The inhibitory effects of Momordica charantia extracts were studied on the uptake of glucose and tyrosine across rat everted gut sacs in vitro. The aqueous extract of the plant was found to inhibit primarily the uptake of glucose in a dose-dependent manner. Uptake of tyrosine was affected at high substrate concentrations only. The extract was also found to decrease the absorptive capacity of fluid across the small intestine and sodium ions. It is hypothesized that the effects of Momordica could involve a washout of glucose from the blood stream.
Nox1, a homologue of gp91phox subunit of the phagocyte NADPH oxidase, is responsible for spontaneous superoxide (O2−) generation in guinea pig gastric mucosal cells (GMC), but involvement of regulatory components (p67phox, p47phox, and Rac) which are essential in phagocytes remains unknown. Here, we aimed to figure out how Nox1 of GMC achieves an active oxidase status. GMC in primary culture show low O2− generation but acquire a 9-fold higher activity when cultured with Helicobacter pylori lipopolysaccharide (LPS), in correlation with a far increased Nox1 expression. Investigation into the O2−-generating ability of LPS-induced Nox1 in cell-free reconstitution assays showed that: 1) Nox1 is unable to generate O2−per se; 2) the combination of Nox1 with GMC cytosol is insufficient for a significant O2− generation; 3) the combination with neutrophil cytosol enables Nox1 to act like gp91phox, i.e., to produce O2− appreciably in response to myristate stimulation; 4) Nox1 prefers NADPH to NADH under the in vitro assay with neutrophil cytosol plus myristate (Km=10.4 μM); 5) substitution of neutrophil cytosol by a set of recombinant cytosolic components (rp67phox, rp47phox, Rac2) is, however, ineffective and still requires GMC cytosol. Thus, Nox1 probably requires an additional cytosolic factor(s). In contrast, GMC cytosol enables cytochrome b558 to generate plenty of O2−, on condition that rp47phox is added. This result suggests that GMC cytosol contains a component with p67phox-ability, and also Rac, but lacks p47phox. These data indicate that GMC Nox1 requires at least a p67phox counterpart and Rac to acquire NADPH oxidase activity.
The thrombopoietin (TPO) receptor (Mpl) belongs to the family of ligand-dependent cytokine receptors and plays a functional role in regulating platelet production. The signaling capacity largely depends on the binding of TPO to the extracellular domains of the TPO receptor (Mpl-EC). Because the expression level of Mpl in human tissue is very low, studies on the functional and spatial characteristics of its ligand-binding sites have been limited. In the present study, we report the expression and purification of Mpl-EC as a fusion with the maltose-binding protein (MBP), designated MBP-Mpl-EC. MBP-Mpl-EC was expressed in the cytoplasm of Escherichia coli as a soluble fusion protein. Specific binding of TPO to purified MBP-Mpl-EC was demonstrated by a dot-blot assay and surface plasmon resonance. We conclude that bacterial expression of MBP-Mpl-EC yields large amounts of protein with correct folding and that it can be used for further structure and function analyses.
Using Ig-β and growth hormone producing cells with liver-derived cells for controls, sensitivity of chromatin to DNase I was measured by real-time PCR at eleven targets in rat Ig-β/growth hormone locus where four cell type-specific genes and two ubiquitously expressed genes are present in a compact 88-kb region. Chromatin situated at the promoter of actively-transcribed gene and placed at cell type-specific DNase I hypersensitive sites with enhancer activity was sensitive to DNase I. In the case of inactive gene, chromatin located in these regions was resistant to DNase I. Unexpectedly, however, chromatin placed in the transcribed intron was resistant to DNase I in two genes. DNase I sensitive chromatin was shown not to distribute locus-widely but rather to localize at the promoter and the enhancer of actively-transcribed genes in this locus.
