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Background Clinical observation suggests that acardiac twinning occurs only in the first trimester. In part, this contradicts our previous analysis (part IV) of Benirschke's concept that unequal embryonic splitting causes unequal embryo/fetal blood volumes and pressures. Our aim is to explain why acardiac onset is restricted to the first trimester. Methods We applied the vascular resistance scheme of two fetuses connected by arterio‐arterial (AA) and veno‐venous (VV) anastomoses, the small VV resistance approximated as zero. The smaller twin has volume fraction α 0.33 and rAA(40)=1.3 mm, modeled survival is >32 weeks. Conclusion Before 13 weeks, embryos with α

Background Acardiac twinning is a complication of monochorionic twin pregnancies. From literature reports, 30 of 41 relatively large acardiac twins with renal tissue produced polyhydramnios within their amniotic compartment. We aim to investigate the underlying mechanisms that cause excess amniotic fluid using an established model of fetal fluid dynamics. Methods We assumed that acardiac onset is before 13 weeks, acardiacs with renal tissue have normal kidney function and produce urine flow from 11 weeks on, and acardiac urine production requires a pressure of half the pump twin's mean arterial pressure. We apply a resistance network with the pump twin's arterio‐venous pressure as source, pump umbilical arteries, placenta, placental arterio‐arterial (AA) anastomoses and acardiac resistances. Acardiac amniotic fluid dynamics excluded acardiac lung fluid secretion, swallowing and the relatively small intramembranous flow. Results In small acardiacs with sufficient urine production, polyhydramnios will occur due to the lack of amniotic fluid resorption. Urine production is dependent upon having sufficient mean arterial pressure, which requires nearly a two‐fold larger resistance within the acardiac as compared to the placental AA resistance. Subphysiologic arterial pressure may result in renal dysgenesis. Conclusion Our findings suggest the potential for prediction of which clinical acardiac cases may or may not develop polyhydramnios based upon noninvasive assessments of renal tissue, blood flow and urine production. This information would be of great value in determining early obstetric interventions as opposed to conservative management. These findings may also contribute to an improved knowledge of the fascinating pathophysiology that surrounds acardiac twinning.

Background A causal genetic mutation is found in 40% of families with dilated cardiomyopathy (DCM), leaving a large percentage of families genetically unsolved. This prevents adequate counseling and clear recommendations in these families. We aim to identify novel genes or modifiers associated with DCM. Methods We performed computational ranking of human genes based on coexpression with a predefined set of genes known to be associated with DCM, which allowed us to prioritize gene candidates for their likelihood of being involved in DCM. Top candidates will be checked for variants in the available whole‐exome sequencing data of 142 DCM patients. RNA was isolated from cardiac biopsies to investigate gene expression. Results PDLIM5 was classified as the top candidate. An interesting heterozygous variant (189_190delinsGG) was found in a DCM patient with a known pathogenic truncating TTN‐variant. The PDLIM5 loss‐of‐function (LoF) variant affected all cardiac‐specific isoforms of PDLIM5 and no LoF variants were detected in the same region in a control cohort of 26,000 individuals. RNA expression of PDLIM5 and its direct interactors (MYOT, LDB3, and MYOZ2) was increased in cardiac tissue of this patient, indicating a possible compensatory mechanism. The PDLIM5 variant cosegregated with the TTN‐variant and the phenotype, leading to a high disease penetrance in this family. A second patient was an infant with a homozygous 10 kb‐deletion of exon 2 in PDLIM5 resulting in early‐onset cardiac disease, showing the importance of PDLIM5 in cardiac function. Conclusions Heterozygous PDLIM5 variants are rare and therefore will not have a major contribution in DCM. Although they likely play a role in disease development as this gene plays a major role in contracting cardiomyocytes and homozygous variants lead to early‐onset cardiac disease. Other environmental and/or genetic factors are probably necessary to unveil the cardiac phenotype in PDLIM5 mutation carriers., A computational ranking‐based method performed in whole exome sequencing data of dilated cardiomyopathy patient put PDLIM5 forward as novel gene implicated in this cardiac disease.

