Biomedical subjects

G M Roomans

Publications and source records attributed to G M Roomans.

At least 19 recordsLinked to original sources

PubMed1992

Ion transport in colon cancer cell cultures studied by X-ray microanalysis.

Three colon cancer cell lines (Colo 205, HT29 and T84) were investigated by X-ray microanalysis with respect to elemental composition and the effect of cAMP on the cellular concentrations of Na, K, and Cl. The cultures were not homogeneous with respect to their elemental composition, but appeared to consist of two sub-groups, low-K cells and high-K cells. In all three cell lines, the low-K cells had, in addition, higher Ca, markedly lower Cl, and somewhat lower P and S concentrations. Differences in Na and Mg concentrations were absent or not consistent. Exposure of cells to cAMP caused a decrease of the cellular Cl and K content in high-K (high-Cl) cells. Changes in Na were not significant. No difference between the three cell lines could be noted. Incubation of the cells with phorbol myristate acetate (PMA), which has been shown to down-regulate the expression of the cystic fibrosis (CF) transmembrane conductance regulator gene and thus confer CF-like characteristics on the cells, significantly decreased the response in the cellular Cl concentration to cAMP stimulation. It is concluded that cAMP initially activates predominantly the apical Cl- channel and the basolateral K+ channel.

Biological Transport↗

PubMed1992

X-ray micro-analysis of cultured respiratory epithelial cells from patients with cystic fibrosis.

X-ray micro-analysis was carried out on cultured respiratory cells from polyps removed from individuals with and without cystic fibrosis (CF). In a first set of experiments, proper experimental conditions were established. Washing the cells with 300 mmol l-1 mannitol in distilled water was found to give the best removal of the culture medium. The elemental concentrations stabilized in about 10 min after the start of the preincubation. Intracellular [Na] and [Cl] increased slightly with increasing passage number, whereas intracellular [K] decreased. Under resting conditions there were no significant differences in elemental content between CF and control cells, and there were no indications for abnormally high total [Ca] in CF cells. In normal cells, stimulation with a cAMP-analogue resulted in a decrease of cellular [Cl], whereas in CF cells an increase was measured. Exposure of both normal and CF cells to ouabain resulted in decreased [K] and increased [Na] and [Cl] level. The calcium ionophore A23187 had a similar effect on normal cells but did not affect CF cells markedly. Application of amiloride to the apical side of the cells resulted in a decrease of cellular [Na] in CF cells, whereas [Na] in control cells was not affected. The results correspond with what is known about the defective cAMP-regulated transepithelial Cl-transport in CF cells. The effect of the calcium ionophore on cellular electrolyte content is more complicated and may be the result of two separate effects: efflux of Cl- via a Ca(2+)-dependent mechanism and inhibition of the Na(+)-K(+)-ATPase by intracellular Ca2+ ions causing an influx of Na+ and Cl- ions.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1992

Elemental changes in guinea pig epidermis at repeated exposure to sodium lauryl sulfate.

Epidermal hyperplasia is the response of the epidermis to external harmful stimuli. The control and regulation of this hyperplasia is not completely understood. It has been proposed that changes in the cellular sodium/potassium ratio are of importance in the regulation of cell proliferation. To evaluate if such a change in the elemental content of epidermal cells can be one factor to consider at irritant contact dermatitis, we performed a quantitative assessment of sodium lauryl sulfate (SLS)-induced contact reactions in the guinea pig. SLS was applied 1, 2 or 3 times and biopsies were obtained at 24 and 84 h after the last application. It was found that repeated exposures to SLS induced a hyperplasia of epidermis at 24 h persisting at 84 h. At 24 h there were significant changes in the sodium and potassium content of the keratinocytes. At 84 h there was still an increased potassium level in the cells and the sodium/potassium ratio was significantly decreased in epidermis exposed three times to SLS. This implies that changes in cellular sodium/potassium ratios occur in epidermal hyperplasia following irritant stimuli.

