[Immunopathomorphology of the human placental barrier in the first trimester of pregnancy complicated by the inflammation of the birth-ways].

The effect of ascending infection of birth-ways on transport of maternal immunoglobulins (Igs) through the placental barrier in humans during the first trimester of pregnancy was studied. The transport of Igs is seen already in 3.5-to 5-week-old embryos, and different cellular and biochemical compounds participate at each stage of this process. Transport of Igs through the trophoblast is carried out due to the secretory component (SC) and, perhaps, to some other receptors. Monocytes together with Igs penetrate into capillaries between the endothelial cells and are transported with the blood all over the body. It seems that SC and other receptors help Igs to penetrate into capillaries through the endothelium. Further, Igs are transported with erythroblasts. In the development without infection the transport of IgG was seen in all cases studied. Inflammation of the birth-ways is accompanied by an increase in transport of all Igs, already in early embryogenesis. Three groups were distinguished: 1) abortions without inflammation; 2) cases with signs of moderate inflammation (endometritis, deciduitis); 3) cases with intensive inflammation with necrosis and leucocytic infiltration. Transport of Igs was seen in 77.8% cases of the first group and in all cases of groups 2 and 3. Transport of IgM was not found in the first group, but was seen in 50% cases of group 2 and 66.7% of group 3.

Expression of p53 protein in rat colon cancer cell lines transfected with Rous sarcoma virus-33.

Expression of the cytoplasmic soluble form of p53 protein in the different rat colon cancer cell lines transfected and non-transfected with Rous sarcoma virus-33 was studied. Concentrations of the p53 protein were detected by commonly used immunochemical methods after its isolation by affinity chromatography columns with the gel fiberglass membranes. The main component of tumor-associated antigens (TAA) eluted from virus-transfected cells was the 53 kDa protein in its cytoplasmic soluble fraction. The non-virogenic colon cancer cells contain a few proteins and concentration of 53 kDa protein was low. Western immunoblotting revealed that the 53 kDa protein isolated from the cell lyzates studied was distinctly recognized by the p53 MAb. ELISA showed that its concentration was markedly higher in the lyzate obtained from the highly virogenic and tumorigenic R9 cell line compared with the non-virogenic cell line RT1. We concluded that the expression of the p53 protein is related to the viral transfection of cancerous cells. The possible role of this phenomenon in the etiology of cancer is discussed.

The role of p53 tumor-associated protein in colon cancer detection and prevention (Review).

The role of the cytoplasmic fraction of p53 protein in the detection and prevention of colon cancer was analyzed by comparing results of the author's studies with data published in the literature. In cancer detection, although immunochemical methods are widely used for diagnostic purposes positive results are not always obtained. Using a new modification of affinity chromatography columns, we isolated the cytoplasmic, soluble form of tumor-associated antigens (TAA) from the serum of colon cancer patients. p53 protein was found to be one of the main components of such TAA. The serum level of p53 antigen was related to tumorigenicity: the correlation and regression coefficients between the serum level of p53 protein and the progress in colon cancer were 0.48 and 0.88, respectively, p<0.01. Thus HPLC-determination of the serum concentration of this protein could serve as a screening tool for cancer detection. The sensitivity and specificity of the method reached 92% and its accuracy was 88%. The method can be used to detect cancer development either as a primary disease or as a recurrent disorder. In cancer prevention, a few attempts have been made to utilize p53 protein as a tumor suppressor. IgG generated against the cytoplasmic p53 antigen from tumor-bearing rats prevents the carcinogenic effect of 1,2-dimethylhydrazine decreasing significantly the number of tumor-bearing rats in vaccinated group compared with non-vaccinated controls. Anti-p53 IgG not only had an antitumor effect but also prevents benign tumors from becoming malignant: the number of malignant tumors in vaccinated rats was half that in controls (29% and 58%, respectively). The antitumor effect of vaccination is accompanied with a significant increase in the serum-level of p53 antigen in vaccinated rats compared with non-vaccinated controls. p53 protein plays an important role in colon cancer: the unique mechanism involved in both cancer development and in cancer prevention appears to include high production of this protein.

The role of dietary factors in prevention of chemically-induced cancers (review).

