Biomedical subjects

G M Roomans

Publications and source records attributed to G M Roomans.

At least 73 records · Page 4Linked to original sources

PubMed1997

Response of myocardial cellular energy metabolism to variation of buffer composition during open-chest experimental cardiopulmonary resuscitation in the pig.

The aim of the present study was to investigate possible relationships in piglets between myocardial energy-related metabolites and intracellular electrolytes during open-chest cardiopulmonary resuscitation (OCCPR) supplemented by the administration of alkaline buffers with varying sodium content. Our hypothesis was that an increasing myocardial intracellular sodium content would decrease the intracellular energy stores. In addition to haemodynamics, acid-base and blood gas variables were analysed, and myocardial biopsies were collected before and during OCCPR as well as after the return of spontaneous circulation. After a period of 4 min of untreated ventricular fibrillation (VF). 25 piglets were randomly allocated to one of four groups: OCCPR with normal saline (n = 5); OCCPR with sodium bicarbonate (SB) (n = 7); OCCPR with Tris buffer mixture (TBM) (n = 7); and a totally untreated control group (n = 6). The results showed that 4 min of untreated VF almost eradicated creatine phosphate (CrP) and that the ATP/ADP ratio decreased to 1.5-2.0. During OCCPR with normal saline, the myocardial content of CrP increased, whereas lactate, ATP and ADP levelled off and AMP decreased, causing an increased ATP/ADP ratio. The adenosine and inosine contents increased, whereas inosine monophosphate was unchanged at a low level, the adenosine and inosine contents being inversely correlated with the total content of adenine nucleotides. In both buffered groups, the increase in most energy-related metabolites (CrP, ATP, ADP, AMP and the ATP/ADP quotient) was less and in lactate more pronounced than in the group not being buffered, with no difference between the groups receiving SB or TBM. Although the intracellular potassium content was unaltered, the sodium, chloride and calcium concentration increased, more so in the group receiving SB. The intracellular content of sodium was correlated with that of calcium. Thus, buffering increased the myocardial AMP degradation during OCCPR by increasing the flux via the 5'-nucleotidase reaction, and SB increased the intracellular contents of sodium and calcium to a greater extent than did TBM.

Adenosine Diphosphate↗

PubMed1997

Methods for X-ray microanalysis of epidermis: the effect of local anaesthesia.

The effect of local anaesthesia on the elemental content of cells in human epidermis was studied by electron probe X-ray microanalysis. Local anaesthesia with lidocaine was given by intracutaneous injection within 1 min prior to taking a skin biopsy. Biopsies taken without local anaesthesia were used as controls. Lidocaine with or without adrenaline caused a significant increase in the concentrations of Na and Cl, and a decrease in the concentration of K in the cells of the stratum basale and the stratum spinosum, compared with the control samples. The presence of adrenalin in the anaesthetic did not change the effect of lidocaine. The effects of local anaesthesia have to be considered in planning and interpretation of clinical applications of X-ray microanalysis.

Adult↗

PubMed1997

Quantitative immunoconfocal analysis of human myometrial gap junction connexin43 in relation to steroid hormone concentrations at term labour.

The aim of this study was to quantify gap junction expression in the human myometrium in relation to progesterone and oestradiol concentrations, and to establish whether oxytocin-resistant dystocia is due to an abnormality in gap junction expression. Three groups of patients were investigated: (i) before labour (at term), (ii) normal labour and (iii) oxytocin-resistant dystocia (eight patients per group). For each patient, the concentrations of oestradiol and progesterone in maternal blood and in myometrial tissue were measured, and the number and area of immunostained connexin43 gap junctions per unit volume of tissue determined by quantitative analysis of digital images obtained by confocal microscopy. No significant difference in connexin43 gap junction content was observed between the three patient groups. When all groups were pooled, there was a significant positive correlation (P < 0.05) between the quantity of immunolabelled gap junctions and the oestradiol:progesterone ratio, but there was no significant difference in this correlation between the groups. Gap junction immunolabelling was not correlated with the progesterone or oestradiol concentration in the maternal blood or the myometrium. These data suggest that in human myometrium: (i) dystocia is not due to a reduced level of immunodetectable connexin43 gap junctions, (ii) onset of labour is not associated with a sudden increase in immunodetectable gap junction protein and (iii) gap junctions can be expressed in the presence of high progesterone concentrations.

