Subchronic oral toxicity of turmeric and ethanolic turmeric extract in female mice and rats.

Subchronic oral toxicity of turmeric and ethanolic turmeric extract was studied in female Swiss mice and Wistar rats fed turmeric (0, 1 and 5%) and ethanolic turmeric extract (0, 0.05 and 0.25%) through diet for 14 and/or 90 days. The administration of a high dose of turmeric (5%) for longer duration (90 days) showed a significant reduction in body weight gain, alterations in absolute and/or relative liver weights, and hepatotoxicity i.e. focal necrosis or focal necrosis with regeneration both in mice and rats. In mice lower doses of turmeric i.e 0.2 or 1% for 14 days also showed hepatotoxicity and they were found to be more vulnerable to turmeric-induced hepatotoxicity than rats.

Lithium inhibits growth in a murine neural precursor cell line.

The influence of lithium on cell growth and cell viability was studied in short-term cultures of a neural precursor cell line (NT) developed from a murine teratocarcinoma. At very low concentrations ranging from 0.1 mM to 1 mM Li2CO3 (equivalent to therapeutic blood concentrations) there was no difference between untreated and treated cultures. 10 mM lithium (Li+) was found to be toxic with 33% of cell death, while there was inhibition of growth without cell death at concentrations of 2.5 mM and 5 mM of Li+. In experiments where 2.5 mM Li+ was added at the time of seeding, there was growth arrest on day 1 followed by recovery on day 2. Flow cytometric analysis revealed that cells treated with Li+ were blocked in S phase. At 5 mM concentration of Li+, the recovery occurred on day 3 and the plating efficiency was significantly low. The ability to form colonies in soft agar was reduced at 2.5 mM and 5 mM concentrations of Li+ to an equal extent. Thus, Li+ has growth inhibitory as well as anchorage-independent growth reducing effects. The NT cell line therefore would be a good model system to study the mechanism of teratogenic effect of Li+.

A neural precursor cell line derived from murine teratocarcinoma.

A cell line NT with phenotypic features of neural precursor cells has been established from an embryo-derived teratocarcinoma in Swiss mouse where, on serial transplantation, the developmental potential becomes restricted to neural pathway. All the cells are positive for nestin (a marker of neuroepithelial stem cells). Many of them are also positive for NFP and/or GFAP. Moreover there is a gradual decrease from 75% to 50% in reactivity for alkaline phosphatase, a marker for EC cells with repeated passages. The bipotential nature, and the probable decline of EC cells suggest that NT is a neural precursor cell line. The cells have doubling time of 12 h with a plating efficiency of 50%. The cells form colonies in soft agar within 7 days and tumorigenicity in syngeneic mice is lost after 70th passage. However, after 70 passages cells do form tumors in nude mice within 5 days and these tumors exhibit better differentiated morphology than the tumors in syngeneic mice. All the other characteristics remain stable. The myc and ras family of oncogenes do not show any alterations in early or late passages. This cell line may therefore be considered as a differentiated cell line derived from teratocarcinoma.

A teratocarcinoma-derived neurotrophic activity.

In a search for nerve growth factor (NGF) in tissue extracts of a murine transplantable teratocarcinoma that harbours immature neural tissue in abundance, a trypsin-sensitive and heat labile neurotrophic activity was identified. The final protein fraction obtained by cation exchange chromatography contained five proteins with mol. wts ranging from 52 to 72 kD. It supported the growth and differentiation of immature neurones in neonatal rat cerebellar cultures but had no effect on embryonic chick dorsal root ganglia which are the classical targets for NGF.

Carcinogenic potential of Indian alcoholic beverage (country liquor).

