Plaque volume measurement by magnetic resonance imaging as an index of remodeling of middle cerebral artery: correlation with transcranial color Doppler and magnetic resonance angiography.

BACKGROUND: The feasibility of a noninvasive evaluation of remodeling of the middle cerebral artery (MCA) by magnetic resonance imaging (MRI) was assessed. The results were correlated with magnetic resonance angiography (MRA) and transcranial color Doppler (TCD) findings. METHODS: 26 patients (13 male and 13 female, age ranged from 46 to 82 years) who presented with symptoms of cerebrovascular accidents had TCD, MRA and MRI assessment of the MCA. The TCD and MRA findings of 40 MCAs accessible by TCD were correlated with the ratio of cross-sectional area of the vessel (VA), luminal area of the vessel (LA) and plaque load (PL). RESULTS: The VA ratio and LA showed no correlation with TCD or MRA results. PL however was shown to be associated with both TCD and MRA. CONCLUSIONS: MRI could be used to assess remodeling in MCAs. PL was found to be associated with TCD and MRA findings.

Modification of doxorubicin-induced cardiotoxicity: manipulation of the dosage schedule.

Modification of the dosing schedule for doxorubicin (DOX) administration represents a possible method of reducing cardiotoxicity from this potent anti-cancer drug, while at the same time maintaining its cytotoxic action. The quantitative effects of modified dosage scheduling have been investigated in a clinically relevant rat model. Cardiotoxicity to DOX was assessed by the degree of reduction in cardiac output at 4-24 weeks after the intravenous administration of DOX. The effects of dose schedules involving three or six small dose administrations, over one and two weeks, were compared with that produced by large single doses of DOX. The total drug dose administered for each schedule was varied in order to establish dose-effect relationships. After a total dose of 3 mg/kg DOX, given as three or six equal small doses, there was a gradual decline in cardiac output in the first 12 weeks after drug administration. Between 12 and 24 weeks, the reduction in cardiac function was relatively stable at between 65% and 85% of that of age-matched controls for three and six equal small doses, respectively. Dose-effect curves for animals showing a > or = 30% reduction in cardiac function after 12 weeks indicated the degree of reduction in cardiac function produced by the modified dose scheduling. Compared with a large single dose, larger total doses were required to produce the same severity of damage. Thus, schedules based on three and six equal small doses resulted in dose modification factor of 1.5 +/- 0.23 and 2.1 +/- 0.28, respectively, when compared with the same effect produced by a large single dose. This appeared to be independent of the severity of cardiac damage, suggesting a simple mathematical relationship between the total acceptable dose of DOX and the dose administered at each intravenous injection. These modifications in the cardiotoxicity of DOX produced by the administration of multiple small doses were of the same order of magnitude as that produced by other methods introduced to reduce anthracycline cardiotoxicity.

Enhanced drug resistance in cells coexpressing ErbB2 with EGF receptor or ErbB3.

Overexpression of ErbB2 has been found in approximately 25-30% of human breast cancers and has been shown to render the cancer cells more resistant to chemotherapy. However, it is not clear whether ErbB2 overexpression renders the cells more resistant to specific anti-cancer drugs or renders the cells more resistant to a broad range of anti-cancer drugs. It is not clear how the function of ErbB2 in drug resistance is related to expression and activation of the other ErbB receptors. In this communication, we showed that several breast cancer cell lines including BT20, BT474, MCF-7, MDA-MB-453, and SKBR-3 cells had a similar pattern of resistance to a broad range of anti-cancer drugs including 5-Fluorouracil, Cytoxan, Doxorubincin, Taxol, and Vinorelbin, suggesting a mechanism of multidrug resistance. High expression of P-glycoprotein and the ErbB receptors contribute to drug resistance of these breast cancer cells; however, overexpression of ErbB2 alone is not a major factor in determining drug resistance. To further determine the role of the ErbB receptors in drug resistance, we selected various NIH 3T3 cell lines that specifically expressed EGF receptor (EGFR), ErbB2, ErbB3, EGFR/ErbB2, EGFR/ErbB3, or ErbB2/ErbB3. A cytotoxicity assay showed that expression of ErbB2 alone did not significantly enhance drug resistance, whereas coexpression of either EGFR or ErbB3 with ErbB2 significantly enhanced drug resistance. Moreover, ErbB2 was highly phosphorylated in NIH 3T3 cells that coexpress ErbB2 with either EGFR or ErbB3, but not in NIH 3T3 cells that express ErbB2 alone. Together, our results suggest that coexpression of EGFR or ErbB3 with ErbB2 induces high phosphorylation of ErbB2 and renders the cells more resistant to various anti-cancer drugs.