Adult↗

PubMed1997

Ion transport in cultured pig tracheal submucosal gland acinar cells studied by X-ray microanalysis.

The submucosal glands of the airway may contribute more to the airway fluid than the surface epithelium. The cellular mechanisms underlying the regulation of electrolyte and water transport in airway submucosal glands are, however, still poorly understood. Therefore, we attempted to establish a cell culture system to facilitate study of this regulation. Acinar cells were isolated by enzymatic disaggregation from pig tracheal submucosal tissue and cultured on a plastic substrate coated with human placental collagen. The fine structure of the cells in confluent culture was studied by conventional transmission electron microscopy. The elemental content in resting cells and stimulated cells grown on a permeable substrate was studied by X-ray microanalysis. The cultured cells retained structural characteristics (microvilli, secretory granules and desmosomes) of in situ epithelia. The total intracellular Cl and K concentrations significantly decreased after stimulation with the cholinergic agonist acetylcholine, the predominantly alpha-adrenergic agonist norepinephrine, or the beta-adrenergic agonist isoproterenol. Both ionomycin and 8-bromoadenosine 3':5'-cyclic monophosphate (8-bromo-cAMP) caused a marked decrease of the intracellular Cl, K and Na concentrations. Cl- secretion induced by acetylcholine was inhibited by Cl- channel blockers anthracene-9-carboxylic acid and 5-nitro-2-(3-phenylpropyl-amino)-benzoic acid (NPPB), but Cl- efflux induced by 8-bromo-cAMP was blocked only by NPPB. The intracellular Cl and Na content significantly increased and the cellular K content markedly decreased after treatment with ouabain. These results indicate that the cultured acinar cells maintained the structural and functional characteristics of in situ tissue and that this system is suitable for studying aspects of ion and water transport by the airway submucosal gland cells.

Animals↗

PubMed1997

Evidence for muscarinic 3 receptor mediated ion transport in HT29 cells studied by X-ray microanalysis.

Changes in elemental content in response to muscarinic drugs in HT29 cells were investigated by X-ray microanalysis. Acetylcholine (ACh) and carbachol (Cch), both agonists binding to muscarinic receptors, induced a decrease of the intracellular Cl and K content. This agrees with the notion that these agonists induce electrolyte and water secretion. Atropine, a non-selective antagonist of muscarinic receptors, inhibited the decrease in K and Cl caused by ACh and Cch, and instead caused an increase of the Cl and K concentrations. A similar inhibition was found in the case of the selective muscarinic 3 receptor antagonist P-F-HHSiD. In contrast, the selective muscarinic 2 receptor antagonist AF-DX 116 did not inhibit Cch-activated secretion of K and Cl. A slight inhibition of ACh induced ion secretion was seen, but this inhibition was weak compared to that caused by P-F-HHSiD. Treatment with U-73122, an inhibitor of phospholipase C, blocked ACh or Cch induced ion secretion. These results suggest that ACh and Cch stimulated secretion of Cl and K is mediated by muscarinic 3 receptors via the inositol-1,4,5-trisphosphate (IP3) dependent pathway.

Acetylcholine↗

PubMed1997

Nebulisation of hypertonic saline causes oedema of the airway wall.

The effect of aerosol challenge with distilled water, and with isotonic and hypertonic saline on the respiratory system of the anaesthetized rabbit was investigated. Nebulisation of hypertonic (3.6%) saline caused an increase in the extravascular lung water without altering the total body weight. Morphometrical investigations revealed an increase of the subepithelial tissue compartment (connective tissue and smooth muscle) of the airway wall. X-ray microanalysis showed higher content of Na, K, and Cl in this compartment already 10 min after nebulisation of hypertonic saline. The formation of oedema was associated with a significant decrease in both compliance and gas exchange.

