Biomedical subjects

C Lee

Publications and source records attributed to C Lee.

At least 289 records · Page 16Linked to original sources

PubMed1999

Malignant progression of rat Nb2 lymphoma cells: chromosomal alterations and metastatic properties.

Cultured rat pre-T Nb2 lymphoma cell lines have provided a useful model for tumor progression of T-cell cancers. Comparative analysis of the non-metastatic, prolactin (PRL)-dependent parental Nb2-U17 line and its PRL-independent and/or metastatic sublines, can be used in a search for progression-related genomic alterations. In the present study, the PRL-dependent, cloned Nb2-11C and PRL-independent Nb2-Sp sublines were used to examine development of metastatic ability and PRL independence relative to chromosomal alterations. Metastatic ability was determined using Noble rats carrying subcutaneous tumor transplants; PRL dependence/autonomy was checked in culture. Nb2-11C tumor transplants quickly gave rise to morbidity, associated with metastases in kidney and liver. Transplants of the slower growing Nb2-Sp cells showed variable tumorigenicity as metastases developed in only 40% of the rats (in lungs, kidney, stomach). G-banded chromosome analysis showed the Nb2-11C culture had the karyotype of the parental Nb2-U17 line plus an extra chromosome 19, thus, indicating an association between the development of metastatic ability in Nb2-11C cells and trisomy 19. The Nb2-Sp subline was not clonal. Its stemline showed two alterations in the parental karyotype: acquisition of an add(7)(q10) and loss of the extra chromosome add(15)(p12). Additional abnormalities, add(6)(q11) and trisomy 19, occurred in 15% and 5% of the Nb2-Sp population, respectively. Passaging of the Nb2-Sp subline in vivo resulted in generation and/or outgrowth of new sublines, a major one of which showed an apparent transient growth requirement for lactogens. Possible mechanisms underlying the PRL independence and in vivo properties of the Nb2-Sp cells are discussed.

Animals↗

PubMed1999

Fragile X premutation is a significant risk factor for premature ovarian failure: the International Collaborative POF in Fragile X study--preliminary data.

The preliminary results of an international collaborative study examining premature menopause in fragile X carriers are presented. A total of 760 women from fragile X families was surveyed about their fragile X carrier status and their menstrual and reproductive histories. Among the subjects, 395 carried a premutation, 128 carried a full mutation, and 237 were noncarriers. Sixty-three (16%) of the premutation carriers had experienced menopause prior to the age of 40 compared with none of the full mutation carriers and one (0.4%) of the controls. Based on these preliminary data, there is a significant association between fragile X premutation carrier status and premature menopause.

Adolescent↗

PubMed1999

Cytotoxic T lymphocytes to an unmutated tumor rejection antigen P1A: normal development but restrained effector function in vivo.

Unmutated tumor antigens are chosen as primary candidates for tumor vaccine because of their expression on multiple lineages of tumors. A critical issue is whether unmutated tumor antigens are expressed in normal cells, and if so, whether such expression imposes special restrictions on cytotoxic T lymphocyte (CTL) responses. In this study, we use a transgenic approach to study the development and effector function of T cells specific for P1A, a prototypical unmutated tumor antigen. We report here that although P1A is expressed at low levels in normal tissues, including lymphoid tissues, the P1A-specific transgenic T cells develop normally and remain highly responsive to the P1A antigen. The fact that transgenic expression of P1A antigen in the thymus induces T cell clonal deletion demonstrates that normal hematopoietic cells can process and present the P1A antigen and that P1A-specific T cells are susceptible to clonal deletion. By inference, P1A-specific T cells must have escaped clonal deletion due to low expression of P1A in the thymus. Interestingly, despite the fact that an overwhelming majority of T cells in the T cell receptor for antigen (TCR)-transgenic mice are specific for P1A, these mice are no more resistant to a P1A-expressing plasmocytoma than nontransgenic littermates. Moreover, when the same TCR-transgenic mice were challenged simultaneously with B7-1(+) and B7-1(-) tumors, only B7-1(+) tumors were rejected. Therefore, even though P1A can be a tumor rejection antigen, the effector function of P1A-specific CTL is restrained in vivo. These results have important implications for the strategy of tumor immunotherapy.

