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Biomedical subjects

C Lau

Publications and source records attributed to C Lau.

At least 91 records · Page 5Linked to original sources
PubMed1993

Effects of 5-fluorouracil on embryonic rat palate in vitro: fusion in the absence of proliferation.

5-Fluorouracil (5-FU) inhibits the enzyme thymidylate synthetase (TS) which results in inhibition of DNA synthesis. 5-FU is teratogenic in many species, inducing cleft palate, limb, and tail defects. In the present study, gestation day (GD) 14 embryonic rat craniofacial explants were exposed to 5-FU in organ culture with increasing concentrations and durations of exposure. Palates exposed to 5-FU were morphologically abnormal and craniofacial shape, size, and palatal fusion pattern were affected with the severity of effects dependent on concentration and duration of exposure. Cleft palate was induced in vitro as opposing palates overlapped in a narrowed oral cavity. Palates exposed to higher levels of 5-FU were growth inhibited, but fused even though proliferation ceased and few cells were available to participate in elevation and fusion. This was demonstrated as a biphasic concentration-response profile for palatal fusion in which 0.05 to 0.15 micrograms 5-FU/ml produced decreasing rates of palatal fusion, while exposure to 0.15 to 3.0 micrograms/ml resulted in progressively increasing rates of fusion. The effects of 5-FU were detected biochemically as a reduction in TS activity which was concentration and time dependent during the first 12 hours, with a return to control levels by 24 hours. During the first day, 5-FU did not alter protein levels, but DNA levels significantly decreased at the high concentration, 2.0 micrograms/ml. After 5 days in culture, both DNA and protein decreased with increasing 5-FU concentration and duration of exposure. Also by the end of the culture period, 3H-TdR incorporation had decreased in a concentration dependent manner. It is concluded that progressive inhibition of proliferation and growth in organ culture results in two different morphological outcomes: cleft palate resulting from a narrowed oral cavity and increased incidence of anterior palatal fusion under conditions of strong growth reduction. This study demonstrates that elevation and fusion can occur in the absence of growth and proliferation. Based on these observations, severe inhibition of growth or proliferation would not necessarily be sufficient to induce cleft palate.

Animals↗
PubMed1993

Utility of the murine erythroleukemic cell (MELC) in assessing mechanisms of action of DNA-active developmental toxicants: application to 5-fluorouracil.

Murine erythroleukemic cells (MELC) exposed to 2'-deoxy-5-azacytidine (D-AZA) or to the active cyclophosphamide (CP) metabolites phosphoramide mustard (PAM) and 4-hydroxycyclophosphamide (OHCP) exhibit cell-cycle perturbations similar to those seen in limb bud nuclei of gestational day (GD) 10 CD-1 mouse embryos exposed in utero to D-AZA or CP, respectively. The similarities in response suggest MELC may be a useful model for determining mechanisms of action of DNA-active developmental toxicants. As such, we used the MELC model to investigate the mechanism of action of 5-fluorouracil (5-FU), an antimetabolite that induced in GD 14 rat fetuses an apparent S-phase accumulation in limb cells 8 hr after in utero exposure, but S-phase depletion in liver cells 24 hr postexposure. MELC timed-recovery and synchronization studies suggest that in proliferative tissues, 5-FU induces an early S-phase accumulation, followed by a synchronous, concentration-dependent delay in progression through the cell cycle. Consequently, it is the tissue-specific rate of delay, rather than different mechanisms of action, that results in apparent tissue-specific perturbations. Moreover, growth and cell-cycle data suggest that cells entering S phase (when TS activity is greatest) are the most sensitive to 5-FU toxicity. Assays of the TS activity of recovering MELC reveal that although the initial extent of TS inhibition does not appear to be concentration-dependent, the time to recovery is, suggesting that the rate of S-phase progression is closely associated with TS activity. Together, the induction of similar cell-cycle perturbations in embryonic/fetal tissues and MELC following exposure to CP (or CP metabolites), D-AZA, or 5-FU, as well as the adaptability of MELC to a variety of kinetic assays suggests that, for those developmental toxicants suspected of inducing cell-cycle perturbations in embryonic/fetal tissues, MELC may prove useful for elucidating mechanisms of action.

Animals↗
PubMed1993

Differential regulation of human progesterone receptor A and B form-mediated trans-activation by phosphorylation.