Juzen-taiho-to, a Kampo formula, originally consists of a mixture of Shimotsu-to and Shikunshi-to formulas together with two other crude ingredients. Juzen-taiho-to is reported to have a preventive effect on endometrial carcinogenesis in mice. Shimotsu-to exerts an inhibitory effect on estrogen-induced expression of c-fos, interleukin (IL)-1α and tumor necrosis factor (TNF)-α in uteri of ovarectomized mice. In the present study, short- and long-term experiments were designed to determine the effects of Juzen-taiho-to and Shimotsu-to on the estrogen-related endometrial carcinogenesis in mouse uteri, associated with the expression of cyclooxygenase (COX)-1 and -2. In the short-term experiment, exposure to Juzen-taiho-to or Shimotsu-to significantly reduced estradiol-17β (E2)-stimulated expressions of COX-2 mRNA (p<0.05) as well as the protein. However, no effects on the expression of COX-1 were observed. Shikunshi-to did not affect COX expression. In the long-term experiment, 90 female ICR mice were given N-methyl-N-nitrosourea (MNU) into their uterine corpora. The animals were divided into four groups as follows: group 1, a diet containing 0.07% Shimotsu-to and 5 ppm E2; group 2, a diet containing 5 ppm E2; group 3, a diet containing 0.07% Shimotsu-to; group 4 served as a control. Exposure of Shimotsu-to reduced the incidence of MNU- and E2-induced endometrial adenocarcinoma and atypical hyperplasia at the termination of the experiment (30 weeks). The above findings and our previous reports suggest that Shimotsu-to is responsible for the preventive effects of Juzen-taiho-to on estrogen-related endometrial carcinogenesis in mice, through the inhibition of estrogen-related COX-2 as well as c-fos, IL-1α and TNF-α expressions.
The effects of ethanol extracts of three Chinese medicinal plants Dahuang (Rheum palmatum L.), Badou (Croton tiglium L.), and Huomaren (Cannabis sativa L.), on ion transport of the rat intestinal epithelia were studied. Rat intestinal epithelia mounted in an Ussing chamber attached with voltage/current clamp were used for measuring changes of the short-circuit current across the epithelia. The intestinal epithelia were activated with current raised by serosal administration of forskolin 5 μM. Ethanol extracts of the three plants all augmented the current additively when each was added after forskolin. In subsequent experiments, ouabain and bumetanide were added prior to ethanol extracts of these medicinal plants to determine their effect on Na+ and Cl− movement. The results suggest that ethanol extracts of the three medicinal plants may affect the Cl− movement more directly than Na+ movement in the intestinal epithelial cells. The results provide evidence for the pharmacologic mechanism of the three Chinese medicinal plants on the intestinal tract.
A possible involvement of inhibitory effects of monochlorobimane (MCB) on the opening of mitochondrial permeability transition (MPT) pore in the cerebroprotection against the ischemic brain injury was examined. MCB (1 mM) inhibited the opening of MPT pore in vitro. Sustained cerebral ischemia was induced by injecting 900 microspheres (48 μm in diameter) into the right hemisphere of rats. At 12 to 72 h after microsphere embolism (ME), the mitochondrial activity was determined histochemically by staining cytochrome c oxidase (COX) and succinate dehydrogenase (SDH) of the brain sections. The COX and SDH stainings in the hippocampus were observed intensively in the pyramidal neurons in the CA2-3 and dentate gyrus rather than those in the CA-1 region. The staining was decreased with time after the embolism. Pretreatment with 10 μg/animal MCB 30 min prior to the embolism significantly attenuated the ME-induced reduction in the staining of COX and SDH in the hippocampus, but not in the pariatal cortex. The results suggest that prevention of the opening of MPT pore by MCB may play an important role in the cerebroprotection against cerebral ischemic injury.
