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Biomedical subjects

R M Clayton

Publications and source records attributed to R M Clayton.

At least 19 recordsLinked to original sources
PubMed1992

Long-term effects of aluminium on the fetal mouse brain.

Potentially noxious substances may act as fetal teratogens at levels far lower than those required to produce detectable effects in adults, and behavioural teratogenicity may occur at levels lower than those which produce morphological teratogenesis. Aluminium (Al) is a potential neurotoxin in adults. Since pregnant women may be exposed to untoward levels of Al compounds under certain conditions, we have examined the long-term effects of treating the pregnant mouse with intraperitoneal or oral aluminium sulphate on brain biochemistry and behaviour of the offspring. The cholinergic system, as evaluated by the activity of choline acetyltransferase (ChAT), was affected differentially in different regions of the brain, and still showed significant effects in the adult. Differences between the intraperitoneal and oral series in the magnitude of effect seen in the regions of the brain probably reflect differences in the effective level of exposure. Growth rate and psychomotor maturation in the pre-weaning mouse were affected in the intraperitoneal series only, showing a marked post-natal maternal effect.

Administration, Oral↗
PubMed1992

Localization of delta-crystallin RNA during lens morphogenesis and differentiation in the normal and talpid3 chick embryo.

Embryonic lens fiber cell differentiation in the chick is marked by the accumulation of delta-crystallin protein. The levels of delta-crystallin RNA are shown here to rise dramatically in the cells of the posterior lens pit prior to their elongation and differentiation as lens fibers. This increase correlates with regional proximity to the underlying optic cup (future retina). This accumulation of delta-crystallin RNA during lens induction operates selectively on the delta 1-crystallin transcripts whereas delta 2-crystallin/argininsosuccinate lyase RNA is detectable at lower levels in all developing ocular tissues throughout this period. The talpid3 mutant forms a flat "bridge" of thickened placode-like cells in the head epithelium between the two lens placodes, and this bridge also accumulates delta 1-crystallin RNA, suggesting that the selective increase in delta 1-crystallin RNA levels over those of delta 2-crystallin represents an early event in cellular commitment to lens fiber differentiation in the chick. The significance of the sequence of temporal changes in inductive sources for lens fiber formation is discussed, and we propose that the role of the optic cup is to provide, bound to its extra-cellular material (ECM), a high local concentration of the same growth factors which act as fiber inducers in the older eye.

Animals↗
PubMed1991

Independent regulation of two coexpressed delta-crystallin genes in chick lens and nonlens tissues.

It is known that delta-crystallin is super-abundant in the early chick lens, but it is found at lower levels in certain other tissues. Ninety-nine percent of the lens delta-crystallin poly(A)+ RNA is from the delta 1-crystallin gene. We report here that the delta 1- and delta 2-crystallin genes are both transcribed in the chick lens and retina throughout embryonic development and that both RNAs are found in embryo adenohypophysis and epiphysis and in day-old posthatch chick tibiofemoral chondrocytes and striated muscle. delta 1-crystallin RNA is more abundant in lens tissues, while delta 2-crystallin RNA is more abundant in all nonlens tissues. However, delta 1-crystallin RNA is processed more efficiently than delta 2-crystallin RNA in all early embryonic tissues examined. A comparison of lens epithelium and fibers established that levels of delta 2-crystallin RNA are the same but those of delta 1-crystallin RNA are over 100-fold higher in fibers compared to epithelial cells. The evidence implies independent regulation both of transcription and of post-transcriptional events for these two genes.

Animals↗
PubMed1991

Evidence for the extralenticular expression of members of the beta-crystallin gene family in the chick and a comparison with delta-crystallin during differentiation and transdifferentiation.

The beta-crystallins are major water soluble proteins of vertebrate lens fibre cells and have previously been regarded as lens-specific proteins: however beta B2-and beta A3/A1-crystallin RNAs are transcribed and beta-crystallin polypeptides are detectable in the developing chick retina. The beta-crystallin RNA is transcribed in a subpopulation of retina cells and the number of transcribing cells and the level of beta-crystallin polypeptides increase during the differentiation of the retina. Several tissues express beta-crystallin polypeptides, but individual tissues are characterised by qualitative and quantitative differences in the beta- and delta-crystallin polypeptides expressed. The expression of beta-crystallins appears to be non-random as defined by tissue distribution, cellular localisation and ontogeny, implying a function for extralenticular beta-crystallins and a complex mechanism for the regulation of their expression.

Animals↗
PubMed1990

Age-related changes in the response of chick lens cells during long-term culture to insulin, cyclic AMP, retinoic acid and a bovine retinal extract.

