Nardostachin, which is an iridoid isolated from Patrinia saniculaefolia, was examined by assessing its effect on the production of tumor necrosis factor-α (TNF-α) and expression of 2 enzymes, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), in lipopolysaccharide (LPS)-stimulated Raw264.7 macrophages. This compound consistently inhibited the production of nitric oxide (NO) and TNF-α production in a dose-dependent manner, with respective IC50 values of 12.3 and 16.2 μM. The decrease in quantity of NO products was accompanied by a decrease in the iNOS protein level, as assessed by Western blotting probed with specific anti-iNOS antibodies. In addition, this compound also reduced the COX-2 protein expression level and the attendant PGE2 production in LPS-stimulated macrophages. These results suggest that nardostachin may be useful for inhibiting the production of inflammatory mediators such as TNF-α, NO and PGE2 in inflammatory diseases.
Screening using a reporter under the control of the hypoxia-response element (HRE) identified several flavonoids and homoisoflavonoids that inhibit the activation of HRE under hypoxic conditions. Among various compounds, isorhamnetin, luteolin, quercetin, and methyl ophiopogonanone B (MOB) were effective at 3 to 9 μg/ml in inhibiting the reporter activity. The expression of vascular endothelial growth factor (VEGF) mRNA during hypoxia was also inhibited by MOB in HepG2 cells, but the effective doses were 10 to 20 μg/ml. MOB caused destabilization of hypoxia-inducible factor (HIF)-1α, as revealed by Western blotting, that was dependent on proteasome activity and the tumor suppressor, p53. The tubular formation and migration of human umbilical vein endothelial cells was also inhibited by MOB. MOB is expected to act as an inhibitor of angiogenesis.
It is known that cholesterol biosynthesis in the liver is inhibited by probucol. This inhibition by probucol is caused at least in part by a decrease in 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase activity. In this study, we examined serum cholesterol and the change in the activity or protein level of mevalonate pyrophosphate decarboxylase (MPD), which is involved in cholesterol biosynthesis, in the livers of rats fed probucol. The results indicated that serum cholesterol, MPD activity and MPD protein were decreased by 70, 50 and 60% by probucol, respectively, as compared with those in rats fed normal chow. These data show for the first time that probucol decreases the level of an enzyme involved in cholesterol biosynthesis other than HMG-CoA reductase.
Tropolone (1) showed strong insecticidal activity on Tyrophagus putrescentiae and Dermatophagoides farinae. The insecticidal effect of 1 on both insects was stronger than that of hinokitiol (2, 4-isopropyltropolone: major component of Thujopsis dolabrata SIEB. et ZUCC. hondai MAKINO). The insecticidal activity of both compounds was higher than that of N,N-diethyl-m-toluamide (DEET), used as a positive control. Compound 1 had potent insecticidal activity against Coptotermes formosanus, although its activity was much lower than that of commercial chloropyrifos. Like 2, 1 showed the inhibitory activity toward metalloproteases such as carboxypeptidase A, collagenase and thermolysin and their inhibitory activities were much higher than that of 1,10-phenanthroline, used as a positive control. The inhibitory activity of 1 on carboxypeptidase A was especially high, its 50% inhibitory concentrations (IC50) being 2.73×10−6 M. This inhibitory activity was as high as that of 2 (IC50: 2.76×10−6 M). Compound 1 inhibited the growth of seven kinds of plant-pathogenic fungi and their minimum inhibitory concentration (MIC) values were in the range of 6.0—50.0 μg/ml. In particular, 1 showed strong antifungal activity on Pythium aphanidermatum IFO-32440 (MIC: 6.0 μg/ml).