Background Benirschke postulated that acardiac twinning occurs when markedly unequal embryonic splitting combines with arterioarterial (AA) and venovenous placental anastomoses. We tested this hypothesis by model simulations and by comparison of outcomes with 18 “pseudo-” (twin fetus with beating heart but otherwise with clear signs of an acardiac) and 3 “normal” acardiac cases. Methods The smaller/larger cell volume ratio at embryonic splitting becomes the smaller/larger embryonic/fetal blood volume ratio (a). From a, we derived nonpulsating blood pressures using normal values (larger twin) and normal values at an appropriate earlier gestational age (smaller twin). These unequal pressure sources were used in a linear resistance fetoplacental network to calculate umbilical venous diameter ratios. Acardiac onset occurs when the smaller twin has 50% left of its normal, singleton placenta. Comparison with clinical cases approximated a by crown-rump-length-ratio to the 3rd power. Input parameters are a and the AA-radius at 40 weeks. Results Acardiacs can be small or large, can occur early or late, earlier at smaller a and larger AA, with larger umbilical venous diameter ratios at smaller a and smaller AA. Comparison with the 21 clinical cases was good, except for 2. Conclusion Our analysis supports Benirschke's hypothesis. The smaller twin has to share its placental perfusion with the larger twin, which is a novel finding. The AA size is essential for the future of both fetuses but complicates easy understanding of (pseudo-)acardiac clinical presentations. Late acardiac onset occurs infrequently. Using nonpulsating circulations may have caused our extensive predictions of late onset. An improved model requires including hypoxemia in the smaller twin from chronic placental hypoperfusion. Birth Defects Research (Part A), 2016. © 2016 Wiley Periodicals, Inc.

Background: Acardiac monochorionic twins lack cardiac function but grow by passive perfusion of the pump twin's deoxygenated arterial blood through placental arterioarterial (AA) and venovenous (VV) anastomoses and by hypoxia-mediated neovascularization. Pump twins therefore must continuously increase their cardiac output which may cause heart failure. Our aims were: to adapt our twin-twin transfusion syndrome model for acardiac twin pregnancies, to simulate pump and acardiac twin development, and to examine the model for early prognostic markers of pump twin survival. Methods: We used an infinite acardiac placental resistance, based on placental dye injection studies and simulations, suggesting the AA-Acardiac-VV series resistance determines the pump twin's excess cardiac output. Pump and acardiac development were expressed by the pump's excess cardiac output versus its normal value, represented by pump/acardiac umbilical venous diameter (UVD) ratios. Results: UVD ratios distinguish between AA-VV anastomoses that do and do not cause hydropic pump twins. Pump twins can handle relative larger acardiac perfusion at later than earlier gestation. Both VV and acardiac resistances are significantly smaller than the AA resistance, based on respectively clinical data and acardiac blood volumetric growth. Conclusion: Our simulations support clinical results which show that UVD ratios aid in the prediction of pump twin risk. The AA anastomosis controls the future of both the pump and the acardiac. Correlation between acardiac size and pump twin risk is secondary to the AA size but remains clinically usable. These factors may aid in the development of methods for pump twin prognosis and the promotion of selective clinical interventions.

Background: We recently published pump/acardiac umbilical venous diameter (UVD) ratios, representing the pump twin's excess cardiac output fraction, of 27 acardiac twin pregnancies. There was a clear separation between the 17 pump twins that had life-threatening complications and the 10 that did not. The hypothesis of this study is that placental chorangioma and sacrococcygeal teratoma (SCT), tumors whose perfusion also causes high-output complications, have the same fetal outcome as pump twins when perfusion of the tumor requires the same excess cardiac output fraction. Methods: We compared the three fetoplacental circulations. Fetuses with a placental chorangioma and acardiac twin pregnancies both have their feeding artery and draining vein located at the placental cord insertion. In contrast, SCT lacks a prescribed feeding artery and draining vein. We, therefore, had to modify our model to assume that the diameter of the hypothetical draining vein is related to the flow difference between inferior vena cava and superior vena cava. The latter flow has been estimated sonographically and is the same as the inferior vena cava flow in the absence of an SCT. Furthermore, a simple modification accounts for the different location of the tumor with respect to the placental cord insertion. Results: We propose to apply the clinical pump/acardiac UVD ratios to pregnancies complicated by placental chorangiomas and the modified pump/acardiac UVD ratios for SCT. Conclusion: Risk prediction of these rare fetal tumors may be possible based on application of data on excess cardiac output fractions from pump/acardiac UVD ratios and will require future clinical validation.

