Expression of cyclooxygenase-2, p53 and Ki-67 in gastric cancer

It has been reported that p53 mutation may contribute to upregulate cyclooxygenase (COX)-2 expression that is observed in malignant tissues. These molecules are involved in carcinogenesis by affecting tumor cell proliferation. The aim of this study was to examine the relationship between COX-2 or p53 expression and clinico-pathological characteristics including tumor cell proliferation in gastric cancer. COX-2 and p53 expressions were investigated with immunostaining, in tissue specimens obtained from 119 patients who underwent surgery for gastric cancer. The Ki-67 labeling index (LI) was counted by Ki-67 immunostaining. COX-2 and p53 expressions correlated significantly with depth of tumor invasion. However, there was no association between COX-2 or p53 expression and survival. p53 expression did not correlate with COX-2 expression. There was no significant difference in various clinicopathological variables between Ki-67 LI subgroups. The mean Ki-67 LI value of COX-2 positive tumors was significantly higher than that of negative tumors. The mean Ki-67 LI value of p53 positive tumors was not significantly higher than that of negative tumors. The mean Ki-67 LI value of both COX-2 and p53 positive tumors was significantly higher than that of both negative tumors. These results imply that COX-2 expression is associated with tumor cell proliferation of gastric cancer.     

Chemoprevention of heterocyclic amine-induced mammary carcinogenesis in rats

2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is one of the most prevalent carcinogenic heterocyclic amines in the environment, targeting the colon, prostate, pancreas, and mammary gland in rodents. Chemopreventive effects of synthetic and naturally occurring compounds on PhIP-induced rat mammary carcinogenesis were investigated in a series of experiments. In a PhIP feeding model, groups of 20-21 female F344 rats each, were treated with 0.02% PhIP alone or PhIP plus 0.5% 1-O-hexyl-2,3,5-trimethylhydroquinone (HTHQ), 1% green tea catechins, 1% alpha-tocopherol, 0.1% ellagic acid, or 1% chlorophyllin, each in the diet, or 0.1% caffeine in drinking water for 52 weeks. To assess the mechanism of HTHQ and caffeine inhibition of PhIP-induced carcinogenesis, effects of these compound on the in vitro metabolic activation of PhIP were examined in the presence of S9 mix. In the next series of experiments, the PhIP intragastric dose model was applied to allow separate investigation of the effects of chemicals during the initiation and postinitiation periods. In these experiments, female Sprague-Dawley rats were given eight intragastric doses of 100 mg/kg body weight during the first 4-8 weeks for initiation. Either during initiation or after initiation, or only after initiation, animals were treated with either corn or perilla oil at doses of 5 and 20%, conjugated fatty acid derived from safflower oil (CFA-S) or perilla oil (CFA-P) at a dose of 1%, arctiin at doses of 0.02 and 0.2% in the diet, or sodium nitrite (NaNO(2)) at a dose of 0.2% in drinking water. In the PhIP feeding model, administration of PhIP alone for 52 weeks induced adenocarcinomas in 40% of rats, but the incidence was remarkably reduced to 5% by the simultaneous treatment with 0.5% HTHQ, a strong lipophilic phenolic antioxidant, or to 10% by 0.1% caffeine. Administration of 1% chlorophyllin exerted similar, albeit weaker, effects. alpha-Tocopherol at a dose of 0.5% only reduced the multiplicity of carcinomas, and 1% green tea catechins only the mean size of mammary tumors. In a metabolic activation study of PhIP, HTHQ and caffeine clearly inhibited the formation of metabolites. In the PhIP gastric dose model, among the naturally occurring compounds examined, a plant lignan arctiin, perilla oil, which contains a large amount of n-6 alpha-linolenic acid, and CFA-S or CFA-P inhibited mammary tumor development, particularly in the postinitiation period, although a clear dose response was not observed. Treatment with 0.2% NaNO(2) in the initiation period was found to lower the volume of mammary tumors. The present results indicate that a number of compounds may be candidate chemopreventive agents against PhIP-induced mammary carcinogenesis, acting through different mechanisms and depending on the stage of carcinogenesis.     

Cyclooxygenase-2 and c-erbB-2 expression in colorectal carcinoma assessed using tissue microarrays.