Animals↗

PubMed1992

Effect of chronic treatment with diuretics on mouse liver: a morphological and microanalytical investigation.

In an attempt to produce an animal model for the disease cystic fibrosis (CF), mice were treated chronically with the diuretics amiloride and furosemide, in order to cause chronic inhibition of transepithelial ion transport. Experiments were carried out on adult mice (2 months treatment); in addition, pregnant mice were treated with diuretics, and tissue from offspring 2 and 7 days post partum was investigated. Since biliary cirrhosis is a common occurrence in CF, hepatocytes in the treated mice were investigated by X-ray microanalysis and by light and electron microscopy. Treatment with amiloride caused a significant decrease in cellular Na concentration in adult animals and in in utero treated mice 2 days after birth. The decrease in Na was paralleled by a decrease in Cl, but K levels were not affected. Furosemide caused a slight increase of cellular Na concentrations, especially in animals aged 7 days. In the adult animals, both amiloride and furosemide caused a significant decrease of the cellular Na and Cl levels. No signs of cirrhosis could be observed. Inconsistent changes in the accumulation of lipid droplets in hepatocytes of adult animals treated with amiloride were observed by electron microscopy. It can be concluded that chronic treatment with diuretics, even though it causes some, possibly pathological, changes of the liver, is only of very limited value for generating an animal model to study liver disease in CF.

Amiloride↗

PubMed1992

Correction for extraneous background in X-ray microanalysis of cell cultures.

Some practical aspects of the X-ray microanalysis of cell cultures have been investigated. Cells were cultured on titanium grids covered with Formvar films and analyzed at 100 kV either in the scanning transmission (STEM) or transmission mode (TEM) of the electron microscope. Different holders, grids and configurations were compared with respect to the relative contribution of different factors to the extraneous background in the X-ray spectrum. When low atomic number holders are used, the contribution to the spectrum of electrons scattered through high angles, may be negligible. In practice this may result in negative values for the contribution of these scattered electrons to the background. Computer programs for correction of the extraneous background should ignore these negative values and replace them by zero. When a brass holder is used, the contribution to the spectrum from electrons scattered through high angles becomes more important than that of the uncollimated radiation. The position of the analyzed cell relative to the grid bars is more important than the choice of grid or holder type. The data show that for the specimens used in the present study the correction for extraneous background is of little importance and can be neglected.

Brain Neoplasms↗

PubMed1992

Effects of chronic and in utero treatment with diuretics on mouse exocrine cells studied by X-ray microanalysis.

In an attempt to develop an animal model for the disease cystic fibrosis, mice were chronically treated with diuretics. In addition, pregnant mice were treated with diuretics and the effect of this treatment in utero on the newborn mice was studied. Pancreas and submandibular gland acinar cells were investigated by X-ray microanalysis and transmission electron microscopy. Long-term treatment with furosemide (up to 13 months) caused transient changes in the elemental content of the pancreatic acinar cells: a decrease in chloride and sulfur, and an increase in phosphorus, potassium and magnesium. All changes normalized with prolonged treatment. Some morphological changes were found in the zymogen granules. Treatment with amiloride or furosemide in utero caused a decrease in cellular sodium and chloride levels, indicative of inhibition of transepithelial ion and fluid transport. Also treatment of adult animals for two months with amiloride caused lower intracellular sodium and chloride levels. In adult animals only minor effects of diuretic treatment on submandibular gland acinar cells were noted. In utero treatment with amiloride caused an increase in sodium and chloride content indicative of cell damage.

Amiloride↗

PubMed1991

Water and ion distributions in myocytes cultured under oxidative stress mimic changes found in the process of aging.

Element concentrations and local water content were measured in cytoplasm, nuclei, mitochondria and lipofuscin granula (LG) of isolated cardiac myocytes of the rat. Cells were cultured for 14 days under either 5%, 20% or 40% ambient oxygen concentration. LG were found to contain less cations and more phosphorous than mitochondria, which might be related to their lower protein and higher lipid content. Under oxidative stress, dehydration of mitochondria occurred, while their cation content remained constant. This is the same pattern of changes as found in cells in situ of aging rats. Therefore, it is concluded that peroxidative damage via oxygen derived free radicals is the reason for the mitochondrial water loss in the process of aging.