Tick populations and tick-borne diseases like Lyme borreliosis have been steadily increasing since the mid-1990s. Realizing the threat that ticks pose to public health, two Dutch citizen science projects have collected tick bite reports since 2006. This unique volunteered geographical dataset, which currently has nearly 35,000 reports, was used to identify environmental and other circumstantial factors associated with tick bites. For this, we first enriched the tick bite reports with temperature, precipitation, vegetation and volunteered data associated with the location of the tick bite. Using this enriched dataset, we then derived a series of features to characterize the environmental and volunteer-related conditions in which each tick bite occurred. Next, we discretized these features using the Jenks Natural Breaks algorithm and, after that, we mined frequent environmental patterns associated with tick bites using the AprioriClose algorithm. Finally, we checked that these patterns are specifically associated with the tick bites by comparing them with the frequent patterns mined from pseudo-random locations. The frequent patterns were visualized using heat maps and ring maps and two representative patterns associated with tick bites were projected into geographic space to study their spatio-temporal distribution. Our results show that factors linked to human activity are more relevant to model tick bites than seasonal accumulations of temperature, vegetation or precipitation. In particular, the number of warm and dry days per season are present in a significant number of patterns and the majority of tick bites are produced within a distance of half a kilometer of a forest, recreational or built-up area. The study of patterns in the time-series revealed that there are several persistent patterns consistently occurring each year and the validation process showed that the volunteer tick bites collection is capturing environmental conditions associated with tick bites, suggesting that these reports have a high scientific value. These results support the creation of a Dutch tick bite risk map that, in turn, will open the door to the design of public health interventions to reduce the incidence of Lyme disease.

Background Acardiac twinning is a rare anomaly of monochorionic twin pregnancies. Acardiac fetuses lack a functional heart but are passively perfused by arterial blood from their pump co-twin. Although four acardiac morphological types have been classified, the various paths of anatomical and circulatory acardiac twin development, and the potential influence of acardiac size and perfusion flow as possible predictors of pump twin morbidity and mortality are poorly understood. This report presents the first high resolution three-dimensional reconstruction of the vasculature of an acardiac twin by cryomicrotome imaging. Case A small, approximately 7.5-cm-diameter ball-shaped acardius amorphous of 30 5/7 weeks had caused pump twin cardiac decompensation that necessitated an emergency cesarian section. The pump twin survived well. The acardiac body had a partially intact vascular system with large diameter arteries and veins and multiple zones that appeared devoid of perfusion. The three-dimensional reconstruction showed neither recognizable organ structures nor identifiable blood vessels except for the umbilical artery and vein. Conclusion Our case showed a small acardiac mass with large diameter vessels and consequential low outflow resistance that caused pump twin complications. This indicates that the development of a method that allows pump twin prognosis is likely more successful if based on the use of acardiac versus pump twin perfusion flows than on body volume ratios. Birth Defects Research (Part A) 106:213–217, 2016. © 2015 Wiley Periodicals, Inc.

Key points Innate collateral arteries provide the biophysical substrate for arteriogenesis. Their distribution and morphology predestine tissue areas salvageable by collateral flow. Fluorescent episcopic cryomicrotome imaging resulted in a three-dimensional representation of the coronary network, in which collateral segments were automatically identified. Innate collateral segments are predominantly present in the subendocardium without preferential connectivity within the left ventricular wall of the dog heart and are preferentially oriented perpendicular to the long axis of the heart in the outer layers and parallel to this axis at the subendocardium. These results suggest that collateral segments are maintained without local hypoxia but because of heterogeneity in pressure gradients in the arterial tree. The high density and long-axis orientation of collateral arteries in the subendocardial region provide the substrate for arterial plexus formation and indicate the need for three-dimensional perfusion assessment in clinical perfusion imaging. Abstract Functional collateral vessels often stem from outward remodelling of pre-existing connections between perfusion territories. Knowledge of the distribution and morphology of innate collateral connections may help in identifying myocardial areas with protection against risk for ischaemia. The coronary network of six healthy canine hearts was investigated with an imaging cryomicrotome. Innate collateral connections ranged from 286 to 1015 μm in diameter. Left ventricular collateral density (number per gram of tissue) was about five in the subendocardium vs. 2.5 in the mid-myocardium (P

