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Biomedical subjects

F Lin

Publications and source records attributed to F Lin.

At least 109 records · Page 6Linked to original sources
PubMed1996

Effects of epidermal growth factor (EGF) and colony-stimulating factor-1 (CSF-1) on expression of c-fos in rat mandibular molars: implications for tooth eruption.

The c-fos gene is expressed in the dental follicle of the first mandibular molar of rats. Molecules that accelerate tooth eruption, epidermal growth factor (EGF) and colony-stimulating factor-1 (CSF-1), enhance the expression of the c-fos gene in vitro in both a time- and concentration-dependent manner. In vivo, EGF enhances c-fos expression in the follicle from day 0-7 postnatally, but by day 9 the follicle is refractory to this stimulus. Immunostaining reveals that the c-fos gene is translated in the cultured dental follicle cells, with staining seen in the nucleus and the perinuclear region of the cytoplasm. In vivo, immunostaining for c-fos is prominent in the dental follicle early postnatally, with little or no staining seen in the stellate reticulum and dental pulp. By day 10 postnatally, staining for c-fos is greatly reduced in the dental follicle. Thus, the presence of c-fos early postnatally in the tissue required for eruption, the dental follicle, as well as the enhancement of c-fos gene expression in the follicle by EGF or CSF-1, suggests that c-fos plays a role in tooth eruption. That role may be either to promote differentiation of mononuclear cells into osteoclasts needed for eruption or to recruit the mononuclear cells into the follicle.

Animals↗
PubMed1996

In vivo and in vitro effects of epidermal growth factor on its receptor gene expression in rat dental follicle cells.

Epidermal growth factor receptor (EGFR) is known to be localized early postnatally in the follicle of rat mandibular molars and to decline later. Here, EGFR mRNA present early postnatally in the follicle was revealed by in situ hybridization and reverse transcription-polymerase chain reaction. Injections of epidermal growth factor (EGF) enhanced the expression of EGFR mRNA but not EGF mRNA through day 5 postnatally in the follicle. By day 7 postnatally, the amount of EGFR mRNA was dramatically reduced and EGF injection at that time or later did not enhance its expression. Cultured dental follicle cells were also shown to contain both EGFR mRNA and EGF mRNA, but only EGFR mRNA expression was enhanced after incubation of the cells with EGF. The EGFR mRNA was apparently translated, because the cultured cells could be immunostained for EGFR. This enhancement of EGFR mRNA in the dental follicle cells by EGF may be a necessary prelude to the mitogenic effect of EGF on the cultured cells because incubating the dental follicle cells in EGF over 6 days resulted in almost a doubling in their number as compared to controls. These results indicate that the tissue necessary for eruption to occur, the dental follicle, contains EGFR mRNA in the early postnatal days. In turn, this mRNA is enhanced by EGF, a molecule known to accelerate eruption if injected early postnatally. Whether or not the mitogenic effect of the EGF on the cultured dental follicle cells is a mechanistic factor that contributes to the subsequent tooth eruption is unknown.

Animals↗
PubMed1996

Selective photosensitizer localization in the human endometrium after intrauterine application of 5-aminolevulinic acid.

OBJECTIVE: Our purpose was twofold: to determine the distribution of the endogenous photosensitizer protoporphyrin IX in the uterus and to ascertain the time interval leading to maximal endometrial fluorescence after intrauterine instillation of 5-aminolevulinic acid. STUDY DESIGN: One milliliter of a 400 mg/ml 5-aminolevulinic acid-Hyskon solution was instilled into the uterine cavity of 27 women before hysterectomy. On frozen sections of uterine samples 5-aminolevulinic acid-induced fluorescence was measured with fluorescence microscopy. RESULTS: 5-Aminolevulinic acid-induced fluorescence could first be detected in the superficial endometrial glands 75 minutes after drug injection. In the endometrial gland stumps fluorescence intensity peaked 4 to 8 hours after 5-aminolevulinic acid instillation and was > 48 times higher than in the underlying myometrium. CONCLUSIONS: Fluorescence in the endometrial glands suggests that selective photodynamic destruction of the endometrium may be possible 4 to 8 hours after intrauterine 5-aminolevulinic acid instillation.

Aminolevulinic Acid↗
PubMed1996

Single-channel currents produced by the serotonin transporter and analysis of a mutation affecting ion permeation.