The role of dietary factors in prevention of chemically-induced cancer was reviewed on two models: i) the role of high fiber diets in prevention of colon cancer and ii) the role of high fat diets in prevention of mammary gland cancer, i) Experiments in colon cancer showed that 20% cellulose content decreased tumor incidence caused by 1,2-dimethylhydrazine (DMH) to 33% compared with 92% of tumors developed in animals fed a fiber-free diet. The tumor-preventive effect of a cellulose diet was accompanied by increased enzyme concentrations, such as ornithine decarboxylase, thymidine kinase and beta-glucuronidase. Corncob fiber (15%), treated with the fungus Pleurotus os., had a significant protective effect against DMH-induced rat colon cancer. This effect was accompanied by activation of some cellular mechanisms, i.e. apoptosis, proliferating cell nuclear antigen (PCNA) and p53 protein synthesis. A high positive correlation was found between tumor grade and p53 protein in the serum (r=0.97) or in the cell cytoplasm (r=0.77), and between tumor grade and PCNA (r=0.81). An inverse relationship was found between tumor grade and apoptosis (r=-0.63). ii) Experiments in mammary gland cancer showed that a 15% olive-oil diet reduced tumor incidence caused by 9,10-dimethyl-1,2-benzanthracene to 30%, compared with 55% in the control group. The antitumor effect of the olive oil diet was connected to its content of monounsaturated fatty acids, such as oleic and palmitic acids. The promotive tumorigenic effects of other high-fat diets (avocado, soybeans) were associated with high content of some polyunsaturated fatty acids (linoleic and alpha-linolenic). We concluded that different diets have different targets. The effect of the same diet depends on its content of anti-tumor substances.

Soluble low-molecular-mass tumor-associated antigens promote the suppression of rat mammary tumors by tamoxifen and prevent its toxic effect.

This study examined whether the soluble tumor-associated antigens (sTAA) of 66 kDa and 51 kDa could promote suppression of chemically-induced rat mammary tumorigenesis by the hormone-related anticancer drug tamoxifen and prevent the drug's toxic side-effects. Dimethylbenzanthracene (DMBA, 10 mg/rat, 3 administrations) was used to induce mammary tumors in 8-week-old Wistar rats. Then, for 13-17 more weeks, preparations of sTAA (50 microg/rat) and tamoxifen (10 mg/rat) were administered, separately or in combination, on a weekly basis. The experiment was continued for 18 weeks and was terminated when the number of dead rats reached 50% in each group. Treatment with tamoxifen inhibited tumor growth and their malignance: the number of rats without malignant tumors significantly increased compared to controls, 27.3% and 5.6%, respectively. Treatment with sTAA resulted in a significant increase in the number of regressed tumors to 10.1% compared to 0% and 1.4% in control and tamoxifen-treated rats, respectively. Moreover, the period of 50% survival increased from 13 weeks in tamoxifen-treated rats to 17 weeks, and as a result, rats treated with sTAA were involved in the experiment for an average 14.3 weeks compared to 10 and 10.4 weeks in control and tamoxifen-treated groups. In rats treated simultaneously with tamoxifen and sTAA, the time of appearance of each new tumor increased from 4.5 weeks to 6.6 weeks with a significant increase to 14.3% in the number of regressed tumors. The period to 50% survival increased to 18 weeks, and these rats were involved in the experiment for up to 16.4 weeks. The number of rats without malignant tumors increased to 22.2% and the time of appearance of malignancy increased to 9.6 weeks, as compared to 7.3 weeks in controls. The results demonstrated that sTAA have tumor-suppressive properties, and also enhance the anticancer effects of tamoxifen and prevent its toxic side-effects.

Soluble low-molecular-mass tumor-associated proteins promote the suppression of mammary tumors by cyclophosphamide.