Curcumin manganese complex (CpCpx) and diacetylcurcumin manganese complex (AcylCpCpx) were determined as to their effect on the nitric oxide (NO) radical scavenging in vitro method using a sodium nitroprusside generating NO system compared with their parent compound and astaxanthin, an extreme antioxidant. All compounds effectively reduced the generation of NO radicals in a dose dependent manner. They exhibited strong NO radical scavenging activity with low IC50 values. The IC50 values of curcumin, diacetylcurcumin, CpCpx and AcylCpCpx obtained are 20.39±4.10 μM, 28.76±1.48 μM, 9.79±1.50 μM and 8.09±0.99 μM, respectively. CpCpx and AcylCpCpx show greater NO radical scavenging than their parent compounds, curcumin and acetylcurcumin, respectively. However, the IC50 values of curcumin and related compounds were found to be less than astaxanthin, an extreme antioxidant, with the lower IC50 value of 3.42±0.50 μM.
Although recent studies have suggested that dentate granule cells play a key role in hippocampal functions, electrophysiological properties in these cells have not been sufficiently explored. In the present study, modification of voltage-gated Ca2+ channels by 4-hydroxynonenal (4HN), a major aldehydic product of membrane lipid peroxidation, in cultured dentate granule cells was examined using the whole-cell patch clamp technique. When whole-cell voltage clamp was applied, the cells exhibited a high-voltage-activated Ca2+ current, which was totally sensitive to 30 μM Cd2+ and partially sensitive to 2 μM nifedipine. 4HN enhanced the Ca2+ current in these cells. When L-type Ca2+ channels were blocked by application of nifedipine, the enhancement was completely canceled, whereas application of ω-conotoxin-GVIA or ω-agatoxin-IVA, blockers of N- and P/Q-type Ca2+ channels, respectively, had no effect. These results suggest that 4HN modulates L-type Ca2+ channels in the dentate granule cells, and thereby plays a role in the physiological and pathophysiological responses of these cells to oxidative stress.
Comparative pharmacokinetic profiles of glycyrrhetic acid (GA), glycyrrhizic acid (GL) and Gancao-Fuzi-Tang (KF) after oral administration of GL and KF were studied. Plasma samples taken from rats were acidified with acetic acid and GA was extracted with isopropanol–ethyl ether (1 : 1). Separation of GA was performed on a C18 column with the detection wavelength set at 254 nm. The mobile phase was methanol–acetonitrile–water–acetic acid (58 : 18 : 24 : 1 v/v). The results showed that the mean residence time and area under the curve of GA in KF-administered rats were 27.6±1.5 h and 122.8±46.7 μg·h/ml respectively, which were significantly different from those in GL-administered rats (15.0±2.0 h and 40.9±9.6 μg·h/ml, respectively). The results suggest the increased effect of GA after oral administration of KF in comparison with GL.
The cardiac toxicity of amitriptyline and the effect of the light schedule on it were studied in chick embryos. Fertilized eggs of White Leghorns were incubated under dark conditions and investigated, on two occasions, in the light phase and in the dark phase. Amitriptyline was injected into the air sac of fertilized eggs on the 16th day of incubation. Electrocardiograms were recorded 0 to 60 min after the injection. After the administration of amitriptyline 1 mg/egg in the light phase, the heart rate did not differ compared with that in controls. However, the heart rate was significantly decreased by the administration of amitriptyline 2.5 mg/egg and 5 mg/egg in the light phase. The heart rate was significantly decreased by the administration of amitriptyline 1 mg/egg under dark conditions. In addition, arrhythmia was produced by the administration of amitriptyline under dark conditions. These findings indicate that manipulation of the light schedule has a marked influence on the toxicity of amitriptyline in chick embryos.
