👤 Yu Sasaki

Expression of intragenic exon rearrangements (IERs) has reportedly been detected in both normal and cancer cells. However, there have been few reports of occurrence of these rearrangements specific to neoplasms including malignant lymphoma. In this study, we detected IERs of ten genes (NBPF8, SOBP, AUTS2, RAB21, SPATA13, ABCC4, WDR7, PHLPP1, NFATC1 and MAGED1) in non-Hodgkin B cell lymphoma (B-NHL) cell line KPUM-UH1 using a high-resolution single nucleotide polymorphism array and reverse transcription polymerase chain reaction using reversely directed divergent primers within exons involved in genomic intragenic gains followed by sequencing analysis. Among them, the IERs involved in SOBP (6q21) exon 2 and 3 and AUTS2 (7q11.22) exon 2-4 were the molecular lesions specific to tumors and were frequently detected in B-NHL samples. These IERs constitute novel genetic alterations of B-NHL, which might be associated with tumorigenesis and be useful as genetic biological markers. Show less

Propranolol is a widely used beta blocker that consists of a racemic mixture of R and S stereoisomers. Only the S stereoisomer has significant activity against the beta-adrenergic receptor. A fortuitous clinical observation was made in an infant who received propranolol for cardiac disease, and regression of a hemangioma of infancy was noted. This has led to the widespread use of propranolol for the treatment of large and life-threatening hemangiomas of infancy. Infants receiving propranolol require monitoring to ensure that they do not suffer from side effects related to beta blockade. The exact mechanism of activity of propranolol in hemangioma of infancy is unknown. In this study, we treated hemangioma stem cells with both beta blockade active S- and inactive R-propranolol and looked for genes that were coordinately regulated by this treatment. Among the genes commonly downregulated, Angiopoietin-like 4 (ANGPTL4) was among the most regulated. We confirmed that propranolol isomers downregulated ANGPTL4 in endothelial cells, with greater downregulation of ANGPTL4 using the beta blockade inactive R-propranolol. ANGPTL4 is present in human hemangiomas of infancy. Finally, R-propranolol inhibited the growth of bEnd.3 hemangioma cells in vivo. The implication of this is that hemangioma growth can be blocked without the side effects of beta blockade. Given that humans have been exposed to racemic propranolol for decades and thus to R-propranolol, clinical development of R-propranolol for hemangiomas of infancy and other angiogenic diseases is warranted. Show less

Long-chain polyunsaturated fatty acids (LCPUFAs) are important constituents of biomembranes. Observation of blood fatty acids indicated that LCPUFA biosynthesis is affected by aging and FADS polymorphisms. This study examined the effects of aging and FADS polymorphisms on LCPUFA biosynthetic capacity via direct quantification using [U- Show less

Rett syndrome (RTT) is a characteristic neurological disease presenting with regressive loss of neurodevelopmental milestones. Typical RTT is generally caused by abnormality of methyl-CpG binding protein 2 ( We performed WES on 77 Pathogenic or likely pathogenic single-nucleotide variants in 28 known genes were found in 39 of 77 (50.6%) patients. WES-based CNV analysis revealed pathogenic deletions involving six known genes (including Our study provides a new landscape including additional genetic variants contributing to RTT-like phenotypes, highlighting the importance of comprehensive genetic analysis. Show less

Genome-wide association studies (GWASs) have identified several susceptibility loci for bipolar disorder (BD) and shown that the genetic architecture of BD can be explained by polygenicity, with numerous variants contributing to BD. In the present GWAS (Phase I/II), which included 2964 BD and 61 887 control subjects from the Japanese population, we detected a novel susceptibility locus at 11q12.2 (rs28456, P=6.4 × 10 Show less