BACKGROUND: Approximately 75% of monozygotic twin pregnancies share one monochorionic placenta where placental anastomoses are virtually always present to connect the two fetoplacental circulations. These anastomoses cause several serious complications such as acardiac twinning. Acardiac twins lack a functional heart but nevertheless show fetal growth because the normal pump twin perfuses the acardiac body through arterioarterial (AA) and venovenous (VV) anastomoses. The widely accepted 1% monochorionic acardiac incidence dates back to 1944 and the associated 1:35,000 pregnancies to 1953. Our aim was to update this analysis. METHODS: We accepted the 1% (actually 1.1%) monochorionic acardiac incidence due to lack of more precise data, included the recently observed 58% early cessation of acardiac development as well as consequences of assisted reproductive technology, and assessed the incidence of acardiac twinning under conditions of AA-VV anastomoses. RESULTS: Early acardiac monochorionic twinning increased from 1.1% to 1.1/(1-0.58) = 2.6%, from 1:35,000 to 1:9,500 to 11,000 pregnancies, depending on number and method of assisted reproductive technology, and occurs in approximately 1:8 AA-VV anastomoses-containing monochorionic placentas. CONCLUSION: Early acardiac twinning is not a rare event. The 1944-based 1% acardiac monochorionic incidence has a weak basis and could therefore be (much) larger. Knowing this incidence more precisely may contribute to our knowledge of embryonic splitting in unequal cell masses. Birth Defects Research (Part A) 103:641-643, 2015. (c) 2015 Wiley Periodicals, Inc.

Background Arrhythmogenic right-ventricular cardiomyopathy (ARVC) can lead to RV dilatation. We hypothesized that electrocardiographic characteristics including QRS amplitudes in the extremity- and precordial leads, the S amplitude in lead V1, and extent of T-wave negativity over the precordial leads are related to RV dilatation in this condition. Methods In 42 ARVC patients and 42 controls, we correlated total QRS amplitude in the extremity leads (∑QRSext), precordial leads (∑QRSprec) and in all leads (∑QRStot : summation of ∑QRSext and ∑QRSprec), S amplitude in lead V1 and the extent of T-wave inversion in the precordial leads (V1 vs. beyond V1) with RV end diastolic diameter (RVEDD) by echocardiography. Results In the ARVC group, the mean age was 46 ± 14 years, 31 patients were male, 28 had an implantable cardioverter defibrillator (ICD), and 7 had a LV ejection fraction (EF) < 55%. The control group was age- and gender matched to the ARVC cohort. In contrast to controls, the ∑QRSext (regression coefficient (RC), −0.29; P = 0.020), ∑QRSprec (RC, −0.20; P = 0.015), and ∑QRStot (RC, −0.14; P = 0.009) were lower with RV dilatation in ARVC. S amplitude in lead V1 was not related to RV diameter (RC, −0.98; P = 0.088). Precordial T-wave inversion beyond lead V1 (V2-V6) was associated with a larger RV diameter (RC, 8.58; P = 0.012). Conclusions Summed QRS amplitudes in the extremity and precordial leads, and T-wave inversion beyond lead V1 are associated with RV dilatation in patients with ARVC.

