In this investigation, we selected PAX3/FKHR and PAX7/FKHR fusion transcript-positive and -negative alveolar rhabdomyosarcomas (ARMSs) and embryonal rhabdomyosarcomas (ERMSs) with and without anaplastic features, to ascertain genomic imbalance differences and/or similarities within these histopathologic and genetic rhabdomyosarcoma (RMS) variants. Comparative genomic hybridization (CGH) and fluorescence in situ hybridization (FISH) studies were performed on 45 rhabdomyosarcoma specimens consisting of 23 ARMSs and 22 ERMSs (12 ERMS cases were included from an earlier study). The anaplastic variant of RMS has not previously been subjected to CGH analysis. Overall, the most prominent imbalances were gain of chromosomes or chromosomal regions 2/2q (40%), 7/7q (31%), 8/8p (53%), 11/11q (31%), 12q13-15 (49%), 13q14 (22%), and 20/20p (31%), and loss of 1p36 (27%), 3p14-21 (22%), 9q21-22 (33%), 10q22-qter (18%), 16q (27%), 17p (22%), and 22 (22%). These gains and losses were distributed equally between ARMS and ERMS histologic subtypes (excluding 7/7q and 11/11q gain that were observed chiefly in ERMS), demonstrating that these entities are similar with respect to recurrent genomic imbalances. Moreover, genomic imbalances were also evenly distributed among the ARMS fusion transcript subtypes, providing evidence for a genetic kinship despite the absence of a fusion transcript in some cases. Genomic amplification was detected in 26% and 23% of the ARMS and ERMS cases, respectively (with nearly all of the latter subset exhibiting anaplastic features). One amplicon, involving 15q25-26, corresponds to the locus of the insulin-like growth factor type I receptor (IGF1R) gene. Amplification of IGF1R was confirmed molecularly in the cases exhibiting a 15q25-26 amplicon. In summary, these results indicate that genomic gains and losses involve alike chromosomes with similar frequencies within the histopathologic and genetic subtypes of rhabdomyosarcoma, that genomic amplification is frequent not only in the alveolar histologic subtype of rhabdomyosarcoma but also in ERMS with anaplasia, and that amplification of IGF1R possibly plays a role in the development or progression of a subset of rhabdomyosarcomas.

The properties and function of Ca(2+)-activated K+ (KCa) and voltage-dependent K+ (IK) currents of rabbit coronary myocytes were studied under whole cell voltage-clamp conditions (22 degrees C). Inhibition of KCa by tetraethylammonium chloride (1-10 mM) or charybdotoxin (50-100 nM) suppressed noisy outward rectifying current elicited by 5-s voltage steps or ramp at potentials > 0 mV, reduced the hump of the biphasic ramp current-voltage relation, and shifted by less than +5 mV the potential at which no net steady-state current is recorded (Enet; index of resting membrane potential). Inhibition of steady-state inward Ca2+ currents [ICa(L)] by nifedipine (1 microM) displaced Enet by -11 mV. Analysis of steady-state voltage dependence of IK supported the existence of a "window" current between -50 and 0 mV. 4-Aminopyridine (2 mM) blocked a noninactivating component of IK evoked between -30 and -40 mV, abolished the hump current during ramps, and shifted Enet by more than +15 mV; hump current persisted during 2-min ramp depolarizations and peaked near the maximum overlap of the steady-state activation and inactivation curves of IK (about -22 mV). A threefold rise in extracellular Ca2+ concentration (1.8-5.4 mM) enhanced time-dependent outward K+ current (6.7-fold at +40 mV) and shifted Enet by -30 mV. It is concluded that, under steady-state conditions, IK and ICa(L) play a major role in regulating resting membrane potential at a physiological level of intracellular Ca2+ concentration, with a minor contribution from KCa. However, elevation of intracellular Ca2+ concentration enhances KCa and hyperpolarizes the myocyte to limit Ca2+ entry through ICa(L).

Nephrotic syndrome (NS) is a genetically heterogeneous group of diseases that are divided into steroid-sensitive NS (SSNS) and steroid-resistant NS (SRNS). SRNS inevitably leads to end-stage kidney disease, and no curative treatment is available. To date, mutations in more than 24 genes have been described in Mendelian forms of SRNS; however, no Mendelian form of SSNS has been described. To identify a genetic form of SSNS, we performed homozygosity mapping, whole-exome sequencing, and multiplex PCR followed by next-generation sequencing. We thereby detected biallelic mutations in EMP2 (epithelial membrane protein 2) in four individuals from three unrelated families affected by SRNS or SSNS. We showed that EMP2 exclusively localized to glomeruli in the kidney. Knockdown of emp2 in zebrafish resulted in pericardial effusion, supporting the pathogenic role of mutated EMP2 in human NS. At the cellular level, we showed that knockdown of EMP2 in podocytes and endothelial cells resulted in an increased amount of CAVEOLIN-1 and decreased cell proliferation. Our data therefore identify EMP2 mutations as causing a recessive Mendelian form of SSNS.

