Refine
Year of publication
Document Type
Language
- English (55)
Is part of the Bibliography
- yes (55)
Keywords
- Epigenetics (5)
- pregnancy (5)
- Fetal programming (4)
- Gestational diabetes (4)
- birth weight (4)
- LC-MS/MS (3)
- Metabolomics (3)
- Placenta (3)
- diabetic nephropathy (3)
- elevated plus-maze (3)
Background: DNA-methylation is a common epigenetic tool which plays a crucial role in gene regulation and is essential for cell differentiation and embryonic development. The placenta is an important organ where gene activity can be regulated by epigenetic DNA modifications, including DNA methylation. This is of interest as, the placenta is the interface between the fetus and its environment, the mother. Exposure to environmental toxins and nutrition during pregnancy may alter DNA methylation of the placenta and subsequently placental function and as a result the phenotype of the offspring. The aim of this study was to develop a reliable method to quantify DNA methylation in large clinical studies. This will be a tool to analyze the degree of DNA methylation in the human placenta in relationship to clinical readouts. Methods: Liquid chromatography-electrospray ionization/multi-stage mass spectrometry (LC-ESI/MS/MS) technique was used for the quantification of the 5dmC/dG ratio in placentas from 248 healthy pregnancies. We were able to demonstrate that this method is a reliable and stable way to determine global placental DNA methylation in large clinical trials. Results/Conclusion: The degree of placental DNA methylation seen in our pilot study varies substantially from 2% to 5%. The clinical implications of this variation need to be demonstrated in adequately powered large studies.
Being born large for gestational age is associated with increased global placental DNA methylation
(2020)
Being born small (SGA) or large for gestational age (LGA) is associated with adverse birth outcomes and metabolic diseases in later life of the offspring. It is known that aberrations in growth during gestation are related to altered placental function. Placental function is regulated by epigenetic mechanisms such as DNA methylation. Several studies in recent years have demonstrated associations between altered patterns of DNA methylation and adverse birth outcomes. However, larger studies that reliably investigated global DNA methylation are lacking. The aim of this study was to characterize global placental DNA methylation in relationship to size for gestational age. Global DNA methylation was assessed in 1023 placental samples by LC-MS/MS. LGA offspring displayed significantly higher global placental DNA methylation compared to appropriate for gestational age (AGA; p<0.001). ANCOVA analyses adjusted for known factors impacting on DNA methylation demonstrated an independent association between placental global DNA methylation and LGA births (p<0.001). Tertile stratification according to global placental DNA methylation levels revealed a significantly higher frequency of LGA births in the third tertile. Furthermore, a multiple logistic regression analysis corrected for known factors influencing birth weight highlighted an independent positive association between global placental DNA methylation and the frequency of LGA births (p=0.001).
Being born large for gestational age is associated with increased global placental DNA methylation
(2020)
Being born small (SGA) or large for gestational age (LGA) is associated with adverse birth outcomes and metabolic diseases in later life of the offspring. It is known that aberrations in growth during gestation are related to altered placental function. Placental function is regulated by epigenetic mechanisms such as DNA methylation. Several studies in recent years have demonstrated associations between altered patterns of DNA methylation and adverse birth outcomes. However, larger studies that reliably investigated global DNA methylation are lacking. The aim of this study was to characterize global placental DNA methylation in relationship to size for gestational age. Global DNA methylation was assessed in 1023 placental samples by LC-MS/MS. LGA offspring displayed significantly higher global placental DNA methylation compared to appropriate for gestational age (AGA; p<0.001). ANCOVA analyses adjusted for known factors impacting on DNA methylation demonstrated an independent association between placental global DNA methylation and LGA births (p<0.001). Tertile stratification according to global placental DNA methylation levels revealed a significantly higher frequency of LGA births in the third tertile. Furthermore, a multiple logistic regression analysis corrected for known factors influencing birth weight highlighted an independent positive association between global placental DNA methylation and the frequency of LGA births (p=0.001).