Epidemiological studies reveal that alcohol consumption is a risk factor for the cancer of the mouth, larynx, esophagus and various other organs. Of the various alcoholic beverages consumed in India, country liquors are widely consumed and that too by the economically weak section of the society. The present paper describes the experiments designed to investigate the effect of one brand of country liquor from Maharashtra State, India (which was found to be more potent in our earlier mutagenicity studies) for its carcinogenicity in two strains of mice and Syrian golden hamsters. The experimental animals received 10% liquor in drinking water from 2 months of age for 16 months. One percent ethanol treated animals served as positive controls. Together with long term bioassays, the transplacental carcinogenic effect of country liquor in the offspring of treated mothers, as well as in the breeders themselves was also investigated. Pregnant mothers were fed 10% liquor through drinking water from 12th day of gestation till weaning of the progeny. Then offspring were allowed to live without further treatment and mothers continued to get liquor treatment. In long term bioassays, liquor caused 22% total tumor incidence in male BALB/c mice and 28% in male Swiss mice. In female Swiss mice and in hamsters, liquor did not show any pronounced effect on tumor incidence. Similar negative results were obtained in case of offspring of treated mothers, but the offspring of liquor treated mothers had higher mortality prior to weaning as compared to those of untreated mothers.

Carcinogenicity studies of tobacco extract in vitamin A-deficient Sprague-Dawley rats.

Long-term carcinogenicity studies were carried out in male Sprague-Dawley rats maintained on vitamin A-sufficient (SLO+) and vitamin A-deficient (SLO-) diets and treated with tobacco extract (TE). Three-week-old rats received by gavage a total dose of 860 mg of TE at a daily dose of 3 mg/rat over a period of 21 months. Besides tumorigenicity, drug-metabolizing phase I and phase II enzymes in lung and liver as well as vitamin A and C levels in plasma and liver were measured at 12 and 21 months of age. The cumulative tumor incidence in TE-treated SLO- rats was significantly higher (77-100%) than that observed in TE-treated SLO+ rats (20-22%). Furthermore, SLO+ rats treated with TE showed lung and forestomach tumors, whereas TE-treated SLO- rats showed a preponderance of pituitary adenomas (87%). It was observed that TE treatment increased the activity of the hepatic and pulmonary phase I enzymes and decreased the glutathione/glutathione S-transferase detoxification system at both time points in SLO- rats. On TE treatment the vitamin A levels in the liver and plasma were significantly decreased with a concurrent increase in vitamin C levels. The data show that a vitamin A-deficient diet renders male Sprague-Dawley rats more susceptible to TE treatment than the vitamin A-sufficient diet, an effect which was associated with the augmented induction of P-450 content and activities and depletion of the glutathione/glutathione S-transferase pathway by TE.

Enhancing effect of malachite green on the development of hepatic pre-neoplastic lesions induced by N-nitrosodiethylamine in rats.

The effects of malachite green (MG) and phenobarbitone (PB) were compared on the development of pre-neoplastic lesions during N-nitrosodiethylamine(DEN)-induced hepatocarcinogenesis in male Wistar rats. Rats were administered 200 p.p.m. DEN in drinking water for a period of 1 month. After an interval of 2 weeks the animals were given either MG (25 p.p.m.) or PB (500 p.p.m.) in drinking water for 2.5 months. The effects were monitored on the basis of the morphological appearance of the liver, histological pattern, gamma-glutamyltranspeptidase (GGT)-positive foci, total GGT activity and the induction of glycogen-deficient islands. Both MG and PB were found to enhance liver carcinogenesis to a significant extent when compared with either their corresponding controls or animals given DEN alone. The enhancing effect of MG at 25 p.p.m. is comparable with PB at 500 p.p.m. An enhancing effect of MG on DEN-induced hepatocarcinogenesis in the rats was demonstrated.

Peripheral nerve sheath tumours: an ultrastructural study of 30 cases.