We have recently demonstrated that Bak Foong Pills (BFP), a well-known Chinese medicine widely used for treating gynecological disorders, stimulates human colonic epithelial anion secretion, which was mediated by intracellular cAMP and Ca2+. The present study further investigated the effect of BFP on exocrine pancreatic-bile secretion using in vivo and in vitro approaches. Duodenal infusion of BFP ethanol extract (1 g/kg) in rats produced increases in the volume and protein output of pancreatic-bile juice, but did not affect its pH. Surgical ablation of vagal neural pathway slightly reduced the effect of BFP on the protein output and volume, indicating that the vagal nerve pathway was not the major player in medicating the effect of BFP on exocrine pancreatic-bile secretion. Using CAPAN-1 cell line, a human pancreatic duct cell line, in conjunction with the short-circuit current (ISC) measurements, we further demonstrated that BFP could directly stimulate pancreatic HCO3− secretion. Basolateral addition of BFP (600 μg/ml) produced averaged charges transported of 2100±382.5 μC/cm2, which was blocked by apical addition of Cl− channel blocker. Removal of HCO3− from the Krebs–Henseleit (K–H) solution inhibited the BFP-induced ISC by more than 95%. The present results suggest that BFP could improve digestive function by stimulating pancreatic protein and HCO3− secretion.
We studied the effect on both platelet aggregation and blood coagulation, known to be major risk factors in thrombogenesis, of proteins from hen egg yolk (EP). EP potently inhibited collagen-induced human platelet aggregation in a dose-dependent manner. Furthermore, EP has a synergistic effect on the inhibition of human platelet aggregation with both molsidomine, an inhibitor of cGMP-specific phosphodiesterase, and theophylline, an inhibitor of cAMP-specific phosphodiesterase. These results indicate that the active mode of EP might be involved in elevation of the levels of both cGMP and cAMP. Prothrombin time and activated partial thromboplastin time were potently prolonged by EP. These data suggest that EP prolongs the time interval between the conversion of fibrinogen to fibrin. Accordingly, these findings demonstrate that EP might have antithrombotic effects by inhibiting platelet aggregation and fibrin formation.
In normal mice, plasma histamine levels were 29.4±10.1 pmol/ml. When 0.1 μg of lipopolysaccharide (LPS) was intravenously injected into Propionibacterium acnes (P. acnes)-primed ICR mice, histamine levels increased remarkably to 61.2±15.9 pmol/ml (p<0.001). An increase was also observed in liver tissues. Oral administration of histidine at 200 mg/kg once daily for 5 d before intravenous LPS injection increased the plasma alanine aminotransferase (ALT) activity to 2936.5±356.3 IU/l, a significant change compared with the controls (2244.8±425.5 IU/l, p<0.05). The 24 h survival rate after LPS injection was 72.7% in the mice treated with 50 mg/kg of ranitidine, in contrast with 50% in the control group although the treatment did not significantly decrease the plasma ALT activity. On the other hand, 50 mg/kg of pyrilamine significantly reduced plasma ALT activity (p<0.001). The results suggested that histamine levels are related to hepatic damage in the P. acnes plus LPS induction of liver injury.
We previously reported that a serotonin precursor 5-hydroxytryptophan (5-HTP) increases serum leptin levels in mice. In this study, we studied the effects of insulin and adrenalectomy on hyperleptinemia induced by 5-HTP. Co-administration of insulin significantly increased hyperleptinemia elicited by 5-HTP. 5-HTP itself increased serum insulin levels. Adrenalectomy, which depletes corticosterone, did not abolish hyperleptinemic effects of 5-HTP. These results suggest that insulin may participate in hyperleptinemic effects of 5-HTP and that the involvement of corticosterone in effects of 5-HTP may be probably small.
This study evaluates whether quercetin (25, 50 and 75 mg/kg body weight) treatment has a protective effect on the pro-oxidant-antioxidant state following chronic ethanol treatment in mice. Pretreatment (quercetin 25, 50 and 75 mg/kg body weight for 15 d+co-treatment of ethanol 18%+quercetin for 15 d and ethanol 18% for the 15 d) increased the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR), and glutathione (GSH) in comparison to the ethanol group. No significant differences from the ethanol group were observed in the group after post-treatment (ethanol 18% for 30 d+quercetin 25, 50 and 75 mg/kg body weight for 15 d) with quercetin. A significant increase in lipid peroxidation (malondialdehyde, MDA) products was observed in liver tissue after administration of ethanol, which was attenuated by pre- and post-treatment with a high dose of quercetin. GSH levels increased and oxidized glutathione (GSSG) levels decreased in groups of ethanol-exposed mice that received quercetin for 15 d prior to ethanol exposure. In conclusion, pre-treatment of quercetin may protect against ethanol-induced oxidative stress by directly quenching lipid peroxides and indirectly by enhancing the production of the endogenous antioxidant GSH. There was no protective effect on post-treatment with quercetin.
