Overexpression of 5-lipoxygenase and its relation with cell proliferation and angiogenesis in 7,12-dimethylbenz(α)anthracene-induced rat mammary carcinogenesis.

The present study was performed to investigate the critical role of 5-lipoxygenase (5-LOX) in 7,12-dimethylbenz(α)anthracene (DMBA)-induced rat mammary inflammation associated carcinogenesis. Female Sprague-Dawley rats at 50 days of age were treated with 7,12-dimethylbenz(α)anthracene (DMBA; 0.5 mg/100 g body weight) by a single tail vein injection, followed by administration of zileuton (2000 mg/kg diet) from week 7 until the termination of the study at 31 wk. 5-LOX protein expression, 5-hydroxyeicosatetraenoic acid (5-HETE), and leukotriene B4 (LTB4 ) production in rat mammary tissue were analyzed at 6, 12, and 24 wk post-DMBA injection. Rate of cell proliferation was analyzed by bromodioxyuridine labeling index (BrdU-LI). Microvessel density, level of VEGF, and MMP-2 were also measured. DMBA induces inflammation in rat mammary gland as early as 6 wk. 5-LOX is upregulated in DMBA treated rats right from 6 wk when compared with their normal counterparts. An overexpression of 5-LOX is accompanied with increase in 5-HETE, LTB4 production and high BrdU-LI with an increase of two key angiogenic factors for tumorigenesis; MMP-2 and VEGF. It was found that 5-LOX specific inhibitor brought about substantial protection against DMBA-induced mammary carcinogenesis. Histological findings showed substantial repair of hyperplastic lesions. There was a significant reduction in the rate of cell proliferation and expression of angiogenic factors, MMP-2 and VEGF. 5-LOX plays an important role in DMBA-induced inflammation associated carcinogenesis via activation of MMP-2 and VEGF. 5-LOX expression can be considered as a critical event in controlling the process of mammary tumor development.

Biological activity of carotenoids: its implications in cancer risk and prevention.

Recently nontoxic natural compounds are getting immense importance for the prevention of diseases of different etiology. Natural product provitamin A "carotenoids", largely α-carotene, ß-carotene, and ß-cryptoxanthin, are typical constituents of orange/red/yellow colored fruits and green vegetables. Different in vitro and in vivo studies have shown that carotenoids possess the capacity to scavenge DNA damaging free radicals, suppress angiogenesis, inhibit cell proliferation and induce apoptosis. Epidemiological reports of case-control studies, nested case-control studies, and cohort studies support significant association between dietary intake and circulating levels of carotenoids and reduction in cancer risk/carcinoma of various organs. However, randomized trials regarding ß-carotene supplementation, alone or in combination with other supplements, have not always well corroborated with this. Of seven trials, one observed a significant benefit on cancer mortality, four reported no significant benefit or harm, while the remaining two trials found an unexpected, but significant increase in lung cancer incidence. This review discusses implications and significance of carotenoids in the field of cancer risk and prevention.

Role of 5-lipoxygenase in resveratrol mediated suppression of 7,12-dimethylbenz(α)anthracene-induced mammary carcinogenesis in rats.

Substantial evidences suggest that lipoxygenase-catalyzed products have a strong influence on the development and progression of human cancers. The dietary phytochemical resveratrol has become a focus of intense research owing to its roles in cancer prevention. A single tail vein injection of 7,12-dimethylbenz(a)anthracene (DMBA) was given at a dose of 0.5mg/0.2 ml oil emulsion/100g body weight at 50 days of age of female Sprague-Dawley rats. Rats were treated with resveratrol from 2 weeks before DMBA injection (5 weeks of animal age) and continued to 24 weeks of the experimentation at a dose of 100 µg/rat in the diet. We observed that resveratrol acts as a potent 5-lipoxygenase (5-LOX) inhibitor obtained from natural sources. Our result indicated that resveratrol is a strong antioxidant in reducing lipid peroxidation and preventing DNA damage. It significantly decreased the extent of DNA strand break, inhibited abnormal cell proliferation as evidenced by BrdU labeling index and also induced apoptosis in carcinogen-challenged rat mammary tissue. Increased TGF-ß1 expression in resveratrol treated rats is thought to be one of the factors inducing apoptosis to suppress DMBA-induced mammary carcinogenesis.