Single-channel activities were observed in outside-out patches excised from oocytes expressing a mammalian 5-hydroxytryptamine (5-HT) transporter. Channel conductance was larger for a mutant in which asparagine177 of the third putative transmembrane domain was replaced by glycine, suggesting that this residue lies within or near the permeation pathway. The N177G mutant enables quantitative single-channel measurements; it displays two conducting states. One state, with conductance of approximately 6 pS, is induced by 5-HT and is permeable to Na+. The other state (conductance of approximately 13 pS) is associated with substrate-independent leakage current and is permeable to both Na+ and Li+. Cl- is not a major current carrier. Channel lifetimes under all conditions measured are approximately 2.5 ms. The single-channel phenomena account for previously observed macroscopic electrophysiological phenomena, including 5-HT-induced transport-associated currents and substrate-independent leakage currents. The channel openings occur several orders of magnitude less frequently than would be expected if one such opening occurred for each transport cycle and therefore do not represent an obligatory step in transport. Nevertheless, single-channel events produced by neurotransmitter transporters indicate the functional and structural similarities between transporters and ion channels and provide a new tool, at single-molecule resolution, for detailed structure-function studies of transporters.

Animals↗
PubMed1996

Bioaccumulation of nickel by intercalation into polycrystalline hydrogen uranyl phosphate deposited via an enzymatic mechanism.

A Citrobacter sp. accumulates uranyl ion (UO2(2+)) as crystalline HUO2PO4.4H2O (HUP), using enzymatically generated inorganic phosphate. Ni was not removed by this mechanism, but cells already loaded with HUP removed Ni2+ by intercalative ion-exchange, forming Ni(UO2PO4)2.7H2O, as concluded by x-ray diffraction (XRD) and proton induced x-ray emission (PIXE) analyses. The loaded biomass became saturated with Ni rapidly, with a molar ratio of Ni:U in the cellbound deposit of approx. 1:6; Ni penetration was probably surface-localized. Cochallenge of the cells with Ni2+ and UO2(2+), and glycerol 2-phosphate (phosphate donor for phosphate release and metal bioprecipitation) gave sustained removal of both metals in a flow through bioreactor, with more extensively accumulated Ni. We propose 'Microbially Enhanced Chemisorption of Heavy Metals' (MECHM) to describe this hybrid mechanism of metal bioaccumulation via intercalation into preformed, biogenic crystals, and note also that MECHM can promote the removal of the transuranic radionuclide neptunium, which is difficult to achieve by conventional methods.

Biomass↗
PubMed1996

Cutaneous pseudosarcomatous polyp: a histological and immunohistochemical study.

Two unusual acquired polypoid skin lesions exhibited prominent histological atypia, but were biologically benign. Both patients were elderly females. The lesions clinically mimicked fibroepithelial polyp or nevus lipomatosus. Both had been present for about 20 years. One lesion was located on the back, the other on the posterior thigh. Each lesion exhibited dilated, hyalinized vessels in the dermis with focal fibrin deposits, myxoid stroma, and a population of bizarre, pleomorphic spindle to stellate cells, some of which were multinucleated. Occasional atypical mitoses were present. One lesion had abundant admixed fat. Immunohistochemical staining was strongly positive only for vimentin. The lesions share features with degenerating angiofibroma and vaginal pseudosarcomatous polyp. As in these lesions, the atypia is most probably reactive and degenerative.

Aged↗
PubMed1996

Role of hydrogen peroxide in the cytotoxic effects of UVA/B radiation on mammalian cells.

Effects of selenium (Se) deficiency on the sensitivity of murine leukemia L1210 cells to broad band UVA/B radiation (310-400 nm) have been investigated. Cells rendered glutathione peroxidase (GPX) deficient by shortterm (2-3 week) growth in 1%, serum/RPMI medium without added Se [L.Se(-) cells] were found to be much less resistant to clonally assessed UVA/B lethality than Se-supplemented controls [L.Se(+) cells]. By contrast, long-term ( > 20 week) Se-deprived [L'.Se(-)] cells whose catalase (CAT) activity was elevated > 100-fold were far more resistant to UVA/B than L.Se(+) cells. Similar trends were observed for cells irradiated in 1% serum/RPMI or Hank's medium. Whereas the CAT inhibitor 3-amino-1,2,4-triazole had no effect on L.Se(+) photosensitivity, it produced a large increase in L'.Se(-) photosensitivity. These findings are consistent with H2O2 intermediacy in photokilling and suggest that L1210 cells depend mainly on GPX for protection against this species but switch to overexpressed CAT after chronic Se deprivation. In agreement with this, steady-state H2O2 levels measured by H2O2 electrode during UVA/B exposure were higher in L.Se(-) than L.Se(+) suspensions but much lower (barely detectable) in L'.Se(-) suspensions. Cytotoxic effects of UVA/B and variations thereof resulting from Se manipulation could be mimicked by treating cells with glucose oxidase in the presence of D-glucose, providing further support for H2O2 involvement. Whether UVA/B-generated H2O2 is directly cytotoxic or gives rise to a more damaging species such as hydroxyl radical (HO) is presently unknown.