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/Aims: Impaired birth outcomes, like low birth weight, have consistently been associated with increased disease susceptibility to hypertension in later life. Alterations in the maternal or fetal metabolism might impact on fetal growth and influence birth outcomes. Discerning associations between the maternal and fetal metabolome and surrogate parameters of fetal growth could give new insight into the complex relationship between intrauterine conditions, birth outcomes, and later life disease susceptibility. Methods: Using flow injection tandem mass spectrometry, targeted metabolomics was performed in serum samples obtained from 226 mother/child pairs at delivery. Associations between neonatal birth weight and concentrations of 163 maternal and fetal metabolites were analyzed. Results: After FDR adjustment using the Benjamini-Hochberg procedure lysophosphatidylcholines (LPC) 14:0, 16:1, and 18:1 were strongly positively correlated with birth weight. In a stepwise linear regression model corrected for established confounding factors of birth weight, LPC 16: 1 showed the strongest independent association with birth weight (CI: 93.63 - 168.94; P = 6.94x10(-11)). The association with birth weight was stronger than classical confounding factors such as offspring sex (CI: - 258.81- -61.32; P = 0.002) and maternal smoking during pregnancy (CI: -298.74 - -29.51; P = 0.017). Conclusions: After correction for multiple testing and adjustment for potential confounders, LPC 16:1 showed a very strong and independent association with birth weight. The underlying molecular mechanisms linking fetal LPCs with birth weight need to be addressed in future studies. (c) 2018 The Author(s) Published by S. Karger AG, Basel
Background/Aims: Impaired birth outcomes, like low birth weight, have consistently been associated with increased disease susceptibility to hypertension in later life. Alterations in the maternal or fetal metabolism might impact on fetal growth and influence birth outcomes. Discerning associations between the maternal and fetal metabolome and surrogate parameters of fetal growth could give new insight into the complex relationship between intrauterine conditions, birth outcomes, and later life disease susceptibility. Methods: Using flow injection tandem mass spectrometry, targeted metabolomics was performed in serum samples obtained from 226 mother/child pairs at delivery. Associations between neonatal birth weight and concentrations of 163 maternal and fetal metabolites were analyzed. Results: After FDR adjustment using the Benjamini-Hochberg procedure lysophosphatidylcholines (LPC) 14:0, 16:1, and 18:1 were strongly positively correlated with birth weight. In a stepwise linear regression model corrected for established confounding factors of birth weight, LPC 16: 1 showed the strongest independent association with birth weight (CI: 93.63 - 168.94; P = 6.94x10(-11)). The association with birth weight was stronger than classical confounding factors such as offspring sex (CI: - 258.81- -61.32; P = 0.002) and maternal smoking during pregnancy (CI: -298.74 - -29.51; P = 0.017). Conclusions: After correction for multiple testing and adjustment for potential confounders, LPC 16:1 showed a very strong and independent association with birth weight. The underlying molecular mechanisms linking fetal LPCs with birth weight need to be addressed in future studies. (c) 2018 The Author(s) Published by S. Karger AG, Basel
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.
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.
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: Blood pressure (BP) regulation during pregnancy is influenced by hormones of placental origin. It was shown that the glucocorticoid system is altered in hypertensive pregnancy disorders such as preeclampsia. Epigenetic mechanism might influence the activity of genes involved in placental hormone/hormone receptor synthesis/action during pregnancy. Method: In the current study, we analyzed the association of 50-C-phosphate-G-30 (CpG) site methylation of different glucocorticoid receptor gene (NR3C1) promoter regions with BP during pregnancy. The study was performed as a nested case-control study (n = 80) out of 1045 mother/ child pairs from the Berlin Birth Cohort. Placental DNA was extracted and bisulfite converted. Nested PCR products from six NR3C1 proximal promoter regions [glucocorticoid receptor gene promotor region B (GR-1B), C (GR-1C), D (GR-1D), E (GR-1E), F (GR-1F), and H (GR-1H)] were analyzed by next generation sequencing. Results: NR3C1 promoter regions GR-1D and GR-1E had a much higher degree of DNA methylation as compared to GR-1B, GR-1F or GR-1H when analyzing the entire study population. Comparison of placental NR3C1 CpG site methylation among hypotensive, normotensive and hypertensive mothers revealed several differently methylated CpG sites in the GR-1F promoter region only. Both hypertension and hypotension were associated with increased DNA methylation of GR-1F CpG sites. These associations were independent of confounding factors, such as family history of hypertension, smoking status before pregnancy and prepregnancy BMI. Assessment of placental glucocorticoid receptor expression by western blot showed that observed DNA methylation differences were not associated with altered levels of placental glucocorticoid receptor expression. However, correlation matrices of all NR3C1 proximal promoter regions demonstrated different correlation patterns of intraregional and interregional DNA methylation in the three BP groups, putatively indicating altered transcriptional control of glucocorticoid receptor isoforms. Conclusion: Our study provides evidence of an independent association between placental NR3C1 proximal promoter methylation and maternal BP. Furthermore, we observed different patterns of NR3C1 promoter methylation in normotensive, hypertensive and hypotensive pregnancy.
