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CXCL12-CXCR4 signaling controls multiple physiological processes and its dysregulation is associated with cancers and inflammatory diseases. To discover as-yet-unknown endogenous ligands of CXCR4, we screened a blood-derived peptide library for inhibitors of CXCR4-tropic HIV-1 strains. This approach identified a 16 amino acid fragment of serum albumin as an effective and highly specific CXCR4 antagonist. The endogenous peptide, termed EPI-X4, is evolutionarily conserved and generated from the highly abundant albumin precursor by pH-regulated proteases. EPI-X4 forms an unusual lasso-like structure and antagonizes CXCL12-induced tumor cell migration, mobilizes stem cells, and suppresses inflammatory responses in mice. Furthermore, the peptide is abundant in the urine of patients with inflammatory kidney diseases and may serve as a biomarker. Our results identify EPI-X4 as a key regulator of CXCR4 signaling and introduce proteolysis of an abundant precursor protein as an alternative concept for chemokine receptor regulation.
Background and objective Whether treatment with vitamin D receptor activators contributes to cardiovascular disease in patients with chronic kidney disease is a matter of debate. We studied mechanisms involved in vitamin D-related vascular calcifications in vivo and in vitro.
Methods Aortic calcifications were induced in subtotally nephrectomized (SNX) rats by treatment with a high dose (0.25 mu g/kg per day) of 1,25-dihydroxyvitamin D-3 (calcitriol) given for 6 weeks. Likewise, primary rat vascular smooth muscle cells (VSMCs) were incubated with calcitriol at concentrations ranging from 10(-11) to 10(-7) mol/l. Immunohistochemistry revealed that the aortic expression of osteopontin, osteocalcin and bone sialoprotein was significantly increased in calcitriol-treated SNX rats compared to untreated SNX controls. In addition, aortic expression of the transient receptor potential vanilloid calcium channel 6 (TRPV6) and calbindin D9k was significantly up-regulated by treatment with calcitriol. Furthermore, calcitriol significantly increased expression of the osteogenic transcription factor osterix. In-vitro studies showed similar results, confirming that these effects could be attributed to treatment with calcitriol.
Conclusions High-dose calcitriol treatment induces an osteoblastic phenotype in VSMC both in SNX rats and in vitro, associated with up-regulation of proteins regulating mineralization and calcium transport, and of the osteogenic transcription factor osterix.
Starch is one of the most popular nutritional sources for both human and animals. Due to the variation of its nutritional traits and biochemical specificities, starch has been classified into rapidly digestible, slowly digestible and resistant starch. Resistant starch has its own unique chemical structure, and various forms of resistant starch are commercially available. It has been found being a multiple-functional regulator for treating metabolic dysfunction. Different functions of resistant starch such as modulation of the gut microbiota, gut peptides, circulating growth factors, circulating inflammatory mediators have been characterized by animal studies and clinical trials. In this mini-review, recent remarkable progress in resistant starch on gut microbiota, particularly the effect of structure, biochemistry and cell signaling on nutrition has been summarized, with highlights on its regulatory effect on gut microbiota.
Aims:This study aims to investigate the relationship between plasma endothelin-1 (ET-1) concentrations, ET-1 gene polymorphisms in loci rs5370, rs1630736, 3A/4A and clinical features of primary nephrotic syndrome (NS) in children.
Materials and methods: Thirty-six children with primary NS were selected as case group, and 94 healthy children were selected as control group. All subjects were genotyped for three single nucleotide polymorphisms (SNPs) (rs5370, rs10478694 [3A4A) and rs 1630736) in the ET-1 gene by gene sequencing. The plasma ET-1 concentrations were measured using a radio-immunoassay.
Key findings: Plasma ET-1 concentrations were higher in NS patients (P = 0.007) as compared to healthy children. The allele frequencies between control and NS patients were significantly different only with respect to the rs10478694 SNP of the ET-1 gene. The allele frequencies between control and NS patients for the rs5370 SNP showed a trend towards difference (P = 0.057). Plasma cholesterol in NS patients is associated with both: the Cl genotype in locus rs5370 and the 3A4A genotype in locus rs10478694 (P < 0.05 in both cases).
Significance: The ET systems might play a disease modifying role in pediatric NS. Plasma cholesterol, a hallmark of NS. seems to be associated with genetic variations within the human ET-1 gene. (C) 2014 Elsevier Inc. All rights reserved.
Background/Aims: A recent study revealed that global overexpression of ET-1 causes a slight reduction in systemic blood pressure. Moreover, heterozygous ET-1 knockout mice are hypertensive. The role of ET-1 in human hypertension was so far not addressed by a strict meta-analysis of published human clinical studies.
Methods: We included studies published between January 1, 1990 and February 28, 2017. We included case control studies analyzing untreated essential hypertension or hypertensive patients where antihypertensive medication was discontinued for at least two weeks. Based on the principle of Cochrane systematic reviews, case control studies (CCSs) in PubMed (Medline) and Google Scholar designed to identify the role of endothelin-1 (ET-1) in the pathophysiological of hypertension were screened. Review Manager Version 5.0 (Rev-Man 5.0) was applied for statistical analysis. Mean difference and 95% confidence interval (CI) were shown in inverse variance (IV) fixed-effects model or IV random-effects models.
Results: Eleven studies fulfilling our in-and exclusion criteria were eligible for this meta-analysis. These studies included 450 hypertensive patients and 328 controls. Our meta-analysis revealed that ET-1 plasma concentrations were higher in hypertensive patients as compared to the control patients [mean difference between groups 1.57 pg/mL, 95%Ci [0.47 similar to 2.68, P = 0.005]. These finding were driven by patients having systolic blood pressure higher than 160 mmHg and diastolic blood pressure higher than 100 mmHg.
Conclusions: This meta-analysis showed that hypertensive patients do have elevated plasma ET-1 concentrations. This finding is driven by those patients with high systolic/diastolic blood pressure. Given that the ET-1 gene did not appear in any of the whole genome association studies searching for hypertension associated gene loci, it is very likely that the elevated plasma ET-1 concentrations in hypertensive patients are secondary to hypertension and may reflect endothelial cell damage.
Objectives: Stroke, frequently a consequence of hypertension, is one of the leading causes of death and neurological disabilities worldwide. In the ischemic brain, levels of endothelin-1, one of the most potent vasoconstrictors, are raised. Anti-inflammatory and neuroprotective effects of endothelin antagonists after stroke have been described in literature. Based on these findings, we investigated the protective effect of the endothelin converting enzyme/neutral endopeptidase blocker, SLV 338, in salt-loaded, stroke-prone, spontaneously hypertensive rats.
Methods: Male, 8-week-old spontaneously hypertensive stroke-prone rats were put on a high salt diet and treated with either 30 mg/kg or 100 mg/kg SLV 338 or vehicle for 27 weeks. Blood pressure, neurological outcome, body weight, and mortality were investigated throughout treatment. In weeks 1 and 9, animals were housed in metabolic cages for collection of urinary and blood samples and assessment of salt water and food intake. In weeks 22 and 27, additional blood samples were taken. At the end of the study, all brains were analyzed using magnetic resonance imaging.
Results: SLV 338 was well tolerated in all animals. Neurological outcome and infarct size were similar in all groups. Albuminuria was considerably delayed and the incidence of stroke significantly lowered in treated animals. In spontaneously hypertensive stroke-prone rats, treatment with SLV 338 significantly (P=0.01) improved survival in comparison to the vehicle treated group in a blood pressure-independent manner.
Discussion: Our data in spontaneously hypertensive stroke-prone rats demonstrate that combined endothelin converting enzyme/neutral endopeptidase inhibition could offer a new therapeutic approach for primary stroke prevention and improvement of mortality. The mechanism seems to be blood pressure-independent.