Animals↗
PubMed1996

Cytochrome P450 4A expression and arachidonic acid omega-hydroxylation in the kidney of the spontaneously hypertensive rat.

20-Hydroxyeicosatetraenoic acid (20-HETE) is a major arachidonate metabolite in the kidney of the spontaneously hypertensive rat (SHR). The increase in its synthesis has been associated with the elevation of blood pressure in the SHR. The omega-hydroxylation of arachidonic acid is an activity associated with members of the CYP4A gene family which, in the rat, comprises three major isoforms: 4A1, 4A2 and 4A3. 20-HETE displays potent and diverse biological activities which can affect pro- and anti-hypertensive mechanisms dependent upon where, when and by which isoform it has been produced. Therefore, it is important to identify and characterize its biosynthetic system. We compared CYP4A mRNA and protein expression to patterns of 20-HETE synthesis in the SHR kidney. The reverse transcription/polymerase chain reaction (RT/PCR) technique was used to amplify CYP4A mRNA in microdissected nephron segments. Southern blot hybridization of PCR products obtained from nephron segments with the CYP4A1 cDNA probe demonstrated strong signals in S2 and S3 segments of the proximal tubule. Immunoblots of nephron segments using a polyclonal anti-rat liver CYP4A1 antibody which cross-reacts with CYP4A2 and CYP4A3, and 14C-arachidonic acid metabolism, confirmed that arachidonic acid omega-hydroxylation, i.e., 14C-20HETE formation, and CYP4A proteins were also localized mainly in the S2 and S3 segments. Correlation also existed between the age-dependent increase in arachidonate omega-hydroxylation in the kidney and CYP4A mRNA levels as measured by Northern hybridization of total RNA using the CYP4A1 cDNA probe. Immunoblot analysis revealed that at 7 weeks, where 20-HETE production is at its maximum, all three proteins are expressed. CYP4A3 and 4A1 immunoreactive proteins appeared at 3 weeks, showed maximum levels at 5 and 7 weeks, respectively, and gradually decreased to lower levels at 13 and 20 weeks, whereas CYP4A2 levels were undetectable at 3, 5 and 7 weeks but appeared at 13-20 weeks. Additional immunoblots indicated that renal cortical CYP4A1 protein levels were higher in SHR compared to Sprague-Dawley and Wistar-Kyoto rats. The increased levels of CYP4A1-immunoreactive band at 7 weeks corresponded to the maximal activity of arachidonate omega-hydroxylation. Thus, CYP4A1 might play a significant role in contributing to the increased cortical/proximal production of 20-HETE seen in 7-week-old SHR. However, given the high homology among members of the CYP4A gene family and the lack of specific tools to discern among these isoforms, additional studies have to be carried out to substantiate our findings.

Aging↗
PubMed1996

Reverse transcription/polymerase chain reaction (RT/PCR) amplification of very small numbers of transcripts: the risk in misinterpreting negative results.

Technical modifications of the reverse-transcription/polymerase chain reaction (RT/PCR) amplification method now permit its use to detect amplified products from as few as one abnormal cell, either isolated or mixed with a larger number of normal cells. We studied the reproducibility of such results using as targets low numbers of cells from chronic myeloid leukaemia (CML) patients and CML cell lines in quintuplicate two-step RT/PCR designed to amplify BCR-ABL sequences. When one K562 or KYO1 cell was diluted in 10(3) non-CML HL60 cells, an amplification product was obtained in each test; at greater dilutions BCR-ABL transcripts were detected erratically. Titration of cDNA synthesised from 5 x 10(7) cells from four CML patients showed that whereas positive BCR-ABL sequences could be amplified in some tests starting with as little as a 1 in 10(7) dilution of cDNA template (corresponding to 5-10 cells), the dilution threshold for reproducible amplification was around 1 to 5 in 10(5) (100-500 cells). Quantitative PCR analysis revealed that reactions from 1 in 10(7) diluted cDNA contained less than 10 BCR-ABL transcripts as the starting template. The stochastic nature of the amplification from such small numbers of transcripts was illustrated by results of 10 replicate PCR tests on cDNA from a patient expressing both b3a2 and b2a2 transcripts: dilutions of cDNA up to 1 in 10(5) yielded dual transcript amplification in all 10 tests, but the 1 in 10(7) cDNA dilution resulted in b3a2 and b2a2 products in three tests, b3a2 only in three, b2a2 only in one and no amplification in three tests. We conclude that this 'sampling effect' may yield false-negative results and thus misinterpretation of data regarding assessment of gene expression when the quantity of target material available for study is very small.