This study examined whether the soluble tumor-associated-antigens (TAA), of 66 kDa and 51 kDa, could promote suppression by anticancer drugs of chemically-induced mammary tumorigenesis. Dimethylbenzanthracene (DMBA, 10 mg/rat, twice) was used to induce mammary tumors. Then, for nine more weeks, the preparation of TAA and cyclophosphamide (CPA), alone or in combination with TAA, were administered in weekly doses. Twenty weeks after DMBA exposure, the mammary tumor yield was 2.4, 2.8 and 2.9 in the experimental groups compared to 3.5 in the controls. Seventy-five percent of the rats in the control group, but only 37% of TAA, 50% of the CPA, and 30% of the CPA and TAA treated animals had malignant tumors. In the experimental groups, 6.5%, 25% and 38%, respectively, of the tumors regressed, compared to 3% in controls. In the groups receiving CPA or TAA, regression was observed in the fifth week of treatment, and in the group receiving combined treatment, already in the first week. The size of the tumors in control rats increased during the last 10 weeks 3.6 times, in the CPA treated rats 1.15 times, but in those receiving CPA plus TAA it decreased by 0.7 times. The results of our experiment demonstrated that TAA have distinct tumor-suppressive properties, and can enhance the anticancer effects of CPA.

The immune system, apoptosis and apoptosis-related proteins in human ovarian tumors (a review).

Our studies on the relationships among the lymphoid system, apoptosis and apoptosis-related proteins (ARP) in human ovarian benign cysts, borderline tumors, and carcinomas are reviewed and analyzed. Fas and Fas ligand are expressed in 50% to 80% of the epithelial cells in all studied tumors. Many bcl-2-positive tumor epithelial cells are seen in benign cysts and they disappear as tumorigenesis progresses, whereas p53 protein is found only in borderline tumors and in carcinomas. Many exceptions to the opinion that bcl-2 inhibits apoptosis and p53 promotes it are encountered. Bcl-2 is lacking in epithelial cells of mucoid tumors of all grades, and its absence does not stimulate their apoptosis. P53 protein is absent from most lymphocytes, macrophages and epithelial tumor cells, nevertheless, they undergo apoptosis. Indeed, in many tumors apoptosis is regulated without the participation of bcl-2 and p53. Different components of the immune system become active during different stages of tumor development. The weak reaction of T-cell killers and macrophages is typical in benign cysts. In borderline tumors, the activity of T-cell killers increases in the parenchyma, and that of T helpers and macrophages in the stroma. In carcinomas with high lymphoid infiltration, a strong reaction of macrophages and T cell killers in the tumoral parenchyma as well high reaction of T helpers and B lymphocytes in the stroma are typical. Apoptosis that should protect against tumor also stimulates apoptotic death of lymphocytes and macrophages, and this has catastrophic consequences, as seen in weakly infiltrated carcinomas. In conclusion, our studies indicate that during malignancy the major task of the immune system is curtailment and control of tumorigenesis.

Gel fiberglass as a new matrix of affinity chromatography columns for isolation of colon cancer-associated antigens.

Gel fiberglass (GFG), a new affinity biosensor, was used to isolate human p53 antigen with rabbit anti-rat p53 IgG. The biosensor was prepared as a membrane from glass fibers covered with oxysilanes. A thin layer of protein, trapped in gel glass during its preparation, is deposited on the surface of a lattice of glass fibers. In such conditions, a maximum number of protein molecules may contact external agents percolated through a membrane. The membranes demonstrate high stability and can be stored in dry conditions or several months at room temperature. Columns for affinity chromatography were prepared from the GFG membranes and were used to isolate various proteins, including the tumor-associated antigens (TAA). The capacity of such columns was calculated as the amount mg of protein isolated from 1 ml of TAA-containing serum. In colon cancer patients, up to 5-6 mg TAA were extracted from 1 mi of sera. Two main components of cytoplasmic TAA isolated in our experiments were p64 and p53 proteins. Their concentration was determined by HPLC. The p53 protein has been isolated from the serum of cancer patients in the highest concentration yet reported, up to 3-4 mg/ml. In our previous studies, isolation of p53 protein was based on its affinity reaction with anti-p53 IgG generated against antigens of the same species. Herein, we report for the first time the capability to isolate human p53 antigen using GFG columns with entrapped anti-rat p53 IgG. Blood levels of p53 antigen isolated were very similar in both experiments. This has both theoretical and practical significance, demonstrating that the GFG membranes have great potential for isolating macromolecules utilizing various ligands. The finding facilitates an easy and highly effective method to isolate antigens from different organs, both animal and human, which can be used for important goals including diagnosis, therapy and generation of specific antibodies.

HPLC determination of serum levels of soluble p53 antigen as a new method for colon cancer detection, and its clinical implication.

