[Liver Fibrosis is Associated with Loss of Skeletal Muscle Mass in Community-Dwelling Older Adults with No History of Liver Diseases].

OBJECTIVES: In this study, we aimed to examine the possible association between liver fibrosis and loss of skeletal muscle mass (SM) in community-dwelling older adults with no history of liver diseases. METHODS: A total of 2,028 older adults (mean age, 69.8 ± 5.2 years) who had not received any treatment for liver diseases and had participated in a comprehensive health survey for community residents in Wakayama, Japan were included in this study. We carried out bioelectrical impedance analysis to estimate the SM of the whole body including the arms, legs, and trunk of the subjects. Liver fibrosis was evaluated by calculating the Fib4 index based on the subject's age, AST level, ALT level, and platelet counts. RESULTS: The subjects were divided into three groups according to cutoff values of the Fib4 index (low: <1.30, medium: 1.30-2.66, high: ≥2.67). The SM index (kg/m2) was the lowest among subjects in the high-Fib4-index category, followed by the medium- and low-Fib4-index categories. This dose-response reduction in the SM index was more pronounced among individuals with lower blood albumin level (low nutrition) and in those with more sedentary behavior (physical inactivity). Among the selected 262 subjects who underwent SM measurement twice with an interval of 3 years, the subjects with a high Fib4 index showed greater reduction in the SM index than those with medium and low Fib4 indices. Multiple regression analysis revealed that the Fib4 index was significantly associated with the SM index, independent of age, sex, albumin level, sedentary behavior, diabetes mellitus, alcohol intake, and smoking status. CONCLUSIONS: The present findings suggest that the potential progression of liver fibrosis is associated with the excessive loss of SM among apparently healthy older adults without any treatment for liver diseases.

Pooled extracellular receptor-ligand interaction screening using CRISPR activation.

Extracellular interactions between cell surface receptors are necessary for signaling and adhesion but identifying them remains technically challenging. We describe a cell-based genome-wide approach employing CRISPR activation to identify receptors for a defined ligand. We show receptors for high-affinity antibodies and low-affinity ligands can be unambiguously identified when used in pools or as individual binding probes. We apply this technique to identify ligands for the adhesion G-protein-coupled receptors and show that the Nogo myelin-associated inhibitory proteins are ligands for ADGRB1. This method will enable extracellular receptor-ligand identification on a genome-wide scale.

A novel growth function incorporating the effects of reproductive energy allocation.

Ontogenetic growth functions provide basic information in biological and ecological studies. Various growth functions classified into the Pütter model have been used historically, regardless of controversies over their appropriateness. Here, we present a novel growth function for fish and aquatic organisms (generalised q-VBGF) by considering an allocation schedule of allometrically produced surplus energy between somatic growth and reproduction. The generalised q-VBGF can track growth trajectories in different life history strategies from determinate to indeterminate growth by adjusting the value of the 'growth indeterminacy exponent' q. The timing of maturation and attainable body size can be adjusted by the 'maturation timing parameter' τ while maintaining a common growth trajectory before maturation. The generalised q-VBGF is a comprehensive growth function in which exponentials in the traditional monomolecular, von Bertalanffy, Gompertz, logistic, and Richards functions are replaced with q-exponentials defined in the non-extensive Tsallis statistics, and it fits to actual data more adequately than these conventional functions. The relationship between the estimated parameter values τ and rq forms a unique hyperbola, which provides a new insight into the continuum of life history strategies of organisms.

A CRISPR Dropout Screen Identifies Genetic Vulnerabilities and Therapeutic Targets in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is an aggressive cancer with a poor prognosis, for which mainstream treatments have not changed for decades. To identify additional therapeutic targets in AML, we optimize a genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) screening platform and use it to identify genetic vulnerabilities in AML cells. We identify 492 AML-specific cell-essential genes, including several established therapeutic targets such as DOT1L, BCL2, and MEN1, and many other genes including clinically actionable candidates. We validate selected genes using genetic and pharmacological inhibition, and chose KAT2A as a candidate for downstream study. KAT2A inhibition demonstrated anti-AML activity by inducing myeloid differentiation and apoptosis, and suppressed the growth of primary human AMLs of diverse genotypes while sparing normal hemopoietic stem-progenitor cells. Our results propose that KAT2A inhibition should be investigated as a therapeutic strategy in AML and provide a large number of genetic vulnerabilities of this leukemia that can be pursued in downstream studies.

On the analytical solution for the Pütter-Bertalanffy growth equation.

This study develops the basic idea of Pütter and Bertalanffy addressing the allometric scaling of anabolism and catabolism on somatic growth dynamics. We proposed a standardized form of the Pütter-Bertalanffy equation (PBE), which is given as the extended model of Richards function, and subsequently solved it. The analytical solution of the PBE was defined by an incomplete beta function and can take a wide range of shapes in its growth curve. The mathematical behavior of PBE due to the change in parameter values was briefly discussed. Most forms of solution consistently hold the implicit functional type with respect to the variable of body size.