False Negative Reactions↗
PubMed1996

Magnetic resonance imaging of primary osteosarcoma of the lumbar vertebra with metastasis to the perirenal fat.

Primary osteosarcoma of the spine not associated with any predisposing factor is rare. The authors describe a case of osteosarcoma of the lumbar spine in a 44-year-old woman with biopsy-proven ulcerative colitis. The patient underwent bone scanning, computed tomography (CT) and magnetic resonance imaging (MRI), and the authors present the first record of the MRI appearance of this disease in a vertebral body. Relative to classical osteosarcoma of long bone, this case involved a higher degree of soft-tissue ossification, which was readily visible with both CT and MRI. The patient had extensive metastasis, including metastasis to the perirenal fat, and died 10 months after presentation.

Adipose Tissue↗
PubMed1996

Reconstitution with Philadelphia chromosome-negative recipient haematopoiesis early after allogeneic BMT for CML.

We report a 33-year-old man with Ph chromosome-positive CML who underwent an allogeneic BMT from an unrelated donor. DNA microsatellite studies showed complete donor chimaerism immediately after BMT followed by mixed chimaerism; by day + 45 haematopoiesis was exclusively of recipient origin. Throughout the first year post-transplant all marrow metaphases were Ph negative but with non-clonal rearrangements consistent with autologous recovery. Cytogenetic relapse of leukaemia was first detected 15 months post-transplant. This case is unusual in that non-malignant stem cells of recipient origin survived the transplant and reconstituted haematopoiesis very early after BMT. Later the leukaemic cells reasserted their 'proliferative' advantage.

Adult↗
PubMed1996

Quantitative molecular methods to monitor the response of CML patients to interferon-alpha.

Interferon-alpha (IFN-alpha) induces cytogenetic responses of variable degree in patients with CML. We sought to establish the relationship between BCR-ABL transcript numbers measured by competitive two-step reverse transcription polymerase chain reaction (RT-PCR) and cytogenetic status in CML patients treated with IFN-alpha. All 305 samples from 133 patients investigated by RT-PCR were positive for BCR-ABL transcripts. In order to standardize results for variability in RNA and cDNA quality, we quantified total ABL transcripts in each sample as an internal control. The BCR-ABL : ABL ratios correlated well with the cytogenetic results. Quantitative nested PCR allowed the detection of residual BCR-ABL transcripts in all complete cytogenetic responders on IFN-alpha. We conclude that competitive PCR with internal controls is a reliable method for monitoring patients on IFN-alpha and reduces the need for repeated marrow investigations.

Antineoplastic Agents↗
PubMed1996

Molecular analysis of transient cytogenetic relapse after allogeneic bone marrow transplantation for chronic myeloid leukaemia.

Serial quantification of residual disease in CML patients after allogeneic BMT is useful for early detection of relapse. However, the fact that some cytogenetic relapses appear to be transient may complicate protocols for early therapeutic intervention based on molecular analysis and could result in the unnecessary treatment of some patients. To determine the frequency and significance of transient cytogenetic relapse, we have studied serial samples from 98 CML patients after allogeneic BMT by conventional cytogenetics and competitive RT-PCR for BCR-ABL mRNA. During the period of study, 26 patients had cytogenetic or haematologic evidence or relapse. In four cases (15% of those who relapsed; 4% of all patients) relapse appeared to be transient; i.e., subsequent marrow samples were completely Ph chromosome-negative despite the fact that there had been no change in treatment, including the level of immunosuppression. BCR-ABL mRNA levels broadly paralleled the cytogenetic findings. Of these four patients, two subsequently progressed to frank haematologic relapse and two remained strongly positive for BCR-ABL transcripts and are therefore presumably still at risk of relapse. Analysis of B cell-enriched, T cell-enriched and lymphoid-depleted fractions for three patients demonstrated that transient relapse was not due to the proliferations of BCR-ABL-positive lymphoid cells. In contrast, BCR-ABL-positive myeloid precursor cells were detected in two of three patients tested. We conclude that transient cytogenetic relapse followed by sustained remission is a relatively infrequent occurrence after current allogeneic transplant regimens.

Adult↗
PubMed1996

Synthesis and characterization of human gene 1 relaxin peptides.