Animals↗

PubMed1999

Differential growth rates in stromal cultures of human prostate derived from patients of varying ages.

BACKGROUND: This study was undertaken to attempt to characterize changes in in vitro growth rates and cellular phenotypes of human prostatic stroma associated with aging and/or development of benign prostatic hyperplasia (BPH). METHODS: Prostate stromal cell strains were established from 12 tissue donors of varying age. Culture growth rate was determined by cell counts over a 6-day period. Cell phenotype was assessed by immunocytochemical staining for smooth muscle alpha-actin, smooth muscle myosin, and prolyl-4-hydroxylase. RESULTS: Growth rates of prostate stromal strains in vitro varied. Stromal cells derived from aged males with BPH had significantly slower growth rates than cells from younger donors. A positive reaction for prolyl-4-hydroxylase, a mesenchymal cell marker, was present in all cell cultures regardless of donor age. Expression of smooth muscle-specific actin, a nonspecific smooth muscle cell marker, was present in 48-79% of prostate stromal cultures. Staining for smooth muscle myosin, a specific smooth muscle cell marker, was found to vary significantly with age. The percentage of smooth muscle myosin-positive cells derived from males aged 15, 45, 57, and 72 years were 0.70 +/- 0.14%, 2.12 +/- 0.30%, 4.20 +/- 0.89%, and 26.25 +/- 1.0%, respectively. The shape and size of actin- and/or myosin-positive stromal cells from a 72-year-old donor culture were also usually larger and polygonal in shape as compared to thin and elongated shapes in 15-year-old donor cultures. The shape of actin- and/or myosin-positive cells from a 45-year-old donor culture demonstrated both phenotypes. CONCLUSIONS: These results suggest that in human prostate stromal cells cultured as described, the growth rate decreases, the percent of smooth muscle cells increases, and the cellular shape changes with increasing donor age and/or development of BPH.

Actins↗

PubMed1999

Intracranial hypotension with parkinsonism, ataxia, and bulbar weakness.

OBJECTIVE: To describe a case of spontaneous intracranial hypotension with a previously unreported constellation of presenting features. DESIGN: Case report. SETTING: Tertiary care center. MAIN OUTCOME AND RESULTS: We describe a patient with intracranial hypotension who presented with a parkinsonian syndrome and later development of ataxia and prominent bulbar symptomatology. Headache was not a feature of her initial presentation and was only reported after repeated questioning during later evaluations. Magnetic resonance imaging of the patient's head revealed findings characteristic of intracranial hypotension. An [18F]fluoro-m-tyrosine positron emission tomographic scan showed normal striatal activity, suggesting intact presynaptic nigrostriatal function. Opening pressure on lumbar puncture was reduced at 40 mm H2O. A source of cerebrospinal fluid leakage was not identified on nuclear cisternography and the patient underwent lumbar epidural blood patching, which resulted in complete resolution of her signs and symptoms as well as in a marked improvement in her imaging findings. CONCLUSIONS: The clinical spectrum of intracranial hypotension can be broadened to include parkinsonism, cerebellar ataxia, and prominent bulbar dysfunction. As with more common manifestations of the disorder, these features may resolve after appropriate treatment.

Ataxia↗

PubMed1999

Cytotoxic sensitivity to tumor necrosis factor-alpha in PC3 and LNCaP prostatic cancer cells is regulated by extracellular levels of SGP-2 (clusterin).

BACKGROUND: SGP-2 is a ubiquitous secreted glycoprotein that prevents cellular apoptosis. This study was carried out to determine the extracellular action of SGP-2 in a model of tumor necrosis factor-alpha (TNF)-induced cytotoxicity using two human prostatic cancer lines, LNCaP and PC3. These two lines were selected because LNCaP cells are highly sensitive to the cytotoxic effect of TNF, while PC3 cells are resistant to TNF at 24 hr. METHODS: Cells were cultured in the presence or absence of TNF (10 ng/ml). LNCaP cells were treated with varying concentrations of exogenous SGP-2, while PC3 cells were treated with antisera to SGP-2 with and without exogenous SGP-2. Following a 24-hr treatment, cultures were assessed by counting of cell number and by the trypan blue exclusion assay. RESULTS: Western blot analysis of conditioned media revealed that PC3 secreted more SGP-2 than did LNCaP. The sensitivity to TNF in LNCaP cells was reduced by the addition of exogenous SGP-2. PC3 cells became sensitive to TNF when SGP-2 antibody was added to the culture. The effect of SGP-2 antibody on PC3 cells was reversed by the addition of exogenous SGP-2 to the culture. CONCLUSIONS: These results suggest that SGP-2 can act as an extracellular mediator of anti-TNF-induced cytotoxicity.

