Total Synthesis and Anti-inflammatory Activity of Asperjinone and Asperimide C.

Total syntheses of two γ-butenolide natural products, asperjinone (1) and asperimide C (2) in both racemic and chiral forms have been accomplished utilizing Basavaiah's one-pot Friedel-Crafts/maleic anhydride formation protocol as a key strategy. Our syntheses verified the revised structure of 1 proposed by Williams et al. and the structure and absolute configuration of 2 reported by the Li group. This work also discloses the unprecedented anti-inflammatory activity of 1. Synthetic 1 exhibited significant anti-inflammatory activity in renal proximal tubular epithelial cells (RPTEC) by suppression of gene expression of pro-inflammatory cytokines TNF-α, IL-1ß and IL-6 under LPS-induced renal inflammation condition and was superior to (S)-1, rac-2, 2, and a positive drug control, indomethacin. Moreover, compound 1 inhibited downstream signaling of inflammation by significantly reducing iNOS and COX-2 gene expression and total NO production. The anti-inflammatory activity of asperjinone (1) renders it a potential and promising candidate for developing novel anti-inflammatory agents against inflammation worsening acute kidney injury.

Enhancing effect of natural adjuvant, panduratin A, on antibacterial activity of colistin against multidrug-resistant Acinetobacter baumannii.

Colistin- and carbapenem-resistant Acinetobacter baumannii is a serious multidrug resistant (MDR) bacterium in clinical settings. Discovery of new antibacterial drugs against MDR is facing multiple challenges in drug development. Combination of known antibiotics with a robust adjuvant might be an alternative effective strategy for MDR treatment. In the study herein, we report an antibiotic adjuvant activity of a natural compound panduratin A from fingerroot (Boesenbergia rotunda) as a potent adjuvant to colistin. The present study investigated the antibiotic adjuvant effect of panduratin A against 10 colistin- and carbapenem-resistant A. baumannii. Antibacterial activities were tested by broth microdilution method. Biofilm assay was used to determine the efficacy of panduratin A in biofilm formation inhibition on two representative strains Aci46 and Aci44. Genomic and transcriptomic analyses of colistin- and carbapenem-resistant A. baumannii strains were used to identify potential resistance and tolerance mechanism in the bacteria. Panduratin A-colistin combination showed an increased effect on antibacterial in the A. baumannii. However, panduratin A did not improve the antibacterial activity of imipenem. In addition, panduratin A improves anti-biofilm activity of colistin against Aci44 and Aci46, the colistin- and carbapenem-resistant A. baumannii. Panduratin A markedly enhances bactericidal and anti-biofilm activity of colistin against colistin- resistant A. baumannii. Based on genome comparisons, single nucleotide polymorphism (SNP) patterns in six genes encoding biofilm and lipid A biosynthesis were shared in Aci44 and Aci46. In Aci44, we identified a partial sequence of pmrB encoding a polymyxin resistant component PmrB, whereas a full length of pmrB was observed in Aci46. RNA-seq analyses of Aci44 revealed that panduratin A-colistin combination induced expression of ribosomal proteins and oxidative stress response proteins, whereas iron transporter and MFS-type transporter systems were suppressed. Panduratin A-colistin combination could promote intracellular reactive oxygen species (ROS) accumulation could lead to the cidal effect on colistin-resistant A. baumannii. Combination of panduratin A and colistin showed a significant increase in colistin efficacy against colistin- resistant A. baumannii in comparison of colistin alone. Genomic comparison between Aci44 and Aci46 showed mutations and SNPs that might affect different phenotypes. Additionally, based on RNA-Seq, panduratin A-colistin combination could lead to ROS production and accumulation. These findings confirmed the potency of panduratin as colistin adjuvant against multidrug resistant A. baumannii.

Activation of farnesoid X receptor retards expansion of renal collecting duct cell-derived cysts via inhibition of CFTR-mediated Cl- secretion.

