Congenital Nail Disorders among Children with Suspected Ectodermal Dysplasias.

We report on a cohort of 204 children referred between January 2017 and January 2022 to the German Center for Ectodermal Dysplasias, Erlangen. The most frequent reasons for referral were tooth malformations and lack of multiple teeth leading to the suspicion of an ectodermal dysplasia. Many patients also suffered from being unable to perspire. Nail abnormalities, in contrast, represented a much rarer finding, albeit the impact on some individuals was large. As ectodermal dysplasias are congenital genetic conditions affecting the development and/or homeostasis of two or more ectodermal derivatives, including hair, teeth, nails, and certain glands, we analyzed congenital nail disorders detected in these patients. Dystrophic or otherwise abnormal nails were evident in 17 of 18 subjects with pathogenic WNT10A or GJB6 variants but in none of 161 children with EDA variants underlying X-linked hypohidrotic ectodermal dysplasia. However, 2 of 17 children who carry mutations in EDAR or EDARADD, two other genes involved in the ectodysplasin A signaling pathway, showed nail abnormalities, such as brittle or hypoplastic nails. TP63 variants were regularly associated with nail disorders. In one girl, anonychia congenita caused by a compound heterozygous variant of the R-spondin-4 gene (RSPO4) was diagnosed. Thus, nail dysplasia is rarer among patients with ectodermal dysplasia than commonly thought.

Hypoxia-Mediated Soluble Fms-Like Tyrosine Kinase 1 Increase Is Not Attenuated in Interleukin 6-Deficient Mice.

The soluble fms-like tyrosine kinase 1 (sFlt-1), known to be increased in the serum of preeclamptic patients, is a relevant factor in causing maternal symptoms like hypertension and proteinuria. In this study, we aimed to reveal whether hypoxia is a cause of increased sFlt-1 levels and inflammation markers in vivo and whether these symptoms can be attenuated by interleukin 6 (IL-6) depletion. For this purpose, pregnant wild-type (wt) mice or IL-6(-/-) mice on embryonic day 16 were placed under either normoxic (20.9% oxygen) or hypoxic (6% oxygen) conditions for 6 hours. This led to a rise of sFlt-1 levels in maternal serum, independent of the IL-6 status of the dam. Increased maternal sFlt-1 serum levels were, however, not due to an increase in sFlt-1 messenger RNA levels in the placenta. Moreover, there was no increase in inflammatory markers in neither wt mice nor IL-6(-/-) mice. This suggests that hypoxia alone does not contribute to the induction of an inflammatory placenta. Also, the hypoxia-induced rise in sFlt-1 levels seems not to be mediated by IL-6 in vivo.

Improved closure of patent ductus arteriosus with high doses of ibuprofen.

The patent ductus arteriosus (PDA) is associated with various complications of prematurity. Cyclooxygenase-inhibitors are the first-line intervention for closure of the PDA. However, the rates of PDA closure still are unsatisfactory. Therefore, an individual trial was performed by changing the strategy for treating neonates with ibuprofen to induce the closure of PDA. In a retrospective study, patients receiving 20, 10, and 10 mg/kg bodyweight ibuprofen (group 1) were compared by chart review with those receiving 10, 5, 5 mg/kg (group 2). The rate of PDA closure, the incidence of side effects related to the use of ibuprofen, and the need for surgical intervention for closure of the PDA were analyzed. A higher rate of closure after three doses in group 1 could be observed (60.9 vs 52.6%; p = 0.75), which was not significant but indicated a clear positive trend. If closure of the PDA was unsuccessful, intravenous ibuprofen was continued for an additional 2 days. After 5 days, 91.3% of PDA in group 1 was closed compared with 68.4% PDA in group 2. In summary, only 8.7% of the group 1 neonates needed surgical closure of PDA after insufficient medicamentous closure compared with 31.6% in group 2 (p = 0.25). Although not statistically significant, a clear positive trend for using the higher-dose medication can be seen. More work dealing with the limitations of a retrospective study must be done. Based on the data from this study, high-dose ibuprofen seems able to increase the rate of effective medicamentous PDA closure without any further unwanted side effects.

Upregulation of leptin-receptor in placental cells by hypoxia.