The mode of action of taxol: apoptosis at low concentration and necrosis at high concentration.

The cytotoxicity of Taxol represents both inhibition of cell proliferation and cell death. The drug blocked cells in the G2/M phase of the cell cycle. It has also been reported that Taxol induced cell apoptosis; however, the mode of action of Taxol is far from clear. In this communication, the cytotoxicity of Taxol in various breast cancer cell lines was carefully examined. We showed that Taxol treatment induced a biphasic decrease of viable cells. While the first phase of decrease occurred over concentrations ranging from 0.005 to 0.05 microM and the second phase of decrease occurred at concentrations ranging from 5 to 50 microM, there was a plateau between these ranges. We determined that the biphasic response was due to two different mechanisms. In the lower concentration range (0.005-0.05 microM), Taxol stabilized the spindle during mitosis, thereby blocking mitosis. This mitotic block led to the inhibition of cell proliferation and the induction of apoptosis. In the higher concentration range (5-50 microM), Taxol mainly increased the polymerization of microtubule and stimulated the formation of microtubule bundles, which blocked entry into S phase. This inhibition of S phase entry led to the inhibition of cell proliferation and the induction of necrosis. These findings may have profound clinical implications.

Endocytosis deficiency of epidermal growth factor (EGF) receptor-ErbB2 heterodimers in response to EGF stimulation.

Epidermal growth factor (EGF) stimulates the homodimerization of EGF receptor (EGFR) and the heterodimerization of EGFR and ErbB2. The EGFR homodimers are quickly endocytosed after EGF stimulation as a means of down-regulation. However, the results from experiments on the ability of ErbB2 to undergo ligand-induced endocytosis are very controversial. It is unclear how the EGFR-ErbB2 heterodimers might behave. In this research, we showed by subcellular fractionation, immunoprecipitation, Western blotting, indirect immunofluorescence, and microinjection that, in the four breast cancer cell lines MDA453, SKBR3, BT474, and BT20, the EGFR-ErbB2 heterodimerization levels were positively correlated with the ratio of ErbB2/EGFR expression levels. ErbB2 was not endocytosed in response to EGF stimulation. Moreover, in MDA453, SKBR3, and BT474 cells, which have very high levels of EGFR-ErbB2 heterodimerization, EGF-induced EGFR endocytosis was greatly inhibited compared with that in BT20 cells, which have a very low level of EGFR-ErbB2 heterodimerization. Microinjection of an ErbB2 expression plasmid into BT20 cells significantly inhibited EGF-stimulated EGFR endocytosis. Coexpression of ErbB2 with EGFR in 293T cells also significantly inhibited EGF-stimulated EGFR endocytosis. EGF did not stimulate the endocytosis of ectopically expressed ErbB2 in BT20 and 293T cells. These results indicate that ErbB2 and the EGFR-ErbB2 heterodimers are impaired in EGF-induced endocytosis. Moreover, when expressed in BT20 cells by microinjection, a chimeric receptor composed of the ErbB2 extracellular domain and the EGFR intracellular domain underwent normal endocytosis in response to EGF, and this chimera did not block EGF-induced EGFR endocytosis. Thus, the endocytosis deficiency of ErbB2 is due to the sequence of its intracellular domain.

Boron neutron capture therapy: long-term effects on the skin and spinal cord of the rat.