Birth weight variation is influenced by fetal and maternal genetic and non-genetic factors, and has been reproducibly associated with future cardio-metabolic health outcomes. In expanded genome-wide association analyses of own birth weight (n = 321,223) and offspring birth weight (n = 230,069 mothers), we identified 190 independent association signals (129 of which are novel). We used structural equation modeling to decompose the contributions of direct fetal and indirect maternal genetic effects, then applied Mendelian randomization to illuminate causal pathways. For example, both indirect maternal and direct fetal genetic effects drive the observational relationship between lower birth weight and higher later blood pressure: maternal blood pressure-raising alleles reduce offspring birth weight, but only direct fetal effects of these alleles, once inherited, increase later offspring blood pressure. Using maternal birth weight-lowering genotypes to proxy for an adverse intrauterine environment provided no evidence that it causally raises offspring blood pressure, indicating that the inverse birth weight-blood pressure association is attributable to genetic effects, and not to intrauterine programming.
Background/Aims: Angiogenesis plays a key role during embryonic development. The vascular endothelin (ET) system is involved in the regulation of angiogenesis. Lipopolysaccharides (LPS) could induce angiogenesis. The effects of ET blockers on baseline and LPS-stimulated angiogenesis during embryonic development remain unknown so far. Methods: The blood vessel density (BVD) of chorioallantoic membranes (CAMs), which were treated with saline (control), LPS, and/or BQ123 and the ETB blocker BQ788, were quantified and analyzed using an IPP 6.0 image analysis program. Moreover, the expressions of ET-1, ET-2, ET3, ET receptor A (ETRA), ET receptor B (ETRB) and VEGFR2 mRNA during embryogenesis were analyzed by semi-quantitative RT-PCR. Results: All components of the ET system are detectable during chicken embryogenesis. LPS increased angiogenesis substantially. This process was completely blocked by the treatment of a combination of the ETA receptor blockers-BQ123 and the ETB receptor blocker BQ788. This effect was accompanied by a decrease in ETRA, ETRB, and VEGFR2 gene expression. However, the baseline angiogenesis was not affected by combined ETA/ETB receptor blockade. Conclusion: During chicken embryogenesis, the LPS-stimulated angiogenesis, but not baseline angiogenesis, is sensitive to combined ETA/ETB receptor blockade. (C) 2018 The Author(s) Published by S. Karger AG, Basel
Background/Aims: Angiogenesis plays a key role during embryonic development. The vascular endothelin (ET) system is involved in the regulation of angiogenesis. Lipopolysaccharides (LPS) could induce angiogenesis. The effects of ET blockers on baseline and LPS-stimulated angiogenesis during embryonic development remain unknown so far. Methods: The blood vessel density (BVD) of chorioallantoic membranes (CAMs), which were treated with saline (control), LPS, and/or BQ123 and the ETB blocker BQ788, were quantified and analyzed using an IPP 6.0 image analysis program. Moreover, the expressions of ET-1, ET-2, ET3, ET receptor A (ETRA), ET receptor B (ETRB) and VEGFR2 mRNA during embryogenesis were analyzed by semi-quantitative RT-PCR. Results: All components of the ET system are detectable during chicken embryogenesis. LPS increased angiogenesis substantially. This process was completely blocked by the treatment of a combination of the ETA receptor blockers-BQ123 and the ETB receptor blocker BQ788. This effect was accompanied by a decrease in ETRA, ETRB, and VEGFR2 gene expression. However, the baseline angiogenesis was not affected by combined ETA/ETB receptor blockade. Conclusion: During chicken embryogenesis, the LPS-stimulated angiogenesis, but not baseline angiogenesis, is sensitive to combined ETA/ETB receptor blockade.
Purpose of reviewIncretin-based therapy with glucagon-like peptide-1 receptor (GLP-1R) agonists and dipeptidyl peptidase-4 (DPP-4) inhibitors is considered a promising therapeutic option for type 2 diabetes mellitus. Cumulative evidence, mainly from preclinical animal studies, reveals that incretin-based therapies also may elicit beneficial effects on kidney function. This review gives an overview of the physiology, pathophysiology, and pharmacology of the renal incretin system.Recent findingsActivation of GLP-1R in the kidney leads to diuretic and natriuretic effects, possibly through direct actions on renal tubular cells and sodium transporters. Moreover, there is evidence that incretin-based therapy reduces albuminuria, glomerulosclerosis, oxidative stress, and fibrosis in the kidney, partially through GLP-1R-independent pathways. Molecular mechanisms by which incretins exert their renal effects are understood incompletely, thus further studies are needed.SummaryThe GLP-1R and DPP-4 are expressed in the kidney in various species. The kidney plays an important role in the excretion of incretin metabolites and most GLP-1R agonists and DPP-4 inhibitors, thus special attention is required when applying incretin-based therapy in renal impairment. Preclinical observations suggest direct renoprotective effects of incretin-based therapies in the setting of hypertension and other disorders of sodium retention, as well as in diabetic and nondiabetic nephropathy. Clinical studies are needed in order to confirm translational relevance from preclinical findings for treatment options of renal diseases.
Observational studies from all over the world continue to find high prevalence rates of vitamin D insufficiency and deficiency in many populations, including pregnant women. Beyond its classical function as a regulator of calcium and phosphate metabolism, vitamin D elicits numerous effects in the human body. Current evidence highlights a vital role of vitamin D in mammalian gestation. During pregnancy, adaptations in maternal vitamin D metabolism lead to a physiologic increase of vitamin D levels, mainly because of an increased renal production, although other potential sources like the placenta are being discussed. A sufficient supply of mother and child with calcium and vitamin D during pregnancy ensures a healthy bone development of the fetus, whereas lack of either of these nutrients can lead to the development of rickets in the child. Moreover, vitamin D insufficiency during pregnancy has consistently been associated with adverse maternal and neonatal pregnancy outcomes. In multitudinous studies, low maternal vitamin D status was associated with a higher risk for pre-eclampsia, gestational diabetes mellitus and other gestational diseases. Likewise, several negative consequences for the fetus have been reported, including fetal growth restriction, increased risk of preterm birth and a changed susceptibility for later-life diseases. However, study results are diverging and causality has not been proven so far. Meta-analyses on the relationship between maternal vitamin D status and pregnancy outcomes revealed a wide heterogeneity of studied populations and the applied methodology in vitamin D assessment. Until today, clinical guidelines for supplementation cannot be based on high-quality evidence and it is not clear if the required intake for pregnant women differs from non-pregnant women. Long-term safety data of vitamin D supplementation in pregnant women has not been established and overdosing of vitamin D might have unfavorable effects, especially in mothers and newborns with mutations of genes involved in vitamin D metabolism. Reliable data from large observational and interventional randomized control trials are urgently needed as a basis for any detailed and safe recommendations for supplementation in the general population and, most importantly, in pregnant women. This is of utmost importance, as ensuring a sufficient vitamin D-supply of mother and child implies a great potential for the prevention of birth complications and development of diseases.
Aims: The nitric oxide and endothelin systems are key components of a local paracrine hormone network in the heart. We previously reported that diastolic dysfunction observed in mice lacking the endothelial nitric oxide synthase (eNOS-/-) can be prevented by a genetic overexpression of ET-1. Sexual dimorphisms have been reported in both ET-1 and NO systems. Particularly, eNOS-/- mice present sex related phenotypic differences.
Main methods: We used the ET-1 transgenic (ET+/+), eNOS-/-, and crossbred ET+/+ eNOS-/- mice, and wild type controls. We measured cardiac function by heart catheterization. Cardiac ventricles were collected for histological and molecular profiling.
