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Biomedical subjects

L Oliver

Publications and source records attributed to L Oliver.

At least 19 recordsLinked to original sources
PubMed1995

The role of exogenous/endogenous basic fibroblast growth factor (FGF2) and transforming growth factor beta (TGF beta-1) on human corneal endothelial cells proliferation in vitro.

Adult human corneal endothelial cells (HCEC) have extremely low turnover rates but undergo rapid division in vitro when stimulated with soluble growth factors. We have investigated the role played by FGF2 and TGF beta-1 in the regulation of HCEC growth stimulation. HCEC from donors who were over 30 years old were cultured and experiments performed on cultures between the 2nd and the 6th passage in the presence of 5% NCS. Cell counts revealed a maximal stimulation of 2.1x for FGF2 and 1.9x for TGF beta-1 compared to control cultures. When both factors were added, a synergistic effect was noticed with a maximal stimulation of the proliferation rate of 4.5x over controls. In addition, endogenous FGF2 produced by HCEC was quantitated in a sensitive EIA assay. After 5 days in culture, 10(6) cells contained 150 ng FGF2 and 35 ng was extracted from trypsin-digested ECM. Two molar NaCl washes of ECM released 15.6 ng FGF2, which induced a slight mitogenic activity (1.5x over control) in HCEC cultures, which was partially inhibited by an anti-FGF2 antibody. Northern blot analysis of HCEC extracts revealed the presence of FGF receptors R1 and R2 mRNA. The bioactive FGFRs were demonstrated by the toxic effect of a mitotoxin FGF2-SAP. These results suggest that FGF2 could participate in the autocrine regulation of HCEC proliferation and survival. The synergy between exogenously added FGF2 and TGF beta demonstrates that a combination of different growth factors may be important to stimulate proliferation of these cells in vivo.

Adult↗
PubMed1994

Up-regulation of aFGF expression in quiescent cells is related to cell survival.

Exogenously administrated acidic FGF modulates the proliferation of several cell types, controls cell differentiation, and promotes cell survival. Most cells that are sensitive to exogenous aFGF are also capable of expressing it at very low levels. Thus in order to establish the role of endogenous aFGF as a mitogenic, differentiation, or survival factor, we studied the regulation of aFGF expression by evaluating the level of mRNA by PCR amplification and the concentration of protein by Enzyme Immuno Assay (EIA). In the lens, the amount of aFGF transcripts in nondividing cells of the central epithelium and in the differentiated fiber cells located at the periphery of the lens is similar, suggesting that endogenous aFGF is not involved with lens differentiation. In cultures, depending on the growth conditions, the endogenous aFGF expressed by Bovine Epithelial Lens (BEL) cells is subject to modulation. Cells arrested either by contact inhibition or by serum deprivation express more aFGF transcripts and protein than in exponentially growing cells, implying that endogenous aFGF has no mitogenic role under these conditions. In serum-deprived cells, the addition of specific aFGF antisense primers inhibits endogenous aFGF expression and leads to the death of these cells. These results associated with the higher expression of aFGF in nondividing BEL cells, suggesting that, contrary to exogenous aFGF, endogenous aFGF is not a mitogenic factor but a survival factor.

Animals↗
PubMed1994

Effects of exogenous FGFs on growth, differentiation, and survival of chick neural retina cells.

Fibroblast growth factors (FGFs) are known to play important roles in various processes including development and differentiation. Chick embryo neural retina cells, capable of transdifferentiation into lentoid bodies and pigmented cells, were used in vitro to examine the effects of exogenous acidic (aFGF) and basic FGF (bFGF) on proliferation and protein accumulation. We demonstrate that both factors increase the proliferation of glial cells and modulate the survival of neurons without affecting protein accumulation within these cells. Moreover, FGFs stimulate the differentiation of the photoreceptors. The rate of proliferation varies over the period of culture, with a maximum occurring after 2 weeks, followed by a decrease concommitant with the appearance of lentoid bodies. The concentration of aFGF was measured using an enzyme immuno assay and showed an accumulation of this protein only in bFGF-treated cultures, suggesting that bFGF positively modulates aFGF synthesis in neural retina cell cultures.

Animals↗
PubMed1993

Acidic fibroblast growth factor is expressed abundantly by photoreceptors within the developing and mature rat retina.