Cell Death↗

PubMed1999

Microbial conversion of indene to indandiol: a key intermediate in the synthesis of CRIXIVAN.

Indene is oxidized to mixtures of cis- and trans-indandiols and related metabolites by Pseudomonas putida and Rhodococcus sp. isolates. Indene metabolism is consistent with monooxygenase and dioxygenase activity. P. putida resolves enantiomeric mixtures of cis-1,2-indandiol by further selective oxidation of the 1R, 2S-enantiomer yielding high enantiomeric purity of cis-(1S, 2R)-indandiol, a potential intermediate in the synthesis of indinavir sulfate (CRIXIVAN), a protease inhibitor used in the treatment of AIDS. Molecular cloning of P. putida toluene dioxygenase in Escherichia coli confirmed the requirement for the dihydrodiol dehydrogenase in resolving racemic mixtures of cis-indandiol. Rhodococcus sp. isolates convert indene to cis-(1S, 2R)-indandiol at high initial enantiomeric excess and one isolate also produces trans-(1R, 2R)-indandiol, suggesting the presence of monooxygenase activity. Scale up and optimization of the bioconversions to these key synthons for chiral synthesis of potential intermediates for commercial manufacture of indinavir sulfate are described.

Drug Design↗

PubMed1999

Role of angiotensin II in bladder smooth muscle growth and function.

Preliminary studies by our group and others indicate that angiotensin II may have an important role in the cellular regulation of smooth muscle growth and collagen production in the bladder. The exact mechanisms in which angiotensin II elicits its cellular effects are not known. Given the available information thus far, we hypothesize the following (see Figure 2): 1) Outlet obstruction of the bladder causes increased cell stretch/strain which in turn induces the local production of angiotensin II. Angiotensin II may also influence cell stretch/strain via its direct effects on bladder tone. 2) Angiotensin II then acts as a trophic factor in the bladder wall to cause smooth muscle cell hypertrophy/hyperplasia and increased collagen production via an autocrine and/or paracrine pathway. 3) The cellular effect(s) of angiotensin II may be mediated by secondary growth factors such as bFGF and TGFb Much more extensive research is certainly needed to reveal whether some part, or all of this hypothesis is correct. If angiotensin II is indeed active in regulating muscle and collagen changes in the pathologic bladder, then the clinical implications are extremely exciting since numerous pharmacologic agents are now available which can either inhibit angiotensin II production and/or block receptor mediated events. These agents may prove to be extremely useful in the clinical management of the neurogenic bladder in which obstructive changes may be prevented and potentially reversed. Despite this, caution must be exercised with regard to the potential use of any medications which alter the systemic renin-angiotensin system in the pediatric population since some research has suggested that an intact system may be necessary for the normal development of some organs, including the kidney.

Angiotensin II↗

PubMed1999

Development of a bioconversion process for production of cis-1S,2R-indandiol from indene by recombinant Escherichia coli constructs.

Recombinant Escherichia coli cells expressing the toluene dioxygenase (TDO) genes from Pseudomonas putida convert indene to cis-1S,2R-indandiol, a potentially important intermediate for the chemical synthesis of the HIV-1 protease inhibitor, Crixivan. A bioconversion process was developed through optimization of medium composition and reaction conditions at the shake-flask and 23-1 fermentor scales. A cis-1,2-indandiol productivity of approx. 1000 mg/l was achieved with construct TDO123, which represents a 50-fold increase over the initial titer. Varying the bioconversion conditions did not change the enantiomeric excess (e.e.) for the 1S,2R enantiomer from about 30%, suggesting that toluene dioxygenase intrinsically converts indene to 1S,2R- and 1R,2S-indandiols at a ratio of 2:1. Further inclusion of the Pseudomonas dehydrogenase gene in construct D160-1 led to the production of chirally pure cis-1S,2R-indandiol (e.e. > 99%) as a result of the selective degradation of the 1R,2S enantiomer, with the overall yield (650 mg/l) proportionally reduced. A single stage process was developed for D160-1 and scaled up to the 23-1 fermentor, achieving a cis-1S,2R-indandiol titer of 1200 mg/l.