To date, mutations in 15 actin- or microtubule-associated genes have been associated with the cortical malformation lissencephaly and variable brainstem hypoplasia. During a multicenter review, we recognized a rare lissencephaly variant with a complex brainstem malformation in three unrelated children. We searched our large brain-malformation databases and found another five children with this malformation (as well as one with a less severe variant), analyzed available whole-exome or -genome sequencing data, and tested ciliogenesis in two affected individuals. The brain malformation comprised posterior predominant lissencephaly and midline crossing defects consisting of absent anterior commissure and a striking W-shaped brainstem malformation caused by small or absent pontine crossing fibers. We discovered heterozygous de novo missense variants or an in-frame deletion involving highly conserved zinc-binding residues within the GAR domain of MACF1 in the first eight subjects. We studied cilium formation and found a higher proportion of mutant cells with short cilia than of control cells with short cilia. A ninth child had similar lissencephaly but only subtle brainstem dysplasia associated with a heterozygous de novo missense variant in the spectrin repeat domain of MACF1. Thus, we report variants of the microtubule-binding GAR domain of MACF1 as the cause of a distinctive and most likely pathognomonic brain malformation. A gain-of-function or dominant-negative mechanism appears likely given that many heterozygous mutations leading to protein truncation are included in the ExAC Browser. However, three de novo variants in MACF1 have been observed in large schizophrenia cohorts. Show less

A comprehensive genome-wide screen of radiosensitization targets in HeLa cells was performed using a shRNA-library/functional cluster analysis and DNMT3B was identified as a candidate target. DNMT3B RNAi increased the sensitivity of HeLa, A549 and HCT116 cells to both γ-irradiation and carbon-ion beam irradiation. DNMT3B RNAi reduced the activation of DNA damage responses induced by γ-irradiation, including HP1β-, γH2AX- and Rad51-foci formation. DNMT3B RNAi impaired damage-dependent H2AX accumulation and showed a reduced level of γH2AX induction after γ-irradiation. DNMT3B interacted with HP1β in non-irradiated conditions, whereas irradiation abrogated the DNMT3B/HP1β complex but induced interaction between DNMT3B and H2AX. Consistent with radiosensitization, TP63, BAX, PUMA and NOXA expression was induced after γ-irradiation in DNMT3B knockdown cells. Together with the observation that H2AX overexpression canceled radiosensitization by DNMT3B RNAi, these results suggest that DNMT3B RNAi induced radiosensitization through impairment of damage-dependent HP1β foci formation and efficient γH2AX-induction mechanisms including H2AX accumulation. Enhanced radiosensitivity by DNMT3B RNAi was also observed in a tumor xenograft model. Taken together, the current study implies that comprehensive screening accompanied by a cluster analysis enabled the identification of radiosensitization targets. Downregulation of DNMT3B, one of the targets identified using this method, radiosensitizes cancer cells by disturbing multiple DNA damage responses. Show less

Recent studies applying high-throughput sequencing technologies have identified several recurrently mutated genes and pathways in multiple cancer genomes. However, transcriptional consequences from these genomic alterations in cancer genome remain unclear. In this study, we performed integrated and comparative analyses of whole genomes and transcriptomes of 22 hepatitis B virus (HBV)-related hepatocellular carcinomas (HCCs) and their matched controls. Comparison of whole genome sequence (WGS) and RNA-Seq revealed much evidence that various types of genomic mutations triggered diverse transcriptional changes. Not only splice-site mutations, but also silent mutations in coding regions, deep intronic mutations and structural changes caused splicing aberrations. HBV integrations generated diverse patterns of virus-human fusion transcripts depending on affected gene, such as TERT, CDK15, FN1 and MLL4. Structural variations could drive over-expression of genes such as WNT ligands, with/without creating gene fusions. Furthermore, by taking account of genomic mutations causing transcriptional aberrations, we could improve the sensitivity of deleterious mutation detection in known cancer driver genes (TP53, AXIN1, ARID2, RPS6KA3), and identified recurrent disruptions in putative cancer driver genes such as HNF4A, CPS1, TSC1 and THRAP3 in HCCs. These findings indicate genomic alterations in cancer genome have diverse transcriptomic effects, and integrated analysis of WGS and RNA-Seq can facilitate the interpretation of a large number of genomic alterations detected in cancer genome. Show less