Key findings: We report here that (i) the level of ET-1 expression in eNOS-/- mice was elevated in males but not in females. (ii) Left ventricular end-diastolic blood pressure was higher in male eNOS-/- mice than in females. (ii) eNOS-/- males but not females developed cardiomyocyte hypertrophy. (iv) Perivascular fibrosis of intra-cardiac arteries developed in female ET+/+ and eNOS-/- mice but not in males. Additionally, (v) the cardiac expression of metalloprotease-9 was higher in eNOS-/- males compared to females. Finally, (vi) cardiac proteome analysis revealed that the protein abundance of the oxidative stress related enzyme superoxide dismutase presented with sexual dimorphism in eNOS-/- and ET+/+ mice.
Significance: These results indicate that the cardiac phenotypes of ET-1 transgenic mice and eNOS knockout mice are sex specific. Since both systems are key players in the pathogenesis of cardiovascular diseases, our findings might be important in the context of gender differences in patients with such diseases. (C) 2013 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
Background The cardiac nitric oxide and endothelin-1 (ET-1) systems are closely linked and play a critical role in cardiac physiology. The balance between both systems is often disturbed in cardiovascular diseases. To define the cardiac effect of excessive ET-1 in a status of nitric oxide deficiency, we compared left ventricular function and morphology in wild-type mice, ET-1 transgenic (ET+/+) mice, endothelial nitric oxide synthase knockout (eNOS(-/-)) mice, and ET(+/+)eNOS(-/-) mice.
Methods and results eNOS(-/-) and ET(+/+)eNOS(-/-) mice developed high blood pressure compared with wild-type and ET+/+ mice. Left ventricular catheterization showed that eNOS(-/-) mice, but not ET(+/+)eNOS(-/-), developed diastolic dysfunction characterized by increased end-diastolic pressure and relaxation constant tau. To elucidate the causal molecular mechanisms driving the rescue of diastolic function in ET(+/+)eNOS(-/-) mice, the cardiac proteome was analyzed. Two-dimensional gel electrophoresis coupled to mass spectrometry offers an appropriate hypothesis-free approach. ET-1 overexpression on an eNOS(-/-) background led to an elevated abundance and change in posttranslational state of antioxidant enzymes (e. g., peroxiredoxin-6, glutathione S-transferase mu 2, and heat shock protein beta 7). In contrast to ET(+/+)eNOS(-/-) mice, eNOS(-/-) mice showed an elevated abundance of proteins responsible for sarcomere disassembly (e. g., cofilin-1 and cofilin-2). In ET(+/+)eNOS(-/-) mice, glycolysis was favored at the expense of fatty acid oxidation.
Conclusion eNOS(-/-) mice developed diastolic dysfunction; this was rescued by ET-1 transgenic overexpression. This study furthermore suggests that cardiac ET-1 overexpression in case of eNOS deficiency causes specifically the regulation of proteins playing a role in oxidative stress, myocytes contractility, and energy metabolism.
Background: Endothelin-1 (ET-1) is a multifunctional peptide, which is implicated in the renal and cardiac physicology as well as in many pathologies of these systems. ET-1's actions take place after the activation of two receptors: ETA and ETB. The expression of these receptors may be modulated during the pathologic process. The analysis of the distribution and level of expression of the receptors in animal models is therefore crucial.
Methods: We developed a protocol for non-radioactive in situ hybridization for the mRNA of the two endothelin receptors on paraffin-embedded tissue using digoxigenin-labeled RNA probes.
Results: In heart and kidney, the staining was reliable and specific. In a mouse model for endothelin/nitric oxide imbalance, cardiac ETB expression was reduced. The distribution of the receptors was in accordance with the actual knowledge. Differences in cell specific expression are discussed.
Conclusions: We developed a protocol for the in situ hybridization of the endothelin receptors in mice. Given that the endothelin system is implicated in the development of many diseases, we believe that this protocol may be useful for a number of future preclinical studies.
Background: To assess the chronic effect of the DPP-4 inhibitor, linagliptin, alone, in combination with exenatide, and during exenatide withdrawal, in diet-induced obese (DIO) rats.
Methods: Female Wistar rats were exposed to a cafeteria diet to induce obesity. Animals were then dosed with vehicle or linagliptin (3 mg/kg PO) orally once-daily for a 28 day period. In a subsequent study, rats received exenatide (either 3 or 30 mu g/kg/day) or vehicle by osmotic mini-pump for 28 days. In addition, groups of animals were dosed orally with linagliptin either alone or in combination with a 3 mu g/kg/day exenatide dose for the study duration. In a final study, rats were administered exenatide (30 mu g/kg/day) or vehicle by osmotic mini-pump for eleven days. Subsequently, exenatide-treated animals were transferred to vehicle or continued exenatide infusion for a further ten days. Animals transferred from exenatide to vehicle were also dosed orally with either vehicle or linagliptin. In all studies, body weight, food and water intake were recorded daily and relevant plasma parameters and carcass composition were determined.
Results: In contrast to exenatide, linagliptin did not significantly reduce body weight or carcass fat in DIO rats versus controls. Linagliptin augmented the effect of exenatide to reduce body fat when given in combination but did not affect the body weight response. In rats withdrawn from exenatide, weight regain was observed such that body weight was not significantly different to controls. Linagliptin reduced weight regain after withdrawal of exenatide such that a significant difference from controls was evident.
Conclusions: These data demonstrate that linagliptin does not significantly alter body weight in either untreated or exenatide-treated DIO rats, although it delays weight gain after exenatide withdrawal. This finding may suggest the utility of DPP-4 inhibitors in reducing body weight during periods of weight gain.
A novel common variant in DCST2 is associated with length in early life and height in adulthood
(2015)
Common genetic variants have been identified for adult height, but not much is known about the genetics of skeletal growth in early life. To identify common genetic variants that influence fetal skeletal growth, we meta-analyzed 22 genome-wide association studies (Stage 1; N = 28 459). We identified seven independent top single nucleotide polymorphisms (SNPs) (P < 1 x 10(-6)) for birth length, of which three were novel and four were in or near loci known to be associated with adult height (LCORL, PTCH1, GPR126 and HMGA2). The three novel SNPs were followed-up in nine replication studies (Stage 2; N = 11 995), with rs905938 in DC-STAMP domain containing 2 (DCST2) genome-wide significantly associated with birth length in a joint analysis (Stages 1 + 2; beta = 0.046, SE = 0.008, P = 2.46 x 10(-8), explained variance = 0.05%). Rs905938 was also associated with infant length (N = 28 228; P = 5.54 x 10(-4)) and adult height (N = 127 513; P = 1.45 x 10(-5)). DCST2 is a DC-STAMP-like protein family member and DC-STAMP is an osteoclast cell-fusion regulator. Polygenic scores based on 180 SNPs previously associated with human adult stature explained 0.13% of variance in birth length. The same SNPs explained 2.95% of the variance of infant length. Of the 180 known adult height loci, 11 were genome-wide significantly associated with infant length (SF3B4, LCORL, SPAG17, C6orf173, PTCH1, GDF5, ZNFX1, HHIP, ACAN, HLA locus and HMGA2). This study highlights that common variation in DCST2 influences variation in early growth and adult height.
