Male B6D2F1 Research Articles

Modulation of nonprotein sulphydryl compounds rhythm with buthionine sulphoximine: relationship with oxaliplatin toxicity in mice.

The relationship between the rhythm in tissue nonprotein sulphydryl groups (NPSH) and that in 1,2-diamine (trans-I)-cyclohexane oxalatoplatinum (1-OHP) toxicity was investigated in a total of 266 male B6D2F1 mice, using buthionine sulphoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase. Mice were synchronized with an alternation of 12 h light (L) and 12 h darkness (D; LD 12:12), and circadian time was expressed in hours after light onset (HALO). NPSH was measured in liver, jejunum and bone marrow at 0, 8 and 16 HALO. Dosing 1-OHP at these times achieved intermediate. high or low toxicity respectively. The physiological circadian rhythm in NPSH content was statistically significant in all tissues studied, with a maximum at the transition from D to L (0 HALO). BSO administration (450 mg/kg i.p., 4 h before sampling) induced a large depletion in liver and jejunum NPSH at their physiological peak (0 HALO), but exerted no significant effect at their trough (8 HALO). As a result, 24 h rhythm was suppressed in liver and jejunum, but remained similar to the physiological one in bone marrow. BSO enhanced 1-OHP-induced mortality and jejunal toxicity, but exerted no significant effect upon bone marrow toxicity. Despite these differences, 1-OHP remained least toxic at 16 HALO, near the middle of the dark span, which corresponds to maximum activity in the circadian rest/activity cycle. Our results show that mean NPSH levels in liver seem to account for the mean level of 1-OHP toxicity, while jejunal NPSH rhythm plays an important role in the intestinal toxicity rhythm of this drug.

Tocolytic effect of magnesium sulfate and/or urinary trypsin inhibitor against lipopolysaccharide-induced preterm delivery in mice.

The purpose of this investigation was to evaluate the effect of magnesium sulfate (magnesium) alone or along with urinary trypsin inhibitor (UTI) in a mouse model in the prevention of preterm delivery. On day 17 of pregnancy, female C3H/HeN mice impregnated by male B6D2F1 mice were given two intraperitoneal injections of lipopolysaccharide (LPS; 50 microg/kg) at a 3-hour interval, which treatment induced a 100% incidence of preterm delivery within 25 hours of the second dose. Magnesium (4, 5, 6, 7, or 8 mg/hr, s.c.), UTI (25 x 10(4) units/kg, i.p.), magnesium (5 mg/hr, s.c.) plus UTI (25 x 10(4) units/kg, i.p.), saline solution (0.3 ml/hr, s.c.), or saline solution (0.1 ml/hr, s.c. and 2.5 ml/kg, i.p.) was administered to pregnant animals on day 18 of gestation. UTI was intraperitoneally given 5 times at 2-hour intervals from 8:00 am to 4:00 pm, and magnesium was infused subcutaneously and constantly from 8:00 am to 6:00 pm. In addition, the preventive effect of magnesium on LPS-induced contraction of uterine muscle strips and that of magnesium, UTI, or magnesium plus UTI on LPS-induced calcium influx in uterine smooth muscle cells were examined using muscle tissue isolated from pregnant mice on day 17 of gestation. The incidence of preterm delivery decreased significantly in a dose-dependent fashion with magnesium treatment, and there was a significant and synergistic decrease after combined treatment with magnesium plus UTI. The in vitro uterine contraction and calcium influx induced by LPS were significantly suppressed by magnesium. The latter was significantly suppressed by UTI and additively reduced by magnesium plus UTI. Combination therapy with magnesium plus UTI may possibly be helpful for preventing preterm delivery in humans without the severe side effects induced by hypermagnesemia.

Biologically active heteroarotinoids exhibiting anticancer activity and decreased toxicity.