The peptide encoded for by one of the two relaxin genes found in the human genome, designated H1, has been synthesized by the Boc-polystyrene solid phase method. The two chains which constitute relaxin, A- and B-, were assembled separately and, after cleavage, deprotection and purification, combined in solution at high pH to form the one intra- and two intermolecular disulfide bonds. Comprehensive chemical characterization including ion spray mass spectrometry of the peptide confirmed both its correct identity and high purity. The synthetic H1 relaxin was analyzed by circular dichroism spectroscopy and shown to possess a greater alpha-helical conformation in water than the corresponding H2 relaxin. The peptide had powerful direct chronotropic and inotropic effects in the isolated rat heart assay as did an analogue of the peptide in which the C-terminus of the B-chain was extended by four residues.

Amino Acid Sequence↗
PubMed1996

New marker for blood vessels in human ovarian and endometrial cancers.

Angiogenesis plays a critical role in tumor biology and may someday be a target for novel therapeutic interventions. To date, however, relatively few markers have been identified that can specifically distinguish between microvessels in benign versus malignant lesions. Here we report that the cationic heme-protein eosinophil peroxidase (EPO) was localized by in situ immunohistochemistry on the vascular endothelial cells and/or connective tissue stroma in 16 of 16 cases of human endometrial carcinoma and in 12 of 15 cases of ovarian carcinoma. Similar deposits of EPO were not detected in normal endometrial tissues or ovaries from five healthy subjects, in adjacent uninvolved tissues from four tumor-bearing subjects, or in any normal organs from five other subjects. These findings imply that eosinophil degranulation is a significant and previously unappreciated component of the interaction between ovarian and endometrial cancers and the host. Moreover, the abundant and highly specific nature of the EPO deposition near and within the microvessels of these cancers suggests that eosinophil degranulation is a new marker for tumor blood vessels that potentially could be exploited to treat these important types of cancers that currently lack highly effective therapies.

Adenocarcinoma↗
PubMed1995

CD and NMR determination of the solution structure of a peptide corresponding to T4 lysozyme residues 38-51.

Solid phase methods have been used to synthesise a peptide corresponding to residues 38-51 of T4 lysozyme. The peptide, LYS(38-51), encompasses helix B in the crystal structure of T4 lysozyme. CD and 1H-NMR analysis showed that the peptide was unstructured in aqueous solution but adopted a helical conformation in the more hydrophobic environment provided by 50% TFE and SDS micelles. The solution structure derived from the NMR data was similar to that of the helix in the X-ray structure, although there was some fraying at the N-terminus.

Bacteriophage T4↗
PubMed1995

Selenoperoxidase-dependent glutathione cycle activity in peroxide-challenged leukemia cells.

Murine leukemia L1210 cells rendered deficient in glutathione peroxidase (GPX) and phospholipid hydroperoxide glutathione peroxidase (PHGPX) by Se deprivation (L.Se(-) cells) were found to be more sensitive to tert-butyl hydroperoxide (t-BuOOH) cytotoxicity than Se-replete controls (L.Se(+) cells). Human K562 cells, which express PHGPX, but not GPX, were also more sensitive to t-BuOOH in the Se-deficient (K.Se(-)) than Se-satisfied (K.Se(+)) condition. In examining the metabolic basis for selenoperoxidase-dependent resistance, we found that glucose-replete Se(-) cells reduce t-BuOOH to t-butanol far more slowly than Se(+) cells, the ratio of the first-order rate constants approximating that of the GPX activities (L1210 cells) or PHGPX activities (K562 cells). Monitoring peroxide-induced changes in GSH and GSSG gave consistent results; e.g., glucose-depleted L.Se(+) cells exhibited a first order loss of GSH that was substantially faster than that of glucose-depleted L.Se(-) cells. Under the conditions used, peroxide-induced conversion of GSH to GSSG could be stoichiometrically reversed by resupplying D-glucose, indicating that no significant lysis or GSSG efflux and/or interchange had taken place. The apparent first-order rate constant for GSH decay increased progressively for L1210 cells expressing a range of GPX activities from approximately 5% to 100%, demonstrating that peroxide detoxification is strictly dependent on enzyme content. The initial rate of 14CO2 release from D-[1-14C]glucose supplied in the medium was much greater for L.Se(+) or K.Se(+) cells than for their respective Se(-) counterparts, consistent with greater hexose monophosphate shunt activity in the former. These results highlight the importance of selenoperoxidase action in the glutathione cycle as a means by which tumor cells cope with hydroperoxide stress.

Animals↗