Effect of hr-IL2 treatment on intestinal P-glycoprotein expression and activity in Caco-2 cells

Caco-2 cells were used to investigate the effect of human recombinant interleukin-2 (IL2) on intestinal P-glycoprotein (P-gp) transporter activity in-vitro. More specifically the efflux function of P-gp was studied by measuring the transepithelial transport of rhodamine-123, a fluorescent substrate of P-gp. Its transport was completely inhibited by two specific P-gp inhibitors, ciclosporin A and GG918, in our experiments. Conversely, these two specific P-gp inhibitors inhibited only 50% of transepithelial transport when [3H]vincristine was used as substrate. After Caco-2 cells were treated with 100 IU mL-1 (6.1 ng mL-1) IL2 for 24 h, a significant diminution (21%) of P-gp transporter function was observed with rhodamine-123 substrate. This effect was also confirmed after 48 and 72 h of exposure to IL2. However, for higher concentrations of IL2 (1000 and 5000 IU mL-1), diminution of P-gp function only occurred after a longer treatment period (48 h and more). The inhibitory effect of IL2 on P-gp activity was found to be independent of tight junction function as demonstrated by constant transepithelial electrical resistance (TEER) measurements for all experimental conditions encountered in this study (time and concentration of IL2 exposure). Furthermore, the MDR1 mRNA level was found to be strongly repressed in Caco-2 cells exposed with 1000 IU mL-1 IL2 for 72 h while the amount of MRP1 mRNA remained unchanged. In conclusion, acute incubation of Caco-2 cells with IL2 induced a decrease of P-gp transporter expression and activity.     

姜黄素对Caco-2细胞上P-糖蛋白活力及MDR1 mRNA表达的影响

目的研究姜黄素对Caco-2细胞上P-糖蛋白(P-gp)功能和表达的影响,并从分子水平研究其对MDR1mRNA表达的影响。方法采用流式细胞仪测定P-gp底物——罗丹明-123在细胞中的浓度,并分析Caco-2细胞上P-糖蛋白的表达量;采用RT-PCR法分析Caco-2细胞上MDR1基因mRNA的表达量。结果低、中、高浓度(0.1、0.5、1.0μmol.L-1)的姜黄素增加了罗丹明-123的外排,其外排量分别增加了6.2%、22.2%和33.8%(P<0.05);0.1～1.0μmol.L-1姜黄素与细胞孵育72h后,细胞膜上的荧光强度随浓度增加而增强(P<0.05),P-gp表达量是空白对照组的4倍,MDR1基因mRNA表达水平上调了1.58倍。结论姜黄素并不是P-gp的抑制剂,它能在较低浓度以及较短时间内(1h)增强Caco-2细胞上P-gp的功能活性,从而减少罗丹明-123在细胞内的蓄积。长时间(72h)应用姜黄素可以诱导P-gp和MDR1mRNA的表达。姜黄素将有可能引起相应的药物-药物/食物的相互作用。     

Caco-2细胞模型研究白芷提取物对P-糖蛋白功能的影响

目的:研究白芷提取物对黄酮类成分葛根素、黄芩苷肠道转运的影响,结合白芷提取物对Caco-2细胞上P-gp功能的影响,探讨白芷提取物对葛根素、黄芩苷肠吸收的影响机制。方法:采用Caco-2细胞模型,研究白芷提取物分别对黄酮类成分葛根素、黄芩苷吸收的影响,以P-gp底物罗丹明123(R-123)为荧光探针,评价不同浓度白芷提取物和P-gp抑制剂维拉帕米对R-123细胞蓄积的影响,细胞内的R-123浓度采用荧光分光光度法检测。结果:白芷提取物对葛根中葛根素、黄芩中黄芩苷的吸收均有明显的促进作用,葛根素的表观渗透系数Papp分别提高1.6～3.6倍,黄芩苷的表观渗透系数Papp分别提高2.3～3.3倍;白芷香豆素(0.15～1.5 mg.mL-1)、白芷挥发油(0.24～6.0μL.mL-1)、白芷香豆素+白芷挥发油均能显著增加细胞内Rh-123的摄取,对P-gp显示出明显剂量依赖性的抑制作用。结论:白芷提取物促进葛根素、黄芩苷等黄酮类成分吸收的原因可能与其抑制P-gp外排有关。     