The effect of β-cryptoxanthin on bone components in the femoral tissues of rats was investigated. β-Cryptoxanthin was isolated from Satsuma mandarin (Citrus unshiu MARC.). Bone tissues were cultured for 48 h in serum-free Dulbecco's modified Eagle's medium containing either vehicle or β-cryptoxanthin (10−7 or 10−6 M). The presence of β-cryptoxanthin (10−7 or 10−6 M) caused a significant increase in calcium content and alkaline phosphatase activity in the femoral-diaphyseal and femoral-metaphyseal tissues. These increases were completely abolished in the presence of cycloheximide (10−6 M), an inhibitor of protein synthesis. Thus β-cryptoxanthin had an anabolic effect on bone calcificantion in vitro. Moreover, β-cryptoxanthin (10, 25, or 50 μg/100 g body weight) was orally administered once daily for 7 d to young male rats. The administration of β-cryptoxanthin (10, 25, or 50 μg/100 g body weight) caused a significant increase in calcium content and alkaline phosphatase activity in the femoral-diaphyseal and femoral-metaphyseal tissues. Femoral-diaphyseal and femoral-metaphyseal DNA contents were significantly increased by the dose of 25 or 50 μg/100 g body weight. A significant increase in metaphyseal DNA content was also seen with the dose of 10 μg/100 g body weight of β-cryptoxanthin. This study demonstrates that β-cryptoxanthin has an anabolic effect on bone components in rats in vitro and in vivo.
We investigated the effect of transglutaminase (TGase) on in guinea pigs and rats. Serum alanine aminotransferase (ALT) level increased 1 d after CCl4 treatment of both in guinea pigs and rats since TGaese activity was greatly higher in guinea pigs than rats. However, serum ALT level in guinea pigs was very much lower than that in rats. Liver TGase activities decreased after CCl4 treatment in both guinea pigs and rats. However, TGase activities in the liver from guinea pigs were higher than that from rats. Decreased TGase activities by CCl4 in the liver from guinea pigs and rats were significantly recovered by retinoic acid treatment that was reported to increase TGase. Degree of recovery of serum ALT level by retinoic acid in rats was larger than in guinea pigs. These results suggested that the distinction of the effect of retinoic acid on serum ALT level in CCl4-treated animals was due to the different TGase activity that increased membrane stability.
We examined the basal adenosine 3′,5′-cyclic monophosphate (cAMP) levels and forskolin-induced cAMP accumulation in cultured Chinese hamster ovary cells (CHO) expressing different levels of human β2-adrenoceptors. Both the basal and forskolin-induced cAMP accumulation in the cells that express higher density of β2-adrenoceptors (CHO-β2/H; 560 fmol/mg protein) were larger than those in the cells that express lower density of β2-adrenoceptors (CHO-β2/L; 270 fmol/mg protein). In addition, isoproterenol-induced cAMP accumulation was also augmented as the number of β2-adrenoceptors was increased. ICI 118,551, a selective β2-adrenoceptor antagonist with inverse agonist properties, decreased all the levels of cAMP observed in both cell lines. These results suggest that the agonist-independent (constitutive) activity of β2-adrenoceptors plays a key role in the control of forskolin-induced cAMP accumulation.
The objective of this study was to investigate the effects of Gelidium amansii, an edible red agar cultivated off the northeast coast of Taiwan, on the growth of two lines of cancer cells, murine hepatoma (Hepa-1) and human leukemia (HL-60) cells, as well as a normal cell line, murine embryo fibroblast cells (NIH-3T3). The potential role of G. amansii on the induction of apoptosis was also examined. The results indicated that all extracts from G. amansii, including phosphate-buffered saline (PBS) and methanol extracts from dried algae as well as the dimethyl sulfoxide (DMSO) extract from freeze-dried G. amansii agar, inhibited the growth of Hepa-1 and NIH-3T3 cells, but not the growth of HL-60 cells. Annexin V-positive cells were observed in methanol and DMSO extract-treated, but not PBS extract-treated Hepa-1 and NIH-3T3 cells, suggesting that the lipid-soluble extracts of G. amansii induced apoptosis. In summary, extracts of G. amansii from various preparations exhibited antiproliferative effects on Hepa-1 and NIH-3T3 cells, and apoptosis may play a role in the methanol and DMSO extract-induced inhibitory effects. However, the antiproliferative effects of PBS extracts was not through apoptosis. Moreover, the growth-inhibitory effects of G. amansii were not specific to cancer cells.