OBJECTIVE: Leptin and its receptor (Ob-R) are co-expressed in human placenta suggesting auto- and paracrine mechanisms of the hormone. So far it is unclear, how changes in the placental environment affect Ob-R expression. Hence, the main purpose of the study was to investigate leptin receptor expression and regulation under hypoxic conditions. The influences of hypoxia and leptin on signal transduction and cell proliferation in chorioncarcinoma cell lines as well as primary villous trophoblasts were determined. RESULTS: We found a time-dependent induction of leptin receptor mRNA and protein in placental cells under hypoxic conditions. In contrast, soluble leptin receptor expression did not change under oxygen deprivation. Leptin treatment neither activated the p42/p44 nor the STAT3 pathway in placental cells, being independent of hypoxic or normoxic conditions. Furthermore, leptin added to the culture medium in high concentrations was unable to interfere with the rate of proliferation. CONCLUSION: Our data demonstrate that hypoxia leads to an increase of Ob-R expression in placental cells. Interestingly, leptin-dependent signal transduction and proliferation remained unaffected. A possible role of the soluble leptin receptor in modulating free leptin levels will be discussed.

Altered leptin secretion in hyperinsulinemic mice under hypoxic conditions.

OBJECTIVE: Hypoxia and insulin are known key players in the activation leptin transcription and translation in vivo and in vitro. These insulin- and hypoxia-dependent effects are leptin transcription are mediated via independent elements on the leptin-promotor, even more coincubation of the two stimuli in vitro results in a supraadditive effect on leptin transcription. The aim of this study was to examine whether hyperinsulinemia is able to interfere with the hypoxia-driven expression of leptin in adipose and extra-adipose tissue in vivo. RESEARCH METHODS AND PROCEDURES: We used the KK/HlJ mouse strain as a model for hyperinsulinemia and C57BL/6J mice as control. These two groups were exposed to hypoxia for 12 h. Serum levels of insulin and leptin were analyzed by ELISA, mRNA expression of leptin was measured via real-time PCR. RESULTS: In the hyperinsulinemic KK/HlJ mice, hypoxia was not able to further increase the amount of leptin in serum. Instead, a significant decrease of insulin levels was detected, while serum leptin and insulin levels increased in C57BL/6J. Analysis of leptin mRNA expression in subcutaneous fat, mesenteric fat and kidney revealed that hypoxia induces leptin transcription in kidneys of C57/BL6 but not in hyperinsulinemic animals. In contrast, leptin expression in adipose tissue was not increased during hypoxia. DISCUSSION: We conclude that leptin regulation during hypoxia in vivo depends at least in part on the modulating role of insulin. The hypoxia driven induction of insulin expression in C57/BL6 animals may be responsible for the stimulation of leptin transcription. In contrast, already hyperinsulinemic animals showed no induction - neither of insulin nor leptin after short-term hypoxia.

Transient growth hormone therapy to rats with low protein-inflicted intrauterine growth restriction does not prevent elevated blood pressure in later life.

Intrauterine growth restriction (IUGR) is a risk factor for the development of hypertension in later life. Insulin-like growth factor I and growth hormone (GH) have the potential to improve metabolic syndrome after IUGR in adult animals. The objective of the present study was to examine whether transient GH treatment of pups after weaning can prevent the development of arterial hypertension in adult rats. IUGR was induced in Wistar rats by isocaloric protein restriction in pregnant dams and litter size was reduced to six male neonates after birth. Recombinant human GH was applied by daily subcutaneous injections at a dose of 3 microg/g body weight between days 24 and 60 of life. Control animals received vehicle treatment (VEH) only. Birth weight was significantly lower in low protein (LP) animals than in normal protein (NP) animals (5.1 +/- 0.3 g vs. 5.9 +/- 0.7 g, p < 0.05). Until weaning at day 23, LP animals reached similar body length, but had reduced body weight compared to NP animals. Intraarterially measured mean arterial blood pressure at day 120 was elevated in LP-VEH compared to NP-VEH animals (113 +/- 6 mmHg vs. 101 +/- 6 mmHg, p < 0.01). However, transient GH-treatment did not prevent arterial hypertension in LP animals (112 +/- 5 mmHg). Our data suggest that GH treatment between days 24 and 60 of life does not or at least not permanently reprogram blood pressure elevation after IUGR.