Inhibitory effect of a bitter melon extract on the P-glycoprotein activity in intestinal Caco-2 cells

Extracts of bitter melon, soybean, dokudami and welsh onion by 40% methanol increased the accumulation of rhodamine-123 by Caco-2 cells, suggesting that these extracts inhibited P-glycoprotein (P-gp). The extract of bitter melon was separated in a tC18 cartridge column and the eluate from 80% acetonitrile most markedly increased the [(3)H]-daunomycin accumulation by Caco-2 cells. The inhibitory compounds in the bitter melon fraction were isolated by HPLC with Pegasil C4 and Pegasil ODS columns. The HPLC fraction having the highest activity was analyzed by (1)H-NMR and FAB-MS, and the active compound was identified as 1-monopalmitin. The inhibitory activities of 1-monopalmitin and its related compounds suggested that the inhibition of P-gp activity was not dependent on the degree of unsaturation of fatty acid in the monoglyceride, but on the chain length. It was also suggested that the monoglyceride structure played an important role in the inhibition of P-gp activity. Monoglycerides could therefore alter the pharmacokinetics of drugs by inhibiting the P-gp-mediated efflux.     

Effect of organophosphate pesticide diazinon on expression and activity of intestinal P-glycoprotein

Organophosphate insecticide diazinon is widely used in agricultural practices, submitting farmers to repeated exposure. Because efflux pumps, as P-glycoprotein (P-gp), serve both as natural defense mechanisms and influence the bioavailability and disposition of drugs, we analyzed the ability of diazinon to act as efflux modulator. Oral administration of diazinon (2-20 mg/kg, 5 days, or 10 mg/kg, 2-12 days) increased intestinal mdr1a mRNA of rats, in both dose- and time-dependent manner, and increased the expression of intestinal P-gp. Using the intestinal cell-line Caco-2, we found that 100 microM diazinon significantly inhibited digoxin and vinblastine secretive flux through the cell monolayers, whereas digoxin and vinblastine absorptive flux increased. The 25 microM diazinon was transported preferentially in basolateral (BL) to apical (AP) direction, suggesting a net secretion. The efflux rate significantly decreased in the presence of metabolic inhibitors sodium azide and 2-deoxy-d-glucose, P-gp inhibitors cyclosporin A and valspodar, but not in the presence of MRPs inhibitor MK571. Repeated exposure of Caco-2 cells to diazinon increased P-glycoprotein expression and activity. These results suggested the involvement of P-gp in the transfer of diazinon, leading to potential consequences for xenobiotic interactions, and showed that repeated exposure to low doses of pesticide may lead to up-regulated P-gp functions in the intestine of mammals.     

Effects of lipopolysaccharide on intestinal P-glycoprotein expression and activity

It is well known that pharmacokinetics is often altered by changing the expression and activity of P-glycoprotein during sepsis. However, there have been few reports about expression and activity of P-glycoprotein in the small intestine during sepsis. We examined the levels of intestinal P-glycoprotein expression and activity using a rat sepsis model induced by lipopolysaccharide (LPS, from Escherichia coli). LPS was administered to male Wistar/ST rats intraperitonealy (i.p.) at 5 mg/kg. The small intestine was excised before and 1, 3 and 7 days after LPS administration, and the intestinal P-glycoprotein expression was determined using Western blot analysis. The activity of P-glycoprotein was evaluated by measuring the efflux of rhodamine-123 (Rho123) in rats using an in situ single perfusion method. The changes of permeability via the paracellular route were evaluated by measuring the amount of fluorescein isothicyanate-dextran 4400 (FD-4) in a similar way. On Day 1 after LPS administration, both the level of P-glycoprotein expression and the total amount of Rho123 excreted into the intestinal lumen decreased significantly, but levels of both AUC2-95 and CLtot were not significantly different as compared with the control group. On Day 3, the total P-glycoprotein, including intestinal P-glycoprotein, might have been induced by sepsis, and then the excretion of P-glycoprotein substrate drugs into the intestinal lumen increased more than that of the control group. On Day 7, all pharmacokinetic parameters returned to the control level. Thus the intestinal P-glycoprotein function recovered within 3 days of LPS administration.     

