Diversity of somatic coliphages in coastal regions with different levels of anthropogenic activity in São Paulo State, Brazil.

Bacteriophages are the most abundant and genetically diverse viruses on Earth, with complex ecology in both quantitative and qualitative terms. Somatic coliphages (SC) have been reported to be good indicators of fecal pollution in seawater. This study focused on determining the concentration of SC and their diversity by electron microscopy of seawater, plankton, and bivalve samples collected at three coastal regions in São Paulo, Brazil. The SC counts varied from <1 to 3.4 × 10(3) PFU/100 ml in seawater (73 samples tested), from <1 to 4.7 × 10(2) PFU/g in plankton (46 samples tested), and from <1 to 2.2 × 10(1) PFU/g in bivalves (11 samples tested). In seawater samples, a relationship between the thermotolerant coliforms and Escherichia coli and SC was observed at the three regions (P = 0.0001) according to the anthropogenic activities present at each region. However, SC were found in plankton samples from three regions: Baixada Santista (17/20), Canal de São Sebastião (6/14), and Ubatuba (3/12). In seawater samples collected from Baixada Santista, four morphotypes were observed: A1 (4.5%), B1 (50%), C1 (36.4%), and D1 (9.1%). One coliphage, Siphoviridae type T1, had the longest tail: between 939 and 995 nm. In plankton samples, Siphoviridae (65.8%), Podoviridae (15.8%), Microviridae (15.8%), and Myoviridae (2.6%) were found. In bivalves, only the morphotype B1 was observed. These SC were associated with enteric hosts: enterobacteria, E. coli, Proteus, Salmonella, and Yersinia. Baixada Santista is an area containing a high level of fecal pollution compared to those in the Canal de São Sebastião and Ubatuba. This is the first report of coliphage diversity in seawater, plankton, and bivalve samples collected from São Paulo coastal regions. A better characterization of SC diversity in coastal environments will help with the management and evaluation of the microbiological risks for recreation, seafood cultivation, and consumption.

Chromosome aberrations in peripheral blood lymphocytes of individuals living in high background radiation areas of Ramsar, Iran.

In order to investigate the biological effects of exposure to low-dose radiation and to assess the dose-effect relationship in residents of high background radiation areas (HBRAs) of Ramsar, cytogenetic investigation of unstable-type aberrations was performed in 15 healthy elderly women in a HBRA of Ramsar, Talesh mahalle, and in 10 elderly women living in a nearby control area with normal background radiation. In total, 77,714 cells were analyzed; 48,819 cells in HBRA residents and 28,895 cells in controls. On average, 3,108 cells per subject were analyzed (range 1,475-5,007 cells). Significant differences were found in the frequency of dicentric plus centric rings in 100 cells (0.207 ± 0.103 vs. 0.047 ± 0.027, p < 0.0005), total chromosome-type aberrations per 100 cells (0.86 ± 0.44 vs. 0.23 ± 0.17, p < 0.0005), and chromatid-type aberrations per 100 cells (3.31 ± 2.01 vs. 1.66 ± 0.63, p = 0.01) by the Mann-Whitney U test between HBRA and the control, respectively. Using chromosomal aberrations as the main endpoint to assess the dose-effect relationship in residents of HBRAs in Ramsar, no positive correlation was found between the frequency of dicentric plus centric ring aberrations and the cumulative dose of the inhabitants estimated by direct individual dosimetry; however, obvious trends of increase with age appeared in the control group. Based on these results, individuals residing in HBRAs of Ramsar have an increased frequency of detectable abnormalities in unstable aberrations.

Construction of syndromic surveillance using a web-based daily questionnaire for health and its application at the G8 Hokkaido Toyako Summit meeting.

We constructed a syndromic surveillance system to collect directly information on daily health conditions directly from local residents via the internet [web-based daily questionnaire for health surveillance system (WDQH SS)]. This paper considers the feasibility of the WDQH SS and its ability to detect epidemics. A verification study revealed that our system was an effective surveillance system. We then applied an improved WDQH SS as a measure against public health concerns at the G8 Hokkaido Toyako Summit meeting in 2008. While in operation at the Summit, our system reported a fever alert that was consistent with a herpangina epidemic. The highly mobile WDQH SS described in this study has three main advantages: the earlier detection of epidemics, compared to other surveillance systems; the ability to collect data even on weekends and holidays; and a rapid system set-up that can be completed within 3 days.

Expression of human beta-defensin -1, -2, and -3 in non-inflamed pseudocyst, mucoceles.