A review of metallothionein isoforms and their role in pathophysiology.

The Metallothionein (MT) is a protein which has several interesting biological effects and has been demonstrated increase focus on the role of MT in various biological systems in the past three decades. The studies on the role of MT were limited with few areas like apoptosis and antioxidants in selected organs even fifty years after its discovery. Now acknowledge the exploration of various isoforms of MT such as MT-I, MT-II, MT-III and MT-IV and other isoforms in various biological systems.Strong evidence exists that MT modulates complex diseases and the immune system in the body but the primary function of MT still remains unknown. This review's main objective is to explore the capability to specifically manipulate MT levels in cells and in animals to provide answers regarding how MT could impact those complex disease scenarios.The experimental result mentioned in this review related among MT, zinc, cadmium, diabetic, heart disease, bone retardation, neuro toxicity, kidney dysfunction, cancer, and brain suggest novel method for exploration and contribute significantly to the growing scientist to research further in this field.

Combined supplementation of vanadium and fish oil suppresses tumor growth, cell proliferation and induces apoptosis in DMBA-induced rat mammary carcinogenesis.

The anti-cancer activity of vanadium and fish oil has been shown in a large number of studies. This study was undertaken to analyze the combined effect of vanadium and fish oil on 7,12-dimethylbenz(α)anthracene (DMBA)-induced mammary carcinogenesis in female Sprague-Dawley rats. The whole experiment was divided into three parts: (1) DNA strand breaks study, (2) morphological analysis, and (3) histological and immunohistochemical study. Rats were treated with DMBA (0.5 mg/0.2 ml corn oil/100 g body weight) by a tail vein injection. Rats received vanadium (w/v) as ammonium monovanadate at a concentration of 0.5 ppm (4.27 µmol/L) in the drinking water and given ad libitum and/or fish oil (0.5 ml/day/rat) by oral gavage. Histology, morphology, DNA strand breaks, cell proliferation, and apoptosis of the mammary tissue were assessed in this study. Treatment with vanadium or fish oil alone significantly reduced the DNA strand breaks, palpable mammary tumors, tumor multiplicity, and cell proliferation but the maximum protection effect was found in the group that received both vanadium and fish oil and the combination treatment offered an additive effect (P < 0.05). Furthermore, vanadium and fish oil significantly increased the TUNEL-positive apoptotic cells (P < 0.05) but the increase was maximal with combination treatment and had an additive effect. These results affirm the benefits of administration of vanadium and fish oil in the prevention of rat mammary carcinogenesis which was associated with reduced DNA strand breaks, palpable mammary tumors and cell proliferation and increased TUNEL-positive apoptotic cells.

Combinatorial effect of fish oil (Maxepa) and 1alpha,25-dihydroxyvitamin D(3) in the chemoprevention of DMBA-induced mammary carcinogenesis in rats.

The present study demonstrates the anti-tumor effects of combined supplementations of dietary fish oil (Maxepa) and 1alpha,25-dihydroxyvitamin D(3) (vitamin D(3)) on 7,12-dimethylbenz(alpha)anthracene (DMBA)-induced rat mammary carcinogenesis. Female Sprague-Dawley rats at 50 days of age were treated with 7,12-dimethylbenz(alpha)anthracene (DMBA; 0.5mg/100g body weight) by a single tail vein injection in an oil emulsion. Both fish oil (rich in EPA and DHA) and vitamin D(3) were administered orally at a dose of 0.5 ml/day/rat and 0.3 microg/100 microL propylene glycol twice a week respectively and continued to 35 weeks after DMBA administration. Fish oil in combination with vitamin D(3) resulted in a significant reduction in incidence, multiplicity and volume of mammary tumors. These supplementation also inhibited DMBA-induced mammary 7-methylguanine DNA adducts formation, which was measured by HPLC-fluorescence assay (at four sequential time points; ANOVA, F=42.56, P<0.0001). Immunohistochemical analysis revealed that the effect of fish oil and vitamin D(3) occurred through suppression of cell proliferation (BrdU-LI: P<0.0001). Fish oil and vitamin D(3) together also reduced the mRNA expression of iNOS (84%, P<0.05). In view of their natural availability, non-toxicity and acceptability; combined supplementation of fish oil and vitamin D(3) might be effective for chemoprevention of mammary carcinogenesis.