Cloning, Molecular↗

PubMed1999

Eye and head coordination in reading: roles of head movement and cognitive control.

Simultaneous recording of eye and head movements during reading revealed that head movements consisted of two components: a modulatory-velocity component coupled to eye saccades, and a constant-velocity component that was independent of eye saccades. Whereas the constant-velocity component increased as subjects repeatedly read the same text, neither the magnitude of the modulatory-velocity component, nor the amplitude of the eye movement, increased. This outcome could be closely simulated when the head movement command was assumed to be stronger, and issued earlier with repeated reading. These results suggest that higher-level processes related to text familiarity modulate eye-head coordination through head movements.

Cognition↗

PubMed1999

Transforming growth factor-beta type II receptor confers tumor suppressor activity in murine renal carcinoma (Renca) cells.

OBJECTIVES: To demonstrate that the introduction of the transforming growth factor-beta (TGF-beta) type II receptor (TbetaR-II) decreases tumorigenicity in an aggressive murine renal carcinoma line, Renca. These cells do not express TbetaR-II. Because the presence of TbetaR-II in benign epithelial cells is ubiquitous, the ability to restore tumor suppressor activity in the Renca cell line with its introduction would elucidate the role of TbetaR-II as a tumor suppressor gene. METHODS: Renca cells were stably transfected with a retrovirus-mediated TbetaR-II expression vector. In vitro sensitivity to growth inhibitory effect of TGF-beta was assessed by the 3H-thymidine incorporation assay. For in vivo testing, xenograft tumors were produced by subcutaneous injection of tumor cells into immunodeficient nude mice. The tumorigenicity of these TbetaR-II transfected cells was tested. Wild-type Renca cells and cells transfected with the control vector were also tested for comparison. RESULTS: Expression of TbetaR-II mRNA was evident in Renca cells after transfection with the TbetaR-II construct. In vitro sensitivity to the growth inhibitory effect of TGF-beta was restored. This effect of TGF-beta was reversible with a neutralizing antibody specific for the extracellular domain of TbetaR-II. Xenografts grown from TbetaR-II transfected cells were significantly smaller, weighed less, and developed tumors later than those developed from wild-type Renca cells and those transfected with the control vector. CONCLUSIONS: We conclude that TbetaR-II is a central mediator of tumorigenicity in Renca cells. As with other tumor suppressor genes, the loss of TbetaR-II expression allows for the development of an aggressive phenotype.

Animals↗

PubMed1999

Determinants of the Permethrin Impregnated Bednets (PIB) in the Republic of Benin: the role of women in the acquisition and utilization of PIBs.

An important aspect of malaria control strategies has been the use of prophylactic measures such as impregnated bednets; however, adoption of this strategy has been slow and uneven. This study considers the factors determining Permethrin Impregnated Bednets (PIB) use in the context of a PIB promotion project in a rural area of Benin undertaken between 1992 and 1995. Quantitative data, on socio-demographic characteristics, malaria knowledge, attitudes, and practices, were gathered from 191 households of PIB users and non-users for comparative purposes using a questionnaire format. Qualitative data were collected from 23 focus group discussion sessions and 16 semi structured interviews. Women's income, men's educational level, and women's participation in communal organizations were the principal variables distinguishing user households from non-user households. Recourse to non-western medicine and, in particular, to medicinal teas which are considered preventive or curative correlates negatively with PIB use. The qualitative data shows that informants consider exposure to the sun, especially while engaged in agriculture work, a principal cause of malaria, and that PIB adoption is not considered justified in a context where there is a quasi chronic shortage of financial resources and where confidence in the efficacy of non-western medicine prevails. Because they have primary responsibility for the health of their families and are more aware of children's vulnerability to malaria, women are more inclined than men to want to buy PIBs. However, because the household head, who is most often male, sets family consumption priorities using family income, women often have to resort to using their own income, which is often considerably lower than that of men, to buy PIBs. Support for community initiatives which are directed to women's work, linked with intensive effort to sensitize men to the mechanisms of malaria transmission and the principal groups at risk, is seen as a means to increase PIB acquisition and use.