Hormone-dependent phosphorylation of progesterone receptors (PRs) plays a functional role in their transcriptional activity. However, hormone-independent phosphorylation has also been shown to modulate the chicken PR-mediated trans-activation in the presence of phosphorylating agents. The present study was designed to investigate the effects of protein kinase A- and protein kinase C-mediated signal transduction pathways on the regulation of the activity of the two forms of human PR (hPRA and hPRB). Similar to chicken PR, hPR was activated by 8-bromo-cAMP (8-Br-cAMP) in the absence of ligand, whereas 8-Br-cAMP synergized with the progestin agonist R5020 to amplify hPRA- and hPRB-mediated reporter activity. Interestingly, the effect of 8-Br-cAMP was much more pronounced on hPRA-induced trans-activation than on hPRB. This differential regulation by 8-Br-cAMP could also be mimicked by okadaic acid. Both mouse mammary tumor virus-thymidine kinase-chloramphenicol acetyl transferase and progesterone response element-thymidine kinase-chloramphenicol acetyl transferase showed a similar response to 8-Br-cAMP in the presence of R5020. Protein kinase C, on the other hand, did not discriminate between hPRA- and hPRB-mediated trans-activation. Unlike 8-Br-cAMP, phorbol 12-myristate 13-acetate did not cause marked ligand-independent trans-activation through either of the two receptor forms. RU486, an antagonist of progestin, preferentially blocked R5020-induced trans-activation compared to R5020 + 8-Br-cAMP synergism. As expected, H-89, a specific inhibitor of protein kinase A was more effective in inhibiting ligand-independent activity. Western analysis of transfected receptors suggested that 8-Br-cAMP and 8-Br-cAMP + R5020 but not R5020 alone down-regulated the level of hPRB in COS-1 cells. Only marginal modulation of hPRA levels was observed with R5020 treatment in the presence and absence of 8-Br-cAMP. These data suggest that R5020 and 8-Br-cAMP mediate PR-dependent transactivation through distinct pathways, and that phosphorylation can differentially regulate the activity of hPRA and hPRB forms, an observation which may be important for selective target gene activation in vivo by progestins.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1993

Effects of diabetes mellitus on lactation in the rat.

Diabetes mellitus adversely affects the process of lactation. Although insulin administration restores lactation, the manner by which it does so is unknown. The goals of this study were to determine which phase of lactation, milk synthesis/supply (MS), and/or milk release (MR) was affected. Inasmuch as insulin, corticosterone, and prolactin, among other hormones, are involved in milk synthesis in vitro, this study investigated their probable roles in the suppression of lactation in diabetes in vivo. Diabetes was induced in rats on Day 3 postpartum with streptozotocin (50-60 mg/kg, ip). Milk synthesis/supply and milk release were indirectly monitored using body weight gain of pups during a timed-feeding period on postnatal Days 8/9 and 13/14. Plasma corticosterone, insulin, C-peptide, and prolactin were measured by radioimmunoassay in control, diabetic, and insulin-replaced diabetic animals. Insulin replacement was provided by means of an osmotic minipump implanted subcutaneously in the nape of the neck. In addition, several parameters of maternal behavior were monitored in order to determine whether diabetes affected maternal behavior and, therefore, whether such changes played a role in the alterations of lactation observed during diabetes. Diabetes significantly suppressed MS and MR. However, the decrease in MR, which was restored after partial insulin replacement, was a reflection of the reduced MS. There were no significant differences between the parameters of maternal behavior monitored in control and diabetic animals. Blood glucose in diabetic dams was significantly increased over that of controls. Although the levels of plasma glucose in the insulin-replaced groups were significantly lower than those of their uncontrolled diabetic counterparts, they also remained higher than that of controls. Corticosterone was not significantly altered after induction of diabetes. Insulin and C-peptide levels were significantly reduced in the uncontrolled diabetics; however, insulin levels were corrected in the insulin-replaced groups. Serum levels of prolactin decreased in all diabetic groups and insulin failed to restore these levels to those of control animals. In conclusion, it appears that diabetes decreases lactation through a suppressive effect on MS rather than on MR, with insulin replacement correcting for such deficiency. In addition, despite the lactogenic importance of glucocorticoids, prolactin and insulin demonstrated in vitro, lactation in vivo can be corrected without returning the levels of all three hormones to normal. The importance of another factor(s), such as normoglycemia, as essential to the observed defects, needs to be investigated.