The transcription factor farnesoid X receptor (FXR) regulates energy metabolism. Specifically, FXR functions to regulate cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl- secretion in intestinal epithelial cells. Therefore, this study aimed to investigate the role of FXR in CFTR-mediated Cl- secretion in renal tubular cells and to further elucidate its effects on renal cyst formation and growth. CFTR-mediated Cl- transport was evaluated via short-circuit current (ISC) measurements in Madin-Darby canine kidney (MDCK) cell monolayers and primary rat inner medullary collecting duct cells. The role of FXR in renal cyst formation and growth was determined by the MDCK cell-derived cyst model. Incubation with synthesized (GW4064) and endogenous (CDCA) FXR ligands reduced CFTR-mediated Cl- secretion in a concentration- and time-dependent manner. The inhibitory effect of FXR ligands was not due to the result of reduced cell viability and was attenuated by cotreatment with an FXR antagonist. FXR activation significantly decreased CFTR protein but not its mRNA. In addition, FXR activation inhibited CFTR-mediated Cl- secretion in primary renal collecting duct cells. FXR activation decreased ouabain-sensitive ISC without altering Na+-K+-ATPase mRNA and protein levels. Furthermore, FXR activation significantly reduced the number of cysts and renal cyst expansion. These inhibitory effects were correlated with a decrease in the expression of protein synthesis regulators mammalian target of rapamycin/S6 kinase. This study shows that FXR activation inhibits Cl- secretion in renal cells via inhibition of CFTR expression and retards renal cyst formation and growth. The discoveries point to a physiological role of FXR in the regulation of CFTR and a potential therapeutic application in polycystic kidney disease treatment.NEW & NOTEWORTHY The present study reveals that farnesoid X receptor (FXR) activation reduces microcyst formation and enlargement. This inhibitory effect of FXR activation is involved with decreased cell proliferation and cystic fibrosis transmembrane conductance regulator-mediated Cl- secretion in renal collecting duct cells. FXR might represent a novel target for the treatment of autosomal dominant polycystic kidney disease.

Altenusin, a fungal metabolite, alleviates TGF-ß1-induced EMT in renal proximal tubular cells and renal fibrosis in unilateral ureteral obstruction.

Renal fibrosis is a pathological feature of chronic kidney disease (CKD), progressing toward end-stage kidney disease (ESKD). The aim of this study is to investigate the therapeutic potential of altenusin, a farnesoid X receptor (FXR) agonist derived from fungi, on renal fibrosis. The effect of altenusin was determined (i) in vitro using the transforming growth factor ß1 (TGF-ß1)-induced epithelial to mesenchymal transition (EMT) of human renal proximal tubular cells and (ii) in vivo using mouse unilateral ureteral obstruction (UUO). The findings revealed that incubation of 10 ng/ml TGF-ß1 promotes morphological change in RPTEC/TERT1 cells, a human renal proximal tubular cell line, from epithelial to fibroblast-like cells. TGF-ß1 markedly increased EMT markers namely α-smooth muscle actin (α-SMA), fibronectin, and matrix metalloproteinase 9 (MMP-9), while decreased the epithelial marker E-cadherin. Co-incubation TGF-ß1 with altenusin preserved the epithelial characteristics of the renal epithelial cells by antagonizing TGF-ß/Smad signaling pathway, specifically a decreased phosphorylation of Smad2/3 with an increased level of Smad7. Interestingly, the antagonizing effect of altenusin does not require FXR activation. Moreover, altenusin could reverse TGF-ß1-induced fibroblast-like cells to epithelial-like cells. Treatment on UUO mice with 30 mg/kg altenusin significantly reduced the expression of α-SMA, fibronectin, and collagen type 1A1 (COL1A1). The reduction in the renal fibrosis markers is correlated with the decreased phosphorylation of Smad2/3 levels but does not improve E-cadherin protein expression. Collectively, altenusin reduces EMT in human renal proximal tubular cells and renal fibrosis by antagonizing the TGF-ß/Smad signaling pathway.

Genipin Analogue (G300) Inhibits Adipogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells through the Suppression of Adipogenic Promoting Factors.