Tramadol is an important spinal drug which produces analgesia following intrathecal injection. It is well known that fatty acids (FAs) play an important role in membrane fluidity of the blood-brain barrier (BBB) tissue, which blocks and/or controls the transportation of toxic substances into the brain. The aim of this study was to investigate the effect of a spinal drug (tramadol) on the concentrations and compositions of fatty acid in BBB tissues of New Zealand male rabbits. The total cellular fatty acid profiles of the tissues in three spinal cord sections (cervical, thoracal and lumbar) and in the brain of rabbits with or without drug administration were determined by gas chromatography using Sherlock Microbial Identification System (MIS) software (Microbial ID, Newark, DE, U.S.A.) with a database of FAME profiles for eukary. The relative percentage of the fatty acid methyl ester (FAME), 24 : 1 ω9c nervonic and 17 : 1 ω8c, did not change with tramadol treatments. However, there was an increase in the concentration of the FA 16 : 0, 18 : 1 ω7c DMA, 18 : 1 ω9c, sum in future 4, sum in future 8, sum in future 9, 18 : 0, 20 : 4 ω6c, sum in future 14, 22 : 4 ω6c, in contrast to a decrease in the percentages of the following FAMEs; 20 : 0, 20 : 1 ω9c. In the brain, there was an increase in the concentration of the FA 18 : 1 ω9c, sum in future 8 and 18 : 0, in contrast to a decrease in the percentages of two FAMEs, 16 : 0, 20 : 4 ω6c and 22 : 6 ω3c. The number of fatty acids were 20 in the spinal cord sections and 8 in the brain tissues of control animals compared to 15—18 fatty acids in the spinal cord section and 7 in the brain tissues of drug administered animals. The overall changes in the concentrations and numbers of FAs suggest that the spinal drug tested in this study has a side effect of disrupting of membrane fluidity of the BBB, which may cause neurotoxicity.
The synthesis and pharmacological evaluation of novel 1-substituted-1,2-dihydro-pyridazine-3,6-diones (4a—l, 5a—j) as potential anticonvulsant agents are described. The compounds were tested in vivo for the anticonvulsant activity. The compound which have maximum protection against MES induced seizures is 1-[3-(2-aminophenylamino)-2-hydroxypropyl)-1,2-dihydro-pyridazine-3,6-dione 4h (ED50=44.7 mg · kg−1 i.p.) 1-[2-hydroxy-3-piperazin1-yl-propyl)-1,2-dihydro-pyridazine-3,6-dione 4c (ED50=72 mg · kg−1 i.p.) and 1-[2-hydroxy-3-imidazol-1-yl-propyl)-1,2-dihydro-pyridazine-3,6-dione 4d (ED50=79 mg · kg−1 i.p.) were also found to have maximum protection against MES induced seizures. Whereas all these compounds failed to protect the animals from subcutaneous pentylenetetrazole (Metrozol) seizure threshold test (sc-Met).