Biphasic regulation of P-glycoprotein function and expression by NO donors in Caco-2 cells

Aim:

To investigate the effects of nitric oxide (NO) donors on the function and expression of P-glycoprotein (P-gp) in Caco-2 cells.

Methods:

Caco-2 cells were exposed to NO donors for designated times. P-gp function and expression were assessed using Rhodamine123 uptake assay and Western blotting, respectively. Intracellular reactive oxygen species (iROS) and intracellular reactive nitrogen species (iRNS) levels were measured using ROS and RNS assay kits, respectively.

Results:

Exposure of Caco-2 cells to 0.1 or 2 mmol/L of sodium nitroprusside (SNP) affected the function and expression of P-gp in concentration- and time-dependent manners. A short-term (4 h) exposure reduced P-gp function and expression accompanied with significantly increased levels of iROS and iRNS. In contrast, a long-term (24 h) exposure stimulated the P-gp function and expression. The stimulatory effects of 2 mmol/L SNP was less profound as compared to those caused by 0.1 mmol/L SNP. The other NO donors SIN-1 and SNAP showed similar effects. Neither the NO scavenger PTIO (2 mmol/L) nor soluble guanylate cyclase inhibitor ODQ (50 μmol/L) reversed the SNP-induced alteration of P-gp function. On the other hand, free radical scavengers ascorbate, glutathione and uric acid (2 mmol/L for each), PKC inhibitor chelerythrine (5 μmol/L), PI3K/Akt inhibitor wortmannin (1 μmol/L) and p38 MAPK inhibitor SB203580 (10 μmol/L) reversed the upregulation of P-gp function by the long-term exposure to SNP, but these agents had no effect on the impaired P-gp function following the short-term exposure to SNP.

Conclusion:

NO donors time-dependently regulate P-gp function and expression in Caco-2 cells: short-term exposure impairs P-gp function and expression, whereas long-term exposure stimulates P-gp function and expression. The regulation occurs via a NO-independent mechanism.     

In vitro potential modulation of baicalin and baicalein on P-glycoprotein activity and expression in Caco-2 cells and rat gut sacs

Context Previous studies have shown that Scutellariae Radix, the dried root of Scutellaria baicalensis Georgi (Labiatae), has a certain inhibitory effect on P-glycoprotein (P-gp), but the effects of its main active constituents on P-gp are still ambiguous.

Objectives In vitro studies were performed to investigate the effects of its main active constituents (baicalin and its aglycone, baicalein) on the activity and expression of P-gp in intestine using Caco-2 cells and rat gut sacs.

Materials and methods In Caco-2 cell experiments, the effects of baicalin and baicalein on P-gp activity were investigated using a P-gp substrate, rhodamine 123 and non-substrate fluorescein Na, by determining their intracellular fluorescence accumulation, and their effects on P-gp expression were determined using flow cytometry. In addition, rat gut sac model was selected to investigate the effects of baicalin and baicalein on the transport of verapamil, a classical P-gp substrate. The gut sacs of male Sprague–Dawley rats were filled with 0.4?mL the test solution contained verapamil (0.2575?mg/mL) and the drugs [baicalin and baicalein, at concentrations of 1/8 IC₅₀ (59.875, 41.5?μg/mL), 1/4 IC₅₀ (119.75, 83?μg/mL) and 1/2 IC₅₀ (239.5, 166?μg/mL)], and then incubated in Tyrode’s solution for a period of time. After termination of the incubation, the incubated solution was processed for the subsequent detection.

