Bromocriptine and suppression of postpartum lactation

The use of bromocriptine for the suppression of postpartum lactation drew widespread attention to a potentially increased risk for cardiovascular and cerebrovascular adverse effects. To estimate the incidence of these events a follow-up study was performed among 2,130 women of 15–44 years of age who were treated with a course of bromocriptine in 1990–1992. None of these women were admitted to the hospital for cardiovascular or cerebrovascular events. However, the incidence of pregnancy hypertension and the use of cardiovascular drugs increased considerably in the last 2 months before delivery. Therefore, cardiovascular or cerebrovascular events can probably be explained by pre-existing morbidity rather than by the use of bromocriptine. We estimated, using a ‘worst-case’ analysis, that fewer than 2 Dutch women each year would develop serious cardiovascular or cerebrovascular events.     

Counteraction by combined treatment with high dose of oestrogen and somatostatin of mammary growth suppression in mice

The interaction between somatostatin (SMS) and high doses of oestrogen on mammary gland growth was examined in C3H/He virgin mice. Mammary gland growth was significantly suppressed by the subcutaneous implantation of pellets of oestradiol benzoate diluted to 1/1000 or 1/500 or by injection twice daily of 50ng SMS 201-995 between 25 and 55 days of age. However, the mammary gland growth of mice receiving SMS and oestrogen in combination was markedly stimulated compared to that of mice given the respective agents. These results indicate that the inhibitory effects of somatostatin and oestrogen on mammary growth were apparently counteracted by the treatment in combination.     

Effect of ethanol/arrack on the lipid metabolism of mammary gland during pregnancy and lactation in rats

Female rats were exposed to arrack (12.0 ml/kg body weight/day) and ethanol (4.0 g/kg body weight/day) before conception and throughout gestation and lactation. On 19th day of gestation and 21st day of lactation there was increase in the cholesterol phospholipids, triglycerides and free fatty acids in the mammary gland of rats administered arrack/ethanol in comparison with the controls. The lipoprotein lipase activity showed significant increase in the treated groups, in which the activity decreased on 21st day in comparison with 19th day. The absolute and relative weight of mammary gland also showed a significant decrease in ethanol/arrack treated group. The biochemical alterations produced in the mammary gland by arrack and its equivalent alcohol were different showing that non-alcoholic portion of arrack interferes with the toxicity induced by alcohol. Arrack was found to be a potent hyperlipidemic agent than ethanol.     

Puromycin's suppression of memory in mice as affected by caffeine.

It has previously been shown that expression of maze-learning in mice is blocked for long periods of time by puromycin injected intracerebrally one or more days after the training experience. Treatment with caffeine after training has now been found to reduce greatly the amnestic effects of puromycin. With a high dose of caffeine (200 mg/kg) this reduction is evident 6 days after treatment with puromycin. With a lower dose of caffeine (25 mg/kg) the effect becomes evident only after a more extended period of time. In view of control experiments, we suggest that caffeine modifies factors necessary for the expression of memory and that this alteration makes puromycin relatively ineffective in blocking memory.     

Charcoal tattooing of mammary carcinoma in mice

We describe a step in the development of a charcoal suspension for tattooing human mammary carcinomas prior to surgery and chemotherapy. Two vehicles (injectable water and 0.9% NaCl) were compared. Migration of the charcoal particles was studied in vitro (gel model) and in vivo (intratumoral injection into nude mice). The charcoal suspension in injectable water was more effective in the detection of tumors.     

Folinic acid protects against suppression of growth by methotrexate in mice.

