Distinct Expression of Cytokines and Mitogenic Inhibitory Factors in Semen of Fertile and Infertile Men

PROBLEM: To assess the effect of seminal plasma (SP) of fertile and infertile men on leukocyte mitogenic response, and the capability of sperm cells to produce IL-1. METHODS: This study included four groups: fertile men (donors, normal), infertile men with azoospermia (azoo), oligo-terato-asthenozoospermia (OTA), and OTA with genital infection (OTA-inf). Mouse spleen cell proliferation in response to lipopolysaccharide (LPS) or Concanavalin-A (Con-A) was examined in the presence of SP from the above four groups. Supernatants (sup) and lysates (lys) of sperm cells from fertile and oligoteratoasthenospermic (OTA) men were evaluated for IL-1 bioactivity by specific bioassay. RESULTS: Seminal plasma (SP) of the four groups were shown to inhibit the mitogenic response of mouse spleen cells to LPS and Con-A. SP of fertile men was significantly more inhibitory than SP from infertile men. Sperm cells from fertile and OTA infertile men constitutively produced IL-1. Sperm cells of both groups produced similar levels of IL-1 as examined in the supernatants and lysates. CONCLUSIONS: Seminal plasma of fertile men had more inhibitory mitogenic activity than that of OTA. Sperm cells constitutively produce IL-1. It is possible that the factors involved in this inhibition are not only anti-proliferative immune factors. Cytokines and inhibitory factors of mitogenesis in the seminal plasma may be involved in the physiology and pathophysiology of sperm functions and thus affect male fertility.     

Spontaneous and ionophore induced acrosome reaction in asthenozoospermic infertile semen.

Spontaneous and ionophore-induced ability of spermatozoa to acrosome-react was examined in asthenozoospermic infertile patients and fertile donors. Spermatozoa were washed free of seminal plasma and capacitated in B2 medium for 2 h at 37 degrees C. Subsequently 5, 10, 20 and 30 microM A23187 (final concentrations) were added to equal aliquots of these samples and incubated for an additional 30 min. The acrosome reaction was then determined by the triple stain technique. The percentage of spontaneous reaction (no ionophore) in asthenozoospermic samples was similar to that in fertile samples (4.2 and 3.8, respectively). However, the ionophore-induced reaction rate remained significantly lower in asthenozoospermic samples than in normozoospermic samples.     

Increased resistance towards oxidative stress accompanies enhancement of metastatic potential obtained by repeated in vivo passage of colon carcinoma cells in syngeneic rats

The colon carcinoma cell line CC531 is metastatic to liver after splenic injection in syngeneic rats. After repeated in vivo passages, a subline was selected that produced liver metastases at a considerably higher rate than the original cell line. These cells were characterized by increased intracellular glutathione, proliferation and ability to restore glutathione after exposure to oxidative stress, thus indicating an elevated resistance to oxidative stress. Furthermore, the increased metastatic ability was also accompanied by increased proliferation rate, adhesion to extracellular matrix proteins and endothelial cells, and secretion of a 60 kD matrix metalloproteinase. When cultured in vitro for a prolonged time (more than 30 trypsinizations), the cells showed a reduced in vivo metastatic ability, reduced secretion of three metalloproteinases including the 60 kD proteinase, and reduced intracellular glutathione. These results indicate that metastatic ability can be influenced through several adaptive mechanisms, and that the cell's ability to resist oxidative stress and maintain intracellular glutathione are of central importance. This revised version was published online in July 2006 with corrections to the Cover Date.     

A randomized controlled trial of lycopene treatment on soluble receptor for advanced glycation end products in seminal and blood plasma of normospermic men

Citation
Oborna I, Malickova K, Fingerova H, Brezinova J, Horka P, Novotny J, Bryndova H, Filipcikova R, Svobodova M. A Randomized controlled trial of lycopene treatment on soluble receptor for advanced glycation end products in seminal and blood plasma of normospermic men. Am J Reprod Immunol 2011; 66: 179–184 Problem The aim of present study was to investigate the effects of antioxidant lycopene on soluble receptor for advanced glycation end products (sRAGE) levels in blood and seminal plasma in normospermic males. Methods Study included 15 fertile volunteers and 13 normospermic male partners from infertile relationships. The treatment was 12‐week administration of 20 mg of lycopene or placebo followed by crossover and treatment for a further 12 weeks. The ELISA kit Quantikine^® was used to determine sRAGE levels. Results Lycopene administration decreased sRAGE levels in seminal plasma in fertile volunteers (controls) as well as in male partners in the infertile relationships group (P = 0.008 and P = 0.012, respectively). No significant effect of lycopene on sRAGE in blood plasma was found in either group, but seminal plasma sRAGE was significantly suppressed. Conclusion Lycopene decreased sRAGE in seminal, but not in blood plasma. This may be because of selective local uptake of lycopene in the male reproductive tract, namely in prostate. Decreased sRAGE may be caused by lycopene suppression of oxidative stressors and explain in part the putative improvement in fertility reported after lycopene treatment.     