In order to further understand the role(s) of fibroblast growth factors (FGFs) in the development, differentiation and function of the central nervous system, we analysed the expression of the mRNA, and the presence and tissue distribution of the translated product, of one member of the FGF family, acidic FGF (aFGF), within the mammalian retina. Firstly, the relative abundance of aFGF mRNA was assayed in embryonic (between 14 and 17 days of gestation), postnatal (between 1 and 17 days after birth) and adult rat retina by quantitative reverse transcription-coupled polymerase chain reaction amplification using specific aFGF oligonucleotides. The level of expression remained uniformly low throughout the embryonic period and until postnatal day 7. Therefore the quantity of aFGF mRNA increased rapidly, reaching 80% of adult levels by eye opening (postnatal day 13). Adult levels were three-fold higher than at early developmental times. In situ hybridization of adult rat retina using specific antisense aFGF riboprobes revealed labelling in all cellular layers. Antisera raised against recombinant human aFGF revealed very little labelling of 4-day postnatal retina, but by postnatal days 8 and 17 immunoreactive aFGF was localized mainly within the photoreceptor cell bodies. Western blots of retinal extracts derived from 17-day embryonic, 4-day postnatal and adult retina probed with the same antibody revealed a single immunoreactive band of the expected molecular weight (18 kDa) in all extracts. Thus aFGF is mostly transcribed and translated within the retina subsequent to the major steps of cell birth, migration and differentiation, and seems to be abundantly expressed by maturing photoreceptor cells.

Animals↗
PubMed1993

Developmental regulation of acidic fibroblast growth factor (aFGF) expression in bovine retina.

Acidic fibroblast growth factor (aFGF) is a signalling molecule implicated in a wide variety of biological processes such as cell growth, differentiation and survival. It has been purified from bovine retina. The present study was carried out to detect which cells in the bovine retina expressed aFGF at the different stages of embryonic and post-natal development. The specific aFGF mRNA and protein were detected by in situ hybridization employing riboprobes and immunocytochemistry using affinity purified polyclonal human recombinant aFGF antibodies respectively. No signal was detected by either technique until 4-5 months and then there was progressive expression of aFGF with terminal morphogenesis of the retina. By 8-9 months of embryonic development, nuclei of the 3 neuronal layers (ganglion cell layer, inner and outer nuclear layers) were all uniformly and intensely labeled. A slight labeling of the pigmented epithelium of the retina was also visible throughout development and maturation. These results showed a good correlation between message and protein expression in these cell types. In contrast, glial cells in the nerve fiber layer and vascular endothelial cells displayed a nuclear immunostaining for the protein in the absence of message. These data suggest that aFGF plays a role in the late steps of retinal differentiation by autocrine and paracrine mechanisms.

Animals↗
PubMed1992

Regulation of proteolytic activity in human bone marrow stromal cells by basic fibroblast growth factor, interleukin-1, and transforming growth factor beta.

Plasminogen activators (PAs) and/or plasmin may be involved in hematopoietic regulation. These enzymes release biologically relevant cytokines such as basic fibroblast growth factor (bFGF) from matrix and cell surfaces. In addition, transforming growth factor beta (TGF beta) and interleukin-1 beta (IL-1 beta) are converted from inactive to active forms by plasmin. Therefore, we studied the regulation of PAs and their specific inhibitors, PA inhibitor 1 (PAI-1) and PA inhibitor 2 (PAI-2), in human bone marrow stromal fibroblasts by IL-1 beta, bFGF, and TGF beta. All three cytokines stimulated PA secretion. IL-1 beta at 10(4) U/mL increased urokinase (u-PA) levels approximately 10-fold, bFGF at 0.2 ng/mL also increased production 10-fold, but increased predominantly tissue PA (t-PA) expression. TGF beta at 0.2 ng/mL increased u-PA production up to 300-fold. PAI-1 and PAI-2 are also regulated by these cytokines. IL-1 beta decreased PAI-1 levels by 50% and stimulated PAI-2 levels sixfold. bFGF had minimal effects on PAI-1 and TGF beta increased PAI-1 levels twofold. Neither of these agents had an effect on PAI-2 levels. Thus, three cytokines relevant to bone marrow physiology regulate PA and inhibitor production by human bone marrow stromal fibroblasts. In this manner PA and plasmin generation in specific microenvironments in the bone marrow may be one of the factors orchestrating the complex series of events, which results in an efficient exquisitely regulated hematopoietic process.