This report deals with the ultrastructural observations of 30 peripheral nerve sheath tumours [PNST], which include 25 schwannomas of acoustic nerve, one schwannoma of cauda equina, one neurofibroma from a case of Von Recklinghausen's disease, one pigmented neurofibroma of spinal nerve root and a malignant schwannoma of frontal region. Interdigitating slender cytoplasmic processes covered with a continuous layer of basal lamina constitute the single most important ultrastructural attribute of Schwann cells. Myelin formation was encountered in the cell processes of four out of 25 acoustic schwannomas. In four cases Microtubular arrays identical to that in an axon were seen in Schwann cells. These two observations require further support by additional cases of PNST studies by electron microscopy. The neurofibroma consisted only of Schwann cells and no ultrastructurally identifiable perineurial cells or fibroblasts were detected. The cells in the pigmented neurofibroma revealed submicroscopic features of both Schwann cell and melanocyte, indicating their common ancestry. A unique case of malignant schwannoma arising from frontal meninges is illustrated and it is emphasized that electron microscopy is mandatory for a correct histogenetic diagnosis of malignant tumours which occur at unexpected anatomical sites.

Carcinogenicity studies of masheri: pyrolysed tobacco product, in vitamin-A-deficient Sprague Dawley rats.

The carcinogenicity of long-term feeding of masheri extract to animals in a vitamin-A-sufficient (SLO+) and deficient (SLO-) state was studied in Sprague Dawley rats by feeding daily dose of 3 mg extract over a period of 21 months. The phase I activating enzymes, the glutathione (GSH)/glutathione S-transferase (GST) detoxification system, and the hepatic and circulating levels of vitamins A and C were also monitored at 12 and 21 months. It was observed that the phase I enzyme activities were significantly higher in SLO+ than in SLO- rats at both 12 months and 21 months. Moreover, the SLO- masheri-treated animals also showed a decreased in the GSH/GST detoxification system while the reverse was observed in SLO+ group. Masheri extract treatment significantly lowered the hepatic and circulating levels of vitamin A while a concurrent increase was observed in the vitamin C level. The extract was found to be tumorigenic in both the SLO+ and SLO- groups. Benign tumours were observed in the SLO+ group while a high incidence of malignant tumours of the lung were observed in the SLO- group upon treatment with masheri extract.

Glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase activities in normal canine mammary gland and in mammary tumours and their correlation with oestrogen receptors.

Glucose-6-phosphate dehydrogenase (G-6-PD) and 6-phosphogluconate dehydrogenase (6-PGD) activities were studied in 10 normal canine mammary glands and in 19 canine mammary tumours, which included seven benign and 12 malignant neoplasms. The malignant tumours were also examined for oestradiol receptors. The mean G-6-PD and 6-PGD activities were 5.17 +/- 1.84 and 1.65 +/- 0.64 IU per g protein in normal glands, 8.8 +/- 2.7 and 3.8 +/- 0.99 IU per g protein in benign and 19.6 +/- 5.2 and 8.5 +/- 2.1 IU per g protein in malignant mammary tumours. The enzyme activities were significantly higher in tumour tissue than in normal glands. Malignant tumours had much greater activity than benign. With the increasing anaplasia, there was a corresponding rise in the activity of the two enzymes. There was an inverse correlation between the oestrogen receptor (ER) status and the enzyme activity. The ER-positive tumours had low and ER-negative tumours had high enzyme activities.

Megaoesophagus in the mouse: histochemical and ultrastructural studies.

Histochemical and ultrastructural studies of the muscle coat of the oesophagus from ICRC/HiCri mice (with megaoesophagus) and DBA/2fNCri mice (normal oesophagus) were carried out. The striking observation from histochemical studies was the presence of smooth muscle in the abdominal segment of the oesophagus from ICRC mouse in contrast to the control strain where smooth muscle was present only in the lowermost portion adjoining the stomach. Ultrastructural studies of the oesophageal wall from 5- and 10-day-old ICRC mice revealed an apparently normal muscle coat. In 3-month-old ICRC mice the upper abdominal segment of the oesophagus showed several abnormalities of smooth muscle fibres and paucity of plexus tissue accompanied by interstitial collagen deposition. The abnormalities were more severe in 1-year-old animals and were seen throughout the abdominal segment. From this study it is suggested that the primary cause of megaoesophagus in ICRC mice is neurogenic and not myogenic.

Comparative pathology of canine mammary tumours.

