Palladium-103 plaque radiation therapy for macular degeneration: results of a 7 year study.

AIM: To report 7 year results of ophthalmic plaque radiotherapy for exudative macular degeneration. METHODS: In a phase I clinical trial, 30 patients (31 eyes) were treated with ophthalmic plaque irradiation for subfoveal exudative macular degeneration. Radiation was delivered to a mean 2 mm from the inner sclera (range 1.2-2.4) prescription point calculated along the central axis of the plaque. The mean prescription dose was 17.62 Gy (range 12.5-24) delivered over 34 hours (range 18-65). Early Treatment Diabetic Retinopathy Study (ETDRS) type standardised visual acuity determinations, ophthalmic examinations, and angiography were performed before and after treatment. Clinical evaluations were performed in a non-randomised and unmasked fashion. RESULTS: At 33.3 months (range 3-4), 17 of 31 (55%) eyes had lost 3 or more lines of vision on the ETDRS chart, five (16%) had improved 3 or more lines, and the remaining nine (29%) were within 2 lines of their pretreatment visual acuity measurement. Overall, 45% of patients were within or improved more than 2 lines of their initial visual acuity. Five eyes developed macular scars, eight developed subsequent neovascularisation or haemorrhage, and three progressed through therapy. Two patients were lost to follow up. The most common finding of patients followed for 6 or more months (n=18 of 29 (62%)) was regression or stabilisation of the exudative process. No radiation retinopathy, optic neuropathy, or cataracts could be attributed to irradiation. CONCLUSION: Ophthalmic plaque radiation can be used to treat exudative macular degeneration. At the dose and dose rates employed, most patients experienced decreased exudation or stabilisation of their maculas. No sight limiting radiation complications were noted during 7 year follow up. Owing to the variable natural course of this disease, a prospective randomised clinical trial should be performed to evaluate the efficacy of plaque radiation therapy for exudative macular degeneration.

Toxicity of alpidem, a peripheral benzodiazepine receptor ligand, but not zolpidem, in rat hepatocytes: role of mitochondrial permeability transition and metabolic activation.

Whereas alpidem is hepatotoxic, zolpidem is not. Despite closely related chemical structures, alpidem, but not zolpidem, is a peripheral benzodiazepine receptor (PBR) ligand, and is also more lipophilic than zolpidem. We compared their effects in isolated rat liver mitochondria and rat hepatocytes. Zolpidem did not affect calcium-induced mitochondrial permeability transition (MPT) in mitochondria, caused little glutathione depletion in hepatocytes, and was not toxic, even at 500 microM. At 250 to 500 microM, alpidem prevented calcium-induced MPT in isolated mitochondria, but caused severe glutathione depletion in hepatocytes that was increased by 3-methylcholanthrene, a cytochrome P4501A inducer, and decreased by cystine, a glutathione precursor. Although cell calcium increased, mitochondrial cytochrome c did not translocate to the cytosol and cells died of necrosis. Cell death was prevented by cystine, but not cyclosporin A, an MPT inhibitor. At low concentrations (25-50 microM), in contrast, alpidem accelerated calcium-induced MPT in mitochondria. It did not deplete glutathione in hepatocytes, but nevertheless caused some cell death that was prevented by cyclosporin A, but not by cystine. Alpidem (10 microM) also increased the toxicity of tumor necrosis factor-alpha (1 ng/ml) in hepatocytes. In conclusion, low concentrations of alpidem increase both calcium-induced MPT in mitochondria, and TNF-alpha toxicity in cells, like other PBR ligands. Like other lipophilic protonatable amines, however, alpidem inhibits calcium-induced MPT at high concentrations. At these high concentrations, toxicity involves cytochrome P4501A-mediated metabolic activation, glutathione depletion, and increased cell calcium, without MPT involvement. In contrast, zolpidem has no mitochondrial effects, causes little glutathione depletion, and is not toxic.

Mechanisms for experimental buprenorphine hepatotoxicity: major role of mitochondrial dysfunction versus metabolic activation.