We have reported that 1-day-old post-hatch chick lens epithelial cells lose the capacity for lentoid body formation and delta-crystallin expression during long-term serial subculture, although they continue to synthesize, but not to accumulate, alpha- and beta-crystallins, even in cells with a transformed phenotype. Here we present evidence that dedifferentiation may reflect an age-related change in the capacity for response to regulatory signals. We have tested the capacity of these cells in serial subcultures to respond to agencies which affect lens cell growth and differentiation in primary culture: retinoic acid (RA), insulin, cAMP and bovine retinal extract (BRE). Secondary cultures responded only to RA and BRE, by an increase in lentoid formation and by alpha- and beta-accumulation, while RA also restored delta-crystallin expression. Later cultures showed no such responses. The results suggest that the process of lens cell dedifferentiation may, at first, be reversible but later becomes irreversible, despite the continuing persistence of low levels of crystallin expression.

Animals↗
PubMed1989

Differential effects of prenatal exposure to phenobarbital on the behaviour and neurochemistry of CBA and C57BL/6J mice.

Pregnant C57BL/6J and CBA mice were administered 60 mg/kg phenobarbital intraperitoneally from days 10 to 16 of gestation. On day 18 of pregnancy half of the control and drug-treated mice were killed and the embryonic brains removed for cell cultures. The remaining mice were allowed to have their litter. After cross-fostering the mice were used for behavioural studies. Pups born to drug-treated CBA mice had birth-weights similar to controls, but their weights had fallen behind controls by day 18 after birth. They were slower at attaining mature responses in tests for sensory motor development and became progressively more hyperactive (three times more active at day 18) compared to controls. Drug-exposed C57 pups also had birth weights similar to controls. After cross-fostering, 19% of control and 31% of drug-exposed pups died, but the remaining drug-exposed pups showed no deficits in weight gain. In contrast to drug-treated CBA pups, drug-exposed C57 pups were slightly quicker in attaining mature responses in some tests. There was no difference in activity between them and their controls. In neurochemical analyses, uptake of neurotransmitters by cerebral cultures from CBA showed that uptake of GABA was increased by 5%, choline by 95%, dopamine 120%, serotonin 165% and noradrenaline by 160% in cultures from drug exposed embryos compared to controls. In cerebral cultures from C57, GABA uptake was reduced by 18%, choline 33%, dopamine 35% and noradrenaline by 25%. Only serotonin uptake was increased by 182% compared to controls. Differences between C57 and CBA were also apparent in the uptake of neurotransmitters by neuronal cultures from the mesencephalon.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗
PubMed1988

The influence of the genotype on the process of ageing of chick lens cells in vitro.

We reported previously that changes in crystallin expression in differentiating long-term primary cultures of lens cells from five different chick genotypes are similar to those which occur in vivo between hatching and the 8-week-old adult. These changes followed a similar program in all genotypes but occurred more rapidly in cells from the fast-growing than from the slow-growing genotypes. The present study examines ageing changes in lens cell populations from the same five genotypes, over a 4-6 month period, using long-term serial subcultures. The capacity for lentoid differentiation was progressively lost, but the rate of loss was inversely related to the intrinsic growth rate of the cells of these genotypes, occurring at the first passage in the slowest-growing strain, while fifth passage cells of the fastest-growing strain still retained some lentoid-forming capacity. The rate of loss of crystallin expression was also inversely related to the genetic growth rate, but the sequence of changes appears to be nonrandom, since it was broadly similar in all genotypes, starting with a preferential loss of delta-crystallin, as occurs in vivo; although alpha- and beta-crystallins were undetectable in late dedifferentiated cultures, the capacity of the cells for their synthesis was still present. Cultures from both fast-growing genotypes eventually showed senescence, but those from all three slow-growing genotypes underwent transformation. The major cell component in late cultures of all genotypes was actin.

Animals↗
PubMed1987

Diuretic drugs as risk factors in cataractogenesis.

The use of diuretic drugs, which previously has been found to be associated with the incidence of cataract, is further investigated to elucidate the nature of the association. Diuretic drugs with different modes of action are considered separately. The degree of association of each correlates with the mean plasma urea level which itself is associated with cataract. However, the association is with cataract in general rather than with any specific type of lens opacity.

Adrenergic beta-Antagonists↗
PubMed1986

Patterns of crystallin expression during differentiation in vitro of several chick genotypes with different effects on lens cell growth rate.