Animals↗
PubMed1993

Comparison of computer interface devices for persons with severe physical disabilities.

This research employed a descriptive case study design to compare subjects' performance using three computer input devices: the Tongue Touch Keypad, the HeadMaster, and the mouthstick. The sample consisted of four students with severe physical disabilities enrolled in an adaptive computer class at a community college. Components of performance examined were input speed, accuracy, and level of perceived exertion. Subjects' acceptance of each of the interface devices was also examined. Results showed input speed to be the fastest with the mouthstick, followed by the HeadMaster and then the Tongue Touch Keypad. Accuracy of input did not vary significantly. Three subjects rated the Tongue Touch Keypad as requiring the lowest rate of perceived exertion, followed by the HeadMaster and then the mouthstick. Overall performance did not necessarily affect subjects' acceptability ratings of the devices. Information from this study will assist therapists in evaluating the effectiveness and desirability of computer interface devices for patients.

Adolescent↗
PubMed1992

Quantitation of human cellular retinoic acid-binding protein II (CRABP-II) RNA from cultured human skin fibroblast cells and human skin biopsies treated with retinoic acid.

A polymerase chain reaction (PCR) method has been validated for the quantitation of retinoic acid (RA) induction of cellular retinoic acid-binding protein II (CRABP-II) RNA from cultured human skin fibroblasts and human skin biopsies. The method utilizes reverse transcription and PCR (RT-PCR) to compare cellular CRABP-II RNA with a known amount of added internal standard RNA generated from a modified CRABP-II cDNA containing a 42 bp deletion. Thus, after RT-PCR of cellular and standard CRABP-II RNA in the same tube, the resulting DNA bands could be distinguished by size on ethidium bromide-stained, nondenaturing polyacrylamide gel. Serial dilutions of cellular RNA were co-amplified with a fixed amount of internal standard CRABP-II RNA, and the ratio of intensities of the two DNA bands was determined by computerized image analysis of the gel photograph. A linear relationship was found between the logs of this ratio and the input RNA. Absolute quantitation of cellular CRABP-II RNA was determined from the 'equivalence point', the dilution at which band intensities from cellular and standard RNAs were identical. Using this quantitative assay, the amount of CRABP-II RNA in cultured fibroblasts was 24 attomoles per microgram total RNA. A 4.2-fold increase in CRABP-II RNA was seen following 24 hours treatment with 10(-6) M RA. CRABP-II RNA content in skin biopsies taken from 3 human subjects ranged from 16 to 25 attomole/micrograms RNA. Topical treatment with 0.1% RA cream resulted in induction ranging from 3.9- to 12-fold over vehicle treatment. The method described here offers a rapid, sensitive and quantitative assay of specific RNAs, and should be especially useful for the measurement of RNA levels from small solid-tissue biopsies.

Base Sequence↗
PubMed1992

A gradient in expression of the Escherichia coli heat-stable enterotoxin receptor exists along the villus-to-crypt axis of rat small intestine.

Binding of Escherichia coli heat-stable enterotoxin to its receptor is critical to the initiation of toxin-induced secretion and diarrheal disease; it is also likely, however, that this receptor binds an endogenous ligand. In order to characterize the expression of the heat-stable enterotoxin receptor in the small intestine, we isolated epithelial cells from villus tip to crypt in rat jejunum and ileum. Binding of radiolabeled toxin was maximal in the villus preparations and gradually decreased along the villus-to-crypt axis, paralleling the decline of sucrase activity. Northern blots of total RNA identified a single heat stable enterotoxin receptor transcript (3.8 kb), predominantly in the villus cell fractions. In situ hybridization demonstrated clear signal in the villus cells with no apparent signal in the crypt cells, lamina propria or muscularis. Expression of this receptor was greatest after enterocytes leave the proliferative cycle and enter villi. This pattern of gene and protein expression may reflect a role of this receptor in binding endogenous ligands which in turn may regulate intestinal ion flux along the villus-to-crypt axis.

Animals↗
PubMed1992

Effects of various stressors on milk release in the rat.