Our studies of the pharmacokinetics of boron focused on the variations in the concentration in blood of Sprague-Dawley rats with time after the administration of single intravenous doses of 50-200 mg/kg of 10B-enriched sodium mercaptoundecahydro-closo-dodecaborate (BSH). After the lowest dose of BSH there was a progressive decline in the boron content of the blood, with a biological half-life (t1/2) of approximately 4.5 h. Higher doses of BSH resulted in slower boron clearance rates. A dose of 100 mg/kg of BSH was the maximum safely tolerated by the rats. The boron content of the skin at this dose of BSH was a factor of 0.6 lower than that in the blood. To determine the dose-related changes in the response of the central nervous system to BNCT-type radiation exposures, a well-established and clinically relevant model, the rat spinal cord, was used. The spinal cords (20 mm field length) of rats, infused with 100 mg/kg of BSH, were irradiated for 3 to 5 h with cold thermal neutrons from the H6 beam on the DIDO reactor (AERE, Harwell). The skin surface neutron flux was 4.8 x 10(8) n/cm2/s. Exposure times of > or = 4 h resulted in vigorous, biphasic skin reactions, indicative of long-term vascular damage in the dermis. Rats were monitored closely for 84 weeks after irradiation. No abnormal neurological responses were observed and there was no histological evidence of lesions in the spinal cord at the end of the study. These findings indicate that the central nervous system has a high tolerance to BNCT-type radiation using BSH as the neutron capture agent.

The protective activity of ICRF-187 against doxorubicin-induced cardiotoxicity in the rat.

The protective activity of the bisdioxopiperazine ICRF-187 against the cardiotoxicity of doxorubicin was evaluated in the rat using both functional and histological assays. Animals that had received a single i.v. dose of doxorubicin (4 mg/kg) alone were compared with those that had been pretreated with a single i.v. injection of saline or ICRF-187 (40 or 60 mg/kg). All rats showed a transient reduction in body weight during the first 3 weeks after drug administration. The greatest reduction (approximately 16%) was observed in animals that had received a combination of ICRF-187 (40 or 60 mg/kg) and doxorubicin. Deaths related to cardiotoxicity were observed only in rats that had received doxorubicin alone and in those treated with saline; most of the deaths occurred at between 8 and 13 weeks after drug administration. Sequential assessments of heart function showed a persistent depression of cardiac output in animals that had received doxorubicin, with or without pretreatment with ICRF-187. The reduction in cardiac output observed in rats that had been pretreated with ICRF-187 (40 or 60 mg/kg) amounted to approximately 15% and approximately 30% after 12 and 20 weeks, respectively, indicating that cardioprotection was only partial. Nevertheless, this represented a marked improvement as compared with the approximately 35% reduction in cardiac output measured at 12 weeks in animals that had received doxorubicin but without pretreatment with ICRF-187. Histological examination of animals that had died during the course of the study and had received doxorubicin after pretreatment with saline revealed severe myocardial lesions typical of doxorubicin-induced damage. In contrast, animals that had been pretreated with ICRF-187 and survived for up to 20 weeks after treatment showed a marked amelioration of these lesions. The present findings may be interpreted as a true cardioprotection or a delay in the onset of the cardiotoxicity of doxorubicin resulting from pretreatment with the bisdioxopiperazine ICRF-187. Although prior and ongoing clinical trials clearly indicate that ICRF-187 protects patients well against doxorubicin-induced heart damage, further investigations are required before high doses of ICRF-187 can be used as a means of increasing the protective activity of this drug against doxorubicin-induced cardiotoxicity.

Reduced cardiotoxicity of doxorubicin given in the form of N-(2-hydroxypropyl)methacrylamide conjugates: and experimental study in the rat.