Background Disturbance in fluid secretion, driven by chloride secretion, might play a role in constipation. However, disturbed chloride secretion in those patients has yet to be evaluated. Therefore, the aim of this study was to compare chloride secretion in rectal biopsies of children with functional constipation (FC) to those without constipation. Methods To measure changes in short circuit current (Isc in μA cm−2) reflecting chloride secretion, intestinal biopsies from children with constipation, to either exclude or diagnose Hirschsprung’s disease, and from children without constipation (controls) undergoing colonoscopy for screening of familial adenomatous polyposis, juvenile polyps or inflammatory bowel disease (IBD), were compared and studied in Ussing chambers. Following electrogenic sodium absorption blockade by amiloride, chloride secretory responses to calcium-linked (histamine, carbachol) and cAMP-linked (IBMX/forskolin) secretagogues were assessed. Key Results Ninety-six patients (46 FC) participated; nine FC patients (n = 1 congenital syndrome and n = 8 technical problems) and 13 controls (n = 6 IBD; n = 7 technical problems) were excluded. No significant difference was found in mean (±SE) basal chloride currents between children with FC and controls (9.6 ± 1.1 vs 9.2 ± 0.8; P = 0.75, respectively). Responses to calcium-linked chloride secretagogues (histamine and carbachol) were significantly higher in controls (33.0 ± 3.0 vs 24.5 ± 2.3; P = 0.03 and 33.6 ± 3.4 vs 26.4 ± 2.7; P = 0.05 following histamine and carbachol, respectively). Conclusions & Inferences Calcium-linked chloride secretion is disturbed in children with FC. Whether this defect occurs at the level of histamine receptors, components of receptor-linked signal transduction pathways or basolateral Ca2+-sensitive K+ channels enhancing the electrical driving force for apical chloride secretion, remains to be explored.

Aliment Pharmacol Ther 2011; 34: 638–648 Summary Background Functional dyspepsia is one of the most prevalent (15–40%) functional gastrointestinal disorders. Antidepressants such as amitriptyline are often used in these patients, but clinical studies are currently lacking. Aim To evaluate the effect of 8 weeks of treatment with amitriptyline on drinking capacity, symptoms evoked by a standardised drink test (primary endpoint) and clinical symptoms (secondary endpoint). Methods Patients meeting the Rome III criteria for functional dyspepsia (FD) were invited to participate in a double blind, randomised, placebo-controlled trial and were treated with either amitriptyline (12.5–50 mg) or placebo during 8 weeks. All included patients underwent a nutrient drink test before and after treatment. Drinking capacity and evoked symptoms were recorded. In addition, dyspeptic symptoms were weekly assessed using PAGI SYM (patient assessment of upper gastrointestinal symptom severity index) questionnaire. Results Thirty-eight patients (amitriptyline n = 18, placebo n = 20; age 41 ± 2 year, 61% F) completed the study. The drinking capacity of liquid meal was not affected by either amitriptyline or placebo treatment. Postprandial symptoms were not significantly different between amitriptyline and placebo. During the entire treatment, total symptom score (0.47 points, P = 0.02) and nausea (0.86 points, P = 0.004) on PAGI SYM were significantly reduced by amitriptyline compared with placebo. Conclusions Amitriptyline did not affect drinking capacity and postprandial symptoms evoked by the drink test in FD patients. However, total clinical symptom score and nausea were reduced during 8 weeks of treatment. Our data suggest that amitriptyline particularly improves nausea in functional dyspepsia, but larger clinical trials are needed to further confirm our findings.

Intestinal manipulation (IM) during abdominal surgery triggers the influx of inflammatory cells, leading to postoperative ileus. Prevention of this local muscle inflammation, using intercellular adhesion molecule-1 (ICAM-1) and leukocyte function-associated antigen-1-specific antibodies, has been shown to shorten postoperative ileus. However, the therapeutic use of antibodies has considerable disadvantages. The aim of the current study was to evaluate the effect of ISIS-3082, a mouse-specific ICAM-1 antisense oligonucleotide, on postoperative ileus in mice. Mice underwent a laparotomy or a laparotomy combined with IM after treatment with ICAM-1 antibodies, 0.1–10 mg kg−1 ISIS-3082, saline or ISIS-8997 (scrambled control antisense oligonucleotides, 1 and 3 mg kg−1). At 24 h after surgery, gastric emptying of a 99mTC labelled semi-liquid meal was determined using scintigraphy. Intestinal inflammation was assessed by myeloperoxidase (MPO) activity in ileal muscle whole mounts. IM significantly reduced gastric emptying compared to laparotomy. Pretreatment with ISIS-3082 (0.1–1 mg kg−1) as well as ICAM-1 antibodies (10 mg kg−1), but not ISIS-8997 or saline, improved gastric emptying in a dose-dependent manner. This effect diminished with higher doses of ISIS-3082 (3–10 mg kg−1). Similarly, ISIS-3082 (0.1–1 mg kg−1) and ICAM-1 antibodies, but not ISIS-8997 or higher doses of ISIS-3082 (3–10 mg kg−1), reduced manipulation-induced inflammation. Immunohistochemistry showed reduction of ICAM-1 expression with ISIS-3082 only. ISIS-3082 pretreatment prevents postoperative ileus in mice by reduction of manipulation-induced local intestinal muscle inflammation. Our data suggest that targeting ICAM-1 using antisense oligonucleotides may represent a new therapeutic approach to the prevention of postoperative ileus. British Journal of Pharmacology (2005) 146, 252–258. doi:10.1038/sj.bjp.0706303