While obesity is a well-known health threatening condition worldwide, effective pharmacological interventions for obesity suppression have been limited due to adverse effects. Therefore, it is important to explore alternative medical treatments for combating obesity. Inhibition of the adipogenesis process and lipid accumulation are critical targets for controlling and treating obesity. Gardenia jasminoides Ellis is a traditional herbal remedy for various ailments. A natural product from its fruit, genipin, has major pharmacological properties; it is anti-inflammatory and antidiabetic. We investigated the effects of a genipin analogue, G300, on adipogenic differentiation in human bone marrow mesenchymal stem cells (hBM-MSCs). G300 suppressed the expression of adipogenic marker genes and adipokines secreted by adipocytes at concentrations of 10 and 20 µM, which effectively reduced the adipogenic differentiation of hBM-MSCs and lipid accumulation in adipocytes. It also improved adipocyte function by lowering inflammatory cytokine secretion and increasing glucose uptake. For the first time, we show that G300 has the potential to be a novel therapeutic agent for the treatment of obesity and its related disorders.

Black ginger extract and its active compound, 5,7-dimethoxyflavone, increase intestinal drug absorption via efflux drug transporter inhibitions.

Black ginger is used as an herbal medicine for self-care and health promotion. Black ginger extract has been shown to alter the function of transporters in several cell types. This study demonstrates the interaction between the extract and 5,7-dimethoxyflavone (DMF) on drug efflux mediated by breast cancer resistance proteins (BCRP) and P-glycoprotein (P-gp) in Caco-2 cells and heterologous cell systems [Madin-Darby canine kidney type II (MDCKII) stably transfected with human BCRP (MDCKII/BCRP) or human P-gp (MDCKII/P-gp)]. The transepithelial flux of 3H-Digoxin and 3H-Estrone sulfate, prototypic substrates of P-gp, and BCRP, respectively, across Caco-2 cell monolayers, MDCKII/BCRP, and MDCKII/P-gp cells were determined. The results demonstrate that black ginger extract (10 µg/ml) significantly increases 3H-Digoxin and 3H-Estrone sulfate transport from the apical to basolateral side while decreasing transport from the basolateral to apical side of Caco-2 cells and MDCKII cell overexpression of BCRP or P-gp. The effect of the extract on 3H-Digoxin and 3H-Estrone sulfate transport was related to a decrease in efflux ratio. Likewise, DMF (5 µM) significantly increased 3H-Digoxin and 3H-Estrone sulfate absorption with a decreased efflux ratio compared to the control. Interestingly, the extract also significantly increased absorption of paclitaxel, an anti-cancer drug, which has poor oral absorption. Taken together, co-administration of drugs as substrates of BCRP and P-gp, with the black ginger extract containing DMF, might alter the pharmacokinetic profiles of the medicine.

Interaction of buspirone and its major metabolites with human organic cation transporters.

Buspirone, a cationic drug, is an anxiolytic and antidepressant drug. However, whether buspirone and its metabolites are interacted with organic cationic transporter remains uncertain. In this study, we examined the interaction of buspirone and its major metabolites 1-(2-pyrimidinyl)piperazine (1-PP) and 6-hydroxybuspirone (6'-OH-Bu) with hOCTs using human hepatocellular carcinoma (HepG2), human colorectal adenocarcinoma (Caco-2) cells, and S2 cells expressing OCT1 (S2hOCT1), 2 (S2hOCT2), or 3 (S2hOCT3). Coadministration of buspirone and fluorescent 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP+ ) was examined using HepG2 cells, and [3 H]-1-methyl-4-phenylpyridinium (MPP+ ) transport was assessed in S2 cell overexpressing hOCTs. The results showed that ASP+ transport was suppressed by buspirone with an IC50 of 26.3 ± 2.9 µM without any cytotoxic effects in HepG2 expressing hOCTs cells. Consistently, buspirone strongly inhibited [3 H]-MPP+ uptake by S2hOCT1, S2hOCT2, and S2hOCT3 cells with an IC50s of 89.0 ± 1.3 µM, 43.7 ± 7.5 µM, and 20.4 ± 1.0 µM, respectively. Nonetheless, 6'-OH-Bu and 1-PP caused weak or no inhibition on ASP+ and [3 H]-MPP+ transport. These findings suggest the potential interaction of buspirone with organic cation drugs that are handled by hOCT3. However, further clinical relevance is needed to support these findings for preventing drug-drug interaction in patients who take prescribed drugs together with buspirone.

Anti-Inflammatory Activity of Panduratin A against LPS-Induced Microglial Activation.