Bedding and Linens↗

PubMed1999

Protective effects of the flavonoid mixture, silymarin, on fetal rat brain and liver.

We investigated the possibility that the flavonoid mixture, silymarin (SY), administered as the compound Silymarin Phytosome (PHYTO), could protect the fetus from maternally ingested EtOH. Seventy-six female rats were randomly assigned to one of seven groups: pair-fed control; chow fed control; EtOH; and four groups receiving EtOH and PHYTO in varying dosages. All groups except the chow-fed control were maintained on a liquid diet. On day 1 of pregnancy the dams began the treatment protocol. On day 21 of pregnancy the rats were sacrificed and the fetuses removed. Gamma glutamyl transpeptidase (GGTP) activity was determined for liver and brain tissue from both the fetuses and the dams. GGTP activity in the EtOH/silymarin treatment groups did not differ significantly from that observed for the pair-fed control group. The observed GGTP activity levels for the EtOH-only group were significantly higher than those attained by the pair-fed control group. Although GGTP activity did not vary significantly with the quantity of PHYTO administered, as PHYTO dose was increased, GGTP activity decreased.

Animals↗

PubMed1999

Leptin differentially regulates NPY and POMC neurons projecting to the lateral hypothalamic area.

Recent studies have reinforced the view that the lateral hypothalamic area (LHA) regulates food intake and body weight. We identified leptin-sensitive neurons in the arcuate nucleus of the hypothalamus (Arc) that innervate the LHA using retrograde tracing with leptin administration. We found that retrogradely labeled cells in the Arc contained neuropeptide Y (NPY) mRNA or proopiomelanocortin (POMC) mRNA. Following leptin administration, NPY cells in the Arc did not express Fos but expressed suppressor of cytokine signaling-3 (SOCS-3) mRNA. In contrast, leptin induced both Fos and SOCS-3 expression in POMC neurons, many of which also innervated the LHA. These findings suggest that leptin directly and differentially engages NPY and POMC neurons that project to the LHA, linking circulating leptin and neurons that regulate feeding behavior and body weight homeostasis.

Animals↗

PubMed1999

Conservation of human gamma-X centromeric satellite DNA among primates with an autosomal localization in certain Old World monkeys.

Gamma-X satellite DNA is a 220-bp tandemly arranged repetitive DNA with specificity for the centromeric region of the human X chromosome. The conservation of this human X centromeric satellite DNA sequence in primate species was evaluated by comparative fluorescence in-situ hybridization to metaphase chromosome preparations of the great apes and three Old World monkeys. Homologous gamma-X DNA were detected at centromeric locations in all six primate species. For the great apes, gamma-X was exclusively localized to the centromeric regions of the X chromosomes. Among the Old World monkeys studied, only the golden monkey exhibited localization to the X chromosome. In the black-and-white colobus and the pig-tailed macaque, human gamma-X sequences were localized to the pericentromeric regions of autosomes 1 and 4, respectively.

Animals↗

PubMed1999

Pharmacology of G-1-64, a new nondepolarizing neuromuscular blocking agent with rapid onset and short duration of action.