4-(N-Hydroxyphenyl)retinamide (also known as 4-HPR or fenretinide), a synthetic amide of all-trans retinoic acid (RA), has been implicated as a promising anticancer agent associated with reducing the toxicity related to RA. However, the low plasma levels of 4-HPR in patients limited clinical trials, leading to a search for derivatives with better efficacy. In this study, we synthesized a series of 4-HPR derivatives in good yields by introducing acetate (compound 1), propionate (2), pyruvate (3), butyrate (4), or stearate (5) to the 4-hydroxylphenyl moiety of 4-HPR. In our initial proliferation assays, we identified compound 3 as the most cytotoxic of the series against four ovarian cancer cell lines (OVCAR-3, PA-1, 2774, and SKOV-3). Dose–response curves yielded IC50 values of 3.75—7.75 μM for AtRA, 2.80—5.50 μM for 9-cis RA, 0.65—4.05 μM for 4-HPR, and 0.25—0.75 μM for compound 3, depending on the cell type treated. Nuclear staining with 4′,6-diamidino-2-phenylindole (DAPI) and DNA fragmentation assays clearly indicated that the antiproliferative effect of compound 3 was mediated by apoptosis. In contrast to natural retinoids, both 4-HPR and compound 3 activated two (RARβ and RARγ) of the three retinoic acid receptor (RAR) subtypes tested, but did not activate any of the three retinoid X receptors (RXRs), as determined by transcription assays in OVCAR-3 cells. However, like natural retinoids, 4-HPR and compound 3 actively suppressed c-Jun transcriptional activity. Thus, compound 3 not only showed more potent antiproliferative activity than any other retinoid derivatives tested, but also effectively inhibited the c-Jun activity that has been implicated in tumor promotion and invasion. These results, together with compound 3's selectivity for RAR subtypes, suggest that compound 3 could be an effective anticancer drug for ovarian cancer, with less toxicity than RA.
This study reports that acidic polysaccharide (PL) isolated from Phellinus linteus alleviated the septic shock induced by high dose lipopolysaccharide (LPS) injection in mice. To examine the origin of this effect, we investigated cytokine production in serum and the expression of MHC II in B cells and macrophages in areas of inflammation. Pretreatment with PL 24 h before LPS administration resulted in a significant inhibition of up to 68% of circulating tumor necrosis factor (TNF)-α, a moderate reduction of 45% of interleukine (IL)-12 and 23% of IL-1β, but no significant reduction in IL-6. In addition, the expression of MHC II in B cells and macrophages was examined. Our results show that LPS-stimulated cytokine release and the level of MHC II can be modulated by in vivo administration of soluble PL in mice. The decrease of IL-1β, IL-12 and TNF-α in sera and the down-modulation of MHC II during septic shock may contribute to the long survival of mice by PL. Administration of PL in vivo decreases IL-2, IFN-γ and TNF-α production in splencotyes and enhances spontaneous cell apoptosis in macrophages and lymphocytes stimulated with LPS in vitro. Thus, part of the anti-inflammatory effects of PL treatment in vivo may result from the enhanced apoptosis of a portion of the activated macrophages and lymphocytes. The ability of PL to significantly reduce the TNF-α production indicates the potential of the polysaccharides in possible therapeutic strategies that are based on down-regulation of TNF-α.
The acute phase response involves changes in serum concentrations of a number of liver-synthesized proteins. Among these are C-reactive protein (CRP), ferritin (FER), transferrin (Trf) and ceruloplasmin (Cp). Determination of serum CRP, FER, Trf, and Cp was performed in 52 patients with inoperable head and neck cancer (n=11), inoperable esophageal cancer (n=10), rectal cancer (n=9; operation was performed=5, inoperable=4), and lung cancer (n=22), all of whom were treated with radical radiotherapy (RT). Post-radiotherapy CRP levels were significantly higher compared to the preradiotherapy levels (p<0.001). We found decreased serum Trf levels during the irradiation period, while acute-phase proteins such as CRP, FER, and Cp levels increased during the RT period. Further studies on the roles of other acute phase reactants and the above mentioned parameters in a large-patients-with cancer group during radiotherapy are required to understand the role of markers, which are altered during radiotherapy.
A marine sponge Petrosia similis afforded two compounds which belongs to bis-quinolizidine alkaloids namely, petrosin (1) and petrosin-A (2), respectively. Petrosin (1) and petrosin-A (2) showed anti-HIV inhibition with IC50 values of 41.3 and 52.9 μm respectively. MAGI cell assays indicated that the compounds inhibited early steps of HIV replication. In extracellular HIV-1 Reverse Transcriptase inhibition assay petrosin and petrosin-A inhibited HIV-1 RT at 10.6 and 14.8 μm. This is the first report of petroisns with anti-HIV activity.