A rat model was used to evaluate the general acute toxicity and the late cardiotoxicity of 4 mg/kg doxorubicin (DOX) given either as free drug or in the form of three N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer conjugates. In these HPMA copolymers, DOX was covalently bound via peptide linkages that were either non-biodegradable (Gly-Gly) or degradable by lysosomal proteinases (Gly-Phe-Leu-Gly). In addition, one biodegradable conjugate containing galactosamine was used; this residue was targeted to the liver. Over the first 3 weeks after the i.v. administration of free and polymer-bound DOX, all animals showed a transient reduction in body weight. However, the maximal reduction in body weight seen in animals that received polymer-bound DOX (4 mg/kg) was significantly lower than that observed in those that received free DOX (4 mg/kg) or a mixture of the unmodified parent HPMA copolymer and free DOX (4 mg/kg; P less than 0.01). Throughout the study (20 weeks), deaths related to cardiotoxicity were observed only in animals that received either free DOX or the mixture of HPMA copolymer and free DOX; in these cases, histological investigations revealed marked changes in the heart that were consistent with DOX-induced cardiotoxicity. Sequential measurements of cardiac output in surviving animals that received either free DOX or the mixture of HPMA copolymer and free DOX showed a reduction of approximately 30% in function beginning at the 4th week after drug administration. The heart rate in these animals was approximately 12% lower than that measured in age-matched control rats (P less than 0.05). Animals that were given the HPMA copolymer conjugates containing DOX exhibited no significant change in cardiac output throughout the study (P less than 0.05). In addition, no significant histological change was observed in the heart of animals that received DOX in the form of HPMA copolymer conjugates and were killed at the end of the study. However, these animals had shown a significant increase in heart rate beginning at 8 weeks after drug administration (P less than 0.01).(ABSTRACT TRUNCATED AT 400 WORDS)

Morphological and functional changes in the rat heart after X irradiation: strain differences.

The hearts of mature male rats of the Wistar and Sprague-Dawley strains were locally irradiated with single doses of 17.5 and 20.0 Gy of X rays, respectively. These two dose levels had previously been shown to result in a comparable latent period between irradiation and the death of rats of these two strains from cardiac failure. Morphological changes in the myocardium and modifications in cardiac function were assessed in the animals at 28, 70, and 100 days after irradiation. The first radiation-induced change which was observed in the myocardium was a rapid decline in capillary density and a loss of alkaline phosphatase activity by the capillary endothelial cells. The capillary density was reduced to approximately 50% of that of unirradiated control values at 28 days and to approximately 40% of the control values between 70 and 100 days after irradiation. The loss of enzyme activity was also detected at 28 days. Examination of histological sections showed an increase by 70 days in the areas with negative enzyme activity up to approximately 70% of the myocardium. The reduction in capillary density and the loss of enzyme activity occurred before any marked pathological changes were seen in the myocardium. The pathological lesions seen in the myocardium at 100 days after irradiation were qualitatively and quantitatively the same in the two strains of rat. Measurements of cardiac output in Sprague-Dawley rats showed a gradual decline in output after irradiation; however, measurements in Wistar rats showed a progressive increase in cardiac output over the same period of time. It was shown by rubidium extraction that there was an increase in the percentage of the total cardiac output distributed to the ventricular muscle of Sprague-Dawley rats, while similar measurements in Wistar rats showed no significant change. In spite of the marked strain differences observed in cardiac output and rubidium extraction, blood perfusion per gram of ventricular muscle was apparently not modified in both strains of rat after irradiation. These findings indicated that the correlation between morphological effects after irradiation and the functional expression of damage is highly complex.

A functional assessment of the relative cardiotoxicity of adriamycin and epirubicin in the rat.

The cardiotoxicity of the two anthracyclines, adriamycin and epirubicin, were studied in 14-week-old Sprague-Dawley rats after the intravenous administration of single doses of 1-4 mg/kg of adriamycin and 2-6 mg/kg of epirubicin. These doses of the two drugs were well tolerated with little acute toxicity. Acute toxicity was characterised by a transient reduction in body weight with recovery within 14 days. The cardiac output of the rats was measured at 4-weekly intervals, for up to 20 weeks, using an external counting technique with the radioactive tracer, 99mTc. The time-related changes in cardiac function in the rat after adriamycin and epirubicin were similar. The time course of the changes in cardiac output were biphasic. There was an initial phase of rapid decline in cardiac output in the first 12 weeks (phase I) and a second phase of persistent depression in cardiac function (phase II) after 12 weeks. The late progression of anthracycline-induced cardiotoxicity could be predicted from the measurements of cardiac output at 12 weeks. The relative cardiotoxicity of adriamycin and epirubicin was evaluated from the reductions in cardiac output and from the incidence of deaths related to cardiotoxicity. Both methods of evaluation showed that adriamycin was approximately twice as cardiotoxic as epirubicin. This relationship was independent both of the dose level of drugs used and of the damage level used for the evaluation. Dose-response curves were steeper for adriamycin than for epirubicin. The present findings agree with the somewhat limited clinical data suggesting that the present rat model is highly predictive and clinically relevant.