Vitamin D, either in its D-2 or D-3 form, is essential for normal human development during intrauterine life, kidney function and bone health. Vitamin D deficiency has also been linked to cancer development and some auto immune diseases. Given this huge impact of vitamin Don human health, it is important for daily clinical practice and clinical research to have reliable tools to judge on the vitamin D status. The major circulating form of vitamin D is 25-hydroxyvitamin D (25(OH)D), although it is not the most active metabolite, the concentrations of total 25-hydroxyvitamin D in the serum are currently routinely used in clinical practice to assess vitamin D status. In the circulation, vitamin D - like other steroid hormones - is bound tightly to a special carrier - vitamin D-binding protein (DBP). Smaller amounts are bound to blood proteins - albumin and lipoproteins. Only very tiny amounts of the total vitamin D are free and potentially biologically active. Currently used vitamin D assays do not distinguish between the three forms of vitamin D - DBP-bound vitamin D, albumin-bound vitamin D and free, biologically active vitamin D. Diseases or conditions that affect the synthesis of DBP or albumin thus have a huge impact on the amount of circulating total vitamin D. DBP and albumin are synthesized in the liver, hence all patients with an impairment of liver function have alterations in their total vitamin D blood concentrations, while free vitamin D levels remain mostly constant. Sex steroids, in particular estrogens, stimulate the synthesis of DBP. This explains why total vitamin D concentrations are higher during pregnancy as compared to nonpregnant women, while the concentrations of free vitamin D remain similar in both groups of women. The vitamin D-DBP as well as vitamin D-albumin complexes are filtered through the glomeruli and re-uptaken by megalin in the proximal tubule. Therefore, all acute and chronic kidney diseases that are characterized by a tubular damage, are associated with a loss of vitamin D-DBP complexes in the urine. Finally, the gene encoding DBP protein is highly polymorphic in different human racial groups. In the current review, we will discuss how liver function, estrogens, kidney function and the genetic background might influence total circulating vitamin D levels and will discuss what vitamin D metabolite is more appropriate to measure under these conditions: free vitamin D or total vitamin D.
Background/Aims: To investigate the renal phenotype under conditions of an activated renal ET-1 system in the status of nitric oxide deficiency, we compared kidney function and morphology in wild-type, ET-1 transgenic (ET+/+), endothelial nitric oxide synthase knockout (eNOS-/-) and ET+/+eNOS-/- mice. Methods: We assessed blood pressure, parameters of renal morphology, plasma cystatin C, urinary protein excretion, expression of genes associated with glomerular filtration barrier and tissue remodeling, and plasma metabolites using metabolomics. Results: eNOS-/- and ET+/+eNOS-/- mice developed hypertension. Osteopontin, albumin and protein excretion were increased in eNOS-/- and restored in ET+/+eNOS-/- animals. All genetically modified mice developed renal interstitial fibrosis and glomerulosclerosis. Genes involved in tissue remodeling (serpinel, TIMP1, Collal, CCL2) were up-regulated in eNOS-/-, but not in ET+/+eNOS-/- mice. Plasma levels of free carnitine and acylcarnitines, amino acids, diacyl phosphatidylcholines, lysophosphatidylcholines and hexoses were descreased in eNOS-/- and were in the normal range in ET+/+eNOS-/- mice. Conclusion: eNOS-/- mice developed renal dysfunction, which was partially rescued by ET-1 overexpression in eNOS-/- mice. The metabolomics results suggest that ET-1 overexpression on top of eNOS knockout is associated with a functional recovery of mitochondria (rescue effect in 13-oxidation of fatty acids) and an increase in antioxidative properties (normalization of monounsaturated fatty acids levels). (C) 2015 The Author(s) Published by S. Karger AG, Basel
The determination of free 25-hydroxyvitamin D (25(OH)D) as compared to the analysis of total 25-hydroxyvitamin D might reflect better the vitamin D status during pregnancy, since vitamin D-binding protein (DBP) concentrations increase throughout pregnancy and the vast majority of 25(OH)D is tightly bound to DBP thus strongly influencing total 25(OH)D. The concentration of the biologically active free 25(OH)D - on the other hand - is much less dependent on the DBP concentrations. The study was conducted in May-June 2016 in 368 Caucasian pregnant healthy women - residents of Northeastern Germany. Free 25(OH)D was either measured directly by commercial ELISA kit or assessed by calculation via total 25(OH)D, DBP, and albumin serum concentrations. Regardless of the detection method, free 25(OH)D lowers in the 3rd trimester comparing to the 1st trimester (by 12% and 21%, p < 0.05 and p < 0.001, for measured and calculated free 25(OH)D, respectively), whereas total 25(OH)D was not decreased in late pregnancy. DBP rises with gestational age. Total 25(OH)D was not correlated with serum calcium (p = 0.251), whereas free 25(OH)D was significantly (p = 0.007 for measured free 25(OH)D and p < 0.001 for calculated free 25(OH)D) positively correlated with calcium. All 25(OH) D isoforms were significantly negatively correlated with bone-specific alkaline phosphatase (BSAP), however the correlation strength was the lowest with total 25(OH)D (rho = -0.108, p = 0.038), whereas both measured and calculated free 25(OH)D revealed better associations with BSAP (rho = -0.203 and rho = -0.211 for measured and calculated free 25(OH)D, respectively, p < 0.001 for both). We established pregnancy trimester specific reference intervals for free measured and calculated 25(OH)D and DBP. Both measured and calculated free 25(OH)D showed better correlations with parameters of the endocrine vitamin D system (calcium and BSAP). Both ways of measuring free 25(OH)D in pregnant women are suitable as novel laboratory parameter for vitamin D status monitoring during human pregnancy and might replace in the future the routine total 25(OH)D assessment.
Dipeptidyl peptidase (DPP)-4 inhibitors delay chronic kidney disease (CKD) progression in experimental diabetic nephropathy in a glucose-independent manner. Here we compared the effects of the DPP-4 inhibitor linagliptin versus telmisartan in preventing CKD progression in non-diabetic rats with 5/6 nephrectomy. Animals were allocated to 1 of 4 groups: sham operated plus placebo; 5/6 nephrectomy plus placebo; 5/6 nephrectomy plus linagliptin; and 5/6 nephrectomy plus telmisartan. Interstitial fibrosis was significantly decreased by 48% with linagliptin but a non-significant 24% with telmisartan versus placebo. The urine albumin-to-creatinine ratio was significantly decreased by 66% with linagliptin and 92% with telmisartan versus placebo. Blood pressure was significantly lowered by telmisartan, but it was not affected by linagliptin. As shown by mass spectrometry, the number of altered peptide signals for linagliptin in plasma was 552 and 320 in the kidney. For telmisartan, there were 108 peptide changes in plasma and 363 in the kidney versus placebo. Linagliptin up-regulated peptides derived from collagen type I, apolipoprotein C1, and heterogeneous nuclear ribonucleoproteins A2/B1, a potential downstream target of atrial natriuretic peptide, whereas telmisartan up-regulated angiotensin II. A second study was conducted to confirm these findings in 5/6 nephrectomy wild-type and genetically deficient DPP-4 rats treated with linagliptin or placebo. Linagliptin therapy in wild-type rats was as effective as DPP-4 genetic deficiency in terms of albuminuria reduction. Thus, linagliptin showed comparable efficacy to telmisartan in preventing CKD progression in non-diabetic rats with 5/6 nephrectomy. However, the underlying pathways seem to be different. Copyright (C) 2016, International Society of Nephrology. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Low birth weight (LBW) is associated with diseases in adulthood. The birthweight attributed risk is independent of confounding such as gestational age, sex of the newborn but also social factors. The birthweight attributed risk for diseases in later life holds for the whole spectrum of birthweight. This raises the question what pathophysiological principle is actually behind the association. In this review, we provide evidence that LBW is a surrogate of insulin resistance. Insulin resistance has been identified as a key factor leading to type 2 diabetes, cardiovascular disease as well as kidney diseases. We first provide evidence linking LBW to insulin resistance during intrauterine life. This might be caused by both genetic (genetic variations of genes controlling glucose homeostasis) and/or environmental factors (due to alterations of macronutrition and micronutrition of the mother during pregnancy, but also effects of paternal nutrition prior to conception) leading via epigenetic modifications to early life insulin resistance and alterations of intrauterine growth, as insulin is a growth factor in early life. LBW is rather a surrogate of insulin resistance in early life - either due to inborn genetic or environmental reasons - rather than a player on its own.
