Essential role of rho kinase in the ca2+ Sensitization of Prostaglandin F2α-Induced Contraction of Rabbit Aortae

Although the prostate gland is a rich source of α1-adreno- (α1-AR) and m1-cholino receptors (m1-AChR), the membrane processes associated with their activation in glandular epithelial cells is poorly understood. We used the whole-cell patch-clamp technique to show that the agonists of the respective receptors, phenylephrine (PHE) and carbachol (CCh), activate cationic membrane currents in lymph node carcinoma of the prostate (LNCaP) human prostate cancer epithelial cells, which are not dependent on the filling status of intracellular IP₃-sensitive Ca²⁺ stores, but directly gated by diacylglycerol (DAG), as evidenced by the ability of its membrane permeable analogue, OAG, to mimic the effects of the agonists. The underlying cationic channels are characterized by the weak field-strength Eisenman IV permeability sequence for monovalent cations ( P _K(25) > P _Cs(4.6) > P _Li(1.4) > P _Na(1.0)), and the following permeability sequence for divalent cations: P _Ca(1.0) > P _Mg(0.74) > P _Ba(0.6) > P _Sr(0.36) > P _Mn(0.3). They are 4.3 times more permeable to Ca²⁺ than Na⁺ and more sensitive to the inhibitor 2-APB than SK&F 96365. RT-PCR analysis shows that DAG-gated members of the transient receptor potential (TRP) channel family, including TRPC1 and TRPC3, are present in LNCaP cells. We conclude that, in prostate cancer epithelial cells, α1-ARs and m1-AChRs are functionally coupled to Ca²⁺-permeable DAG-gated cationic channels, for which TRPC1 and TRPC3 are the most likely candidates.     

Vinyl polymerization. 294. Decomposition of tetramethyltetrazene in the presence of p-substituted benzyl chlorides

A study was made of the polymerization of acrylonitrile in dimethylformamide (DMF) initiated by the binary systems of tetramethyltetrazene (TMT) and p-substituted benzyl chlorides. The polymerization rate increased linearly with the σ-constants of substituents as electron-releasing groups were introduced to the phenyl ring of benzyl chloride. In order to elucidate the initiation mechanism of the polymerization, a kinetic investigation was also undertaken of the decomposition of TMT in the presence of p-substituted benzyl chlorides in DMF. The decomposition rate was first-order in TMT and first-order in p-substituents benzyl chloride. The decomposition rate also increased with the σ constants of substituents as electron-releasing groups were introduced. On the basis of the results, the initiation mechanism for the polymerization was discussed.     

Late-onset obesity in mice transgenic for the human renin gene

We previously generated a strain of transgenic mice carrying the human renin gene, hRN8-12, in the background of C57BL/6j. In this study, we discovered that hRN8-12 male mice, but not females, developed obesity starting at 15 weeks of age. The body weight of 60-week-old male transgenic mice was 2 times higher than that of age-matched wild-type mice. Interestingly, male mice heterozygous for the human renin gene showed moderate weight gain compared with transgenic and wild-type mice. Obese hRN8-12 mice exhibited hyperglycemia, hyperinsulinemia, hyperleptinemia, and hyperlipidemia, and increase in weight in the adipose tissue, liver, heart, and kidneys. Histological analysis demonstrated that fatty hRN8-12 mice developed hypertrophy of pancreatic islets and fatty liver. These results suggested that hRN8-12 mice are associated with obesity dependent on the transgene dosage and should be a genetic model for late-onset obesity.     

Studies on epidermal growth factor receptor signaling in vertebrate limb patterning.

