| 1. |
- Ågren, Johan
(författare)
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Water transport through perinatal skin : Barrier function and aquaporin water channels
- 2003
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- While constituting a well functioning interface with the aqueous environment in utero, the skin offers a poor barrier after very preterm birth. As a result, transepidermal water loss (TEWL) is high, a fact which has important clinical consequences in these infants. To investigate the transport of water through perinatal skin and the potential role of aquaporin (AQP), a water channel protein, in this process, we determined TEWL in a group of extremely preterm infants, and in an experimental rat model we analyzed the expression and distribution of AQP in perinatal skin in relation to TEWL, skin surface hydration and water content. The effects of antenatal corticosteroids (ANS) and of restricted intake of fluids and nutrients on barrier characteristics of the perinatal skin and its AQP expression were also studied.In infants born at 24 and 25 weeks of gestation TEWL was very high in the first days after birth and decreased with increasing postnatal age. At a postnatal age of 4 weeks, TEWL was still twice as high as previously reported in infants born at a gestational age of 25-27 weeks and four times higher than in infants born at term. In the rat model, immunohistochemical analysis revealed that AQP1 and AQP3 are abundantly expressed in the skin. AQP1 was expressed exclusively in dermal capillaries and AQP3 in basal layers of the epidermis. AQP1 and AQP3 mRNA as assessed by semiquantitative RT-PCR was higher in fetal than in adult skin. As in infants, TEWL and skin surface hydration were inversely related to gestational age in the rat. In preterm rat pups exposed to ANS, TEWL and skin surface hydration were lower than in unexposed controls, and AQP3 expression was selectively induced by ANS. In term newborn rat pups, restriction of fluid and nutrient intake resulted in a higher skin water content and higher TEWL early after birth, while at an age of 7 days TEWL was lower in fasting rat pups than in controls, although skin water content was still higher.To conclude, TEWL is very high in extremely preterm infants early after birth and then decreases at a slower rate than previously reported for a group of slightly more mature infants. This is the first time that the distribution and gene expression of AQP1 and AQP3 have been demonstrated in perinatal skin. The localization and expression of AQP in the skin might indicate that these water channels are involved in the regulation of skin hydration and transepidermal water transport in the fetus and newborn infant.
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| 2. |
- Allen, Adrian, et al.
(författare)
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Gastroduodenal mucus bicarbonate barrier: protection against acid and pepsin.
- 2005
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Ingår i: Am. J. Physiol. Cell Physiol.. ; 288:Jan, s. C1-C19
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Tidskriftsartikel (refereegranskat)abstract
- Secretion of bicarbonate into the adherent layer of mucus gel creates a pH gradient with a near neutral pH at the epithelial surfaces in stomach and duodenum, providing the first line of mucosal protection against luminal acid. The continuous adherent mucus layer is also a barrier to luminal pepsin thereby protecting the underlying mucosa from proteolytic digestion. Here we review the current state of the gastroduodenal mucus bicarbonate barrier two decades later from when the first supporting experimental evidence appeared. The primary function of the adherent mucus gel layer is a structural one to create a stable unstirred layer to support surface neutralisation of acid and act as a protective physical barrier against luminal pepsin. The emphasis therefore on mucus in this review is on the form and role of the adherent mucus gel layer. The primary function of the mucosal bicarbonate secretion is to neutralise acid diffusing into the mucus gel layer and for there quantitatively to be sufficient secretion to maintain a near neutral pH at the mucus-mucosal surface interface. The emphasis on mucosal bicarbonate in this review is on the mechanisms and control of its secretion and the establishment of a surface pH gradient. Evidence suggests that under normal physiological conditions the mucus bicarbonate barrier is sufficient for protection of the gastric mucosa against acid and pepsin and, even more so, in the duodenum
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| 3. |
- Ammoun, Sylwia, 1968-
(författare)