Bone Marrow↗
PubMed1992

Endogenous aFGF expression and cellular changes after a demyelinating lesion in the spinal cord of adult normal mice: immunohistochemical study.

Fibroblast growth factors (FGFs) are known to act on glial cells in vitro. At the present time, their involvement in the remyelinating process of the adult central nervous system (CNS) is still unknown. In the present study, using immunohistochemistry (IHC), we investigated the evolution in time and space of acidic FGF (aFGF) expression and CNS cell changes occurring after a chemically induced demyelinating lesion. In a first early period, aFGF immunostaining was shown to decrease around the demyelinated area. A dramatic increase was then observed and was accompanied by an increase of cell density around and inside the lesion. This was correlated with the beginning of remyelination. Late after demyelination, while remyelination was still in progress, aFGF immunostaining of the lesion and unlesioned spinal cord were comparable. A role of aFGF in remyelination is proposed.

Animals↗
PubMed1992

Acidic fibroblast growth factor (aFGF) in developing normal and dystrophic (mdx) mouse muscles. Distribution in degenerating and regenerating mdx myofibres.

Affinity purified polyclonal antibodies directed against human recombinant acidic FGF (aFGF), were used in immunofluorescence studies to localize this growth factor in several normal and dystrophic (mdx) mouse skeletal muscles. The expression of aFGF was detected throughout the life of both the control and mdx mice. In striated muscles, examined up to 3 weeks postnatal, aFGF was localized around the myofibres and this pattern was consistent in both mdx and the normal counterpart strain. However, the intensity of the signal was much stronger in the mdx strain. In mdx mouse skeletal muscles, examined during the acute phase of degeneration and regeneration (3-14 weeks) aFGF was localized around the myofibres, in approximately 60% of the nuclei of newly formed or regenerated myofibres and also in the pockets of necrosis which represented actively degenerating myofibres. In normal mouse skeletal muscles, studied over the same period, the antibodies localized aFGF mainly to the periphery of the muscle fibres. The augmentation of aFGF observed by immunofluorescence in mdx mouse muscles was confirmed by enzyme immunoassay (EIA) analysis of the same muscles over the same period of time. The data from the EIA indicated a 3.5-fold increase in aFGF in mdx as compared to normal muscles at 3 weeks, and an approximate 26-fold increase during the period of active degeneration-regeneration. This increased concentration of aFGF noted in the mdx muscles suggests that this endogenous aFGF may participate in the high level of regenerative activity observed in mdx mouse.

Animals↗
PubMed1992

Immunolocalization of acidic and basic fibroblast growth factors during mouse odontogenesis.

Acidic and basic fibroblast growth factors (aFGF and bFGF), are both known to bind to extracellular matrix components, particularly proteoheparin sulfates, and to regulate in vitro proliferation, differentiation and morphology of cells of neuroectodermal and mesodermal origins. Their patterns of distribution were studied during mouse odontogenesis by means of indirect immunofluorescence and immunoperoxidase histochemistry on frozen fixed sections and after Bouin's fixative and paraffin embedding. Localization of aFGF on frozen fixed sections was observed in the oral epithelium, dental lamina and oral mesenchyme (day-12 of gestation), the stellate reticulum and oral epithelium (day-14), the stratum intermedium and at the basal and apical poles of preameloblasts at bell stage. After birth aFGF epitopes were localized within the predentin-dentin area, the stratum intermedium and at the secretory pole of ameloblasts. There was no staining with anti-aFGF antibodies after Bouin's fixative and paraffin embedding. In contrast, using this protocol, intense stainings were found with anti-bFGF antibodies predominantly within dental and peridental basement membranes and mesenchyme: staining of the dental basement membranes was transient (bud and cap stage) and discontinuous; a preferential concentration of bFGF epitopes in the condensed dental mesenchyme of incisors (cap stage) and the dental papillae mesenchymal cells of molars (bell stage) was observed in the posterior and the cervical part of tooth germs. An intense immunostaining of the stellate reticulum with anti-bFGF antibodies was also found on paraffin sections from bud to bell stage.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Acidic fibroblast growth factor (aFGF) is expressed in the neuronal and glial spinal cord cells of adult mice.

Fibroblast growth factors (FGFs) are known to be synthesized in the central nervous system (CNS) and to act on CNS cells in vitro, but less is known about their synthesis, expression, and role in vivo. In this work, using specific anti-acidic fibroblast growth factor (aFGF) antibodies, we have shown for the first time, by immunohistochemistry, that aFGF is expressed in spinal cord cells of young adult normal mice. This expression is predominant in the cell nucleus. Using immunohistochemical double staining procedures, we identified the cell type expressing aFGF as neurons, astrocytes, and oligodendrocytes, but for each type, cells were not all positively immunostained.