We have recently reported that chick lens cells during differentiation in long-term culture show a programme of change in crystallin expression which mimics events during lens development in vivo. The aim of the present work was to examine the stability of the programme by testing the response to genetic influences and exposure to a carcinogen. Five genetically distinct inbred strains of chick, differing in the intrinsic growth rates of lens epithelial cells in vitro, were used to study the effects of the rate of mitosis on crystallin expression, both during lens development and in long-term cell culture. The time of appearance of lentoids, their size and abundance and the rate of change in crystallin expression were all modified in a genotype-specific way, related to the rate of mitosis, but the programme of changes in crystallin expression was the same for all genotypes. Genetic differences were also found in the patterns of response to treatment of cultures during the logarithmic growth phase with a nitrosoguanidine compound known to affect cell differentiation in lens cultures and several other systems. The changes in crystallin expression and fibre differentiation were delayed, but cultures of the faster growing genotypes were least affected. With further culture, crystallin expression tended to recover to control values although levels of fibre differentiation and cell growth remained depressed. The results indicate that genetic differences in intrinsic growth rate moderate but do not change the programme of crystallin expression shown by lens epithelial cells in culture, and that this programme shows resistance to change.

Animals↗
PubMed1986

Alterations in crystallin gene expression during subculture of chick lens cells.

We report here on the changes in crystallin gene expression during serial subculture of lens epithelial cells derived from day-old post-hatch chicks. Total cellular RNA from mass cultures were analysed by in vitro cell-free translation and by RNA blot (Northern) hybridization using a cloned delta-crystallin cDNA. Our results indicate that following subculture, lens epithelial cells which still retain the capacity for lens fibre differentiation (lentoid body formation) show a selective loss of delta-crystallin synthesis, and that this is related to the loss of delta-crystallin mRNA. The data suggest that older epithelial-cell populations give rise to lentoid bodies which in terms of crystallin gene expression closely resemble the later-formed cortical fibres of the adult chick lens. Tertiary cultures had an accelerated growth rate, formed no lentoids, contained no translatable alpha- or delta-crystallin mRNAs but still contained translatable beta-crystallin mRNAs.

Animals↗
PubMed1986

Developmental changes in membrane protein expression by chick lens cells in vivo and in vitro and the detection of main intrinsic polypeptide (MIP).

We have compared the long-term developmental changes in water-insoluble protein expression by chick lens cells in vitro and in vivo. Crude membrane fractions were prepared by alkali treatment of the urea-insoluble protein fraction, and the proteins analysed by sodium dodecyl sulphate-polyacrylamide (SDS-PAGE) gel electrophoresis. The major component present in the urea-insoluble fraction of chick lens fibres, a 25,000 MW polypeptide (MIP-25K) was more abundant in adult (8 weeks) than day-old post-hatch chick lens fibre masses. MIP-25K was detected in differentiated but not predifferentiated lens cell cultures, and indirect immunofluorescence using anti-bovine MIP antiserum indicated that MIP-25K was localized in the lentoid bodies. Our findings indicate that the urea-insoluble protein profiles of long-term well-differentiated chick lens cell cultures are qualitatively very similar to the profiles of the lens fibres. The data also confirm that the expression of MIP-25K, rather than the expression of water-soluble crystallin protein, is a marker for lens cell differentiation, and confirm earlier reports, which have been disputed, that delta-crystallin (but not alpha-or beta-crystallin) is specifically associated with chick lens fibre membranes.

Aging↗
PubMed1985

Effects of anticonvulsant drugs on brain cultures from chick embryos: a comparison with cultures from embryos treated in ovo.

The effects of anticonvulsant drugs phenytoin, phenobarbitone, and sodium valproate on neurons and glia from embryonic chicken brain have been tested. These effects have been compared with those produced in neuronal and glial cultures established from embryos that were injected with the drugs in ovo. Choline acetyl transferase activity and accumulation of gamma-aminobutyric acid were measured in neuronal cultures, and carbonic anhydrase and 2,'3'-cyclic nucleotide 3' phosphohydrolase activities were measured in glial cultures. Similarities have been observed in the morphological and biochemical changes brought about by the in vitro and in vivo treatment with the drugs. The use of in vitro cell culture systems for screening drugs for potential behavioural teratogenic effects is discussed.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1985

Cellular heterogeneity in the expression of the delta-crystallin gene in non-lens tissue.

RNA transcripts of the delta-crystallin genes, which code for the major chicken lens protein, have been detected at low levels in many non-lens tissues. Here it is demonstrated by in situ hybridisation that these transcripts are concentrated at a high level in small, infrequent clusters of cells in many non-lens tissues. While the nuclei of these cells are very heavily labelled, there is only light labelling of the cytoplasm. The unlabelled cells surrounding the labelled clusters are of similar morphology and staining properties as the labelled cells, and all have the characteristic morphology of cells of the embryonic tissue used. With the exception of neural retina, it is not yet known whether the labelled clusters are found in specific locations in the tissues, or whether they arise at random.