Multidrug resistance protein 2 (MRP2, ABCC2) is localized to the apical membrane of hepatocytes and played an important role in the biliary excretion of a broad range of endogenous and xenobiotic compounds and drugs, such as pravastatin. However, the effects of statins on MRP2 in the liver and the precise mechanisms of their actions have been obscure. The goal of this study was to determine the regulatory molecular mechanism for statin-induced MRP2 expression in hepatocytes. In vitro and in vivo studies suggested that pitavastatin increased MRP2 expression. Pitavastatin promoted liver X receptor (LXR) α/β translocation from the cytosol to nuclei, resulting in LXR activation. Deletion and mutational analysis suggested that the potential sterol regulatory element (SRE) played a major role in the observed modulation of MRP2 expression by pitavastatin. Furthermore pitavastatin increased the protein-DNA complex, and when SRE was mutated, stimulation of the protein-DNA complex by pitavastatin was decreased. It was demonstrated that pitavastatin upregulated MRP2 expression by an SREBP regulatory pathway in hepatocytes and that the actions of statins may lead to improve the biliary excretion of MRP2 substrates. Show less

The lens of the eye is composed of fiber cells, which differentiate from epithelial cells and undergo programmed organelle degradation during terminal differentiation. Although autophagy, a major intracellular degradation system, is constitutively active in these cells, its physiological role has remained unclear. We have previously shown that Atg5-dependent macroautophagy is not necessary for lens organelle degradation, at least during the embryonic period. Here, we generated lens-specific Atg5 knock-out mice and showed that Atg5 is not required for lens organelle degradation at any period of life. However, deletion of Atg5 in the lens results in age-related cataract, which is accompanied by accumulation of polyubiquitinated and oxidized proteins, p62, and insoluble crystallins, suggesting a defect in intracellular quality control. We also produced lens-specific Pik3c3 knock-out mice to elucidate the possible involvement of Atg5-independent alternative autophagy, which is proposed to be dependent on Pik3c3 (also known as Vps34), in lens organelle degradation. Deletion of Pik3c3 in the lens does not affect lens organelle degradation, but it leads to congenital cataract and a defect in lens development after birth likely due to an impairment of the endocytic pathway. Taken together, these results suggest that clearance of lens organelles is independent of macroautophagy. These findings also clarify the physiological role of Atg5 and Pik3c3 in quality control and development of the lens, respectively. Show less

Semaphorin3A (Sema3A) exerts a wide variety of biological functions by regulating reorganization of actin and tubulin cytoskeletal proteins through signaling pathways including sequential phosphorylation of collapsin response mediator protein 1 (CRMP1) and CRMP2 by cyclin-dependent kinase-5 and glycogen synthase kinase-3β (GSK3β). To delineate how GSK3β mediates Sema3A signaling, we here determined the substrates of GSK3β involved. Introduction of either GSK3β mutants, GSK3β-R96A, L128A, or K85M into chick dorsal root ganglion (DRG) neurons suppressed Sema3A-induced growth cone collapse, thereby suggesting that unprimed as well as primed substrates are involved in Sema3A signaling. Axin-1, a key player in Wnt signaling, is an unprimed substrate of GSK3β. The phosphorylation of Axin-1 by GSK3β accelerates the association of Axin-1 with β-catenin. Immunocytochemical studies revealed that Sema3A induced an increase in the intensity levels of β-catenin in the DRG growth cones. Axin-1 siRNA knockdown suppressed Sema3A-induced growth cone collapse. The reintroduction of RNAi-resistant Axin-1 (rAxin-1)-wt rescued the responsiveness to Sema3A, while that of nonphosphorylated mutants, rAxin S322A/S326A/S330A and T485A/S490A/S497A, did not. Sema3A also enhanced the colocalization of GSK3β, Axin-1, and β-catenin in the growth cones. The increase of β-catenin in the growth cones was suppressed by the siRNA knockdown of Axin-1. Furthermore, either Axin-1 or β-catenin RNAi knockdown suppressed the internalization of Sema3A. These results suggest that Sema3A induces the formation of GSK3β/Axin-1/β-catenin complex, which regulates signaling cascade of Sema3A via an endocytotic mechanism. This finding should provide clue for understanding of mechanisms of a wide variety of biological functions of Sema3A. Show less