Cortisol-cortisone ratios in small for gestational age (SGA) children without postnatal catch-up growth.

OBJECTIVE: Low birthweight is a risk factor for metabolic and cardiovascular disorders in later adult life. Changes in the activity of 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2) and the consequent disequilibrium between cortisol (F) and cortisone (E) are thought to be a key mechanism for these effects. We investigated whether prenatal programming leads to alterations in F/E ratios on a systemic level. DESIGN, PATIENTS AND METHODS: In a cross-sectional, retrospective study we analysed sera of 132 children born small for gestational age (SGA) (aged 2-13 years) with persistent short stature [< -2 standard deviation score (SDS)] and of 25 children born appropriate for gestational age (AGA) (aged 4-11 years) with normal body height. Thirty-one per cent of the SGA and 44% of the AGA children were born preterm. Serum E and F concentrations were measured using tandem mass spectrometry. To exclude species-specific effects, we studied the 11beta-HSD system by measuring the ratio of corticosterone (B) to dehydrocorticosterone (11OH-B) in rats that were born SGA after protein restriction of the female dams during pregnancy. RESULTS: F, E and the F/E ratio in serum did not differ in these children when comparing SGA to children who were born AGA and had normal height. The concentrations were independent of weight and length SDS at birth as well as gestational age. In rats born SGA, the B/11OH-B ratio was not different to that in normal control animals at 6, 11 and 15 weeks of life. CONCLUSION: We found no alterations in systemic cortisol-cortisone conversion either in short children born SGA or in SGA rats. However, local modifications of the 11beta-HSD system may be possible.

Fusiogenic endogenous-retroviral syncytin-1 exerts anti-apoptotic functions in staurosporine-challenged CHO cells.

Fusiogenic glycoprotein syncytin-1, expressed in human placenta, is a promising candidate for acquiring a basic knowledge of placental syncytialization. However, its cellular mode of action is unidentified. We investigated whether syncytin-1 may exert influence on apoptotic processes. Therefore, we incubated CHO cells after stable transfection with syncytin-1 (CHO-52) in the presence or absence of staurosporine (STS), a kinase inhibitor well characterized to induce apoptosis. When testing the phenotype of CHO-52 cells, we could demonstrate that the induction of apoptosis by STS was delayed over a period of up to 24 h. Furthermore, the cell death rate was decreased by approx 75% following transfection of syncytin-1 in CHO-52 compared to mock-treated cells. In detail, after 18h of incubation with 500 nM STS, 64 +/- 2% of CHO-52 cells were viable compared to 16 +/- 1% of CHO-mocks, after 24 h 43 +/- 3% vs 5 +/- 2%, respectively. CHO-52 cells exhibited a lower expression of active caspase 3 and anti-apoptotic Bcl-2 was found to be increased in CHO-52 cells at baseline and following STS treatment. Our study provides first evidence that syncytin-1 serves anti-apoptotic function under certain conditions. A lessened activation of caspase 3 and an increased expression of Bcl-2 are possible mechanisms.

Hypertonic-hyperoncotic solutions improve cardiac function in children after open-heart surgery.