This study investigated the effect of four stimuli on milk release (MR), namely, sound, nociception, novelty, and restraint. The role of the ensuing adrenocortical response in the suppression of MR was also evaluated. Plasma corticosterone (CORT) levels were measured at 0 (basal), 15, 30, and 60 min during the suckling sessions to determine whether elevated CORT normally associated with stress could be inhibitory to MR. Compared to nonstressed lactators, dams exposed to sound demonstrated no suppression in MR, but a significant increase in plasma CORT. Pain did not alter milk yield and elevated CORT only at the end of the first hour of exposure. During novelty, MR was suppressed and again CORT was only elevated at the end of the sampling period. Restraint decreased milk yield and increased CORT. During novelty, MR appeared to be regulated by an adrenal factor, which remains to be identified. The peripheral opiates seem to be partially involved during restraint. In conclusion, not all types of aversive stimuli interfere with MR. Of those which do, different mechanisms seem to be implicated depending upon the nature of the stressor. Furthermore, reduced MR during stress is not a direct consequence of increased CORT.

Acoustic Stimulation↗
PubMed1992

Spermine dialdehyde, a novel ex vivo purging agent for both allogenic and autologous bone marrow transplantations.

The use of ex vivo purging agents has shown to be beneficial for both allogeneic and autologous bone marrow transplantations. We have shown previously that spermine dialdehyde (SDA), an oxidized product of spermine, when used in phosphate-buffered saline (PBS) preferentially inhibits T-cell proliferation while sparing myeloid cells. Lethally irradiated mice were rescued by reconstitution with SDA-treated allogeneic marrow and showed no sign of graft-vs-host disease (GVHD). In this paper, we show by HPLC analysis that SDA degraded rapidly in PBS but remained intact in saline. Administered in saline, SDA was more inhibitory on leukemic cell lines than normal myeloid or T-cells. Lethally irradiated mice receiving a syngeneic bone marrow leukemic cell mixture treated ex vivo with SDA in saline did not manifest leukemia. Thus, the preferential inhibitory effect of SDA and its degraded products in different buffers suggest that SDA could act as a novel purging agent for both allogeneic and autologous bone marrow transplantations.

Acrolein↗
PubMed1992

Behavioral development following daily episodes of mother-infant separation in the rat.

Use of dermal or inhalation routes of maternal exposure during the postnatal period in rodent developmental neurotoxicity evaluations would be most practical if dams could be separated from their pups during the exposure period. However, this procedure raises questions concerning the effects of mother-infant separation itself on neurotoxicity endpoints. In the present study, Sprague-Dawley rat pups were either maternally deprived in warm incubators for 6 hr each day (7:00 AM-1:00 PM) or left with their dams (control), from Postnatal Day 4-20 (PND4-20), and were tested on a range of endpoints commonly used in developmental neurotoxicology. These included motor activity (PND13, 17, 19, 21, 29, 60), olfactory learning (PND18) and retention (PND25), T-maze delayed alternation (PND23, 24), acoustic startle response (PND23, 62), and auditory thresholds (PND62). None of the behavioral measures were affected by daily separation. Apparently, interrupting the mother-infant interaction for 6 hr/day has little or no effect by itself on behavioral development, as assessed by these measures.

Aging↗
PubMed1992

General practitioners' use of non-steroidal anti-inflammatory drugs in Tayside and Fife regions.

The objectives of this study were to assess the prescribing of non-steroidal anti-inflammatory drugs (NSAIDs) by general practitioners and to determine their attitudes to problems caused by this class of drugs. The study consisted of two parts. The first was a questionnaire survey among general practitioners in Fife and Tayside, and the second was an analysis of NSAID prescribing over 12 months among the doctors in the Carnoustie Health Centre, using duplicate prescriptions. In the questionnaire survey 61% of the general practitioners responded. The three most preferred drugs were buprofen (56%), naproxen (20%) and mefenamic acid (7%); choice of drug was determined by efficacy and personal experience. Gastrointestinal side effects were most frequently encountered, although there was little consensus amongst respondents as to their management. The duplicate prescription study showed that 14% of patients (1607 individuals) received at least one NSAID prescription in the year of study. Ibuprofen (31%), naproxen (20%) and piroxicam (15%) were most frequently prescribed and up to 16% of the patients were co-prescribed a gastroprotective agent; ranitidine (75%) was the most commonly prescribed. Despite the introduction of newer NSAIDs, ibuprofen and naproxen are still the most commonly prescribed drugs. Furthermore, although gastrointestinal side effects are commonly encountered, there is some uncertainty about their management.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1992

Repeated maternal separation in the neonatal rat: cellular mechanisms contributing to brain growth sparing.