Suppression of the inflammatory cascade is implicated in resveratrol chemoprevention of experimental hepatocarcinogenesis.

PURPOSE: Resveratrol, present in grapes and red wine, has been found to prevent diethylnitrosamine (DENA)-initiated rat liver tumorigenesis, though the chemopreventive mechanisms are not completely elucidated. The current study was designed to explore whether the antiinflammatory properties of resveratrol play a role in its antihepatocarcinogenic action. METHODS: Liver samples were harvested from a 20-week chemopreventive study in which resveratrol (50, 100 and 300 mg/kg) was shown to inhibit DENA-induced hepatocyte nodules in Sprague-Dawley rats in a dose-responsive manner. Hepatic preneoplastic and inflammatory markers, namely heat shock protein (HSP70), cyclooxygenase-2 (COX-2) and nuclear factor-kappaB (NF-kappaB), were studied using immunohistochemical as well as Western blot techniques. RESULTS: Resveratrol dose-dependently suppressed DENA-induced increased expressions of hepatic HSP70 and COX-2. Resveratrol also attenuated the DENA-mediated translocation of NF-kappaB p65 from the cytosol to the nucleus with stabilization of inhibitory kappaB. CONCLUSION: The present findings indicate that resveratrol exerts chemoprevention of hepatocarcinogenesis possibly through antiinflammatory effects during DENA-evoked rat liver carcinogenesis by suppressing elevated levels of HSP70, COX-2 as well as NF-kappaB. These beneficial effects combined with an excellent safety profile encourage the development of resveratrol for chemoprevention and intervention of human HCC that remains a devastating disease.

Vanadium in the detection, prevention and treatment of cancer: the in vivo evidence.

Vanadium, a dietary micronutrient, is yet to be established as an essential part of the human diet. Over the past century, several biological effects of vanadium, such as insulin-mimetic action as well as amelioration of hyperlipidemia and hypertension, have been discovered. This transition element is known to influence a battery of enzymatic systems, namely phosphatases, ATPases, peroxidases, ribonucleases, protein kinases and oxidoreductases. Multiple biochemical and molecular actions of vanadium have been implicated in its inhibitory effects on various tumor cells of human origin. Successful in vitro studies over the past few decades have advanced the anticancer research on vanadium into the preclinical stage. Vanadium in several animal cancer models provides protection against all stages of carcinogenesis--initiation, promotion, and progression. This review focuses on the current advances in cancer prevention and treatment as well as early detection by vanadium compounds in preclinical animal models while pointing to possible mechanisms of such diverse beneficial effects. Clinical pharmacokinetic and potential toxicity studies on vanadium are also highlighted in this review. Supporting and challenging evidence as well as future directions of vanadium research exploring the possibility of using this dietary agent for detection, prevention and treatment of human cancers are critically discussed.

Fish oil regulates cell proliferation, protect DNA damages and decrease HER-2/neu and c-Myc protein expression in rat mammary carcinogenesis.

BACKGROUND & AIMS: The aim of this study is to assess the effect of dietary fish oil (MaxEPA) on DNA-strand breaks, cell proliferation and anti-apoptotic protein expressions in rat mammary carcinogenesis. METHODS: Eighty-one female Sprague-Dawley rats were divided into two parts, one for DNA-strand breaks study and the other for immunohistochemical study. Female Sprague-Dawley rats were treated with 7,12-dimethylbenz(alpha)anthracene (DMBA) (0.5 mg/0.2 ml corn oil/100 g body weight) by a tail vein injection. Rats were fed either fish oil or corn oil (0.5 ml/day/rat) by oral gavage. RESULTS: Fish oil-treated group showed significant protection against generation of single-strand breaks (SSBs) (56.1%, P < 0.05) but increased effect (72.3%, P < 0.05) was found in the corn oil-treated group when compared to DMBA control group. Furthermore, fish oil-treated group exhibited substantial decrease in Ki-67 (P < 0.05), HER-2/neu (P < 0.05) and c-Myc (P < 0.05) immunolabelling indices when compared to carcinogen counterpart. However, corn oil treatment resulted in significant increase in the above parameters. CONCLUSIONS: The above data support the role of n-3 PUFA as a preventive agent for DNA damages and a potential to inhibit mammary carcinogenesis.