The epidermal growth factor receptor (EGFR) regulates multiple patterning events in Drosophila limb development, but its role in vertebrate limb morphogenesis has received little attention. The EGFR and several of its ligands are expressed in developing vertebrate limbs in manners consistent with potential patterning roles. To gain insight into functions of EGFR signaling in vertebrate limb development, we expressed a constitutively active EGFR in developing chick limbs in ovo. Expression of activated EGFR causes pre- and postaxial polydactyly, including mirror-image-type digit duplication, likely due to induction of ectopic expression and/or modulation of genes involved in anterior-posterior (AP) patterning such as Sonic hedgehog (Shh), dHand, Patched (Ptc), Gli3, Hoxd13, Hoxd11, bone morphogenetic protein 2 (Bmp2), Gremlin, and FGF4. Activation of EGFR signaling dorsalizes the limb and alters expression of the dorsal-ventral (DV) patterning genes Wnt7a, Lmx, and En1. Ectopic and/or extended FGF8 expressing apical ectodermal ridges (AERs) are also seen. Interdigital regression is inhibited and the digits fail to separate, leading to syndactyly, likely due to antiapoptotic and pro-proliferative effects of activated EGFR signaling on limb mesoderm, and/or attenuation of interdigital Bmp4 expression. These findings suggest potential roles for EGFR signaling in AP and DV patterning, AER formation, and cell survival during limb morphogenesis.     

Preparation and characterization of fluorine-containing aromatic condensation polymers, 4. Preparation and characterization of fluorine-containing aromatic polyazomethines and copolyazomethines from perfluoroisopropylidene group-containing aromatic diamines and/or isopropylidene group-containing aromatic diamines and phthaladehydes

A series of novel fluorine-containing aromatic homopolyazomethines and copolyazomethines were synthesized by solution polycondensation of perfluoroisopropylidene (perfluoropropane-2,2-diyl) group-containing aromatic diamines, 2,2-bis[4-(4-aminophenoxy)phenyl]-1,1,1,3,3,3-hexafluoropropane (4,4′-[1,1,1,3,3,3-hexafluoropropane-2,2-diylbis(1,4-phenyleneoxy)]dianiline ( 1a )) and 2,2-bis(4-aminophenyl)-1,1,1,3,3,3-hexafluoropropane (4,4′-(1,1,1,3,3,3-hexafluoropropane-2,2-diyl)dianiline ( 1b )), and/or the corresponding analogous isopropylidene group-containing aromatic diamines, 2,2-bis[4-(4-aminophenoxy)phenyl]propane (4,4′-[isopropylidenebis(1,4-phenyleneoxy)]dianiline ( 1c )) and 2,2-bis(4-aminophenyl)propane (4,4′-isopropylidenedianiline ( 1d )), with terephthalaldehyde ( 2a ) and isophthaladehyde ( 2b ). The effect of fluorine substitution on the synthesis and properties of these polymers was investigated by comparison with those of the corresponding homopolyazomethines but without fluorine. Film-forming fluorine-containing polyazomethines with reduced viscosities up to η_red = 0,44 dL · g^?1 were obtained in high yields by using m-cresol as reaction medium. Incorporation of perfluoroisopropylidene groups in the polyazomethine backbone greatly enhanced the solubility of the polymers in various organic solvents including chloroform and tetrahydrofuran. Their thermal stability was also successfully improved by the introduction of fluorine, and both the temperatures of 5% and 10% weight loss in air increased monotonically with the increase of the fluorine content. The surface properties of the films or the contact angles formed by water and diiodomethane were, however, scarcely affected by fluorine substitution. The mechanical properties of the films, such as tensile strength and initial modulus, fairly decreased by the modification.     

Flavonoids such as luteolin, fisetin and apigenin are inhibitors of interleukin-4 and interleukin-13 production by activated human basophils

BACKGROUND: We have previously shown that fisetin, a flavonol, inhibits IL-4 and IL-13 synthesis by allergen- or anti-IgE-antibody-stimulated basophils. This time, we investigated the inhibition of IL-4 and IL-13 production by basophils by other flavonoids and attempted to determine the fundamental structure of flavonoids related to inhibition. We additionally investigated whether flavonoids suppress leukotriene C4 synthesis by basophils and IL-4 synthesis by T cells in response to anti-CD3 antibody. METHODS: Highly purified peripheral basophils were stimulated for 12 h with anti-IgE antibody alone or anti-IgE antibody plus IL-3 in the presence of various concentrations of 18 different kinds of flavones and flavonols. IL-4 and IL-13 concentrations in the supernatants were then measured. Leukotriene C4 synthesis was also measured after basophils were stimulated for 1 h in the presence of flavonoids. Regarding the inhibitory activity of flavonoids on IL-4 synthesis by T cells, peripheral blood mononuclear cells were cultured with flavonoids in anti-CD3-antibody-bound plates for 2 days. RESULTS: Luteolin, fisetin and apigenin were found to be the strongest inhibitors of both IL-4 and IL-13 production by basophils but did not affect leukotriene C4 synthesis. At higher concentrations, these flavonoids suppressed IL-4 production by T cells. Based on a hierarchy of inhibitory activity, the basic structure for IL-4 inhibition by basophils was determined. CONCLUSIONS: Due to the inhibitory activity of flavonoids on IL-4 and IL-13 synthesis, it can be expected that the intake of flavonoids, depending on the quantity and quality, may ameliorate allergic symptoms or prevent the onset of allergic diseases.     