Nonoxidized, biologically active parathyroid hormone determines mortality in hemodialysis patients
(2013)
Background: It was shown that nonoxidized PTH (n-oxPTH) is bioactive, whereas the oxidation of PTH results in a loss of biological activity.
Methods: In this study we analyzed the association of n-oxPTH on mortality in hemodialysis patients using a recently developed assay system.
Results: Hemodialysis patients (224 men, 116 women) had a median age of 66 years. One hundred seventy patients (50%) died during the follow-up period of 5 years. Median n-oxPTH levels were higher in survivors (7.2 ng/L) compared with deceased patients (5.0 ng/L; P = .002). Survival analysis showed an increased survival in the highest n-oxPTH tertile compared with the lowest n-oxPTH tertile (chi(2), 14.3; P = 0008). Median survival was 1702 days in the highest n-oxPTH tertile, whereas it was only 453 days in the lowest n-oxPTH tertile. Multivariable-adjusted Cox regression showed that higher age increased odds for death, whereas higher n-oxPTH reduced the odds for death. Another model analyzing a subgroup of patients with intact PTH (iPTH) concentrations at baseline above the upper normal range of the iPTH assay (70 ng/L) revealed that mortality in this subgroup was associated with oxidized PTH but not with n-oxPTH levels.
Conclusions: The predictive power of n-oxPTH and iPTH on the mortality of hemodialysis patients differs substantially. Measurements of n-oxPTH may reflect the hormone status more precisely. The iPTH-associated mortality is most likely describing oxidative stress-related mortality.
Fatty acid synthase (FASN) catalyzing the terminal steps in the de novo biogenesis of fatty acids is correlated with low survival and high disease recurrence in patients with bladder cancer. Pyruvate kinase M2 (PKM2) regulates the final step of glycolysis levels and provides a growth advantage to tumors. However, it is unclear whether the change of PKM2 has an effect on FASN and what is the mechanisms underlying. Here we describe a novel function of PKM2 in control of lipid metabolism by mediating transcriptional activation of FASN, showing the reduced expression of sterol regulatory element binding protein 1c (SREBP-1c). We first discovered that PKM2 physically interacts with the SREBP-1c using biochemical approaches, and downregulation of PKM2 reduced the expression of SREBP-1c by inactivating the AKT/mTOR signaling pathway, which in turn directly suppressed the transcription of major lipogenic genes FASN to reduce tumor growths. Furthermore, either PKM2 inhibitor-Shikonin or FASN inhibitor-TVB-3166 alone induced a strong antiproliferative and anticolony forming effect in bladder cancer cell line. The combination of both inhibitors exhibits a super synergistic effect on blocking the bladder cancer cells growth. It provides a new target and scientific basis for the treatment of bladder cancer.
To identify genetic variants associated with head circumference in infancy, we performed a meta-analysis of seven genome-wide association studies (GWAS) (N = 10,768 individuals of European ancestry enrolled in pregnancy and/or birth cohorts) and followed up three lead signals in six replication studies (combined N = 19,089). rs7980687 on chromosome 12q24 (P = 8.1 x 10(-9)) and rs1042725 on chromosome 12q15 (P = 2.8 x 10(-10)) were robustly associated with head circumference in infancy. Although these loci have previously been associated with adult height(1), their effects on infant head circumference were largely independent of height (P = 3.8 x 10(-7) for rs7980687 and P = 1.3 x 10(-7) for rs1042725 after adjustment for infant height). A third signal, rs11655470 on chromosome 17q21, showed suggestive evidence of association with head circumference (P = 3.9 x 10(-6)). SNPs correlated to the 17q21 signal have shown genome-wide association with adult intracranial volume(2), Parkinson's disease and other neurodegenerative diseases(3-5), indicating that a common genetic variant in this region might link early brain growth with neurological disease in later life.
Background: The goal was to study lipid profiles (TG, TC, LDL, HDL), effects on serum leptin, and fat tissue adiponectin, and resistin as well as body weight effects of Shan He Jian Fei Granules (SHJFG) in rats on a high fat diet.
Methods: Rats were randomly divided into five groups: normal control group fed with normal fat diet, rats on high fat diet receiving low dosage, middle dosage, high dosage of Shan He Jian Fei Granules (SHJFG) as well as a high fat diet group receiving placebo. Rats were treated for 8 weeks. Body weight and naso-anal length of each rat were recorded and Lee's index was calculated. Serum TG, TC, LDL, HDL and leptin concentrations were analyzed. The gene expressions of adiponectin and resistin in adipose tissues were tested by RT-PCR.
Results: Compared to the high-fat diet group, body weights, Lee's indexes, weight of fat tissues and serum TG, TC, LDL and leptin of SHJFG groups significantly decreased (p<0.05), whereas mRNA expressions of adiponectin and resistin of SHJFG groups significantly increased (p<0.05).
Conclusions: SHJFG could significantly lower body weight and serum TG, TC, and LDL of obese rats. The effects of SHJFG in lowering leptin synthesis and raising mRNA expression of adiponectin and resistin in fat tissues may act as part of the mechanisms in lowering body weight of obese rats. Further studies are needed to demonstrate whether SHJFG may also reduce overall cardiovascular morbidity and mortality like other lipid lowering drugs.
Direct stimulation of soluble guanylate cyclase (sGC) is emerging as a potential new approach for the treatment of renal disorders. sGC catalyzes the formation of cyclic guanosine monophosphate (cGMP), deficiency of which is implicated in the pathogenesis of chronic kidney disease (CKD). Therefore, new classes of drugs sGC stimulators and activators are being investigated in preclinical models under conditions where nitric oxide is deficient. In preclinical models with different etiologies of CKD, the sGC stimulators BAY 41-2272, BAY 41-8543, BAY 60-4552, riociguat and vericiguat and the sGC activators cinaciguat, ataciguat, BI 703704 and GSK2181236A have shown consistently renoprotective effects. Clinical trials are required to confirm these findings in humans, and to ascertain whether these agents could provide a future alternative to guideline-recommended treatments.
Background: Despite the beneficial effects of type 4 dipeptidyl peptidase (DPP-4) inhibitors on glucose levels, its effects on diabetic nephropathy remain unclear.
Method: This study examined the long-term renoprotective effects of DPP-4 inhibitor linagliptin in db/db mice, a model of type 2 diabetes.
Results were compared with the known beneficial effects of renin-angiotensin system blockade by enalapril. Ten-week-old male diabetic db/db mice were treated for 3 months with either vehicle (n = 10), 3 mg linagliptin/kg per day (n = 8), or 20 mg enalapril/kg per day (n = 10). Heterozygous db/m mice treated with vehicle served as healthy controls (n = 8). Results: Neither linagliptin nor enalapril had significant effects on the parameters of glucose metabolism or blood pressure in diabetic db/db mice. However, linagliptin treatment reduced albuminuria and attenuated kidney injury. In addition, expression of podocyte marker podocalyxin was normalized. We also analysed DPP-4 expression by immunofluorescence in human kidney biopsies and detected upregulation of DPP-4 in the glomeruli of patients with diabetic nephropathy, suggesting that our findings might be of relevance for human kidney disease as well.
