DICER1 mutations in childhood cystic nephroma and its relationship to DICER1-renal sarcoma

The pathogenesis of cystic nephroma of the kidney has interested pathologists for over 50 years. Emerging from its initial designation as a type of unilateral multilocular cyst, cystic nephroma has been considered as either a developmental abnormality or a neoplasm or both. Many have viewed cystic nephroma as the benign end of the pathologic spectrum with cystic partially differentiated nephroblastoma and Wilms tumor, whereas others have considered it a mixed epithelial and stromal tumor. We hypothesize that cystic nephroma, like the pleuropulmonary blastoma in the lung, represents a spectrum of abnormal renal organogenesis with risk for malignant transformation. Here we studied DICER1 mutations in a cohort of 20 cystic nephromas and 6 cystic partially differentiated nephroblastomas, selected independently of a familial association with pleuropulmonary blastoma and describe four cases of sarcoma arising in cystic nephroma, which have a similarity to the solid areas of type II or III pleuropulmonary blastoma. The genetic analyses presented here confirm that DICER1 mutations are the major genetic event in the development of cystic nephroma. Further, cystic nephroma and pleuropulmonary blastoma have similar DICER1 loss of function and ‘hotspot' missense mutation rates, which involve specific amino acids in the RNase IIIb domain. We propose an alternative pathway with the genetic pathogenesis of cystic nephroma and DICER1-renal sarcoma paralleling that of type I to type II/III malignant progression of pleuropulmonary blastoma

Crucial Roles of the Protein Kinases MK2 and MK3 in a Mouse Model of Glomerulonephritis

<div><p>Elevated mitogen-activated protein kinase p38 (p38 MAPK) signaling has been implicated in various experimental and human glomerulopathies, and its inhibition has proven beneficial in animal models of these diseases. p38 MAPK signaling is partially mediated through MK2 and MK3, two phylogenetically related protein kinases that are its direct substrates. The current study was designed to determine the specific roles of MK2 and MK3 in a mouse model of acute proliferative glomerulonephritis, using mice with disrupted MK2 and/or MK3 genes. We found that the absence of MK3 alone worsened the disease course and increased mortality slightly compared to wild-type mice, whereas the absence of MK2 alone exhibited no significant effect. However, in an MK3-free background, the disease course depended on the presence of MK2 in a gene dosage-dependent manner, with double knock-out mice being most susceptible to disease induction. Histological and renal functional analyses confirmed kidney damage following disease induction. Because the renal stress response plays a crucial role in kidney physiology and disease, we analyzed the stress response pattern in this disease model. We found that renal cortices of diseased mice exhibited a pronounced and specific pattern of expression and/or phosphorylation of stress proteins and other indicators of the stress response (HSPB1, HSPB6, HSPB8, CHOP, eIF2α), partially in a MK2/MK3 genotype-specific manner, and without induction of a general stress response. Similarly, the expression and activation patterns of other protein kinases downstream of p38 MAPK (MNK1, MSK1) depended partially on the MK2/MK3 genotype in this disease model. In conclusion, MK2 and MK3 together play crucial roles in the regulation of the renal stress response and in the development of glomerulonephritis, which can potentially be exploited to develop novel therapeutic approaches to treat glomerular disease.</p> </div

Effect of MK2 and MK3 genotypes on proteinuria in response to the AMC serum.

<p>(A) Scatter plots show the urinary protein/creatinine ratios from samples collected from all mice at day 0 prior to AMC serum injection and at days 4, 8, and 12 following AMC serum injection. Horizontal bars indicate the means and error bars represent S.D. Asterisks indicate significant (<i>P</i><0.05) differences between means, as compared to the wild-type group at the same day. At days 8 and 12, all means were significantly greater than baseline proteinuria values at day 0 of the same genotypic group, with exception of the MK2/MK3 double knock-out (<i>MK2^−/−MK3^−/−</i>) mice at day 8. Given the high degree of variability within each experimental group, potential differences in mean proteinuria values among all other groups failed to reach statistical significance. Urine samples of mice selected for electrophoretic protein analysis as shown in (B) are labeled by numbers in the panels of days 4, 8, and 12. Note that in some instances the amount of collected urine was not sufficient for protein determination (i.e. the number of dots is less than the number of surviving mice as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054239#pone-0054239-g002" target="_blank">Figure 2</a>). (B) Urinary serum albumin excretion of selected mice (numbered 1 - 20) from the different <i>MK2</i> and <i>MK3</i> genotypes as visualized on Coomassie-stained SDS gels. Some of the selected mice survived throughout the entire experiment, while others died after day 4. The mouse numbers correspond to the numbered proteinuria values as indicated in (A). Consistent with the protein/creatinine ratios shown in (A), at day 0 mice of all genotypes had negligible albuminuria. Following AMC serum treatment, massive albuminuria was detected in most of the mice, with some variation in its extent and onset.</p

Expression and phosphorylation of various MKs in renal cortices in response to the AMC serum.

<p>Extracts of renal cortices were processed for SDS-PAGE from untreated mice (day 0; baseline control) and AMC serum-treated mice (day 8 of treatment). Expression and phosphorylation (activation) of MK5 (A), MNK1 (B), and MSK1 (C) before and after AMC serum treatment are shown. The dot in (B) marks the correct MNK1 band (upper band; ∼48 kDa), whereas the lower band (∼44 kDa) probably results from an unspecific cross-reaction of the antibody. (D) Expression of GAPDH served as a loading control.</p