OBJECTIVES AND DESIGN: The expressions of human beta defensin-1 (HBD-1), -2 (HBD-2) and -3 (HBD-3) in non-inflamed pseudocysts such as mucoceles were investigated immunohistochemically in this study. MATERIALS AND METHODS: Mucocele specimens were obtained from 21 patients. The expression of HBDs was studied immunohistochemically by using antibodies directed against HBD-1, -2, and -3. Statistical analyses were carried out on serial sections stained with antibodies. RESULTS: Cells expressing HBDs were found in mucoceles. The expression of HBD-2 was observed in floating cells in all the specimens, whereas HBD-1 and HBD-3-expressing cells were detected in 93% and 73% of the mucoceles, respectively. The HBD-2 signal was the most intense and the HBD-3 signal intensity was weaker than that of HBD-1. HBDs were expressed in neutrophils and in other floating cells. Interestingly, the signal intensity and the population of positive cells located close to the centers of cysts were higher than those located in the peripheral areas of cysts. CONCLUSION: The expression of HBDs was found even in non-inflamed pseudocysts such as mucoceles. These results suggest that an unknown mechanism not involved in biophylaxis for the expression of HBDs may exist.

Hypoxic regulation of stability of connective tissue growth factor/CCN2 mRNA by 3'-untranslated region interacting with a cellular protein in human chondrosarcoma cells.

Connective tissue growth factor (CTGF/CCN2) can be induced by various forms of stress such as exposure to high glucose, mechanical load, or hypoxia. Here, we investigated the molecular mechanism involved in the induction of ctgf/ccn2 by hypoxia in a human chondrosarcoma cell line, HCS-2/8. Hypoxia increased the ctgf/ccn2 mRNA level by altering the 3'-untranslated region (UTR)-mediated mRNA stability without requiring de novo protein synthesis. After a series of extensive analyses, we eventually found that the cis-repressive element of 84 bases within the 3'-UTR specifically bound to a cytoplasmic/nuclear protein. By conducting a UV crosslinking assay, we found the cytoplasmic/nuclear protein to be a 35 kDa molecule that bound to the cis-element in a hypoxia-inducible manner. These results suggest that a cis-element in the 3'-UTR of ctgf/ccn2 mRNA and trans-factor counterpart(s) play an important role in the post-transcriptional regulation by determining the stability of ctgf/ccn2 mRNA.

Design of stable biologically active recombinant lutropin analogs.

Glycoprotein hormones are noncovalent heterodimers comprised of a common alpha subunit and a hormone-specific beta subunit. Secretion and biologic action of these hormones are dependent on the formation of the heterodimer. The human LH beta subunit is unique among the other beta subunits in that it assembles inefficiently with the alpha subunit. To bypass this rate-limiting step, we constructed the LH single chains where the carboxy terminus of beta was fused to the amino terminus of alpha subunit through a linker. Compared to the human LH heterodimer, the extent of secretion was greater for the tethers although the rate was dependent on the nature of the linker. The LH single chains were biologically active even though there was loss of recognition by a LH-specific monoclonal antibody. This suggests that receptor binding of the single chains is not impaired by changes in the heterodimeric configuration resulting from tethering the subunits. In addition, single chains exhibited a remarkably greater in vitro stability than the heterodimer, implying that these analogs will be useful as diagnostic reagents and that their purification will be facilitated.

Radiation induction of endogenous type C virus from mouse cells transformed in vitro by murine sarcoma virus.

Cell lines from AKR and BALB/c mouse embryos were compared for their sensitivity to X-ray induction of endogenous type C virus. K-Balb cells, a Balb/3T3 cell line nonproductively transformed by Kirsten murine sarcoma virus, were found to be sensitive to X-irradiation. At a dose as low as 50 R, X-rays induced virus expression in K-Balb cells, and the induction frequency increased with increasing dose of X-rays up to 400 R. Among two classes of inducible endogenous viruses carried by K-Balb cells, only Balb:virus-2 was activated by X-irradiation, whereas both Balb:virus-1 and Balb:virus-2 were activated after the cells were treated with 5-iodo-2'-deoxyuridine. UV light and 4-nitroquinoline 1-oxide were also shown to induce virus expression in K-Balb cells. The virus-induction frequency for these physical and chemical carcinogens was much lower (approximately 3 times 10(-4)) than that for 5-bromo-2'-deoxyuridine (approximately 1 times 10(-1)).

