Differential PGE2 and cAMP production by allogeneic and syngeneic pregnant mice uteri.

Prostaglandin E2 (PGE2) and cAMP production in allogeneic and syngeneic pregnant mice uteri, were measured in relation to the ratio of plasma estrogen/progesterone levels. PGE2 generation by allopregnant uteri varied with the days of pregnancy. The increment of the prostanoid coincided with the increase in plasma estrogen concentration, whereas the decrement of its production was in parallel with the increment of plasma progesterone. The syngeneic pregnant uterus was unable to increase the release of PGE2 above basal values during the whole pregnancy. The rise of PGE2 production by the allogeneic pregnant uterus was correlated with an increase in cAMP levels. It is proposed that the pregnant mouse uterus increases its ability to release PGE2 in response to an ovarian steroids.

Apoptosis and expression of Bcl-2 and Bax in eutopic endometrium from women with endometriosis.

OBJECTIVE: To evaluate and compare spontaneous apoptosis and Bcl-2 and Bax expression in eutopic endometrium from women with and without endometriosis. DESIGN: Apoptosis and Bcl-2 and Bax expression were examined in eutopic endometrium from women with and without endometriosis. SETTING: Instituto de Biología y Medicina Experimental-CONICET, Department of Gynecology and Department of Gynecological Pathology, Clínicas University Hospital, Buenos Aires, Argentina. PATIENT(S): Women with untreated endometriosis (n = 14) and controls (n = 16). INTERVENTION(S): Collection of endometrial samples during diagnostic or therapeutic laparoscopy. MAIN OUTCOME MEASURE(S): Apoptotic cells were detected with use of the dUTP nick-end labeling (TUNEL) assay; Bcl-2 and Bax expressions were assessed with use of immunohistochemical techniques. RESULT(S): Spontaneous apoptosis was significantly lower in eutopic endometrium from patients with endometriosis, compared with healthy controls (2.26 +/- 0.53 and 9.37 +/- 1.69 apoptotic cells/field, respectively) and was independent of cycle phase. An increased expression of Bcl-2 protein was found in proliferative eutopic endometrium from patients with endometriosis. Bax expression was absent in proliferative endometrium, whereas there was an increase in its expression in secretory endometrium from both patients and controls. CONCLUSION(S): Women with endometriosis show decreased number of apoptotic cells in eutopic endometrium. The abnormal survival of endometrial cells may result in their continuing growth into ectopic locations.

Effects of sexual steroid hormones on the functionality of murine peritoneal macrophages.

In the present work, we studied the possible effect of steroid hormones, estradiol, progesterone, and 5 alpha-dihydrotestosterone, on different phenotypic and functional characteristics of peritoneal adherent mononuclear cells. We used female and male mice of Balb/c strain, normal, gonadectomized, and gonadectomized with hormonal replacement. We found that gonadectomy in both sexes produced a significant decrease in the functionality of membrane receptors for the complement and in phagocytic activity of Candida albicans-anti-C albicans system. In addition, the percentages of cells that reduced nitroblue tetrazolium were diminished in castrated animals. Ovariectomized females injected with estradiol presented normal levels of phagocytic and metabolic capacities, but the expression of membrane receptors for complement remained decreased. In contrast, progesterone treatment of ovariectomized animals had the opposite effect. Simultaneous treatment with estradiol plus progesterone gave results similar to those observed with estradiol only. Dihydrotestosterone per se did not affect any of the parameters measured in the conditions used here. These results suggest that female steroids affect macrophage functionality, probably by regulating surface receptors that are involved in phagocytic activity.

In vitro secretion of cytokines and prostaglandin-E2 by monocytes from lung cancer patients.

Monocytes (MO) from cancer patients present functional abnormalities, such as an altered secretion of soluble factors. In the present study, our aim was to evaluate the levels of interleukin-1beta (IL-1beta), tumour necrosis factor-alpha (TNF-alpha) and prostaglandin-E2 (PGE2) secreted in vitro by MO from lung cancer patients (LCP), spontaneously or after stimulation with lipopolysaccharide (LPS). Results showed that cytokine secretion was higher for MO from LCP than for MO from healthy controls, while in the 25% of the patients analysed, an absence of response to LPS treatment was found.

Interleukin-1 beta levels in human embryo culture supernatants and their predictive value for pregnancy.