Previously, we have described a new modification of affinity chromatography columns for isolation of the cytoplasmic, soluble form of tumor-associated antigens (TAA) from the serum of colon cancer patients (Oncol Rep 2: 679-683, 1995). In this communication, we have shown that the main proteins of these TAA were p64 and p53. The correlation coefficient between each of these proteins and the total amount of TAA or total serum protein ranged from 0.55 to 0.93. The serum level of p53 antigen was shown to be related to the tumorigenicity: the correlation and regression coefficients between the serum level of p53 protein and the progress in colon cancer were 0.48 and 0.88, respectively, p<0.001. Therefore, the determination of serum concentration of this protein can serve as a screening tool for cancer detection. The serum level of p53 protein ranges between 0.24 to 0.94 mg/ml in patients with non cancer diseases, and between 1.0 to 2.0 mg/ml in patients with polyposis and in a high risk group, respectively, increases over 2.0 mg/ml in primary colon cancer patients and up to 5.0 mg/ml in cancer patients with metastases. The sensitivity and specificity of our method achieved 92% and 96%, respectively, and accuracy 88%. The presence of p53 protein in the cytoplasm of cells from patients with non cancer diseases may explain why p53 antigen is presented in their sera. Our method can be useful to detect cancer development either as a primary illness or as a recurrent disorder. It is possible to follow up patients with chronic diseases and to detect transformation of these diseases into cancer, or to follow up former cancer patients in order to detect as early as possible incidence of recurrent cancer. It should also be emphasized that our method allows the detection of patients with polyposis or those of high risk groups who exhibit a tendency to develop colon cancer.

Variability of serum tumor-associated antigens and their significance in cancer-diagnosis (review).

The significance of serum tumor markers in the diagnosis of several of the most widespread types of cancer (lung, gastric, colorectal, breast, endometrial and ovarian) has been reviewed. The high heterogeneity of obtained results demonstrated in this review suggests that none of the present markers can be utilized alone as a significant marker of neoplasia and that only their combined use can give objective information. The role of individual variability of various markers in the detection of different stages of neoplastic transformations has also been revised. It was concluded that none of the markers studied can be absolutely positive without false results, and that objective information regarding early stages of malignancy may be obtained only with a combined analysis utilizing many laboratory methods. A new approach should be developed in oncological diagnostic practice.

The clinical relevance of p53 oncoprotein determination in cancer-diagnosis and prognosis.

The role of p53 overexpression in tumorous tissues and in blood of cancerous patients is reviewed. It has been shown that neither immunohistochemical nor immunochemical methods can be presently regarded as being highly significant in diagnosis or prognosis of cancer. The data of the literature and our preliminary studies showed that overexpression of p53 not only has a role in oncogenesis but can also be regarded as a protein which accumulates in cells under stress conditions. The overexpression of p53 protein can be found not only in cancerous patients but also in patients with non-cancerous diseases. Further investigations should be undertaken to better understand the diagnostic and prognostic role of p53 in clinical practice.

Isolation of tumor-associated protein from sera of colon tumor-bearing rats using polyclonal antibodies entrapped in gel fiberglass columns.

A new, effective support for the isolation of proteins has been invented utilizing a gel fiberglass (GFG) (R. Zusman, Patent application 1992, 1993). We describe the utilization of this support to isolate tumor-associated proteins (TAP) from sera of colon tumor-bearing rats. Sera were percolated through GFG columns with entrapped anti-rat colon cancer IgG from rabbits. TAP were eluted in a large amount, up to 3 mg/ml sera/column. Their affinity to antitumorous polyclonal antibodies was detected by ELISA. Western immunoblotting with commercial monoclonal antibodies has identified the isolated antigens as p21 and p53. Their concentrations were much higher (up to 1,000 times) in sera from tumor-bearing rats compared to control rats. The method was highly reproducible. It is suggested that data obtained can be considered as a basis for the further improving of quantitative methods of diagnosis.

Transplacental tumor-preventive effects of polyclonal antibodies generated against the soluble 53 kDa antigen on mammary tumorigenesis in offspring.