The c-erbB-2 and cyclooxygenase (COX) pathways are involved in the pathogenesis of colorectal carcinoma, but the relationship is not fully understood. This study evaluated the significance of c-erbB-2, COX-2, and Ki-67 protein expression in 185 patients with colorectal carcinoma using tissue microarrays. Only one case expressed cerbB-2 protein. COX-2 expression was noted in 166 of 176 cases (94.3%), and the Ki-67 expression rate averaged 5.9%. There was no relationship among c-erbB-2, COX-2, and Ki-67 protein expression, and COX-2 protein expression was not related to tumor stage, differentiation, size, depth of invasion, lymphatic or vascular invasion, or patient survival. While the contribution of c-erbB-2 to colorectal carcinogenesis may be of little quantity, COX-2 may be deeply involved in colorectal carcinogenesis.     

Chemopreventive effect of the non-psychotropic phytocannabinoid cannabidiol on experimental colon cancer

Colon cancer affects millions of individuals in Western countries. Cannabidiol, a safe and non-psychotropic ingredient of Cannabis sativa, exerts pharmacological actions (antioxidant and intestinal antinflammatory) and mechanisms (inhibition of endocannabinoid enzymatic degradation) potentially beneficial for colon carcinogenesis. Thus, we investigated its possible chemopreventive effect in the model of colon cancer induced by azoxymethane (AOM) in mice. AOM treatment was associated with aberrant crypt foci (ACF, preneoplastic lesions), polyps, and tumour formation, up-regulation of phospho-Akt, iNOS and COX-2 and down-regulation of caspase-3. Cannabidiol-reduced ACF, polyps and tumours and counteracted AOM-induced phospho-Akt and caspase-3 changes. In colorectal carcinoma cell lines, cannabidiol protected DNA from oxidative damage, increased endocannabinoid levels and reduced cell proliferation in a CB(1)-, TRPV1- and PPARγ-antagonists sensitive manner. It is concluded that cannabidiol exerts chemopreventive effect in vivo and reduces cell proliferation through multiple mechanisms.     

Reduced levels of the cell-cycle inhibitor p27Kip1 in epithelial dysplasia and carcinoma of the oral cavity.

Recent studies have shown that the cyclin-dependent kinase (cdk) inhibitors play important roles in cell cycle progression in normal cells. Alterations in the cdk inhibitors also appear to be important in cancer development in a number of human tumors. p27Kip1 is a member of the CIP/KIP family of cdk inhibitors that negatively regulates cyclin-cdk complexes. Reduced levels of p27Kip1 protein have been identified in a number of human cancers, and in some cases reduced p27Kip1 is associated with an increase in proliferative fraction. In the present study, we examined p27Kip1 protein by immunohistochemistry in 10 normal and 36 dysplastic epithelia and in 8 squamous cell carcinomas from one anatomical site within the oral cavity, the floor of the mouth. Proliferative activity was assessed in serial sections by determining the expression of the cell cycle proteins Ki-67 and cyclin A. p27kip1 protein was significantly reduced in oral dysplasias and carcinomas compared with that in normal epithelial controls. In addition, there was a significant reduction in p27Kip1 protein between low- and high-grade dysplasias, suggesting that changes in p27Kip1 expression may be an early event in oral carcinogenesis. There was increasing expression of Ki-67 and cyclin A proteins with increasingly severe grades of dysplasia compared with normal controls. Although there was a strong correlation between Ki-67 and cyclin A scores (r2= 0.61) for all categories of disease, there was a weak negative correlation between Ki-67 and p27Kip1 levels (r2 = 0.29) and between cyclin A and p27Kip1 levels (r2 = 0.25). In conclusion, this study has found that a reduction in the proportion of cells expressing p27Kip1 protein is frequently associated with oral dysplasia and carcinoma from the floor of the mouth. Furthermore, reductions in p27Kip1 levels are associated with increased cell proliferation, although other changes likely contribute to altered cell kinetics during carcinogenesis at this site.     