Interleukin 1 (IL-1) is possibly one factor produced by the embryo that might have a role in the maternal immunological recognition of pregnancy. The purpose of this study was to identify an embryo-related factor suitable for prediction of pregnancy during IVF procedures. For this purpose, IL-1 beta levels were measured in 21 samples of human embryo culture-conditioned media. The average number of embryos per sample was 5 +/- 1. Simultaneously, 16 cell culture media containing 10% autologous serum but no embryos were tested as controls. IL-1 beta levels were measured using the ELISA technique, and the biological activity of IL-1 was measured by means of a C3H/HeJ mice thymocyte proliferation assay. The average IL-1 beta level +/- S.E.M. was 49 +/- 7 pg/ml in embryo culture-conditioned media and 12 +/- 2 pg/ml in controls (P < 0.001). The average IL-1 beta level in embryo culture-conditioned media from viable pregnancy cycles was 82 +/- 6 pg/ml (n = 8), while in those cases that did not result in viable pregnancies the IL-1 beta level was significantly lower (28 +/- 4 pg/ml, n = 13, P < 0.001). The IL-1 activity of embryo culture-conditioned media, measured by [3H]thymidine incorporation in thymocytes was increased, compared with control media (442 +/- 51 counts/min vs. 337 +/- 13 counts/min, P < 0.05), and the highest values corresponded to media containing those embryos that resulted in pregnancies (589 +/- 41 counts/min, P < 0.01 vs. controls). We conclude that the determination of the levels of this cytokine in embryo culture-conditioned media might be a predictive parameter for pregnancies in patients undergoing IVF-ET.

Regulation of HLA-DR antigen in monocytes from colorectal cancer patients by in vitro treatment with human recombinant interferon-gamma.

During the immune response, a great number of cytokines that modulate the function of mononuclear phagocytes are produced. Since interferons are one of the most important cytokines, the aim of this study was to evaluate the expression of HLA-DR antigen after an 18-h culture with human recombinant interferon-gamma (hrIFN-gamma) on freshly isolated peripheral blood monocytes from 16 colorectal cancer patients and 16 healthy donors, using an indirect immunofluorescence method. The results obtained showed that there was a decreased percentage of HLA-DR+ monocytes in the colorectal cancer patents (51 +/- 3%, p <0.01) compared with the healthy donors (77 +/- 2%). Treatment for 18 h with hrIFN-gamma increased the percentages of monocytes expressing HLA-DR: 71 +/- 3% for the cancer patients and 84 +/- 2% for the healthy donors (p <0.01 and <0.001, respectively). Our results demonstrate that after in vitro treatment with hrIFN-gamma, functionally altered monocytes from colorectal cancer patients can reach major histocompatibility complex II antigen expression values similar to those of healthy donors, thus improving the host's cellular immune response against the tumor.

Interferon-alpha 2b modulation of doxorubicin sensitivity in a multidrug resistant cell line.

We evaluated the effect of interferon-alpha2b as a chemosensitiser on HCT-15 cell line in treatment with doxorubicin. Chemosensitivity was determined by [3H]-thymidine incorporation and tetrazolium assays. The levels of expression of P-glycoprotein, Bcl-2 oncoprotein and HLA-ABC complex, and cell cycle/apoptosis analysis were determined by flow cytometry. Dox 50 ng/ml - IFN alpha 2b 500 IU/ml treatment inhibited cell proliferation (47.2 +/- 1.4%, p < 0.0001; MTT assay: 40.6 +/- 1.2%, p < 0.0001) and augmented the expression of P-170, Bcl-2 and HLA-ABC, while it didn't exert apoptosis, producing a slight G2/M arrest. A concentration of IFN-alpha2b, that by itself is not cytotoxic, can potentiate the efficacy of the anticancer drug. This effect is not due to a down-modulation of P-170. The absence of apoptosis and augmented levels of Bcl-2 expression suggests that this could be one of the mechanisms of drug resistance exerted by these cells.

Tumor necrosis factor-alfa production by monocytes from lung and colorectal cancer patients.

Tumour necrosis factor-alpha (TNF-alpha) is a monocyte (MO)-derived cytokine that plays an essential role in the immunological system. In the present study our aim was to evaluate the levels of TNF-alpha secreted by MO from cancer patients. Blood MO were obtained from 10 lung cancer patients (LCP), 10 colorectal cancer patients (CCP) and 10 healthy donors (HD). TNF-alpha levels in MO culture supernatants spontaneously (sp) secreted or after stimulation with LPS treatment were evaluated using a commercial ELISA kit (sensibility: 10-1000 pg/ml). Mean values, expressed as pg/ml were: LCP: sp= 452.6+/-107.2, LPS= 589.5+/-126.7); CCP: sp= 84.1+/-25.0, LPS= 437.3+/-93.2; HD: sp= 74.2+/-21.5, LPS= 573.5+/-87.1. We concluded that MO from LCP secrete high levels of TNF-alpha spontaneously (p< 0.003 versus HD) and it was also observed an absence of response to LPS treatment in the 33% of the cases in these patients.