DPP-4 inhibition with linagliptin delays the progression of diabetic nephropathy in db/db mice
(2012)
DPP4 inhibition prevents AKI
(2017)
Background: As the prevalence of diabetes rises, its complications such as diabetic nephropathy affect an increaseing number of patients. Consequently, the need for biomarkers in rodent models which reflect the stage and course of diabetic nephropathy is high. This article focuses on Heart-type fatty acid binding protein (H-FABP), osteopontin (OPN), nephrin, and Neutrophil gelatinase-associated lipocalin (NGAL) in urine, and kidney injury molecule (KIM)-1, clusterin, and tissue inhibitior of metalloproteinases (TIMP) 1 in plasma in uni-nephrectomized rats with streptocotozin-induced type 1 diabetes mellitus, a common animal model to explore renal impairment in the setting of diabetes mellitus.
Methods: 23 male Wistar rats were uni-nephrectomized and subsequently divided into two study groups. The diabetic group received streptozotocin (STZ) via tail-vein injection, the non-diabetic group received citrate buffer without STZ. Subsequently, blood glucose, body weight, and blood pressure were checked regularly. After 18 weeks, animals were placed in metabolic cages, blood and urine obtained and subsequently organs were harvested after sacrifice.
Results: Blood glucose levels were highly increased in diabetic animals throughout the experiment, whereas systolic blood pressure did not differ between the study groups. At study end, classical biomarkers such as urinary albumin and protein and plasma cystatin c were only slightly but not significantly different between groups indicating a very early disease state. In contrast, urinary excretion of H-FABP, OPN, nephrin, and NGAL were highly increased in diabetic animals with a highly significant p-value (p<0.01 each) compared to non-diabetic animals. In plasma, differences were found for calbindin, KIM-1, clusterin, TIMP-1, and OPN. These findings were confirmed by means of the area under the receiver operating characteristic curve (ROC-AUC) analysis.
Conclusions: In summary, our study revealed elevated levels of new plasma and urinary biomarkers (urinary osteopontin, urinary nephrin, urinary NGAL, urinary H-FABP, plasma KIM-1, plasma TIMP-1) in uni-nephrectomized diabetic rats, an established rat model of diabetic nephropathy. These biomarkers appeared even before the classical biomarkers of diabetic nephropathy such as albuminuria and urinary protein excretion. The new biomarkers might offer advantage to urinary albumin and plasma cystatin c with respect to early detection.
Background/Aims: Carbon monoxide (CO) interferes with cytochrome-dependent cellular functions and acts as gaseous transmitter. CO is released from CO-releasing molecules (CORM) including tricarbonyl-dichlororuthenium (II) dimer (CORM-2), molecules considered for the treatment of several disorders including vascular dysfunction, inflammation, tissue ischemia and organ rejection. Cytochrome P450-sensitive function include formation of 1,25-dihydroxyvitamin D-3 (1,25(OH)(2)D-3) by renal 25-hydroxyvitamin D-3 1-alpha-hydroxylase (Cyp27b1). The enzyme is regulated by PTH, FGF23 and klotho. 1,25(OH)(2)D-3 regulates Ca2+ and phosphate transport as well as klotho expression. The present study explored, whether CORM-2 influences 1,25(OH)(2)D-3 formation and klotho expression. Methods: Mice were treated with intravenous CORM-2 (20 mg/kg body weight). Plasma 1,25(OH)(2)D-3 and FGF23 concentrations were determined by ELISA, phosphate, calcium and creatinine concentrations by colorimetric methods, transcript levels by quantitative RT-PCR and protein expression by western blotting. Fgf23 mRNA transcript levels were further determined in rat osteosarcoma UMR106 cells without or with prior treatment for 24 hours with 20 mu M CORM-2. Results: CORM-2 injection within 24 hours significantly increased FGF23 plasma levels and decreased 1,25(OH)(2)D-3 plasma levels, renal Cyp27b1 gene expression as well as renal klotho protein abundance and transcript levels. Moreover, treatment of UMR106 cells with CORM-2 significantly increased Fgf23 transcript levels. Conclusion: CO-releasing molecule CORM-2 enhances FGF23 expression and release and decreases klotho expression and 1,25(OH)(2)D-3 synthesis.