乳腺导管内增生性病变中ER、Ki-67和cyclin D1的表达

目的 探讨ER、Ki-67和cyclin D1在乳腺导管内增生性病变中的表达及意义。方法 采用免疫组化和免疫荧光双标记法对56例乳腺导管内增生性病变进行ER、 Ki-67和cyclin D1染色标记。结果 正常乳腺组织中仅有散在的少数上皮细胞呈ER阳性表达。在普通型导管增生（usual ductal hyperplasia,UDH）中ER表达比正常乳腺组织增加,但ER阳性细胞呈不连续分布,阳性细胞间有较多的阴性细胞。非典型性导管增生（atypical ductal hyperplasia,ADH）和低级别原位导管癌（ductal carcinoma in situ,DCIS）中ER表达比UDH明显增加（P〈0．05）,ER阳性细胞呈连续的片状分布,阳性细胞间较少或没有ER阴性细胞。ADH和低级别DCIS中ER表达较高级DCIS显著（P〈0．01）。DCIS中Ki-67和cyclin D1表达高于UDH（P〈0．05）,并与UDH、ADH和DCIS的组织学分组呈正相关（r=0．352,P〈0．05和r=0．390,P〈0．05）。正常乳腺组织中上皮细胞内无ER和Ki-67同时表达。在UDH中有极少数上皮细胞ER和Ki-67同时表达,而在ADH和DCIS中ER和 Ki-67同时表达的细胞明显增加。结论 从正常乳腺组织到UDH、ADH、低级DCIS的恶性转化过程中伴有ER表达的逐渐增高。ER过度表达及ER和Ki-67在上皮细胞内同时表达可能是某些乳腺癌发生过程中的早期事件。     

PTEN regulates apoptotic cell death through PI3-K/Akt/GSK3β signaling pathway in DMH induced early colon carcinogenesis in rat

Phosphatidylinositol 3-kinase (PI3-K) and Akt (protein kinase B), are both essential signaling molecules that are up-regulated in various cancers. Here, we examined the molecular mechanisms by which PI3-K and Akt expression are regulated by glycogen synthase kinase-3β (GSK-3β) and the phosphatase and tensin homolog deleted on chromosome 10 (PTEN) in the early stages of experimental colon carcinogenesis. 1,2-dimethylhydrazine (DMH) was utilized for the induction of colon cancer while piroxicam, a traditional non-steroidal anti-inflammatory drug and c-phycocyanin, a biliprotein from Spirulina platensis (cyanobacterium) as the chemopreventive agents. Western blotting and immunofluorescence results indicated that the expression of PI3-K and Akt was promoted in the DMH group while least apoptosis was detected in this group as analyzed by Hoechst 33342-propidium iodide co-staining. DMH group further detected lower GSK-3β and PTEN expression as compared to other groups. Piroxicam and c-phycocyanin treatment resulted significant apoptotic cell death while showing low PI3-K and Akt expressions. Mitochondrial membrane potential (ΔΨ(M)) alterations (examined by JC-1 and rhodamine 123 labeling of colonocytes) and fluorescence intensity measurement of ROS level, were also analyzed showing the raised ΔΨ(M) while reduced ROS levels in DMH group, however piroxicam and c-phycocyanin treatment resulted in falling of ΔΨ(M) although both stimulated the ROS production as analyzed by flow cytometry. The present study thus identified that piroxicam, a traditional NSAID and c-phycocyanin, a newly discovered COX-2 selective inhibitor, constitute remarkable chemopreventive targets in mediating apoptosis in the DMH induced early rat colon carcinogenesis via regulating PI3-K/Akt/GSK-3β/PTEN signaling pathways. Further, a combination of the two drugs provides a better therapeutic option, than the monotherapy regimen.     

