currently, tissue engineering offers the possibility of performing a suitable therapeutic wound coverage after early burn excision by using cultured keratinocyte sheets supported by a dermal layer. The aim of this study was to develop and characterize a skin substitute composed of both epidermal and dermal elements. For this purpose we grew keratinocytes and fibroblasts separately for 15 days within two different types of biomaterials. Cells then were co-cultured for an additional period of 15 days, after which samples were taken and processed with either classic or immunohistochemical stainings. Results showed that (1) human fibroblasts and keratinocytes can be cultured on hyaluronic acid-derived biomaterials and that (2) the pattern of expression of particular dermal-epidermal molecules is similar to that found in normal skin. The data from this study suggest that our skin equivalent might be useful in the treatment of both burns and chronic wounds.

Biocompatible Materials↗
PubMed1997

Cerebrospinal-fluid ciliary neurotrophic factor in neurological patients.

We developed a double sandwich immunoassay for the dosage of ciliary neurotrophic factor (CNTF) in cerebrospinal fluid (CSF). The detection limit was 100 pg/ml. This assay was applied to human CSF samples from 14 normal subjects, 26 patients with multiple sclerosis (MS), 17 with Guillain-Barré syndrome (GBS) or chronic inflammatory demyelinating polyneuropathy (CIDP), and 22 with tumours of the central nervous system (CNS) or leucaemic meningosis (LM). Samples from normal control subjects and from patients with tumours did not contain detectable CNTF. Only 2 patients with LM were positive, and all the patients with inflammatory diseases of the CNS and peripheral nervous system were positive. The MS group presented a mean value of 240 pg/ml CNTF and the GBS/CIDP group a value of 430 pg/ml.

Astrocytoma↗
PubMed1995

Effects of keratinocyte-secreted soluble factors on spreading, number of dendrites and cell-cell contacts of human epidermal melanocytes and dermal fibroblasts: a quantitative analysis.

We previously demonstrated that spreading and clustering of in vitro cultured human keratinocytes are autocrine-induced phenomena, mediated by keratinocyte-secreted soluble factor. In this paper, the effects of this factor on spreading, number of dendrites and cell-cell contacts of the two cellular components of skin, melanocytes and fibroblasts have been studied 24 h after plating cells on uncoated plastic surfaces in MCDB153 serum-free medium or in the same medium conditioned by keratinocytes (KCM). Spreading of melanocytes present in the epidermal cell population remained constant at increasing cell density, while that of keratinocytes showed a statistically significant increase. Moreover, time-course experiments showed that the rate of spreading was faster for melanocytes. At increasing epidermal cell density, a statistically significant increase in number of dendrites and cell-cell contacts of melanocytes was observed. Similar results were obtained when melanocytes were plated both in coculture with keratinocytes (as epidermal cell cultures) or as a pure cell population in keratinocyte conditioned medium (KCM), suggesting that the observed phenomena are due to keratinocyte-secreted soluble factors and not to direct keratinocyte-melanocyte interactions. The addition of nerve growth factor (NGF) to fresh medium or addition of an inactivating anti-NGF monoclonal antibody (alpha D11) to KCM did not affect the number of dendrites or cell-cell contacts of melanocytes. Keratinocyte-secreted soluble factor(s) present in KCM also dramatically influenced morphology and cell-cell contacts of human dermal fibroblasts.

Cell Adhesion↗
PubMed1994

Characterization of an established human, malignant, glioblastoma cell line (GBM) and its response to conventional drugs.

A cell line, GBM, was established from a human malignant glioblastoma and was characterized with particular reference to its response to conventional drugs. The GBM cell line exhibited a 73 +/- 7 h doubling time in monolayer cultures. Expression of glial fibrillary acidic and S-100 proteins was observed. Karyotype analysis of GBM cells at early passages revealed the presence of two near-triploid clones (A and B) with multiple chromosome rearrangements; a 100% frequency for clone B was observed in the established cell line. GBM cells had tumorigenic properties, since the s.c. injection of cultured cells into nude mice gave rise to slowly growing tumors. The morphology of GBM cells was retained during in vitro and in vivo passages, as judged by light microscopy. GBM cells were relatively resistant to most conventional drugs; among the tested drugs, only taxol exhibited a marked cytotoxic effect comparable to that found in cells of a different tumor type. GBM cells were found positive for the epidermal growth factor receptor, HER2-neu and P-glycoprotein by flow cytometry of cells labelled with monoclonal antibodies. In spite of the expression of relatively high gamma-glutamyltransferase activity, the intracellular glutathione level was comparable to that of other chemosensitive tumor cells. This glioblastoma cell line is a suitable model for the identification and preclinical studies of new agents and provides an additional system to explore the molecular basis of the intrinsic drug resistance of glioblastoma.