Estrogen suppresses MLK3-mediated apoptosis sensitivity in ER+ breast cancer cells.

Little knowledge exists about the mechanisms by which estrogen can impede chemotherapy-induced cell death of breast cancer cells. 17beta-Estradiol (E(2)) hinders cytotoxic drug-induced cell death in estrogen receptor-positive (ER(+)) breast cancer cells. We noted that the activity of the proapoptotic mixed lineage kinase 3 (MLK3) kinase was relatively higher in estrogen receptor-negative (ER(-)) breast tumors, suggesting that E(2) might inhibit MLK3 activity. The kinase activities of MLK3 and its downstream target, c-Jun NH(2)-terminal kinase, were rapidly inhibited by E(2) in ER(+) but not in ER(-) cells. Specific knockdown of AKT1/2 prevented MLK3 inhibition by E(2), indicating that AKT mediated this event. Furthermore, MLK3 inhibition by E(2) involved phosphorylation of MLK3 Ser(674) by AKT, attenuating the proapoptotic function of MLK3. We found that a pan-MLK inhibitor (CEP-11004) limited Taxol-induced cell death and that E(2) accentuated this limitation. Taken together, our findings indicate that E(2) inhibits the proapoptotic function of MLK3 as a mechanism to limit cytotoxic drug-induced death of ER(+) breast cancer cells.

Caspase-mediated cleavage of beta-catenin precedes drug-induced apoptosis in resistant cancer cells.

A delicate balance between cell death and survival pathways maintains normal physiology, which is altered in many cancers, shifting the balance toward increased survival. Several studies have established a close connection between the Wnt/beta-catenin pathway and tumorigenesis, aberrant activation of which might contribute toward increased cancer cell growth and survival. Extensive research is underway to identify therapeutic agents that can induce apoptosis specifically in cancer cells with minimal collateral damage to normal cells. Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can induce apoptosis specifically in tumor cells, many cancer cells develop resistance, which can be overcome by combinatorial treatment with other agents: for example, peroxisome proliferator-activated receptor gamma (PPARgamma) ligands. To identify the molecular target mediating combinatorial drug-induced apoptosis, we focused on beta-catenin, a protein implicated in oncogenesis. Our results show that co-treatment of TRAIL-resistant cancer cells with TRAIL and the PPARgamma ligand troglitazone leads to a reduction of beta-catenin expression, coinciding with maximal apoptosis. Modulation of beta-catenin levels via ectopic overexpression or small interference RNA-mediated gene silencing modulates drug-induced apoptosis, indicating involvement of beta-catenin in regulating this pathway. More in-depth studies indicated a post-translational mechanism, independent of glycogen synthase kinase-3beta activity regulating beta-catenin expression following combinatorial drug treatment. Furthermore, TRAIL- and troglitazone-induced apoptosis was preceded by a cleavage of beta-catenin, which was complete in a fully apoptotic population, and was mediated by caspases-3 and -8. These results demonstrate beta-catenin as a promising new target of drug-induced apoptosis, which can be targeted to sensitize apoptosis-resistant cancer cells.

Vesicular flavonoid in combating diethylnitrosamine induced hepatocarcinoma in rat model.

Reactive oxygen species (O2(*-), OH(-), H2O2) are known to play an important role in tumor initiation in hepatocarcinoma. Hepatocarcinoma was developed in the Swiss Albino rats by administration three doses of diethylnitrosamine (DEN) (200 mg/kg b. wt.) (i.p.) at 15 days interval. Quercetin (QC), herbal polyphenolic compound, is a potent anticancer drug. Clinical trials are difficult for its hydrophobic nature. To overcome this problem, our study was aimed to formulate soluble liver specific, galactosylated liposomal QC and to investigate its efficacy against hepatocarcinoma in rat model. Galactosylated liposomal QC was formulated and the suspension was introduced intravenously to rats (8.98 microM/kg) once in a week for 16 weeks. Hepatocarcinoma in rat model and its pathological improvement were evaluated histopathologically, histochemically and electron microscopically. Severe oxidative damage was noticed in the whole liver and its microsomal fraction of DEN treated rats. Huge numbers of hyperplastic nodules (HNs) with pre-neoplastic lesions appeared in rat liver by DEN administration. Galactosylated liposomal QC injections prevented DEN mediated development of hepatocarcinoma and oxidative damage in rat liver. Quercetin in liver specific galactosylated liposomal drug delivery system may be recommended as a potent therapeutic formulation against DEN-induced hepatocarcinoma.