BACKGROUND/AIMS: Although sublingual buprenorphine is safely used as a substitution drug in heroin addicts, large overdoses or intravenous misuse may cause hepatitis. Buprenorphine is N-dealkylated to norbuprenorphine by CYP3A. METHODS: We investigated the mitochondrial effects and metabolic activation of buprenorphine in isolated rat liver mitochondria and microsomes, and its toxicity in isolated rat hepatocytes and treated mice. RESULTS: Whereas norbuprenorphine had few mitochondrial effects, buprenorphine (25-200 microM) concentrated in mitochondria, collapsed the membrane potential, inhibited beta-oxidation, and both uncoupled and inhibited respiration in rat liver mitochondria. Both buprenorphine and norbuprenorphine (200 microM) underwent CYP3A-mediated covalent binding to rat liver microsomal proteins and both caused moderate glutathione depletion and increased cell calcium in isolated rat hepatocytes, but only buprenorphine also depleted cell adenosine triphosphate (ATP) and caused necrotic cell death. Four hours after buprenorphine administration to mice (100 nmol/g body weight), hepatic glutathione was unchanged, while ATP was decreased and serum transaminase increased. This transaminase increase was attenuated by a CYP3A inducer and aggravated by a CYP3A inhibitor. CONCLUSIONS: Both buprenorphine and norbuprenorphine undergo metabolic activation, but only buprenorphine impairs mitochondrial respiration and ATP formation. The hepatotoxicity of high concentrations or doses of buprenorphine is mainly related to its mitochondrial effects.

Hepatitis after intravenous buprenorphine misuse in heroin addicts.

BACKGROUND: Sublingual buprenorphine is used as a substitution drug in heroin addicts. Although buprenorphine inhibits mitochondrial function at high concentrations in experimental animals, these effects should not occur after therapeutic sublingual doses, which give very low plasma concentrations. CASE REPORTS: We report four cases of former heroin addicts infected with hepatitis C virus and placed on substitution therapy with buprenorphine. These patients exhibited a marked increase in serum alanine amino transferase (30-, 37-, 13- and 50-times the upper limit of normal, respectively) after injecting buprenorphine intravenously and three of them also became jaundiced. Interruption of buprenorphine injections was associated with prompt recovery, even though two of these patients continued buprenorphine by the sublingual route. A fifth patient carrying the hepatitis C and human immunodeficiency viruses, developed jaundice and asterixis with panlobular liver necrosis and microvesicular steatosis after using sublingual buprenorphine and small doses of paracetamol and aspirin. CONCLUSIONS: Although buprenorphine hepatitis is most uncommon even after intravenous misuse, addicts placed on buprenorphine substitution should be repeatedly warned not to use it intravenously. Higher drug concentrations could trigger hepatitis in a few intravenous users, possibly those whose mitochondrial function is already impaired by viral infections and other factors.

Urinary bladder hyporeflexia and reduced pain-related behaviour in P2X3-deficient mice.

Extracellular ATP is implicated in numerous sensory processes ranging from the response to pain to the regulation of motility in visceral organs. The ATP receptor P2X3 is selectively expressed on small diameter sensory neurons, supporting this hypothesis. Here we show that mice deficient in P2X3 lose the rapidly desensitizing ATP-induced currents in dorsal root ganglion neurons. P2X3 deficiency also causes a reduction in the sustained ATP-induced currents in nodose ganglion neurons. P2X3-null mice have reduced pain-related behaviour in response to injection of ATP and formalin. Significantly, P2X3-null mice exhibit a marked urinary bladder hyporeflexia, characterized by decreased voiding frequency and increased bladder capacity, but normal bladder pressures. Immunohistochemical studies localize P2X3 to nerve fibres innervating the urinary bladder of wild-type mice, and show that loss of P2X3 does not alter sensory neuron innervation density. Thus, P2X3 is critical for peripheral pain responses and afferent pathways controlling urinary bladder volume reflexes. Antagonists to P2X3 may therefore have therapeutic potential in the treatment of disorders of urine storage and voiding such as overactive bladder.