Uncontrolled and excessive microglial activation is known to contribute to inflammation-mediated neurodegeneration. Therefore, reducing neurotoxic microglial activation may serve as a new approach to preventing neurodegeneration. Here, we investigated the anti-inflammatory effects of panduratin A against microglial activation induced by lipopolysaccharides (LPS) in the SIMA9 microglial cell line. We initially examined the anti-inflammatory properties of panduratin A by measuring LPS-induced nitric oxide (NO) production and the levels of pro-inflammatory cytokines (TNF-α, IL-1ß, and IL-6). Panduratin A significantly reduced NO levels and pro-inflammatory cytokines' production and secretion. In addition, panduratin A enhanced the production of anti-inflammatory cytokines IL-4 and IL-10. The anti-inflammatory effects of panduratin A are related to the suppression of the NF-κB signaling pathway. Together, these results demonstrate the anti-inflammatory properties of panduratin A against LPS-induced microglial activation, suggesting panduratin A has the potential to be further developed as a new agent for the prevention of neuroinflammation-associated neurodegenerative diseases.

Pharmacological Effects of Panduratin A on Renal Cyst Development in In Vitro and In Vivo Models of Polycystic Kidney Disease.

Renal cyst expansion in polycystic kidney disease (PKD) involves abnormalities in both cyst-lining-cell proliferation and fluid accumulation. Suppression of these processes may retard the progression of PKD. Evidence suggests that the activation of 5' AMP-activated protein kinase (AMPK) inhibits cystic fibrosis transmembrane conductance regulator (CFTR)-mediated chloride secretion, leading to reduced progression of PKD. Here we investigated the pharmacological effects of panduratin A, a bioactive compound known as an AMPK activator, on CFTR-mediated chloride secretion and renal cyst development using in vitro and animal models of PKD. We demonstrated that AMPK was activated in immortalized normal renal cells and autosomal dominant polycystic kidney disease (ADPKD) cells following treatment with panduratin A. Treatment with panduratin A reduced the number of renal cyst colonies corresponding with a decrease in cell proliferation and phosphorylated p70/S6K, a downstream target of mTOR signaling. Additionally, panduratin A slowed cyst expansion via inhibition of the protein expression and transport function of CFTR. In heterozygous Han:Sprague-Dawley (Cy/+) rats, an animal model of PKD, intraperitoneal administration of panduratin A (25 mg/kg BW) for 5 weeks significantly decreased the kidney weight per body weight ratios and the cystic index. Panduratin A also reduced collagen deposition in renal tissue. Intraperitoneal administration of panduratin A caused abdominal bleeding and reduced body weight. However, 25 mg/kg BW of panduratin A via oral administration in the PCK rats, another non-orthologous PKD model, showed a significant decrease in the cystic index without severe adverse effects, indicating that the route of administration is critical in preventing adverse effects while still slowing disease progression. These findings reveal that panduratin A might hold therapeutic properties for the treatment of PKD.

Pinostrobin inhibits renal CFTR-mediated Cl- secretion and retards cyst growth in cell-derived cyst and polycystic kidney disease rats.

Cystic fibrosis transmembrane conductance regulator (CFTR) plays crucial role in renal cyst expansion via increase in fluid accumulation. Inhibition of CFTR has been proposed to retard cyst development and enlargement in polycystic kidney disease (PKD). Pinostrobin, a bioactive natural flavonoid, possesses several pharmacological effects. The present study investigated pharmacological effects of pinostrobin on CFTR-mediated Cl- secretion and renal cyst expansion in in vitro and in vivo models. Pinostrobin (10 and 50 µM) reduced number of MDCK cell-derived cyst colonies and inhibited cyst expansion via inhibition of cell proliferation and CFTR-mediated Cl- secretion. The inhibitory effect of pinostrobin was not due to the decrease in cell viability and activity of Na+-K+-ATPase. We also investigated the natural analogue pinocembrin as well as the synthetic analogue pinostrobin oxime. Both pinocembrin and pinostrobin oxime did not reduce CFTR-mediated Cl- secretion. In PKD rats, oral administration of pinostrobin (40 mg/kg/day) exhibited a decreasing in cystic area compared to vehicle-treated rats. Pinostrobin treatment inhibited renal expression of CFTR protein in PKD rats. Our findings highlighted the potential therapeutic application of pinostrobin in PKD.