In liver, glucose utilization and lipid synthesis are inextricably intertwined. When glucose availability exceeds its utilization, lipogenesis increases, leading to increased intrahepatic lipid content and lipoprotein secretion. Although the fate of three-carbon metabolites is largely determined by flux rate through the relevant enzymes, insulin plays a permissive role in this process. But the mechanism integrating insulin receptor signaling to glucose utilization with lipogenesis is unknown. Forkhead box O1 (FoxO1), a downstream effector of insulin signaling, plays a central role in hepatic glucose metabolism through the regulation of hepatic glucose production. In this study, we investigated the mechanism by which FoxO1 integrates hepatic glucose utilization with lipid synthesis. We show that FoxO1 overexpression in hepatocytes reduces activity of carbohydrate response element binding protein (Chrebp), a key regulator of lipogenesis, by suppressing O-linked glycosylation and reducing the protein stability. FoxO1 inhibits high glucose- or O-GlcNAc transferase (OGT)-induced liver-pyruvate kinase (L-PK) promoter activity by decreasing Chrebp recruitment to the L-PK promoter. Conversely, FoxO1 ablation in liver leads to the enhanced O-glycosylation and increased protein level of Chrebp owing to decreased its ubiquitination. We propose that FoxO1 regulation of Chrebp O-glycosylation is a mechanism linking hepatic glucose utilization with lipid synthesis. Show less

ATP-binding cassette transporters (ABC) A1 and G1 are key molecules in cholesterol efflux from macrophages, which is an initial step of reverse cholesterol transport (RCT), a major anti-atherogenic property of high-density lipoprotein (HDL). Astaxanthin is one of the naturally occurring carotenoids responsible for the pink-red pigmentation in a variety of living organisms. Although astaxanthin is known to be a strong antioxidant, it remains unclear through what mechanism of action it affects cholesterol homeostasis in macrophages. We therefore investigated the effects of astaxanthin on cholesterol efflux and ABCA1/G1 expressions in macrophages. Astaxanthin enhanced both apolipoprotein (apo) A-I- and HDL-mediated cholesterol efflux from RAW264.7 cells. In supporting these enhanced cholesterol efflux mechanisms, astaxanthin promoted ABCA1/G1 expression in various macrophages. In contrast, peroxisome proliferator-activated receptor γ, liver X receptor (LXR) α and LXRβ levels remained unchanged by astaxanthin. An experiment using actinomycin D demonstrated that astaxanthin transcriptionally induced ABCA1/G1 expression, and oxysterol depletion caused by overexpression of cholesterol sulfotransferase further revealed that these inductions in ABCA1/G1 were independent of LXR-mediated pathways. Finally, we performed luciferase assays using human ABCA1/G1 promoter-reporter constructs to reveal that astaxanthin activated both promoters irrespective of the presence or absence of LXR-responsive elements, indicating LXR-independence of these activations. In conclusion, astaxanthin increased ABCA1/G1 expression, thereby enhancing apoA-I/HDL-mediated cholesterol efflux from the macrophages in an LXR-independent manner. In addition to the anti-oxidative properties, the potential cardioprotective properties of astaxanthin might therefore be associated with an enhanced anti-atherogenic function of HDL. Show less