Recurrent malignant chondroid syringoma of the foot: a case report and review of the literature.

Malignant chondroid syringoma, or mixed tumor of the skin, salivary gland type, is an uncommon neoplasm believed to originate in sweat glands. This neoplasm occurs mostly in women and is typically seen in the extremities and torso. A case of recurrent malignant chondroid syringoma of the right foot in a man aged 34 years is described with a review of pertinent literature. The surgically excised neoplasm was evaluated by routine histology, immunohistochemistry, and transmission electron microscopy. The malignant chondroid syringoma showed microscopic dermal satellite tumor nodules. Immunohistochemical staining was positive for keratin and S100 and negative for actin and p53. Ki-67 showed <10% positive staining. Ultrastructurally, the neoplasm was composed of epithelial cells with tonofilaments, cell junctions, and electron-dense amorphous keratin-like substance in the intercellular spaces. No evidence of myoepithelial differentiation was noted. Given the tumoral size, acral location, and histologic findings, the neoplasm was classified as a malignant chondroid syringoma. After reviewing the literature, it became apparent that wide surgical excision, adjuvant radiation therapy as well as patient education are critical in facilitating long-term survival.

Opening of the mitochondrial permeability transition pore causes matrix expansion and outer membrane rupture in Fas-mediated hepatic apoptosis in mice.

Although Fas stimulation has been reported to cause outer mitochondrial membrane rupture in Jurkat cells, the mechanism of this effect is debated, and it is not known if outer membrane rupture also occurs in hepatocyte mitochondria. We studied the in vivo effects of Fas stimulation on ultrastructural lesions and mitochondrial function in mice. Four hours after administration of an agonistic anti-Fas antibody (8 microg/animal), caspase activity increased 5.4-fold. Nuclear DNA showed internucleosomal fragmentation, whereas supercoiled mitochondrial DNA was replaced by circular and linear forms. Mitochondrial cytochrome c was partly released into the cytosol. Ultrastructurally, mitochondrial lesions were observed in both apoptotic hepatocytes (with nuclear chromatin condensation/fragmentation) and nonapoptotic hepatocytes (without nuclear changes). In nonapoptotic cells, outer mitochondrial membrane rupture allowed herniation of the inner membrane and matrix through the outer membrane gap. In apoptotic hepatocytes, the matrix became electron-lucent and no longer protruded through the outer membrane gap. Mitochondria clustered around the nucleus, whereas rough endoplasmic reticulum cisternae became peripheral. In liver mitochondria isolated after Fas stimulation, the membrane potential decreased, whereas basal respiration increased. Pretreatment with either z-VAD-fmk (an inhibitor of caspases) or cyclosporin A (a permeability transition inhibitor) totally or mostly prevented mitochondrial outer membrane rupture, membrane potential decrease, cytochrome c release, and apoptosis. In conclusion, in vivo Fas stimulation causes caspase activation, mitochondrial permeability transition (decreasing the membrane potential and increasing basal respiration), mitochondrial matrix expansion (as shown by matrix herniation), outer mitochondrial membrane rupture, and cytochrome c release.

Diode-light transillumination for ophthalmic plaque localization around juxtapapillary choroidal melanomas.

PURPOSE: An evaluation of plaque-mounted diode-light transillumination (DLT) for localization of episcleral plaques beneath juxtapapillary tumors. METHODS AND MATERIALS: Two patients scheduled for radiotherapy for juxtapapillary melanomas were offered DLT as an additional method of ophthalmic plaque localization. Plaques were constructed by affixing 4 non-heat producing, light-emitting diodes with their apertures flush with the episcleral outer surface of the plaque's rim. Bio-implantable epoxy was used to encapsulate the electronic components. Then the plaques were loaded with 103Pd seeds. After the eye-plaques were sewn to the episclera covering the base of the intraocular tumors; the diode-lights were illuminated, viewed and recorded. Photodocumentation of the relative position of the 4 lights around tumor's base was obtained in both cases. RESULTS: Digital images of plaque-mounted diode retro-transillumination were obtained. No evidence of diode-light toxicity was noted. Both tumors were found to be covered by the ophthalmic plaques. CONCLUSION: Juxtapapillary tumors are often difficult or impossible to visualize with standard transillumination techniques and have been associated with poor local control rates. We have developed plaque-mounted DLT in an effort to improve ophthalmic plaque localization. Retrobulbar transillumination was viewed by indirect ophthalmoscopy and recorded with video-imaging. This technique provides unique photographic documentation of episcleral plaque localization beneath juxtapapillary tumors.