Nephroprotective potential of Panduratin A against colistin-induced renal injury via attenuating mitochondrial dysfunction and cell apoptosis.

Colistin is a last-resort polypeptide antibiotic widely used to treat against multidrug-resistant Gram-negative bacterial infections. However, this treatment is associated with nephrotoxicity. The aim of this study was to examine the potential protective effect of panduratin A, a bioactive compound of Boesenbergia rotunda, on colistin-induced nephrotoxicity in both in vivo and in vitro models. Intraperitoneal injection of 15 mg/kg colistin for 7 days markedly promoted renal tubular degeneration, increased blood urea nitrogen (BUN) levels, and upregulated the expression of renal injury biomarker and apoptosis proteins. In addition, treatment with colistin increased oxidative stress and apoptosis in mice kidney tissues. Interestingly, these defects were attenuated when co-administered of colistin with panduratin A (2.5 or 25 mg/kg). The underlying mechanisms of panduratin A attenuating colistin toxicity was investigated in human renal proximal tubular cells (RPTEC/TERT1). The mechanisms by which colistin-triggered cytotoxicity was determined by analysis of cell death, reactive oxygen species (ROS) levels, mitochondria function as well as the expression of proteins related to apoptosis pathway. Colistin treatment (200 µg/ml) significantly increased cell apoptosis, elevated ROS production, reduced mitochondrial membrane potential, and decreased anti-apoptotic protein (Bcl-2) expression. These effects were notably suppressed by co-treatment with panduratin A (5 µM). Collectively, panduratin A exerts as a novel nephroprotective agent to protect against colistin-induced renal injury by attenuating mitochondrial damage and renal cell apoptosis.

Panduratin A Derivative Protects against Cisplatin-Induced Apoptosis of Renal Proximal Tubular Cells and Kidney Injury in Mice.

BACKGROUND: Panduratin A is a bioactive cyclohexanyl chalcone exhibiting several pharmacological activities, such as anti-inflammatory, anti-oxidative, and anti-cancer activities. Recently, the nephroprotective effect of panduratin A in cisplatin (CDDP) treatment was revealed. The present study examined the potential of certain compounds derived from panduratin A to protect against CDDP-induced nephrotoxicity. METHODS: Three derivatives of panduratin A (DD-217, DD-218, and DD-219) were semi-synthesized from panduratin A. We investigated the effects and corresponding mechanisms of the derivatives of panduratin A for preventing nephrotoxicity of CDDP in both immortalized human renal proximal tubular cells (RPTEC/TERT1 cells) and mice. RESULTS: Treating the cell with 10 µM panduratin A significantly reduced the viability of RPTEC/TERT1 cells compared to control (panduratin A: 72% ± 4.85%). Interestingly, DD-217, DD-218, and DD-219 at the same concentration did not significantly affect cell viability (92% ± 8.44%, 90% ± 7.50%, and 87 ± 5.2%, respectively). Among those derivatives, DD-218 exhibited the most protective effect against CDDP-induced renal proximal tubular cell apoptosis (control: 57% ± 1.23%; DD-218: 19% ± 10.14%; DD-219: 33% ± 14.06%). The cytoprotective effect of DD-218 was mediated via decreases in CDDP-induced mitochondria dysfunction, intracellular reactive oxygen species (ROS) generation, activation of ERK1/2, and cleaved-caspase 3 and 7. In addition, DD-218 attenuated CDDP-induced nephrotoxicity by a decrease in renal injury and improved in renal dysfunction in C57BL/6 mice. Importantly, DD-218 did not attenuate the anti-cancer efficacy of CDDP in non-small-cell lung cancer cells or colon cancer cells. CONCLUSIONS: This finding suggests that DD-218, a derivative of panduratin A, holds promise as an adjuvant therapy in patients receiving CDDP.

Colistin and Carbapenem-Resistant Acinetobacter baumannii Aci46 in Thailand: Genome Analysis and Antibiotic Resistance Profiling.