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Orexin Receptors in Recombinant CHO Cells : Signaling to Short- and Long-Term Cell Responses
- 2005
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Recently discovered neuropeptides orexins (orexin-A and -B) act as endogenous ligands for G-protein-coupled receptors called OX1 and OX2 receptors. Our previous studies have established model systems for investigation of the pharmacology and signaling of these receptors in recombinant CHO cells. OX1 receptor-expressing CHO cells were mainly utilized in this thesis.Orexin-A and -B activate both OX1 and OX2 receptors. However, orexin-B is less potent in activating OX1 receptors than orexin-A, whereas the peptides are equipotent on OX2 receptors. We have performed mutagenesis on orexin-A to investigate the basis for this selectivity. We show that OX2 receptor is generally less affected by the mutations and thus OX2 receptor appears to have less strict requirements for ligand binding, likely explaining the lack of difference in affinity/potency between orexin-A and orexin-B on OX2 receptor.The other studies focus on orexin receptor signaling. OX1 receptors are shown to regulate adenylyl cyclase both in positive and negative manner, activate different MAP-kinases (ERK1/2 and p38) and induce cell death after long-lasting stimulation. Adenylyl cyclase regulation occurs likely through three different G-protein families, Gi, Gs and Gq. For ERK1/2, several downstream pathways, such as Ras, Src, PI3-kinase and protein kinase C (PKC) are implicated. OX1 receptor-mediated activation of ERK is suggested to be cytoprotective whereas p38 MAP-kinase induces programmed cell death. Three particularly interesting findings were made. Firstly, novel PKC δ (delta) is suggested to regulate adenylyl cyclase, whereas conventional and atypical PKCs are involved in activation of ERK. Secondly, adenylyl cyclase and ERK activation is fully dependent on extracellular Ca2+. Further experiments suggest that the previously discovered receptor-operated Ca2+ influx is not affecting the downstream effectors of orexin receptors but that it instead enables orexin receptors to couple to several signal cascades. Thirdly, upon inhibition of caspases, classical mediators of programmed cell death, OX1 receptor-mediated cell death is not reversed, but instead the pathways to death are altered so de novo gene transcription is no longer required for cell death.
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| 4. |
- Bengtsson, Magnus W., et al.
(författare)
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Duodenal bicarbonate secretion in rats : Stimulation by intra-arterial and luminal guanylin and uroguanylin
- 2007
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Ingår i: Acta Physiologica. - : Wiley. - 1748-1708 .- 1748-1716. ; 191:4, s. 309-317
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Tidskriftsartikel (refereegranskat)abstract
- Aim: Uroguanylin and guanylin are endogenous ligands for guanylate cyclase C, an upstream regulator of the cystic fibrosis transmembrane resistance (CFTR) anion channel, and both peptides increase intestinal anion export in vitro. We have compared the effects of close intra-arterial and luminal administration of uroguanylin and guanylin on duodenal bicarbonate secretion in vivo and studied the interactions with melatonin and cholinergic stimulation. Methods: Lewis × Dark Agouti rats were anaesthetized and a segment of the proximal duodenum with intact blood supply was cannulated in situ. Mucosal bicarbonate secretion (pH stat) was continuously recorded and peptides were infused intra-arterially or added to the luminal perfusate. Results: Intra-arterial (50–1000 pmol kg−1 h−1) as well as luminal administration (50–500 nmol L−1) of guanylin or uroguanylin caused dose-dependent increases in the duodenal secretion. Luminal administration induced more rapidly appearing rises in secretion and the two peptides induced secretory responses of similar shape and magnitude. The melatonin MT2-selective antagonist luzindole (600 nmol kg−1) significantly depressed the response to intra-arterial guanylins but did not affect secretion induced by luminal guanylins. Similarly, the muscarinic antagonist atropine (0.75 μmol kg−1 followed by 0.15 μmol kg−1 h−1) abolished the response to intra-arterial uroguanylin but caused only slight suppression of the response to luminal uroguanylin. Conclusions: Intra-arterial as well as luminal uroguanylin and guanylin are potent stimuli of duodenal mucosal bicarbonate secretion in vivo. The response to luminal guanylins reflects an action at apical receptors. Stimulation by parenteral guanylins, in contrast, is under cholinergic influence and interacts with melatonin produced by mucosal enteroendocrine cells.