Animals↗
PubMed1994

Nerve growth factor (NGF) in cerebrospinal fluid (CSF) from patients with various neurological disorders.

It has been recently shown that NGF is not only involved in the survival and development of sympathetic and neural crest-derived sensory neurons, but also in some mechanisms of the immune system. For this reason, we studied the content of NGF in CSF samples from patients with diseases in which neuroimmunological mechanisms seem to be involved (multiple sclerosis, amyotrophic lateral sclerosis, Alzheimer disease, chronic relapsing polyradiculoneuritis, Guillain-Barré syndrome, and tumors of the nervous system), as well as from a number of normal control subjects. We setup an ELISA aimed at the beta subunit of NGF, obtaining good validation tests and a detection limit of 28 pg beta NGF per ml. None of the samples was found to contain detectable levels of NGF and, when a concentration method for sample enrichment was used, only one patient was NGF-positive. This suggests that NGF is probably not involved in the neuroimmunological mechanisms underlying some inflammatory and degenerative diseases of the nervous system.

Antibody Specificity↗
PubMed1994

Establishment and characterization of a CHO cell line producing a secreted form of human ciliary neurotrophic factor: neuroprotective effects of the recombinant protein.

Human ciliary neurotrophic factor (CNTF) was inserted into a mammalian expression vector linked to the prepro sequence of human nerve growth factor. A Chinese hamster ovary cell line was established by resistance to neomycin and the plasmid integrated DNA was amplified using the metallothionein gene. This cell line contained several hundred copies of the human CNTF gene and produced an NH2 terminal truncated form of human CNTF (22 kDa) which was secreted into the medium. Although the copy number of the human CNTF gene was high and its mRNA was actively transcribed, the recombinant protein secreted into the medium constituted only 35-40% of the total amount of human CNTF synthesized by these cells. Both wild-type human CNTF produced in bacterial cells and the human CNTF obtained by forced secretion were effective in protecting hippocampal pyramidal neurons from injury induced by glucose deprivation, a form of excitotoxic neurodegeneration.

Amino Acid Sequence↗
PubMed1993

Effect of modulation of protein kinase C activity on cisplatin cytotoxicity in cisplatin-resistant and cisplatin-sensitive human osteosarcoma cells.

The effect of modulation of protein kinase C (PKC) activity by 12-O-tetradecanoylphorbol-13-acetate (TPA) on cisplatin cytotoxicity was examined in a human osteosarcoma U2-OS cell line and in a U2-OS variant (U2-OS/Pt) selected after continuous exposure to increasing concentrations of cisplatin. U2-OS/Pt cells showed a 7.5-fold resistance to the drug. A 24 h exposure of cells to TPA caused a potentiation of cisplatin cytotoxicity in sensitive and in resistant cells; under these conditions, PKC activity was shown to be down-regulated. In contrast, a short-term exposure of cells to TPA did not affect cisplatin cytotoxicity in U2-OS or in U2-OS/Pt cells. These results support the involvement of PKC in cellular response to cisplatin. However, this enzyme is probably not directly implicated in the mechanisms of acquired resistance in this cell system.

Cisplatin↗
PubMed1993

Production and characterization of monoclonal antibodies to human ciliary neurotrophic factor with defined epitope recognition.

Ten mouse hybridoma lines producing monoclonal antibodies (Mabs) against recombinant human ciliary neurotrophic factor (rhCNTF) have been obtained. Two monoclonal antibodies belonging to the IgG1 class were selected and characterized. Their specificity was established by ELISA and Western blotting. Epitopes recognized by the two Mabs were investigated with ELISA and Western blotting by using rhCNTF mutants, rhCNTF fragments and synthetic peptides mimicking different portions of the CNTF molecule. The carboxy-terminal part of the CNTF and particularly the sequence between aa 150 and 159 appeared to constitute the immunodominant group. The fact that certain amino acid sequences of CNTF are conserved among species was utilized to examine the crossreactivity patterns of the two Mabs with rat sciatic nerve CNTF by Western blotting and immunohistochemistry. These antibodies will be useful for studying the distribution of CNTF in the nervous system and in developing an enzyme-linked immunosorbent assay for the quantitative determinations of CNTF in various neuropathologies.