Dietary fish oil associated with increased apoptosis and modulated expression of Bax and Bcl-2 during 7,12-dimethylbenz(alpha)anthracene-induced mammary carcinogenesis in rats.

The present study investigated the chemopreventive effect of dietary fish oil (Maxepa), rich in eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on induction of apoptosis in mammary carcinogenesis model. Mammary carcinogenesis was initiated by a single, tail vein injection of 7,12-dimethylbenz(alpha)anthracene (DMBA) (0.5mg/0.2ml corn oil/100g body weight) at 7 weeks of animal age. Ninety female Sprague-Dawley rats were divided into two parts: part one was used for histology and immunohistochemical study and part two for morphological analysis. Each part consists of three experimental groups having 15 animals, i.e., Group A (DMBA control), Group B (DMBA+fish oil) and Group C (DMBA+corn oil). Rats were fed either fish oil or corn oil (0.5ml/day/rat) by oral gavage, 2 weeks prior to DMBA injection. Treatment was continued 25 weeks, studying histopathology, expression of Bax and Bcl-2 proteins by immunohistochemistry and apoptosis by TUNEL assay and morphological study at 36 weeks. Results showed that the fish oil-treated group exhibited a substantial increase in Bax (p<0.05) immunolabelling and a reduction of Bcl-2 immunopositivity (p<0.05), and increased TUNEL-positive apoptotic cells (p<0.05); however, corn oil treatment did not show these beneficial effects toward mammary preneoplasia. We conclude that fish oil has the potential to play a significant role in limiting mammary tumourigenesis in vivo.

Vanadium and 1, 25 (OH)2 vitamin D3 combination in inhibitions of 1,2, dimethylhydrazine-induced rat colon carcinogenesis.

The current investigation demonstrates the antitumor effects of combined supplementations of vanadium (V) (4.27 micromol/L drinking water ad libitum) and 1alpha, 25-dihydroxy vitamin D(3) (Vitamin D(3)) (0.3 mug/100 muL propylene glycol per os twice a week) on 1, 2 dimethylhydrazine (DMH) (20 mg/kg body weight) induced rat colon carcinogenesis. There was a significant reduction in incidence (70%), multiplicity (P<0.0001) and volume (P<0.01) of colon tumors. HPLC-fluorescence assay detected the combinatorial actions of V and Vitamin D(3) against DMH-induced colonic O(6)-methylguanine DNA adducts formation (at four sequential time points; ANOVA, F=13.56, P<0.01). Simultaneous inhibition of DNA single strand breaks (P<0.001) indicates the potency of the combination regimen in limiting the initiation event of colon carcinogenesis. Immunohistochemical analysis revealed that the effect of V and vitamin D(3) occurred through suppression of cell proliferation (BrdU-LI: P<0.001) along with an induction of apoptosis (TUNEL-LI: P<0.01). The immunoexpression of tumor suppressor p53 and downregulation of antiapoptotic protein BCl-2 in subsequent immunofluorescence assay further provide strong evidence for the combinatorial inhibitory actions of vanadium and vitamin D(3) against DMH-induced rat colon carcinogenesis.

Protective effects of vanadium against DMH-induced genotoxicity and carcinogenesis in rat colon: removal of O(6)-methylguanine DNA adducts, p53 expression, inducible nitric oxide synthase downregulation and apoptotic induction.