Lack of chemopreventive efficacy of DL-selenomethionine in colon carcinogenesis

Epidemiologic observations and laboratory research have suggested that dietary selenium reduces the risk of colon cancer. Selenium-enriched brewer's yeast as a dietary supplement reduces the incidence of and mortality from cancer of the colon in humans. It is not clear whether the observed inhibitory effect is due to selenomethionine, or to other forms of selenium, or to a mixture of the selenium compounds present in selenium-enriched brewer's yeast. Therefore, bioassay described in this study examined the chemopreventive efficacy of 10 and 15 ppm selenomethionine, equivalent to 3.6 and 5.4 ppm as selenium, against azoxymethane (AOM)-induced colon carcinogenesis. At five weeks of age, groups of male F344 rats were fed diets containing 0 (control diet), 10 or 15 ppm selenomethionine. At seven and eight weeks of age, all rats except those in vehicle-treated groups received s.c. injections of AOM at a dose rate of 15 mg/kg body wt. The rats were maintained on their respective diets for 52 weeks and were then sacrificed. Colon tumors were processed and evaluated histopathologically. Colon tumor incidence and multiplicity were analyzed statistically. No obvious toxic effects were observed following dietary administration of 10 or 15 ppm selenomethionine as indicated by body weight gain. Administration of 10 or 15 ppm selenomethionine had no significant effect on colon tumor incidence and multiplicity. This study suggests that i) selenomethionine lacks chemopreventive efficacy against AOM-induced colon carcinogenesis and ii) other forms of selenium or a mixture of selenium compounds present in selenium-enriched brewer's yeast need to be evaluated for their chemopreventive efficacy.     

Induction of pulmonary carcinogenesis in Wistar rats by a single dose of 9, 10 Dimethylbenz(a)anthracene (DMBA) and the chemopreventive role of Diclofenac

Objectives

To evaluate the chemopreventive efficacy of Diclofenac, a preferential cyclooxygenase-2 (COX-2) inhibiting non steroidal anti-inflammatory drug (NSAID) in the 9, 10 Dimethylbenz(a)anthracene (DMBA) induced experimental lung carcinogenesis.

Methods

Animals were divided into 4 groups. Control group received normal saline intratratracheally. DMBA group was given DMBA (20 mg/kg of body weight) in the similar manner. DMBA + Diclofenac group was given daily oral dose of Diclofenac (8 mg/kg of body weight) in addition to DMBA while the last group received Diclofenac only. Animals were sacrificed after 24 weeks. COX-2 expression was studied by immunohistochemistry (IHC) and Western immunoblotting. For apoptosis study DNA fragmentation on agarose gel and florescent staining of alveolar macrophages were done.

Results

The incidence and burden of tumor were reduced by the Diclofenac treatment. Diclofenac caused the reduction in the COX-2 levels which were increased in the DMBA treated group. It also caused the induction of apoptosis as seen by both techniques.

Conclusion

From all these results it can be concluded that Diclofenac might have a chemopreventive role for lung carcinogenesis which is mediated by suppression of COX-2 enzyme and induction of apoptosis.     

Hesperidin alleviates oxidative stress and downregulates the expressions of proliferative and inflammatory markers in azoxymethane-induced experimental colon carcinogenesis in mice

Objective

Colon cancer is a common malignant neoplasm causing huge morbidity and mortality worldwide. Current therapeutic interventions are unsatisfying, which necessitates novel chemopreventive strategies. The present study was intended to elucidate the chemopreventive efficacy of hesperidin against azoxymethane (AOM)-induced mouse colon carcinogenesis.

Materials and methods

Swiss albino mice were subjected to intraperitoneal injections of AOM once a week for 3 consecutive weeks. Hesperidin treatments were provided in the initiation or post-initiation phases. The number and multiplicity of aberrant crypt foci (ACF), tumor incidence and antioxidant status were determined. Histopathological analyses, proliferating cell nuclear antigen (PCNA) index and modulations in the expression of inflammatory markers such as nuclear factor kappa B (NF-κB), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were studied.

Results

Hesperidin treatments significantly inhibited the number and multiplicities of AOM-induced ACF and tumor incidence. Hesperidin reduced oxidative stress parameters and enhanced antioxidant status. A marked decrease in the PCNA index was evident on hesperidin administration. Hesperidin treatments caused a prominent downregulation of NF-κB and its target molecules iNOS and COX-2, thereby combating inflammation.

Conclusion

This study proves the chemopreventive efficacy of hesperidin against the deleterious traits of colon carcinogenesis including accelerated proliferation, inflammation and persistent oxidative stress.     