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| 5. |
- Bengtsson, Magnus Wilhelm, 1977-
(författare)
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Effects of Orexins, Guanylins and Feeding on Duodenal Bicarbonate Secretion and Enterocyte Intracellular Signaling
- 2008
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The duodenal epithelium secretes bicarbonate ions and this is regarded as the primary defence mechanism against the acid discharged from the stomach. For an efficient protection, the duodenum must also function as a sensory organ identifying luminal factors. Enteroendocrine cells are well-established intestinal “taste” cells that express signaling peptides such as orexins and guanylins. Luminal factors affect the release of these peptides, which may modulate the activity of nearby epithelial and neural cells.The present thesis considers the effects of orexins and guanylins on duodenal bicarbonate secretion. The duodenal secretory response to the peptides was examined in anaesthetised rats in situ and the effects of orexin-A on intracellular calcium signaling by human as well as rat duodenal enterocytes were studied in vitro.Orexin-A, guanylin and uroguanylin were all stimulants of bicarbonate secretion. The stimulatory effect of orexin-A was inhibited by the OX1-receptor selective antagonist SB-334867. The muscarinic antagonist atropine on the other hand, did not affect the orexin-A-induced secretion, excluding involvement of muscarinic receptors. Orexin-A induced calcium signaling in isolated duodenocytes suggesting a direct effect at these cells. Interestingly, orexin-induced secretion and calcium signaling as well as mucosal orexin-receptor mRNA and OX1-receptor protein levels were all substantially downregulated in overnight fasted rats compared with animals with continuous access to food. Further, secretion induced by Orexin-A was shown to be dependent on an extended period of glucose priming.The uroguanylin-induced bicarbonate secretion was reduced by atropine suggesting involvement of muscarinic receptors. The melatonin receptor antagonist luzindole attenuated the secretory response to intra-arterially administered guanylins but had no effect on secretion when the guanylins were given luminally. In conclusion, the results suggest that orexin-A as well as guanylins may participate in the regulation of duodenal bicarbonate secretion. Further, the duodenal orexin system is dependent on the feeding status of the animals.
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| 6. |
- Bengtsson, Magnus W., et al.
(författare)
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Food-induced expression of orexin receptors in rat duodenal mucosa regulates the bicarbonate secretory response to orexin-A
- 2007
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Ingår i: American Journal of Physiology - Gastrointestinal and Liver Physiology. - : American Physiological Society. - 0193-1857 .- 1522-1547. ; 293:2, s. G501-G509
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Tidskriftsartikel (refereegranskat)abstract
- Presence of appetite-regulating peptides orexin-A and orexin-B in mucosal endocrine cells suggests a role in physiological control of the intestine. Our aim was to characterize orexin-induced stimulation of duodenal bicarbonate secretion and modulation of secretory responses and mucosal orexin receptors by overnight food deprivation. Lewis x Dark Agouti rats were anesthetized and proximal duodenum cannulated in situ. Mucosal bicarbonate secretion (pH stat) and mean arterial blood pressure were continuously recorded. Orexin-A was administered intra-arterially close to the duodenum, intraluminally, or into the brain ventricles. Total RNA was extracted from mucosal specimens, reverse transcribed to cDNA and expression of orexin receptors 1 and 2 (OX1 and OX2) measured by quantitative real-time PCR. OX1 protein was measured by Western blot. Intra-arterial orexin-A (60–600 nmol·h–1·kg–1) increased (P < 0.01) the duodenal secretion in fed but not in fasted animals. The OX1 receptor antagonist SB-334867, which was also found to have a partial agonist action, abolished the orexin-induced secretory response but did not affect secretion induced by the muscarinic agonist bethanechol. Atropine, in contrast, inhibited bethanechol but not orexin-induced secretion. Orexin-A infused into the brain ventricles (2–20 nmol·kg–1·h–1) or added to luminal perfusate (1.0–100 nM) did not affect secretion, indicating that orexin-A acts peripherally and at basolateral receptors. Overnight fasting decreased mucosal OX1 and OX2 mRNA expression (P < 0.01) as well as OX1 protein expression (P < 0.05). We conclude that stimulation of secretion by orexin-A may involve both receptor types and is independent of cholinergic pathways. Intestinal OX receptors and secretory responses are markedly related to food intake.
