ABCG2 Dysfunction Increases the Risk of Renal Overload Hyperuricemia

ATP-binding cassette transporter, sub-family G, member 2 (ABCG2/BCRP) is identified as a high-capacity urate exporter, and its dysfunction has an association with serum uric acid levels and gout/hyperuricemia risk. Generally, hyperuricemia has been classified into urate “overproduction type,” “underexcretion type,” and “combined type” based on only renal urate excretion, without considering an extra-renal pathway such as gut excretion. In this study, we investigated the effects of ABCG2 dysfunction on human urate handling and the mechanism of hyperuricemia.

Clinical parameters for urate handling including urinary urate excretion (UUE) were examined in 644 Japanese male outpatients with hyperuricemia. The severity of their ABCG2 dysfunction was estimated by genotype combination of two common ABCG2 variants, nonfunctional Q126X (rs72552713) and half-functional Q141K (rs2231142).

Contrary to the general understanding that ABCG2 dysfunction leads to decreased renal urate excretion, UUE was significantly increased by ABCG2 dysfunction (P = 3.60 × 10^?10). Mild, moderate, and severe ABCG2 dysfunctions significantly raised the risk of “overproduction” hyperuricemia including overproduction type and combined type, conferring risk ratios of 1.36, 1.66, and 2.35, respectively.

The present results suggest that common dysfunctional variants of ABCG2 decrease extra-renal urate excretion including gut excretion and cause hyperuricemia. Thus, “overproduction type” in the current concept of hyperuricemia should be renamed “renal overload type,” which is caused by two different mechanisms, “extra-renal urate underexcretion” and genuine “urate overproduction.”

Our new concept will lead to a more accurate diagnosis and more effective therapeutic strategy for hyperuricemia and gout.     

Identification of a Hypouricemia Patient with SLC2A9 R380W,A Pathogenic Mutation for Renal Hypouricemia Type 2

Hypouricemia is characterized by low serum uric acid (SUA) levels (≤3.0 mg/dL) with complications such as urolithiasis and exercise-induced acute renal failure. We have previously reported that urate transporter 1 (URAT1/SLC22A12) and glucose transporter 9 (GLUT9/SLC2A9) are causative genes for renal hypouricemia type 1 (RHUC1) and renal hypouricemia type 2 (RHUC2), respectively. In the series of experiments, two families have been revealed to have RHUC2 due to GLUT9 missense mutations R198C or R380W, respectively. Thus far, however, no studies have reported other RHUC2 families or patients with these pathogenic mutations. This study is aimed to find other cases of RHUC2.

We performed mutational analyses of GLUT9 exon 6 (for R198C) and exon 10 (for R380W) in 50 Japanese hypouricemia patients. Patients were analyzed out of a collection of more than 2000 samples from the Japan Multi-Institutional Collaborative Cohort Study (J-MICC Study).

We identified a novel male patient with heterogeneous RHUC2 mutation R380W. The SUA of this hypouricemia patient was 2.6 mg/dL, which is similar to that of our previous report (SUA: 2.7 mg/dL).

This is the second report indicating RHUC2 patient due to GLUT9 mutation R380W. This mutation occurs in highly conserved amino acid motifs and is reported to be an important membrane topology determinant. R380W is a dysfunctional mutation which completely diminishes the urate transport activities of GLUT9. Our study revealed a second hypouricemia patient with GLUT9 R380W, a pathogenic mutation of RHUC2, which may help to expand our understanding of RHUC pathogenesis.     

Detection and Quantitation of Circulating Tumor Cell Dynamics by Bioluminescence Imaging in an Orthotopic Mammary Carcinoma Model

Circulating tumor cells (CTCs) have been detected in the bloodstream of both early-stage and advanced cancer patients. However, very little is know about the dynamics of CTCs during cancer progression and the clinical relevance of longitudinal CTC enumeration. To address this, we developed a simple bioluminescence imaging assay to detect CTCs in mouse models of metastasis. In a 4T1 orthotopic metastatic mammary carcinoma mouse model, we demonstrated that this quantitative method offers sensitivity down to 2 CTCs in 0.1–1mL blood samples and high specificity for CTCs originating from the primary tumor, independently of their epithelial status. In this model, we simultaneously monitored blood CTC dynamics, primary tumor growth, and lung metastasis progression over the course of 24 days. Early in tumor development, we observed low numbers of CTCs in blood samples (10–15 cells/100 µL) and demonstrated that CTC dynamics correlate with viable primary tumor growth. To our knowledge, these data represent the first reported use of bioluminescence imaging to detect CTCs and quantify their dynamics in any cancer mouse model. This new assay is opening the door to the study of CTC dynamics in a variety of animal models. These studies may inform clinical decision on the appropriate timing of blood sampling and value of longitudinal CTC enumeration in cancer patients.     

Prosocial Behaviors during School Activities among Child Survivors after the 2011 Earthquake and Tsunami in Japan: A Retrospective Observational Study

Background

The 2011 Japan massive tsunami traumatized many children. The aim of this study was to assess changes in strengths and difficulties experienced in home and school by among surviving children after the 2011 tsunami, in comparison with published normal Japanese data.

Methods

In November 2012 (20 months after the disaster) and September 2013 (30 months after the disaster), the Strengths and Difficulties Questionnaire (SDQ), a questionnaire on children''s strengths and difficulties in home and school activities, were distributed to 12,193 and 11,819 children, respectively. An effective response of children 20 months and 30 month after the disaster was obtained in 10,597 children (86.9%), and 10,812 children (91.4%), respectively. The SDQ scores evaluated by parents and teachers were compared with published normal Japanese SDQ scores.