Pioglitazone, a peroxisome proliferator-activated receptor γ (PPARγ) agonist, reportedly reduces cardiovascular events in diabetic patients. ATP cassette binding transporters (ABC) A1 and G1 are pivotal molecules for cholesterol efflux (ChE) from macrophages and high density-lipoprotein biogenesis, and the A1 transporter is regulated by a PPARγ-liver receptor X (LXR) pathway. Also, pioglitazone induces ABCG1 expression, though the exact mechanism remains unclear. We therefore investigated the effects of pioglitazone on ABCA1/G1 expression in vitro and ex vivo. The effects of pioglitazone on ChE and ABCA1/G1 expressions in macrophages were assessed. Then, mRNA was quantified in macrophages when PPARγ/LXR inhibition by siRNA or overexpression of oxysterol sulfotransferase was performed. ABCA1/G1 promoter activity with mutated LXR-responsive elements was also measured. As an ex vivo study, 15 type 2 diabetic patients were administered pioglitazone or placebo, and ChE assays and protein expressions were determined using macrophages cultured with the corresponding sera. Pioglitazone increased LXRα/ABCA1/G1 expressions, which enhanced ChE from macrophages. Inhibition of PPARγ/LXR pathways revealed that LXR was primarily involved in pioglitazone's transactivation of ABCA1 but only partially involved for ABCG1. Promoter assays showed that ABCG1 was regulated more by the promoter in intron 4 than that upstream of exon 1 but both promoters were responsive to LXR activation. Sera obtained after pioglitazone treatment promoted ChE and ABCA1/G1 expressions in macrophages. Pioglitazone enhanced ChE from macrophages by increasing ABCA1/G1 in LXR-dependent and -independent manners. Our comparable in vitro and ex vivo results shed new light on pioglitazone's novel anti-atherogenic property. Show less

The study shows constitutive activation of the Notch pathway in various types of malignancies. However, it remains unclear how the Notch pathway is involved in the pathogenesis of osteosarcoma. We investigated the expression of the Notch pathway molecules in osteosarcoma biopsy specimens and examined the effect of Notch pathway inhibition. Real-time PCR revealed overexpression of Notch2, Jagged1, HEY1, and HEY2. On the other hand, Notch1 and DLL1 were downregulated in biopsy specimens. Notch pathway inhibition using gamma-secretase inhibitor and CBF1 siRNA slowed the growth of osteosarcomas in vitro. In addition, gamma-secretase inhibitor-treated xenograft models exhibited significantly slower osteosarcoma growth. Cell cycle analysis revealed that gamma-secretase inhibitor promoted G1 arrest. Real-time PCR and western blot revealed that gamma-secretase inhibitor reduced the expression of accelerators of the cell cycle, including cyclin D1, cyclin E1, E2, and SKP2. On the other hand, p21(cip1) protein, a cell cycle suppressor, was upregulated by gamma-secretase inhibitor treatment. These findings suggest that inhibition of Notch pathway suppresses osteosarcoma growth by regulation of cell cycle regulator expression and that the inactivation of the Notch pathway may be a useful approach to the treatment of patients with osteosarcoma. Show less

Activation of the Wnt signaling pathway is frequently observed in hepatocellular carcinoma (HCC), though mutation of three of its components, CTNNB1, AXIN1, and AXIN2, is observed substantially less often. We examined the relationship between Wnt signaling and epigenetic alteration of secreted frizzled-related protein (SFRP) genes in HCC. We frequently detected the active form of beta-catenin and accumulation of nuclear beta-catenin in liver cancer cell lines. We detected methylation of SFRP family genes in liver cancer cell lines (SFRP1, 9/12, 75%; SFRP2, 7/12, 58%; SFRP4, 3/12, 25%; SFRP5, 7/12, 58%) and primary HCCs (SFRP1, 9/19, 47%; SFRP2, 12/19, 63%; SFRP5, 8/19, 42%), though methylation of SFRP4 was not found in primary HCCs. SFRP methylation also was detected in hepatitis B or C virus-associated chronic hepatitis (SFRP1, 6/37, 16%; SFRP2, 14/37, 38%; SFRP5, 5/37, 14%) and liver cirrhosis (SFRP1, 10/28, 36%; SFRP2, 9/28, 32%; SFRP5, 3/28, 11%), suggesting that methylation of these genes is an early event in liver carcinogenesis. Ectopic expression of SFRPs downregulated T-cell factor/lymphocyte enhancer factor (TCF/LEF) transcriptional activity in liver cancer cells, while overexpression of a beta-catenin mutant and depletion of SFRP1 using siRNA synergistically upregulated TCF/LEF transcriptional activity. Our results confirm the frequent methylation and silencing of Wnt antagonist genes in HCC, and suggest that their loss of function contributes to activation of Wnt signaling during hepatocarcinogenesis. Show less