The objective of this study was to investigate whether folinic acid supplementation would protect young mice against suppression of growth by methotrexate (MTX). Four equal groups of Balb/c young male mice (5 animals in each group; mean+/-SD body weight 9.64+/-0.85 g, in their rapid growth phase) were subjected to the following drug treatment: One group was given MTX (3.5 mg/kg body weight) intraperitoneally on every 2nd day, another received folinic acid (7.0 mg/kg body weight) intraperitoneally every 2nd day. The third group was given both of these drugs (MTX on every 2nd day and folinic acid 8 h post-MTX injection). The fourth group was injected with physiological saline every other day to serve as a control group. Total body weight, food and water consumption by animals in each group were monitored every second day for a period of 3 weeks. After this period mice were sacrificed and liver, spleen and kidneys were excised, weighed and analyzed for MTX and dihydrofolate reductase activity. A small segment of the proximal part of small intestine and small pieces of liver and kidney were also removed to study morphological changes. Compared to the groups, which received folinic acid alone, folinic acid plus MTX or physiological saline, mean increase in body weight (6.8+/-0.8 g) of mice over a period of 3 weeks was minimal in the group receiving MTX alone (one-way ANOVA p=0.0001). The mean weights of liver and kidney in this group receiving MTX alone were also found to be significantly less than the mean weights of these organs in the 3 groups (p<0.001). The negative effect on growth of animals appears not only due to malabsorption but inhibition of pathway of de novo DNA synthesis may also be involved. This is supported by loss of villous pattern in small intestine of mice treated with MTX alone and increased accumulation of free MTX and decreased dihydrofolate reductase in the liver of the group receiving MTX alone as compared with the group receiving MTX plus folinic acid. The data indicate that the administration of folinic acid protects mice against suppression of growth by MTX. On the basis of these observations it can be deduced that patients suffering from juvenile rheumatoid arthritis or acute lymphoblastic leukaemia receiving MTX over a long period of time might be at a risk of experiencing short-term suppression of growth, however they could benefit from supplementation with folinic acid.     

Bisphosphonates. Pharmacology and use in the treatment of tumour-induced hypercalcaemic and metastatic bone disease.

The geminal bisphosphonates are a new class of drugs characterised by a P-C-P bond. Consequently, they are analogues of pyrophosphate, but are resistant to chemical and enzymatic hydrolysis. The bisphosphonates bind strongly to hydroxyapatite crystals and inhibit their formation and dissolution. This physicochemical effect leads in vivo to the prevention of soft tissue calcification and, in some instances, inhibition of normal calcification. The main effect is to inhibit bone resorption, but in contrast to the effect on mineralisation, the mechanism involved is cellular. These various effects vary greatly according to the structure of the individual bisphosphonate. The half-life of circulating bisphosphonates is very brief, in the order of minutes to hours. 20% to 50% of a given dose is taken up by the skeleton, the rest being excreted in the urine. The half-life in bone is far longer and depends upon the turnover rate of the skeleton itself. Bisphosphonates are very well tolerated; the relatively few adverse events that have been associated with their use are specific for each compound. Bisphosphonates have been used to treat various clinical conditions, namely ectopic calcification, ectopic bone formation, Paget's disease, osteoporosis and increased osteolysis of malignant origin. The three compounds commercially available for use in tumour-induced bone disease are in order of increasing potency, etidronate, clodronate and pamidronate. Most data have been obtained with the latter two agents. By inhibiting bone resorption, they correct hypercalcaemia and hypercalciuria, reduce pain, the occurrence of fractures, as well as the development of new osteolytic lesions, and in consequence improve the quality of life. In view of these actions, of their excellent tolerability and of the fact that they are active for relatively long periods, these compounds are, after rehydration, the drugs of choice in tumour-induced bone disease and an excellent auxiliary to the drugs used in oncology.     

Cadmium pathways during gestation and lactation in control versus metallothoinein 1,2-knockout mice.

Effects of metallothionein (MT) on cadmium absorption and transfer pathways during gestation and lactation in mice were investigated. Female 129/SvJ metallothionein-knockout (MT1,2KO) and metallothionein-normal (MTN) mice received drinking water containing trace amounts of (109)CdCl(2) (0.15 ng Cd/ml; 0.074 micro Ci (109)Cd/ml). (109)Cd and MT in maternal, fetal, and pup tissues were measured on gestation days 7, 14, and 17 and lactation day 11. In dams, MT influenced both the amount of (109)Cd transferred from intestine into body (two- to three-fold higher in MT1,2KO than MTN dams) and tissue-specific (109)Cd distribution (higher liver/kidney ratio in MT1,2KO dams). Placental (109)Cd concentrations in MT1,2KO dams were three- and seven-fold higher on gestation days 14 and 17, respectively, than in MTN dams. Fetal (109)Cd levels were low in both mouse types, but at least 10-fold lower in MTN fetuses. MT had no effect on the amount of (109)Cd transferred to pups via milk; furthermore, 85-90% of total pup (109)Cd was recovered in gastrointestinal tracts of both types, despite high duodenal MT only in MTN pups. A relatively large percentage of milk-derived intestinal (109)Cd was transferred to other pup tissues in both MT1,2KO and MTN pups (14 and 10%, respectively). These results demonstrate that specific sequestration of cadmium by both maternal and neonatal intestinal tract does not require MT. Although MT decreased oral cadmium transfer from intestine to body tissues at low cadmium exposure levels, MT did not play a major role in restricting transfer of cadmium from dam to fetus via placenta and to neonate via milk.     