Results

The SDQ scores (emotional problems, conduct problems, hyperactivity/inattention, peer relationship problems, and total difficulty score) evaluated by parents of children in the 4th to 9th grade who were evaluated after 30 and 20 months were significantly high compared with the published normal data of children without traumatic experiences (all P<0.001). The SDQ scores (prosocial behavior) evaluated by teachers of children in the 4th to 9th grade who were evaluated after 30 and 20 months were significantly low compared with the published normal data of children without traumatic experiences (all P<0.001).

Conclusions

This study showed that the experience of the disaster affected those children with prosocial behaviors towards teachers and friends at school. However, no significant changes (in their prosocial attitude) had been seen at home, where they continued to keep their respect and caring feelings for parents. These results indicate that for accurate diagnosis, clinicians should not only evaluate these children''s daily activities at home but also try to objectively assess their daily activities at school.     

Mathematical model of nucleotide regulation on airway epithelia. Implications for airway homeostasis

In the airways, adenine nucleotides support a complex signaling network mediating host defenses. Released by the epithelium into the airway surface liquid (ASL) layer, they regulate mucus clearance through P2 (ATP) receptors, and following surface metabolism through P1 (adenosine; Ado) receptors. The complexity of ASL nucleotide regulation provides an ideal subject for biochemical network modeling. A mathematical model was developed to integrate nucleotide release, the ectoenzymes supporting the dephosphorylation of ATP into Ado, Ado deamination into inosine (Ino), and nucleoside uptake. The model also includes ecto-adenylate kinase activity and feed-forward inhibition of Ado production by ATP and ADP. The parameters were optimized by fitting the model to experimental data for the steady-state and transient concentration profiles generated by adding ATP to polarized primary cultures of human bronchial epithelial (HBE) cells. The model captures major aspects of ATP and Ado regulation, including their >4-fold increase in concentration induced by mechanical stress mimicking normal breathing. The model also confirmed the independence of steady-state nucleotide concentrations on the ASL volume, an important regulator of airway clearance. An interactive approach between simulations and assays revealed that feed-forward inhibition is mediated by selective inhibition of ecto-5'-nucleotidase. Importantly, the model identifies ecto-adenylate kinase as a key regulator of ASL ATP and proposes novel strategies for the treatment of airway diseases characterized by impaired nucleotide-mediated clearance. These new insights into the biochemical processes supporting ASL nucleotide regulation illustrate the potential of this mathematical model for fundamental and clinical research.     

Establishment of axenic endosymbiotic strains of Japanese Paramecium bursaria and the utilization of carbohydrate and nitrogen compounds by the isolated algae

Cells of the green paramecium, Paramecium bursaria, contain several hundred endosymbiont cells. The properties of the symbionts are considered to vary depending on the collection site of the host. Difficulties in achieving axenic cells and maintenance of axenic strains for long periods have been reported for symbiotic algae from P. bursaria isolated in Japan. To establish axenic algal strains from such Japanese P. bursaria, symbionts were isolated carefully, and isolated axenic strains were grown on an agar medium containing organic nitrogen compounds. Symbiotic algal strains were obtained from three Japanese P. bursaria strains and their axenicity was confirmed by DAPI staining, cultural tests of bacterial contamination, and DGGE-PCR. These axenic strains have been maintained for over 2 years. Utilization of carbohydrates and nitrogen compounds by symbionts was examined. Monosaccharides (glucose and fructose) increased the growth of the symbiont but disaccharides (maltose and sucrose) did not. Japanese axenic symbionts were able to use ammonia and amino acids, but not nitrate or nitrite. While potent nitrite reductase activity was stimulated by nitrate induction, nitrate reductase activity was not. Nitrate utilization of Japanese symbionts differed from that reported for European and American symbionts.     

Molecular data do not support a southern hemisphere base of Nothofagus powdery mildews

Three powdery mildew species present on Nothofagus (viz. Erysiphe magellanica, E. nothofagi and E. patagoniaca) are endemic to South America and have unique ascomatal appendages that are not found in powdery mildews of the northern hemisphere. We determined the nucleotide sequences of the rDNA internal transcribed spacer regions and D1/D2 domains of the 28S rDNA of these three powdery mildew species to reveal their phylogenetic relationships with powdery mildews of the northern hemisphere. Although the molecular phylogenetic analyses indicated that the three Nothofagus powdery mildews are closely related to each other they did not group into one clade in either the ITS or 28S trees. Kishino-Hasegawa, Shimodaira-Hasegawa and Templeton tests could not significantly reject the constrained trees that were constructed based on the assumption that the Nothofagus powdery mildews would form a single clade. Based on this result and the evidence that all Nothofagus powdery mildews are endemic to South America and have similar morphological characteristics, it is likely that these three species diverged from a single ancestor present on Nothofagus. Calibration of evolutionary events with molecular clocks suggested that the Nothofagus powdery mildews split from the northern hemisphere relatives 22-16 million y ago (Ma) in the middle Miocene, and divergence among the Nothofagus powdery mildews occurred 17-13 Ma. These results do not support a southern hemisphere base of the Nothofagus powdery mildews.     

Increase in urinary markers during the acute phase reflects the degree of chronic tubulointerstitial injury after ischemia-reperfusion renal injury

Context: Acute kidney injury (AKI) could lead to progressive chronic kidney disease (CKD). Objectives: To demonstrate that urinary markers in AKI are associated with the degree of persistent renal injury. Material and methods: Human L-FABP chromosomal transgenic (T_g) mice were subjected to ischemia-reperfusion (I/R) clamping renal pedicle for 20?min or 30?min. Kidneys were obtained at one and 40 days after I/R. Results: Urinary L-FABP, NGAL, Kim-1 and albumin levels increased during the acute phase and were significantly correlated with the degree of tubulointerstitial fibrosis during the chronic phase. Discussion and conclusion: These markers could detect higher risk of progression to CKD.