Previous studies have shown that dietary micronutrient vanadium can protect neoplastic development induced by chemical carcinogens. Current investigation is an attempt to evaluate the role of vanadium (4.27 micro mol/l) in inhibiting 1,2 dimethyhydrazine (DMH) (20 mg/kg body weight) induced rat colon carcinogenesis. We investigated the effect of vanadium against the formation of DMH-induced O(6)-methylguanine (O(6)-Meg) DNA adduct, a potent cytotoxic and mutagenic agent for colon cancer. Supplementation of vanadium significantly reduced the hepatic (P<0.05), and colonic (at three sequential time points; ANOVA, F=4.96, P<0.05) O(6)-Meg DNA adduct levels in rats, indicating vanadium's potency in limiting the initiation event of colon carcinogenesis. Removal of initiated and damaged precancerous cells by apoptosis can prevent tumorigenesis and further malignancy. DNA fragmentation study revealed the vanadium-mediated apoptotic induction in colon tumors. The increased value of apoptotic index (AI) (62.27%; P<0.01) in subsequent TUNEL assay further confirmed the apoptosis induction by vanadium. This paralleled the nuclear immunoexpression of p53. A significant positive correlation between p53 immunoexpression and AI (P=0.0026, r=0.83, r(2)=0.69) links its association with vanadium-mediated apoptotic induction. Vanadium treatment also abated the mRNA expression of iNOS (54.03%), reflecting its protective effect against nitric oxide-mediated genotoxicity and colon tumorigenesis. These studies cumulatively provide strong evidence for the inhibitory actions of vanadium against DMH-induced genotoxicity and carcinogenesis in rat colon.

Acanthus ilicifolius plant extract prevents DNA alterations in a transplantable Ehrlich ascites carcinoma-bearing murine model.

AIM: To investigate the chemopreventive efficacy of the Indian medicinal plant Acanthus ilicifolius L Acanthaceae in a transplantable Ehrlich ascites carcinoma (EAC)-bearing murine model. METHODS: Male Swiss albino mice were divided into four groups: Group A was the untreated normal control; Group B was the EAC control mice group that received serial, intraperitoneal (ip) inoculations of rapidly proliferating 2 x 10(5) viable EAC cells in 0.2 mL of sterile phosphate buffered saline; Group C was the plant extract-treated group that received the aqueous leaf extract (ALE) of the plant at a dose of 2.5 mg/kg body weight by single ip injections, once daily for 10, 20 and 30 consecutive days following tumour inoculation (ALE control); and Group D was the EAC + ALE-treatment group. The chemopreventive potential of the ALE was evaluated in a murine model by studying various biological parameters and genotoxic markers, such as tumour cell count, mean survival of the animals, haematological indices, hepatocellular histology, immunohistochemical expression of liver metallothionein (MT) protein, sister-chromatid exchanges (SCEs), and DNA alterations. RESULTS: Treatment of the EAC-bearing mice with the ALE significantly (P < 0.001) reduced viable tumour cell count by 68.34% (228.7 x 10(6) +/- 0.53) when compared to EAC control mice (72.4 x 10(6) +/- 0.49), and restored body and organ weights almost to the normal values. ALE administration also increased (P < 0.001) mean survival of the hosts from 35 +/- 3.46 d in EAC control mice to 83 +/- 2.69 d in EAC + ALE-treated mice. Haematological indices also showed marked improvement with administration of ALE in EAC-bearing animals. There was a significant increase in RBC count (P < 0.001), hemoglobin percent (P < 0.001), and haematocrit value (P < 0.001) from 4.3 +/- 0.12, 6.4 +/- 0.93, and 17.63 +/- 0.72 respectively in EAC control mice to 7.1 +/- 0.13, 12.1 +/- 0.77, and 30.23 +/- 0.57 respectively in EAC + ALE-treated group, along with concurrent decrement (P < 0.001) in WBC count from 18.8 +/- 0.54 in EAC control to 8.4 +/- 0.71 in EAC + ALE. Furthermore, treatment with ALE substantially improved hepatocellular architecture and no noticeable neoplastic lesions or foci of cellular alteration were observed. Daily administration of the ALE was found to limit liver MT expression, an important marker of cell proliferation with concomitant reduction in MT immunoreactivity (62.25 +/- 2.58 vs 86.24 +/- 5.69, P < 0.01). ALE was also potentially effective in reducing (P < 0.001) the frequency of SCEs from 14.94 +/- 2.14 in EAC control to 5.12 +/- 1.16 in EAC + ALE-treated group. Finally, in comparison to the EAC control, ALE was able to suppress in vivo DNA damage by abating the generations of 'tailed' DNA by 53.59% (98.65 +/- 2.31 vs 45.06 +/- 1.14, P < 0.001), and DNA single-strand breaks (SSBs) by 38.53% (3.14 +/- 0.31 vs 1.93 +/- 0.23, P < 0.01) in EAC-bearing murine liver. CONCLUSION: Our data indicate that, ALE is beneficial in restoring haematological and hepatic histological profiles and in lengthening the survival of the animals against the proliferation of ascites tumour in vivo. Finally, the chemopreventive efficacy of the ALE is manifested in limiting MT expression and in preventing DNA alterations in murine liver. The promising results of this study suggest further investigation into the chemopreventive mechanisms of the medicinal plant A. ilicifolius in vivo and in vitro.