A series of retinoids, containing heteroatoms in a cyclic ring and called heteroarotinoids, were synthesized, and their biological activity was evaluated using tissue culture lines that have measurable responses to trans-retinoic acid (t-RA). Transglutaminase (TGase) was assessed in the human erythroleukemia cell line (GMO6141A) as an indicator of differentiation and apoptosis. Proliferation was evaluated in a human cervical cell line, CC-1, which exhibits dose-dependent alterations in growth rate in response to treatment with trans-retinoic acid. Activation of nuclear retinoic acid receptors was determined in a reporter cell line established from CC-1. The reporter line, called CC-B, contains a reporter gene controlled by a retinoic acid responsive element (RARE) and a thymidine kinase (tk) promoter. Treatment of the CC-B line with the heteroarotinoids resulted in a dose-responsive and retinoid-dependent regulation of reporter gene expression. The heteroarotinoids exhibited activity in all assays and correlated in a statistically significant manner between assays. RARE transactivation activity in CC-B cells correlated with induction of TGase in GMO6141A (R = 0.96) and with a decrease in the growth rate of CC-1 cells (R = -0.90). The ability of the selected heteroarotinoids to induce differentiation, inhibit proliferation, and activate nuclear receptors demonstrates the chemotherapeutic potential of these agents. In view of the biological activity cited, an in vivo toxicity study was conducted on male B6D2F1 mice with three heteroarotinoids, namely 8 [(2E,4E,6E)-3,7-dimethyl-7-(1,2,3,4-tetrahydro-4,4-dimeth ylthiochroman-6-yl)-2,4,6-heptatrienoic acid], 10 [(2E,4E,6E)-3,7-dimethyl-7-(1,2,3,4-tetrahydro-4,4-dimeth ylchroman-6-yl)-2, 4,6-heptatrienoic acid], and 13 [(E)-p-[2-(4,4-dimethylchroman-6-yl)propenyl]benzoic acid]. The mice were used with gavage of heteroarotinoids in corn oil [0.1, 0.2, 0.4, or 0.8 mg/kg] and with 0.01 or 0.05 mg/kg of TTNPB (5) [(E)-4-[2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1- propenyl]benzoic acid] as reference controls. The target organs affected in the mice by the three heteroarotinoids were those typically associated with t-RA (1) toxicity. The maximum tolerated dose (MTD) of 13 was 9.4 mg/kg/day, which was equal in toxicity to that of t-RA (1) and 1000-fold less toxic than TTNPB (5). The MTDs of 8 and 10 were 34 and 32 mg/kg/day, respectively, which is 3-fold less toxic than t-RA (1) and 3000-fold less toxic than TTNPB (5). The 3000-fold reduced toxicity, compared with only a 27% reduction biological activity of 8 and 10 with respect to that of TTNPB, observed in our assays indicates a good therapeutic ratio of these heteroarotinoids over the parent compound. The biological activity and reduced toxicity of these heteroartinoids demonstrate the potential efficacy as anticancer agents.

Preventive effect of ritodrine hydrochloride and/or urinary trypsin inhibitor against lipopolysaccharide-induced preterm delivery in mice.

The purpose of this study was to confirm the preventive effect of ritodrine hydrochloride (ritodrine) alone or ritodrine plus urinary trypsin inhibitor (UTI) in a mouse model of preterm delivery. On day 17 of pregnancy, female C3H/HeN mice impregnated by male B6D2F1 mice were given two intraperitoneal injections of lipopolysaccharide (LPS) (50 micrograms/kg) at a 3-hour interval, which induced a 100% incidence of preterm delivery within 25 hours of the second dose. Ritodrine (1, 3, or 10 mg/kg, p.o.), UTI (25 X 10(4) units/kg, i.p.), ritodrine (3 mg/kg, p.o.) plus UTI (25 x 10(4) units/kg, i.p.), distilled water (10 ml/kg, p.o.), or distilled water (10 mg/kg, p.o.) plus saline solution (10 ml/kg, i.p.) were administered to the pregnant animals 10 times at 1-hour intervals from 8:00 AM to 5:00 PM on day 18 of pregnancy. In addition, the preventive effect of ritodrine, UTI, or ritodrine plus UTI was examined on LPS-induced contraction of uterine muscle strips isolated from pregnant mice on day 17 of gestation. The incidence of preterm delivery decreased significantly in a dose-dependent fashion with ritodrine treatment, and there was a significant and synergistic decrease after combined treatment with ritodrine plus UTI. The in vitro uterine contraction induced by LPS was significantly suppressed by both ritodrine and UTI. Combination therapy with ritodrine plus UTI may be helpful for preventing preterm delivery in humans without the cardiovascular side effects that often accompany treatment with ritodrine alone.