The pathological features of 52 canine mammary tumours were studied and compared with those of human breast neoplasms. In many of the former, the constituent cells were both epithelial and myoepithelial. Cartilaginous and osseous metaplasia of the stromal tissue was striking in a few tumours. Carcinosarcomas were also encountered. On a careful search, virus-like particles were observed in five tumours. The similarities and differences between the human and canine mammary tumours were discussed with a view to assessing the suitability of dog mammary tumour as an animal model for human breast tumours.

Carcinogenicity studies on the two tobacco-specific N-nitrosamines, N'-nitrosonornicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone.

The tobacco-specific N-nitrosamines (TSNA) have been implicated in oral cancer. However, except for one study using rats, no study has shown the ability of TSNA in inducing oral tumours in experimental animals. We have studied the carcinogenic potentials of N'-nitrosonornicotine (NNN) and 4-methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in mice and hamsters, wherein the nitrosamines were administered on the tongues of the mice and the cheek pouches of the hamsters to simulate the exposure conditions of humans. It was observed that in Swiss and BALB/c male mice, both NNN and NNK induced tumours of lung, forestomach and liver. However, no oral tumours were induced in mice. The effect of vitamin A depletion was tested in Swiss male mice. It was found that a low vitamin A status did not alter the percentage incidence of tumours induced by both nitrosamines to a significant extent. In the studies using Syrian golden hamsters, long-term treatment of NNK to hamster cheek pouch induced tumours in the lung, liver, stomach and cheek pouch. Subsequently, the effect of hydrogen peroxide (H2O2) on NNK-induced carcinogenicity in hamsters was studied. It was observed that simultaneous administration of NNK and H2O2 to the animals increased the incidence of cheek pouch tumours. Another pertinent observation was that even when a small initiator dose of NNK was given followed by the application of H2O2, a very significant increase in the tumour incidence was observed. This observation suggests that H2O2 could act as a promoter to NNK-induced carcinogenesis. In conclusion it may be stated that both NNN and NNK do not show any strain or species specificity. They failed to produce tumours at the site of application in mice but in hamsters few cheek pouch tumours were seen or were induced when NNK was applied alone. The cheek pouch tumour incidence increased when H2O2 was given concurrently or when applied for a long period after a low initiator dose of NNK was administered in the cheek pouch.

Anticarcinogenic effect of betel leaf extract against tobacco carcinogens.

Epidemiological studies have implicated that betel quid offers some protection to tobacco induced carcinogenesis. Earlier studies in our laboratory have shown betel leaf extract (BLE) to be antimutagenic against standard mutagens and tobacco-specific N'-nitrosamines (TSNA), N'-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). In the present study, we have tested the anticarcinogenic effect of BLE using Swiss male mice. Two protocols of study were used to test this effect. In the first protocol, the effect of BLE was tested against the standard carcinogen benzo[a]pyrene (BP) using Wattenberg's stomach tumor model, Cancer Res., 41 (1981) 2820-2823. In this protocol, BLE inhibited the tumorigenicity of BP to a significant extent. In the second protocol, the effect of BLE against the two tobacco-specific nitrosamines, NNN and NNK was studied using long-term studies on Swiss male mice. The nitrosamines were administered on the tongues of the mice, while the BLE was supplied in drinking water. Two doses of NNN (22 mg and 72 mg) and one dose of NNK (22 mg) were used. In this study, it was observed that the number of tumor bearing animals decreased, but the difference was significant only in the group treated with the low dose of NNN in combination with BLE. However, in all the BLE treated animals, irrespective of the dose of nitrosamine, the hepatic vitamin A and C levels were elevated significantly as compared to the corresponding nitrosamine-treated controls. These results indicate that BLE has a promising anticarcinogenic role to play in tobacco induced cancer.(ABSTRACT TRUNCATED AT 250 WORDS)

Diethylnitrosamine and acetoxymethyl methylnitrosamine-induced carcinogenesis in mice and vitamin A deficiency.