Suppression of early stages of neoplastic transformation in a two-stage chemical hepatocarcinogenesis model: supplementation of vanadium, a dietary micronutrient, limits cell proliferation and inhibits the formations of 8-hydroxy-2'-deoxyguanosines and DNA strand-breaks in the liver of sprague-dawley rats.

Previous studies from our laboratory have demonstrated the potential anticarcinogenicity of vanadium, a dietary micronutrient in rat liver, colon, and mammary carcinogenesis models in vivo. In this paper, we have investigated further the antihepatocarcinogenic role of this essential trace element by studying several biomarkers of chemical carcinogenesis with special reference to cell proliferation and oxidative DNA damage. Hepatocarcinogenesis was induced in male Sprague-Dawley rats by chronic feeding of 2-acetylaminofluorene (2-AAF) at a dose of 0.05% in basal diet daily for 5 days a week. Vanadium in the form of ammonium metavanadate (0.5 ppm equivalent to 4.27 micromol/l) was supplemented ad lib to the rats. Continuous vanadium administration reduced relative liver weight, nodular incidence (79.99%), total number and multiplicity (P < 0.001; 68.17%) along with improvement in hepatocellular architecture when compared to carcinogen control. Vanadium treatment further restored hepatic uridine diphosphate (UDP)-glucuronosyl transferase and UDP-glucose dehydrogenase activities, inhibited lipid peroxidation, and prevented the development of glycogen-storage preneoplastic foci (P < 0.01; 63.29%) in an initiation-promotion model. Long-term vanadium treatment also reduced BrdU-labelling index (P < 0.02) and inhibited cell proliferation during hepatocellular preneoplasia. Finally, short-term vanadium exposure abated the formations of 8-hydroxy-2'-deoxyguanosines (P < 0.001; 56.27%), length:width of DNA mass (P < 0.01), and the mean frequency of tailed DNA (P < 0.001) in preneoplastic rat liver. The study indicates the potential role of vanadium in suppressing cell proliferation and in preventing early DNA damage in vivo. Vanadium is chemopreventive against the early stages of 2-AAF-induced hepatocarcinogenesis in rats.

Glycogen synthase kinase-3beta induces neuronal cell death via direct phosphorylation of mixed lineage kinase 3.

Mixed lineage kinase 3 (MLK3) is a mitogen-activated protein kinase kinase kinase member that activates the c-Jun N-terminal kinase (JNK) pathway. Aberrant activation of MLK3 has been implicated in neurodegenerative diseases. Similarly, glycogen synthase kinase (GSK)-3beta has also been shown to activate JNK and contribute to neuronal apoptosis. Here, we show a functional interaction between MLK3 and GSK-3beta during nerve growth factor (NGF) withdrawal-induced cell death in PC-12 cells. The protein kinase activities of GSK-3beta, MLK3, and JNK were increased upon NGF withdrawal, which paralleled increased cell death in NGF-deprived PC-12 cells. NGF withdrawal-induced cell death and MLK3 activation were blocked by a GSK-3beta-selective inhibitor, kenpaullone. However, the MLK family inhibitor, CEP-11004, although preventing PC-12 cell death, failed to inhibit GSK-3beta activation, indicating that induction of GSK-3beta lies upstream of MLK3. In GSK-3beta-deficient murine embryonic fibroblasts, ultraviolet light was unable to activate MLK3 kinase activity, a defect that was restored upon ectopic expression of GSK-3beta. The activation of MLK3 by GSK-3beta occurred via phosphorylation of MLK3 on two amino acid residues, Ser(789) and Ser(793), that are located within the C-terminal regulatory domain of MLK3. Furthermore, the cell death induced by GSK-3beta was mediated by MLK3 in a manner dependent on its phosphorylation of the specific residues within the C-terminal domain by GSK-3beta. Taken together, our data provide a direct link between GSK-3beta and MLK3 activation in a neuronal cell death pathway and identify MLK3 as a direct downstream target of GSK-3beta. Inhibition of GSK-3 is thus a potential therapeutic strategy for neurodegenerative diseases caused by trophic factor deprivation.