Molecular dynamics study on helium nanobubbles in water.

Interfacial properties of helium nanobubbles in water at normal conditions have been investigated using large-scale molecular dynamics simulations for systems including over one million atoms. The surface tension of a helium nanobubble is a convex function with respect to the bubble radius, and is estimated to vanish at a critical radius of approximately 1 nm.

Nano bubbles in liquid of a noble-gas mixture.

Large-scale molecular dynamics (MD) simulations with over one million atoms are used to investigate nano bubbles in Ar-Ne liquid. The simulations demonstrate cavitations in the stretched liquid, and bubble creation and collapse. We find that a small cavity created in the stretched liquid spontaneously transforms into a nano bubble with the homogeneous vapor region. The equilibrium spherical bubble of 11.4 nm in radius is obtained after the long-time MD run. The surface tension of the nano bubble is found to be larger than that of the flat surface.

Synthesis and evaluation of 1-arylsulfonyl-3-piperazinone derivatives as factor Xa inhibitors V. A series of new derivatives containing a spiro[imidazo[1,2-a]pyrazine-2(3H),4'-piperidin]-5(1H)-one scaffold.

We have already reported unique compounds containing a N,O-spiro acetal structure as an orally active factor Xa (FXa) inhibitor. This time, we described a N,N-spiro acetal structure as an analogue of the N,O-spiro acetal structure for an orally active FXa inhibitor. The synthesis of these analogues could be achieved in a similar fashion to the N,O-spiro acetal synthesis. Consequently, FXa inhibitory activity was increased and more active compounds could be found (M58163: IC50 = 0.61 nM, M58169: IC50 = 0.58 nM). Additionally, the absolute configuration could be determined by X-ray crystallography analysis (M58169: (R)-config.).

What kinds of substrates show P-glycoprotein-dependent intestinal absorption? Comparison of verapamil with vinblastine.

The influence of P-glycoprotein (P-gp) on intestinal absorption of drugs was investigated by comparison of the uptakes of two P-gp substrates, verapamil and vinblastine, using intestinal segments of wild-type and mdr1a/1b gene-deficient (mdr1a/1b(-/-)) mice, and Caco-2 cells. When [(3)H]vinblastine was injected into intestinal segments of wild-type mice, vinblastine was absorbed from duodenum and ileum, but not from jejunum. This difference among intestinal regions could not be explained by segmental differences of mdr1a mRNA expression. In Caco-2 cells, it was found that vinblastine had a high value of efflux/influx ratio (an index of affinity for P-gp) of 12.1, and a low permeability of less than 1 x 10(-6) cm/sec. The corresponding values for verapamil were 4.9 and 10.6 x 10(-6) cm/sec, respectively. After oral administration of [(3)H]vinblastine to mice, the maximum concentration (C(max)) and the area under the plasma concentration time-curve from time 0 to 24 hr (AUC(0-24 hr)) for mdr1a/1b(-/-) mice were 1.5 times greater than those for wild-type mice, while these parameters were not significantly different between the two strains in the case of [(3)H]verapamil. Therefore, P-gp substrates may be classified into at least two types, i.e., verapamil-type, for which the intestinal absorption is unaffected by P-gp, and vinblastine-type, for which the intestinal absorption is influenced by P-gp. Vinblastine-type P-gp substrates, with low permeability and high affinity for P-gp, would be unfavorable candidates for oral drugs.

Improving effect of ethyl eicosapentanoate on statin-induced rhabdomyolysis in Eisai hyperbilirubinemic rats.

The effect of ethyl eicosapentanoate (EPA-E) on statin-induced rhabdomyolysis was investigated by co-administration of EPA-E and pravastatin (PV), as a typical statin, to Eisai hyperbilirubinemic rats (EHBR). It was confirmed that the plasma PV concentration was not affected by simultaneous administration of EPA-E, and there was no cumulative increase of PV during prolonged co-administration of EPA-E and PV. Muscular degeneration was prominent (incidence 5/5; average grade 3.5 (range 2-4)) in EHBR treated with PV alone at 200 mg/kg/day for 14 days, but co-administration of EPA-E at doses of 100, 300, and 1000 mg/kg/day decreased the average grades to 1.4 (range 0.3-3.0), 0.5 (0.2-1.0), and 0.6 (0.0-1.7), respectively. Creatine phosphokinase (CPK) and myoglobin levels in plasma were well correlated with the grade of skeletal muscle degeneration. Thus, EPA-E appears to reduce the severity of statin-induced rhabdomyolysis.