Trophoblastic apoptosis in mice with preterm delivery and its suppression by urinary trypsin inhibitor

To investigate histopathologic changes of the placenta in mice with preterm delivery induced by lipopolysaccharide and the effect of urinary trypsin inhibitor. Female C3H/HeN mice impregnated by male B6D2F1 mice were treated with lipopolysaccharide (50 micrograms/kg, intraperitoneally) or lipopolysaccharide plus urinary trypsin inhibitor (25 x 10(4) U/kg, intraperitoneally). At 3, 6, 9, and 18 hours after the second dose of lipopolysaccharide, and at delivery in the control and urinary trypsin inhibitor-treated groups, the concentrations, of interleukin-1 alpha and tumor necrosis factor-alpha were determined in serum and amniotic fluid. Subsequently, the placentas were examined. In the same manner, we examined mice treated with interleukin-1 alpha (250 micrograms/kg, subcutaneously) on day 15 of pregnancy and intact mice on days 15 and 18 of pregnancy as well as at delivery. To assess the direct action of cytokines, we cultured placental slices with tumor necrosis factor-alpha, interleukin-1 alpha, or tumor necrosis factor-alpha plus urinary trypsin inhibitor and examined them morphologically. Control mice were characterized by trophoblastic apoptosis and increased serum levels of tumor necrosis factor-alpha and interleukin-1 alpha. In contrast, urinary trypsin inhibitor-treated mice showed suppression of apoptosis and lower cytokine levels. Interleukin-1 alpha induced trophoblastic apoptosis and increased the serum level of tumor necrosis factor-alpha. The in vitro study showed that tumor necrosis factor-alpha directly induced trophoblastic apoptosis in placental slices. We demonstrated that trophoblastic apoptosis occurs in the placentas of a mouse model with preterm delivery induced by lipopolysaccharide. We postulated that apoptosis may lead to placental abruption, and its development may be prevented by treatment with urinary trypsin inhibitor.

Circadian rhythm in toxic effects of cystemustine in mice: Relevance for chronomodulated delivery

Cystemustine is a new nitrosourea with high anti-tumor activity and a short plasma half-life in mice. The influence of circadian dosing time upon its toxicities was first investigated in a total of 368 synchronized male B6D2F1 mice. Late survival rate varied from 4% in mice receiving a single dose of cystemustine (conventional lethal dose 50%) at 7 hours after 1ight onset (HALO) up to 88% in mice treated at 15 or at 19 HALO. Target organ toxicities (bone marrow, circulating blood cells, spleen, colon and duodenum) were studied following a single slightly lower dose of cystemustine. Leukopenia was the major hematologic effect encountered. Leukocyte count nadir occurred 7 days after injection and was lowest following cystemustine at 7 HALO as compared to 13 or 19 HALO. Recovery was faster after cystemustine at 19 HALO as compared to other dosing times. Bone-marrow necrotic lesions were more pronounced I day after cystemustine at 7 HALO than after cystemustine at 19 HALO. Thus, a large-amplitude circadian rhythm characterized the toxicity of this nitrosourea in mice. The lowest cystemustine toxicity was found near the middle of the active span of the rest-activity circadian cycle of mice. © 1996 Wiley-Liss, Inc.