Conclusion: Treatment with DPP-4 inhibitor linagliptin delays the progression of diabetic nephropathy damage in a glucose-independent and blood-pressure-independent manner. The observed effects may be because of the attenuation of podocyte injury and inhibition of myofibroblast transformation.
Background: Acute kidney injury (AKI) as well as chronic renal failure are associated with a huge mortality/morbidity. However, so far no drugs have been approved for the treatment of acute kidney failure and only a few for the treatment of chronic kidney disease (CKD). We analysed the effect of SLV 338, a neutral endopeptidase (NEP)/endothelin converting enzyme (ECE)-inhibitor in animal models of acute kidney failure as well as chronic renal failure.
Methods: Acute renal failure was induced in male Wistar rats by uninephrectomy and clamping of the remaining kidney for 55 minutes. SLV338 (total dose: 4.9 mg/kg) or vehicle was continuously infused for 2 hours (starting 20 minutes prior to clamping). Sham operated animals served as controls. Plasma creatinine was measured at baseline and day 2 and 8 after renal ischemia-reperfusion.
Hypertensive renal damage was induced in male Sprague Dawley rats by nitric oxide deficiency using L-NAME (50 mg/kg per day, added to drinking water for 4 weeks). One group was treated over the same time period with SLV338 (30 mg/kg per day, mixed with food). Systolic blood pressure was monitored weekly. At study end, urine and blood samples were collected and kidneys were harvested.
Results: Acute renal ischemia-reperfusion caused a 5-fold plasma creatinine elevation (day 2), which was significantly attenuated by more than 50 % in animals treated with SLV338 (p < 0.05). Renal failure was accompanied by a 67 % mortality in vehicle-treated rats, but only 20 % after SLV338 treatment (p = 0.03 compared to sham controls).
Chronic L-NAME administration caused hypertension, urinary albumin excretion, glomerulosclerosis, renal arterial remodelling, and renal interstitial fibrosis. Treatment with SLV338 did not significantly affect blood pressure, but abolished renal tissue damage (interstitial fibrosis, glomerulosclerosis, renal arterial remodelling (p < 0.05 versus L-NAME group in each case).
Conclusions: The dual ECE/NEP inhibitor SLV338 preserves kidney function and reduces mortality in severe acute ischemic renal failure. Moreover, combined ECE/NEP inhibition prevents hypertensive renal tissue damage in a blood pressure independent manner in L-NAME-treated rats.
Objectives The nitric oxide-soluble guanylate cyclase (sGC)-cGMP signal transduction pathway is impaired in different cardiovascular diseases, including pulmonary hypertension, heart failure and arterial hypertension. Riociguat is a novel stimulator of soluble guanylate cyclase (sGC). However, little is known about the effects of sGC stimulators in experimental models of hypertension. We thus investigated the cardio-renal protective effects of riociguat in low- renin and high-renin rat models of hypertension. Methods The vasorelaxant effect of riociguat was tested in vitro on isolated saphenous artery rings of normal and nitrate tolerant rabbits. The cardiovascular in-vivo effects of sGC stimulation were evaluated in hypertensive renin-transgenic rats treated with the nitric oxide-synthase inhibitor N- nitro-L-arginine methyl ester (L-NAME) (high-renin model) and in rats with 5/6 nephrectomy (low-renin model). Results In both animal models, riociguat treatment improved survival and normalized blood pressure. Moreover, in the L-NAME study part, riociguat reduced cardiac target organ damage as indicated by lower plasma ANP, lower relative left ventricular weight and lower cardiac interstitial fibrosis, and reduced renal target organ damage as indicated by lower plasma creatinine and urea, less glomerulosclerosis and less renal interstitial fibrosis. In the 5/6 nephrectomy study part, riociguat reduced cardiac target organ damage as indicated by lower plasma ANP, lower relative left ventricular weight, lower myocyte diameter and lower arterial media/lumen ratio, and reduced renal target organ damage as indicated by improved creatinine clearance and less renal interstitial fibrosis. Conclusion We demonstrate for the first time that the novel sGC stimulator riociguat shows in two independent models of hypertension a potent protection against cardiac and renal target organ damage. J Hypertens 28: 1666-1675 (c) 2010 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.
DPP-4 inhibition with linagliptin delays the progression of diabetic nephropathy in db/db mice
(2012)
Context: Relations between fibroblast growth factor-23 (FGF-23), soluble alpha-klotho (s-alpha-klotho), and kidney function in chronic kidney disease (CKD) are still unclear. Especially the role of s-alpha-klotho requires further study.
Objectives: Our objectives were to analyze the relation of s-alpha-klotho to estimated glomerular filtration rate (eGFR), FGF-23, and other parameters of calcium-phosphate metabolism and to investigate the response of s-alpha-klotho to cholecalciferol.
Patients, Design, and Setting: Twenty-four CKD (stage 1-5) patients participated in this 8-week randomized controlled trial (vitamin D and chronic renal insufficiency).
Interventions: Interventions included 40 000 IU cholecalciferol or placebo weekly.
Main Outcome Measure: S-alpha-klotho was determined by ELISA with antihuman klotho antibodies 67G3 and 91F1.
Results: For all patients, s-alpha-klotho concentrations did not differ between CKD stages. When patients were subdivided based on FGF-23 concentrations, a positive association of s-alpha-klotho with eGFR became apparent in patients with lower than median FGF-23 concentrations but not in those above median value. Patients with s-alpha-klotho below 204 pg/mL showed higher age, lower phosphate clearance, and lower bone-specific alkaline phosphatase compared with patients with higher s-alpha-klotho. Treatment with cholecalciferol significantly increased 1,25-dihydroxyvitamin D. The increase of FGF-23 had only borderline significance. There was no significant effect of high-dose cholecalciferol administration for 8 weeks on plasma s-alpha-klotho.
Conclusions: CKD patients with s-alpha-klotho below 204 pg/mL had higher age, lower phosphate clearance, and lower bone-specific alkaline phosphatase. An association of s-alpha-klotho with eGFR was observed only in the presence of close to normal, but not high, FGF-23 concentrations. Cholecalciferol treatment did not change s-alpha-klotho concentrations.
Background: The ONTARGET trial revealed an association of ACEI/ARB combination treatment (telmisartan and ramipril) with adverse renal outcome versus respective monotherapy; preclinical evidence regarding renal outcome in ACEI/ARB combination treatment is scarce.
Methods: Spontaneously hypertensive stroke prone rats (SHR-SP) rats on a salt-rich diet were randomly allocated to 4 groups: SHR (untreated, n = 24), SHR + telmisartan (SHR-T, 2.39 +/- 0.69 mg/kg bw; n = 27), SHR + ramipril (SHR-R, 6.28 +/- 3.48 mg/kg bw; n = 27) and combination treatment (SHR-TR, 0.51 +/- 0.14 mg/kg bw; same dose for telmisartan and ramipril; n = 26). Study duration was 12 weeks, blood pressure was assessed weekly and doses were adjusted to maintain equal blood pressure. Finally, blood and urine samples were obtained and kidneys were harvested for histological studies.
Results: Blood pressure in untreated rats rose to a maximum of 239 mmHg, whereas in all treatment groups it remained stable betvveen 140 and 150 mmHg. Mortality was 50% in the untreated group, whereas all treatment groups survived completely. Renal function - as indicated by plasma urea and cystatin c - was significantly worse in SHR-TR animals compared to all other groups. With plasma creatinine a similar trend was observed. All treatment options significantly decreased albuminuria. Renal glomerulosclerosis was decreased by monotherapy, whereas combination therapy failed to have a significant effect. Interstitial fibrosis was decreased to a similar extent by all treatment options.