This study was designed to determine whether immunizing females with polyclonal antibodies generated against the soluble 53 kDa tumor-associated antigen (s53 TAA) has a tumor-preventive effect on their progeny and whether this effect is manifested in some biochemical characteristics. Rat females were immunized before mating with anti-s53 IgG (50 microg/rat in Freund's complete and incomplete adjuvant, three times during a month) and their 5-week-old offspring was exposed to the carcinogen (dimethylbenz(a)antracene, 10 mg/rat). Results of these experiments were studied 4 months later. Vaccination of mothers decreased the tumorigenic effects of DMBA on their offspring. Blood levels of soluble TAA were analyzed in offspring of different groups. Two TAA were isolated from the blood, with molecular masses of 64 and 53 kDa. Their concentrations differed in offspring obtained from different maternal groups. Vaccination itself resulted in a marked increase in the blood levels of TAA, not only in the mothers but also in their offspring, however, this increase was not significant in tumor-bearing animals. In offspring from non-vaccinated mothers, tumorigenesis resulted in high overexpression of s53. In offspring from vaccinated mothers, a high blood level of s53 was shown even in tumor-free animals, probably due to maternal vaccination. We conclude that maternal vaccination before pregnancy increases immunoreactivity in offspring and can reduce risk of tumors in those progeny.

Mammary tumors in splenectomized rats.

The objective of this study was to examine how splenectomy affects the immune response, particularly T cells, in chemically-induced mammary tumors. Female rats were splenectomized and then exposed to 9,10-dimethyl-1,2-benz(a)anthracene (DMBA) to induce mammary tumors. Splenectomy significantly decreased the rate of tumor appearance and their malignant transformation. The tumor latency period in splenectomized rats was 12.0+/-0.9 weeks compared to 9.7+/-0.5 wk in intact controls, and malignancy appeared in 45% of splenectomized rats, compared to 70% in controls. By the end of the experiment, the total number of tumors and their size were similar in both groups. Blood CD4+ and CD8+ T cell concentrations were similar in tumor-bearing and tumor-free splenectomized animals, but in both groups CD4- and CD8- lymphocytes decreased sharply compared to control animals. In tumor-bearing rats, splenectomy also resulted in significantly more circulating natural killer cells. The spleens of tumor-bearing control rats had significantly fewer CD4+ and CD8+ lymphocytes and more CD4- and CD8- lymphocytes and natural killer cells than did their blood. In conclusion, splenectomy inhibits the early stages of tumorigenesis and reduces the rate of malignant transformation of benign tumors, but does not prevent the progress of carcinogenesis. Differences between splenectomized (operated) and intact rats to the effect of DMBA can be explained by an increase in non-specific resistance of splenectomized rats as a result of operation.

Effects of a 15% orange-pulp diet on tumorigenesis and immune response in rats with colon tumors.

This study evaluated whether the feeding of rats with a 15% orange-pulp diet affects the lymphatic system and the tumorigenic response in rats exposed to a high dose of carcinogen. Five-week-old Sprague Dawley rats were divided into 2 groups fed a control chow diet or the same diet with 15% orange pulp. All rats were injected with 1,2-dimethylhydrazine (DMH) (20 mg/kg) weekly for 6 weeks. At 8 months, tumors, spleens and descending colon were taken from each group for analyses. Feeding rats the 15% orange-pulp diet did not reduce the tumor number but modified the number of adenocarcinomas found in the orange-pulp group compared to controls: 66.7% vs. 93.7%. The number of endophytic tumors was also significantly lower in the experimental group: 6.3% vs. 32.3% in controls. DMH affected the size of the splenic structures. The size of follicles and germinal centers decreased significantly in tumor-bearing rats compared to tumor-free rats. This effect was changed in rats fed the orange-pulp diet. In tumor-bearing rats from this group, only the area of the marginal zone decreased and the red pulp increased compared to tumor-free rats. The size of germinal centers significantly increased compared to tumor-bearing rats in controls. The total number of lymphoid cells decreased in germinal centers of spleens obtained from control tumor-bearing rats compared to tumor-free rats. DMH alone significantly increased the total number of cells in the colon mucosa of the rats fed the control diet. In tumor-bearing rats exposed to the carcinogen and fed the 15% orange-pulp diet, the total number of cells and the number of Ki-67+ cells increased in the depth of tumors whereas the number of CD8+ T cells increased in the colon mucosa, at the border of tumors and its depth. The caspase-3 protein a cysteine protease was elevated in tumors from rats fed the orange-pulp diet. Although the 15% orange-pulp diet did not change the number of tumors in the tumor-bearing rats, feeding rats orange pulp significantly decreased the number of endophytic tumors and increased the number of exophytic tumors. Increased activity of T cell killers in tumors and higher level of proteins involved with apoptosis following consumption of the orange pulp indicate a clear tumor suppressor effect of these dietary fibers.