Circadian rhythm in toxic effects of cystemustine in mice: relevance for chronomodulated delivery.

Cystemustine is a new nitrosourea with high anti-tumor activity and a short plasma half-life in mice. The influence of circadian dosing time upon its toxicities was first investigated in a total of 368 synchronized male B6D2F1 mice. Late survival rate varied from 4% in mice receiving a single dose of cystemustine (conventional lethal dose 50%) at 7 hours after light onset (HALO) up to 88% in mice treated at 15 or at 19 HALO. Target organ toxicities (bone marrow, circulating blood cells, spleen, colon and duodenum) were studied following a single slightly lower dose of cystemustine. Leukopenia was the major hematologic effect encountered. Leukocyte count nadir occurred 7 days after injection and was lowest following cystemustine at 7 HALO as compared to 13 or 19 HALO. Recovery was faster after cystemustine at 19 HALO as compared to other dosing times. Bone-marrow necrotic lesions were more pronounced 1 day after cystemustine at 7 HALO than after cystemustine at 19 HALO. Thus, a large-amplitude circadian rhythm characterized the toxicity of this nitrosourea in mice. The lowest cystemustine toxicity was found near the middle of the active span of the rest-activity circadian cycle of mice.

L-deprenyl treatment in aged mice slightly increases life spans, and greatly reduces fecundity by aged males.

Male and female B6D2F1 (C57BL/6J x DBA/2J)F1 and B6CBAF1 (C57BL/6J x CBA/CaHT6J)F1 mice were injected subcutaneously 3 times a week with L-deprenyl (0.25 mg/kg) starting at mean ages of 26 months and 18.5 months, respectively. Life spans of aging mice were increased 6-9% by the drug. While none of the life span effects were significant for a single genotype and gender, life spans were significantly longer in L-deprenyl-treated animals (p = .011) when all data were combined. L-deprenyl-injected mice consumed about the same amounts of food as controls: L-deprenyl 3.1 g/day, control 3.3 g/day, after 7 months of treatment. There were no significant effects of L-deprenyl on measures of changes with age in the following biological systems: activity, excitement, red blood cell mass, collagen denaturation rate, and wound healing rate. L-deprenyl-treated B6CBAF1 males and females were significantly heavier than controls after 4-6 months of treatment. To measure fecundity, B6CBAF1 males at an average age of 750 days were each caged with two young B6 females; 10 of 17 L-deprenyl-injected males sired an average of 31.3 pups per male, while 14 of 24 controls sired 82.1 pups per male.

In Situ Radiation Sensitivity of Recombinant Human Granulocyte Colony-Stimulating Factor–Recruited Murine Circulating Blood and Bone Marrow Progenitors (Colony-Forming Unit [CFU]–Granulocyte-Macrophage and CFU-Megakaryocyte): Evidence for Possible Biologic Differences Between Mobilized Blood and Bone Marrow