Germination of seeds proceeds in general in two phases, an initial imbibition phase and a subsequent growth phase. In grasses like barley, the latter phase is evident as the emergence of the embryonic root (radicle). The hormone abscisic acid (ABA) inhibits germination because it prevents the embryo from entering and completing the growth phase. Genetic and physiological studies have identified many steps in the ABA signal transduction cascade, but how it prevents radicle elongation is still not clear. For elongation growth to proceed, uptake of osmotically active substances (mainly K(+)) is essential. Therefore, we have addressed the question of how the activity of K(+) permeable ion channels in the plasma membrane of radicle cells is regulated under conditions of slow (+ABA) and rapid germination (+fusicoccin). We found that ABA arrests radicle growth, inhibits net K(+) uptake and reduces the activity of K(+) (in) channels as measured with the patch-clamp technique. In contrast, fusicoccin (FC), a well-known stimulator of germination, stimulates radicle growth, net K(+) uptake and reduces the activity of K(+) (out) channels. Both types of channels are under the control of 14-3-3 proteins, known as integral components of signal transduction pathways and instrumental in FC action. Intriguingly, 14-3-3 affected both channels in an opposite fashion: whereas K(+) (in) channel activity was fully dependent upon 14-3-3 proteins, K(+) (out) channel activity was reduced by 14-3-3 proteins by 60%. Together with previous data showing that 14-3-3 proteins control the activity of the plasma membrane H(+)-ATPase, this makes 14-3-3 a prime candidate for molecular master regulator of the cellular osmo-pump. Regulation of the osmo-pump activity by ABA and FC is an important mechanism in controlling the growth of the embryonic root during seed germination.

Elevated levels of circulating monocytes (MCs) have been correlated with collateral development in coronary artery disease patients. The aim of this study was to employ a model of progressive coronary artery occlusion (CO), in order to compare the temporal response of MC upregulation in the systemic circulation vs. their infiltration into ischemic myocardium. An ameroid constrictor was implanted on the left coronary artery of New Zealand White rabbits (CO after 7-10 days). Peripheral blood MCs were isolated, fluorescently labelled and administered intravenously, 9 (n=4),16 (n=4) or 28 (n=5) days after surgery. Coronary vessels were perfused with fluorescent cast. Hearts were processed with an imaging cryomicrotome, generating 3D reconstructions of the vascular network and MCs. Peripheral blood MCs increased 83.47±55.18% 9 days after CO surgery, while 16 and 28 days after CO surgery showed 39.96±65.27 and -13.82±24.17 % change, respectively. MC subset ratios (classical/non-classical) showed the largest per...

Some species, including humans, have structural collateral vessels, CV, that can further develop in the presence of coronary stenosis by increasing pressure difference across them. Question is how ...

We previously reported activation of an inhibitory adrenergic and a non-adrenergic non-cholinergic (NANC) pathway during abdominal surgery relaxing the rat gastric fundus. In the present study, we investigated the possible role of nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) in the NANC part of the surgery-induced fundic relaxation. The effect of the NO biosynthesis inhibitor NG-nitro-L-arginine (L-NOARG), the non-selective VIP receptor antagonist [D-p-Cl-Phe6,Leu17]-VIP and the selective VIP1 receptor antagonist [Acetyl-His1,D-Phe2,Lys15,Arg16,Leu17]-VIP was investigated on the non-adrenergic fundic relaxation induced by manipulation of the small intestine followed by resection of the caecum. Guanethidine partly reduced the manipulation-induced fundic relaxation. Addition of L-NOARG reduced this non-adrenergic component, whereas the non-selective VIP receptor antagonist had no significant effect. Combination of L-NOARG and the non-selective VIP antagonist however further reduced the relaxation to manipulation. The selective VIP1 receptor antagonist reduced the mean and maximal relaxation induced by abdominal surgery in the presence of guanethidine. When combined with L-NOARG, the relaxation of the gastric fundus was almost completely abolished. The VIP1 receptor antagonist alone had no significant effect on the mean and maximal relaxation, but enhanced recovery of fundic tone. In conclusion, as VIP1 receptors are not present in the rat gastric fundus, these results suggest that the NANC inhibitory pathway activated during abdominal surgery involves VIP1 receptors, most likely in the afferent limb. The inhibitory neurotransmitters released at the level of the gastric fundus smooth muscle are NO and a substance different from VIP. British Journal of Pharmacology (2000) 131, 705–710; doi:10.1038/sj.bjp.0703625