Results According to the results of MTT assay, the IC₅₀ values of verapamil, baicalin and baicalein were 104, 479, 332?μg/mL, respectively. The obtained results from the two models were confirmed mutually. As a result, baicalin exhibited no obvious effect on intracellular accumulation of Rh-123, and almost had no effect on P-gp expression and verapamil transportation, while baicalein significantly increased intracellular accumulation of Rh-123 (p?<?0.01), down-regulated P-gp expression (p?<?0.01) and increased the transport of verapamil (p?<?0.05).

Discussion and conclusion The results indicated that baicalein may be a P-gp inhibitor, which presented obvious inhibitory effects on P-gp activity and expression level. A comparison of the structures of baicalin and baicalein indicates that the existence of glucosyl plays a decisive role in influencing the activity and expression of P-gp.     

Influence of tumor necrosis factor-alpha on the expression and function of P-glycoprotein in an immortalised rat brain capillary endothelial cell line,GPNT

Drug cerebral pharmacokinetics may be altered in the case of inflammatory diseases. This may be due to a modification of drug transport through the blood-brain barrier, in particular through drug interaction with the membrane efflux transporter, P-glycoprotein. The objective of this study was to investigate the influence of the inflammatory cytokine, tumor necrosis factor (TNF)-alpha, on the functionality and expression of P-glycoprotein, and on mdr1a and mdr1b mRNA expression in immortalised rat brain endothelial cells, GPNT. Cells were treated with TNF-alpha for 4 days. Levels of mdr1a and mdr1b mRNAs were quantitated using real-time RT-PCR analysis and expression of P-glycoprotein was analyzed by Western blot. The functionality of P-glycoprotein was studied by following the accumulation of [3H]vinblastine in the cells without and with a pre-treatment with a P-glycoprotein inhibitor, GF120918. TNF-alpha increased the levels of mdr1a and mdr1b mRNAs while no effect was observed on protein expression. TNF-alpha increased [3H]vinblastine accumulation indicating a time and concentration-dependent decrease of P-glycoprotein activity. This effect was eliminated when the cells were pre-treated with GF120918. Our observation of a decrease in P-glycoprotein activity could suggest that in the case of inflammatory diseases, brain delivery of P-glycoprotein-dependent drugs can be enhanced.     

Effects of monoglycerides on P-glycoprotein: modulation of the activity and expression in Caco-2 cell monolayers

The purpose of this study was to analyze the effects of two common monoglyceride components of lipid excipients, 1-monoolein and 1-monostearin, on the activity and expression of P-glycoprotein (P-gp) in Caco-2 cells. Non-cytotoxic concentrations of 1-monoolein and 1-monostearin were determined by assessing membrane permeability and mitochondrial activity in Caco-2 cells, a human colon adenocarcinoma cell line. Concentrations of 500 and 100 microM were used to evaluate P-gp activity through Rh123 accumulation and bifunctional transport studies. The P-gp protein expression levels were quantified through the use of immunoblots. The changes in cell membrane fluidity and nuclear membrane integrity upon the addition of monoglycerides were analyzed by fluorescence anisotropy using DPH and TMA-DPH as the fluorescent labels and by using increasing salt concentrations to release the nuclear contents, respectively. The absorptive flux (apical to basolateral) in the bifunctional transport studies was not found to be statistically significant for the non-cytotoxic concentrations of 1-monoolein and 1-monostearin. However, treatments of 500 and 100 microM of 1-monoolein or 1-monostearin displayed statistically lowered efflux (basolaterial to apical, P < 0.05) compared to the controls (7.9 +/- 0.8, 12.9 +/- 2.6 x 10 (6) cm/s for 1-monoolein or 11.1 +/- 2.0, 11.4 +/- 2.3 x 10 (6) cm/s for 1-monostearin, respectively, compared to the untreated control, 21.1 +/- 2.9 x 10 (6) cm/s, n = 5). Rh123 accumulation was also found to be enhanced upon 24 h incubation with both concentrations of the monoglycerides; however, only concentrations of 500 muM of the monoglycerides were shown to significantly reduce the P-gp protein expression. The results from this study suggest that these two monoglycerides, common components in various lipid excipients, are inhibitors of P-gp.     