Increasing evidence especially stemming from peripheral blood progenitor transplantation studies points to a possible biologic difference between mobilized blood and bone marrow progenitor cells. The objective of this study was to compare the in situ radiation sensitivity of recombinant human granulocyte colony-stimulating factor (rhG-CSF)-recruited circulating granulopoietic (blood colony-forming unit-granulocyte-macrophage [CFU-GM(blood)]) and megakaryocytopoietic (blood CFU-megakaryocyte [CFU-Meg(blood)]) progenitors, with the nonmobilized fraction remaining in the bone marrow (CFU-GM(femur) and CFU-Meg(femur)). Splenectomized male B6D2F1 mice received 50 micrograms/kg/d rhG-CSF daily for 8 days to induce high levels of circulating progenitors, followed by either total body X-irradiation (TBI) or X-irradiation of the chest (CI) with 62.5, 125, 250, or 500 cGy. Progenitor cells were assayed 24 hours after irradiation. Circulating CFU-GM and CFU-Meg in the blood were decreased in a dose-dependent fashion by both TBI and CI, with TBI causing greater damage than CI. Average D0 values for TBI were 53 cGy for CFU-GM(blood) and 40 cGy for CFU-Meg(blood) D0 values for CI were 90 cGy for CFU-GM(blood) and 140 cGy for CFU-Meg(blood). As seen for blood progenitor cells, TBI caused a dose-dependent decrease of both CFU-GM(femur) (D0, 136 cGy) and CFU-Meg(femur) (D0, 148 cGy). However, radiation-induced bone marrow progenitor cell kill was significantly lower when compared with blood progenitors. Despite the fact that circulating blood elements only received a fraction of the total dose administered as Cl, the extent of blood progenitor kill caused by Cl was higher than the effects of identical TBI doses on bone marrow CFU. The results of this study showed that rhG-CSF-recruited CFU-Meg(blood) and CFU-GM(blood) were considerably more radiosensitive than femoral progenitors, thereby providing novel evidence for a biologic difference between rhG-CSF-recruited peripheral blood progenitors and the nonrecruited bone marrow CFU.

Modulation of cisplatin chronotoxicity related to reduced glutathione in mice

Intracellular reduced glutathione (GSH) concentrations were measured according to the tissue sampling-time along the 24 h scale in male B6D2F1 mice. A significant circadian rhythm in GSH content was statistically validated in liver, jejunum, colon and bone-marrow (P < or = 0.02) but not in kidney. Tissue GSH concentration increased in the dark-activity span and decreased in the light-rest span of mice. The minimum and maximum of tissue GSH content corresponded respectively to the maximum and minimum of cisplatin (CDDP) toxicity. The role of GSH rhythms with regard to CDDP toxicity was investigated, using a specific inhibitor of GSH biosynthesis, buthionine sulfoximine (BSO). Its effects were assessed on both tissue GSH levels and CDDP toxicity at three circadian times. BSO resulted in a 10-fold decrease of the 24 h-mean GSH in kidney. However a moderate GSH decrease characterized liver (-23%) and jejunum (-30%). BSO pretreatment largely enhanced CDDP toxicity which varied according to a circadian rhythm. Although BSO partly and/or totally abolished the tissue GSH rhythms, it did not modify those in CDDP toxicity. We conclude that GSH have an important influence on CDDP toxicity but not in the circadian mechanism of such platinum chronotoxicity.

Levels of serum steroids, aromatase activity, and estrogen receptors in preoptic area, hypothalamus, and amygdala of B6D2F1 male house mice that differ in the display of copulatory behavior after castration.

Most male B6D2F1 hybrid house mice continue to copulate after castration (continuers), whereas others do not (noncontinuers). Copulation in continuers appears estrogen dependent. Serum testosterone (T), 17 beta-estradiol (E2), and dihydrotestosterone (DHT), as well as aromatase activity (AA) and estrogen receptor (ER) levels in preoptic area (POA), hypothalamus (HYP), and amygdala (AM) were measured to determine if continuers and noncontinuers differ in estrogen physiology. In general, continuers and noncontinuers did not differ in serum steroid levels, AA, or ER levels. Castration reduced AA in the POA, HYP, and AM. Castration did not affect nuclear ER levels in the POA and HYP but reduced nuclear ER in AM. The data demonstrate that castrated B6D2F1 male mice continue to be under the influence of circulating nongonadal E2 that is important for copulation.

Use of multiple drug resistance modifiers to overcome tumor resistance to cytostatics in anin vivo system

Quinine and halidor, modifiers of multiple drug resistancein vitro, did not abolish tumor resistance to doxorubicinin vivo, as was shown in male hybrid BDF1 mice with transplanted leukemia P 388 sensitive to doxorubicin and with induced resistance to it. Induced hyperglycemia did not lead to the manifestation of a therapeutic effect of doxorubicin in mice with leukemia P 388 sensitive to the antibiotic and with induced resistance to it.