OBJECTIVES: Hypertonic-hyperoncotic solutions are used for the improvement of micro- and macrocirculation in various types of shock. In pediatric intensive care medicine, controlled, randomized studies with hypertonic-hyperoncotic solutions are lacking. Hypertonic-hyperoncotic solutions may improve cardiac function in children. The primary objective of this controlled, randomized, blinded study was to evaluate the hemodynamic effects and safety of hypertonic-hyperoncotic solution infusions in children shortly after open-heart surgery for congenital cardiac disease. The secondary objective was to determine whether the administration of hypertonic-hyperoncotic solutions could be a potential and effective therapeutic option for preventing a probable capillary leakage syndrome that frequently occurs in children after open-heart surgery. METHODS: The children were randomly assigned to 2 groups of 25. The hypertonic-hyperoncotic solution group received Poly-(O-2)-hydroxyethyl-starch 60.0 g, with molecular weight of 200 kDa, Na+ 1232 mmol/L and osmolality of 2464 mOsmol/L (7.2% sodium chloride with 6% hydroxyethyl-starch 200 kDa). The isotonic saline solution group received isotonic saline solution (0.9% sodium chloride). Atrial and ventricular septal defects were corrected using a homograft patch. Monitoring consisted of an arterial, a central venous, and a thermodilution catheter (PULSIOCATH). Cardiac index, extravascular lung water index, stroke volume index, mean arterial blood pressure, and systemic vascular resistance index were measured (Pulse Contour Cardiac Output technique). Immediately after surgery, patients were loaded either with hypertonic-hyperoncotic solution or with isotonic saline solution (4 mL/kg). Blood samples (sodium concentration, osmolality, thrombocyte count, fibrinogen, and arterial blood gases) were drawn directly before; immediately after; 15 minutes after; and, 1, 4, 12, and 24 hours after the end of volume loading. Hemodynamic parameters were registered at the same time. The total amount of dobutamine required was documented, as well as the 24- and 48-hour fluid balances. RESULTS: In the hypertonic-hyperoncotic solution group, cardiac index was 3.6 +/- 0.26 L/min per m2 before volume administration and increased to 5.96 +/- 0.27 after the administration of the study solution (64%). Fifteen and 60 minutes after administration, the cardiac index remained significantly elevated (5.55 +/- 0.29 L/min per m2 and 4.65 +/- 0.18 L/min per m2, respectively) and returned to preadministration values after 4 hours. In the isotonic saline solution group, the cardiac index did not change during the entire observation period (3.39 +/- 0.21 before and 3.65 +/- 0.23 L/min per m2 after isotonic saline solution). The systemic vascular resistance index decreased in the hypertonic-hyperoncotic solution group after administration from 1396 +/- 112 to 868 +/- 63 dyn/sec per cm(-5)/m2. The decrease of systemic vascular resistance index in the hypertonic-hyperoncotic solution group was transiently significant within 60 minutes after administration but stayed lower than before volume load (999 +/- 70 dyn/sec per cm-(5)/m2). In the isotonic saline solution group, we found no statistically relevant change in systemic vascular resistance index. Stroke volume index significantly increased after hypertonic-hyperoncotic solution infusion (53.9 +/- 3.0 mL/m2 directly after, 48.8 +/- 2.46 mL/m2 15 minutes after, and 41.4 +/- 2.2 mL/m2 60 minutes after) when compared with stroke volume index before administration (32.4 +/- 2.6 mL/m2). In the hypertonic-hyperoncotic solution group, an increase in mean arterial blood pressure remained transiently significant within 60 minutes after administration when compared with the isotonic saline solution group, in which the mean arterial blood pressure remained unchanged. Both central venous pressure and heart rate were unchanged during the whole time of observation in both groups. In the hypertonic-hyperoncotic solution group, extravascular lung water index decreased from 10.6 +/- 1.2 to 5.6 +/- 1.2 mL/kg and remained significantly decreased 15 minutes after (6.5 +/- 1.2 mL/kg) when compared with before volume administration. In the isotonic saline solution group, extravascular lung water index increased from 12.3 +/- 1.1 mL/kg to 18.1 +/- 1.7 mL/kg directly after administration and remained elevated for 60 minutes after volume loading (15.6 +/- 1.5 mL/kg). In all patients, no hypoxia (Pa(O2)<60 mm Hg) or hypercapnia (Pa(CO2) >60 mm Hg) was observed. Arterial blood gas analysis showed pH and base excess within physiologic range, and this did not change throughout the whole period of observation. After infusion of hypertonic-hyperoncotic solution, sodium concentration increased from 139.2 +/- 0.7 to 147.5 +/- 0.7 mmol/L. The maximum sodium concentration was 153 mmol/L, measured immediately after hypertonic-hyperoncotic solution in 1 patient. The total amount of fluid infused was similar in both groups. The postoperative need for infused dobutamine in the patients in the hypertonic-hyperoncotic solution group was decreased compared with the isotonic saline solution group (46.9 +/- 8.8 microg/kg vs 308.2 +/- 46.6 microg/kg). No patient presented with severe bleeding. Short- and long-term cardiac and neurologic outcome was not reduced and all patients left the hospital in a clinically sufficient state. DISCUSSION: This study demonstrates a profound increase of cardiac index after the administration of hypertonic-hyperoncotic solution in children after uncomplicated open-heart surgery, suggesting a positive inotropic effect. The total amount of catecholamine was lower, assuming that hypertonic-hyperoncotic solution reduces the need for positive inotropic support. The observed positive cardiac effect of hypertonic-hyperoncotic solution may even be intensified by the decreased afterload (decreased systemic vascular resistance index). According to the Frank-Starling relation, an effective tool in the treatment of low cardiac output are an elevated preload while afterload is diminished. Therefore, we postulate that hypertonic-hyperoncotic solution may be helpful in preventing or attenuating low cardiac output failure in childhood. Capillary leakage syndrome also is a frequent problem after cardiopulmonary bypass. For quantification of edema formation, extravascular lung water index measurement is a useful tool. Using this approach, we provided evidence that the infusion of hypertonic-hyperoncotic solution is transiently able to reduce extravascular lung water index. This reduction was transient but might prevent the triggering of a clinically relevant capillary leakage syndrome. This is in line with in vitro studies demonstrating that hypertonic-hyperoncotic solution improves microcirculation by reducing vascular permeability. The single administration of hypertonic-hyperoncotic solution infusion was safe, and no adverse effects, such as hemostatic disturbances, were observed. CONCLUSIONS: A single infusion of hypertonic-hyperoncotic saline solution after cardiac surgery is safe despite the hypertonicity and the colloid component of the hypertonic-hyperoncotic saline solution. In children after cardiopulmonary bypass surgery, the administration of hypertonic-hyperoncotic saline solution increased cardiac index by elevating stroke volume index in combination with a lowered systemic vascular resistance index. Extravascular lung water index transiently decreased, suggesting that hypertonic-hyperoncotic saline solution effectively counteracts the capillary leakage that often occurs after cardiac surgery in children. Additional investigations might elucidate whether the temporary effects of hypertonic-hyperoncotic saline solution are beneficial in the treatment of severe capillary leakage after complicated cardiac surgery. It has to be shown that hypertonic-hyperoncotic saline solution is a long-lasting, effective treatment strategy for low cardiac output failure in children that is caused by sepsis, multiorgan failure, and endothelial edema. We have provided evidence to pediatric intensive care clinicians that the single administration of hypertonic-hyperoncotic saline solution might be a useful and safe treatment in the amelioration of contractility, inotropy, and the possible treatment of early-onset capillary leakage.