Resistance to the last-line antibiotics against invasive Gram-negative bacterial infection is a rising concern in public health. Multidrug resistant (MDR) Acinetobacter baumannii Aci46 can resist colistin and carbapenems with a minimum inhibitory concentration of 512 µg/mL as determined by microdilution method and shows no zone of inhibition by disk diffusion method. These phenotypic characteristics prompted us to further investigate the genotypic characteristics of Aci46. Next generation sequencing was applied in this study to obtain whole genome data. We determined that Aci46 belongs to Pasture ST2 and is phylogenetically clustered with international clone (IC) II as the predominant strain in Thailand. Interestingly, Aci46 is identical to Oxford ST1962 that previously has never been isolated in Thailand. Two plasmids were identified (pAci46a and pAci46b), neither of which harbors any antibiotic resistance genes but pAci46a carries a conjugational system (type 4 secretion system or T4SS). Comparative genomics with other polymyxin and carbapenem-resistant A. baumannii strains (AC30 and R14) identified shared features such as CzcCBA, encoding a cobalt/zinc/cadmium efflux RND transporter, as well as a drug transporter with a possible role in colistin and/or carbapenem resistance in A. baumannii. Single nucleotide polymorphism (SNP) analyses against MDR ACICU strain showed three novel mutations i.e., Glu229Asp, Pro200Leu, and Ala138Thr, in the polymyxin resistance component, PmrB. Overall, this study focused on Aci46 whole genome data analysis, its correlation with antibiotic resistance phenotypes, and the presence of potential virulence associated factors.

Protective Effect of Panduratin A on Cisplatin-Induced Apoptosis of Human Renal Proximal Tubular Cells and Acute Kidney Injury in Mice.

BACKGROUND: Cisplatin is an effective chemotherapy but its main side effect, acute kidney injury, limits its use. Panduratin A, a bioactive compound extracted from Boesenbergia rotunda, shows several biological activities such as anti-oxidative effects. The present study investigated the nephroprotective effect of panduratin A on cisplatin-induced renal injury. METHODS: We investigated the effect of panduratin A on the toxicity of cisplatin in both mice and human renal cell cultures using RPTEC/TERT1 cells. RESULTS: The results demonstrated that panduratin A ameliorates cisplatin-induced renal toxicity in both mice and RPTEC/TERT1 cells by reducing apoptosis. Mice treated with a single intraperitoneal (i.p.) injection of cisplatin (20 mg/kg body weight (BW)) exhibited renal tubule injury and impaired kidney function as shown by histological examination and increased serum creatinine. Co-administration of panduratin A (50 mg/kg BW) orally improved kidney function and ameliorated renal tubule injury of cisplatin by inhibiting activation of extracellular signal-regulated kinase (ERK)1/2 and caspase 3. In human renal proximal tubular cells, cisplatin induced cell apoptosis by activating pro-apoptotic proteins (ERK1/2 and caspase 3), and reducing the anti-apoptotic protein (Bcl-2). These effects were significantly ameliorated by co-treatment with panduratin A. Interestingly, panduratin A did not alter intracellular accumulation of cisplatin. It did not alter the anti-cancer efficacy of cisplatin in either human colon or non-small cell lung cancer cell lines. CONCLUSIONS: The present study highlights panduratin A has a potential protective effect on cisplatin's nephrotoxicity.

Tiliacora triandra (Colebr.) Diels Leaf Aqueous Extract Inhibits Hepatic Glucose Production in HepG2 Cells and Type 2 Diabetic Rats.

This study investigated the effects of Tiliacora triandra (Colebr.) Diels aqueous extract (TTE) on hepatic glucose production in hepatocellular carcinoma (HepG2) cells and type 2 diabetic (T2DM) conditions. HepG2 cells were pretreated with TTE and its major constituents found in TTE, epicatechin (EC) and quercetin (QC). The hepatic glucose production was determined. The in vitro data were confirmed in T2DM rats, which were supplemented daily with 1000 mg/kg body weight (BW) TTE, 30 mg/kg BW metformin or TTE combined with metformin for 12 weeks. Results demonstrate that TTE induced copper-zinc superoxide dismutase, glutathione peroxidase and catalase genes, similarly to EC and QC. TTE decreased hepatic glucose production by downregulating phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) and increasing protein kinase B and AMP-activated protein kinase phosphorylation in HepG2 cells. These results correlated with the antihyperglycemic, antitriglyceridemic, anti-insulin resistance, and antioxidant activities of TTE in T2DM rats, similar to the metformin and combination treatments. Consistently, impairment of hepatic gluconeogenesis in T2DM rats was restored after single and combined treatments by reducing PEPCK and G6Pase genes. Collectively, TTE could potentially be developed as a nutraceutical product to prevent glucose overproduction in patients with obesity, insulin resistance, and diabetes who are being treated with antidiabetic drugs.