Gastric juvenile polyposis associated with germline SMAD4 mutation

We treated a 39-year-old woman with hypoproteinemia and anemia who had profuse gastric polyposis. Radiographic and endoscopic examination showed numerous polyps restricted to the stomach. The patient had pulmonary arteriovenous malformations in the left lung. Histological examination of the resected stomach revealed the gastric polyposis to be composed of cystic dilatation of the glands with small areas of adenocarcinoma. These findings were compatible with gastric juvenile polyposis (GJP) accompanied by gastric cancer. Analysis of genomic DNA revealed that the patient had truncating mutation of SMAD4, a responsible gene for juvenile polyposis (JP). Our case suggests that SMAD4 is possibly a responsible gene for GJP.     

Ultrastructural cytochemistry of proteoglycans associated with calcification of shark cartilage

Proteoglycans (PGs) as well as sulfated glycosaminoglycans (GAGs) are closely associated with cartilage calcification. An inner zone of endoskeletal tesserae of sharks is composed of a unique calcified hyaline cartilage. Initial calcification can be seen in the cartilage close to the inner zone. We have ultrastructurally examined shark, Triakis scyllia, noncalcifying, calcifying, and calcified cartilage using the tannic acid-ferric chloride (TA-Fe), the high iron diamine (HID), and the HID-thiocarbohydrazide-silver proteinate (HID-TCH-SP) methods for localization of sulfated complex carbohydrates. In noncalcifying cartilage, TA-Fe and HID strongly stained matrix granules which were round, ovoid, elongated, or irregularly shaped and presumably represented PG monomers. The size and staining intensity of the reactive matrix granules progressively decreased in calcifying cartilage toward the calcification front of the calcified cartilage. Similarly, a progressive decrease in the size of the HID-TCH-SP stain deposits in the matrix granules was observed in the calcifying cartilage close to the calcification front and was interpreted as a decrease in length of sulfate containing GAG chains. In the calcified cartilage, the highly calcified areas were often localized in the calcification front and contained few or no small HID-TCH-SP stain deposits, whereas the weakly calcified regions contained more stain deposits. These results indicate that partial and complete degradation of sulfated GAGs and/or PGs may be a requisite for calcification of shark cartilage.     

Deproteinizing effects on resin-tooth bond structures

This study investigated the effects of NaOCl on resin-tooth bonds to simulate the situations of long-term durability and caries invasion. Resin-tooth bonded specimens were produced with the use of two resin adhesives (Excite and One-Bond). Resin-tooth bonded beams (adhesive area; 0.9 mm2) were serially sectioned and the specimens were immersed in 10% NaOCl medium for 0 (control), 2, 4, and 6 h after being stored in water for 24 h. After immersion, microtensile bond tests were performed. SEM fractography was conducted to calculate each failure mode by image analysis. In addition, the adhesive interface was examined with the use of TEM. In the control specimens, enamel bond strengths had no difference between Excite (45.6 +/- 15.0) and One-Bond (56.9 +/- 12.9). On the other hand, dentin bond strengths had significant difference between Excite (80.6 +/- 21.2) and One-Bond (50.7 +/- 11.2). The bond strengths decreased with increased storage time for both systems with enamel and dentin bonds. The deteriorated mineralized dentin of beams resulted in bond-strength reduction for resin-enamel bonds. For dentin bonding, the adhesive interface was gradually dissolved from the outer to the center portion of the beam. The depletion of collagen fibrils within the demineralized dentin or hybrid layer deformation was found under SEM and TEM examinations. These morphological changes are responsible for bond strength reduction of resin-dentin bonds.