Animals↗
PubMed1985

A comparison of the changing patterns of crystallin expression in vivo, in long-term primary cultures in vitro and in response to a carcinogen.

Changes in the differentiation of day-old chick lens epithelium in long-term primary culture conditions were investigated by sodium dodecyl sulphate-polyacrylamide electrophoresis, using integrating densitometry to assess the relative levels of accumulated crystallin and non-crystallin polypeptides and fluorography to assess their relative levels of synthesis. The main changes during the culture period included a relative decline in the proportion of actin and other non-crystallins, an initial increase in 48K delta-crystallin expression followed by a decline and a shift in beta-crystallin expression from a relative preponderance of the 24K and 23K polypeptides to a relative preponderance of the 24K and 22K polypeptides. At all stages the level of the 19K alpha-crystallin was higher than that of the 20K alpha-crystallin polypeptide. In general, the changes in the pattern of expression of these polypeptides in culture were similar to those observed in vivo in the post-hatch chick, suggesting an intrinsic programme of crystallin expression. The changes in gene expression were also tested indirectly by brief exposure of the cells in vitro to a carcinogen, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) which is known to produce, in some systems, effects related to the status of the cell at the time of treatment. The effects were found to depend on the stage of differentiation of the culture at the time of treatment. Treatment on day 1 of culture prevented later lentoid formation and severely reduced the expression of all crystallins with the exception of the 34K beta-crystallin polypeptide. Actin was the most abundant soluble cell component, and a proportion of the cells acquired a fibroblast-like morphology. Treatment with MNNG on day 7 led to a delay in lentoid formation and a differential reduction of the synthesis of crystallin polypeptides, whereas the treatment of already differentiated cultures on day 18 and to lesser extents on days 27, 45 and 55, respectively, led to an increase in crystallin synthesis relative to controls. These results suggest that this programme of crystallin gene expression becomes more resistant to change with increasing epithelial differentiation.

Animals↗
PubMed1985

The penetration of detergents into adult and infant eyes: possible hazards of additives to ophthalmic preparations.

In studies of surfactant penetration into the eye, radiolabelled detergent was found to penetrate the rabbit cornea, to accumulate readily in ocular tissues and to be released slowly. Repeated applications led to increasing binding of the detergent. Permeability, ocular and systemic uptake and retention were all greater in juvenile than in adult rabbits. Preliminary tests of the effects of surfactants on ocular cells in vitro using concentrations found in vivo, suggest that some low-level effects might occur, particularly in juveniles.

Age Factors↗
PubMed1985

The expression of chick alpha A2-crystallin RNA during lens development and transdifferentiation.

During development of the vertebrate lens, the lens epithelium undergoes a final stage of differentiation into lens fibre cells, during which the major lens proteins, the crystallins, are synthesised. Lentoids, comprising clusters of lens fibre cells can also be produced by transdifferentiation from certain non-lens tissues, including neural retina and pigmented retinal epithelium. We have isolated an alpha A2-crystallin genomic probe and used it to study the transcription and processing of alpha A2-crystallin mRNA both during lens development and transdifferentiation. We relate these results to earlier measurements in this laboratory of delta-crystallin transcription and alpha- and delta-crystallin protein synthesis, to compare the expression of these two sets of genes. Tissue specific differences in gene expression were found. delta-crystallin mRNA is transcribed before alpha A2-crystallin RNA in the lens, but after it in transdifferentiating neural retina and pigmented epithelia.

Animals↗
PubMed1985

Comparative studies of normal and genetically hyperplastic lens epithelia II: lectin-induced cell agglutination and 125I-lectin uptake by cells.

The reactions of lectins with dissociated and cultured epithelial cells from hyperplastic lenses of two unrelated chick strains, Hy-1 and Hy-2, characterised by hyperplasia of the lens epithelium, and lenses of a normal genotype (N) were investigated using four different lectins. Three methods of monitoring lectin-binding to cell surfaces were employed. Each of the four lectins used showed an individual pattern of reactivity to separated membrane components. Data obtained with the three labelling methods showed the same trend viz: increased agglutinability and high affinity for binding of all four lectins by Hy-1 and Hy-2 cells. These results suggest that Hy-1 and Hy-2 lens epithelial cells are characterised by alteration in their cell surfaces.

Agglutination↗