Cisplatin containing chemotherapy influences HLA-DR expression on monocytes from cancer patients.

The effect of cisplatin-containing chemotherapy regimen was evaluated on the expression of HLA-DR antigen in peripheral blood monocytes from patients with lung (LCP) and colorectal (CCP) cancer. Chemotherapeutic schedules employed in patients were etoposide and cisplatin for LCP and 5-fluorouracil and cisplatin for CCP. The results obtained showed a diminished percentage of monocytes expressing HLA-DR antigen in LCP (52.4 +/- 2.6, p < 0.004) and CCP (50.1 +/- 2.1, p < 0.001) respectively versus healthy donors (71.0 +/- 1.1%), and that their values increased during chemotherapy, raising them up to control level after the second cycle of treatment, independently of the course of the cancer growth. We conclude that both modalities of treatment allowed an increase of monocytes expressing HLA-DR antigen, suggesting that this effect may be due to cisplatin action.

Bone marrow fibroblasts in patients with advanced lung cancer.

In a previous study we demonstrated that the incidence of fibroblast colony-forming units (CFU-F) was very low in bone marrow primary cultures from the majority of untreated advanced non-small lung cancer patients (LCP) compared to normal controls (NC). For this reason, we studied the ability of bone marrow stromal cells to achieve confluence in primary cultures and their proliferative capacity following four continuous subcultures in consecutive untreated LCP and NC. We also evaluated the production of interleukin-1beta (IL-1beta) and prostaglandin E2 (PGE2) by pure fibroblasts. Bone marrow was obtained from 20 LCP and 20 NC. A CFU-F assay was used to investigate the proliferative and confluence capacity. Levels of IL-1beta and PGE2 in conditioned medium (CM) of pure fibroblast cultures were measured with an ELISA kit and RIA kit, respectively. Only fibroblasts from 6/13 (46%) LCP confluent primary cultures had the capacity to proliferate following four subcultures (NC = 100%). Levels of spontaneously released IL-1beta were below 10 pg/ml in the CM of LCP, while NC had a mean value of 1,217 +/- 74 pg/ml. In contrast, levels of PGE2 in these CM of LCP were higher (77.5 +/- 23.6 pg/ml) compared to NC (18.5 +/- 0.9 pg/ml). In conclusion, bone marrow fibroblasts from LCP presented a defective proliferative and confluence capacity, and this deficiency may be associated with the alteration of IL-1beta and PGE2 production.

Antitumor and cytotoxic screen of 5,6,7-trisubstituted flavones from Gomphrena martiana.

An in vivo antitumor screening of extracts of Gomphrena martiana indicated positive activity in the petroleum ether extract, and its further bioactivity-directed fractionation resulted in a lipophilic flavonoid fraction. Upon inoculation of various doses of 5,6,7-trisubstituted flavones on two murine tumor lines, Sarcoma 180 and Ehrlich's carcinoma, a decrease of tumor growth was observed. An in vitro KB cultured cell screen indicated cytotoxicity.

Murine peritoneal macrophages in syngeneic and allogeneic pregnancies.

We have previously observed that some functional characteristics of peritoneal macrophages (MOp) are altered during syngeneic murine pregnancy. To determine if these alterations are related to the immunological stimulation that the embryo produces on the mother, we evaluated MOp activity in allogeneic pregnancy. We also compared expression of the la antigen, ability to phagocyte and reduce nitro blue tetrazolium (NBT) and to produce interleukin-1 (IL-1) in allogeneic and syngeneic pregnancies. We observed that at Day 7 of pregnancy the increment in MOpIa(+) percentages was more evident in allogeneic (P < 0.05) than syngeneic pregnancies, and that these values remained high during the second week of gestation. We also observed a significant decrease in the macrophages that reduced NAT during the first week both in allogeneic and syngeneic pregnancies. Yet, in the former, the percentages of MOpNBT(+) were still low in the last week of pregnancy (P < 0.05). No differences were found in IL-1 production or in estradiol and progesterone levels between the 2 types of pregnancies. Thus, it is possible to postulate that during the first week of pregnancy the strong antigenic challenge that the embryo represents may activate MOp and that this activation could be augmented when major antigenic differences between mother and embryo are present.