Separation of neonates from their dam has been shown to evoke acutely a variety of biochemical responses, typified by depression of ornithine decarboxylase (ODC) activity. In the current study where rat pups were removed from their nursing dams for 6 h, ODC activities in the liver, heart, kidney and lung were markedly suppressed, but the enzyme in the brain was not altered during the early postnatal ages. These data suggest that the brain was protected from maternal separation insults, a homeostatic response mediated in part, by an increase of circulating corticosterone and glycogen mobilization from peripheral tissues, particularly the liver. In addition, we examined whether these responses were extended to pups who were subject to repeated episodes of maternal deprivation, and whether this stress paradigm might be associated with corresponding changes of cellular growth and maturation. Pups were removed from their dams for 6 h daily beginning at 4 days of age until weaning at 21 days. Plasma corticosterone levels of the deprived pups were elevated significantly at the end of each stress episode but returned to basal (control) levels subsequently. The repeating stress paradigm did not influence the magnitude of this hormonal response at the ensuing ages. Consistent with findings observed in the single episodes of stress, ODC activities in the peripheral tissues were significantly depressed in pups subject to repeated maternal deprivation, but the enzyme appeared to recover to control levels 18 h after each insult. In contrast, brain ODC activity did not exhibit any change throughout the period examined. Moreover, while ontogenetic gains of DNA and protein in the peripheral tissues of the deprived rats lagged slightly but consistently behind those of controls, these macromolecules in the brain were not affected appreciably. These results thus suggest that brain growth was, by and large, spared from insults associated with repeated maternal separation; but this stressful paradigm did produce marked, though reversible biochemical and physiological responses in the peripheral tissues of neonates, which cumulatively led to a lag of cellular development.

Animals↗
PubMed1992

Expression of sucrase-isomaltase mRNA along the villus-crypt axis in the rat small intestine.

Sucrase-isomaltase has been used as a marker enzyme to study cell differentiation along the intestinal villus-crypt axis. Previous studies are in agreement that sucrase activity is confined to villus epithelial cells. However, immunoreactivity data are at conflict, with some studies reporting sucrase antigen in crypts as well as villi. To resolve this discrepancy, our goal was to determine the distribution of sucrase-isomaltase mRNA. A cDNA clone representing 3.0 kb of rat sucrase-isomaltase, including the sucrase active site, was characterized. Northern analysis of 12 tissues demonstrated a 6 kb transcript only in the small intestine. Jejunal cell fractions prepared by a washing technique showed declining levels of both sucrase activity and sucrase-isomaltase mRNA as well as increasing levels of thymidine kinase activity from early to later fractions. Since later fractions did not yield pure crypt cells, in situ hybridization using an 35S-labeled sucrase-isomaltase riboprobe was performed. The transition from zero to intense signal at the crypt-villus junction leads us to conclude that in the adult rat, sucrase-isomaltase gene expression is initiated only after cells leave the proliferative cycle and migrate onto the villi.

Amino Acid Sequence↗
PubMed1990

Trophic control of the ornithine decarboxylase/polyamine system in neonatal rat brain regions: lesions caused by 6-hydroxydopamine produce effects selective for cerebellum.

Norepinephrine has been hypothesized as a trophic factor influencing postnatal development of the cerebellum. In the current study, neonatal rats were given 6-hydroxydopamine (6-OHDA) to destroy noradrenergic projections and the effects on the ornithine decarboxylase (ODC)/polyamine system were evaluated; ODC initiates the synthesis of polyamines, which are known to control cellular development in the cerebellum, and neonatal ODC activity is regulated in part by beta 2-adrenergic receptors. Intracisternal administration of 6-OHDA resulted in complete and permanent depletion of cerebellar norepinephrine and a deficit in ODC, polyamine levels and cerebellar growth. Subcutaneous administration of 6-OHDA, which caused only a small initial reduction in cerebellar norepinephrine, did not affect ODC and had only minor effects on tissue growth. Indeed, levels of the polyamines tended to be elevated after subcutaneous 6-OHDA, associated with postweaning elevations in norepinephrine, results which are probably indicative of axonal regeneration. In contrast to the effects of 6-OHDA on cerebellar development, neither the intracisternal nor subcutaneous drug treatment had any effect on cerebral cortical polyamines or growth, although the intracisternal treatment did impair ODC activity early in development. These data suggest that postnatal noradrenergic input, acting through the ODC/polyamine pathway, plays a selective role in cerebellar development.

