Disruption of renal tubular mitochondrial quality control by Myo-inositol oxygenase in diabetic kidney disease.

Diabetic kidney disease (DKD) is associated with oxidative stress and mitochondrial injury. Myo-inositol oxygenase (MIOX), a tubular-specific enzyme, modulates redox imbalance and apoptosis in tubular cells in diabetes, but these mechanisms remain unclear. We investigated the role of MIOX in perturbation of mitochondrial quality control, including mitochondrial dynamics and autophagy/mitophagy, under high-glucose (HG) ambience or a diabetic state. HK-2 or LLC-PK1 cells subjected to HG exhibited an upregulation of MIOX accompanied by mitochondrial fragmentation and depolarization, inhibition of autophagy/mitophagy, and altered expression of mitochondrial dynamic and mitophagic proteins. Furthermore, dysfunctional mitochondria accumulated in the cytoplasm, which coincided with increased reactive oxygen species generation, Bax activation, cytochrome C release, and apoptosis. Overexpression of MIOX in LLC-PK1 cells enhanced the effects of HG, whereas MIOX siRNA or d-glucarate, an inhibitor of MIOX, partially reversed these perturbations. Moreover, decreasing the expression of MIOX under HG ambience increased PTEN-induced putative kinase 1 expression and the dependent mitofusin-2-Parkin interaction. In tubules of diabetic mice, increased MIOX expression and mitochondrial fragmentation and defective autophagy were observed. Dietary supplementation of d-glucarate in diabetic mice decreased MIOX expression, attenuated tubular damage, and improved renal functions. Notably, d-glucarate administration also partially attenuated mitochondrial fragmentation, oxidative stress, and apoptosis and restored autophagy/mitophagy in the tubular cells of these mice. These results suggest a novel mechanism linking MIOX to impaired mitochondrial quality control during tubular injury in the pathogenesis of DKD and suggest d-glucarate as a potential therapeutic agent for the amelioration of DKD.

Inhibition of P-glycoprotein-mediated transport by terpenoids contained in herbal medicines and natural products.

Terpenoids form a large and structurally diverse family of natural products and are ingredients of various herbal medicines. We have investigated possible interactions between herbal medicines and conventional medicines, and recently reported that monoterpenoids contained in Zanthoxyli Fructus can be potent inhibitors of P-glycoprotein (P-gp). In the present study, the influence of 70 kinds of terpenoids present in natural products on P-gp-mediated efflux transport was investigated. LLC-GA5-COL150 cells transfected with human MDR1 cDNA encoding P-gp were used to screen the terpenoids. Large increases in the intracellular accumulation of [(3)H]digoxin were observed in the presence of (R)-(+)-citronellal, (S)-(-)-beta-citronellol, alpha-terpinene, terpinolene, (-)-beta-pinene, abietic acid, ophiobolin A, cucurbitacin I, and glycyrrhetic acid. A study of the concentration-dependency revealed that the IC(50) of ophiobolin A, glycyrrhetic acid, (R)-(+)-citronellal, abietic acid, and cucurbitacin I was smaller than that of verapamil. The transcellular transport of [(3)H]digoxin across Caco-2 cell monolayers was then examined in the presence of (R)-(+)-citronellal, abietic acid, and glycyrrhetic acid. Significant increases in the apical-to-basolateral transport and decreases in the basolateral-to-apical transport and efflux ratio were demonstrated. These findings suggest that some natural products containing these terpenoids may inhibit P-gp-mediated transport and interact with P-gp substrates in the intestinal absorption process.

New acylated clionasterol glycosides from Valeriana officinalis.

The chloroform extract of Valeriana officinalis led to the isolation of clionasterol-3-O-beta-D-glucopyranoside and a mixture of 6'-O-acyl-beta-D-glucosyl-clionasterols. The acyl moieties were identified as hexadecanoyl, 8 E,11 E-octadecadienoyl and 14-methylpentadecanoyl by alkaline hydrolysis followed by GC-MS analysis. The isolated compounds did not exhibit any anti-inflammatory, anticancer or cytotoxic activity when tested in a variety of in vitro cell-based assays.

Antioxidative activity of furanocoumarins isolated from Angelicae dahuricae.

The methylene chloride extract of the root of Angelicae dahuricae showed high protective activity against 2,2'-azobis (2-aminodinopropane) dihydrochloride (AAPH)-induced cellular damage. From this extract, 11 furanocoumarins were isolated, namely oxypeucedanin hydrate, 9-hydroxy-4-methoxypsoralen, byakangelicin, pabulenol, alloisoimperatorin, neobyakangelicol, byakangelicol, oxypeucedanin, imperatorin, phellotorin and isoimperatorin, respectively. Among these 11 furanocoumarins, 9-hydroxy-4-methoxypsoralen and alloisoimperatorin displayed potent antioxidant effects against the DPPH radical and against renal epithelial cell injury by using AAPH to generate peroxyl radicals in vitro.

Acquired resistance to rechallenge injury with uranyl acetate in LLC-PK1 cells.

This study was conducted to determine whether cultured renal cells exposed to previous uranium injury would be resistant to subsequent insult, and if so, the mechanisms for this resistance. The addition of a toxic dose of uranyl acetate (UA) (1 X 10(-3) mol/L) for 48 hours to the culture medium significantly enhanced the release of lactate dehydrogenase (LDH) from LLC-PK1 cells, expressed as LDH activity in the medium corrected by protein content of the cells, compared with control conditions (31.5 +/- 3.6 vs 5.5 +/- 0.6 Wroblewski unit/microg protein, p < 0.01). Pretreatment with a toxic dose of UA (1 x 10(-3) mol/L for 24 hours) or heat stress (42 degrees C for 30 minutes) significantly lessened the extent of increase in LDH after exposure to toxic doses of UA for 48 hours (15.2 +/- 1.4 and 7.6 +/- 0.6 Wroblewski unit/microg protein, respectively). Pretreatment with a nontoxic dose of UA (3 x 10(-4) mol/L for 24 hours) had no effect on the release of LDH after a toxic dose of UA treatment (38.6 +/- 7.0 Wroblewski unit/microg protein). Quercetin (100 microm ) and staurosporine (0.1 microg/ml), both known to inhibit the development of thermotolerance, hindered acquisition of the resistance to rechallenge with UA (42.5 +/- 1.1 and 38.9 +/- 1.8 Wroblewski unit/microg protein, respectively). Quercetin did not modify the UA-induced increase in the release of LDH in cells not pretreated with UA or heat stress. It follows from these findings that LLC-PK1 cells previously exposed to a toxic dose of UA are resistant to rechallenged insult and that mechanisms similar to those for thermotolerance might contribute to this acquired resistances.