Amino Acid Sequence↗
PubMed1990

Large scale purification and immunological characterization of human placental nerve growth factor.

Nerve growth factor (NGF) is a protein which plays a critical role in the development and survival of not only peripheral neurons, but possibly also cholinergic brain neurons. The present study describes a procedure for large scale isolation of human NGF of placental origin, and its immunological characterization. A protein species of approximately 26 kDa was obtained, which cross-reacted with antibodies to mouse NGF. Polyclonal and monoclonal anti-mouse NGF antibodies appeared to recognize different bands within this human NGF preparation. Although these polyclonal antibodies recognized both the dimeric and monomeric forms of mouse NGF, the monoclonal antibody recognized only a band corresponding to the dimeric form of mouse NGF.

Antibodies↗
PubMed1990

Different interaction of cisplatin and etoposide on in vivo and in vitro tumor systems.

The effect of cisplatin (CDDP) in combination with etoposide (VP16) was examined on four human cell lines (one colon adenocarcinoma, LoVo; one ovarian carcinoma, IGROV-1; two small cell lung cancers, NCI-H146 and NCI-N592; and two murine leukemias, P388 and L1210). Simultaneous exposure to CDDP and subtoxic concentrations of VP16 for 1 h produced a cell killing in all cell lines comparable to that achieved by CDDP alone. Sequential exposure of NCI-H146 and NCI-N592 to CDDP for 1 h followed by VP16 for 96 h again produced an additive effect. When two of these cell lines were treated in in vivo models (i.p. P388 leukemia, s.c. NCI-N592) with suboptimal doses of the two drugs, a potentiation of the antitumor effects of the two drugs in simultaneous combination was evidenced by the increase in survival time and in the number of 'cures' in P388 leukemia bearing mice and by the inhibition of tumor size in NCI-N592. This comparative study, using the same cell lines in vitro and in vivo, indicates that the CDDP-VP16 potentiation observed in vivo does not reflect a specific interaction at the cellular-biochemical level. The results support the therapeutic interest of this combination (presumably as a result from favorable pharmacological interactions) even despite the lack of potentiation at cellular level, under comparable conditions of treatment.

Animals↗
PubMed1990

Effect of tumor necrosis factor on human tumor cell lines sensitive and resistant to cytotoxic drugs, and its interaction with chemotherapeutic agents.

We investigated the cytotoxic effects of recombinant tumor necrosis factor (TNF) alone and in combination with interferon-gamma (IFN-gamma) and/or cytotoxic drugs on a variety of human tumor cell lines (U937, IGROV-1, HT29, LoVo, MCF7 and U20S), including cell lines with in vitro acquired resistance (LoVo/DX and MCF7/DX selected for resistance to doxorubicin (DX) and characterized by pleiotropic drug resistance; U20S selected for resistance to cisplatin (CDDP], using MTT assay. U937 and MCF7 were sensitive to the cytotoxic effect of TNF, whereas all the other cells were insensitive up to 1000 U/ml (the maximum tested dose). Surprisingly, TNF was cytotoxic (30-40% cytotoxicity) against two resistant lines (LoVo/DX and U20S/Pt) but not against the parent sensitive lines. Treatment with increasing doses of TNF after 6 h incubation with a subtoxic concentration of IFN-gamma produced a synergistic effect in four cell lines (U937, HT29, LoVo/DX and MCF7), whereas in the other five the cell killing of the combination was comparable with that achieved by TNF alone. The combination of subtoxic doses of TNF and increasing doses of drugs targeted at DNA topoisomerase II (i.e. DX, actinomycin D and VP16) produced an additive cytotoxic effect in all cell lines. The same results were obtained combining TNF and CDDP, except in U20S/Pt cells in which TNF synergistically increased CDDP cytotoxicity. The combination of TNF and IFN-gamma enhanced cytotoxicity about 20-fold for DX and 6-fold for CDDP, evaluated in terms of the modification index, against LoVo/DX and U20S/Pt cells respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Antineoplastic Agents↗
PubMed1989

An exponential-Gompertzian description of LoVo cell tumor growth from in vivo and in vitro data.

The data of nine monolayer cultures, 48 multicellular spheroids, and 19 s.c. xenografts of LoVo cells were fitted, on an individual basis, by exponential and Gompertzian equations, respectively. The mean growth parameters alpha 0 (initial growth rate) and beta (retardation factor) of the three experimental systems presented a strong linear correlation alpha 0 = 6.88 beta + 0.56, r = 0.9843. This implies that, at the particular tumor size of 155 microns in diameter (mean +/- SD = 50-310 microns), the tumor growths described by Gompertzian curves (from spheroids as well as from s.c. xenografts) have the same growth rate as monolayer cultures. This occurrence strongly supports an exponential-Gompertzian growth model, where an exponential monolayer-like phase changes to a Gompertzian growth, controlled by environmental conditions, when the tumor size has reached a critical value.