Two glucuronides (4′-O-, and 7-O-) and a glucuronyl (7-O-) sulfate (4′-O-) of genistein, two glucuronides (4′-O-, and 7-O-) and a glucuronyl (7-O-) sulfate (4′-O-) of daidzein, 7-O-glucuronides of glycitein, dihydrodaidzein and O-desmethylangolensin were isolated from the urine of volunteer subjects fed soy bean curds (Tofu). The estrogenic activities, i.e., i) the effect on the estrogen-dependent growth of MCF-7 cells, ii) the binding ability to human estrogen receptors (hERs) α and β, and iii) the effect on hER-dependent β-galactosidase induction, of these isoflavone metabolites were examined. Two synthetic isoflavone aglycones (dihydrodaidzein and O-desmethylangolensin) and four synthetic sulfates (4′-O- and 4′-, 7-di-O-) of genistein and daidzein were also studied for their estrogenic activities for the purpose of comparison. With respect to estrogenic acivity, the tested isoflavone metabolites were classified into three groups. The first group shows a very poor stimulatory effect toward the growth of MCF-7 cells, binding activity, and β-galactosidase induction. The sulfates belong to this group. The second group shows a moderate binding activity but poor stimulation and β-galactosidase induction. Some glucuronyl conjugates belong to this group. The last group shows a moderate stimulation and β-galactosidase induction but poor binding activity. A mixed type of conjugates having glucuronyl and sulfony moieties belong to this group.
Previously we demonstrated that chlorophyllin suppressed the genotoxicities of many carcinogens. However, the genotoxicity of IQ (2-amino-3-methylimidazo[4,5-f]quinoline), a carcinogenic heterocyclic amine, was not suppressed in Drosophila. On the contrary, it has been reported that chrolophyllin suppressed the genotoxicity of IQ in rodents, rainbow trout and Salmonella. We demonstrated that the chlorophyllin-induced suppression of MeIQx (2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline)-genotoxicity was associated with a decrease in MeIQx-DNA adduct formation in Drosophila larval DNA. MeIQx represents another type of heterocyclic amine and is similar to IQ in structure. In this study we utilized 32P-postlabeling to examine whether chlorophyllin reduced IQ-DNA adduct formation in Drosophila DNA in the same way as MeIQx. The results revealed that the formation of IQ-DNA adducts was unaffected by treatment with chlorophyllin. This was consistent with the absence of any inhibitory effect on genotoxicity as observed in the Drosophila repair test. These results suggest that IQ-behavior in Drosophila is not affected by chlorophyllin, indicating that the process of IQ-DNA adduct formation followed by expression of genotoxicity in Drosophila may be different from that in other organisms.
Substitution of 2′,6′-dimethyltyrosine (Dmt) for the N-terminal Tyr in opioid peptides has recently been shown to be a promising tool for improving opioid receptor affinity and biological activity. We have also demonstrated that another unnatural amino acid, 2′,6′-dimethylphenylalanine (Dmp), is not only an excellent substitute for Phe at position 3 but also can mimic the aromatic N-terminal Tyr residue in a μ opioid receptor-selective dermorphin analogue (YRFB: Tyr-D-Arg-Phe-βAla-NH2). To further evaluate the value of Dmp in opioid peptides, we investigated Dmp1-substituted analogues of the δ receptor ligands, deltorphin II (DLT: Tyr-D-Ala-Phe-Glu-Val-Val-Gly-NH2) and enkephalin (ENK: Tyr-Gly-Gly-Phe-Leu). In the receptor binding assay, both [Dmp1]DLT and [Dmp1]ENK bound to the δ-receptor with high affinity and selectivity, and were nearly as effective as the parent peptides. The potency of the Dmp1-peptides on the MVD and GPI assays correlated well with the receptor binding affinity data. These results are in contrast to the tendency of corresponding Dmt1-analogues to have poor receptor selectivity. Taken together with the results with YRFB, we conclude that the Dmp1-peptide is superior to the corresponding Dmt1-peptide in its receptor selectivity. [Dmp1]DLT and [Dmp1]YRFB may serve as pharmacological tools for the studies of ligand recognition and opioid receptor signal transduction.