The relative toxicity of intravenous and intraperitoneal doses of epirubicin.

The toxicity of single doses of the anthracycline epirubicin was compared in the rat after either the intravenous (i.v.; 2-6 mg/kg) or intraperitoneal (i.p.; 4-8 mg/kg) administration of the drug. These doses produced comparable acute toxicity that was characterised by a dose-dependent, transient reduction in body weight (less than 15%) in the first 2 weeks after drug administration. Sequential measurements of cardiac output in animals that received i.v. doses of epirubicin showed that the time-related changes in cardiac function were biphasic. There was an initial decline in cardiac output in the first 12 weeks, which was followed by a phase of persistent depression in cardiac output between 12 weeks and 20 weeks. The time-related changes in cardiac function after i.p. doses of the drug were more variable, although the trend of changes, as after i.v. administration, appeared to be dose-dependent. Recovery of cardiac function occurred at 20 weeks after an i.p. dose of 4 mg/kg; however, after 6 mg/kg, cardiac function was significantly depressed after greater than or equal to 16 weeks. For both routes of administration, the likelihood of late cardiac complications was dependent on the level of the reduction in cardiac output at 12 weeks. A study of the impairment of cardiac output and the incidence of cardiac-related mortality demonstrated that epirubicin was more cardiotoxic after i.v. administration. Equivalent cardiotoxic doses of epirubicin after i.v. and i.p. administration were highly linearly correlated (r = 0.998), although there appeared to be a threshold dose of 3.33 mg/kg after i.p. administration of the drug. Thus, the relative cardiotoxicity between the two routes of administration was found to be dependent on the drug dose and, hence, the level of effect. The difference in the effect was less for high drug doses. The LD50 for deaths due to cardiotoxicity at 20 weeks was 4.42 +/- 0.42 mg/kg after i.v. administration, which was significantly lower than the value of 6.28 +/- 0.41 mg/kg obtained after i.p. administration of the drug (P less than 0.01). There was no qualitative difference in the histological lesions induced in the myocardium after the i.v. vs i.p. administration of epirubicin.

Time- and dose-related changes in the white matter of the rat brain after single doses of X rays.

Following the local irradiation of the rat brain with single doses of 17.5-25 Gy of X rays, necrosis of the white matter was seen after a latent interval of greater than 26 weeks. At 39 weeks and 52 weeks after irradiation the incidence of necrosis was dose-related. The doses associated with a 50% incidence of necrosis in the white matter (ED50) at these times were 23.45 +/- 0.49 and 20.98 +/- 0.91 Gy, respectively. At both these times the incidence of necrosis was higher in the fimbria than in the capsula interna and the corpus callosum. This reflects a variation in the latency time for the appearance of necrosis. Necrosis occurs earlier in the fimbria. In the corpus callosum and the capsula interna the latency times for the appearance of necrosis were also dose-dependent. In the latent period prior to the onset of necrosis of the fimbria, a number of dose-related changes were seen in the vasculature and the associated astroglial cells. These changes, which included blood vessel dilation, blood vessel wall thickening, endothelial cell nuclear enlargement and the hypertrophy of perivascular astrocytes, were highly correlated and when combined appeared to represent a "unit of tissue injury". The incidence and severity of this "unit of tissue injury" apparently increased with time after irradiation until necrosis ensued. These dose-related vascular/glial changes were preceded by a reduction in the endothelial cell and vascular density. No early changes were seen in the number of glial parenchymal cells.

Time- and dose-related modifications in cardiac function in rats after single intravenous doses of epirubicin.