Eisai hyperbilirubinemic rat (EHBR) as an animal model affording high drug-exposure in toxicity studies on organic anions.

The Eisai hyperbilirubinemic rat (EHBR) should be a useful animal model for studies on the toxicity of organic anions which are substrates of multidrug resistance-associated protein 2 (Mrp2), since the systemic exposure to these compounds is expected to be increased in EHBR. In this study, we tested the value of EHBR for this purpose, using pravastatin (PV) and methotrexate (MTX) as model compounds. In the case of a single oral dose of PV (200 mg/kg), C(max) in plasma was 4.0-fold higher and AUC(0-infinity) was 3.6-fold larger than those of normal Sprague-Dawley rats (SDR), respectively. When multiple doses of PV were given to EHBR without co-administration of any other compound, drug-induced skeletal muscle toxicity (myopathy/rhabdomyolysis) and increased creatine phosphokinase (CPK) level were observed, whereas a control experiment using SDR did not show any toxic change. When a single dose of MTX (0.6 mg/kg) was given to EHBR orally, C(max) was 1.7-fold higher and AUC(0-infinity) was 1.6-fold larger than those of SDR, respectively. When multiple doses of MTX were given to EHBR, the changes in bone marrow, spleen and intestines were more severe than those in SDR. These findings support the view that EHBR would be a valuable animal model for toxicity studies on organic anion compounds which are substrates of Mrp2.

Synthesis and evaluation of 1-arylsulfonyl-3-piperazinone derivatives as factor Xa inhibitors IV. A series of new derivatives containing a spiro[5H-oxazolo[3,2-a]pyrazine-2(3H),4'-piperidin]-5-one skeleton.

In the course of development of factor Xa (FXa) inhibitor in an investigation involving the synthesis of 1-arylsulfonyl-3-piperazinone derivatives, we found new compounds containing a unique spiro skeleton. Among such compounds, (-)-7-[(6-chloro-2-naphthalenyl)sulfonyl]tetrahydro-8a-(methoxymethyl)-1'-(4-pyridinyl)-spiro[5H-oxazolo[3,2-a]pyrazine-2(3H),4'-piperidin]-5-one (28, M55529) had activity more favorable than those of previously reported compounds. The inhibitory activity of M55529 for FXa is IC(50)=2 nM, with high selectivity for FXa over thrombin and trypsin.

Synthesis and evaluation of 1-arylsulfonyl-3-piperazinone derivatives as factor Xa inhibitors III. Effect of ring opening of piperazinone moiety on inhibition.

Compounds containing an ethylenediamine structure in place of the piperazine ring of M55113 (1) and M55551 (2) were synthesized to investigate the effects of a piperazine moiety and evaluated for activity as factor Xa (FXa) inhibitors. Most such compounds, however, exhibited lower activity (1/10-1/100) than that of M55113 and M55551 as FXa inhibitors.

Functional characterization of active transport of progesterone to adrenal cells.

The characterization of the transport mechanism of progesterone, which is one of the neutral steroids in the adrenal cells, has been studied by the examination of progesterone uptake into the monolayers of SW-13 cells (a human adrenal adenocarcinoma cell line). The uptake of [(3)H]progesterone at a tracer concentration (1 nM) exhibited temperature, pH and sodium dependency. According to kinetic analysis of the concentration dependence, the uptake of progesterone involves saturable and non-saturable processes. The uptake for the saturable process, which gave K(t) values (half-saturation concentration) of 4.7 +/- 8.7 microM, was inhibited by metabolic inhibitors and amino-acid modifiers but not by endocytosis inhibitors or substrates for known transporters. The uptake of progesterone was also inhibited by several neutral steroids but not by anionic steroids. The inhibition by both beta-estradiol and estriol was competitive. The uptake of progesterone by the adrenal cells might be at least partially accounted for by a specific carrier-mediated transport mechanism generated by sodium ions and an electrochemical mechanism.

Synthesis and evaluation of 1-arylsulfonyl-3-piperazinone derivatives as a factor Xa inhibitor II. Substituent effect on biological activities.

Intravascular clot formation is an important event in a number of cardiovascular diseases. The prevention of blood coagulation has become a major target for new therapeutic agents. Factor Xa (FXa) is a trypsin-like serine protease that plays a key role in the blood coagulation cascade and represents an attractive target for anticoagulant drug development. We have investigated substituents in the central part of a lead compound (3: M55113), and discovered that compound M55551 (34: (R)-4-[(6-Chloro-2-naphthalenyl)sulfonyl]-6-oxo-1-[[1-(4-pyridinyl)-4-piperidinyl]methyl]-2-piperazinecarboxylic acid) is a potent inhibitor of FXa (IC(50)=0.006 microM), with high selectivity for FXa over trypsin and thrombin. The activity of this compound is ten times more powerful than the lead compound.