Calcifying nested stromal epithelial tumor of the liver in a patient with Klinefelter syndrome: a case report and review of the literature

Background

Calcifying nested stromal epithelial tumor (CNSET) is a primary neoplasm of the liver, characterized by well-demarcated nests consisting of spindle and epithelioid cells with calcification and bone formation. An association of Cushing syndrome with CNSET has drawn attention, but the origin of CNSET has not been clarified.

Case presentation

We report here the case of a 20-year-old male with Klinefelter syndrome who underwent liver resection for an increasing liver tumor that was pathologically diagnosed with CNSET. He was postoperatively followed up and received several examinations, and recurrences and extrahepatic lymph node metastases were detected on the 64th day after surgery. Chemoembolization and chemotherapy were not effective, leading to tumor progression with development of progressive liver failure, and the patient finally died 164 days after hepatectomy.

Conclusions

This case suggests that an imbalance of hormones affects the genesis and progression of CNSET, and indicates the importance of closely following patients with CNSET by imaging with attention to hepatic recurrence and extrahepatic metastases.

     

Expression,purification, and characterization of highly active endo-α-N-acetylgalactosaminidases expressed by silkworm-baculovirus expression system

The O-glycosidase, endo-α-N-acetylgalactosaminidase from Enterococcus faecalis (endoEF) catalyzes the cleavage of core 1 and core 3 type O-linked disaccharides between GalNAc and serine or threonine residues from glycoproteins. The endoEF has broad substrate specificity and thus is extensively utilized for the structural and functional analysis of the O-linked glycans. In this study, we expressed and purified the recombinant endoEF (rEndoEF) by using the silkworm-baculovirus expression vector system (Silkworm-BEVS) and confirmed the deglycosylation activity of rEndoEF targeting reporter glycoproteins, which was equivalent to the commercial O-glycosidase. Thus, our study provides important clues to produce highly active rEndoEF O-glycosidases employing silkworm-BEVS as an alternative.     

Cooperative interaction between hepatocyte nuclear factor 4 alpha and GATA transcription factors regulates ATP-binding cassette sterol transporters ABCG5 and ABCG8

Cholesterol homeostasis is maintained by coordinate regulation of cholesterol synthesis and its conversion to bile acids in the liver. The excretion of cholesterol from liver and intestine is regulated by ATP-binding cassette half-transporters ABCG5 and ABCG8. The genes for these two proteins are closely linked and divergently transcribed from a common intergenic promoter region. Here, we identified a binding site for hepatocyte nuclear factor 4alpha (HNF4alpha) in the ABCG5/ABCG8 intergenic promoter, through which HNF4alpha strongly activated the expression of a reporter gene in both directions. The HNF4alpha-responsive element is flanked by two conserved GATA boxes that were also required for stimulation by HNF4alpha. GATA4 and GATA6 bind to the GATA boxes, coexpression of GATA4 and HNF4alpha leads to a striking synergistic activation of both the ABCG5 and the ABCG8 promoters, and binding sites for HNF4alpha and GATA were essential for maximal synergism. We also show that HNF4alpha, GATA4, and GATA6 colocalize in the nuclei of HepG2 cells and that a physical interaction between HNF4alpha and GATA4 is critical for the synergistic response. This is the first demonstration that HNF4alpha acts synergistically with GATA factors to activate gene expression in a bidirectional fashion.     

A mitochondrial exopolyphosphatase activity modulated by phosphate demand in Rhipicephalus (Boophilus) microplus embryo

This study describes Exopolyphosphatases (PPX) activity in mitochondria of Rhipicephalus microplus embryos. Mitochondria were isolated by differential centrifugation and PPX activity was analyzed through the hydrolysis of the substrate Polyphosphate (Poly P(15)). We investigated the influence of NADH, NAD+, Pi and ADP in a concentration range of 0.1-2.0 mM. Poly P hydrolysis was stimulated about two-fold by NADH and strongly inhibited by Pi. The PPX activity also increased in the presence of the respiratory substrates pyruvic and succinic acids, and this stimulatory effect disappeared upon addition of KCN. Mitochondrial respiration was activated by ADP using poly P as the only source of Pi. Endogenous poly P content changed following PPX activity during embryogenesis from the first up to 18th day of development. The data describe exopoly P as being modulated by Pi demand and related to energy supply during embryogenesis of hard ticks.     

Abnormal expression of ENaC and SGK1 mRNA induced by dietary sodium in Dahl salt-sensitively hypertensive rats

Epithelial sodium channel (ENaC) plays a crucial role in controlling sodium reabsorption in the kidney keeping the normal blood pressure. We previously reported that the expression of ENaC mRNA in the kidney of Dahl salt-sensitive (DS) rats was abnormally regulated by aldosterone, however it is unknown if dietary sodium affects the expression of ENaC and serum and glucocorticoid-regulated kinase 1 (SGK1), which plays an important role in ENaC activation, in DS rats. In the present study, we investigated whether dietary sodium abnormally affects the expression of ENaC and SGK1 mRNA in DS rats. DS and Dahl salt-resistant (DR) rats (8 weeks old) were divided into three different groups, respectively: (1) low sodium diet (0.005% NaCl), (2) normal sodium diet (0.3% NaCl), and (3) high sodium diet (8% NaCl). The high sodium diet for 4 weeks in DS rats elevated the systolic blood pressure, but did not in any other groups. The expression of alpha-ENaC mRNA in DS rats was abnormally increased by high sodium diet in contrast to DR rats, while it was normally increased by low sodium diet in DS rats similar to DR rats. The expression of beta- and gamma-ENaC mRNA in DS rats was also abnormally increased by high sodium diet unlike DR rats. The expression of SGK1 mRNA was elevated by high sodium diet in DS rats, but it was decreased in DR rats. These observations indicate that the expression of ENaC and SGK1 mRNA is abnormally regulated by dietary sodium in salt-sensitively hypertensive rats, and that this abnormal expression would be one of the factors causing salt-sensitive hypertension.     