银杏叶提取物对大鼠实验性结肠炎肿瘤坏死因子-α表达的影响

目的观察银杏叶提取物对三硝基苯磺酸灌肠诱导大鼠实验性结肠炎肿瘤坏死因子-α(TNF-α)的影响及其作用机制。方法大鼠随机分为正常对照组、三硝基苯磺酸模型组、阳性药物对照组、EGB组4组。用三硝基苯磺酸灌肠诱导大鼠实验性结肠炎,评估结肠组织大体形态和组织学评分;生化法检测大鼠肠组织谷胱苷肽过氧化物酶(GSH-Px)活性及一氧化氮(NO)含量;免疫组化检测肠组织TNF-α,核因子-κBp65(NF-κBp65)蛋白表达。逆转录聚合酶链反应(RT-PCR)检测肠组织诱生型一氧化氮合酶(iNOS)表达。结果EGB组大体形态和组织学评分较模型组明显下降,GSH-Px活性明显增高,NO含量明显减少,结肠黏膜TNF-α,NF-KBp65和iNOS表达明显降低。结论EGB可能通过抑制TNF-α的表达和NF-κBp65及iNOS的激活从而抑制炎症级联来保护TNBS诱导的实验性结肠炎。     

卡介菌多糖核酸对肺结核病人外周血细胞因子mRNA表达的调节

目的 :观察活动性肺结核病人外周血单个核细胞 (PBMC)细胞因子信息核糖核酸 (mRNA)表达特点以及卡介菌多糖核酸的调节作用。方法 :活动性肺结核病人和健康志愿者各 15例 ,分离单个核细胞。分 3组进行培养 ,A组为不加刺激物的对照组 ,B组加入结核菌素纯蛋白衍生物 (PPD) ,C组加入卡介菌多糖核酸 (BCG PSN )。检测PBMCγ 干扰素 (IFN γ) ,肿瘤坏死因子 (TNF α)和白细胞介素10 (IL 10 )mRNA表达。结果 :活动性肺结核病人PBMC在A组IFN γmRNA的表达明显低于健康者 ,P <0 .0 1,TNF α与IL 10mRNA的表达与健康人无差异。在B组IFN γmRNA和TNF αmRNA表达与健康人无差异 ;IL 10mRNA的表达增强 ,与健康人比较 ,P <0 .0 1。在C组 ,IFN γmRNA的表达增强 ,仍低于健康人 ,P <0 .0 5 ;TNF αmRNA的表达有增加 ,与健康人比较 ,P >0 .0 5 ,IL 10mR NA的表达无改变。结论 :活动性肺结核病人保护性细胞因子IFN γ减低 ;BCG PSN刺激可提高免疫细胞IFN γmRNA ,但不增高IL 10mRNA的生成     