Effect of giant hepatomas on lymphocyte production and secretion of apoptosis-related proteins in the rat spleen.

The effect of extremely large hepatomas on splenic lymphoid elements and apoptosis-related proteins in rats were studied. Hepatoma cells were inoculated subcutaneously into 6-week-old rats, and 4 months later the quantities of T and B cells, macrophages, and cells positive for Fas, Fas ligand (FasL) and interleukin-2 (IL-2) were immunohistochemically evaluated in spleens. Grafting of hepatoma cells caused hyperplasia of the spleen and development of giant tumors that could reach one-third of the rat's body weight. A 7-fold increase in the weight of the spleen was mainly due to proliferation of B lymphocytes and macrophages in the red pulp, while the relative quantity of CD4+ and CD8+ T cells decreased. Extremely small amount of Fas+ and FasL+ lymphocytes were present in the marginal zone, the follicles, red pulp, and occasionally in the PALS. All the splenic zones were abundant with IL-2+ cells, while macrophages and siderophages were present mainly in the red pulp and in the marginal zone of the white pulp. We suggest that all these changes are compensatory processes of the host's lymphatic system.

High tumor-preventive effects of polyclonal IgG generated against p53 tumor-associated protein obtained from benign-tumor bearing rats.

We have shown the different effects of rabbit IgG generated against various types of p53 tumor-associated protein on chemically induced colon cancer in rats. p53 protein was isolated in the form of cytoplasmic, soluble, protein from sera obtained from: a) rats with colon cancer and b) rats with benign colon tumors. The isolation was performed using the affinity chromatography columns with,eel fiberglass membranes. Anti-p53 IgG were obtained from rabbits vaccinated with the above mentioned types of p53. Sprague Dawley rats were vaccinated with anti-p53 IgG (100 mu g/rat) at two-week intervals for 2 months and then monthly for 3 months. The induction of colon cancer was caused by weekly injections with 1,2-dimethylhydrazine (20 mg/kg) for 7 weeks and was initiated 8 weeks after the start of the vaccination. Results of experiments were evaluated 6 months after the start of cancer induction. It was found that vaccination of rats with IgG generated against the p53 protein isolated from cancer-bearing rats did not exhibit significant protective effect. Only IgG generated against p53 protein from benign tumor-bearing rats significantly prevented the carcinogenic effect of DMH. The number of tumor-bearing rats in vaccinated group decreased to 44% as compared with 93% in the control group. In vaccinated rats, the number of tumors/rat was 0.8 as compared to 9.3 in controls. The number of malignant tumors in vaccinated rats was half that in controls: 29% and 58%, respectively. In the controls, metastases were found in 6 of 45 rats (13%). Anti-p53 IgG not only has an anti-tumor effect but also prevented benign tumors from becoming malignant. We suggest that the anticancer role of a vaccine generated against p53 protein from benign tumor-bearing rats is related to a wild-type p53 protein. Further studies will be performed to clarify this hypothesis.

Transplacental effect of a 15% olive-oil diet on functional activity of immune components in the spleen and colon tumors of rat offspring.