Mouse lung tumors exhibit specific Ki-ras mutations following transplacental exposure to 3-methylcholanthrene.

A pharmacogenetic mouse model was utilized to determine the role of Cyp1a1 expression on the formation of Ki-ras mutations in lung tumors following transplacental exposure to polycyclic aromatic hydrocarbons (PAHs). A backcross between Ah responsive male B6D2F1 mice and nonresponsive female DBA mice resulted in a litter in which both responsive and nonresponsive fetuses resided in the same nonresponsive maternal environment. Pregnant mothers received a single i.p. injection of either 10 or 30 mg/kg of 3-methylcholanthrene (MC) or olive oil vehicle on day 17 of gestation. At the higher dose of MC, the responsive offspring of both sexes had significantly (P < 0.05) higher incidences of lung tumors than their nonresponsive littermates. The male responsive mice also exhibited a significantly increased liver tumor incidence over the nonresponsive mice at the P < 0.05 level. Administration of 10 mg/kg of MC caused a very low incidence of lung tumors and did not result in the appearance of macroscopically visible liver tumors. Exons 1 and 2 of the Ki-ras gene were amplified from paraffin-embedded tissue samples. The PCR products were screened by allele-specific oligonucleotide hybridization (ASO). Thirteen of 16 lung tumors (81%) screened exhibited point mutations in the 12th or 13th codon, including seven tumors that contained GGT-->GTT (GLY12-->VAL12) transversions, four which exhibited GGT-->TGT (GLY12-->CYS12) transversions, and two which contained GGC-->CGC (GLY13-->ARG13) transversions. None of the tumors had mutations at codon 61. The results obtained by ASO were confirmed by cloning and sequence analysis of the PCR products from four of these tumors. Within the subset of 16 tumors examined in this study, the same types of mutations in the Ki-ras gene were generally present in both responsive and nonresponsive mice, although G-->C transversions were found in two tumors from a single responsive female mouse. Interestingly, while both males and females exhibited the GGT-->GTT mutations at codon 12, the GGT-->TGT transversion was only found in male mice. These results are consistent with a key role for Cyp1a1 in modulating individual susceptibility to cancer formation through the formation of reactive intermediates that bind to DNA and result in activating mutations in key regulatory molecules.

Age and Sex Dependent Regulation of the Factor IX Gene in Mice

Plasma factor IX and liver factor IX mRNA levels in two normal mouse strains (B6D2F1 and BALB/CJNIA) were determined in relation to aging and sex of the animals. With male B6D2F1 mice, mean plasma factor IX activity levels for the 14 and 21 approximately 22 month-old animals were found to be 124% and 226%, respectively, of the 5 month-old group. Similarly, liver factor IX mRNA levels for the same age animal groups were 145% and 227%, respectively, of the reference group. Mean plasma factor IX levels for the same age female animals were 132% and 175%, respectively, and were accompanied by similarly elevated liver factor IX mRNA levels, 119 and 175%, respectively, of the 5 month-old female group. Factor IX activity and mRNA levels for the 5, 14 and 21 approximately 22 month-old female animal groups were lower than those of the corresponding male age groups by 25, 20 and 37%, and 20, 36 and 38%, respectively. With BALB/CJNIA mice, similar correlation was observed between the advancing age and substantial elevations in the factor IX mRNA level as well as on the unequal factor IX mRNA levels in females and males. These results indicate that the plasma factor IX level in both male and female mice is greatly elevated with aging, in general agreement with a similar phenomenon observed for human populations, and that this increase is due to a similar elevation in the factor IX mRNA level in the liver. In mice, both factor IX activity and mRNA levels are significantly higher in males than in females, which has not been described for humans.

Levels of serum steroids, aromatase activity, and estrogen receptors in preoptic area, hypothalamus, and amygdala of B6D2F1 male house mice that differ in the display of copulatory behavior after castration.