An alcoholic binge causes massive degradation of hepatic mitochondrial DNA in mice.

BACKGROUND & AIMS: Ethanol causes oxidative stress in the hepatic mitochondria of experimental animals and mitochondrial DNA deletions in alcoholics. We postulated that ethanol intoxication may cause mitochondrial DNA strand breaks. METHODS: Effects of an intragastric dose of ethanol (5 g/kg) on hepatic mitochondrial DNA levels, structure, and synthesis were determined by slot blot hybridization, Southern blot hybridization, and in vivo [3H]thymidine incorporation, respectively. RESULTS: Two hours after ethanol administration, ethane exhalation (an index of lipid peroxidation) increased by 133%, although hepatic lipids were unchanged. Mitochondrial DNA was depleted by 51%. Its supercoiled form disappeared, whereas linearized forms increased. Long polymerase chain reaction evidenced lesions blocking polymerase progress on the mitochondrial genome. Mitochondrial transcripts decreased. Subsequently, [3H]thymidine incorporation into mitochondrial DNA increased, and mitochondrial DNA levels were restored. In contrast, nuclear DNA was not fragmented and its [3H]thymidine incorporation was unchanged. Liver ultrastructure only showed inconstant mitochondrial lesions. Ethanol-induced mitochondrial DNA depletion was prevented by 4-methylpyrazole, an inhibitor of ethanol metabolism, and attenuated by melatonin, an antioxidant. CONCLUSIONS: After an alcoholic binge, ethanol metabolism causes oxidative stress and hepatic mitochondrial DNA degradation in mice. DNA strand breaks may be involved in the development of mitochondrial DNA deletions in alcoholics.

Identification and characterization of a myristylated and palmitylated serine/threonine protein kinase.

We report the molecular cloning and initial characterization of a novel fatty acid acylated serine/threonine protein kinase. The putative open reading frame is predicted to encode a 305 amino acid protein possessing a carboxy-terminal protein kinase domain and amino-terminal myristylation and palmitylation sites. The protein kinase has been accordingly denoted as the myristylated and palmitylated serine/threonine protein kinase (MPSK). Human and mouse MPSKs share approximately 93% identity at the amino acid level with complete retention of acylation sites. Radiation hybridization localized the human MPSK gene to chromosome 2q34-37. Northern analysis demonstrated that the human MPSK 1.7 kilobase mRNA is widely distributed. Epitope tagged human MPSK was found to be acylated by myristic acid at glycine residue 2 and by palmitic acid at cysteines 6 and/or 8. Palmitylation of MPSK in these experiments was found to require an intact myristylation site. While epitope tagged MPSK in immune complexes or purified human glutathione S transferase-MPSK was found to autophosphorylate at one or more threonine residues, the enzyme was not found to phosphorylate several other common exogenous substrates. Indeed, only PHAS-I was identified as an exogenous substrate which was found to be phosphorylated on threonine and serine residues.

Interleukin-2 overexpresses c-myc and down-regulates cytochrome P-450 in rat hepatocytes.