We have shown that Protease N treated Royal Jelly (ProRJ) and peptides from ProRJ (Ile-Tyr (IY), Val-Tyr (VY), Ile-Val-Tyr (IVY)) inhibited angiotensin I-converting enzyme (ACE) activity and they have an antihypertensive effect in repeated oral administration for 28 d on spontaneously hypertensive rats (SHR). We investigated the contributive ratio of these peptides in ProRJ for antihypertensive effect in single oral administration on SHR. In single oral administration of each peptide and peptides mixture (MIX; IY, VY and IVY) at doses of 0.5, 1 and 10 mg/kg, systolic blood pressure (SBP) of SHR was reduced dose-dependently. This antihypertensive effect was held for 8 h. These results suggest that peptides contributed to the antihypertensive effect of ProRJ. And the contributive ratio of MIX in ProRJ for antihypertensive effect was computed to be about 38%. Therefore it is considered that intake of peptides, as a functional food would be beneficial for improving blood pressure in people with hypertension.
The constituents of the aerial parts of Physalis angulata (Solanaceae) were investigated based on the plant's trypanocidal activity against epimastigotes of Trypanosoma cruzi, the etiologic agent for Chagas' disease. Four new withanolides were isolated, along with six known ones, from the active fraction. Their structures were determined by spectroscopic analysis. Trypanocidal activity against trypomastigotes, an infectious form of T. cruzi, was also estimated, as well as cytotoxic activity against human uterine carcinoma (HeLa) cells in vitro. Evaluation of trypanocidal activity using the colorimetric reagent Cell Counting Kit-8 was also examined.
The antidiabetic activity of Momordica charantia L. (Cucurbitaceae) with exercise was investigated in KK-Ay mice, an animal model with type 2 diabetes with hyperinsulinemia. The water extract of the fruit of Momordica charantia L. (MC) with exercise reduced the blood glucose of KK-Ay mice 5 weeks after oral administration (p<0.001), and also significantly lowered the plasma insulin of KK-Ay mice under similar conditions (p<0.01). The blood glucose of MC with exercise is lower than that of MC only or exercise only 5 weeks after the administration. MC with exercise decreased blood glucose in a glucose tolerance test. These results suggest that MC with exercise is useful for type 2 diabetic cure.
Eighteen main compounds, including four norsesquiterpenoids (1—4) and 14 phenolic compounds (5—18) isolated previously from Phyllanthus emblica, together with a main constituent, proanthocyanidin polymers (19) identified at this time from the roots, were estimated for their antiproliferative activities against MK-1 (human gastric adenocarcinoma), HeLa (human uterine carcinoma), and B16F10 (murine melanoma) cells using an MTT method. All of the phenolic compounds including the major components 5—8 from the fruit juice, 8, 9, and 12 from the branches and leaves, and 19 from the roots showed stronger inhibition against B16F10 cell growth than against HeLa and MK-1 cell growth. Norsesquiterpenoid glycosides 3 and 4 from the roots exhibited significant antiproliferative activities, although their aglycon 1 and monoglucoside 2 showed no inhibitory activity against these tumor cells.
We found lymphocyte proliferating substances in the water-soluble and non-dialyzable fraction prepared from the crude drug Senso (Chan Su). The effect of this fraction was increased by affinity chromatography using the concanavalin A–agarose. By analyzing the fraction using sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) and lectin blotting, we estimated that one of the active substances of this fraction is a glycoprotein that has about 13 kDa of molecular weight and D-mannose within the molecule. The purified fraction increased the IL-2 and the IL-12 level in the supernatant of spleen cell culture, and increased the natural killer activity of spleen lymphocyte in C3H/HeN mice. These results show that Senso contains immunopotentiating substances that may serve as an immunomodulator in an organism.