Animals↗
PubMed1991

Hepatocyte growth factor in human placenta and trophoblastic disease.

The hepatocyte growth factor (HGF) is an acidic protein with a strong mitogenic effect on hepatocytes. Hepatocyte growth factor mRNA recently was cloned from a placental cDNA library. Here we demonstrate the purification of HGF from human placenta with heparin-agarose chromatography and TSK-heparin high-pressure liquid chromatography and describe the distribution of placental HGF by immunohistochemistry using a polyclonal antibody to HGF. The yield of HGF from the placenta was approximately 100 to 200 times greater than that previously obtained from human plasma. Placental HGF was expressed strongly in the villous syncytium, extravillous trophoblast, and amnionic epithelium, and, to a lesser degree in endothelial cells and villous mesenchyme. Hepatocyte growth factor also was identified in the trophoblast of complete hydatidiform moles, choriocarcinomas, and a case of blighted ovum. The presence of HGF in an organ characterized by rapid cell proliferation during gestation and in trophoblastic tumors strongly suggests that the growth-regulating effect of HGF is not limited to hepatocytes.

Abortion, Spontaneous↗
PubMed1990

Identification and partial characterization of receptor binding sites for HGF on rat hepatocytes.

Hepatocyte Growth Factor (HGF) (also known as Hepatopoietin A [HPTA] (1-9) is a heterodimeric heparin-binding polypeptide mitogen for hepatocytes distinct from other well-known growth factors. In this study, biologically active radioiodinated HGF was used to identify binding sites on intact hepatocytes in culture. The results show the presence of relatively low affinity binding sites due to the presence of heparin or heparin-like molecules and high affinity specific receptor binding sites on the cell surface of intact hepatocytes. Scatchard analysis of binding data indicates an apparent dissociation constant (Kd) of 3.5 nM with 120,000 sites per hepatocyte for the cell-surface receptor. Analysis of affinity cross-linked 125I-HGF-receptor complex by SDS-PAGE under non-reducing conditions reveals the presence of a distinct band with apparent Mr of 230,000. These data show that HGF exerts its biological effect on hepatocytes (stimulation of DNA synthesis) through a specific and unique cell-surface receptor.

Animals↗
PubMed1990

Urinary dysfunction in transverse myelitis.

Six men and 2 women with a history of transverse myelitis and persistent lower urinary tract symptoms underwent neurourological evaluation. Of the patients, 4 were neurologically intact, while the remainder had residual neurological deficits. Urodynamic studies revealed detrusor-external sphincter dyssynergia in 6 patients. Two patients had detrusor hyperreflexia, of whom 1 also had an incompetent sphincter. Erectile or ejaculatory dysfunction was reported by 3 men. We conclude that prolonged bladder and sexual dysfunction, caused by spinal cord inflammatory insult, may persist despite a systemic neurological recovery. Therefore, bladder management guided by initial and followup urodynamics is recommended.

Adult↗
PubMed1990

Thermoluminescence dosimetry in the Prince of Wales Hospital, Sydney.

Thermoluminescence dosimeters (TLDs) were introduced in the Radiotherapy Department of the Prince of Wales Hospital in 1975. They are used in three different types of applications: 1. to confirm doses in complicated geometries (e.g. axilla, ear, nose), 2. to measure the dose delivered to critical organs (e.g. lens, scrotum) and 3. to monitor certain treatments like total body irradiations (TBI). Using four LiF chips (TLD 100; size: 3.1mm x 3.1mm x 0.9mm) per measuring point the accuracy of each measurement is +/- 5%. Approximately 200 TLD measurements are requested from the planning radiographers per year. Lens dose determination (32% of all cases) and total body irradiations are the main applications. Since 1988 the planning radiographers have been asked to state their expectation of the dose where it could be estimated. In 14% of these cases (7% of all requests) the expected and the measured dose differed by more than 20%. This was usually followed by revising the treatment set-up and a repeated TLD measurement. For about one third of all patients more than one dose measurement is required either to monitor the treatment or to investigate changes in the delivered dose after changes in the treatment set-up. Several changes to treatment modalities (e.g. boli in various treatments, a changed scrotal shield) were made due to TLD results. This is reflected in the still increasing number of TLD requests which demonstrates the benefit of TLDs in clinical practice.