Novel Potential Application of Chitosan Oligosaccharide for Attenuation of Renal Cyst Growth in the Treatment of Polycystic Kidney Disease.

Chitosan oligosaccharide (COS), a natural polymer derived from chitosan, exerts several biological activities including anti-inflammation, anti-tumor, anti-metabolic syndrome, and drug delivery enhancer. Since COS is vastly distributed to kidney and eliminated in urine, it may have a potential advantage as the therapeutics of kidney diseases. Polycystic kidney disease (PKD) is a common genetic disorder characterized by multiple fluid-filled cysts, replacing normal renal parenchyma and leading to impaired renal function and end-stage renal disease (ESRD). The effective treatment for PKD still needs to be further elucidated. Interestingly, AMP-activated protein kinase (AMPK) has been proposed as a drug target for PKD. This study aimed to investigate the effect of COS on renal cyst enlargement and its underlying mechanisms. We found that COS at the concentrations of 50 and 100 µg/mL decreased renal cyst growth without cytotoxicity, as measured by MTT assay. Immunoblotting analysis showed that COS at 100 µg/mL activated AMPK, and this effect was abolished by STO-609, a calcium/calmodulin-dependent protein kinase kinase beta (CaMKKß) inhibitor. Moreover, COS elevated the level of intracellular calcium. These results suggest that COS inhibits cyst progression by activation of AMPK via CaMKKß. Therefore, COS may hold the potential for pharmaceutical application in PKD.

Germacrone Reduces Cisplatin-Induced Toxicity of Renal Proximal Tubular Cells via Inhibition of Organic Cation Transporter.

Cisplatin is a widely used chemotherapy for solid tumors; however, its benefits are limited by serious nephrotoxicity, particularly in proximal tubular cells. The present study investigated the renoprotective effect and mechanisms of germacrone, a bioactive terpenoid compound found in Curcuma species on cisplatin-induced toxicity of renal cells. Germacrone (50 and 100 µM) attenuated apoptosis of human renal proximal tubular cells, RPTEC/TERT1 following treatment with 50 µM cisplatin and for 48 h. Co-treating RPTEC/TERT1 cells with cisplatin and germacrone significantly reduced cellular platinum content compared with cisplatin treatment alone. The effect of germacrone on organic cation transporter 2 (OCT2) which is a transporter responsible for cisplatin uptake was determined. Germacrone showed an inhibitory effect on OCT2-mediated methyl-4-phenylpyridinium acetate (3H-MPP+) uptake with IC50 of 15 µM with less effect on OCT1. The germacrone's protective effect on cisplatin-induced cytotoxicity was not observed in cancer cells; cisplatin's anti-cancer activity was preserved. In conclusion, germacrone prevents cisplatin-induced toxicity in renal proximal tubular cells via inhibition OCT2 transport function and reducing cisplatin accumulation. Thus germacrone may be a good candidate agent used for reducing cisplatin-induced nephrotoxicity.

Farnesoid X Receptor Activation Stimulates Organic Cations Transport in Human Renal Proximal Tubular Cells.