Although mutations of APC, CTNNB1 (beta-catenin) and AXIN1 are rare in oral squamous cell carcinoma (OSCC), activation of the Wnt signaling pathway is thought to play an important role in oral carcinogenesis. In the present study, we examined the relationship between Wnt signaling and epigenetic alteration of the secreted frizzled-related protein (SFRP) genes in OSCC. We frequently detected loss of membrane localization of beta-catenin and its cytoplasmic or nuclear accumulation in OSCC cell lines, although these cell lines showed no APC or CTNNB1 (beta-catenin) mutations and no methylation of CDH1 (E-cadherin). By contrast, we frequently detected methylation of SFRP1 (7/17, 41%) SFRP2 (16/17, 94%) and SFRP5 (14/17, 82%) in a panel of OSCC cell lines, as well as in specimens of primary tumors collected from 44 OSCC patients (SFRP1, 10/42, 24%; SFRP2, 16/44, 36%; SFRP5, 7/43, 16%). We also observed that OSCC cell lines express various Wnt ligands, and that ectopic expression of SFRPs inhibited cancer cell proliferation. Our results confirm the frequent methylation and silencing of SFRP genes in OSCC, and suggest that their loss of function contributes to activation of Wnt signaling that leads to cell proliferation during oral carcinogenesis. Show less

Extracellular signal-regulated kinase 5 (ERK5), the newest member of the mitogen-activated protein (MAP) kinase family, is regulated differently than the other MAP kinases. Emerging evidence suggest the role of ERK5 signaling in promoting cell proliferation, differentiation, neuronal survival, and neuroprotection. The present study investigates whether suicide brain is associated with alterations in components of the ERK5 signaling cascade. In the prefrontal cortex (PFC) and hippocampus of suicide subjects (n=28) and nonpsychiatric control subjects (n=21), we examined the catalytic activities and protein levels of ERK5 and upstream MAP kinase kinase MEK5 in various subcellular fractions; mRNA levels of ERK5 in total RNA; and DNA-binding activity of myocyte enhancer factor (MEF)2C, a substrate of ERK5. In the hippocampus of suicide subjects, we observed that catalytic activity of ERK5 was decreased in cytosolic and nuclear fractions, whereas catalytic activity of MEK5 was decreased in the total fraction. Further, decreased mRNA and protein levels of ERK5, but no change in protein level of MEK5 were noted. A decrease in MEF2C-DNA-binding activity in the nuclear fraction was also observed. No significant alterations were noted in the PFC of suicide subjects. The observed changes were not related to a specific psychiatric diagnosis. Our findings of reduced activation and/or expression of ERK5 and MEK5, and reduced MEF2C-DNA-binding activity demonstrate abnormalities in ERK5 signaling in hippocampus of suicide subjects and suggest possible involvement of this aberrant signaling in pathogenic mechanisms of suicide. Show less

We studied the association of six common polymorphisms of four genes related to lipid metabolism with serum lipid levels. We selected single-nucleotide polymorphisms (SNPs) in the genes for cholesteryl ester transfer protein (CETP), lipoprotein lipase (LPL), hepatic lipase (LIPC), and apolipoprotein CIII (APOC3), and studied 2267 individuals randomly selected from the participants of Serum Lipid Survey 2000. There was a significant association of CETP polymorphism (D442G, Int14 +1 G --> A, and TaqIB), LPL polymorphism (S447X), and LIPC polymorphism (-514 --> CT) with HDL-cholesterol levels. We also found a significant association of LPL polymorphism (S447X) and APOC3 polymorphism (SstI) with triglyceride levels. This is the largest database showing the association of common genetic variants in lipid metabolism with serum lipid levels in the general Japanese population. Further study is necessary to elucidate the role of these gene polymorphisms in cardiovascular events. Show less

