Nathan J Cherrington

Deltamethrin [(S)-alpha-cyano-3-phenoxybenzyl-cis-(1 R,3R)-3(2,2-dibromovinyl)(2,2-dimethyl-cyclopropane-carboxylate] and permethrin [3-phenoxybenzyl(1RS)-cis,trans-3-(2,2-dichlorovinyl)-2,2-dimethyl-cyclopropanecarboxylate] are pyrethroid insecticides used in agriculture, public health and military deployments. Pyrethroids are known to be capable of inducing cytochrome P450 (CYP) 2B1/2B2, CYP1A1 and overall CYP content in rat liver. The objectives of this study were to evaluate the potential of deltamethrin and permethrin to cause cytotoxicity and to induce CYP isoforms in human hepatocytes. Permethrin and deltamethrin showed dose-dependent effects on adenylate kinase activity in HepG2 cells, in which 50 and 100 microM doses, respectively, induced a 3-5 fold increase in activity, and also induced adenylate kinase activity in primary human hepatocytes. An approximately 3-fold induction was noted at 200 microM deltamethrin and a 4-fold induction at 100 microM permethrin. Cytotoxicity was noted in HepG2 cells following 48-72 h exposure to 100 or 200 microM deltamethrin and permethrin, respectively. Dose-dependent induction of caspase-3/7 was initiated by 12.5 microM deltamethrin or by 3.125 microM permethrin. Actinomycin D, a positive control for induction of caspase 3/7, induced caspase-3/7, an effect completely abrogated by the specific inhibitor Z-DEVD-FMK. At 100 microM deltamethrin 2-3 fold induction of CYP1A1 and CYP2B6 mRNA was observed, while at the same time an approximately 25-fold induction of CYP3A4 was noted. Permethrin-mediated CYP induction was much less potent, 4-fold or less for CYP1A1, CYP3A4, CYP3A5, CYP2B6 and CYP2A6. It has also been shown that these pyrethroids are ligands for the pregnane X receptor (PXR).

It is hypothesized that during cholestasis, the liver, kidney, and intestine alter gene expression to prevent BA accumulation; enhance urinary excretion of BA; and decrease BA absorption, respectively. To test this hypothesis, mice were subjected to either sham or bile-duct ligation (BDL) surgery and liver, kidney, duodenum, ileum, and serum samples were collected at 1, 3, 7, and 14 days after surgery. Serum total BA concentrations were 1-5 mumol/l in sham-operated mice and were elevated at 1, 3, 7, and 14 days after BDL, respectively. BDL decreased liver Ntcp, Oatp1a1, 1a5, and 1b2 mRNA expression and increased Bsep, Oatp1a4, and Mrp1-5 mRNA levels. In kidney, BDL decreased Oatp1a1 and increased Mrp1-5 mRNA levels. In intestine, BDL increased Mrp3 and Ibat mRNA levels in ileum. BDL increased Mrp1, 3, 4, and 5 protein expression in mouse liver. These data indicate that the compensatory regulation of transporters in liver, kidney, and intestine is unable to fully compensate for the loss of hepatic BA excretion because serum BA concentration remained elevated after 14 days of BDL. Additionally, hepatic and renal Oatp and Mrp genes are regulated similarly during extrahepatic cholestasis, and may suggest that transporter expression is regulated not to remove bile constituents from the body, but instead to remove bile constituents from tissues.

Nonalcoholic fatty liver disease (NAFLD) is categorized into nonalcoholic fatty liver (NAFL) and nonalcoholic steatohepatitis (NASH) and has emerged as a risk factor for more critical clinical conditions. However, the underlying mechanisms of NAFLD pathogenesis are not fully understood. In this study, expression of proteins associated with endoplasmic reticulum (ER) stress, apoptosis and autophagy were analyzed in normal, NAFL and NASH human livers by western blotting. Levels of some ER stress-transducing transcription factors, including cleaved activating transcription factor 6, were higher in NASH than in the normal tissues. However, the expression of a majority of the ER chaperones and foldases analyzed, including glucose-regulated protein 78 and ER protein 44, was lower in NASH than in the normal tissues. Levels of apoptosis markers, such as cleaved poly (ADP-ribose) polymerase, were also lower in NASH tissues, in which expression of some B-cell lymphoma-2 family proteins was up- or down-regulated compared to the normal tissues. The level of the autophagy substrate p62 was not different in NASH and normal tissues, although some autophagy regulators were up- or down-regulated in the NASH tissues compared to the normal tissues. Levels of most of the proteins analyzed in NAFL tissues were either similar to those in one of the other two types, NASH and normal, or were somewhere in between. Together, these findings suggest that regulation of certain important tissues processes involved in protein quality control and cell survival were broadly compromised in the NAFLD tissues.