Effect of caffeine on induction of endogenous type C virus in mouse cells in vitro.

The effect of caffeine on the expression of murine endogenous virus in mouse cells induced by radiation and chemicals was studied. Postirradiation treatment of K-BALB cells with caffeine enhanced cell killing as well as the induction of xenotropic virus after ultraviolet light irradiation. The degree of enhancement for the virus induction was comparable to that for cell killing. On the other hand, colony-forming ability and the expression of xenotropic virus of K-BALB cells after X-irradiation were unaffected by caffeine. These data suggest a linear relationship between the degree of endogenous virus expression and the amount of lethal damages after irradiation. For induction by halogenated pyrimidines, a 24-hr incubation of AKR2B cells with caffeine after 5-iodo-2'-deoxyuridine treatment resulted in marked suppression of the expression of ecotropic virus. On the contrary, in K-BALB cells, caffeine exerted only a small effect on 5-iodo-2'-deoxyuridine-induced expression of ecotropic and xenotropic viruses. These results indicate that, although using the same inducing agent, the pathway of endogenous virus induction may be different for AKR2B cells and for K-BALB cells.

Comparison of drug sensitivity among tumor cells within a tumor, between primary tumor and metastases, and between different metastases in the human tumor colony-forming assay.

The human tumor colony-forming assay was used to compare chemosensitivity among tumor cells within a primary tumor, between primary tumor and metastases, and between different metastases. No significant differences in cloning efficiency were found in any of the three comparison studies. However, considerable differences in chemosensitivities were observed between different parts of the same tumor and between the primary tumor and metastases. Two different parts of the same tumor were comparably assayed for nine primary tumors. In nine paired samples which allowed in vitro drug sensitivity testing, there was no satisfactory correlation of sensitivity to cytostatic drugs. Cell suspensions were prepared from 28 primary tumors and from metastases taken from the same patient. In 14 paired samples which formed sufficient colonies for determination of drug effect, the data showed no satisfactory correlation of chemosensitivity between a primary tumor and its metastases. Both tumor samples from different metastatic sites of the same patient formed sufficient colonies in seven of eight instances. In the seven paired samples, there was strong association of chemosensitivity (p less than 0.005). The results indicate that the reported discrepancies of in vitro and in vivo results in clinical trials using the tumor colony-forming assay for predicting resistance or sensitivity to cytostatic drugs may be due to therapeutic heterogeneity among tumor colony-forming units within a primary tumor and between a primary tumor and its metastases.

The mode of actions of lysozyme as an immunoglobulin production stimulating factor.

As we demonstrated before, hen egg white lysozyme stimulates immunoglobulin production by a human-human hybridoma line, HB4C5 cells and human peripheral blood lymphocytes. Then, the mode of actions of lysozyme as an immunoglobulin production stimulating factor was investigated. The immunoglobulin production stimulating activity of lysozyme was inactivated by trypsin digestion, even though the enzymatic activity was completely preserved. This fact suggests that the immunoglobulin production stimulating effect of lysozyme is irrelevant to its enzymatic function. Furthermore, this means that the effect is a novel function of this enzyme. Lysozyme enhanced IgM production by transcription-suppressed HB4C5 cells treated with actinomycin D. However, the enzyme was ineffective to accelerate IgM production by translation-suppressed HB4C5 cells treated with cycloheximide or sodium fluoride. In addition, the intracellular IgM content of HB4C5 cells treated with monensin for suppression of the post-transcription activity was obviously increased by lysozyme, although the secretion of IgM was inhibited. These findings suggest that lysozyme accelerates the translation process to enhance immunoglobulin productivity.

A novel function of enolase from rabbit muscle; an immunoglobulin production stimulating factor.

Rabbit muscle enolase stimulates immunoglobulin production by a human hybridoma line, HB4C5 cells under serum-free condition. IgM productivity of HB4C5 cells was enhanced more than 20-fold by this enzyme at 220 micro/ml. Human peripheral blood lymphocytes were also facilitated their IgM and IgG productivity in the serum-free medium. However, baker's yeast enolase was ineffective to accelerate immunoglobulin production by HB4C5 cells, in spite of the same specific enzymatic activity as rabbit muscle enolase. There were differences in sensitivities against heat treatment and trypsin digestion between IPSF and enzymatic activities of enolase. These results imply that the immunoglobulin production stimulating effect of rabbit muscle enolase is irrelevant to its enzymatic function and reaction products. This fact also means that this enzyme has another function other than enzymatic one in glycolysis. Rabbit muscle enolase enhanced IgM production of transcription-suppressed HB4C5 cells treated with actinomycin D. Cycloheximide treatment of HB4C5 cells was useless to inhibit the expression of immunoglobulin production stimulating activity. However, inhibition of post-transcriptional process by monensin invalidated the activity of enolase. These findings suggest that enolase from rabbit muscle accelerates the steps between translation and post-translational processes to enhance immunoglobulin productivity. In addition, laser confocal microscopic analysis revealed that enolase from rabbit muscle was subsequently incorporated by HB4C5 cells.