Cell Cycle Regulation and Differentiation in the Human Podocyte Lineage

Mature podocytes are regarded as growth-arrested cells with characteristic phenotypic features that underlie their function. To determine the relationship between cell cycle regulation and differentiation, the spatiotemporal expression of cyclin A, cyclin B1, cyclin D1, the cyclin-dependent kinase inhibitors (CKIs) p27 and p57, and markers of differentiating podocytes in developing human kidneys was investigated by immunohistochemistry. In S-shaped body stage, Ki-67, a cell proliferation marker that labels the G1/S/G2/M phase, was expressed in the majority (more than 80%) of presumptive podocytes, along with cyclin A (~20% of the Ki-67-positive cells) and cyclin B1 (less than 5% of Ki-67-positive cells) expression. Among these cells, cyclin D1 and CKIs were markedly down-regulated. At the capillary-loop stage, by contrast, CKIs and cyclin D1 were intensely positive in podocytes, whereas no Ki-67, cyclin B1, or cyclin A expression was seen. Moreover, double-immunolabeling and serial-section analysis provided evidence that CKIs and markers specific for differentiating podocytes, namely PHM-5 (podocalyxin-like protein in humans), synaptopodin (a foot process-related protein), and C3b receptor, were co-expressed at the capillary-loop stage. Podocytes were the only cells within the glomeruli that expressed CKIs at immunohistochemically detectable levels. Furthermore, bcl-2 (an apoptosis inhibitory protein) showed a reciprocal expression pattern to that of CKI. These results suggest that 1) the cell cycle of podocytes is regulated by cyclin and CKIs, 2) CKIs may act to arrest the cell cycle in podocytes at the capillary-loop stage, and 3) the specific cell cycle system in podocytes may be closely correlated with their terminal differentiation in humans.     

Possible prognostic significance of p53, cyclin D1 and Ki-67 in the second primary malignancy of patients with double primary malignancies

Patients with two types of primary cancers are rare. In this study, we investigated the expression of p53, cyclin D1, and Ki-67 in the second primary malignancy. Tissue samples were obtained from the second primary cancer site of 43 patients who met the diagnostic criteria for double primary cancer. p53, cyclin D1 and Ki-67 were determined using immunohistochemistry. Categorical variables were compared using the Chi-squared test; correlation between data scores and histology was calculated using the Spearman’s rank-order correlation. The expression rates of p53, cyclin D1 and Ki-67 in the second primary malignancy site were 60.5%, 30.2% and 65.1% respectively. p53 expression showed statistically significant association with tumor occurrence interval, pathological grading and nodal metastasis (p < 0.05). Positive correlation was detected between the expression of cyclin D1 and Ki-67 and the expression of p53 (r = 0.313, p = 0.041; r = 0.319, p = 0.037, respectively). High-expressing p53 or cyclin D second primary malignancies were associated with decreased overall survival (p = 0.040 and p = 0.043, respectively). Ki-67 expression levels did not exhibit statistically significant differences in survival. In conclusion, elevated protein expression of p53, cyclin D1 and Ki-67 in the second primary malignancy is an indicator of more aggressive malignant behavior of the secondary tumor. These markers may have prognostic value in the clinical setting.     

Nuclear beta-catenin expression is rare and its potential association with short survival in colorectal signet-ring cell carcinoma.

Colorectal signet-ring cell carcinoma (SRCC) is a rare cancer and the prognosis is usually very poor. The biologic pathways involved in its oncogenesis are unknown. beta-catenin, a key target in the Wnt-signaling pathway, is recognized to play an important role in the carcinogenesis in conventional colorectal carcinoma. This study explores the involvement of Wnt-signaling molecules beta-catenin and cyclin D1, cell cycle regulators cyclin D3, proliferative index Ki-67, apoptotic index, and angiogenic indicator CD31 in 20 colorectal SRCC paraffin-embedded specimens. Results showed that there were 2 specimens with nuclear beta-catenin and higher expression of cyclin D1 than the remaining 18 specimens. Surprisingly, those 2 patients had a much shorter survival of 6 months than the remaining 15 patients, who had around 24 months. Moreover, all colorectal SRCC specimens had an overexpression of cyclin D1, cyclin D3, and Ki-67, as well as much more angiogenesis and apoptosis than adjacent normal epithelial tissues. The authors make the preliminary comment that nuclear beta-catenin is a rare phenomenon in colorectal SRCC, but the involvement of it may indicate a worse prognosis with shorter survival than colorectal SRCC without nuclear beta-catenin expression. Besides, overexpression of cyclin D1, cyclin D3, Ki-67, and increased angiogenesis and apoptosis may play a vital role in promoting colorectal SRCC development.     

