Oxidative drug metabolism in liver microsomes from uremic rats

Oxidative drug metabolism in liver microsomes from uremic rats. Uremia was achieved by subtotal nephrectomy in 50 to 70 day old male rats weighing 252.0 ± 20.1 g. Nephrectomized rats and sham-operated controls were sacrificed six days later. After liver microsomes had been isolated, microsomal cytochrome P-450 and cytochrome b5 content as well as microsomal specific activity for N-demethylation of aminopyrine, O-demethylation of p-nitroanisole and p-hydroxylation of acetanilide were measured. Serum urea concentration rose to 370.0 ± 80.0 mg/100 ml. The serum creatinine rose to 4.8 ± 1.2 mg/100 ml. In sham-operated controls serum urea and creatinine concentrations were approximately 50 and 0.6 mg/100 ml, respectively. Uremic rats, in contrast to sham-operated controls, lost about 25 % of their initial body wt. In uremic rats and in sham-operated controls with caloric deficiency there was a significant decrease of total microsomal protein to 51 % of the value measured in sham-operated controls fed ad libitum. Absolute and relative liver wet wt decreased more in caloric deficient controls than in uremic rats, whereas microsomal protein content per g liver was lower in uremic rats. In comparison with sham-operated controls, there was a pronounced decrease in the specific activity of liver microsomes from uremic rats for the demethylation of aminopyrine and p-nitroanisole and for the p-hydroxylation of acetanilide. The microsomal cytochrome P-450 content was also significantly diminished, whereas the microsomal cytochrome b5 content was not influenced, but there was stimulation of the O-demethylation of p-nitroanisole to 125.1% when compared to controls fed ad libitum. Four intraperitoneal injections of 6 mg δ-aminolevulinic acid/kg body given to uremic rats normalized the cytochrome P-450 content and significantly stimulated the specific activity for demethylation of p-nitroanisole, whereas demethylation of aminopyrine, p-hydroxylation of acetanilide and cytochrome b5 content were not altered. In sham-operated controls none of the measured parameters were influenced by δ-aminolevulinic acid pretreatment. It is assumed that the reduction of cytochrome P-450 content in liver microsomes from uremic rats is caused by a deficiency of δ-aminolevulinic acid which leads to disturbances in cytochrome synthesis. The observed decrease in the ability of liver microsomes from uremic rats to metabolize aminopyrine, p-nitroanisole and acetanilide is not merely due to reduced cytochrome P-450 content

Closing in on the link between apoptosis and autophagy

While there is a clear connection between apoptosis and autophagy, the mechanisms that regulate the interaction have been difficult to identify. The initial clue to the link was the observation that Bcl-2 was located at the endoplasmic reticulum (ER), where it could prevent some forms of apoptosis and also bind to the autophagy regulatory protein Beclin-1. However, both of these enigmatic observations have been united with the discovery of the nutrient-deprivation autophagy factor-1 (NAF-1) protein. As an ER-localized protein that enhances the interaction of Bcl-2 and Beclin-1 and that also binds to the pro-apoptotic protein Bik, NAF-1 is perfectly placed to be a central regulator of the switch between autophagy and apoptosis

BH3-only proteins: Orchestrators of apoptosis

AbstractThe BH3-only proteins of Bcl-2 family are essential initiators of apoptosis that propagate extrinsic and intrinsic cell death signals. The interaction of BH3-only proteins with other Bcl-2 family members is critical for understanding the core machinery that controls commitment to apoptosis by permeabilizing the mitochondrial outer membrane. BH3-only proteins promote apoptosis by both directly activating Bax and Bak and by suppressing the anti-apoptotic proteins at the mitochondria and the endoplasmic reticulum. To prevent constitutive cell death, BH3-only proteins are regulated by a variety of mechanisms including transcription and post-translational modifications that govern specific protein–protein interactions. Furthermore, BH3-only proteins also control the initiation of autophagy, another important pathway regulating cell survival and death. Emerging evidence indicates that the interaction of BH3-only proteins with membranes regulates binding to other Bcl-2 family members, thereby specifying function. Due to the important role of BH3-only proteins in the regulation of cell death, several promising BH3-mimetic drugs that are active in pre-clinical models are currently being tested as anti-cancer agents. This article is part of a Special Issue entitled Mitochondria: the deadly organelle

Previous attentional set can induce an attentional blink with task-irrelevant initial targets