Animals↗

PubMed1997

X-ray microanalysis of uterine secretion in the mouse.

In an attempt to characterize ion transport mechanisms in the uterine epithelium the effect of diuretics on the elemental composition of the uterine secretion was investigated by X-ray microanalysis. Mice were treated with a variety of diuretics and Sephadex beads were incubated in situ in the uterus where they absorbed the uterine secretion. In animals treated with furosemide, bumetanide, or amiloride, the concentration of Na, K, and Cl in the uterine secretion was significantly decreased. After treatment with acetazolamide, only the K concentration was significantly decreased. Treatment with diuretics also decreased the concentration of P and S in the uterine secretion, indicating that less mucus was absorbed to the Sephadex beads. Transmission electron microscopy confirmed that treatment with amiloride and furosemide decreased the amount of mucus covering the epithelium. A possible explanation is that because of lack of water, the uterine mucus cannot leave the lumen of the uterine glands and reach the surface of the epithelium where the beads are. The data strongly suggest that Na+, K+, and Cl- transport in the uterine epithelium is dependent on a basolaterally situated Na(+)-(K+)-Cl(-)-cotransport mechanism. Disturbance of normal ion and water transport apparently affects the composition and consistency of the mucus, which may help explain abnormalities in cervical mucus in patients with cystic fibrosis.

Acetazolamide↗

PubMed1997

Effects of nicotine on the uterine epithelium studied by X-ray microanalysis.

The composition of the uterine fluid is of importance for the development of the embryo. The ionic composition of the uterine fluid is determined by transport processes in the uterine epithelium. The aim of the present study was to investigate whether nicotine can affect the ion transport processes and ultrastructure of the uterine epithelium in mouse and rat. The effects of nicotine on the uterine epithelium were compared with those on kidney and liver. Nicotine was administered to mice as an intraperitoneal injection of a 1 microgram/l nicotine solution twice daily for 8 days, or in the drinking water (108 microM nicotine) for 15 days, whereas rats received nicotine in their drinking water for 10 days (54 or 108 microM nicotine). In addition, primary cultures of endometrial cells were exposed to nicotine in vitro. Changes in morphology were examined by transmission electron microscopy and changes in elemental content by X-ray microanalysis. Nicotine caused a significant decrease in the concentrations of Na, K, and Cl in the uterine fluid and in the endometrial cells in situ. Similar changes in elemental concentrations were observed in endometrial cells in vitro. Ultrastructural changes in endometrial cells included swollen and damaged mitochondria. In liver and kidney much smaller effects of nicotine on the elemental composition of the cells were observed. However, marked ultrastructural effects were seen in the kidney: a reduction in mitochondrial size and an increase in lysosomal volume. It is concluded that nicotine directly interferes with fluid transport across the uterine epithelium.

Animals↗

PubMed1996

X-ray microanalysis in cell biology and cell pathology.

Electron probe X-ray microanalysis has been used for the last 25 years by biologists to obtain information about the distribution of elements at the cell and tissue level. During this period, progress has mainly been made through the development of more adequate techniques for specimen preparation (mainly low temperature techniques) and quantitative analysis, so that accurate analysis of the physiologically important cellular ions can be carried out. Use of in vitro systems and cell cultures may further increase the number of problems to which X-ray microanalysis can be applied. Among the numerous applications of X-ray microanalysis in cell biology and cell pathology, applications in the areas of epithelial ion transport, the role of calcium in secretory and contractile cells, and the role of ions in cell proliferation and cancer are discussed in more detail.

Animals↗

PubMed1996

Chloride secretion in the submandibular gland of adult and early postnatal rats studied by X-ray microanalysis.