Desmodium gangeticum is herbal species which is widely used in the indigenous system of medicine and is reported to contain flavone and isoflavanoid glycosides. In view of its wide use and it's chemical composition, this study was aimed at examining the antioxidant activity of the extract of D. gangeticum. The extract was studied for diphenyl picryl hydrazyl (DPPH), nitric oxide, ferryl-bipyridyl and hypochlorous acid scavenging activity along with lipid peroxidation. Nitric oxide was generated using sodium nitroprusside and was studied using Griess reagent. In order to study the iron chelating capacity of the extract, the percentage ferryl-bipyridyl inhibition was studied. Hypochlorous acid scavenging activity was tested by measuring the inhibition of 5-thio-2-nitrobenzoic acid oxidation. The extract was also studied for lipid peroxidation assay by thiobarbituric acid-reactive substances (TBARS) method using rat brain homogenate. The results indicate that D. gangeticum extract has potent antioxidant activity.
Neuronal apoptosis may contribute to pathologic neuronal loss in certain disease states such as neurodegenerative diseases. Staurosporine (ST), a nonselective protein kinase inhibitor, has been shown to induce apoptosis in a variety of cells including nerve cell lines. In this study, we investigated the neuroprotective effect of sauchinone, which is a unique lignan from Saururus chinensis, on ST-induced apoptosis in C6 rat glioma cells. Sauchinone attenuated ST-induced apoptosis of C6 glioma cells as evidenced by DNA fragmentation. We also provide evidence that the inhibitory effect of sauchinone on ST-induced apoptosis involves a dose-dependent upregulation of an antiapoptotic protein, Bcl-2. Mounting evidence shows that the activation of caspases, especially caspase-3, triggers the apoptotic process. The activity of caspase-3 of ST-pretreated cells was significantly decreased upon sauchinone treatment in a dose-dependent manner. Taken together, the data demonstrate that sauchinone protects C6 glioma cells from ST-induced apoptosis in a caspase-3 dependent manner. Our findings may be critical for developing a strategy to protect nerve cells from apoptosis, suggesting the potential development of sauchinone as a neuroprotective agent.
Sixteen cardenolides, two hemiterpenoids, two phenylpropanoids and a phenylethanoid isolated from the roots of Streptocaulon juventas (LOUR.) MERR. were examined for their antiproliferative activity toward three human-derived (HT-1080 fibrosarcoma, lung A549 adenocarcinoma, cervix HeLa adenocarcinoma) and three murine-derived (colon 26-L5 carcinoma, Lewis lung carcinoma, B16-BL6 melanoma) cell lines. The cardenolides selectively and strongly inhibited proliferation of the HT-1080 (IC50 values, 0.054—1.6 μM) and A549 (IC50, 0.016—0.65 μM) cell lines. The characteristic morphological changes and ladder-like DNA fragmentation in those cells treated with the cardenolides indicated the antiproliferative activity was due to the induction of apoptosis.
As an attempt to search for bioactive natural constituents exerting antinociceptive and antiinflammatory activities, we examined the potency of the extract of Rubus coreanus fruits by the activity-guided fractionation. The EtOAc- and BuOH fraction and those alkaline hydrolysates showed significant antinociceptive effects as assessed by writhing-, hot plate- and tail flicks tests in mice and rats as well as antiinflammatory effect in rats with carrageenan-induced edema. BuOH extract was subjected to column chromatography to obtain a large amount of niga-ichigoside F1 (1,23-hydroxytormentic acid 28-O-glc), which was again hydrolyzed in NaOH solution to yield an aglycone 23-hydroxytormentic acid (1a). The aglycone, 23-hydroxytormentic acid, was much more potent in both antinociceptive and antiinflammatory tests than the glycoside, niga-ichigoside F1. The antiinflammatory effects of these compounds were further supported by the reduction of carrageenan-induced lipid peroxidation and hydroxyl radical in serum. These results suggested that 23-hydroxytormentic acid might be an active moiety of niga-ichigoside F1 present in R. coreanus.