Allergen-induced bronchial inflammation is associated with decreased levels of surfactant proteins A and D in a murine model of asthma

Background Increasing evidence suggests that pulmonary surfactant protein A (SP‐A) and D (SP‐D) participate in the lung defence against pathogens. However, the role of surfactant proteins in the pathogenesis of allergen‐induced airway inflammation has not been elucidated. In this study we examined the levels and distributions of SP‐A and SP‐D in a dust mite (Dermatophagoides pteronyssinus, Der p) allergen‐induced murine model of asthma. Methods The concentration of SP‐A and SP‐D in the bronchoalveolar lavage fluid (BALF) and the distribution of surfactant proteins in the lung were assayed by ELISA and immunohistochemistry methods, respectively. The effect of surfactant proteins on allergen‐induced pulmonary lymphocyte proliferation was also studied. Results We demonstrated that there were marked reductions of SP‐A and SP‐D levels in the BALF of Der p‐sensitized BALB/c mice at 48–72 h after allergen challenge (AC). Both purified SP‐A and SP‐D were able to suppress, in a dose dependent manner, Der p‐stimulated intrapulmonary lymphocyte proliferation of naïve mice with saline or allergen challenge, or of Der p‐sensitized mice with saline challenge. On the contrary, this suppressive effect was mild (< 9%) on lymphocytes from sensitized mice after AC. Conclusion These results indicated the involvement of pulmonary surfactant proteins in the allergic bronchial inflammation of sensitized mice.     

Detection of Monocyte Chemotactic and Activating Factor (MCAF) and Interleukin (IL)-6 in Human Seminal Plasma and Effect of Leukospermia on These Cytokine Levels

PROBLEM : To demonstrate whether monocyte chemotactic and activating factor (MCAF) and interleukin-6 (IL-6) are present in the seminal plasma, and whether these presence is modulated by leukospermia. METHODS : Semen samples from 53 men were obtained by masturbation and examined for the presence of MCAF and IL-6 by enzyme immunoassay (EIA). Semen samples were obtained from 28 infertile men without leukospermia, 16 infertile men with leukospermia, and nine proven-fertile men. The correlation between the amount of MCAF in the seminal plasma with some spermiogram parameters and other cytokines such as IL-6 and IL-8 was statistically evaluated. RESULTS : Immunoreactive MCAF was detected in the seminal plasmas of all 53 subjects. The MCAF titer in the seminal plasma of patients with leukospermia (11.19 ± 2.75 μg/1) was significantly higher than that in the seminal plasma of the patients without leukospermia (3.24 ± 0.53 μg/1) and the fertile men (2.78 ± 0.35 μg/1) (P < 0.001). The IL-6 titer in the seminal plasma of the patients with leukospermia (21.05 ± 4.49 ng/1) was also significantly higher than that in the seminal plasma of the patients without leukospermia (8.77 ± 1.92 ng/1) and the fertile men (6.94 ± 1.27 ng/1) (P < 0.01). There was a high degree of correlation among the levels of MCAF, IL-6 and IL-8 in the seminal plasma. CONCLUSIONS : These findings demonstrated the presence of MCAF and IL-6 in the seminal plasma, and that the levels of these cytokines were elevated in the seminal plasma of the infertile patients with leukospermia.     

Intimal plaque development and oxidative stress in cholesterol‐induced atherosclerosis in New Zealand rabbits

Although oxidative stress is well known in atherogenesis, the origin, nature and kinetics of free radicals involved have not been well described till now. Here, we correlated parameters of oxidative stress with cellular components during induction and stabilization of aortic intimal lesions which were induced in rabbits by feeding a cholesterol‐enriched diet for 6 weeks and a normal diet for further 68 weeks. Plasma lipids, aortic plaque size and composition (macrophages, smooth muscle cells, oxidized LDL by morphometry), as well as aortic radical production (by luminol‐enhanced chemiluminescence and TEMPO‐9AC fluorescence) were measured after various time points. The parameters of oxidative stress were correlated with the different cellular components of the aortic plaques. The plaques increased until week 21, no significant regression was found until week 74, plasma cholesterol was maximal at week 6. Macrophages, oxidized LDL and generation of different species of free radicals were increased during plaque development, yet with different time kinetics. Whereas chemiluminescence correlated only weakly with the amount of intimal macrophages, strong correlations were found between TEMPO fluorescence and smooth muscle cells (r = 0.4778, P < 0.001) and between macrophages and oxidized LDL (r = 0.5896, P < 0.0001). Different indicators of oxidative stress were increased during plaque progression and stabilization. However, the various correlations show, that distinct types of reactive species secreted probably from macrophages and smooth muscle cells contribute to oxidative stress in the different phases of plaque development.     