We studied whether feeding pregnant female rats a 15% olive-oil diet affects the activity of lymph cells in the spleen and tumors in offspring with chemically-induced colon tumors. Rat mothers were fed either a 7% corn-oil or a 15% olive-oil diet. Five-week-old male offspring were divided into 3 groups. A control group was fed the 7% corn-oil diet similar to their mothers. The experimental group I was fed the 7% corn-oil diet whereas their mothers were fed the 15% olive-oil diet. The experimental group II was fed the same 15% olive-oil diet as their mothers. Experimental rats were injected weekly for 8 weeks with the carcinogen, 1,2-dimethylhydrazine (DMH), 20 mg/kg b.w. Results of experiments were studied 6 months later. The area of zones in the spleen responsible for producing B and T lymphocytes were measured and the number of cells counted. The activity of lymphoid elements of the spleen and of tumors were studied using immunohistochemical methods for evaluating the synthesis of CD8(+) lymphocytes and proliferative activity of lymphocytes in spleens and tumors. Feeding pregnant or lactating mothers with the 15% olive-oil diet had no marked tumor-protective effect on chemically-induced colon cancer in offspring. Diet-dependent changes were found at the cellular level. In the spleen of control offspring, the presence of a tumor was accompanied by an increase in the number of Ki-67(+) cells and CD8(+) lymphocytes in the red pulp. In experimental group I, DMH significantly increased the total cell number and the number of CD8(+) lymphocytes in the red pulp of the spleen in both tumor-bearing and tumor-free rats. In experimental group II, the total number of lymph cells and the number of CD8(+) lymphocytes increased compared to offspring fed a control diet. Tumor formation activated the proliferative activity of lymph elements. The total number of cells in infiltrates of the colon mucosa decreased in tumor-bearing rats compared to tumor-free counterparts, and this was seen in all three dietary groups of rats. In tumors from offspring of experimental group II, only the number of CD8(+) lymphocytes increased compared to those in offspring of experimental group I. The findings indicate that feeding mothers the 15% olive-oil diet had a cancer-inhibiting role in offspring, predominantly changes at the cellular level.

Comparative effects of dimethylbenz(a)anthacene and a 15% olive-oil diet on cellular components and expression of apoptosis-related proteins in the spleen and mammary gland tumors of rats.

We compared effects of a high fat diet and a carcinogen on cellular elements of the spleen and mammary gland tumors in rats. Animals were fed a 15% olive-oil diet and a group of them were exposed to a carcinogen, dimethylbenz(a)anthacene (DMBA), in two doses of 10 mg/rat. Results of the experiments were evaluated after 4 months. We studied changes in the areas of different zones of the spleen related to production of B and T lymphocytes and also the number of cells in the spleen and tumors with positive reaction to receptors related to manifestation of apoptosis (FasL and p53) and receptors related to inhibition of apoptosis (bcl-2). In the spleen, dietary fats as well as DMBA alone decreased the zones related to production of B lymphocytes and increased the number of T lymphocytes. The combined effect of a carcinogen and a high fat diet manifested in an increase in the number of lymphoid cells and macrophages. In tumors from rats fed a low-fat diet, an extremely high number of lymphoid cells was seen in the border of tumors with T cell killers as a main component of these infiltrates. In tumors from rats fed a 15% olive-oil diet, the main component of the infiltrates were macrophages. High levels of p53+ and bcl-2+ cells were found in the spleen of rats exposed to a carcinogen. The combined effect of a carcinogen and the 15% olive-oil diet inhibited production of FasL and p53 receptors and stimulated synthesis of bcl-2 protein. In tumors, a carcinogen alone stimulated the high expression of FasL and p53 proteins, but in combination with the 15% olive-oil diet synthesis of these receptors decreased while production of bcl-2 protein increased sharply. This observation may serve as an additional proof of tumor-promoter effects of a high fat diet.

Tissue-specific expression of the p53 tumor-suppressor gene in the intestine of transgenic mice exposed to DMH and p53 antibodies.

We studied the tissue-specific expression of the p53 gene in different parts of the intestine of mice treated with low doses of a carcinogen and exposed to different p53 antibodies. The human p53 promoter-CAT transgenic mice were immunized with different p53 antibodies (monoclonal - PAb 421 and DO1, and polyclonal - H-p53 and anti-soluble p53 IgG) and then exposed to low doses of dimethylhydrazine (DMH). Enzymatic CAT activity was determined in the ileum and colon 8 weeks later after the final injection of DMH. Expression of the p53 transgene in the normal ileum was twice as high as in the colon. Treatment with DMH significantly decreased the expression of the p53 transgene both in the ileum (from 18% to 100%) and in the colon (from 10% to 52%). Vaccination of mice protected at least in part such a decrease. The most effective results were found after exposure of mice to polyclonal H-p53 and to a lesser extent to anti-p53 IgG. No difference was found in the effects of antibodies on the small and large intestines. We concluded that polyclonal antibodies were more effective than monoclonal ones in protection against anti-p53 action of DMH. The observation of these effects may make it possible to explain the higher antitumor activity of polyclonal antibodies.