David W. Bombick

TCDD (2,3,7,8-tetrachlorodibenzo-P-dioxin) causes increases in protein kinases particularly protein kinase C in the hepatic plasma membrane of the rat and the guinea pig

To study the cause of TCDD-evoked changes in the functions of plasma membrane constituents TCDDs effects on protein kinase activities in the liver of rats and guinea pigs were investigated. TCDD was found to cause a sharp increase in both c-AMP independent and dependent protein kinase activities in plasma membrane preparations from rat liver within 48 hours from the time of administration. Such effects reached maxima around day 20, and were quite noticeable even 40 days after a single administration of TCDD. As a result of SDS-polyacrylamide gel-electrophoresis (SDS-PAGE) analysis several substrate proteins for these increased protein kinases were observed. Among them are 170 K - 150 K bands, representing EGF receptor protein. TCDD was found to particularly stimulate protein kinase C which is known to influence many enzyme and receptor functions through protein phosphorylation. The possible significance of such an action of TCDD is discussed.

Alteration of rat hepatic plasma membrane functions by 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD)

In vivo administration of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to rats produced significant alterations in Na-K, Ca, Mg ATPase and protein kinase activity, and receptor binding activity to insulin, concanavalin A, glucagon and epidermal growth factor in the canaliculi-rich plasma membrane fraction of the hepatocytes. Some of the changes are dose-related, and generally follow the time course of the sign of TCDD poisoning in the rat (e.g., the loss in body weight). Such TCDD-induced changes in membrane proteins and subsequent alteration in membrane functions appear to be extensive.

Rabbit serum Hypertriglyceridemia after administration of 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin (TCDD)

TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) caused a dose-dependent decrease of adipose tissue lipoprotein lipase (LPL) activity and caused a concomitant increase in serum triglyceride concentration in the rabbit 10 d after single ip administration of either 1 or 50 micrograms/kg. Hepatic low-density lipoprotein (LDL) binding was markedly depressed and serum cholesterol concentrations were modestly increased relative to pair-fed control animals. Serum glucose concentrations were significantly lower in the rabbit administered TCDD compared to ad libitum or pair-fed control animals, although little change was observed in serum insulin concentration. Electron microscopic examination of aortic arches 20 d after a single ip administration of 50 micrograms TCDD/kg revealed ruffling, denudation, and sloughing off of the cell surface and the appearance of macrophage-like structures in the intima and media of the endothelial cells. These alterations resemble preatherosclerotic lesions typical in animals with hyperlipidemia. It is proposed that TCDD causes hyperlipidemia in the rabbit through suppression of LPL activity and LDL receptor binding.

TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin) causes reduction in the low density lipoprotein (LDL) receptor activities in the hepatic plasma membrane of the guinea pig and rat.

Administration of 1 micrograms/kg (single intraperitoneal injection and studied after 10 days) of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) to young male guinea pigs was found to cause a significant reduction in binding of low-density lipoprotein (LDL) to its receptor on the hepatic plasma membrane. This reduction in LDL binding is not caused by the decrease of food intake by treated animals since pair-fed control animals had significantly higher LDL binding than treated animals. It was also found that primary hepatocytes from treated animals had a reduced ability to internalize LDL than controls. Such a change in the plasma membrane function may explain the resulting hyperlipidemia particularly hypercholesterolemia which occurs in this species as a result of TCDD administration.

TCDD (2, 3, 7, 8-Tetrachlorodibenzo-p-dioxin) causes an increase in protein tyrosine kinase activities at an early stage of poisoning in vivo in rat hepatocyte membranes

TCDD was found to cause a significant rise in protein tyrosine kinase levels at an early stage of poisoning in rat liver membrane preparations. The results of sephadex G-150 column chromatographic analysis on rat hepatocyte membranes indicate that there are at least three tyrosine kinases of which activities increase as a consequence of TCDD treatment in vivo. The TCDD-evoked rise in such protein tyrosine kinase activity precedes the down-regulation of the EGF (epidermal growth factor) receptor in the plasma membrane in vivo.

Stimulation of c-ras expression by 2,3,7,8-tetrachlorodibenzo-p-dioxin

A series of in vivo and in vitro experiments were conducted to determine the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) administered on the expression of c-ras. Differences in c-ras expression between control and TCDD treated groups were determined by immunoassay of p21ras protein, or indirectly measured by the specific binding of 3H-GTP to hepatic plasma membrane preparations. Intraperitoneal injection of sublethal doses of TCDD significantly elevated (P<0.05, Student t test) levels of hepatic p21ras protein in Sprague-Dawley rats and TCDD sensitive C57BL/6J mice. Such an increase occurred at an early stage of poisoning in the C57BL/6J mice. The earliest increase was detectable 6 h after dosing, and the difference became statistically significant by 12 and 24 h after dosing. TCDD treatment increased levels of a 21Kd protein found in the in vitro translation products of RNA purified from guinea pig liver. This protein was identified as a c-ras protein based upon its ability to bind GTP, precipitation by a polyclonal antibody against the rasHa and Ki proteins and subsequent SDS-PAGE which showed a single protein band of ≈21Kd.