Hypoxia-induced leptin production in human trophoblasts does not protect from apoptosis.

OBJECTIVE: The ob-gene product, leptin, is an important regulator of placental and fetal development during pregnancy. Leptin, being induced by hypoxia in the placenta, is a known pro-apoptotic molecule in adipose tissue but is also known to inhibit apoptosis in other tissues like neuroblastoma cells. Based on these findings, we investigated if leptin has a pro- or anti-apoptotic effect on a trophoblastic cell line (JAr cells) in the presence or absence of oxygen. METHODS AND RESULTS: Measurement of leptin in the supernatant by using ELISA showed hypoxia-induced leptin production in JAr cells in vitro. This could be confirmed by a leptin-specific RT-PCR. By analyzing leptin and/or hypoxia exposed cells with FACS cytometry we found that JAr cells can cope with hypoxia down to oxygen tensions of 1%. At this level, only a small number of cells underwent apoptosis. Interestingly, leptin added to the culture medium in high concentrations was not able to interfere with the rate of proliferation or apoptosis in these cells independent of the oxygen tension. Finally, an anti-caspase-3 and anti-caspase-9 Western blot was performed. Again, no difference in the expression of caspase-3 and -9 under the conditions tested was seen. CONCLUSIONS: These results show that leptin, produced by placental cells after hypoxia in vitro, has no influence on the rate of proliferation of these cells. Furthermore, it does not influence apoptotic pathways in the trophoblastic cell line tested under hypoxic and non-hypoxic conditions.

Alterations of leptin and ghrelin serum concentrations in renal disease: simple epiphenomena?