Carcinoma↗
PubMed1989

Changes in the three-dimensional organization of LoVo cells associated with resistance to doxorubicin.

LoVo cells and a derived subline resistant to doxorubicin were compared in the spheroids system. The resistant line, unlike the parent one, was unable to grow as spheroids, but formed irregular loose aggregates. Moreover treatment of the resistant cells with membrane-active agents able to reverse pleiotropic drug resistance had no effect on the capability of these cells to grow as spheroids. The results indicate that the inability of resistant cells to form spheroids is not related to the resistance mechanism.

Adenocarcinoma↗
PubMed1988

Development and survival of neurons in dissociated fetal mesencephalic serum-free cell cultures: I. Effects of cell density and of an adult mammalian striatal-derived neuronotrophic factor (SDNF).

The use of CNS cultures for detection and quantification of neuronotrophic activity in the CNS has been analyzed. In particular the development, i.e., neurotransmitter uptake characteristics, and survival of dopaminergic and GABAergic neurons in fetal mouse (E13)-dissociated mesencephalic cells cultured in serum-free, hormone-supplemented medium have been assessed as a function of culture time and cell density. At all times, more than 98% of the cells were classified as neurons on the basis of immunocytochemical criteria. Results indicate that the increase of cell density in vitro significantly enhances specific high-affinity dopamine uptake per dopaminergic cell and cell survival. This effect is not limited to the dopaminergic cells and suggests that the development of neurotransmitter-related traits and cell survival are influenced by cell density-derived trophic signals. The above-mentioned cultures and parameters have also been used to detect neuronotrophic activity in adult mammalian brain extracts or more purified preparations. In particular, bovine striatal extracts contain activity capable of increasing high-affinity neurotransmitter uptake parameters and cell survival of at least the dopaminergic and GABAergic neurons present in the culture system. The neuronotrophic activity from bovine striatum has been partially purified and is associated with a fraction whose main component is a basic protein of approximately 14 kDa.

Animals↗
PubMed1986

Formation, growth and morphology of multicellular tumor spheroids from a human colon carcinoma cell line (LoVo).

LoVo human colon carcinoma cells cultured by a liquid overlay technique form and grow as multicellular tumor spheroids. The growth curve of LoVo spheroids exhibits Gompertzian growth kinetics, i.e., exponential growth for 10 days, followed by exponential retardation in the rate of growth. Doubling time in the exponential growth phase is longer than in monolayer cultures (5 days for LoVo spheroids vs. 37 h for monolayers). When LoVo spheroids reach a diameter of about 300 microns, a necrotic core appears in their center and continuously increases with spheroid growth. The cell ultrastructure and organization in spheroids closely resemble those of the same cells when grown as tumors in vivo or as monolayer, i.e. intestinal epithelium, desmosomes, intracytoplasmic lumina and acinar structures. Individual cells from spheroids can be obtained by trypsinization and assayed for colony formation. LoVo spheroids provide a model which can be readily manipulated and appears to be suitable for evaluation of anticancer drugs. A comparison of LoVo spheroids exposed to doxorubicin with the same cells grown in monolayers emphasized the role of cell organization in determining drug resistance.

Cell Line↗
PubMed1986

Lack of effect of glutathione depletion on cytotoxicity, mutagenicity and DNA damage produced by doxorubicin in cultured cells.

Since endogenous glutathione (GSH), the main non-protein intracellular thiol compound, is known to provide protection against reactive radical species, its depletion by diethylmaleate (DEM) was used to assess the role of free radical formation mediated by doxorubicin in DNA damage, cytotoxicity and mutagenicity of the anthracycline. Subtoxic concentrations of DEM that produced up to 75% depletion of GSH did not increase doxorubicin cytotoxicity in a variety of cell lines, including Chinese hamster ovary (CHO) and lung (V-79) cells, LoVo human carcinoma cells and P388 murine leukemia cells. Similarly, the number of doxorubicin-induced DNA single strand breaks in CHO cells and the mutation frequency in V-79 cells were not affected by GSH depletion. The results obtained suggest that mechanisms other than free radical formation are responsible for DNA damage, cytotoxicity and mutagenicity of anthracyclines.

Animals↗