Most male B6D2F1 hybrid house mice continue to copulate after castration (continuers), whereas others do not (noncontinuers). Copulation in continuers appears estrogen dependent. Serum testosterone (T), 17 beta-estradiol (E2), and dihydrotestosterone (DHT), as well as aromatase activity (AA) and estrogen receptor (ER) levels in preoptic area (POA), hypothalamus (HYP), and amygdala (AM) were measured to determine if continuers and noncontinuers differ in estrogen physiology. In general, continuers and noncontinuers did not differ in serum steroid levels, AA, or ER levels. Castration reduced AA in the POA, HYP, and AM. Castration did not affect nuclear ER levels in the POA and HYP but reduced nuclear ER in AM. The data demonstrate that castrated B6D2F1 male mice continue to be under the influence of circulating nongonadal E2 that is important for copulation.

Effects of rhG-CSF, 5-fluorouracil and extramedullary irradiation on murine megakaryocytopoiesis in vivo.

The aim of this study was to systematically characterize possible rhG-CSF effects on the murine megakaryocyte-platelet system (untreated and recovering from chemotherapy or extramedullary irradiation). In untreated, splenectomized male B6D2F1 mice, rhG-CSF treatment (50 micrograms/kg/d for up to 8 d) markedly decreased femoral megakaryocytopoiesis. CFU-Meg, small acetylcholinesterase-positive (SAChE) cells, and megakaryocytes were significantly reduced to 35-70%; platelets, however, were not affected. Peripheral CFU-Meg and CFU-GM increased up to 200-fold. Following a single injection of 5-FU (150 mg/kg) on day 0, rhG-CSF (50 micrograms/kg/d) on days 1-8 suppressed the megakaryocytopoietic recovery as indicated by significantly lower platelet numbers on day 9. Granulopoietic recovery was accelerated by rhG-CSF. When rhG-CSF treatment was started on day 5, no beneficial effect on granulopoietic recovery was observed, but again platelet levels were significantly lower on day 9, indicating that within the first 4 d of rhG-CSF application, recruitment or lineage competition was not a critical event. To test for the effects of extramedullary irradiation on circulating progenitors, mice pretreated with 50 micrograms/kg/d of rhG-CSF for 8 d received irradiation to the chest with 500 cGy resulting in a substantial kill of circulating CFU-Meg and CFU-GM of up to 99%. However, this striking decrease of blood progenitors did not significantly affect their total body contents. This study indicates that rhG-CSF treatment can impair bone marrow megakaryocytopoiesis, which might be an important consideration for those clinical situations that carry a high potential for treatment-induced thrombocytopenia.

Circadian changes in mitoxantrone toxicity in mice: Relationship with plasma pharmacokinetics

Circadian time-dependent differences in clinical toxicity of several anti-cancer agents were predicted from murine studies. Mitoxantrone is an anthracenedion (an anthracycline-related class of compounds) of increased clinical use, which may benefit from selective circadian timing. In 3 consecutive studies, a total of 428 male B6D2F1 mice aged from 8 to 10 weeks were synchronized by an alternation of 12 hr of light and 12 hr of darkness (LD 12:12). They received a single i.v. injection of mitoxantrone at one of 6 or 4 circadian stages differing by 4 or 6 hr. A dose-response relationship characterized body-weight loss and survival rate. Dose-toxicity relationship further closely depended upon circadian dosing time. Thus, a dose of 16 mg/kg killed 100% of the mice injected at 3 hr after light onset (HALO), and none of them at 11 or at 15 HALO (p from chi 2 < 0.001). Body-weight loss varied from 41% at 3 HALO to 32% at 15 HALO (p from ANOVA < 0.0001). Least hematologic toxicity and fastest recovery of a normal circulating leukocyte count corresponded to mitoxantrone injection near the middle of the dark-activity span, at 16 HALO. Similar findings characterized colonic and splenic lesions. Moreover, mitoxantrone was both distributed and eliminated faster after injection at 16 HALO. If such data apply to cancer patients, as was the case for other drugs investigated with this methodology, an afternoon infusion should enable high-dose mitoxantrone to be well tolerated.