Cadmium-induced disturbances in lactating mammary glands of mice

Previous studies in various species have demonstrated that cadmium levels in milk are low and that cadmium is retained in the mammary glands of lactating rodents. The objective of the present study was to examine mammary glands following cadmium exposure during peak lactation. Mice were given 5 microg, 100 microg or 2000 microg cadmium/kg body weight subcutaneously on lactation days 8-10 and mammary glands were dissected on lactation day 11 for histology, quantitative gene expression and mineral analyses. Cadmium exposure induced morphological changes in the lactating mammary gland. A remodelling of the lactating mammary tissue including an increase in fat content, a less active feature of the mammary alveolar epithelial cells and more condensed appearance of the milk alveoli were observed. Although these changes were most prominent in the animals exposed to the highest cadmium dose similar morphological alterations were indicated at the two lower doses. A significant negative dose-response relationship between beta-casein gene expression and exposure of cadmium was demonstrated, while mRNA levels of alpha-lactalbumin were not affected by cadmium treatment. Furthermore, reduced levels of calcium in the mammary glands of the dams exposed to the highest cadmium dose and a positive correlation between calcium and beta-casein were demonstrated. No other differences were detected among the cadmium dose groups in mammary levels of calcium, zinc, iron or copper. In conclusion, our results indicate that cadmium disturbs the function of the lactating mammary gland, which consequently may impair the development of the suckling offspring.     

Role of glutathione and reactive oxygen intermediates in 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced immune suppression in C57Bl/6 mice.

Recent developments in basic immunology have revealed the importance of glutathione (GSH) and cellular redox balance in the generation of an immune response. In the liver, it has been shown that exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) alters cellular GSH and reactive oxygen intermediate (ROI) production. We have tested the hypothesis that TCDD mediates the suppression of the cytotoxic T lymphocyte (CTL) response to alloantigen by increasing oxidative stress. Total cellular GSH, GSSG, and GSH-protein adducts were analyzed by HPLC. Changes in intracellular GSH and ROI were simultaneously measured in isolated hepatocytes and individual subpopulations of spleen cells (CD4+, CD8+, B220+, and Mac-1+) following in vivo exposure to TCDD and antigenic challenge with P815 mastocytoma cells. Monochlorobimane was utilized to measure GSH levels, and two fluorescent probes were used to evaluate ROI levels: dichlorofluoroscein diacetate to monitor peroxides and dihydroethidine to assess superoxide anion. In hepatocytes, in vivo treatment with TCDD resulted in a transient, 2-fold increase in GSH, a 50% decrease in peroxide levels and a small (20-40%) decrease in superoxide anion levels. Although alloantigen challenge resulted in increased GSH and peroxide in spleen cells, in vivo exposure to TCDD had no effect on splenic ROI levels, nor did it consistently alter GSH levels in any subpopulation of spleen cells examined. Moreover, in vivo treatment with the antioxidant N-acetyl cysteine failed to affect the immune suppression caused by TCDD. These results suggest to us that although TCDD perturbs cellular redox balance in the liver, it does not exacerbate or diminish the normal increased GSH and ROI which occur in the spleen in response to antigenic challenge.     