PMID: 12151624;Abstract:

The intestinal absorption of cadmium (Cd) increases when the body iron (Fe) stores are depleted. The depletion of Fe upregulates the expression of divalent metal transporter 1 (DMT1), which is located at the apical membrane of enterocytes lining the small intestine. DMT1 has been shown to transport Fe and other divalent metal ions in vitro. However, it is not known whether DMT1 mediates the intestinal absorption of Cd. To investigate DMT1 involvement in Cd absorption, rats were fed a diet for 4 weeks either deficient in Fe (FeD diet, 2-6 mg Fe/kg) or supplemented with Fe (FeS diet, 120 mg Fe/kg), followed by a single oral administration of ¹⁰⁹CdCl2. Body Fe status, hemoglobin, and tissue Cd concentration were determined at 48 h after Cd administration. Also, DMT1 mRNA levels were quantified in duodenum, kidney, and liver by the branched DNA signal amplification method. Animals fed the FeD diet exhibited a reduced body weight gain, depletion of body Fe, and Fe deficiency anemia. Tissue Cd concentration was significantly higher in FeD than in FeS diet-fed rats, especially in the duodenum. The amount of Cd retained in the body was 10-fold higher in rats fed the FeD diet than in those fed the FeS diet. DMT1 mRNA was highly expressed in duodenum and was 15-fold higher in the FeD diet group. The levels of DMT1 mRNA were significantly lower in kidney and liver than in duodenum, but were 30 and 40% higher, respectively, in rats fed the FeD diet than in rats fed the FeS diet. These findings suggest that functional DMT1 protein is likely upregulated in the small intestine at the mRNA level by body iron depletion and increases Cd uptake from the gastrointestinal tract with subsequent transfer of Cd to the circulation and body tissues. Furthermore, the data from this study may indicate that DMT1 is a nonspecific metal transporter, which can transport not only Fe, but probably the toxic metal as well.

Alterations in transporter expression may represent a compensatory mechanism of damaged hepatocytes to reduce accumulation of potentially toxic compounds. The present study was conducted to investigate the expression of hepatobiliary efflux transporters in livers from patients after toxic acetaminophen (APAP) ingestion, with livers from patients with primary biliary cirrhosis (PBC) serving as positive controls. mRNA and protein expression of multidrug resistance-associated protein (MRP) 1-6, multidrug resistance protein (MDR) 1-3/P-glycoprotein (P-gp), and breast cancer resistance protein (BCRP) in normal (n = 6), APAP overdose (n = 5), and PBC (n = 6) human liver samples were determined by branched DNA and Western blot analysis, respectively. Double immunohistochemical staining of P-gp and proliferating cell nuclear antigen (PCNA), a marker of proliferation, was performed on paraffin-embedded tissue sections. Compared with normal liver specimens, MRP1 and MRP4 mRNA levels were elevated after APAP overdose and in PBC. Up-regulation of MRP5, MDR1, and BCRP mRNA occurred in PBC livers. Protein levels of MRP4, MRP5, BCRP, and P-gp were increased in both disease states, with MRP1 and MRP3 protein also being induced in PBC. Increased P-gp protein was confirmed immunohistochemically and was found to localize to areas of PCNA-positive hepatocytes, which were detected in APAP overdose and PBC livers. The findings from this study demonstrate that hepatic efflux transporter expression is up-regulated in cases of APAP-induced liver failure and PBC. This adaptation may aid in reducing retention of byproducts of cellular injury and bile constituents within hepatocytes. The close proximity of P-gp and PCNA-positive hepatocytes during liver injury suggests that along with cell regeneration, increased efflux transporter expression is a critical response to hepatic damage to protect the liver from additional insult.