Submandibular acinar cells of 1-day-old, 7-day-old, and adult rats were analyzed with X-ray microanalysis after stimulation with carbachol for different time periods (2-7 min). In unstimulated animals, marked differences in elemental content between compartments could be observed: secretory granules had a higher Ca and lower P and K content than other cell compartments. Comparison between different age groups showed significant differences for Ca, which increased with age in all compartments; Mg increased with age in the secretory granules and the apical cytoplasm. Only the glands from adult animals showed a significant effect of cholinergic stimulation: a transient decrease in Cl and K. The Cl concentration in the secretory granules decreased to 60% of the control value, which suggests that the granules release Cl upon stimulation. In young animals, no or little change in elemental distribution was observed after stimulation. This may indicate that Cl- secretion mechanisms are much less prominent in young animals. The ultrastructure of submandibular secretory granules depends on the preparation method: condensed and electrondense in freeze-substituted unfixed tissue, decondensed and more translucent in aldehyde-fixed tissue. This may indicate that the granules can transport water, and swell during the process of aldehyde fixation.

Aging↗

PubMed1996

Effects of progesterone and oxytocin on intracellular elemental composition of term human myometrium in vitro.

OBJECTIVE: To study the effects of progesterone and oxytocin on contractile activity related to the intracellular elemental composition of human pregnant myometrium. STUDY DESIGN: Myometrial strips were mounted in tissue baths and superfused with plain buffer, oxytocin or progesterone. Progesterone was initially added to the tissue bath or at the onset of spontaneous contraction of the strips. The myometrial contractile activities were recorded isometrically and the results were analyzed by a specific computer program. The effects of oxytocin and progesterone on the intracellular elemental composition were studied by X-ray microanalysis in an electron microscope. RESULTS: The contractile activity in terms of frequency and tonus of the muscle strips was increased by oxytocin and progesterone. This increase was more pronounced if progesterone was added to the bath at the start of the experiment. After 120 min of incubation with oxytocin and progesterone the total intracellular concentration of calcium was significantly higher (P < 0.0001 and P < 0.002, respectively) compared to buffer. CONCLUSIONS: (i) Oxytocin increases total intracellular calcium concentration concomitant with an increase in uterine contractile activity. (ii) Progesterone increases the myometrial tone and frequency of contractions, simultaneously with an increase in total intracellular calcium.

Calcium↗

PubMed1996

cAMP-induced chloride transport in NCL-SG3 sweat gland cells.

cAMP-induced ion transport in a human sweat gland cell line, NCL-SG3, was investigated by X-ray microanalysis and patch-clamp technique. Stimulation with cAMP caused a decrease in the cellular Cl and K. cAMP had no significant effect on the intracellular Na and Ca. Chloride channel blockers (9-AC, DPC and NPPB) inhibited the cAMP-induced chloride efflux. In patch-clamp experiments the inward current increased over a period of 5-15 min on addition of membrane-permeable cAMP in 66% of the attempts when the cell was held at 0 mV and pulsed to negative membrane potentials. The inward current was completely blocked by chloride channel blockers. Washout reversed the effect of cAMP. The inward current was not diminished by substitution of impermeable cations for Na in the bath and was insensitive to TEA (tetraethylammoniumchloride). It is concluded that the inward current is mainly a chloride current. Cystic fibrosis transmembrane regulator (CFTR) could not be demonstrated in the NCL-SG3 cells. It is therefore possible that the chloride efflux is mediated by other types of chloride channels.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1996

Progesterone, oestradiol and oxytocin and their in vitro effect on maintaining the number of gap junction plaques in human myometrium at term.