Aging↗

PubMed1991

X-ray microanalysis of cAMP-induced ion transport in cystic fibrosis fibroblasts.

cAMP-induced ion transport in normal and cystic fibrosis (CF) fibroblasts was investigated by X-ray microanalysis. Stimulation with cAMP causes an increase in cellular Na content and a decrease in cellular Cl and K content. No significant difference in response between CF and normal cells was noted. In this respect, fibroblasts differ from epithelial cells, where cAMP-induced Cl- efflux blocked in CF patients. Isoproterenol produced similar changes in Na and K content as cAMP, but did not effect Cl content.

Cells, Cultured↗

PubMed1991

Cryopreparation of tissue for clinical applications of X-ray microanalysis.

A number of diseases is associated with changes in ion and/or water distribution at the tissue or cell level, and X-ray microanalysis can be used to investigate the relationship between the disease process and the changes in elemental distribution. For analysis of diffusible elements by X-ray microanalysis, the tissue has to be prepared by cryotechniques. To carry out studies of this kind in a clinical environment poses a number of problems. Some of these problems occur already before the tissue is frozen, others are caused by the practical and ethical limitations that are imposed on the freezing method itself when human tissue is to be used. The use of cryostat sections for analysis at the cellular level, and of in vitro systems and cell cultures in which sampling and cryopreparation can be separated in time and place can be useful alternatives.

Animals↗

PubMed1990

Plasma calcitonin levels in patients with cystic fibrosis.

Plasma levels of calcitonin were investigated in patients with cystic fibrosis and compared with age- and sex-matched healthy controls. About forty per cent of the cystic fibrosis patients had plasma calcitonin levels that were markedly higher than normal. Elevated levels were found predominantly in female patients. In general, patients with marked pulmonary involvement had high plasma calcitonin levels. Immunoextraction and gel chromatography, carried out on plasma of one of the patients demonstrated monomer-like forms of calcitonin similar to those found in the healthy subjects.

Adolescent↗

PubMed1990

Changes in rat and mouse salivary glands and pancreas after chronic treatment with diuretics: a potential animal model for cystic fibrosis.

Defective transepithelial chloride and water transport is thought to be the cellular basis of the disease cystic fibrosis (CF). Therefore, it was attempted to develop an animal model for this disease by chronically inhibiting transepithelial chloride transport in experimental animals by long term treatment with high doses of diuretics. In the present study, changes in the salivary glands and pancreas after such treatment were investigated by X-ray microanalysis and electron microscopy. Treatment of rats for one month with diuretics caused a significant decrease in chloride and an increase in calcium in the acinar cells of the submandibular gland. This increase was due to accumulation of mucus in the cells. The strongest effect was obtained after combined treatment with furosemide and acetazolamide. Only minor changes were noted in the parotid gland and the pancreas. Treatment of mice for three months with diuretics caused similar changes in the submandibular glands. In addition, marked changes in the pancreas were observed. The chloride content of the pancreatic acinar cells was decreased. In many acinar cells, only very few zymogen granules were present. The morphological and microanalytical results point to severe dysfunction of the exocrine pancreas. These changes parallel those found in patients with CF, and the chronically furosemide-treated mouse thus could serve as an animal model for this disease.

Acetazolamide↗

PubMed1990

Essential fatty acid deficient rats in the study of cystic fibrosis: an X-ray microanalytical and ultrastructural study in chronically reserpinized rats.