Animals↗
PubMed1990

Purification, characterization, and structural elucidation of the active moiety of the previously called "suppressor activating factor (SAF)".

Upon extensive purification of the serum-free supernatant produced by a mutant T cell line (6T-CEM), an immunosuppressive activity was found to reside in an oxidized product of spermine, spermine dialdehyde (SDA). The activity was purified to homogeneity from a serum-free supernatant by using gel filtration chromatography and reverse-phase C18 HPLC. Fast Atom Bombardment (FAB) mass spectral analysis revealed its MW to be 202 and Electron Impact (EI) analysis of the acetylated material identified the purified molecule to be spermine. In the presence of human or rodent plasma, spermine exhibited no immunosuppressive activity up to 2 mg/ml. However, when assayed in the presence of FCS, which contains polyamine oxidase (PAO), spermine is oxidized to its corresponding dialdehyde which is active at 0.1 microM/ml. We have previously described a high molecular weight suppressor activating factor (SAF) found in the serum-containing supernatant of the 6T-CEM cell line. Our preliminary biological data suggest that SDA is probably responsible for the immunosuppressive activities previously observed for the SAF. The strong affinity of SDA for proteins and thiocompounds may account for the apparent high MW previously reported for SAF.

Aldehydes↗
PubMed1990

Ex vivo spermine dialdehyde treatment prevents lethal GVHD in a murine bone marrow transplantation model.

Spermine dialdehyde (SDA), an oxidized product of spermine which irreversibly suppresses T cell and NK cell activities, was evaluated as an ex vivo purging agent in the prevention of graft-versus-host disease (GVHD) in mice transplanted with SDA-treated allogeneic bone marrow. In this model, lethally irradiated C3H (H-2k) mice received BALB/c (H-2d) bone marrow and spleen cell mixtures which had been treated ex vivo for 10 min with SDA. Mice receiving SDA-treated cells survived past 100 days whereas mice in the control group died between days 25-35 suffering from severe GVHD. Surviving mice from the SDA-treated groups exhibited full chimerism at day 120. In vitro assays indicated that SDA inhibited T cell and NK cell activities at concentrations that spared myeloid cell growth. When a minimum number of bone marrow cells were used for reconstitution, SDA-treated marrow reconstituted lethally irradiated mice as effectively as control marrow suggesting that SDA had little impact on early myeloid cells which are required for engraftment. SDA may have clinical application as a purging agent in allogeneic bone marrow transplantation.

Animals↗
PubMed1989

Suppression of lymphocyte proliferation by hexamethylene diamine.

The antiproliferative potential of hexamethylene diamine (HMDA) for mitogen-stimulated splenic lymphocytes was evaluated in vitro at final concentrations of 0.1-16 mM. Addition at the start of culture or after 24 or 48 h of culture decreased the proliferative response to T and B cell mitogens. However, the concentration of HMDA required to cause suppression increased with incubation time. Removal of diamine after 24 h allowed cells to proliferate normally upon reculture with mitogen. Mitogenic responses of cultures containing the potent ornithine decarboxylase (ODC) inhibitor alpha-difluoromethylornithine (DFMO) were also inhibited in a time and dose dependent fashion. ODC activity, which was much greater in cultures stimulated with Con A than LPS, was markedly decreased by inclusion of diamine or DFMO in the culture medium. Addition of putrescine to cultures did not reverse the suppressive effects of diamine on proliferation but did restore DFMO-containing cultures to control levels of activity. These results indicate that HMDA does suppress lymphocyte proliferation in vitro by alteration of ODC and polyamine activity. However, comparison of results obtained with DFMO and HMDA suggests that HMDA may act via multiple pathways, only one of which involves inhibition of ODC activity.

Animals↗