The ability of preferredoxin to inactivate a 50-pS anion channel of the chloroplast inner membrane in the presence of an energy source was investigated using single-channel recordings. It was found that preferredoxin cannot inactivate the channel when GTP is the only energy source present. From this it is concluded that the precursor has to interact with the , translocon of the inner membrane of chloroplasts (Tic) complex to be able to inactivate the 50-pS anion channel. The ability of two mutants of preferredoxin with deletions in their transit sequence to inactivate the channel was also tested. Both mutants have been shown to have a similar binding affinity for the chloroplast envelope, but only one is able to fully translocate. The mutants were both able to inactivate the channel in a similar manner. From this it is concluded that full translocation is not necessary for the inactivation of the channel. It is also shown that preferredoxin is capable of inactivating the 50-pS anion channel in the chloroplast-attached configuration as was previously found in the inside-out configuration. From this it is concluded that stromal factors do not influence the protein-import-induced inactivation of the 50-pS anion channel of the chloroplast inner membrane. Finally the effect of the anion channel blocker 4,4′-diisothiocyanostilbene-2,2′-disulfonate (DIDS) on the channel activity and on protein import was investigated. It was found that DIDS blocked the channel. Furthermore the addition of the channel blocker reduces the efficiency of import to 52%. This leads to the conclusion that correct functioning of the channel is important for protein import.

An ex vivo model system was developed to investigate melanocyte migration. Within this model system, melanocytes migrate among other epidermal cells in the epibolic outgrowth of skin explants. This process is initiated by loss of contact inhibition of epidermal cells at the rim of the explants and by locally produced chemotactic factors. Punch biopsies provided explants of reproducible diameter. Optimal culture conditions include medium consisting of Dulbecco's Minimal Essential Medium containing 10% inactivated normal human serum and placement of explants epidermal side up at the air-liquid interphase. Within 7 days, epidermal cells completely surround the explant. Approximately 3 days after the onset of keratinocyte migration, melanocytes distribute themselves within the newly formed epidermis. Throughout the 7-day culture period, melanocytes and keratinocytes show maintenance of subcellular morphology, and the dermo-epidermal junction remains intact. Melanocyte migration was quantified using immunoperoxidase staining in combination with light microscopy and computer-aided image analysis. Preliminary results using the model system to compare migration in control and nonlesional vitiligo skin indicate that no inherent migration defect is responsible for impaired repigmentation of vitiligo lesions. The organotypic culture model system allows for investigations on melanocytes within their environment of autologous epidermal and dermal components, closely resembling in vivo circumstances in human skin.

Vitiligo is an acquired melanin pigmentary disorder manifesting itself by expanding depigmented lesions of the skin. To date, the etiopathomechanism of vitiligo has not been convincingly elucidated and a number of seemingly mutually opposed hypotheses with equal likelihood still coexist. Concurrent theories on vitiligo etiology, together with supportive evidence, are reviewed here. Due to the observed variation in clinical manifestations of the disease, it seems likely that the etiology of vitiligo may differ among patients. Therefore several theories on vitiligo etiopathogenesis have been combined to formulate a convergence theory for vitiligo, also presented in this article. This theory states that stress, accumulation of toxic compounds, infection, autoimmunity, mutations, altered cellular environment and impaired melanocyte migration and/or proliferation can all contribute to vitiligo etiopathogenesis in varying proportions.