The carboxy-terminal region of the beta-subunits of luteinizing hormone and chorionic gonadotropin differentially influence secretion and assembly of the heterodimers.

One of the major structural differences between the LH beta and CG beta subunits is the carboxy-terminal region: beyond amino acid 114, LH beta has a hydrophobic heptapeptide stretch, while CG beta contains a 31-amino acid hydrophilic carboxy-terminal peptide (CTP) that is O-glycosylated. The CG beta subunit is secreted quantitatively as a monomer and assembles efficiently whereas secretion and assembly of LH beta is inefficient. We previously implicated the carboxy-terminal heptapeptide as a determinant for the different intracellular behavior manifested by the LH beta subunit compared with the CG beta subunit. Here we tested the function of the heptapeptide and CTP domains by fusing them to their counterparts at amino acid 114 of CG beta or LH beta subunits. The secretion and assembly of these chimeras were examined in transfected Chinese hamster ovary cells. Removal of the heptapeptide enhanced the amount of LH beta subunit secreted 4-fold compared with intact LH beta. Fusion of this heptapeptide to CG beta 114, i.e. CG beta lacking the CTP, decreased the amount of secreted subunit 2-fold compared with wild type human CG beta. Similar experiments reveal that although deleting the CTP from the CG beta subunit did not significantly alter secretion, the combination efficiency of the truncated subunit was reduced to 60%. Perturbing the native carboxy-terminal sequence of either subunit increased the heterogeneity of the secreted forms. This result suggests that these regions are also involved in the posttranslational processing of the asparagine-linked oligosaccharides of the beta-subunits. Fusion of the LH beta heptapeptide to the truncated CG beta subunit decreased combination with the alpha-subunit. These data further support the hypothesis that the carboxy-terminal regions of LH beta and CG beta subunits play a role in the intracellular behavior of the corresponding heterodimers.

Fusing the carboxy-terminal peptide of the chorionic gonadotropin (CG) beta-subunit to the common alpha-subunit: retention of O-linked glycosylation and enhanced in vivo bioactivity of chimeric human CG.

The hCG beta-subunit contains a carboxy-terminal extension bearing four serine-linked oligosaccharides [carboxy-terminal peptide (CTP)], which is important for maintaining its longer half-life compared with the other glycoprotein hormones. Previously, we enhanced the in vivo half-life of FSH by fusing the CTP to the carboxy end of FSH beta coding sequence. The alpha-subunit is common to the glycoprotein family. We constructed alpha-subunit CTP chimeras, since such analogs with the appropriate O-linked glycosylation and conformation would increase the in vivo stability of the entire glycoprotein hormone family. Two chimeras were constructed using overlapping polymerase chain reaction mutagenesis: a variant with CTP at the carboxy end and another analog with the CTP at the N-terminal region of the subunit, between amino acids 3 and 4. The latter design was based on models showing that the amino-terminal region of alpha is not involved in assembly with the beta-subunit, nor is it essential for receptor binding and signal transduction. These chimeras were cotransfected with the hCG beta gene into Chinese hamster ovary cells. The chimeras were secreted and combined efficiently with the CG beta-subunit, comparable to the wild type alpha-subunit. CG dimers containing the alpha-subunit chimera with CTP at the carboxy end of the subunit had a much lower binding affinity for the hLH-hCG receptor in vitro, whereas the binding of the dimer containing the CTP at the amino-terminal end of the subunit was similar to wild type hCG. Furthermore, the in vivo activity of this analog was enhanced significantly. Moreover, regardless of the two insertion points in the alpha-subunit, the CTP sequence was O-glycosylated. These data suggest that the entire signal for O-glycosylation is primarily contained within the CTP sequence and is not dependent on the flanking regions of the recipient protein. The transfer of CTP to the alpha-subunit of hCG results in an agonist with prolonged biological action in vivo. These data further support the rationale for using the CTP as a general target to increase the potency of bioactive glycoproteins.