The interaction of interleukin-2 (IL-2) with its receptor (IL-2R) decreases cytochrome P-450 (CYP) expression in rat hepatocytes. Because IL-2 increases c-Myc in lymphocytes and because c-myc overexpression represses several genes, we postulated that the IL-2/IL-2R interaction may increase c-Myc and thereby down-regulate CYP in hepatocytes. Cultured rat hepatocytes were exposed for 24 h to IL-2 (350 U/ml) and other agents. IL-2 increased c-myc mRNA and protein but decreased total CYP and the mRNAs and proteins of CYP2C11 and CYP3A. The IL-2-mediated c-myc overexpression and CYP down-regulation were prevented by 1) genistein (a tyrosine kinase inhibitor that blocks the initial transduction of the IL-2R signal), 2) retinoic acid, butyric acid, or dimethyl sulfoxide (three agents that block c-myc transcription), or 3) an antisense c-myc oligonucleotide (which may cause rapid degradation of the c-myc transcript). It is concluded that IL-2 causes the overexpression of c-myc and the down-regulation of CYPs in rat hepatocytes. Block of c-myc overexpression, at three different levels with five different agents, prevents CYP down-regulation, suggesting that c-myc overexpression may directly or indirectly repress CYP in hepatocytes.

Transcriptional analysis of the PTEN/MMAC1 pseudogene, psiPTEN.

PTEN/MMAC1 is a recently characterized tumor suppressor. A pseudogene derived from the human PTEN/MMAC1 phosphatase, psiPTEN, has been reported. Recent analysis of the pseudogene revealed conflicting results about the expression of psiPTEN. In this study, we show that the PTEN/MMAC1 pseudogene is actively transcribed in all cells and tissues examined. In some cases, pseudogene transcripts were found to represent as much as 70% of the total PTEN/MMAC1 RNA. As psiPTEN is transcribed, there is a potential for misinterpretation of PTEN/MMAC1 mutations when RT-PCR techniques are used, as well as potential for a psiPTEN-encoded translation product. Although we were unable to detect a pseudogene protein product in the cell lines examined, a baculovirus produced GST pseudogene fusion protein exhibited phosphatase activity comparable to wild type. The results of this study, taken together, indicate the potential complication of PTEN/MMAC1 molecular analysis caused by the expression of psiPTEN.

Decrease in hepatic cytochrome P450 after interleukin-2 immunotherapy.

Interleukin-2 (IL-2) has been shown to decrease cytochrome P450 (CYP) mRNAs and proteins in cultured rat hepatocytes, and IL-2 administration decreases CYPs in rats. Although high doses of IL-2 are administered to cancer patients, the effect on human CYPs has not yet been determined. Patients with hepatic metastases from colon or rectum carcinomas were randomly allocated to various daily doses of human recombinant IL-2 (from 0 to 12.10(6) units/m(2)). IL-2 was infused from day 7 to day 3 before hepatectomy and the conservation of a non-tumorous liver fragment in liquid nitrogen. Hepatic CYPs and monooxygenase activities were not significantly decreased in 5 patients receiving daily doses of 3 or 6 10(6) IL-2 units/m2, compared to 7 patients who did not receive IL-2. In contrast, in 6 patients receiving daily doses of 9 or 12 x 10(6) IL-2 units/m2, the mean values for immunoreactive CYP1A2, CYP2C, CYP2E1, and CYP3A4 were 37, 45, 60 and 39%, respectively, of those in controls; total CYP was significantly decreased by 34%, methoxyresorufin O-demethylation by 62%, and erythromycin N-demethylation by 50%. These observations suggest that high doses of IL-2 may decrease total CYP and monooxygenase activities in man.

Radiation therapy for age-related macular degeneration.

Neovascular age-related macular degeneration is the most common cause of severe irreversible blindness in the Western world in people older than age 50. Laser photocoagulation is the only proven treatment for this disease; however, fewer than 20% of patients are eligible for this treatment because the majority of choroidal neovascularization membranes are not visible by ophthalmoscopy or angiography. In addition, many patients elect not to undergo this treatment because laser treatment of subfoveal neovascular membranes results in immediate and permanent central visual loss. Several treatments are under investigation, including external-beam radiation therapy. There are multiple publications of early trials using radiation therapy, but to date there is only one randomized published study. This article reviews these trials and summarizes the status of radiation therapy as a treatment for macular degeneration.