Parathion-induced suppression of humoral immunity in inbred mice

Numbers of IgM plaque-forming cells (PFC) were reduced by 65% in C57B1/6 mice given parathion 2 days after immunization with sheep red blood cells (SRBC). The immunosuppressive dose (16 mg/kg, p.o.) caused signs of cholinergic poisoning and 20% mortality. Survivors appeared to have recovered fully from the cholinergic crisis at the time of immunologic assay. However, these animals had reduced tissue cholinesterase (ChE) activities and decreased numbers of nucleated spleen cells. Immunosuppression was apparent on day 4 but not on day 3 or days 5–8 of the primary IgM response. Reduction of serum hemagglutinin titers coincided with reduction of the number of splenic PFC. A lower dose of insecticide (4 mg/kg, p.o.) did not produce signs of poisoning and was not immunosuppressive. The number of 8-day IgG PFC was reduced by 45% when parathion (16 mg/kg, p.o.) was given 6 days after immunization, but not when parathion was given 2 days after immunization. The data suggest that cholinergic stimulation and/or the associated toxic chemical stress may be involved in parathion-induced immunosuppression.     

Histamine in C3H/W mice carrying spontaneous tumors of the mammary gland

Adenocarcinoma mammae, a spontaneously growing mammary cancer in C3H/W mice, contrary to many transplanted tumors does not evoke any rise in histamine level either in the tumor or in distant tissues. On the other hand, the histamine level is reduced by 90% in the tumor in comparison with the healthy gland. This seems to be a consequence of the fall of histidine decarboxylase activity to below a detectable level. There is also a significant reduction in histamine N-methyltransferase activity to one-fifth of the control level. The healthy mammary gland contains a high concentration of histamine and catabolizes it exclusively through the methylation pathway.     

Inhibition of lactation.

The mechanism and hormonal regulation of lactation is explained and illustrated with a schematic representation. Circulating estrogen above a critical amount seems to be the inhibitory factor controlling lactation during pregnancy. Once delivery occurs, the level of estrogen falls, that of prolactin rises, and lactation begins. Nonsuckling can be used to inhibit lactation. Estrogens can also be used to inhibit lactation more quickly and with less pain. The reported association between estrogens and puerperal thromboembolism cannot be considered conclusive due to defects in the reporting studies. There is no reason not to use estrogens in lactation inhibition except for women over 35 who experienced a surgical delivery. Alternative therapy is available for these women. The recently-developed drug, brom-ergocryptine, may replace other methods of lactation inhibition.     

A role played by sigma receptors in the conditioned suppression of motility in mice

Mice exhibited marked suppression of motility (conditioned suppression) when placed in the same environment in which they had previously received an electric foot-shock. The conditioned suppression was attenuated by cyclazocine and N-allylnormetazocine (SKF-10047), sigma agonists. The effect of these drugs was reduced by chronic pretreatment with cyclazocine. This behavioral change was related to the change in binding activity of [³H]-phencyclidine to sigma receptors (defined using non-radioactive SKF-10047). In the chronic vehicle-pretreated conditioned suppression group, the K_d and B_max values of [³H]-phencyclidine binding at the high affinity site were increased when compared to those in the chronic vehicle-pretreated control group. The increased values were restored to the control levels by acute treatment with cyclazocine and SKF-10047. On the other hand, in the chronic cyclazocine-pretreated conditioned suppression group, acute cyclazocine and SKF-10047 treatment failed to change the increased values of K_d and B_max at the high affinity site. The present behavioral and receptor-binding experiments suggest that the activation of nervous system mediated by sigma receptors may be responsible for the attenuation of conditioned suppression of motility.     

Chemotherapy of mammary carcinomas arising in ras transgenic mice

Transgenic female mice carrying the V-Ha-ras transgene linked to the MMTV promoter, which developed mammary carcinomas, were treated with selected cancer chemotherapy drugs. Agents were administered i.p. on a daily × 9 schedule when mice developed tumors that were 50–100 mg in size. Drugs which are clinically effective against breast cancer were quite efficacious in the transgenic model at their maximum tolerated dose. Doxorubicin produced excellent responses in tumor-bearing transgenic mice, with several mammary carcinomas undergoing tumor shrinkage. Two anthrapyrazoles, DuP 937 and DuP 941, novel anticancer drugs with phase 2 activity against breast cancer, were as effective as doxorubicin in the oncomice. Mitoxantrone, a synthetic agent with some properties similar to the anthracyclines, also had antitumor activity, but not as pronounced as obtained with doxorubicin or the anthrapyrazoles. Cisplatin, a drug with limited use in human breast cancer, only caused modest antitumor responses. A computerized data analysis method based on the area under the tumor growth curve was developed to better quantitate the data and provide statistical information. This quantitative analysis confirmed the high statistical significance of the activity of doxorubicin or the anthrapyrazoles in the ras transgenic model, and defined an excellent dose response relationship for each drug tested. Our results suggest that the ras transgenic model may be useful for identifying drugs that have efficacy for breast cancer in women.Abbreviations MMTV Mouse Mammary Tumor Virus - AUC Area Under Curve - MGR Mean Growth Rate     