The hypothesis that alterations of serum concentrations of the anorexigenic adipose tissue-derived hormone leptin or the orexigenic peptide ghrelin might help to regain appetite and fight malnutrition in patients with chronic renal failure cannot be confirmed at present. For the future, however, strategies interfering with signal transduction of these peptides in the hypothalamus might be more promising and should be investigated and further developed.

Regulation and signal transduction of toll-like receptors in human chorioncarcinoma cell lines.

PROBLEM: Toll-like receptors (TLRs) are expressed on placental cells. The aim of this study is to analyze signaling components activated in placenta cells after TLR ligand engagement. METHODS OF STUDY: In chorioncarcimoma cell lines the regulation of TLRs was determined by real time polymerase chain reaction as well as by fluorescence-activated cell sorter analysis. Activation of NF-kappaB was determined in a reporter assay system and the activation of the mitogen-activated protein kinase signaling pathways by immunoblot analysis. RESULTS: Both lipopolysaccharide (LPS) and DNA oligonucleotides containing unmethylated CpG motifs (CpG) induced the enhanced expression of TLR2 mRNA as well as a TLR2 surface protein expression. Functionally, incubation of JAR cells with microbial stimuli such as LPS activated NF-kappaB, as well as the phosphorylation of ERK1/2 and p38 MAP kinases and secretion of interleukin-8. CONCLUSION: The functional expression of TLRs on placental cells may play an important role in the initiation of an immune response in the developing fetus.

Differential regulation of leptin synthesis in rats during short-term hypoxia and short-term carbon monoxide inhalation.

Leptin is a circulating hormone that is secreted primarily by adipose tissue. However, recent studies have demonstrated leptin production by other tissues, including placenta, stomach, kidney, liver, and lung, a process not only activated by stimuli such as insulin or corticosteroids, but also by hypoxia, which is mediated by the hypoxia inducible factor-1. In contrast to this fact, smokers have lower plasma leptin levels. The purpose of this study was to determine whether tissue hypoxygenation [induced by lack of oxygen] or inhalation of carbon monoxide (CO) are sufficient to up-regulate leptin in fat cells as well as in peripheral organs such as lung, liver, and kidney of rats. In hypoxic rats, leptin expression was unchanged or even reduced in adipose tissue. In contrast, in liver, kidney, and lung we observed an increase in leptin expression compared with normoxic controls, whereas plasma levels were unchanged. When animals were exposed to CO, generating a functional anemia known to activate the HIF-1-dependent transcription, a significant decrease in leptin gene expression in adipose tissue and in all organs tested was observed. Plasma leptin concentrations after CO exposure were significantly diminished compared with those in control animals. These findings suggest that tissue hypoxygenation up-regulates leptin expression in nonadipose tissue. However, this is not sufficient to raise plasma leptin levels in rats. Inhalation of CO leads to a significant decrease in leptin mRNA and protein concentration in the plasma of the animals, suggesting a negative effect of CO on leptin transcription.

Transcriptional effects of hypoxia on fusiogenic syncytin and its receptor ASCT2 in human cytotrophoblast BeWo cells and in ex vivo perfused placental cotyledons.

OBJECTIVE: The purpose of this study was to test the hypothesis that hypoxia down-regulates placental syncytin, which could play a role in altered placentogenesis; we investigated the influence of hypoxia on syncytin and its receptor ASCT2 gene expression in BeWo cells and in ex vivo perfused human cotyledons. STUDY DESIGN: BeWo cells were incubated with deferoxamine or cobalt chloride under normoxia and hypoxia. Additionally, a model of dually ex vivo perfused cotyledons was applied. Under hypoxic and cobalt chloride stimuli syncytin, ASCT2, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta-actin, and beta(2)-microglobulin (beta(2)-MG) messenger RNAs were analyzed with the use of real-time polymerase chain reaction. RESULTS: Hypoxia, deferoxamine, and cobalt chloride markedly decreased syncytin messenger RNA in BeWo cells, whereas ASCT2 messenger RNA was not altered significantly. In isolated perfused cotyledons, hypoxia also reduced syncytin (P<.05) but not ASCT2 messenger RNA. CONCLUSION: Our data provide first evidence that syncytin gene expression is down-regulated by hypoxia, which strengthens the hypothesis that syncytin is reduced in disturbed pregnancies in the course of placental hypoxia.