BACKGROUND: Chances are slim that a clinically useful ultra-short-acting neuromuscular blocking agent of rapid onset will emerge from the benzylisoquinolinium or the aminosteroid series to which all currently popular relaxants belong. G-1-64 is a promising prototype of a new series of bis-quaternary ammonium salt of bistropinyl diester derivatives we have synthesized and studied in the laboratory. METHODS: Neuromuscular block (NMB) and autonomic and cardiovascular side effects were studied on appropriate preparations of anesthetized rats, rabbits, cats, ferrets, pigs and monkeys. Neuromuscular blocking characteristics, cumulativeness, and pharmacological reversibility were determined. Cardiac vagal block was evaluated by the inhibition of the bradycardic response to stimulation of the vagus in the cat, rat, ferret, and pig. Sympathetic ganglion block was evaluated on the cat's superior cervical ganglion/nictitating membrane preparation. Arterial blood pressure and heart rate were determined in all species. RESULTS: G-1-64 produced nondepolarizing NMB with train-of-four (TOF) and tetanic fades, and reversible with anticholinesterases. Its ED50 ranged between 60 and 800 microg/kg in these species. It showed a significantly faster onset (0.9-2.1 min) and/or shorter duration of action (5-12 min) than either atracurium or mivacurium. It did not show cumulativeness on repeated doses or infusion. A varying degree of cardiac vagal block was present, dependent on the species at doses exceeding the ED80 for NMB. Cardiovascular changes, ganglion block or signs suggesting histamine release were absent. CONCLUSION: With favorable neuromuscular blocking characteristics and modest side effects, G-1-64 and similar derivatives may have clinical potential.

Androstanols↗

PubMed1999

Repression of the gene encoding the TGF-beta type II receptor is a major target of the EWS-FLI1 oncoprotein.

Chromosomal translocations resulting in the expression of chimaeric transcription factors are frequently observed in tumour cells, and have been suggested to be a common mechanism in human carcinogenesis. Ewing sarcoma and related peripheral primitive neuroectodermal tumours share recurrent translocations that fuse the gene EWSR1 (formerly EWS) from 22q-12 to FLI1 and genes encoding other ETS transcription factors (which bind DNA through the conserved ETS domain). It has been shown that transduction of the gene EWSR1-FLI1 (encoding EWS-FLI1 protein) can transform NIH3T3 cells, and that mutants containing a deletion in either the EWS domain or the DNA-binding domain in FLI1 lose this ability. This indicates that the EWS-FLI1 fusion protein may act as an aberrant transcription factor, but the exact mechanism of oncogenesis remains unknown. Because ETS transcription factors regulate expression of TGFBR2 (encoding the TGF-beta type II receptor, TGF-beta RII; Refs 9,14), a putative tumour suppressor gene, we hypothesized that TGFBR2 may be a target of the EWS-FLI1 fusion protein. We show here that Ewing sarcoma [corrected] (ES) cell lines with the EWSR1-FLI1 fusion have reduced TGF-beta sensitivity, and that fusion-positive ES cells and primary tumours both express low or undetectable levels of TGFBR2 mRNA and protein product. Co-transfection of FLI1 and the TGFBR2 promoter induces promoter activity, whereas EWSR1-FLI1 leads to suppression of TGFBR2 promoter activity and FLI1-induced promoter activity. Introduction of EWSR1-FLI1 into cells lacking the EWSR1-FLI1 fusion suppresses TGF-beta RII expression, whereas antisense to EWSR1-FLI1 in ES cell lines positive for this gene fusion restores TGF-beta RII expression. Furthermore, introduction of normal TGF-beta RII into ES cell lines restores TGF-beta sensitivity and blocks tumorigenicity. Our results implicate TGF-beta RII as a direct target of EWS-FLI1.

Animals↗

PubMed1999

Creation of a novel donor splice site in intron 1 of the factor VIII gene leads to activation of a 191 bp cryptic exon in two haemophilia A patients.

We have constructed a confidential U.K. database of haemophilia A mutations and pedigrees by characterizing the gene defect of one index patient in each U.K. family. Mutations were identified by screening all coding regions of the factor VIII (FVIII) mRNA, using solid-phase fluorescent chemical cleavage of mismatch and examining additional non-coding regions of the gene. Here we report two haemophilia A patients (UK 114 FVIII:C 2% and UK 243 FVIII:C < 1%) with an abnormal FVIII mRNA due to an A to G point mutation, 1.4 kb downstream from exon 1 in the FVIII gene. This mutation creates a new donor splice site in intron 1 and leads to insertion of a 191 bp novel exon in the mRNA. Haplotype analysis suggests that the mutation may have originated in a common ancestor of the two patients, who further illustrate how mRNA analysis allows higher efficiency of haemophilia A mutation detection, because their mutation would not have been identified by direct analysis of the factor VIII gene.

Base Sequence↗