146 research outputs found
Analysis of localization and protein abundance of SCD1 in healthy and UAKD-affected kidneys.
<p>(<b>A</b>) SCD1 was detected in the cytoplasmic compartment selectively of proximal tubular cells (in the straight S3 segment). In the kidney of the homozygous <i>Umod</i><sup>C93F</sup> mutant mouse, SCD1 appeared to be abundant in a larger fraction of proximal tubular cells compared to the kidney of the wild-type mouse, and the average staining intensity of SCD1 positive cells appeared to be more prominent. Age of mice analyzed: four months. <i>Umod</i><sup>wt</sup>: wild-type mouse; <i>Umod</i><sup>C93F</sup>: homozygous <i>Umod</i><sup>C93F</sup> mutant mouse. Uromodulin immunohistochemistry enabled identification of TALH segments. Proximal tubule segment are morphologically characterized by luminal microvilli. Chromogen: DAB for SCD1, Vector RED for UMOD; nuclear staining: hemalum. (<b>B</b>) Protein abundance of SCD1 in whole kidney lysate of homozygous <i>Umod</i> mutant mice of both lines was increased compared to wild-type mice. Signal intensities of SCD1 were corrected for GAPDH signal intensities of the same PVDF-membrane. Mean of protein abundance of wild-type mice was set on a value of 1 [mean (wild-type)  = 1]. One-way-ANOVA with Tukey's Multiple Comparison Post hoc Test: <i>p</i> vs. wild-type, **, <i>p</i><0.01; ***, p<0.001. Age of mice analyzed: four months.</p
Functional classification of differentially expressed genes in kidney of <i>Umod</i><sup>A227T</sup> mutant line.
<p>Functional classification of differentially expressed genes in kidney of <i>Umod</i><sup>A227T</sup> mutant line.</p
List of primer sequences used for RT-qPCR.
<p>cDNA-specific primers for amplification of mouse angiopoietin-like 7 (<i>Angptl7</i>), hemoglobin alpha adult chain 1 (<i>Hba-a1</i>), ornithine decarboxylase structural 1 (<i>Odc1</i>), stearoyl-Coenzyme A desaturase 1 (<i>Scd1</i>), WAP four-disulfide core domain 15B (<i>Wfdc15b</i>), and the housekeeping genes ribosomal protein L13A (<i>Rpl13a</i>), succinate dehydrogenase complex subunit A flavoprotein (<i>Sdha</i>) and TATA box binding protein (<i>Tbp</i>).</p><p>List of primer sequences used for RT-qPCR.</p
Verification of DEGs, identified by transcriptome profiling of whole kidneys, by RT-qPCR.
<p>Data are shown as scatter dot plot with mean (n = 5 per group). Age of mice analyzed: four months. One-way-ANOVA with Tukey's Multiple Comparison Post hoc Test: <i>p</i> vs. wild-type, *, <i>p</i><0.05; **, <i>p</i><0.01; ***, p<0.001.</p
Analysis of localization and protein abundance of ANGPTL7 in healthy and UAKD-affected kidneys.
<p>(<b>A</b>) ANGPTL7 was predominantly detected in the cytoplasmic compartment of tubular cells, predominantly in TALH cells, of wild-type mice. Compared to the staining intensity of ANGPTL7 in TALH cells of wild-type mice, TALH cells of <i>Umod</i> mutant mice exhibited a lower cytoplasmic staining intensity of ANGPTL7. Age of mice analyzed: four months. <i>Umod</i><sup>wt</sup>: wild-type mouse; <i>Umod</i><sup>C93F</sup>: homozygous <i>Umod</i><sup>C93F</sup> mutant mouse. UMOD immunohistochemistry enabled identification of TALH segments. Serial kidney sections were used for ANGPTL7 and uromodulin immunohistochemistry and corresponding kidney regions are shown. C: renal cortex; P: renal papilla. Chromogen: DAB; nuclear staining: hemalum. (<b>B</b>) Protein abundance of ANGPTL7 in the outer medulla of kidneys of homozygous <i>Umod</i> mutant mice of both lines was decreased compared to wild-type mice. Signal intensities of ANGPTL7 were corrected for GAPDH signal intensities of the same PVDF-membrane. Mean of protein abundance of wild-type mice was set on a value of 1 [mean (wild-type)  = 1]. One-way-ANOVA with Tukey's Multiple Comparison Post hoc Test: <i>p</i> vs. wild-type, **, <i>p</i><0.01; ***, p<0.001. Age of mice analyzed: four months.</p
<i>RCO015</i> mutants and identification of the underlying mutation.
<p>a) A heterozygous (left) and homozygous (middle) <i>RCO015</i> mutant mouse at the age of 9 months compared to a wild type (right). The homozygous mutants are smaller and have rough hair and small eyes. b) Haplotype analysis defines the critical interval between the markers <i>116J6</i>.<i>1</i> and <i>D7Mit294</i> at mouse chromosome 7.</p
Histology.
<p>Histological sections from embryonic day 17.5 (E.17.5) till postnatal day 14 (P14) are given in an overview (left panel) and in a higher magnification of the anterior/equatorial region in the right panel of wild types, heterozygous and homozygous mutants. Cataract formation starts after birth in the homozygous mutants only. The first signs are vacuoles in the posterior cortical region of newborn mice (P1, red arrows), which are compressed later by the growing lens (P7, red arrow). Finally, they give rise to the sharp boundaries in the cortical region (P14, red arrow) being observed in the isolated lenses (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0125304#pone.0125304.g002" target="_blank">Fig 2A</a>). The bars indicate the magnification of the individual figures; the lenses of the homozygous mutants are shown in an even higher magnification than the heterozygotes to demonstrate the pathological processes in more detail.</p
DNA repair after ionizing radiation.
<p>The remaining DNA damage after irradiation of isolated peripheral mouse lymphocytes with 1 Gy (<sup>137</sup>Cs) is given. 77–95 cells of each mouse were counted manually for remaining γH2AX foci 6 hrs after irradiation. It is obvious that the heterozygous mutant show significantly more foci than the wild types indicating that the repair of DNA double-strand breaks in the mutants is not as efficient as in the wild types (p = 0.006602). The columns represent the means of foci per nucleus from 3 males of each phenotype; bars indicate the standard error of the mean (SEM).</p
Immuno-histochemistry ERCC2 expression.
<p>Using antibodies against ERCC2 and Alexa488 (green) as secondary antibody, we demonstrate that ERCC2 is expressed weakly in the ocular lens around birth; expression is increasing from P7 onward. Cell nuclei are counterstained by DAPI (blue). ERCC2 is mainly expressed in the cytosol.</p
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