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| 7. |
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| 8. |
- Bengtsson, Magnus W., et al.
(författare)
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Short food deprivation inhibits orexin receptor 1 expression and orexin-A induced intracellular calcium signaling in acutely isolated duodenal enterocytes
- 2009
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Ingår i: American Journal of Physiology - Gastrointestinal and Liver Physiology. - : American Physiological Society. - 0193-1857 .- 1522-1547. ; 296:3, s. G651-G658
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Tidskriftsartikel (refereegranskat)abstract
- Bengtsson MW, Makela K, Herzig KH, Flemstrom G. Short food deprivation inhibits orexin receptor 1 expression and orexin-A induced intracellular calcium signaling in acutely isolated duodenal enterocytes. Am J Physiol Gastrointest Liver Physiol 296: G651-G658, 2009. First published December 31, 2008; doi:10.1152/ajpgi.90387.2008.-Close intra-arterial infusion of the appetite regulating peptide orexin-A stimulates bicarbonate secretion from the duodenal mucosa. The aim of the present study was to elucidate the ability of orexin-A to induce intracellular calcium signaling in acutely isolated duodenal enterocytes. Freshly isolated clusters of enterocytes, obtained from rat duodenal mucosa or human duodenal biopsies, were loaded with fura 2-AM and mounted in a perfusion chamber. Cryptlike enterocytes were selected (caged), and changes in intracellular calcium concentration ([Ca2+](i)) were evaluated by fluorescence imaging. Total RNA was extracted from pellets of enterocytes and reverse transcribed to cDNA, and expression of orexin receptors 1 and 2 (OX1R and OX2R) was measured by quantitative real-time PCR. Orexin-A at all concentrations tested (1-100 nM) increased [Ca2+](i) in enterocytes isolated from continuously fed rats, and the OX1R-antagonist SB-334867 (10 nM) attenuated the response. The primary [Ca2+](i) response was a slow increase to a sustained plateau persisting after orexin-A removal, and a similar response was observed in enterocytes from human biopsies. In contrast to orexin-A, the OX2R agonist (Ala(11), D-Leu(15))orexin-B (1-10 nM) did not induce calcium signaling. There were no significant [Ca2+](i) responses in enterocytes from animals food deprived overnight, and overnight fasting decreased (P < 0.01) enterocyte OX1R as well as OX2R mRNA. Induction of intracellular calcium signaling in isolated duodenal enterocytes is thus mediated primarily by OX1R receptors. Short (overnight) food deprivation markedly depresses receptor expression and inhibits orexin-A induced increases in [Ca2+](i). Studies of enterocyte signaling and intestinal secretion requires particular evaluation regarding feeding status.
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| 9. |
- Berg, Anna, 1975-
(författare)
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Effects of nitric oxide on gastric acid secretion in human gastric mucosa : functional and morphological studies
- 2005
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Hydrochloric acid (HCI) is secreted in high amounts by parietal cells in the human gastric mucosa and the resulting low pH constitutes an important factor for creating a suitable environment for the digestion. The normal gastric mucosa is equipped with an arsenal of protective mechanisms against the extreme chemical environment which the gastric acid creates. There are situations when the barrier function of the gastric mucosa is disrupted and gastric acid becomes potentially deleterious. Understanding the regulatory mechanisms by which the secretion of gastric acid is controlled under physiological conditions may improve future treatment in peptic ulcers, gastritis and other gastric inflammatory disorders.Nitric oxide (NO) has previously been found to regulate gastric acid secretion in animals. Immunohistochemical investigation of normal human gastric mucosa revealed that hitherto unknown endocrine cells in the oxyntic mucosa express nitric oxide synthase (NOS). These cells were found located in close contact with parietal cells, which suggests a paracrine effect of NO on parietal cell function.Functional studies of the effects of exogenous and endogenous NO on stimulated gastric acid secretion were performed on isolated human gastric glands. Indirect determination of gastric acid secretion by using the 14C-labeled aminopyrine (AP) technique was used. Stimulation was induced by administration of histamine or db-cAMP. Secretagogue-induced AP-accumulation in gastric glands treated with NO-donor was significantly decreased compared with untreated glands. This indicates that exogenously