[Effect of BCG on the growth rate of Sarcoma in normal and splenectomized mice]. / Efecto del BCG sobre el crecimiento del Sarcoma 180 en ratones normales y esplenectomizados.

A study was carried out on the growth of Sarcoma 180 alone or mixed with BCG in normal and splenectomized BALB/c mice, with or without previous immunization with BCG intradermally (id), 40 days before tumor challenge. No survival was observed in non splenectomized mice inoculated with Sarcoma 180, both in normal mice and in those previously treated with BCG, whereas the inoculation of Sarcoma 180 mixed with BCG produced a 37 % increase in the survival rate in normal animals and an 83 % increase in the group previously treated with BCG. In splenectomized mice the survival rate was 56 % in those inoculated with Sarcoma 180 alone, rising to 100 % in the group inoculated with Sarcoma 180 mixed with BCG. Previous immunization with BCG in splenectomized animals did not affect tumor incidence significantly. These results suggest the participation of the spleen in the mechanism of tumoral rejection favoured by BCG inoculation.

[Evaluation of the interleukin 1 production by adherent cells in patients with diabetes type II]. / Evaluación de la producción de interluequina 1 por células adherentes en pacientes con diabetes tipo II.

We have studied the comitogenic activity of IL 1 produced by cultures of mononuclear adherent cells obtained from Diabetes Mellitus (DM) type II or non insulin dependent diabetic patients. This activity was measured by the incorporation of 3H Thymidine into cultures of C3H/HeJ mice thymocytes in the presence of phytohemagglutinin (PHA). We have observed that IL 1 from patients with DM type II did not produce a synergistic effect with PHA, since the lymphoproliferation levels were similar to those obtained in the absence of this lectin. This lack of comitogenic activity (P less than 0.001) in relation to the response obtained with IL 1 from normal subjects plus PHA, could be due to the release of one or several soluble substances capable of blocking glycosylated receptors to mitogens or of impairing the cellular activation process.

[Production of interleukin 1 during tumor growth]. / Producción de interleuquina 1 durante el crecimiento tumoral.

The production of IL-1 by splenic mononuclear adherent cells (MAC) from BALB/c mice inoculated with Sarcoma 180 (S180) was examined as a possible mechanism underlying the immunosuppression observed in tumor bearing mice. Two different inducers of IL-1 were used to stimulate MAC. A natural polysaccharide PCj3 (1, 5, 10 micrograms/ml) and a lipopolysaccharide (LPS) of E. coli (1, 5, 10, 20 micrograms/ml). The IL-1 activity was assayed by the capacity of the supernatants of MAC cultures in different dilutions (1/5, 1/10, 1/20) to enhance the mitogenic response of murine thymocytes to phytohemagglutinin (PHA). Both stimulants induced comparable levels of IL-1 in normal and 10 day tumor bearing mice. Normal MAC were able to elaborate IL-1 spontaneously in the presence of 1% FCS: the optimum LPS concentration was 20 micrograms/ml in the 1/5 dilution. With regard to PCj3, the optimum concentration was 5 and 10 micrograms/ml in the 1/5 dilution (p less than 0.001 vs control). The maximum activity of IL-1 for MAC of 10 day tumor bearing mice was given by the concentration of 5 micrograms/ml for both stimulants. When MAC were incubated with LPS, the IL-1 production was dose dependent while PCj3 seemed to have reached a saturation level between 5 and 10 micrograms/ml. The increase in tumor size (day 20-30) was associated with a significant decrease in IL-1 production by MAC in response to both stimulants. Therefore, the immunosuppression associated with the late stages of tumor growth may be due to inhibition of IL-1 production.

[Cancer and BCG]. / Cáncer y BCG.

In the present paper the modification that BCG induces on experimental tumor growth and its possible human medical applications are analyzed. The mechanisms by which the BCG inoculated mixed with tumor cells enhance its rejection involve the participation of different cell populations, such as sensitized lymphocytes, activated macrophages and the spontaneous cytotoxicity effector cells (N. K. cells). The tumor rejection depends on the BCG doses and the inoculation modes. In this regard, high doses induce toxic effects and can activate suppressor mechanisms mediated by cells or serum factors. The action of BCG on tumoral growth is apparent only under certain circumstances where the host-tumor system should be carefully selected.