Gastrointestinal absorption of cadmium in mice during gestation and lactation: II. Continuous exposure studies

The effect on cadmium retention of continuous exposure to drinking water containing low levels of cadmium during pregnancy and lactation was studied in mice. Female mice were provided drinking water ad libitum containing ¹⁰⁹CdCl₂ (0.03 μCi ¹⁰⁹Cd/ml, 0.11 ppb total cadmium) throughout either gestation, lactation, or a combined period of pregnancy and lactation. Nonpregnant control mice were exposed to the same cadmium solution for similar time periods. Dams in all three experimental groups retained two to three times more cadmium (expressed as percentage of ingested dose) than did nonpregnant controls. The ¹⁰⁹Cd contents of liver, kidney, mammary tissue, and duodenum increased strikingly in all three groups. Increases in kidney and mammary tissue were particularly apparent during lactation, with increases of fivefold for kidney and at least ninefold for mammary tissue, compared to levels in nonpregnant controls. Increases in ¹⁰⁹Cd retention by the duodenum were fivefold during gestation and three- to fourfold during lactation. The kidneys of dams exposed during lactation retained 53% of the whole body ¹⁰⁹Cd, while kidneys of nonpregnant controls retained only 27%. Results indicate that pregnant and lactating mice absorb and subsequently retain substantially more cadmium from their diets than do nonpregnant mice.     

Studies on the molecular pharmacology of GR63178A. A novel pentacyclic pyrolloquinone anticancer drug.

GR63178A (NSC D611615) is the second pentacyclic pyrolloquinone to be evaluated clinically as an anticancer drug. Its mechanism of action is unknown but may be related either to its quinone group or planar ring system. In this report we have investigated the ability of GR63178A to bind non-covalently to DNA, inhibit topoisomerase II and undergo reduction to reactive free radical species. Using two DNA duplexes, a 12-mer oligonucleotide which is a preferred sequence for minor groove binders and a hexamer which is a preferred sequence for intercalators, no evidence of significant binding with GR63178A was found. Neither GR63178A nor GR54374X (its 9-hydroxy metabolite) inhibited purified human topoisomerase II in a decatenation assay. Free radical chemistry was studied by both pulse radiolysis and ESR spectroscopy as well as by in vitro drug incubations with NADPH-fortified rat liver microsomes and purified cytochrome P450 reductase. The one-electron reduction potential of GR63178A was -207 mV +/- 10 which is much more positive than other quinone-containing anticancer drugs such as doxorubicin, mitomycin C and mitozantrone. GR63178A underwent enzyme-catalysed quinone reduction more readily than doxorubicin but produced significantly fewer reactive oxygen species. No evidence was detected of drug-induced, radical-mediated DNA damage in vitro using pBR322 plasmid DNA. Disproportionation of the GR63178A semi-quinone free radical proceeded with a rate constant of 1 x 10(9) M-1 sec-1 under anaerobic conditions, one order of magnitude faster than doxorubicin. The preferential disproportionation of the semi-quinone may explain our inability to detect a free radical signal by ESR. The hydroquinone of GR63178A was stable and exhibited strong visible absorption with a bathochromic shift of 120 nm over the parent drug. These unusual properties may be due to the hydroquinone undergoing a form of keto-enol tautomerization. Thus, GR63178A free radical formation does not appear to result in significant drug activation. In conclusion, GR63178A is unlikely to mediate its antitumour activity by DNA binding, topoisomerase II inhibition or free radical formation in direct contrast to similar anthracycline- and anthraquinone-based anticancer drugs.