In an HBV-immunocompetent mouse model, non-alcoholic hepatic steatosis inhibited HBV replication, as indicated by the reduction of HBV DNA and HBV-related antigens. HBV replication did not alter lipid metabolism in mice.

Studies implicate an important role for the mixed lineage leukemia (Mll) gene in hematopoiesis, mainly through maintaining Hox gene expression. However, the mechanisms underlying Mll-mediated hematopoiesis during embryogenesis remain largely unclear. Here, we investigate the role of mll during zebrafish embryogenesis, particularly hematopoiesis. Mll depletion caused severe defects in hematopoiesis as indicated by a lack of blood flow and mature blood cells as well as a significant reduction in expression of hematopoietic progenitor and mature blood cell markers. Furthermore, mll depletion prevented the differentiation of hematopoietic progenitors. In addition, we identified the N-terminal mini-peptide of Mll that acted as a dominant negative form to disrupt normal function of mll during embryogenesis. As expected, mll knockdown altered the expression of a subset of Hox genes. However, overexpression of these down-regulated Hox genes only partially rescued the blood deficiency, suggesting that mll may target additional genes to regulate hematopoiesis. In the mll morphants, microarray analysis revealed a dramatic up-regulation of gadd45␣a. Multiple assays indicate that mll inhibited gadd45␣a expression and that overexpression of gadd45␣a mRNA led to a phenotype similar to the one seen in the mll morphants. Taken together, these findings demonstrate that zebrafish mll plays essential roles in hematopoiesis and that gadd45␣a may serve as a potential downstream target for mediating the function of mll in hematopoiesis.

GADD45 gene has been implicated in cell cycle arrest, cell survival or apoptosis in a cell type specific and context-dependent manner. Members of GADD45 gene family have been found differentially expressed in several podocyte injury models, but their roles in podocytes are unclear. Using an in vivo zebrafish model of inducible podocyte injury that we have previously established, we found that zebrafish orthologs of gadd45b were induced upon the induction of podocyte injury. Podocyte-specific overexpression of zebrafish gadd45b exacerbated edema, proteinuria and foot-process effacement, whereas knockdown of gadd45b by morpholino-oligos in zebrafish larvae ameliorated podocyte injury. We then explored the role of GADD45B induction in podocyte injury using in vitro podocyte culture. We confirmed that GADD45B was significantly upregulated during the early phase of podocyte injury in cultured human podocytes and that podocyte apoptosis induced by TGF-β and puromycin aminonucleoside (PAN) was aggravated by GADD45B overexpression but ameliorated by shRNA-mediated GADD45B knockdown. We also showed that ROS inhibitor NAC suppressed PAN-induced GADD45B expression and subsequent activation of p38 MAPK pathway in podocytes and that inhibition of GADD45B diminished PAN-induced p38 MAPK activation. Taken together, our findings demonstrated that GADD45B has an important role in podocyte injury and may be a therapeutic target for the management of podocyte injury in glomerular diseases.

AIMTo evaluate the differences in acute kidney injury (AKI) between acute-on-chronic liver failure (ACLF) and decompensated cirrhosis (DC) patients.METHODSDuring the period from December 2015 to July 2017, 280 patients with hepatitis B virus (HBV)-related ACLF (HBV-ACLF) and 132 patients with HBV-related DC (HBV-DC) who were admitted to our center were recruited consecutively into an observational study. Urine specimens were collected from all subjects and the levels of five urinary tubular injury biomarkers were detected,including neutrophil gelatinase-associated lipocalin (NGAL), interleukin-18 (IL-18), liver-type fatty acid binding protein (L-FABP), cystatin C (CysC), and kidney injury molecule-1 (KIM-1). Simultaneously, the patient demographics, occurrence and progression of AKI, and response to terlipressin therapy were recorded. All patients were followed up for 3 mo or until death after enrollment.RESULTSAKI occurred in 71 and 28 of HBV-ACLF and HBV-DC patients, respectively (25.4% vs 21.2%, P = 0.358). Among all patients, the levels of four urinary biomarkers (NGAL, CysC, L-FABP, IL-18) were significantly elevated in patients with HBV-ACLF and AKI (ACLF-AKI), compared with that in patients with HBV-DC and AKI (DC-AKI) or those without AKI. There was a higher proportion of patients with AKI progression in ACLF-AKI patients than in DC-AKI patients (49.3% vs 17.9%, P = 0.013). Forty-three patients with ACLF-AKI and 19 patients with DC-AKI were treated with terlipressin. The response rate of ACLF-AKI patients was significantly lower than that of patients with DC-AKI (32.6% vs 57.9%, P = 0.018). Furthermore, patients with ACLF-AKI had the lowest 90 d survival rates among all groups (P < 0.001).CONCLUSIONAKI in ACLF patients is more likely associated with structural kidney injury, and is more progressive, with a poorer response to terlipressin treatment and a worse prognosis than that in DC patients.

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