Objective: Curcumin is a wide-spectrum cellular protector with antiinflammatory, antioxidizant, and antifibrotic effects. This study was conducted to investigate its effects on myocardial collagen remodeling in pressure overloaded rabbits. Methods and Results: Pressure overloaded rabbits were established by partial abdominal aorta ligation. The rabbits were divided into the sham-operation group, vehicle group and curcumin group. Curcumin was administered orally at a dose of 100 mg/kg·d in 10 ml of 2.5% polyethylene glycol solution and the other 2 groups were given the same dose of polyethylene glycol solution. Compared with the vehicle group, left ventricular function in the curcumin group was significantly ameliorated, as indicated by decreased left ventricular end-diastolic pressure, left ventricle weight to body weight ratio, and the left ventricular posterior wall thickness. The collagen volume fraction in the curcumin group was also reduced. Myocardial tumor necrosis factor (TNF)-α and matrix metalloproteinase (MMP)-2 expression were significantly overexpressed in the vehicle group and markedly suppressed in the curcumin group at both the 4th and 8th weeks. At the end of the 8th week, the ejection fraction in the curcumin group was increased compared with that in the vehicle group. Conclusion: Curcumin improved left ventricular function in pressure overloaded rabbits. This might be duo to inhibition of collagen remodeling associated with suppression of myocardial expression of tumor necrosis factor-α, and matrix metalloproteinase-2.
Although alcohol consumption is a factor in which the bioavailability of saquinavir (SQV) are retarded, the cause for this phenomenon remains to be uncertain. In the presence study, we examined factors to decrease plasma concentration of SQV in ethanol-treated rats. The ethanol-treated rats were prepared by making them freely access to 15% ethanol solution for 14 d (Day 14 rats). The exsorption clearance of SQV from the blood circulation to the jejunal lumen in the Day 14 rats increased by 6-fold as compared to ethanol non-treated (NT) rats. In the presence of 25 μM ketoconazole (KCZ) or 10 μM cyclosporin A (CsA) in the jejunal lumen, the plasma concentration of SQV in the portal vein increased significantly, and this effect of 10 μM CsA was superior to that of 25 μM KCZ. The biliary excretion clearance of SQV in Day 14 rats also increased by 1.8-fold as compared to that in the NT rats. The metabolic clearance rate (Vmax/Km) of SQV in the intestinal microsomes from the Day 14 rats increased significantly, while in the liver microsomes the Vmax/Km did not change. The phase II metabolism processes in the Day 14 rats based on UDP-glucuronosyltransferases and gultathion-S-tnrasferase activities were activated, however, they were not likely to be one of factors to decrease the bioavailability of oral SQV, because CYP3A activity in the liver and intestine was not activated to such an extent and SQV itself was not conjugated. These observations suggest that a main possible factor to explain the reducing effect on the SQV oral bioavailability during ethanol consumption is an enhanced efflux of SQV at the intestine and liver, where it is suggested that functional enhancement or excessive expression of P-glycoprotein is caused by ethanol consumption.