小檗碱对高果糖饲养诱导大鼠胰岛素抵抗和肝脏TNF-α表达的影响

目的研究小檗碱对高果糖饲养诱导胰岛素抵抗大鼠的作用。方法高果糖饲料喂养SD大鼠6wk后,分为3组,模型组,小檗碱组(187.5mg.kg-1.d-1灌服)、二甲双胍组(184mg.kg-1.d-1灌服)作为对照,继续高果糖饮食。实验同时设立正常对照组,普通饮食喂养。4wk后处死大鼠;测定血糖、血清胰岛素、胰岛素抵抗指数及血脂的变化,用RT-PCR方法观察肝脏TNF-αmRNA的表达。结果模型组胰岛素、胰岛素抵抗指数、游离脂肪酸(free fatty acids,FFA)及甘油三酯(triglycerides,TG)水平均明显升高;肝脏TNF-αmRNA的表达与正常对照组比较明显升高。小檗碱降低了胰岛素抵抗大鼠的血糖、血清胰岛素、胰岛素抵抗指数、TG、FFA及TNF-αmRNA的表达。二甲双胍降低了大鼠的血糖、血清胰岛素、胰岛素抵抗指数及TG。结论小檗碱可以改善胰岛素抵抗,其机制可能包括抑制肝脏TNF-αmR-NA的表达与改善脂代谢紊乱。     

Role of tumor necrosis factor-alpha in down-regulation of hepatic cytochrome P450 and P-glycoprotein by endotoxin

We investigated the role of tumor necrosis factor-alpha (TNF-alpha) in the down-regulation of hepatic P-glycoprotein and cytochrome P450 (CYP) by endotoxin, using TNF-alpha gene-deficient (TNF-alpha-/-) mice. In the case of P-glycoprotein, endotoxin (10 mg/kg) significantly decreased the expression of hepatic P-glycoprotein in wild-type mice 6 h, but not 24 h, after intraperitoneal injection, with no significant differences in the constitutional expression of P-glycoprotein between wild-type mice and TNF-alpha-/- mice. However, endotoxin had no effect on the expression of P-glycoprotein in TNF-alpha-/- mice either 6 or 24 h after injection. When doxorubicin was administered intravenously to TNF-alpha-/- mice treated 6 h earlier with and without endotoxin, no significant differences in the plasma concentrations of doxorubicin 3 h after injection were observed between endotoxin-treated and untreated TNF-alpha-/- mice. These results suggest that TNF-alpha plays a pivotal role in the down-regulation of P-glycoprotein by endotoxin. In the case of CYP, the constitutive expression of hepatic CYP3A2 and CYP2C11 had a tendency to decline in TNF-alpha-/- mice compared with that in wild-type mice. Endotoxin significantly decreased the expression of hepatic CYP3A2 and CYP2C11 in wild-type mice 24 h after injection, and that decreased expression was significantly greater in TNF-alpha-/- mice than wild-type mice. When antipyrine was administered intravenously to wild-type mice and TNF-alpha-/- mice treated 24 h earlier with endotoxin, the plasma concentrations of antipyrine in TNF-alpha-/- mice 3 h after injection were significantly higher than those in wild-type mice. These findings suggest that TNF-alpha plays a key role in endotoxin-induced down-regulation of hepatic P-glycoprotein, as well as plays a protective role in the regulation of hepatic CYP3A2 and CYP2C11 against endotoxin-induced acute inflammatory response. In TNF-alpha-/- mice, other cytokines appear to function as compensation for the lack of endogenous TNF-alpha.     

Increased percentages of tumor necrosis factor-alpha+/interferon-gamma+ T [corrected] lymphocytes and calprotectin+/tumor necrosis factor-alpha+ monocytes in patients with acute Kawasaki disease