Farnesoid X receptor (FXR) is a ligand-activated transcription factor highly expressed in the liver and kidneys. Activation of FXR decreases organic cation transporter (OCT) 1-mediated clearance of organic cation compounds in hepatocytes. The present study investigated FXR regulation of renal clearance of organic cations by OCT2 modulation and multidrug and toxin extrusion proteins (MATEs). The role of FXR in OCT2 and MATEs functions was investigated by monitoring the flux of 3H-MPP+, a substrate of OCT2 and MATEs. FXR agonists chenodeoxycholic acid (CDCA) and GW4064 stimulated OCT2-mediated 3H-MPP+ uptake in human renal proximal tubular cells (RPTEC/TERT1 cells) and OCT2-CHO-K1 cells. The stimulatory effect of CDCA (20 µM) was abolished by an FXR antagonist, Z-guggulsterone, indicating an FXR-dependent mechanism. CDCA increased OCT2 transport activity via an increased maximal transport rate of MPP+. Additionally, 24 h CDCA treatment increased MATEs-mediated 3H-MPP+ uptake. Moreover, CDCA treatment increased the expression of OCT2, MATE1, and MATE2-K mRNA compared with that of the control. OCT2 protein expression was also increased following CDCA treatment. FXR activation stimulates renal OCT2- and MATE1/2-K-mediated cation transports in proximal tubules, demonstrating that FXR plays a role in the regulation of OCT2 and MATEs in renal proximal tubular cells.

High affinity of 4-(4-(dimethylamino)styryl)-N-methylpyridinium transport for assessing organic cation drugs in hepatocellular carcinoma cells.

Human organic cation transporter 1 (hOCT1) and human organic cation transporter 3 (hOCT3) are highly expressed in hepatocytes and play important roles in cationic drug absorption, distribution, and elimination. A previous study demonstrated that downregulation of hOCT1 and hOCT3 mRNA was related to hepatocellular carcinoma (HepG2) prognosis and severity. Whether these transporters expressed in HepG2 cells serve for cationic drug delivery has not been investigated. Besides radioactive transport, options for assessing hOCTs in hepatocytes are limited. This study clarified the significant roles of hOCTs in HepG2 by comparing cationic fluorescent 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP+ ) with traditional [3 H]-1-methyl-4-phenylpyridinium (MPP+ ). The results showed ASP+ was preferably transported into HepG2 compared to [3 H]-MPP+ with high affinity and a high maximal transport rate. Selective transport of ASP+ mediated by hOCTs was influenced by extracellular pH, temperature, and membrane depolarization, corresponding to hOCT1 and hOCT3 expressions. Furthermore, transport of cationic drugs, metformin, and paclitaxel in HepG2 cells was blunted by OCT inhibitors, suggesting that hOCT1 and hOCT3 expressed in HepG2 cells exhibit notable impacts on cationic drug actions. The fluorescent ASP+ -based in vitro model may also provide a rapid and powerful analytical tool for further screening of cationic drug actions and interactions with hOCTs, particularly hOCT1 and hOCT3 in hepatocellular carcinoma.

Soluble (pro)renin receptor regulation of ENaC involved in aldosterone signaling in cultured collecting duct cells.

We have previously shown that activation of (pro)renin receptor (PRR) induces epithelial Na+ channel (ENaC) activity in cultured collecting duct cells. Here, we examined the role of soluble PRR (sPRR), the cleavage product of PRR in ENaC regulation, and further tested its relevance to aldosterone signaling. In cultured mpkCCD cells, administration of recombinant histidine-tagged sPRR (sPRR-His) at 10 nM within minutes induced a significant and transient increase in the amiloride-sensitive short-circuit current as assessed using the Ussing chamber technique. The acute ENaC activation was blocked by the NADPH oxidase 1/4 inhibitor GKT137892 and siRNA against Nox4 but not the ß-catenin inhibitor ICG-001. In primary rat inner medullary collecting duct cells, administration of sPRR-His at 10 nM for 24 h induced protein expression of the α-subunit but not ß- or γ-subunits of ENaC, in parallel with upregulation of mRNA expression as well as promoter activity of the α-subunit. The transcriptional activation of α-ENaC was dependent on ß-catenin signaling. Consistent results obtained by epithelial volt ohmmeter measurement of equivalent current and Ussing chamber determination of short-circuit current showed that aldosterone-induced transepithelial Na+ transport was inhibited by the PRR decoy inhibitor PRO20 and PF-429242, an inhibitor of sPRR-generating enzyme site-1 protease, and the response was restored by the addition of sPRR-His. Medium sPRR was elevated by aldosterone and inhibited by PF-429242. Taken together, these results demonstrate that sPRR induces two phases of ENaC activation via distinct mechanisms and functions as a mediator of the natriferic action of aldosterone.