Essential fatty acid (EFA) deficiency has been proposed as a major pathogenic mechanism for cystic fibrosis (CF) and EFA-deficient animals have been proposed as an animal model for CF. In the present study, the elemental composition and ultrastructure of the acinar cells of the submandibular and parotid gland of the pancreas of EFA-deficient rats were investigated by X-ray microanalysis and electron microscopy. The effects of EFA-deficiency were compared to changes in these exocrine glands in chronically reserpine-treated rats, an established animal model for CF. EFA-deficiency did not cause any significant changes in the elemental composition of the acinar cells of the submandibular or parotid gland, or of the pancreas. The changes in elemental composition induced by reserpine treatment were only slightly modified by EFA-deficiency, mainly towards normalization. EFA-deficiency resulted in the presence of abnormal, electron translucent, zymogen granules in the parotid gland and in a reduction of the number of zymogen granules in pancreatic acinar cells. Since EFA-deficiency in rats only causes minor changes in structure and elemental composition of salivary glands and pancreas, and does not potentiate the effect of chronic reserpine treatment on these tissues, it is concluded that EFA-deficiency is likely to be of minor importance in the exocrine gland disturbances in CF.

Animals↗

PubMed1989

Mechanism of action of calcitonin on secretion in rat submandibular gland.

Calcitonin (CT) was found to reduce the initial flow of pilocarpine-stimulated saliva from the submandibular glands in the rat. Although there was a concomitant increase of the concentration of calcium and protein in the saliva, the calcium/protein ratio was not significantly affected. CT also caused a significant increase of the potassium concentration in submandibular saliva. Both in vivo and in vitro, CT inhibited the production of cyclic AMP (cAMP) both in the absence and in the presence of forskolin. This decrease in intracellular cAMP levels could result in an inhibition of mucus secretion, which would explain the previously observed calcitonin-induced intracellular accumulation of mucus in the submandibular gland acinar cells. CT did not affect the cytoplasmic free Ca2+ concentration (as measured with fura 2) in isolated submandibular acini either in the absence or in the presence of cholinergic or adrenergic agonists. These results indicate that the inhibition of fluid secretion in the submandibular gland by calcitonin must be located distal to changes in [Ca2+]i. It can be concluded that CT affects both mucus and fluid secretion in the submandibular gland, but that only the inhibition of mucus secretion can as yet be explained by an effect at the level of the second messenger.

Adenylyl Cyclases↗

PubMed1989

Effects of thyroxine and dexamethasone on rat submandibular glands.

Glucocorticoids and thyroxine are known to have a marked effect on the flow rate and protein composition of rat parotid saliva in hormonally intact animals. In the present study, the effects of a one-week treatment of male rats with dexamethasone and thyroxine were studied by electron microscopy and x-ray micro-analysis, and by measurement of the flow rate and determination of the chemical composition of pilocarpine-induced submandibular saliva. Thyroxine had the most extensive effects on the submandibular gland. The acinar cells were enlarged and filled with mucus; the cellular calcium concentration was significantly increased. The flow rate of the submandibular saliva was significantly reduced compared with that in saline-injected control animals. Thyroxine caused an increase in the concentrations of protein, total calcium, and potassium in the saliva. Dexamethasone had no significant effects on gland ultrastructure or on the elemental composition of the acinar cells; flow rate was not affected, but the concentrations of protein, calcium, and potassium were significantly increased. The effects of dexamethasone and thyroxine on the flow rate and protein composition of pilocarpine-induced rat submandibular saliva differ from those reported earlier for rat parotid saliva after simultaneous stimulation with pilocarpine and isoproterenol.

Analysis of Variance↗

PubMed1989

Strontium and bromide as tracers in X-ray microanalysis of biological tissue.