Effects of Carotenoid Administration on Bladder Cancer Prevention

Bladder cancer was induced in male B6D2F1 strain mice by the administration of N-butyl-N-(4-hydroxybutyl)nitrosamine. Mice supplemented with beta-carotene for 5 weeks before receiving the carcinogen and maintained on beta-carotene for an additional 26 weeks developed significantly fewer tumors than did unsupplemented mice. Mice receiving canthaxanthin for the same time period showed no protection against the development of bladder cancer.

Inhibition of Tumor Growth and Metastasis by Chronic Intravenous Infusion of Prostaglandin E1

The role of prostaglandins and their synthesis inhibitors in malignant disease is undefined. The following studies were done to determine the effects of continuous intravenous prostaglandin E1 (PGE1) or a prostaglandin synthesis inhibitor, indomethacin, on tumor growth and metastasis in mice bearing Lewis lung carcinoma. Male B6D2F1 mice underwent tumor implantation in the right axilla on day 0. After 10 days of tumor growth, mice underwent intravenous (IV) catheterization and were infused with either PGE1 at 3 micrograms/kg/minute (PG-LOW), PGE1 at 6 micrograms/kg/minute (PG-HIGH), indomethacin (INDO) at 1 microgram/kg/minute, or normal saline (NS). After 10 days of infusion, tumor volume, tumor weight, and the number of metastases greater than 2 mm in diameter were significantly decreased, and tumor doubling time was significantly prolonged in the PG-HIGH group compared to NS controls. None of the other experimental groups showed differences in these parameters. A second experiment with a similar experimental design was done infusing PGE1 at 6 micrograms/kg/minute and at 12 micrograms/kg/minute to determine the maximum dose response of IV PGE1. Again a decrease in tumor volume, tumor weight, and metastatic rates were identified when compared to saline control, but there were no significant difference between the two doses of PGE1.

Pharmacological and toxicological properties of arotinoids SMR-2 and SMR-6 in mice

Studies were conducted to define primary pharmacological and toxicological properties of two arotinoids, SMR-2 and SMR-6, in male B6D2F1 mice. Mice were gavaged daily for up to 22 days with retinoids in corn oil (0.1, 0.2, or 0.4 mg/kg day SMR-2 or SMR-6 or 2.5, 10, or 30 mg/kg all- trans-retinoic acid as a reference control). Toxicological and biochemical endpoints were assayed after 8, 15, and 22 days. At toxic doses, i.e., those inducing weight loss, morphological changes were observed in skin, lymph nodes, spleen, bone marrow, liver, thymus, forestomach, adrenal, bone, and testes. Biochemical alterations included elevated serum alkaline phosphatase, corticosterone, and interleukins-1, -2, and -3. Additional immune alterations included increased responsiveness of spleen cells to both thymus-dependent and thymus-independent mitogens and increases in the total number of B cells in the spleen. At doses not inducting weight loss, target organ effects included the appearance of plasma cells and infiltration of polymorphonuclear cells in lymph nodes; myeloid cell hypercellularity in bone marrow; hematopoiesis in spleen; subacute inflammation in forestomach; and periportal cytoplasmic vacuolization in liver. At the low doses, SMR-2 resulted in decreased responsiveness of spleen cells to mitogens and SMR-6 caused increased responsiveness. SMR-6 also increased interleukin-1 and -2 production at low doses. Biochemical effects included reduced activities of liver aryl hydrocarbon hydroxylase (AHH) and soluble brain protein kinase C. Overall, the results suggest that leukcopoiesis and reduced liver AHH and reduced soluble protein kinase C activities are the primary and initial pharmacological and toxicological effects of retinoids.