Aralia elata Seemann is an edible mountain vegetable in Korea containing saponin, alkaloid, palmitic acid, linoleic acid, methyl eicosanoate and hexacosol, and is known to manifest an effect on cardiac infarction, gastric ulcer, colitis, and enervation. This study has examined the effects of Aralia elata Seemann ethanol extract on antioxidant enzyme systems and lipid metabolism in rats along with benzo(α)pyrene (B(α)P) administration. Rats were divided into four groups: control (C), an extract fed group (CE), a B(α)P fed group (CB), and a B(α)P and extract fed group (CBE). The ethanol extracts of Aralia elata Seemann (50 mg/kg body weight) were fed to the rats for 4 weeks by stomach tubing. Extract administration increased the antioxidant activities of glutathione sulfur transferase (GST). Total superoxide dismutase (SOD) and Cu,Zn-SOD activities were stimulated. Catalase activities were increased by 50% with extract feeding. Cu,Zn-SOD was greatly enhanced from 0.10 unit to 0.18 unit and catalase activity also was increased. Serum α-tocopherol was markedly increased by the extracts. The ethanol fraction of Aralia elata Seemann decreased total serum cholesterol. However, serum HDL-cholesterol was increased by 35% (p<0.05). The results indicate that Aralia elata Seemann exerts antioxidant and strong hypocholesterolemic and hypolipidemic effects in vivo with the administration of B(α)P.
A 35% EtOH extract of flowers of Impatiens textori MIQ. showed an inhibitory effect on blood pressure decrease in response to platelet activating factor (PAF) measured with a blood pressure monitoring system. Bioassay-guided fractionation of the 35% EtOH extract (IT) led to isolation of the flavones apigenin (1) and luteolin (3), which significantly inhibited blood pressure decrease in response to PAF. Their compounds and apigenin 7-glucoside (2), chrysoeriol (4), quercetin (5), quercetin 3-glucoside (6), kaempferol (7), kaempferol 3-glucoside (8) and kaempferol 3-rhamnosyldiglucoside (9) were also isolated from the flowers of I. textori for the first time. This study revealed that the flowers of I. textori might be a possible anti-allergy agent.
To determine the usefulness of the monkey as an animal model, which can predict in vivo performance of humans, the major gastrointestinal physiological parameters of this animal were evaluated. The pH of gastric juice collected by a fiberscope from the stomach in fasted cynomolgus monkeys showed a high acidity level, which ranged from 1.2 to 4.3. The gastric emptying time of oral dosage forms (solution, granules and tablets) showed that the larger size dosage forms seemed to be emptied more slowly, and three dosage forms were prolonged by feeding. The gastrointestinal agitation intensity of monkeys was estimated using controlled-release tablets of acetoaminophen, which showed a slow erosion rate. The in vivo release amount-time profiles of the tablet in fasted monkeys corresponded to their in vitro profiles with paddle agitation conditions of between 10 rpm and 50 rpm of the paddle method; this result was smaller than in dogs (100 rpm) but equivalent to that in humans (10 rpm). Further, the small intestinal transit time (SITT), estimated using a double marker method, ranged from 2.2 to 4.2 h in the fasting state and from 2.2 to 3.2 h in the fed state; the SITT was not significantly delayed by feeding. Comparison with the published data about dogs and humans showed these gastrointestinal physiological parameters of monkeys to be more similar to those of humans. Consequently, it is assumed that the monkey is useful as an animal model for bioavailability studies of oral dosage forms.
We examined the 4′-hydroxylation of flurbiprofen in rat hepatocytes and liver microsomes in order to know whether the metabolism of flurbiprofen is changed on its administration to experimental animals after overnight fasting, because starvation and fasting change both the composition of cytochrome P450s (CYPs) and metabolic activity. CYPs involved in the hydroxylation were determined by various CYP inhibitors and inhibitory antibodies against rat CYP2C11 and CYP2E1 using the microsomes in fasting and feeding. The results provided a possibiliy that the 4′-hydroxylation might be regulated by CYP2C11, but not by CYP2E1, at fasting rather than feeding.