The modulating influence of vitamin A deficiency on carcinogenesis induced by two potent carcinogens, diethylnitrosamine (DEN) and acetoxymethyl methylnitrosamine (AMMN), was studied in BALB/c mice. DEN was administered intragastrically every 30 days at a total dose of 200 mg/kg body weight, split into four doses. AMMN was applied continuously every 14 days on the tongue, at a dose of 2 mg/kg body weight. AMMN and DEN treated animals fed the vitamin A deficient diet had a significantly higher tumor incidence that mice fed the normal diet (p less than 0.05). Studies on the levels of vitamins A, C, B2 and folic acid in the liver and plasma of mice treated with the two carcinogens revealed that both the carcinogens increased vitamin C in both tissues, decreased folic acid and had no effect on vitamin A, while hepatic vitamin B2 was lowered by treatment with AMMN by not by DEN.

Rat renal mesenchymal tumor as an experimental model for human congenital mesoblastic nephroma: I. Induction.

The effect of low-dose (2 Gy) radiation on ethylnitrosourea (ENU)-induced neoplasms was studied in Sprague-Dawley and Holtzman strains of rats. With a 60 mg/kg dose of ENU administered on day 1 in Sprague-Dawley rats, 18.4% of the neoplasms induced were found in the kidney. When the same dose of ENU was given on day 10, the incidence of kidney tumors fell to 2.8%. Prior (2 Gy) radiation on day 9 enhanced kidney tumor induction to 16.1%, a trend also observed in the case of ENU-induced neural tumors. In Holtzman rats, 40 mg/kg ENU induced more kidney tumors (12.5%) when given on day 4 than on day 0, and prior irradiation enhanced the ENU-induced kidney tumors even though the interval between irradiation and carcinogen administration was fairly long--4 days.

Rat renal mesenchymal tumor as an experimental model for human congenital mesoblastic nephroma: II. Comparative pathology.

The morphological features of kidney neoplasms induced by ethylnitrosourea (ENU) with or without prior irradiation were examined with a view to comparing them with human renal tumors of childhood. The rat renal tumors consisted of poorly differentiated, highly mitotic mesenchymal cells frequently attempting to differentiate along fibroblastic and myofibroblastic lineages. Though the host renal tubules were frequently entrapped in these tumors, immature tubules and islands of epithelial cells occasionally were seen to form an integral part of the neoplasms. Rarely, adenomatous areas surrounded by mesenchymal proliferation were observed. None of the tumors had the blastemal component that is the hallmark of nephroblastoma. The rat mesenchymal tumors resembled the human congenital mesoblastic nephroma.

Weak carcinogenic effect of masheri, a pyrolysed tobacco product, in mouse skin tumor models.

Two commonly used varieties of masheri, pyrolysed tobacco products, were tested for their skin carcinogenicity in Swiss mice and the more sensitive strain of Swiss bare mice. In Swiss mice, painting of brown and black varieties of masheri extract did not show any tumorgenic effect; however, a marginal synergistic effect of 7,12-dimethylbenz[a]anthracene (DMBA) and black masheri extract was observed when DMBA was used as an initiator. In Swiss bare mice, black masheri extract induced tumors in 20%-35% animals at both the doses tested. In an initiation/promotion protocol with DMBA as an initiator, induction of tumors in 50%-52% Swiss bare mice and a slight synergistic effect of black masheri extract were observed with a low dose of DMBA, suggesting a synergistic effect.

Carcinogenicity studies of masheri, a pyrolysed product of tobacco.

The carcinogenicity of two commonly used brown and black varieties of masheri, a pyrolysed tobacco product, was studied by feeding the masheri through the diet at a 10% level to three different animal species of both sexes. In Sprague-Dawley rats, only brown masheri was used, while in Swiss mice and Syrian golden hamsters both varieties were used. In all the three species, forestomach papillomas were induced as a result of masheri treatment. In rats, 37% of animals showed forestomach papillomas while in mice and hamsters the incidence was 42-47% and 25-43%, respectively. No malignant changes were observed in any of the groups except 2/23 male hamsters showed forestomach carcinoma in the black masheri diet group.