Effect of ENU (ethylnitrosourea) mutagenesis in cleft lip and/or palate pathogenesis in mice.

Cleft lip and/or palate (CL/P) are caused by many factors. The aim of this study was to investigate the effects of genetic point mutations in CL/P pathogenesis. ICR and AJ strain mice were used. Ethylnitrosourea (ENU) was injected into 10-week-old male mice (G0) intraperitoneally at a dose of 250 mg/kg. The males were bred with two untreated virgin females of the same strain on day 100 after injection. The uterine contents (G1) of one female were examined on day 18.5 of pregnancy. From the other female, the offspring were delivered naturally, and F3 mice (G3) were also examined in the same way. In ICR strain mice, cleft palate only (CPO) was increased in both the G1 and G3. The frequency was significantly higher in the G3 than the G1 generation. Cleft lip was not observed. In AJ strain mice, CL/P increased in both the G1 and G3. In the G3, the frequency of CPO was increased significantly. Genes related to CPO may be recessive in phenotype. CPO and cleft lip differ from a genetic viewpoint. Point mutations play a significant role in cleft lip and palate.

The mode of actions of glyceraldehyde-3-phosphate dehydrogenase identified as an immunoglobulin production stimulating factor.

Immunoglobulin production stimulating factor-II alpha, which enhances immunoglobulin production of human and mouse hybridomas was purified from cell lysate of human Burkitt's lymphoma, Namalwa cells, and identified as glyceraldehyde-3-phosphate dehydrogenase. The enhancement of immunoglobulin production with this enzyme was not linked with its enzymatic activity. The enzyme enhanced immunoglobulin productivity of transcription-suppressed hybridomas, but did not enhance that of translation-suppressed hybridomas. From these results, it is suggested that this enzyme takes part in the post-translational control or the enhancement of translation activity to stimulate immunoglobulin production of hybridomas.

Expression of osteoclast differentiation factor and osteoclastogenesis inhibitory factor in rat osteoporosis induced by immunosuppressant FK506.

Immunosuppressant drugs are currently required by transplant recipients for the remainder of their lives, despite the many adverse effects associated with these therapies. Acute osteoporosis is one such effect, and a reproducible osteoporosis model has been established through the administration of the immunosuppressant drug FK506 in rats. The cause of this osteoporosis has been shown to be abnormal osteoclast proliferation, altering the process of bone remodeling. However, the reasons why FK506 induces osteoclast proliferation and whether this process is mediated by cytokine changes or an increase in bone resorption factors have been unclear. An investigation was therefore conducted focusing on the recent discoveries of osteoclast differentiation factor (ODF) and osteoclastogenesis inhibitory factor (OCIF). These factors led to elucidation of the osteoclast differentiation-maturation mechanism. An osteoporosis model was produced in rats utilizing intramuscular FK506 injection (1 mg/kg) for 28 consecutive days. Trabecular bone resorption was observed inferior to enchondral ossification in the FK506 group, and tartrate resistant acid phosphatase (TRAP) staining revealed a clear increase in osteoclasts at the site of enchondral ossification, relative to the control group. Real-time PCR and in situ hybridization (ISH) demonstrated minimal differences in OCIF expression between control and the treatment groups. However, Real-time PCR revealed clearly increased ODF expression in the treatment group. ODF expression was also shown to be increased in the treatment group using ISH. This was histologically consistent with a region of osteoclast proliferation inferior to enchondral ossification. The results of this study support the hypothesis that FK506-mediated osteoporosis occurs by action of the drug on osteoclasts, promoting expression of ODF messenger ribonucleic acid (mRNA) and thus prompting osteoclast differentiation and maturation.

Differential action on cancer and normal tissue by adrenochrome monoaminoguanidine methanesulfonate and cytochrome C combined with radiotherapy.