Conclusions: ACEI/ARB combination treatment failed to render significant additional benefits on renal outcome in hypertensive rats when compared to monotherapy. Instead our data indicate that dual RAAS blockade might have an adverse effect on kidney function and histology when compared to monotherapy in salt-loaded SHR-SP.
Background. Endothelin 1 contributes to renal blood flow control and pathogenesis of kidney diseases. The differential effects, however, of endothelin 1 (ET-1) on afferent (AA) and efferent arterioles (EA) remain to be established.
Methods. We investigated endothelin type A and B receptor (ETA-R, ETB-R) functions in the control of AA and EA. Arterioles of ETB-R deficient, rescued mice [ETB (-/-)] and wild types [ETB(+/+)] were microperfused.
Results. ET-1 constricted AA stronger than EA in ETB (-/-) and ETB(+/+) mice. Results in AA: ET-1 induced similar constrictions in ETB(-/-) and ETB(+/+) mice. BQ-123 (ETA-R antagonist) inhibited this response in both groups. ALA-ET-1 and IRL1620 (ETB-R agonists) had no effect on arteriolar diameter. L-NAME did neither affect basal diameters nor ET-1 responses. Results in EA: ET-1 constricted EA stronger in ETB(+/+) compared to ETB(-/-). BQ-123 inhibited the constriction completely only in ETB(-/-). ALA-ET-1 and IRL1620 constricted only arterioles of ETB(+/+) mice. L-NAME decreased basal diameter in ETB(+/+), but not in ETB(-/-) mice and increased the ET-1 response similarly in both groups. The L-NAME actions indicate a contribution of ETB-R in basal nitric oxide (NO) release in EA and suggest dilatory action of ETA-R in EA.
Conclusions. ETA-R mediates vasoconstriction in AA and contributes to vasoconstriction in EA in this mouse model. ETB-R has no effect in AA but mediates basal NO release and constriction in EA. The stronger effect of ET-1 on AA supports observations of decreased glomerular filtration rate to ET-1 and indicates a potential contribution of ET-1 to the pathogenesis of kidney diseases.
Pulmonary arterial hypertension (PAH) is a fatal disease that eventually results in right heart failure and death. Current pharmacologic therapies for PAH are limited, and there are no drugs that could completely cure PAH. Enhanced activity of endothelin system has been implicated in PAH severity and endothelin receptor antagonists have been used clinically to treat PAH. However, there is limited experimental evidence on the direct role of enhanced endothelin system activity in PAL-I. Here, we investigated the correlation between endothelin-1 (ET-1) and PAH using ET-1 transgenic (ETTG) mice. Exposure to chronic hypoxia increased right ventricular pressure and pulmonary arterial wall thickness in ETTG mice compared to those in wild type mice. Of note, ETTG mice exhibited modest but significant increase in right ventricular pressure and vessel wall thickness relative to wild type mice even under normoxic conditions. To induce severe PAH, we administered SU5416, a vascular endothelial growth factor receptor inhibitor, combined with exposure to chronic hypoxia. Treatment with SU5416 modestly aggravated hypoxia-induced pulmonary hypertension, right ventricular hypertrophy, and pulmonary arterial vessel wall thickening in ETTG mice in association with increased interleukin-6 expression in blood vessels. However, there was no sign of obliterative endothelial cell proliferation and plexiform lesion formation in the lungs. These results demonstrated that enhanced endothelin system activity could be a causative factor in the development of PAH and provided rationale for the inhibition of endothelin system to treat PAH. (C) 2015 Elsevier Inc. All rights reserved.
Objectives: In an experimental heart failure model, we tested the hypothesis that furosemide causes excess mortality.
Background: Post-hoc analysis of large clinical heart failure trails revealed that furosemide treatment might be associated with worsening of morbidity and even mortality in heart failure patients.
Methods and results: Myocardial infarction was induced in 7 +/- 1 week old male Wistar rats by ligation of the left coronary artery. In study 1, animals were randomly assigned to treatment with furosemide (10 mg/kg/d via drinking water, n = 33) or placebo (n = 33) starting 18 days after surgery. In study 2, animals received furosemide from day 18 and were then randomized to ongoing treatment with either furosemide only (n = 38) or furosemide plus ACE-inhibitor Ramipril (1 mg/kg/d, n = 38) starting on day 42. In study 1 survival rate in the furosemide group was lower than in the placebo group (hazard ratio {HR} 3.39, 95% confidence interval {CI} 1.14 to 10.09, p = 0.028). The furosemide group had a lower body weight (-6%, p = 0.028) at the end of the study and a higher sclerosis index of the glomeruli (+9%, p=0.026) than the placebo group. Wet lung weight, infarct size, and cardiac function were similar between the groups. In study 2, the furosemide group had a higher mortality rate than the furosemide + ramipril group (HR 4.55, 95% CI 2.0 to 10.0, p = 0.0003).
Conclusion: In our rat model of heart failure furosemide, provided at a standard dose, was associated with increased mortality. This increased mortality could be prevented by additional administration of an ACE-inhibitor.
BACKGROUND AND PURPOSE Results regarding protective effects of dipeptidyl peptidase 4 (DPP4) inhibitors in renal ischaemia-reperfusion injury (IRI) are conflicting. Here we have compared structurally unrelated DPP4 inhibitors in a model of renal IRI. EXPERIMENTAL APPROACH IRI was induced in uninephrectomizedmale rats by renal artery clamping for 30 min. The shamgroup was uninephrectomized but not subjected to IRI. DPP4 inhibitors or vehicle were given p. o. once daily on three consecutive days prior to IRI: linagliptin (1.5 mg.kg(-1).day(-1)), vildagliptin (8mg.kg(-1).day(-1)) and sitagliptin (30 mg.kg(-1).day(-1)). An additional group received sitagliptin until study end (before IRI: 30 mg.kg(-1).day(-1); after IRI: 15mg.kg(-1).day(-1)). KEY RESULTS Plasma-active glucagon-like peptide type 1 (GLP(-1)) increased threefold to fourfold in all DPP4 inhibitor groups 24 h after IRI. Plasma cystatin C, a marker of GFR, peaked 48 h after IRI. Compared with the placebo group, DPP4 inhibition did not reduce increased plasma cystatin C levels. DPP4 inhibitors ameliorated histopathologically assessed tubular damage with varying degrees of drug-specific efficacies. Renal osteopontin expression was uniformly reduced by all DPP4 inhibitors. IRI-related increased renal cytokine expression was not decreased by DPP4 inhibition. Renal DPP4 activity at study end was significantly inhibited in the linagliptin group, but only numerically reduced in the prolonged/dose-adjusted sitagliptin group. Active GLP(-1) plasma levels at study end were increased only in the prolonged/dose-adjusted sitagliptin treatment group. CONCLUSIONS AND IMPLICATIONS In rats with renal IRI, DPP4 inhibition did not alter plasma cystatin C, a marker of glomerular function, but may protect against tubular damage.
Endothelin receptor antagonists (ETRAs) are approved for the treatment of pulmonary hypertension and scleroderma-related digital ulcers. The efforts to approve this class of drugs for renal indications, however, failed so far. Preclinical studies were promising. Transgenic overexpression of ET-1 or ET-2 in rodents causes chronic renal failure. Blocking the ET system was effective in the treatment of renal failure in rodent models. However, various animal studies indicate that blocking the renal tubular ETAR and ETBR causes water and salt retention partially mediated via the epithelial sodium transporter in tubular cells. ETRAs were successfully tested clinically in renal indications in phase 2 trials for the treatment of diabetic nephropathy. They showed efficacy in terms of reducing albumin excretion on top of guideline based background therapy (RAS blockade). However, these promising results could not be translated to successful phase Ill trials so far. The spectrum of serious adverse events was similar to other phase III trials using ETRAs. Potential underlying reasons for these failures and options to solve these issues are discussed. In addition preclinical and clinical studies suggest caution when addressing renal patient populations such as patients with hepatorenal syndrome, patients with any type of cystic kidney disease and patients at risk of contrast media induced nephropathy. The lessons learned in renal indications are also important for other potential promising indications of ETRAs like cancer and heart failure. (C) 2014 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).