The pharmacokinetic parameters of lopinavir (LPV) were examined by administering Kaletra (LPV+ritonavir) to 8 healthy Japanese volunteers both in the fasting and postprandial conditions. LPV showed a biphasic decline, which was slower in the initial phase and became more rapid in the later phase. The behavior of LPV in the initial phase could be modeled using a one-compartment model with first-order absorption. In the fasting study, calculations based on the pharmacokinetic model revealed that the time to reach the maximum concentration (Tmax), maximum concentration (Cmax), half-life (T1/2), lag time, apparent volume of distribution (Vd/F) and oral clearance (Cl/F) were 3.2±1.0 h, 6.9±1.9 μg/ml, 10.0±3.7 h, 0.71±0.32 h, 51.0±12.4 l and 4.2±2.6 l/h, respectively. On the other hand, in the postprandial study, the calculated Tmax, Cmax, T1/2, lag time, Vd/F and Cl/F were 5.6±2.0 h, 7.6±1.8 μg/ml, 16.7±7.0 h, 2.35±0.78 h, 48.0±15.9 l and 2.1±0.6 l/h, respectively. The values for the area under the curve for data collected over a 24-h period (AUC24 h) in the fasting and postprandial studies were 86.0±27.7 and 102.1±31.0 μg·h/ml, respectively. The T1/2 had a tendency to be prolonged after food intake, but there were 2 cases with shortened T1/2. Food intake prolonged the lag time 3-fold and as a result, the postprandial Tmax was 2 times longer.
Aberrant wound healing, either causing scarring or chronic wounds, is a significant cause of morbidity. There is therefore, considerable interest in agents which can modulate certain aspects of the wound healing process. Fucoidans, sulphated polyfucose polysaccharides which may be extracted from Fucus spp., have been shown to modulate the effects of a variety of growth factors through mechanisms thought to be similar to the action of heparin. We investigated the interaction between two commercial preparations of fucoidan and transforming growth factor (TGF)-β1. These preparations of fucoidan, as well as heparin, inhibited fibroblast proliferation at concentrations from 0.01 to 100 mg/ml. The anti-proliferative effects of 1 ng/ml TGF-β1 on dermal fibroblasts were abrogated by fucoidan preparation F7 when used at concentrations over 1 mg/ml. In a three dimensional in vitro model of wound repair, the fibroblast populated collagen lattice or “dermal equivalent”, TGF-β1 reduced the rate of fibroblast repopulation of a wound defect created by punch biopsy. Addition of fucoidan to the model in the presence of TGF-β1 increased the rate of fibroblast repopulation of the wound and at 10 mg/ml of fucoidan the number of cells which had migrated into the wounded defect was similar to that of control cultures. These data suggest that fucoidan has properties which may be beneficial in the treatment of wound healing.
Sweet potato acid phosphatase was covalently coupled with glutaraldehyde to aminopropyl controlled-pore glass, and used as a pre-column enzyme reactor. The immobilized enzyme reactor (IMER) was continuously operated using an automated chromatographic detection system we developed. Functional evaluation of the IMER was carried out by injecting ten samples on the same day at an injection amount of 1.25 nmol (62.5 nmol per ml) using riboflavin sodium phosphate (FMNs) as a substrate, and by prolonged use for ten months. The IMER exhibited decreased activity after repeated use for a total of 3000 samples, but about 75% of its original activity remained. The conversion rate of FMNs to riboflavin by IMER was increased from 89 to 97% by adding citrate, ethylenediaminetetraacetic acid disodium salt, etc., but especially by adding citrate. The increased conversion of FMNs to riboflavin due to the addition of citrate was probably not due to the chelation of heavy metal ions by citrate. We also investigated complex formation of acid phosphatase with the substrate FMNs using surface plasmon resonance to determine the effect of citrate on the processes of association and/or dissociation between the enzyme and substrate. Enzyme fatigue was also observed during the course of prolonged and repeated use.
For purpose of reducing renal accumulation of radioactivity of a known radiopharmaceutical agent, i.e., 111In-DTPA (diethylenetriaminepentaacetic acid)-D-Phe1-octreotide, a derivative in which p-carboxy-L-phenylalanine is substituted for D-Phe1 was synthesized. Biodistribution study of the resultant compound having carboxy-substituted L-Phe1 revealed that the renal accumulation was significantly lower than that of control compound having unsubstituted L-Phe1, demonstrating that the presence of negative charge on the N-terminal amino acid of octreotide is effective to reduce the renal accumulation. This effect can be attributed to the reduction of lipophilicity and also the repulsive force arisen from the negative charge of renal brush border membrane.