PURPOSE: The possibility that radioprotective effects on potent natural killer (NK) cells by adrenochrome monoaminoguanidine methanesulfonate+cytochrome C during radiotherapy (RT) for lung cancer might result in the radiosensitization of human lung cancer cells in vivo is examined. METHODS AND MATERIALS: Human lung cancer xenografts in the right hind legs of KSN mice (10 weeks old) were locally irradiated with 20 Gy of X ray. Adrenochrome monoaminogluanidine methanesulfonate (AMM) (10 mg/kg/day) and/or cytochrome C (CCC) (5 mg/kg/day) were given intraperitoneally immediately before or after RT, followed by daily administration for 4 days. Natural killer activities of host splenocytes were also tested with the standard 51Cr releasing assay with YAC-1 cells as target cells. In a clinical study, 65 patients with lung cancer were treated with more than 50 Gy of RT with or without combination with AMM+CCC, OK-432 or AMM+CCC+OK-432. Before and after RT, lymphocyte subsets in the peripheral blood were examined with dichromatic analysis using an Ortho Spectrum IIIFCM system and fluorescent MABs. In this study, the change in the absolute number of each subset was investigated. RESULTS: Adrenochrome monoaminoguanidine methanesulfonate+cytochrome C augumented NK activity in KSN nude mice, protected potent NK cells in patients with lung cancer against RT and sensitized the human lung cancer xenografts to RT. CONCLUSION: Adrenochrome monoaminoguanidine methanesulfonate+cytochrome C may have the potential as a differential modulator of radiosensitivity of normal tissues and of tumors.

Effects of inhibitors of radiation-induced potentially lethal damage repair on chemotherapy in murine tumors.

Enhancement of various antitumor drugs effects by inhibitors of radiation-induced potentially lethal damage (PLD) repair was studied in three murine tumors (EMT-6, RIF-1 and SQ-1). In EMT-6 tumors, PLD repair inhibitors, 3'-deoxyguanosine (3'-dG) and 7904 (a derivative of 3'-deoxyadenosine) showed a marked enhancement of tumor growth inhibition by anticancerous drugs (FT-207 (a derivative of 5-FU), bleomycin, Ara-C, ACNU). However, the effects of mitomycin-C and vincristine were not potentiated by the inhibitors. In SQ-1 carcinomas, another repair inhibitor, ara-A (1-beta-D-arabinofuranosyladenine) (32 mg/kg) potentiated the effect of ACNU. In RIF-1 sarcomas, in which a low PLD repair function has been reported after ionizing radiation exposure, the potentiation was not so marked as in EMT-6 or SQ-1 tumors. Thus, as a possibility, the potentiation by inhibitors of radiation-induced PLD repair might be a result of the inhibition of chemical-induced PLD repair. The study of this field may contribute to the improvement of cancer treatment not only by radiotherapy but also by chemotherapy.

Dephosphorylation of WR-2721 with mouse tissue homogenates.

Mouse liver homogenate had an optimum pH of 8.6 to 8.8 for dephosphorylation of WR-2721 in the analyzed pH range from 5.2 to 10.0. At this optimum pH condition, the dephosphorylation activities of six mouse tissue homogenates were analyzed. Kidney, liver and small intestine homogenates showed higher dephosphorylation activities (935, 336 and 314 nmoles/mg protein/hr, respectively) than spleen and lung homogenates (86, 49 nmoles/mg protein/hr, respectively). Furthermore, serum did not show any dephosphorylation activity. The high activity found in liver homogenate agrees well with our previous data with mouse L cells. However, optimum pH from 8.6 to 8.8 in liver homogenate is quite different from the data reported by using Ehrlich ascites tumor cells (optimum pH was 5.6). Therefore, it is suggested that WR-2721 administered into mouse is efficiently dephosphorylated in certain tissues such as liver to its active form with the enzyme(s) different from that found in Ehrlich ascites tumor cells.

Effects of combination of immunomodulators and an adrenochrome derivative on survival of irradiated mice.

PURPOSE: The combined effects of immunomodulators (lithium or OK432) and an adrenochrome derivative (AMM), an agent found to activate granulocyte-macrophage colony stimulating activity, on the survival of irradiated ddY mice is described. METHODS AND MATERIALS: ddY mice at 4-5 weeks old were whole body irradiated with X rays at 8.5 Gy. Sole injection and combined injection of AMM and/or one of the immunomodulators were performed before or after irradiation. Then, survival was monitored daily for 30 days after irradiation. RESULTS: Lithium at 60 mg/kg had no radioprotective effect; rather it accelerated radiation induced death. Sole treatment with AMM (100 mg/kg) had no effect on survival of irradiated mice. However, combination of both drugs caused a slight radioprotection. OK432 (25 KE/kg), which activates a variety of cellular effector cells had radioprotective effect. When combined with AMM, however, it totally lost radioprotective effect. CONCLUSION: Lithium chloride cannot be used as a radioprotector because of its adverse effect. Combination with AMM showed slight radioprotection, but the extent thereof may not be clinically useful. OK432 was proved to be a potent radioprotector. However, combination with AMM should be avoided, since the radioprotection was totally eliminated.