Palladium-103 plaque radiotherapy for choroidal melanoma: results of a 7-year study.

OBJECTIVE: To describe the first clinical experience with palladium-103 (103Pd) ophthalmic plaque radiotherapy for choroidal melanoma. DESIGN: Phase-I (nonrandomized) clinical trial. PARTICIPANTS: Eighty patients with uveal melanomas were diagnosed by clinical examination, found to be negative for metastatic disease, and offered 103Pd radioactive plaque treatment. Nine patients were concurrently treated with microwave hyperthermia. INTERVENTION: Palladium-103 ophthalmic plaque radiotherapy was employed for each patient. Eye plaques were sewn to the episclera to cover the base of the intraocular tumor, radiation was continuously delivered over 5 to 7 days, and then the plaques were removed. A mean apical dose of 81 Gy was delivered. MAIN OUTCOME MEASURES: The authors evaluated the ease of use of 103Pd seeds within standard gold eye plaques. Patient-related outcomes were control of tumor growth, change in visual acuity, the development of radiation damage (retinopathy, optic neuropathy, and cataract), and metastatic disease. RESULTS: From September 1990 to December 1997, 80 patients were treated with 103Pd and followed for an average of 38 months. Two patients were lost to follow-up. During this time, the authors found that 103Pd seeds were equivalent to iodine-125 (125I) with respect to plaque manufacture and ease of dosimetric calculations. Two patients in this series were treated for tumor recurrence after 125I plaque radiotherapy. They both failed secondary 103Pd treatment and were enucleated. When 103Pd was used as a primary treatment, it controlled the growth of 75 of 78 tumors (96%). Overall, there have been six enucleations: three failures of primary treatment, two failures of retreatment, and one for neovascular glaucoma. Visual acuity evaluations at the 36-month follow-up visit (including the enucleated patients) revealed that 38% of eyes had decreased 3 or more lines of vision, and 77% were 20/200 or better. CONCLUSION: Palladium-103 plaque radiotherapy can be used to treat uveal melanomas. Compared with 125I, computerized dosimetry suggests a more favorable dose distribution with 103Pd. Treatment of most patients resulted in tumor shrinkage and preservation of functional vision. The authors have noted no complications that might preclude the use of 103Pd ophthalmic plaque radiotherapy for choroidal melanoma.

Ophthalmic plaque radiotherapy for age-related macular degeneration associated with subretinal neovascularization.

PURPOSE: To evaluate ophthalmic plaque radiotherapy for the treatment of subretinal neovascularization associated with age-related macular degeneration. METHODS: In a prospective phase I clinical trial, we treated 23 patients (23 eyes) with ophthalmic plaque radiotherapy for subfoveal exudative macular degeneration. Palladium 103 ophthalmic plaque brachytherapy was delivered to a retinal apex dose of 1,250 to 2,362 cGy (rad). Early Treatment Diabetic Retinopathy Study type visual acuity determinations, ophthalmic examinations, and angiography were performed before and after treatment. Clinical evaluations were performed in a nonrandomized and unmasked fashion. RESULTS: Patients were followed up for a mean (+/-SD) of 19 +/- 10.7 months (range, 3 to 37 months). Six months after radiation therapy, three (16%) of 19 eyes had lost 3 or more lines of best-corrected visual acuity; 12 months after radiation therapy, four eyes (31% of 13 eyes), and 24 months after radiation therapy, only two (22% of nine eyes) lost 3 or more lines of visual acuity. No eye suffered sudden irreversible loss of central vision. No radiation retinopathy, optic neuropathy, or cataract could be attributed to radiotherapy within this follow-up period. CONCLUSION: Ophthalmic plaque radiotherapy can be used to treat neovascular age-related macular degeneration. In contrast to external beam radiotherapy, ophthalmic plaque radiotherapy is a unilateral treatment, which allows a larger dose to be delivered to the macula with less irradiation of normal ocular structures. We have found no sight-limiting complications at the doses, dose rates, and follow-up evaluated in this study.