administered NO inhibits stimulated gastric acid secretion in humans. Inhibition of endogenous NO-production by the use of NOS-inhibitors caused an increase in AP-accumulation, which suggests that NO released from cells within the glandular epithelium exerts a physiological effect in acting as an inhibitor of stimulated gastric acid secretory activity in humans.Further functional and morphological investigations showed that exogenously administered cGMP induced a concentration-dependent inhibition of AP-accumulation in isolated human gastric glands similar to that induced by NO-donors. When soluble guanylate cyclase (sGC), a common target enzyme for NO, was blocked NO failed to induce inhibition. Biochemical analysis of the cGMP concentrations in isolated gastric glands after treatment with NO-donor revealed that inhibition of AP-accumulation due to NO is accompanied by an increase in glandular cGMP content. This increase was localized by immunohistochemistry to the parietal cells. These results indicates that NO inhibits secretagogue-induced gastric acid secretion in isolated human gastric glands via activation of sGC, which results in an increased concentration of cGMP in the parietal cells.In order to determine the cGMP-dependent mechanisms leading to diminished output of gastric acid, parietal cells were investigated with emphasis on the cytological transformations associated with stimulation of acid secretion. Isolated human gastric glands were treated with NO-donor prior to administration of histamine. The cytoskeletal rearrangement as well as the translocation and incorporation of H+/K+-ATPase into the apical membrane was studied using con focal and electron microscopy techniques. Results showed that histamine-induced F-actin rearrangement as well as the translocation of H+/K+-ATPase rich tubulovesicles to the canalicular membrane, and their fusion with the same, was unaffected by NO. The secretory canaliculi, which swell to great size as a result of histamine-treatment, were however small and unexpanded in response to treatment with NO-donor. The unexpanded canaliculi reflected the NO-induced inhibition of secretion of HCI observed in the functional studies.In conclusion, these results show that NO may be a physiological regulator of stimulated gastric acid secretion in humans and that this inhibition is a cGMP-dependent mechanisms which diminishes output of HCI from parietal cells without affecting stimuli-induced cytological transformations.
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| 10. |
- Flemström, Gunnar, 1941-, et al.
(författare)
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Apelin stimulation of duodenal bicarbonate secretion : feeding-dependent and mediated via apelin-induced release of enteric cholecystokinin
- 2011
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Ingår i: Acta Physiologica. - Oxford : Wiley-Blackwell. - 1748-1708 .- 1748-1716. ; 201:1, s. 141-150
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Tidskriftsartikel (refereegranskat)abstract
- Aim: Apelin peptides is the endogenous ligand of the G protein-coupled receptor APJ. Proposed actions include involvement in control of cardiovascular functions, appetite and body metabolism. We have investigated effects of apelin peptides on duodenal bicarbonate secretion in vivo and the release of cholecystokinin (CCK) from acutely isolated mucosal cells and the neuroendocrine cell line STC-1. Methods: Lewis x Dark Agouti rats had free access to water and, unless fasted overnight, free access to food. A segment of proximal duodenum was cannulated in situ in anesthetized animals. Mucosal bicarbonate secretion was titrated (pH stat) and apelin was administered to the duodenum by close intra-arterial infusion. Total RNA was extracted from mucosal specimens, reverse transcripted to cDNA and expression of the APJ receptor measured by quantitative real-time PCR. Apelin-induced release of CCK was measured using (i) cells prepared from proximal small intestine, and (ii) STC-1 cells. Results: Even the lowest dose of apelin-13 (6 pmol kg-1 h-1) caused a significant rise in bicarbonate secretion. Stimulation occurred only in continuously fed animals and even a 100-fold greater dose (600 pmol kg-1 h-1) of apelin was without effect in overnight food deprived animals. Fasting also induced a 8-fold decrease in the expression of APJ receptor mRNA. Apelin induced significant release of CCK from both mucosal and STC-1 cells, and the CCKA receptor antagonist devazepide abolished bicarbonate secretory responses to apelin. Conclusions: Apelin-induced stimulation of duodenal electrolyte secretion is feeding dependent and mediated by local mucosal release of CCK
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