Reactive nitrogen species in colon carcinogenesis

The role of reactive nitrogen species (RNS) in colon carcinogenesis is multifactorial and affects diverse processes, such as proliferation, apoptosis, differentiation, tumorigenesis, and metastases. This review describes the stages in colon carcinogenesis where nitric oxide (NO) and inducible NO synthase (NOS2) may influence the progression of a normal mucosa to overt metastatic cancer. Overexpression of NOS2 and an increase in the generation of NO and other RNS may lead to apoptosis resistance, DNA damage, mutation, up-regulation of COX-2, increased proliferation, an increase in oxidative stress and an increase in tumor vascularity and metastatic potential. Therefore, future goals are to establish mechanistically based biomarkers to assess individuals at risk for colon cancer and to implement chemopreventive and dietary strategies that reduce colon cancer risk. An understanding of NO signaling pathways in colon epithelial cells should provide the basis for novel biomarker development. Colon cancer prevention may be achieved effectively by chemically interfering with key components of the NO signaling pathways, changing dietary habits to reduce fat and increase antioxidant-containing vegetables, and dietary supplementation to increase DNA repair.     

Potential chemoprotective effects of the coffee components kahweol and cafestol palmitates via modification of hepatic N-acetyltransferase and glutathione S-transferase activities

Coffee drinking has been associated with reduced incidence of colorectal cancer, possibly via chemoprotection/modification of the metabolism of dietary heterocyclic amine carcinogens such as 2-amino-1-methyl-6-phenylimidazo-[4,5-b]pyridine (PhIP) by kahweol and cafestol palmitates (K/C), two components of unfiltered coffee. Using the PhIP-exposed male Fisher F344 rat as a model, K/C have been shown to reduce colonic PhIP-DNA adducts by > 50%. We have used the male F344 rat to investigate the effects of dietary K/C (0.02-0.2% as a 1:1 mixture) on the metabolism of PhIP by N-acetyltransferase- (NAT), sulfotransferase- (SULT), and glutathione-dependent pathways. K/C decreased hepatic NAT-dependent PhIP activation by up to 80% in a dose-dependent manner. Conversely, hepatic glutathione S-transferase (GST) activity/expression increased, e.g., 3-4 fold toward 1-chloro-2,4-dinitrobenzene (total activity), up to 23-fold toward 4-vinylpyridine (rGSTP1), and approximately 7-fold for rGSTA2 protein. These effects had fully developed after 5 days of the test diet and persisted for at least 5 days after withdrawal of K/C. Hepatic glutathione increased two- to threefold and this increase was more short-lived than other changes. K/C did not modify hepatic SULT activity or colon NAT and GST activities. Benzylisothiocyanate and black tea, which have also been shown to reduce the formation of PhIP-DNA adducts in this model, had little effect on hepatic NAT, SULT, GST, or GSH. In primary culture of rat hepatocytes, both kahweol and cafestol palmitates reduced NAT activity by 80%. In summary, the unique potential of K/C to convert rapid acetylators to a slow acetylator phenotype, accompanied by GST induction, might contribute to chemoprevention against cancers associated with heterocyclic amines.     

Studies on mammary carcinogenesis induced by a heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine, in mice and rats

Ten heterocyclic amines (HCAs) that are produced by heating amino acids, proteins, or proteinaceous food such as fish and meat were examined for carcinogenicity in rats and mice. Three of them, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), have been shown to induce mammary cancer in female F344 and/or SD rats, but none of the HCAs induced mammary cancer in CDF(1) mice. This report reviews our recent studies on mammary carcinogenesis of PhIP in various strains of mice and on the roles of genomic instability in the rat mammary carcinogenesis of PhIP. We demonstrated that the survival time from mammary adenocarcinomas was shorter in PhIP-treated BALB/c mice than that in the untreated control, and with a significantly higher incidence in the C.B-17 strain of mice compared with that of the control. To clarify mechanisms of mammary carcinogenesis, we examined genomic instability in rat mammary cancer induced by PhIP. Mammary cancers were induced in F344 x SD F(1) rats harboring the lacI transgene, and two cell lines were established from two adenocarcinomas. They showed a greater than 10-fold higher frequency of spontaneous mutations than that of the primary culture of normal mammary epithelial cells, in the lacI transgene and the hprt endogenous gene during cell replication. Nucleotide sequencing revealed that almost all types of mutations were increased, with a remarkable increase of A:T --> C:G mutation. This genomic instability was not attributed either to alterations of mismatch-repair enzymes or to p53. These mutational characteristics were also observed in the original tumors. Single-nucleotide instability (SNI) might be implicated in the mammary cancer induced by PhIP.     