Previous studies by electron microscopy and laser confocal microscopy of immunohistochemically stained sections have shown that during pregnancy the extent of gap junction formation in human myometrium is low, but that an increase occurs in the active stages of labour despite a high concentration of progesterone in maternal blood. The present investigation focused on the effect of in vitro exposure of isolated myometrial tissue to progesterone, oestradiol and oxytocin, on the number of gap junction plaques in human myometrium at term. Myometrial biopsies were obtained at term from 13 pregnant women who had an elective caesarean section in the 37th or 40th week of pregnancy. The biopsies were immersed immediately in Hepes buffer and buffer containing 0.5, 5.0 micrograms mL-1 of progesterone, and 0.1 micrograms mL-1 of oestradiol. The muscle biopsies were trimmed under a stereo microscope into strips along the bundles of smooth muscle cells and mounted in tissue baths, superfused with Hepes buffer supplemented with glucose (0.01 mM); subsequently the strips were exposed to buffer containing different concentrations of progesterone, oestradiol and oxytocin. Incubation of the muscle strips for 180 min resulted in a significant decrease of the number of gap junctions (P < 0.01). Neither oestradiol or oxytocin, alone or in combination, had a significant effect on the maintenance of the number of gap junctions. The progesterone concentration of 5.0 micrograms mL-1, combined with oxytocin, and with or without oestradiol has a significantly positive effect on the number of gap junction plaques in strips of human myometrium at term (P < 0.05 vs. buffer alone). The high concentration of progesterone in the superfusion medium during in vitro experiments may be responsible for the maintenance of high numbers of gap junction complexes in term human myometrium. This finding is of interest in the light of findings of persisting high progesterone levels in maternal blood during labour.

Estradiol↗

PubMed1996

Use of primary cell cultures and intact isolated glandular epithelia for X-ray microanalysis.

Changes in the elemental composition of cells during isolation of glandular epithelia were studied by electron probe X-ray microanalysis. Fine chopping of rat submandibular gland followed by enzymatic treatment for 15 min caused marked increases in Na and Cl and a decrease in K concentrations in acinar cells. After enzymatic treatment for 50 min, Na, Cl and K concentrations returned to close to the control level. Mechanical disaggregation of the acinar clumps following enzymatic treatment resulted again in minor increases in Na and Cl and a marked decrease in K concentration. Exposure of isolated acini to cholinergic stimulation in vitro resulted in secretion of Cl and K from the acinar cells. Dissection of the sweat gland from human skin caused a decrease in the K/Na ratio. Incubation of the gland for 30-45 min with collagenase gave rise to a gradual decrease in the K/Na ratio. After mechanical separation of the gland into the secretory coil and reabsorptive duct, a further reduction of the K/Na ratio was seen. However, the duct cells had a much lower K/Na ratio and higher Ca concentration than the coil cells. In primary cultures, the K/Na ratios of the coil and duct cells returned to the in situ level. The elemental composition of sweat gland cells incubated in collagenase-containing medium was no different from that in cells incubated in collagenase-free medium. In the intact collagenase-isolated tissue, Cl- secretion in the coil was elicited by carbachol but not by cAMP, whereas in the duct cells the reverse was the case. In primary cell cultures, Cl- efflux in both coil and duct cells could be elicited by both carbachol and cAMP. In conclusion, although changes in elemental composition of gland cells during the isolation procedure occur, physiological responses can be detected. When primary cell cultures are used, it should be borne in mind that cultured cells may have physiological properties different from those of the intact tissue.

Animals↗

PubMed1996

X-ray microanalysis applied to the study of salivary glands.

X-ray microanalysis can be used to study the distribution of elements in salivary glands at the cellular and subcellular level in a variety of physiological and pathophysiological conditions. The acinar cells of the adult rat and mouse submandibular gland contain high concentrations of calcium due to the presence of calcium-rich secretory granules. The calcium concentration of the acinar cells can be increased under a variety of pathophysiological conditions, e.g., in animal models for cystic fibrosis, due to an increase in the number of secretory granules and changes in their calcium-binding properties. Cholinergic stimulation results in a decrease in the concentrations of Cl and K in all cell compartments in adult animals, but not in newborn rats. Also the secretory granules in the glands in adult animals lose Cl and K, indicating that ion (and water) transport mechanisms may be present in the membrane of the granules. During chemical fixation, the secretory granules in the adult glands swell and their morphology is different from that in freeze-substituted glands. The characteristics of the secretory granules in the acinar cells of newborn rats are different from those of adult animals.