Preventive effect of Ninjin-yoei-to,a Kampo medicine,on amyloid β1-42-induced neurodegeneration via intracellular Zn2+ toxicity in the dentate gyrus

Ninjin-yoei-to (NYT), a Kampo medicine, has ameliorative effects on cognitive dysfunction via enhancing cholinergic neuron activity. To explore an efficacy of NYT administration for prevention and cure of Alzheimer’s disease, here we examined the effect of NYT on amyloid β_1-42 (Aβ_1-42)-induced neurodegeneration in the dentate gyrus. A diet containing 3% NYT was administered to mice for 2 weeks and human Aβ_1-42 was intracerebroventricularly injected. Neurodegeneration in the dentate granule cell layer of the hippocampus, which was determined 2 weeks after the injection, was rescued by administration of the diet for 4 weeks. Aβ staining (uptake) was not modified in the dentate granule cell layer by pre-administration of the diet for 2 weeks, while Aβ_1-42-induced increase in intracellular Zn²⁺ was reduced, suggesting that pre-administration of NYT prior to Aβ injection is effective for reducing Aβ_1-42-induced Zn²⁺ toxicity in the dentate gyrus. As a matter of fact, Aβ_1-42-induced neurodegeneration in the dentate gyrus was rescued by pre-administration of NYT. Interestingly, the level of metallothioneins, intracellular Zn²⁺-binding proteins, which can capture Zn²⁺ from Zn-Aβ_1-42 complexes, was elevated in the dentate granule cell layer by pre-administration of NYT. The present study suggests that pre-administration of NYT prevents Aβ_1-42-mediated neurodegeneration in the dentate gyurs by induced synthesis of metallothioneins, which reduces intracellular Zn²⁺ toxicity induced by Aβ_1-42.     

MECP2 Isoform-Specific Vectors with Regulated Expression for Rett Syndrome Gene Therapy

Background

Rett Syndrome (RTT) is an Autism Spectrum Disorder and the leading cause of mental retardation in females. RTT is caused by mutations in the Methyl CpG-Binding Protein-2 (MECP2) gene and has no treatment. Our objective is to develop viral vectors for MECP2 gene transfer into Neural Stem Cells (NSC) and neurons suitable for gene therapy of Rett Syndrome.

Methodology/Principal Findings

We generated self-inactivating (SIN) retroviral vectors with the ubiquitous EF1α promoter avoiding known silencer elements to escape stem-cell-specific viral silencing. High efficiency NSC infection resulted in long-term EGFP expression in transduced NSC and after differentiation into neurons. Infection with Myc-tagged MECP2-isoform-specific (E1 and E2) vectors directed MeCP2 to heterochromatin of transduced NSC and neurons. In contrast, vectors with an internal mouse Mecp2 promoter (MeP) directed restricted expression only in neurons and glia and not NSC, recapitulating the endogenous expression pattern required to avoid detrimental consequences of MECP2 ectopic expression. In differentiated NSC from adult heterozygous Mecp2^tm1.1Bird+/− female mice, 48% of neurons expressed endogenous MeCP2 due to random inactivation of the X-linked Mecp2 gene. Retroviral MECP2 transduction with EF1α and MeP vectors rescued expression in 95–100% of neurons resulting in increased dendrite branching function in vitro. Insulated MECP2 isoform-specific lentiviral vectors show long-term expression in NSC and their differentiated neuronal progeny, and directly infect dissociated murine cortical neurons with high efficiency.

Conclusions/Significance

MeP vectors recapitulate the endogenous expression pattern of MeCP2 in neurons and glia. They have utility to study MeCP2 isoform-specific functions in vitro, and are effective gene therapy vectors for rescuing dendritic maturation of neurons in an ex vivo model of RTT.     

Boophilus microplus: its saliva contains microphilin, a small thrombin inhibitor

Saliva of the cattle tick Boophilus microplus contains two thrombin inhibitors, BmAP and microphilin. This work presents the purification and characterization of microphilin. It was purified from the saliva by gel filtration, ultrafiltration through a 3 kDa cut-off membrane and affinity chromatography in a thrombin-Sepharose column. Analysis by mass spectrometry showed a molecular mass of 1770 Da. Microphilin is the smallest salivary thrombin inhibitor peptide known to date. It inhibits fibrinocoagulation and thrombin-induced platelet aggregation with an IC(50) of 5.5 microM, is temperature resistant and its inhibitory activity was abolished by protease K treatment. Microphilin did not inhibit the amidolytic activity of the enzyme upon a small chromogenic substrate, but inhibited the hydrolysis of a substrate that binds both catalytic site and exosite I. Therefore, we propose that microphilin blocks thrombin at exosite I.