Hepatic dysfunction due to flutamide administration has been reported and this side effect often limits the use of the agent. The prediction of flutamide-induced hepatotoxicity is attributed to the proper use of the antiandrogen. In this study, we investigated whether hepatic dysfunction could be assessed by the metabolite profile in serum from patients receiving this drug. Serum samples were obtained from 15 patients with prostate cancer, 12 patients with no sign of hepatotoxicity and 3 patients with slight hepatic dysfunction during long-term flutamide treatment. We analyzed the metabolite profiles by LC/MS in selected ion monitoring mode and detected a new metabolite (M3) that was an oxidation product of flutamide. However, there were no consistent differences in the serum flutamide metabolites between patients with normal function and those suffering hepatic dysfunction. The metabolite profiles in the β-glucuronidase-treated serum showed a similar pattern between normal functioning and dysfunctional groups. Thus, the profile of flutamide metabolites determined in serum may not contribute to the risk prediction of flutamide-related hepatotoxicity.
The intradermal delivery of an antisense oligonucleotide was examined by iontophoresis. In this experiment, the antisense sequence of [32P]-labeled phosphodiester oligonucleotide ([32P]D-oligo, 18-mer) hybridizing to mouse interleukin 10 (IL-10) mRNA was used as a model D-oligo. In in vitro iontophoretic experiments, isolated hairless mouse skin was used with a horizontal diffusion cell. The enhancing effect of pulse depolarization (PDP) iontophoresis on the [32P]D-oligo permeation through the skin was better, and the skin irritation was less, than those of constant direct current (CDC) iontophoresis. The apparent fluxes of [32P]D-oligo were enhanced with the increasing current densities and [32P]D-oligo concentrations in the donor solution, whereas the enhanced flux decreased with the increasing NaCl concentrations in the donor solution. An optimum electric current was observed for the intradermal delivery of [32P]D-oligo, and intact [32P]D-oligo was detected within the skin after iontophoresis for 6 h. These results suggest that PDP iontophoresis may be useful for the intradermal delivery of antisense oligonucleotides.
We previously reported the substantial synergic effects of electroporation and electrolytes, particularly those containing CaCl2 on the skin permeation of the model low-molecular weight compound, calcein. We then investigated the effects of electroporation (300 V, 10 ms×10 times) and 150 mM NaCl or CaCl2 on skin permeation of higher molecular weight compounds, fluorescein isothiocyanate (FITC)-dextrans (FD-4, FD-10 and FD-40; average molecular weight, 4.4, 9.6 and 35.6 kDa, respectively) using excised hairless rat skin. The observed steady state flux of FD-4 was 1.3 pmol/cm2/h after electroporation without NaCl or CaCl2. The flux did not differ greatly from that without electroporation. In contrast, a much higher steady state flux was observed after electroporation with NaCl or CaCl2 (2.5 and 8.2 pmol/cm2/h, respectively). For FD-10 and FD-40, no flux was detected with electroporation in water (without electrolytes) or without electroporation. On the other hand, high skin permeation was observed after electroporation in NaCl or CaCl2 solution (FD-10: 7.5 and 18.2 pmol/cm2/h, FD-40: 4.5 and 9.3 pmol/cm2/h in NaCl and CaCl2, respectively). The effects of CaCl2 on FD permeation were greater than those of NaCl. The present finding suggests that electroporation application in the presence of electrolytes, particularly CaCl2, was very effective in increasing transdermal delivery of water-soluble macromolecules.