Cell proliferation and hepatocarcinogenesis in rat initiated by diethylnitrosamine and promoted by phenobarbital: potential roles of early DNA damage and liver metallothionein expression.

Cell proliferation plays an important role in multistage chemical carcinogenesis. Again, several reports demonstrated that upregulation of metallothionein (MT) expression is associated with increased cell proliferation that may contribute to the pathogenesis of preneoplastic phenotype to frank malignancy. In this study, we evaluated the roles of early DNA damage, altered expressions of liver MT and Ki-67 nuclear antigen, and altered hepatic levels of zinc (Zn) and copper (Cu) on cell proliferation and the progression of hepatocarcinogenesis through premalignant, late premalignant and malignant transformation phases in male Sprague-Dawley rats. We have further studied the association between MT expression and cell proliferation in hepatocarcinogenesis. There was substantial induction of DNA single-strand breaks (SSBs) (P<0.001) and development of hepatocellular premalignant lesions along with significant decrease in hepatic levels of Zn and increase in Cu content following a single, necrogenic, intraperitoneal (i.p.) injection (200 mg/Kg body weight) of diethylnitrosamine (DEN) at week 4 of the experimental protocol. Moreover, DEN+phenobarbital (PB)-treatment significantly elevated MT-, Ki-67-, and BrdU-immunoexpressions along with their immunolabeling indices. Furthermore, positive correlations between MT- and Ki-67- labeling (P=0.0006) at various time intervals, as well as, between MT immunoreactivity and 5'-bromo-2'-deoxyuridine-labeling index (BrdU-LI) (P=0.0007) indicate that, MT expression might be associated with Ki-67 expression and cell proliferation thereby. The study suggests that DEN treatment may lead to alteration of Zn and Cu levels resulting in early DNA damage along with elevation of MT expression that may ultimately lead to hepatic cell proliferation. The results thus provide evidence in support of the role of MT as a potential positive regulator of cell growth during the early stages of hepatocellular transformation in rats.

Protective role of fish oil (Maxepa) on early events of rat mammary carcinogenesis by modulation of DNA-protein crosslinks, cell proliferation and p53 expression.

BACKGROUND: Fish oil is known to protect from many types of cancers of the colon, liver, breast, prostate and lung 123. The objective of the present study was to evaluate the role of fish oil [Maxepa, supplemented at a dose of 0.5 ml is equivalent to 90 mg eicosapentaenoic acid (EPA) and 60 mg docosahexaenoic acid (DHA)] on cell proliferation, expression of p53 tumor suppressor protein and DNA protein crosslinks (DPCs) in a defined model of chemical rat mammary carcinogenesis. Mammary carcinogenesis was initiated by a single, intravenous (i.v.) tail vein injection of 7,12 dimethylbenz(alpha)anthracene (DMBA) at a dose of 5 mg DMBA/2 ml corn oil/kg body weight in female Sprague-Dawley rats at 7 weeks of age. Fish oil supplementation was started daily, 2 weeks prior to DMBA injection and continued for 24 (31 weeks of animal age) weeks and 35 (42 weeks of animal age) weeks of post DMBA injection, for histopathological and immunohistochemical and for morphological studies, respectively. RESULTS: Our results indicate the chemopreventive effect of fish oil (Maxepa) on DMBA-induced rat mammary carcinogenesis. Administration of fish oil further showed a prominent reduction of cell proliferation (24.34%, P = 0.001); DPCs (25%, P < 0.001) and an increased expression of p53 protein (4.636 +/- 0.19, P < 0.001) in preneoplastic mammary tissue when compared to carcinogen control counterpart. Histopathological and morphological analyses were carried out as end-point biomarkers. CONCLUSION: Our study thus provides evidence for the anticarcinogenic effect of fish oil (Maxepa) in limiting mammary preneoplasia in Sprague-Dawley rats.