Animals↗

PubMed1996

Steroid hormone effects on intercellular communication between term pregnant human myometrial cells before labor.

The appearance of gap junctions (GJs) between myometrial smooth muscle cells is one of the major events associated with the onset of labor. We have employed dye-coupling and electrical-current injection techniques to study the mechanisms by which steroid hormones regulate GJs in term pregnant myometrium of women before labor. Progesterone (P4) did not alter the input resistance (Ro) of the tissues when added to Tyrode's solution, which was used as control treatment. Octanol, the putative gap junctional uncoupling agent, increased the Ro of the cells compared to the control and P4-treated groups. The membrane potential (Em) did not differ between these groups. However, when P4 was applied after the tissue was perfused with estradiol (E2), the results changed dramatically: the Em hyperpolarized, and the Ro increased. Octanol increased the Ro in E2-treated tissues, but did not affect the Em. Consecutive application of E2, octanol, E2, and P4 resulted in rapid changes in the Ro of the cells. Dye-coupling was mostly detected between cells from controls and E2-treated tissues. These results indicate that P4 exerts its effects in the presence of E2 and that P4 has rapid effects on the intercellular communication between human myometrial cells.

Cell Communication↗

PubMed1996

Different pathways in irritant contact eczema? Early differences in the epidermal elemental content and expression of cytokines after application of 2 different irritants.

The epidermal response to 2 different irritants, nonanoic acid (NAA) and sodium lauryl sulfate (SLS), was investigated with 2 different methods. NAA 80% and SLS 4% were applied under occlusion for up to 24 h. Elemental changes were determined in cryosections by x-ray microanalysis. Compared to unexposed skin a significantly higher sodium/potassium ratio was found after 6 h in NAA-exposed skin and a lower ratio in SLS-exposed. At 24 h both substances had induced similar changes, compatible with a cell injury. The findings demonstrate a time-dependent NAA and SLS response. With reverse transcription polymerase chain reaction, the mRNA expression of interleukin-1 alpha (IL-1 alpha), -1 beta (IL-1 beta), -6 (IL-6), and -8 (IL-8), tumor necrosis factor alpha (TNF alpha) and granulocyte macrophage colony stimulating factor (GM-CSF) in shave biopsies from irritated and unexposed skin was studied at 0. 4. 8 and 24 h. NAA, but not SLS, induced an increase in mRNA expression for IL-6 mRNA-expression for GM-CSF was increased after SLS exposure, but not after NAA. These findings indicate a time and substance dependent difference in the up-regulation of mRNA for different cytokines in epidermis during the first 24 h of the irritant reaction. This might be the effect of differences in the irritants action on the cell membranes, which is also reflected by the differences found in the elemental content at 6 h.

Adult↗

PubMed1996

X-ray microanalysis of myometrium in parturient women at term.

The purpose of this study was to investigate the elemental composition of myometrial cells from term parturient women with normal and dysfunctional labour. Myometrial biopsies were obtained from forty-four term pregnant women undergoing Cesarean section. The patients were categorized according to uterine activity as follows: before labour, normal labour, labour successfully augmented by oxytocin, oxytocin-resistant labour, and labour arrested by terbutaline. X-ray microanalysis of freeze-dried sections (16 mu m thick) of the myometrium was carried out. An increase in intracellular phosphorus level (p < .01) was noted in the normal labour group compared to before labour. In patients with normal labour, higher phosphorus (p < .009) and potassium (p < .005) were found compared to oxytocin resistant labour. Patients with oxytocin resistant labour had lower intracellular potassium (p < .0006) and phosphorus (p < .02), and higher chloride (p < .05) and sodium (p < .03) compared to levels found in patients who responded to oxytocin treatment. In dysfunctional (oxytocin-resistant) labour the ion distribution in the myometrial cells might be disturbed. The reduced level of potassium and phosphorus together with the high sodium and chloride levels found in patients with oxytocin resistant labour may be connected to an impairment in sodium-potassium pump and muscle dysfunction, clinically diagnosed as dystocia.

Biopsy↗