Objective:To investigate the effects of linagliptin alone and in combination with the angiotensin II receptor blocker (ARB), telmisartan on blood pressure (BP), kidney function, heart morphology and oxidative stress in rats with renovascular hypertension.Methods:Fifty-seven male Wistar rats underwent unilateral surgical stenosis of the renal artery [2-kidney-1-clip (2k1c) method]. Animals were randomly divided into four treatment groups (n=14-18 per group) receiving: telmisartan (10mg/kg per day in drinking water), linagliptin (89ppm in chow), combination (linagliptin 89ppm+telmisartan 10mg/kg per day) or placebo. An additional group of 12 rats underwent sham surgery. BP was measured one week after surgery. Hypertensive animals entered a 16-week dosing period. BP was measured 2, 4, 8, 12 and 16 weeks after the initiation of treatment. Blood and urine were tested for assessment of kidney function and oxidative stress 6, 10, 14 and 18 weeks after surgery. Blood and urine sampling and organ harvesting were finally performed.Results:Renal stenosis caused an increase in meanSD systolic BP as compared with the sham group (157.7 +/- 29.3 vs. 106.2 +/- 20.5mmHg, respectively; P<0.001). Telmisartan alone and in combination with linagliptin, normalized SBP (111.1 +/- 24.3mmHg and 100.4 +/- 13.9mmHg, respectively; P<0.001 vs. placebo). Telmisartan alone and in combination with linagliptin significantly prevented cardiac hypertrophy, measured by heart weight and myocyte diameter. Renal function measured by cystatin C was not affected by 2k1c surgery. Telmisartan significantly increased plasma concentration of cystatin C. 2k1c surgery initiated fibrosis in both kidneys. Telmisartan promoted further fibrotic changes in the clipped kidney, as measured by protein expression of Col1a1 and histology for interstitial fibrosis and glomerulosclerosis. In non-clipped kidneys, telmisartan demonstrated antifibrotic properties, reducing Col1a1 protein expression. Plasma levels of oxidized low-density lipoprotein were higher in the placebo-treated 2k1c rats as compared to sham-operated animals. The increase was abolished by linagliptin alone (P=0.03 vs. placebo) and in combination with telmisartan (P=0.02 vs. placebo). Combination therapy also significantly reduced plasma concentration of carbonyl proteins (P=0.04 vs. placebo).Conclusion:Inhibition of type 4 dipeptidyl peptidase with linagliptin did not counter BP-lowering effects of ARB in 2k1c rats. Linagliptin reduced lipid and protein oxidation in 2k1c rats, and this effect was BP-independent.
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/).
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
Background: Recent studies show that preterm birth is associated with hypertension in later life. The renin-angiotensin system (RAS) during pregnancy influences fetal growth and development. In the current study, we investigated the impact of fetal as well as maternal angiotensin (1-7) [Ang (1-7)] and angiotensin II (Ang II) plasma concentrations on the risk of preterm birth.
Methods: Three hundred and nine pregnant women were prospectively included into the study. The pregnant women were divided into two groups, for example, preterm birth of lower than 37 gestational weeks (n = 17) and full-term birth of 37 gestational weeks or more (n = 292). Maternal and neonatal plasma Ang (1-7) and Ang II concentrations were analyzed at birth from maternal venous blood and umbilical cord blood, respectively. Risk factors for premature birth were determined by multiple logistic regression analysis.
Results: Fetal and maternal plasma Ang (1-7) concentrations in the preterm group were lower than those of the term group fetal Ang (1-7) preterm birth: 486.15 +/- 337.34 ng/l and fetal Ang (1-7) term birth: 833.84 +/- 698.12 ng/l and maternal Ang (1-7) preterm birth: 399.86 +/- 218.93 ng/l; maternal Ang (1-7) term birth: 710.34 +/- 598.22 ng/l. Multiple logistic regression analysis considering confounding factors revealed that preeclampsia (P < 0.001), premature rupture of membranes (P = 0.001), lower concentration of maternal Ang (1-7) (P = 0.013) and fetal plasma Ang (1-7) (P = 0.032) were independently associated with preterm birth. We could furthermore demonstrate that the maternal Ang (1-7)/Ang II ratio is independently associated with gestational hypertension or preeclampsia, factors causing preterm birth.
Conclusions: Lower concentrations of maternal and fetal Ang (1-7) are independently associated with preterm birth - a risk factor of hypertension in later life.