The concept of developmental origins of diseases has gained a huge interest in recent years and is a constantly emerging scientific field. First observations hereof originated from epidemiological studies, linking impaired birth outcomes to adult chronic, noncommunicable disease. By now there is a considerable amount of both epidemiological and experimental evidence highlighting the impact of early life events on later life disease susceptibility. Albeit far from being completely understood, more recent studies managed to elucidate underlying mechanisms, with epigenetics having become almost synonymous with developmental programming. The aim of this review was to give a comprehensive overview of various aspects and mechanisms of developmental origins of diseases. Starting from initial research foci mainly centered on a nutritionally impaired intrauterine environment, more recent findings such as postnatal nutrition, preterm birth, paternal programming and putative interventional approaches are summarized. The review outlines general underlying mechanisms and particularly discusses mechanistic explanations for sexual dimorphism in developmental programming. Furthermore, novel hypotheses are presented emphasizing a non-mendelian impact of parental genes on the offspring's phenotype.
Background: Gestational diabetes mellitus (GDM) is associated with adverse pregnancy outcomes. It is known that GDM is associated with an altered placental function and changes in placental gene regulation. More recent studies demonstrated an involvement of epigenetic mechanisms. So far, the focus regarding placental epigenetic changes in GDM was set on gene-specific DNA methylation analyses. Studies that robustly investigated placental global DNA methylation are lacking. However, several studies showed that tissue-specific alterations in global DNA methylation are independently associated with type 2 diabetes. Thus, the aim of this study was to characterize global placental DNA methylation by robustly measuring placental DNA 5-methylcytosine (5mC) content and to examine whether differences in placental global DNA methylation are associated with GDM. Methods: Global DNA methylation was quantified by the current gold standard method, LC-MS/MS. In total, 1030 placental samples were analyzed in this single-center birth cohort study. Results: Mothers with GDM displayed a significantly increased global placental DNA methylation (3.22 +/- 0.63 vs. 3.00 +/- 0.46 %; p = 0.013; +/- SD). Bivariate logistic regression showed a highly significant positive correlation between global placental DNA methylation and the presence of GDM (p = 0.0009). Quintile stratification according to placental DNA 5mC levels revealed that the frequency of GDM was evenly distributed in quintiles 1-4 (2.9-5.3 %), whereas the frequency in the fifth quintile was significantly higher (10.7 %; p = 0.003). Bivariate logistic models adjusted for maternal age, BMI, ethnicity, recurrent miscarriages, and familiar diabetes predisposition clearly demonstrated an independent association between global placental DNA hypermethylation and GDM. Furthermore, an ANCOVA model considering known predictors of DNA methylation substantiated an independent association between GDM and placental DNA methylation. Conclusions: This is the first study that employed a robust quantitative assessment of placental global DNA methylation in over a thousand placental samples. The study provides large scale evidence that placental global DNA hypermethylation is associated with GDM, independent of established risk factors.
DPP4 inhibition prevents AKI
(2017)
Background/Aims: Contrast induced acute kidney injury (CI-AKI) remains a serious complication of contrast media enhanced procedures like coronary angiography. There is still a lack of established biomarkers that help to identify patients at high risk for short and long-term complications. The aim of the current study was to evaluate plasma kynurenine as a predictive biomarker for CI-AKI and long-term complications, measured by the combined endpoint "major adverse kidney events" (MAKE) up to 120 days after CM application.
Methods: In this prospective cohort study 245 patients undergoing coronary angiography were analyzed. Blood samples were obtained at baseline, 24h and 48h after contrast media (CM) application to diagnose CI-AKI. Patients were followed for 120 days for adverse clinical events including death, the need for dialysis, and a doubling of plasma creatinine. Occurrence of any of these events was summarized in the combined endpoint MAKE.
Results: Preinterventional plasma kynurenine was not associated with CI-AKI. Patients who later developed MAKE displayed significantly increased preinterventional plasma kynurenine levels (p<0.0001). ROC analysis revealed that preinterventional kynurenine is highly predictive for MAKE (AUC=0.838; p<0.0001). The optimal cutoff was found at >= 3.5 mu mol/L. Using this cutoff, the Kaplan-Meier estimator demonstrated that concentrations of plasma kynurenine >= 3.5 mu mol/L were significantly associated with a higher prevalence of MAKE until follow up (p<0.0001). This association remained significant in multivariate Cox regression models adjusted for relevant factors of long-term renal outcome.
Conclusion: Preinterventional plasma kynurenine might serve as a highly predictive biomarker for MAKE up to 120 days after coronary angiography.
Background/Aims: Cardiovascular disease partially originates from poor environmental and nutritional conditions in early life. Lack of micronutrients like 25 hydroxy vitamin D-3 (25OHD) during pregnancy may be an important treatable causal factor. The present study explored the effect of maternal 25OHD deficiency on the offspring. Methods: We performed a prospective observational study analyzing the association of maternal 25OHD deficiency during pregnancy with birth outcomes considering confounding. To show that vitamin D deficiency may be causally involved in the observed associations, mice were set on either 25OHD sufficient or insufficient diets before and during pregnancy. Growth, glucose tolerance and mortality was analyzed in the F1 generation. Results: The clinical study showed that severe 25OHD deficiency was associated with low birth weight and low gestational age. ANCOVA models indicated that established confounding factors such as offspring sex, smoking during pregnancy and maternal BMI did not influence the impact of 25OHD on birth weight. However, there was a significant interaction between 25OHD and gestational age. Maternal 25OHD deficiency was also independently associated with low APGAR scores 5 minutes postpartum. The offspring of 25OHD deficient mice grew slower after birth, had an impaired glucose tolerance shortly after birth and an increased mortality during follow-up. Conclusions: Our study demonstrates an association between maternal 25OHD and offspring birth weight. The effect of 25OHD on birth weight seems to be mediated by vitamin D controlling gestational age. Results from an animal experiment suggest that gestational 25OHD insufficiency is causally linked to adverse pregnancy outcomes. Since birth weight and prematurity are associated with an adverse cardiovascular outcome in later life, this study emphasizes the need for novel monitoring and treatment guidelines of vitamin D deficiency during pregnancy.
Dealing with large sample sizes: comparison of a new one spot dot blot method to western blot
(2014)
Background: Western blot is the gold standard method to determine individual protein expression levels. However, western blot is technically difficult to perform in large sample sizes because it is a time consuming and labor intensive process. Dot blot is often used instead when dealing with large sample sizes, but the main disadvantage of the existing dot blot techniques, is the absence of signal normalization to a housekeeping protein.
Methods: In this study we established a one dot two development signals (ODTDS) dot blot method employing two different signal development systems. The first signal from the protein of interest was detected by horseradish peroxidase (HRP). The second signal, detecting the housekeeping protein, was obtained by using alkaline phosphatase (AP).
Results: Inter-assay results variations within ODTDS dot blot and western blot and intra-assay variations between both methods were low (1.04 - 5.71%) as assessed by coefficient of variation.
Conclusions: ODTDS dot blot technique can be used instead of western blot when dealing with large sample sizes without a reduction in results accuracy.