In vivo exposure to microorganisms resident in the oral cavity is considered as a possible cause of Kawasaki disease (KD), and some epitopes derived from streptococci display homology with Factor H of Complement. Additionally, calprotectin, a major calcium binding protein released by neutrophils and activated monocytes, could be directly involved in endothelial damage occurring in KD. The aim of our study is to evaluate the percentages of IFN-gamma+ and/or TNF-alpha+ lymphocytes and double positive calprotectin/TNF-alpha monocytes (CD14+) after in vitro stimulation with streptococcal- and/or Factor H-derived peptides, in patients with acute KD. Peripheral Blood Mononuclear Cells (PBMCs) obtained from KD patients and febrile controls were stimulated in vitro with peptides. After culture, cells were collected, stained with fluorochrome-labelled monoclonal antibodies against CD3, CD14, calprotectin, IFN-gamma and TNF-alpha, and cytofluorimetric analyses were performed. Our results showed increased percentages of TNF-alpha+/IFN-gamma+ lymphocytes in KD patients in respect to controls when PBMCs were stimulated with streptococcal or Factor H-derived epitopes. In addition, also calprotectin+/TNF-alpha+ monocytes from KD patients were activated after PBMC in vitro stimulation. These findings lead us to speculate that some peptides, derived from oral streptococci and cross-reactive with the human Factor H of Complement, could induce lymphocyte and monocyte activation potentially involved in the pathogenesis of KD. Our results should be confirmed by further studies enrolling more patients and controls than those analyzed in our study.     

beta-adrenoceptor agonists downregulate adiponectin, but upregulate adiponectin receptor 2 and tumor necrosis factor-alpha expression in adipocytes

Recently, the insulin-sensitizing adipokine adiponectin and the insulin resistance-inducing adipokine tumor necrosis factor-alpha (TNF-alpha) were reported to inhibit each other's production in adipocytes. We investigated the effects of two beta(3)-adrenoceptor agonists, 5-[(2R)-2-[[(2R)-2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]-1,3-benzodioxole-2,2-dicarboxylate (CL-316,243) and (+/-)-(R(*),R(*))-[4-[2-[[2-(3-chlorophenyl)-2-hydroxyethyl]amino]propyl]phenoxy]acetic acid (BRL37344), on the gene expression of adiponectin, two adiponectin receptors, and TNF-alpha in adipose tissues of C57BL/6J mice. CL-316,243 and BRL37344 downregulated adiponectin, but upregulated adiponectin receptor 2 (not receptor 1) in epididymal or/and subcutaneous white adipose tissues and in brown adipose tissue. TNF-alpha expression was upregulated only in epididymal adipose tissue. To further explore these effects, we treated differentiated 3T3-L1 adipocytes with the non-selective beta-adrenoceptor agonist isoproterenol. As a result, adiponectin receptor 2 (but not receptor 1) gene expression and TNF-alpha protein expression increased, but gene expression and secretion of adiponectin decreased. The upregulation of adiponectin receptor 2 by isoproterenol is most likely via beta(2),beta(3)-adrenoceptors, adenylyl cyclases, and protein kinase A (PKA). However, the accompanying activation of AMP-activated protein kinase (AMPK) may inhibit this upregulation. Our results suggest that upregulation of TNF-alpha and downregulation of adiponectin by beta-adrenoceptor activation may contribute to the pathogenesis of catecholamine-induced insulin resistance, and that upregulation of adiponectin receptor 2 may be a feedback result of reduced adiponectin.     

Anti-inflammatory activity of aucubin by inhibition of tumor necrosis factor-alpha production in RAW 264.7 cells

To elucidate a possible mechanism for the anti-inflammatory action of iridoid glycosides, the effects of both aucubin (AU) and its hydrolyzed product (H-AU) by beta-glucosidase treatment were studied on the production of TNF-alpha in RAW 264.7 cells. H-AU suppressed the production of both mRNA for TNF-alpha and subsequent TNF-alpha protein in the culture, but AU did not. The production of TNF-alpha protein was inhibited in a dose-dependent manner with an IC (50) of 9.2 microM. In addition, treatment with H-AU blocked both the I-kappa B alpha degradation and the translocation of NF-kappa B from the cytosol fraction to the nuclear fraction (55 % inhibition) in the culture. However, treatment with H-AU did not affect the intracellular level of cAMP formed by forskolin treatment in human monocytes U937 culture, implying that there is no influence on the cAMP level in other cell systems. The present study indicates a possible justification for those medicinal plants containing iridoid glycoside that have been used for the treatment of inflammation.