Automated determination of 8-OHdG in cells and tissue via immunofluorescence using a specially created antibody.

Despite powerful DNA repair systems, oxidative damage/modification to DNA is an inevitable side effect of metabolism, ionizing radiation, lifestyle habits, inflammatory pathologies such as type-2 diabetes or metabolic syndrome, cancer and natural aging. One of the most common oxidative DNA modifications is 8-OHdG (8­hydroxy-2'-deoxyguanosine), which is the most widely used marker in research and clinical diagnostics. 8-OHdG is easily and specifically detectable in various samples such as urine, plasma, cells and tissues via a large variety of methods like ELISA, HPLC, chromatographic methods, and immunochemistry. Formed by oxidation of guanine and being representative for the degree of DNA damage, 8-OHdG can be also used as biomarker for risk assessment of various cancers as well as degenerative diseases. Here, we present a highly specific, self-developed 8-OHdG antibody in successful comparison to a commercially one, tested in cells (FF95, HCT116, and HT22) and intestinal tissue, focusing on automatized evaluation via fluorescence/confocal microscopy.

Global Longitudinal Strain at Rest as an Independent Predictor of Mortality in Liver Transplant Candidates: A Retrospective Clinical Study.

Speckle tracking echocardiography enables the detection of subclinical left ventricular dysfunction at rest in many heart diseases and potentially in severe liver diseases. It could also possibly serve as a predictor for survival. In this study, 117 patients evaluated for liver transplantation in a single center between May 2010 and April 2016 with normal left ventricular ejection fraction were included according to clinical characteristics of their liver disease: (1) compensated (n = 29), (2) clinically significant portal hypertension (n = 49), and (3) decompensated (n = 39). Standard echocardiography and speckle tracking echocardiography were performed at rest and during dobutamine stress. Follow-up amounted to three years to evaluate survival and major cardiac events. Altogether 67% (78/117) of the patients were transplanted and 32% (31/96 patients) died during the three-year follow-up period. Global longitudinal strain (GLS) at rest was significantly increased (became more negative) with the severity of liver disease (p < 0.001), but reached comparable values in all groups during peak stress. Low (less negative) GLS values at rest (male: >-17/female: >-18%) could predict patient survival in a multivariate Cox regression analysis (p = 0.002). GLS proved valuable in identifying transplant candidates with latent systolic dysfunction.

Homozygous YME1L1 mutation causes mitochondriopathy with optic atrophy and mitochondrial network fragmentation.

Mitochondriopathies often present clinically as multisystemic disorders of primarily high-energy consuming organs. Assembly, turnover, and surveillance of mitochondrial proteins are essential for mitochondrial function and a key task of AAA family members of metalloproteases. We identified a homozygous mutation in the nuclear encoded mitochondrial escape 1-like 1 gene YME1L1, member of the AAA protease family, as a cause of a novel mitochondriopathy in a consanguineous pedigree of Saudi Arabian descent. The homozygous missense mutation, located in a highly conserved region in the mitochondrial pre-sequence, inhibits cleavage of YME1L1 by the mitochondrial processing peptidase, which culminates in the rapid degradation of YME1L1 precursor protein. Impaired YME1L1 function causes a proliferation defect and mitochondrial network fragmentation due to abnormal processing of OPA1. Our results identify mutations in YME1L1 as a cause of a mitochondriopathy with optic nerve atrophy highlighting the importance of YME1L1 for mitochondrial functionality in humans.

Morphological and behavioral characterization of adult mice deficient for SrGAP3.

SrGAP3 belongs to the family of Rho GTPase proteins. These proteins are thought to play essential roles in development and in the plasticity of the nervous system. SrGAP3-deficient mice have recently been created and approximately 10 % of these mice developed a hydrocephalus and died shortly after birth. The others survived into adulthood, but displayed neuroanatomical alteration, including increased ventricular size. We now show that SrGAP3-deficient mice display increased brain weight together with increased hippocampal volume. This increase was accompanied by an increase of the thickness of the stratum oriens of area CA1 as well as of the thickness of the molecular layer of the dentate gyrus (DG). Concerning hippocampal adult neurogenesis, we observed no significant change in the number of proliferating cells. The density of doublecortin-positive cells also did not vary between SrGAP3-deficient mice and controls. By analyzing Golgi-impregnated material, we found that, in SrGAP3-deficient mice, the morphology and number of dendritic spines was not altered in the DG. Likewise, a Sholl-analysis revealed no significant changes concerning dendritic complexity as compared to controls. Despite the distinct morphological alterations in the hippocampus, SrGAP3-deficient mice were relatively inconspicuous in their behavior, not only in the open-field, nest building but also in the Morris water-maze. However, the SrGAP3-deficient mice showed little to no interest in burying marbles; a behavior that is seen in some animal models related to autism, supporting the view that SrGAP3 plays a role in neurodevelopmental disorders.

SrGAP3 knockout mice display enlarged lateral ventricles and specific cilia disturbances of ependymal cells in the third ventricle.

In several mouse models of mental retardation, ventricular enlargements have been observed. Mutation in the SrGAP3 gene residing on chromosome 3p25 has previously been associated with intellectual disability in humans. In addition, SrGAP3 is related to Rho-GAPs signaling pathways, which play essential roles in the development and plasticity of the nervous system. About 10 % of postnatal homozygous SrGAP3-deficient mice die due to hydrocephalus, whereas the remaining mice survive into adulthood but display enlarged ventricles. We analyze the ventricular enlargement of these mice by performing a post-mortem MRI approach. We found a more than 15-fold enlargement of the lateral ventricles of homozygous SrGAP3-deficient mice. Moreover, we demonstrate that this phenotype was not accompanied by a stenosis of the aqueduct. Instead, SrGAP3 knockout mice displayed reduced densities of cilia of ependymal cells in These third ventricle compared to age-matched controls. This results indicate that the ventricular enlargement may be due to ciliopathy.