Time-related changes in cardiac output have been studied in rats after the intravenous administration of a range of single doses of epirubicin. There was an initial decline in cardiac function in the first 4-12 weeks after drug treatment at a rate that appeared to be dose-related. After 12 weeks, an average cardiac output of approximately 70%, relative to control values, was maintained in animals that survived until the end of the study. This effect was independent of the drug dose, reflecting the death of animals in the higher dose groups. The incidence of drug-induced cardiotoxic deaths, the majority of which (approximately 62%) occurred within the first 12 weeks, was dose-related, and suggested a LD50 for cardiac-related mortality of 5.48 +/- 0.39 mg/kg. There appeared to be a relationship between the reduction in cardiac output at 12 weeks and the probability of a further deterioration in function. Animals showing a greater than 40% reduction in cardiac output at 12 weeks accounted for greater than 90% of all the additional deaths between 12 and 20 weeks. Rats showing a less than 40% reduction in cardiac output at 12 weeks had a very high probability of surviving without clinical signs, although with a persistently depressed cardiac function. These findings are similar to the trends demonstrated in the sparse clinical data and this supports the view that the present simple animal model is suitable for the investigation of cardiotoxicity after the administration of cardiotoxic cytotoxic drugs.

Dose-dependent and time-dependent changes in the choroid plexus of the irradiated rat brain.

A histological assessment has been made of both time- and dose-related changes in the choroid plexus after the local irradiation of the rat brain with single doses of 17.5-25 Gy of X rays. These investigations involved the serial killing of animals 1-52 weeks after irradiation and the quantitative and semiquantitative evaluation of histological sections. Counts of the relative number of cells in the choroid plexus of the lateral ventricles showed an atrophy of the epithelial layer after 13 weeks. However, this was not as marked as the reduction in the number of endothelial cells in the wall of blood vessels. Moreover, the epithelium had recovered by 39 weeks after irradiation, while the dose-related depletion in endothelial cells tended to be progressive. A highly correlated group of changes in the vascular-connective tissue was used to produce a numerical "factor". This represented a combined score of radiation damage which was both time- and dose-related. These data suggest that, as an expression of late radiation damage to the choroid plexus, the effect on the endothelium was more important than that to the epithelial cells.

Radiation induced damage in the choroid plexus of the rat brain: a histological evaluation.

Early and late histological changes have been investigated in the choroid plexus after the local irradiation of the brain of rats with single doses of 17 X 5-25 Gy of X-rays. Changes were seen in epithelial cells within 1 week of irradiation. This was characterized by vacuolation, blebbing and partial lysis of the cytoplasm. Oedema in the underlying connective tissue was also seen after 1 week. A marked reduction in the number of endothelial cells, lining the walls of blood vessels, was found after 12 weeks; the severity was dose related and increased with time after irradiation. Slight atrophy of the epithelium was seen after 12 weeks. The reduction in endothelial cell number was associated with an attempt at regeneration as indicated by endothelial nuclear enlargement, nuclear pairs or groups and by the appearance of mitotic figures. Evidence or epithelial regeneration was not seen until week 26. Interstitial fibrosis was seen from 26 weeks after irradiation with the severity being both dose and time related. These late fibrotic changes were associated with basal membrane thickening, degenerative changes in the tunica media of arterioles and the appearance of thrombi. These changes are compared and contrasted with those seen in other normal tissues after irradiation and are discussed in terms of their relevance to the development of late radiation damage to the central nervous system.

Effects of single doses of radiation on cardiac function in the rat.

The hearts of rats were irradiated locally to single doses of X-ray in the range of 20-50 Gy. The mean latency period between treatment and death ranged from 550 +/- 12 days to 260 +/- 13 days for animals irradiated with doses of 20 and 50 Gy. The growth rate and cardiac output was followed for a period of 10 months after treatment. In irradiated animals, there was a delay in growth and the rate of growth plateaued at approximately 6 months after treatment. Cardiac output was measured by an external counting technique. There was a general decline in cardiac output at time intervals from 4 to 10 months after irradiation. This reduction in cardiac output was dose-dependent. A single dose of 30 Gy reduced the cardiac output to approximately 70% of the control value, 6 months after irradiation.