Humans↗
PubMed1986

Two-dimensional characterization of prostatic acid phosphatase, prostatic specific antigen and prostate binding protein in expressed prostatic fluid.

Specimens of pooled prostatic fluid, collected by rectal massage from men under 50 years of age with no apparent prostatic disorders, were subjected to two-dimensional gel electrophoresis to study the composition of its proteins. In a preliminary study, a total of 57 major protein groups were detected. In the present study, we attempted to identify, in the two-dimensional gels, those that are related to prostate-associated proteins, i.e., prostatic acid phosphatase (PAP), prostatic specific antigen (PSA), and prostate binding protein (PBP). Individual proteins were recognized by the procedure of Western Blot using specific antisera with peroxidase-antiperoxidase as the staining reagent. Each protein spot in the two-dimensional gel was expressed, along the abscissa, by its isoelectric point (pI) and, along the ordinate, by the molecular weight (MW). PAP consisted of a train of more than ten protein spots that occupied an area in the gel from pI 7.0, MW 45,000 to pI 6.0, MW 50,000. Four protein spots with a MW of 34,000 and a pI range of 8.2-8.8 were identified as PSA. PBP was observed as having three protein spots that were located at pI 5.6-6.6 with a single MW of 15,000. For PAP and PSA, additional protein spots with lower MWs also stained positively with the specific antisera, suggestive of the presence of degradative products of these proteins. Following the removal of the serum-related proteins by an extensive absorption with anti-human serum antibody by affinity chromatography, the prostatic fluid contained 27 major groups of non-serum proteins. These non-serum proteins in the prostatic fluid included PAP, PSA, PBP, and their related smaller molecular species. These results indicate that the prostatic fluid contains PAP, PSA, PBP and that their presence and the patterns of their distribution in the two-dimensional gels should be considered as the characteristic property of the prostatic secretions.

Acid Phosphatase↗
PubMed1985

Chromosomal localization of the human elastin gene.

mRNA isolated from fetal human aorta was used to synthesize cDNA that was cloned into the PstI site of pBR322. The recombinant clones were screened with an authentic sheep elastin cDNA, and one human clone that hybridized strongly was isolated and characterized. The 421-base pair (bp) insert of this human clone was sequenced by the dideoxy method, and the DNA sequence showed strong homology to the nontranslated portion of the sheep elastin cDNA. This result unequivocally identified the human clone, designated pcHEL1, as an elastin clone. Plasmid pcHEL1 labeled with [3H] nucleotides was used in in situ hybridization experiments utilizing normal metaphase chromosomes and also with cells carrying a balanced translocation between chromosomes 1 and 2: 46,XY,t(1;2)(p36;q31). The results strongly suggest that the elastin gene is localized to the q31----qter region of chromosome 2.

Animals↗
PubMed1984

Characterization of a sheep elastin cDNA clone containing translated sequences.

mRNA, isolated from the ligamentum nuchae of fetal sheep by guanidine HCl extraction and oligo(dT) cellulose chromatography, was used to synthesize blunt-ended cDNA molecules by the successive application of AMV reverse transcriptase, DNA polymerase and S1 nuclease. The cDNA was centrifuged on a 15-30% sucrose gradient and molecules greater than 700 bp were tailed with dCTP and cloned into the PstI site of pBR322 which had been tailed with dGTP. Ampicillin-sensitive and tetracycline-resistant colonies were screened by in situ hybridization with elastin-enriched mRNA that had been terminally labeled with 32p. Recombinant plasmids prepared from strongly hybridizing colonies were characterized by restriction mapping and the plasmid with the largest insert (1300 bp) thought to contain elastin sequences was characterized in more detail. The nick-translated cDNA hybridized to a single 3.5 kb mRNA species upon blot hybridization, a size identical to that previously identified for chick elastin mRNA (Burnett et al. (1982) J. Biol. Chem. 259, 1569-1572). Nucleotide sequencing of the 5' end of the cDNA demonstrated a sequence which was extremely GC rich and which corresponded to an amino acid sequence partially homologous to that previously identified in porcine tropoelastin (Foster et al. (1973) J. Biol. Chem. 248, 2876-2879). This is the first report of the identification of a plasmid containing sequences complementary to a translated region of elastin mRNA.

Amino Acid Sequence↗