Phenotypic analysis of human glioma cells expressing the MMAC1 tumor suppressor phosphatase.

MMAC1, also known as PTEN or TEP-1, was recently identified as a gene commonly mutated in a variety of human neoplasias. Sequence analysis revealed that MMAC1 harbored sequences similar to those found in several protein phosphatases. Subsequent studies demonstrated that MMAC1 possessed in vitro enzymatic activity similar to that exhibited by dual specificity phosphatases. To characterize the potential cellular functions of MMAC1, we expressed wild-type and several mutant variants of MMAC1 in the human glioma cell line, U373, that lacks endogenous expression. While expression of wild-type MMAC1 in these cells significantly reduced their growth rate and saturation density, expression of enzymatically inactive MMAC1 significantly enhanced growth in soft agar. Our observations indicate that while wild-type MMAC1 exhibits activities compatible with its proposed role as a tumor suppressor, cellular expression of MMAC1 containing mutations in the catalytic domain may yield protein products that enhance transformation characteristics.

Does preventive vaccination with engineered tumor cells work in cancer-prone transgenic mice?

The use of genetically modified tumor cells as vaccines has been successful in numerous animal models of grafted syngenic tumors and has provided the groundwork for many clinical trials of gene therapy in cancer patients. To investigate the real efficacy of ex vivo gene therapy-based vaccines, we used transgenic mice that express the SV40 large T and small t antigens under the control of hepatic antithrombin III (ASV-B)-regulatory sequences. These mice systematically develop hepatocarcinoma. Hepatoma cells, derived from ASV-B transgenic mice, were gene-transduced to express either interleukin-2, interleukin-4, the granulocyte-macrophage colony-stimulating factor, or the T-cell costimulatory molecule B7.1. First, we demonstrated the vaccine potential of engineered hepatoma cells by immunizing nontransgenic mice with these cells, which prevented the growth of subsequent grafted nontransduced hepatoma cells. However, vaccination of pretumoral transgenic animals with various combinations of engineered hepatoma cells failed to inhibit hepatoma onset and progression. Rather, tumor development in ASV-B mice appears to be dependent on the immune system, since neonatal induction of immunotolerance to tumor in ASV-B mice cells was associated with a moderate, but significant, acceleration of tumor development. These results seriously call into question the efficacy of this strategy of active vaccinotherapy against natural tumors.

Steatohepatitis-inducing drugs cause mitochondrial dysfunction and lipid peroxidation in rat hepatocytes.

BACKGROUND & AIMS: 4,4'-Diethylaminoethoxyhexestrol (DEAEH), amiodarone, and perhexiline cause steatohepatitis in humans. The mechanisms of these effects are unknown for DEAEH and have not been completely elucidated for amiodarone and perhexiline. The aim of this study was to determine these mechanisms. METHODS: Rat liver mitochondria, cultured rat hepatocytes, or rats were treated with these drugs, and the effects on mitochondrial respiration, beta-oxidation, reactive oxygen species formation, and lipid peroxidation were determined. RESULTS: DEAEH accumulated in mitochondria and inhibited carnitine palmitoyl transferase I and acyl-coenzyme A dehydrogenases; it decreased beta-oxidation and caused lipid deposits in hepatocytes. DEAEH also inhibited mitochondrial respiration and decreased adenosine triphosphate (ATP) levels in hepatocytes. DEAEH, amiodarone, and perhexiline augmented the mitochondrial formation of reactive oxygen species and caused lipid peroxidation in rats. CONCLUSIONS: Like amiodarone and perhexiline, DEAEH accumulates in mitochondria, where it inhibits both beta-oxidation (causing steatosis) and respiration. Inhibition of respiration decreases ATP and also increases the mitochondrial formation of reactive oxygen species. The latter oxidize fat deposits, causing lipid peroxidation. We suggest that ATP depletion and lipid peroxidation may cause cell death and that lipid peroxidation products may account, in part, for other steatohepatitis lesions.