To clarify the genetic aberrations involved in the development and progression of hepatitis C virus-associated hepatocellular carcinoma (HCV-HCC), we investigated DNA copy number aberrations (DCNAs) in 19 surgically resected HCCs by conventional CGH and array CGH. Conventional CGH revealed that increases of DNA copy number were frequent at 1q (79% of the cases), 8q (37%), 6p (32%), and 10p (32%) and that decreases were frequent at 17p (79%), 16q (58%), 4q (53%), 13q (42%), 10q (37%), 1p (32%), and 8p (32%). In general, genes that showed DCNAs by array CGH were usually located in chromosomal regions with DCNAs detected by conventional CGH analysis. Increases in copy numbers of the LAMC2, TGFB2, and AKT3 genes (located on 1q) and decreases in copy numbers of FGR/SRC2 and CYLD (located on 1p and 16q, respectively) were observed in more than 30% of tumors, including small, well-differentiated carcinomas. These findings suggest that these genes are associated with the development of HCV-HCC. Increases of MOS, MYC, EXT1, and PTK2 (located on 8q) were detected exclusively in moderately and poorly differentiated tumors, suggesting that these alterations contribute to tumor progression. In conclusion, chromosomal and array CGH technologies allow identification of genes involved in the development and progression of HCV-HCC. Show less

The Wnt signaling pathway is essential for development and organogenesis. Wnt signaling stabilizes beta-catenin, which accumulates in the cytoplasm, binds to 1-cell factor (TCF; also known as lymphocyte enhancer-binding factor, LEF) and then upregulates downstream genes. Mutations in CTNNB1 (encoding beta-catenin) or APC (adenomatous polyposis coli) have been reported in human neoplasms including colon cancers and hepatocellular carcinomas (HCCs). Because HCC5 tend to show accumulation of beta-catenin more often than mutations in CTNNB1, we looked for mutations in AXIN1, encoding a key factor for Wnt signaling, in 6 HCC cell lines and 100 primary HCC5. Among the 4 cell lines and 87 HCC5 in which we did not detect CTNNB1 mutations, we identified AXIN1 mutations in 3 cell lines and 6 mutations in 5 of the primary HCCs. In cell lines containing mutations in either gene, we observed increased DNA binding of TCF associated with beta-catenin in nuclei. Adenovirus mediated gene transfer of wild-type AXINI induced apoptosis in hepatocellular and colorectal cancer cells that had accumulated beta-catenin as a consequence of either APC, CTNNB1 or AXIN1 mutation, suggesting that axin may be an effective therapeutic molecule for suppressing growth of hepatocellular and colorectal cancers. Show less

A comparative assessment has been made regarding efficacy and safety of the double filtration plasmapheresis (DFPP), thermofiltration (TFPP), and low-density lipoprotein (LDL) adsorptive (PA) methods by making a crossover test on heterozygous familial hypercholesterolemia patients. Treatments by DFPP, TFPP (secondary membrane Evalux 5A), and PA (Liposorber LA-40) were carried out 5 times each, with a 2-week interval, in 5 patients with heterozygous familial hypercholesterolemia. The same plasma separator (Plasmacure PS-60, polysulfone) was used in all cases, and the volume of plasma processed was set at 4 L. High removal rates were obtained of total cholesterol, LDL cholesterol, triglycerides TG, and apolipoprotein B (apoB) by all three methods, and no differences were observed. Lipoprotein (a), apoA-2, apoC-3, fibrinogen, and immunoglobulin M (IgM) showed significantly high removal rates by the DFPP and TFPP methods compared with the PA method. The sieving coefficient of albumin and high-density lipoprotein (HDL) cholesterol at 2 and 4 L of plasma processed exhibited high permeabilities using all three methods. Supplementing albumin was not necessary. An increase of the transmembrane pressure was observed in 1 case treated by DFPP but was not observed when using the TFPP or PA method. No changes were observed in serum interleukin 1beta (IL-1beta) or tumor necrosis factor-alpha (TNF-alpha) before and after treatment by any of the three methods. No remarkable side effects were observed using either the DFPP or TFPP method. The DFPP and TFPP methods showed efficacy and safety that was not inferior to the PA method in conventional LDL apheresis, and the dead-end method of the filter operation without the discarding of plasma was shown to be possible. Show less