An efficient and rapid protocol for in vitro induction and complete plant regeneration of Polygonum multiflorum THUNB has been developed. Nodal explants were grown in vitro on Murashige and Skoog's (MS) basal medium containing different concentrations of α-naphthaleneacetic acid (NAA) and benzyladenine (BA). The nodal explants (97%) produced multiple shoots (4.7 shoots per explant) on MS basal medium supplemented with 0.2 mg/l NAA and 2.0 mg/l BA after 6 weeks of culture. Eighty-eight percent to 100% of the shoots (1.0 cm in length) elongated (about 3.02—4.28 cm) and rooted on MS basal medium supplemented with NAA or indole-3-butyric acid (IBA). All the rooted shoots were transferred to pots containing autoclaved soil, vermiculite, and peat moss (1 : 1 : 1). The plantlets were successfully acclimatized under greenhouse conditions with high humidity before transferring to the field. The anthraquinone contents were determined using HPLC. Analysis revealed that the contents of the major medicinal compounds–emodin and physcion in the 6 weeks old in vitro grown shoots and three month old in vitro propagated plants grown in greenhouse were higher than those of the marketed crude drug (processed underground or stem parts of P. multiflorum).
Rheumatoid arthritis (RA) is characterized by chronic inflammation of the synovial membrane in the joint, which leads to the progressive destruction of articular cartilage, ligament and bone. Several cytokines such as tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), and interleukin 6 (IL-6) have been implicated in the pathological mechanisms of synovial tissue proliferation, joint destruction and programmed cell death in rheumatoid joint. In the Korean traditional medicine, Hominis placenta (HP) as an herbal component of herb-acupuncture has been widely used to treat chronic inflammatory diseases such as RA. To study the therapeutic effects of HP injection into the ST36 acupoint (HP herb-acupuncture) on the inflammatory responses of a subchondral region of rheumatoid joint, the polyarthritis-induced Sprague–Dawley (SD) rat was developed as a rheumatoid arthritis model by the intradermal injection of dried cells of Mycobacterium tuberculosis emulsified in squalene to the base of tail. After the onset stage (11 d after adjuvant injection) of polyarthritis, a fixed volume of HP extract was daily injected to Zusanli (ST36) acupoint on the rat's leg for 2 weeks. The body weight, paw volume of the knee joint and articular index were exploited as an assessment method addressing arthritic symptoms, and the expression profiles of TNF-α, IL-1β and IL-6 at the subchondral bone of the joint were analyzed using an immunohistochemistry. After the treatment of arthritic rats with HP, the body weights and paw volumes of arthritic rats were almost restored to the levels of normal rats whereas the evaluation by the articular index was not remarkable. The TNF-α, IL-1β and IL-6 positive cells in the immunohistological sections of subchondral bone region of the joint significantly decreased in HP-treated (ST36 acupoint) arthritic group as compared with those in non-treated or HP-treated (non-acupoint) ones, which was coincident with the behavioral studies. In conclusion, the HP herb-acupuncture was found to be effective to alleviate the arthritic symptoms in adjuvant-induced arthritis rats as regards the body weight, joint appearance and the expression profiles of inflammatory cytokines.
Flavone and its derivatives had very weak antibacterial effects on methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive S. aureus, but dramatically intensified MRSA's susceptibility to β-lactams. We named these compounds “ILSMR (intensifier of β-lactam-susceptibility in MRSA).” We also found discrepancies among MRSA strains in their responses to flavone; some strains showed phenotypic susceptibility to methicillin while others showed phenotypic resistance to it. To understand the mechanism underlying this discrepancy, we characterized 20 MRSA strains in detail, analyzed their conventional and molecular typings, and examined each strain's resistance to β-lactams, with COL serving as a reference. Neither SCCmec typing nor coagulase typing explained the diverse effects of flavone on the β-lactam MICs of these strains. Likewise, changes in pulsed-field gel electrophoresis (PFGE) type were not associated with the profiles of ILSMR effects. However, the present observations suggest that the ILSMR effects on MRSA is strain-specific, and that this effect depends on an as-yet unknown mechanism that is essential for the expression of the phenotype conferring β-lactam resistance to MRSA strains, independently of an interaction with the mecA-encoded penicillin-binding protein 2a or with the β-lactamase.