Studies with the azoxymethane-rat preclinical model for assessing colon tumor development and chemoprevention

During recent years, multidisciplinary studies in epidemiology and molecular biology have contributed to our understanding of the etiology of colorectal cancer; more importantly they have enabled us to approach its prevention. An impressive body of epidemiological data suggests an inverse relationship between colorectal cancer risk and consumption of diets rich in omega (omega)-3 fatty acids (n-3 PUFAs) or the regular use of nonsteroidal antiinflammatory drugs (NSAIDs), including aspirin. The development of strategies for the chemoprevention of colorectal cancer have been facilitated by the use of relevant animal models mimicking the neoplastic processes that occur in humans, including similarities in histopathology and molecular and genetic lesions during both the early and promotion/progression stages of carcinogenesis. Studies with the azoxymethane-F344 rat model indicate that diets rich in n-3 PUFAs, NSAIDs, selective cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) inhibitors, and curcumin can reduce the incidence of colon cancer. Advances in the knowledge of the mechanisms by which chemopreventive agents act offer opportunities to use combinations of specific chemopreventive agents, having clinically beneficial aggregate activity with minimal toxicity. This approach is extremely important when a promising chemopreventive agent demonstrates apparent efficacy but may produce toxic effects at high doses. Our studies show that a combination of very low doses of piroxicam (NSAID) and difluoromethylornithine, a specific inhibitor of ornithine decarboxylase, or very low doses of COX-2 and HMG-CoA reductase inhibitors are more effective in inhibiting colon carcinogenesis than administration of these compounds as single agents even at higher levels. The natural history of colorectal cancer, from dysplastic aberrant crypts to adenomas and adenocarcinomas, offers multiple opportunities for assessment and intervention. Of further importance is to identify whether the molecular targets that are critical in the growth and survival of the malignant colorectal cell are modulated by n-3 PUFAs, NSAIDs, or COX-2 and iNOS inhibitors.     

Documentation of Ultrastructural Changes in Nucleus and Microvilli by Fish Oil in Experimental Colon Carcinogenesis

Fish oil (FO) exerts a chemopreventive effect by regulating apoptosis in colon carcinogenesis. The present study reports the ultrastructural changes in various organelles on supplementation of FO in experimental colon carcinogenesis. The carcinogen treatment led to abnormal nuclear shape and alteration in microvilli number indicating cancer establishment. On the other hand, different ratios of FO and corn oil increased chromatin condensation along with an extensive loss of microvilli in a dose- and time-dependent manner which depicts an increase in apoptosis. The associated ultrastuctural alterations support the facilitation of apoptosis by FO as a mechanism for its beneficial effect in colon carcinogenesis.     

Reduced expression of p27/Kip1 is associated with the development of pulmonary adenocarcinoma

p27/Kip1 (p27), a negative regulator of cell proliferation, is a powerful prognostic marker in non-small cell lung carcinoma. To clarify the significance of p27 aberrations in the tumourigenesis of lung adenocarcinoma, p27 expression was investigated by immunohistochemistry in lung adenocarcinoma and its precursor lesion, atypical adenomatous hyperplasia (AAH), and correlated with the expression of Ki-67, cyclin D1, and cyclin E. The p27 labelling index decreased in parallel with tumour progression (24.0% to 4.5%) and was found to be lower in neoplastic lesions than in normal bronchiolar epithelial cells (48.8%). There was a negative correlation between p27 and Ki-67 expression (rho=-0.384, p<0.001). Cyclin E-positive lesions (with labelling index >/=5%) were found only in overt adenocarcinomas. The Ki-67 labelling index of cyclin E-positive, high (>/=10%) p27 expressers was lower than that of cyclin E-positive, low (<10%) p27 expressers (16.8% vs. 42.6%; p=0. 046) and was similar to that of cyclin E-negative adenocarcinomas (15.0%). These results indicate that reduced p27 expression is associated with and may play a role in progression during the development of pulmonary adenocarcinoma.