Since energy dispersive X-ray microanalysis cannot distinguish between isotopes of the same element, alternative methods have to be used to get information similar to that obtained in experiments with radioactive tracers. In the present study, strontium was used as a tracer for calcium, and bromide as a tracer for chloride. Rats were injected with strontium chloride in vivo, and the uptake of strontium in the acinar cells of the submandibular gland was studied. Eventually a Sr/Ca ratio of 0.3 was reached. In some animals, secretion of mucus had been elicited by stimulation with isoproterenol 4 h prior to injection of strontium chloride. Exchange of calcium for strontium was enhanced by prior injection with isoproterenol. In a second experiment, rats were injected with sodium bromide, and the uptake of bromide by the submandibular acinar cells was followed in time, both in pilocarpine-stimulated and unstimulated glands. Under the experimental conditions, bromide was rapidly taken up by the cells, and the cellular Br/Cl ratio was close to that found in serum. Submandibular glands take up Br somewhat faster than other tissues (liver, heart muscle, skeletal muscle). The uptake of Br in pancreatic acinar cells was studied in vitro. These experiments showed a 1:1 ratio (molar) exchange of Cl for Br.

Animals↗

PubMed1989

X-ray microanalysis of mammalian salivary glands.

X-ray microanalysis was used to study the elemental composition of major salivary glands in rat and man. The elemental composition of the acinar cells is influenced by a variety of factors. Stimulation of fluid secretion by cholinergic, alpha-adrenergic or peptidergic agonists leads to a sustained increase of the Na/K ratio. The elemental composition and ultrastructure of the acinar cells of rat submandibular gland, as well as flow rate and composition of pilocarpinestimulated submandibular saliva are significantly affected by thyroxine, but less by the synthetic glucocorticoid dexamethasone. Hypercalcemia (induced by chronic vitamin D treatment) as well as hypocalcemia (induced by calcitonin) cause accumulation of intracellular mucus in the submandibular acinar cells, with concurrent increase in the cellular calcium concentration, and a significantly decreased flow rate after pilocarpine stimulation. Several animal models have been proposed for the generalized exocrinopathy cystic fibrosis (CF), which may be caused by a defective regulation of chloride transport in epithelial cells. A potential new model is proposed: the chronically furosemide-treated rat, which shows several abnormalities in salivary gland structure and function that parallel the abnormalities found in the human disease. Another experimental model of potential interest in the study of CF is short-term treatment of experimental animals with the adenylate-cyclase inhibitor alloxan.

Animals↗

PubMed1989

Sodium lauryl sulfate enhances nickel penetration through guinea-pig skin. Studies with energy dispersive X-ray microanalysis.

The effect of sodium lauryl sulphate (SLS), a common ingredient of detergents, on the penetration of nickel through the stratum corneum in the guinea-pig skin model was studied with energy dispersive X-ray microanalysis (EDX) to evaluate the barrier-damaging properties of this common detergent. The EDX technique allows a simultaneous determination of physiologically important elements, e.g., Na, Mg, P, Cl, K, Ca and S in addition to Ni at each point of measurement in epidermal cell strata. Our results show that SLS reduces the barrier function to Ni-ion penetration of the stratum corneum. In addition we have shown that EDX allows analysis of the influence of different factors involved in nickel penetration through the skin by giving data on the physiological effects on the epidermal cells caused by the applied substances.

Animals↗

PubMed1989

Distribution of diffusible ions in the aging rat lens. An X-ray microanalytical investigation.

The elemental content of the anterior and posterior cortex and of the nucleus of the rat eye lens, and the changes occurring as a function of age, were investigated by energy-dispersive X-ray microanalysis of cryostat sections. In contrast to most other cells investigated by this technique, the main element detectable in the lens was sulfur, not phosphorus. This is likely to be due to the lack of cell organelles and of nucleic acids in the lens cells. Up to 19 months of age, the concentrations of Na, Cl and K are much lower in the nucleus than in the cortex, whereas the concentration of S is highest in the cortex. At 25 months the differences in elemental content between nucleus and cortex, while still present, are less pronounced. The age-related changes in ion content are rather complex and appear to be different for different parts of the lens. In general, the ion content of the lens increases with age. In the nucleus, the concentration of K increases gradually with age, whereas the Na concentration only appears to increase after 19 months. The Na/K ratio in the lens nucleus is lower in the oldest groups. The Ca concentration increases with age in the nucleus and the posterior cortex, but not in the anterior cortex.

Aging↗