Identification of a second target is often impaired by the requirement to process a prior target in a rapid serial visual presentation (RSVP). This is termed the attentional blink. Even when the first target is task-irrelevant an attentional blink may occur providing this first target shares similar features with the second target (contingent capture). An RSVP experiment was undertaken to assess whether this first target can still cause an attentional blink when it did not require a response and did not share any features with the following target. The results revealed that such task-irrelevant targets can induce an attentional blink providing that they were task-relevant on a previous block of trials. This suggests that irrelevant focal stimuli can distract attention on the basis of a previous attentional set

Bcl-XL is qualitatively different from and ten times more effective than Bcl-2 when expressed in a breast cancer cell line

BACKGROUND: Bcl-2 and Bcl-XL are anti-apoptotic paralogues that inhibit apoptosis elicited by a wide variety of stimuli, and play critical roles in cancer development and resistance to treatment. Many clinical studies have indicated that expression of these anti-apoptotic proteins in tumours is associated with poor prognosis. It has therefore been assumed that in cells the essential difference between Bcl-2 and Bcl-XL involves regulation of expression and that they are otherwise functionally similar. To examine this issue, we have compared the function of the proteins and of mutants of Bcl-2 and Bcl-XL specifically targeted to different subcellular sites. METHODS: We generated clones of the human breast cancer line MCF-7 stably expressing known amounts of Bcl-2, or Bcl-XL as determined by quantitative immunoblotting. Clones expressing equivalent amounts of wild-type and mutants of Bcl-2 and Bcl-XL with subcellular localization restricted to the cytoplasm, endoplasmic reticulum or outer mitochondrial membrane were studied in both MCF-7 and Rat-1 fibroblasts. In MCF-7 cells we measured the functional activities of these proteins in preventing apoptosis induced by four different agents (doxorubicin, ceramide, thapsigargin, TNF-α). Etoposide and low serum were used to compare the effect of Bcl-2, Bcl-XL and mutants located at the endoplasmic reticulum on induction of apoptosis in fibroblasts. RESULTS: We noted both qualitative and quantitative differences in the functional activity of these two anti-apoptotic proteins in cells: Bcl-2 localized to the endoplasmic reticulum inhibits apoptosis induced by ceramide and thapsigargin but not by doxorubicin or TNFα, while Bcl-XL at the endoplasmic reticulum is active against all four drugs. In fibroblasts Bcl-2 localized to the ER did not prevent cell death due to etoposide whereas Bcl-XL in the same location did. Finally in MCF-7 cells, Bcl-XL is approximately ten times more active than Bcl-2 in repressing apoptosis induced by doxorubicin. This difference can be manifest as a large difference in clonal survival. CONCLUSION: When examined in the same cellular context, Bcl-2 and Bcl-XL differ substantially in the potency with which they inhibit apoptosis, mediated in part by differences in the inhibition of specific subcellular pathways

Direct costs and cost-effectiveness of dual-source computed tomography and invasive coronary angiography in patients with an intermediate pretest likelihood for coronary artery disease

The study aims to determine the direct costs and comparative cost-effectiveness of latest-generation dual-source computed tomography (DSCT) and invasive coronary angiography for diagnosing coronary artery disease (CAD) in patients suspected of having this disease

Bcl-XL Inhibits Membrane Permeabilization by Competing with Bax

Although Bcl-XL and Bax are structurally similar, activated Bax forms large oligomers that permeabilize the outer mitochondrial membrane, thereby committing cells to apoptosis, whereas Bcl-XL inhibits this process. Two different models of Bcl-XL function have been proposed. In one, Bcl-XL binds to an activator, thereby preventing Bax activation. In the other, Bcl-XL binds directly to activated Bax. It has been difficult to sort out which interaction is important in cells, as all three proteins are present simultaneously. We examined the mechanism of Bax activation by tBid and its inhibition by Bcl-XL using full-length recombinant proteins and measuring permeabilization of liposomes and mitochondria in vitro. Our results demonstrate that Bcl-XL and Bax are functionally similar. Neither protein bound to membranes alone. However, the addition of tBid recruited molar excesses of either protein to membranes, indicating that tBid activates both pro- and antiapoptotic members of the Bcl-2 family. Bcl-XL competes with Bax for the activation of soluble, monomeric Bax through interaction with membranes, tBid, or t-Bid-activated Bax, thereby inhibiting Bax binding to membranes, oligomerization, and membrane permeabilization. Experiments in which individual interactions were abolished by mutagenesis indicate that both Bcl-XL–tBid and Bcl-XL–Bax binding contribute to the antiapoptotic function of Bcl-XL. By out-competing Bax for the interactions leading to membrane permeabilization, Bcl-XL ties up both tBid and Bax in nonproductive interactions and inhibits Bax binding to membranes. We propose that because Bcl-XL does not oligomerize it functions like a dominant-negative Bax in the membrane permeabilization process