Combined Polymorphisms in Oxidative Stress Genes Predict Coronary Artery Disease and Oxidative Stress in Coronary Angiography Patients

Oxidative stress has been implicated in all stages of atherosclerosis, but how inherited variations in oxidative stress genes influence the severity of cardiovascular disease is not known. We tested associations between polymorphisms in candidate oxidative stress genes, plasma oxidative stress biomarkers, and cardiovascular mortality in an angiography cohort. Single nucleotide polymorphisms (SNPs) across 15 genes were selected by linkage disequilibrium tagging. Genotyping was performed using customized arrayed primer extension micro‐arrays, with automated genotype calling methods. Effects of SNPs and haplotypes on plasma oxidative stress and coronary artery disease (CAD) were estimated using a stochastic estimation maximization algorithm. Proportionate hazards analyses were used to determine effects of single and combined genetic markers on cardiovascular mortality risk, and on the following oxidative stress biomarkers: myeloperoxidase (MPO), nitrotyrosine, oxidized low‐density lipoprotein, and antioxidant capacity. Oxidative stress gene SNPs associated with CAD were combined into an oxidative stress risk allele score, which predicted disease presence (1.5‐fold risk increase per allele, P < 0.001). Combined risk alleles were also associated with elevated plasma MPO (P < 0.003), an oxidative stress biomarker that predicts cardiovascular mortality. Genetic markers that represent lifetime oxidative stress burden may implicate specific oxidative stress pathways in the pathogenesis of atherosclerosis, and offer therapeutic opportunities.     

Characterization of Latent Transforming Growth Factor-β From Human Seminal Plasma

PROBLEM : Human seminal plasma is known to exhibit immunosuppressive activity. Transforming growth factor β (TGF-β) has been identified as an immunosuppressive factor in human seminal plasma. Biologically active TGF-β represents a family of 25-kDa homodimeric proteins linked with disulfide bonds. TGF-β associates with high molecular weight proteins noncovalently to form a type of latency that is biologically inactive. Quantitative distribution of active form of TGF-β versus inactive latent form of TGF-β, and mechanism of the TGF-β activation in human seminal plasma remain to be elucidated. PURPOSE : To characterize seminal plasma latent form of TGF-β, including its concentration, and the mechanism underlying the activation of TGF-β. METHOD : Gel filtrations on ACA-34 and Biogel P-60 were used to fractionate seminal plasma. TGF-β was measured by enzyme immunoassay using antibodies specific for TGF-β₁ and TGF-β₂, respectively. Radioreceptor assay with recombinant human [¹²⁵I]-TGF-β₁ was applied to qualitatively identify TGF-β₁. Kinetic experiments with various pH, temperature and time, along with protease inhibitors, were performed to delineate the activation mechanism of latent TGF-β. RESULTS : Human seminal plasma contained both TGF-β₁ and TGF-β₂, predominantly in latent form. The total concentration of TGF-β₁ averaged 238 ng/ml versus an average of 18 ng/ml for TGF-β₂. The in vitro activation or release of TGF-β₁, from latent TGF-β₁ was achieved only at acidic pH of <4.0, and was time and temperature dependent. At pH 3.7 and 37°C, a significant activation of latent TGF-β₁ was achieved after an incubation of only 15 min, reached the maximum at 120 min, and the activated TGF-β₁ remained relatively stable for at least 24 h. The activation was not inhibitable by a series of protease inhibitors examined, alone or in combination (e.g., phenylmethylsulfonyl fluoride, E-64, pepstatin, leupeptin, ethylenediamine tetraacetic acid). Competitive radioreceptor assay established the functional identity of TGF-β₁ in human seminal plasma with recombinant human TGF-β₁. CONCLUSION : Human seminal plasma TGF-β is biologically activated from high molecular weight latent TGF-β by acid pH. The acidic environment of female lower genital tract could represent an in vivo physiological condition for activation of seminal plasma TGF-β that may immunologically protect the integrity of sperm.     

Autoimmune T cell responses to seminal plasma in chronic pelvic pain syndrome (CPPS)

The aetiology of chronic prostatitis is not understood. The aim of this study is to investigate an autoimmune hypothesis by looking for T cell proliferation in response to proteins of the seminal plasma. We studied peripheral blood mononuclear cell proliferation from 20 patients with chronic prostatitis and 20 aged-matched controls in response to serial dilutions of seminal plasma (SP) from themselves (autologous SP) and from a healthy individual without the disease (allo-SP). We found that the patients have a statistically greater lymphocyte proliferation to autologous SP at the 1/50 dilution on day 6 compared to controls (P = 0 x 01). They also have a greater proliferation to allo-SP on both day 5 (P = 0 x 001) and day 6 (P = 0 x 01) at the same dilution. Using a stimulation index (SI) of 9 to either autologous SP or allo-SP on day 6 at the 1/50 dilution as a definition of a proliferative response to SP, then 13/20 patients as compared to 3/20 controls showed a proliferative response to SP (P = 0 x 003, Fishers exact test). These data support an autoimmune hypothesis for chronic prostatitis.     

Transforming Growth Factor β as an Immunosuppressive Protein in Human Seminal Plasma

PROBLEM: Human seminal plasma is known to exhibit immunosuppressive activity in seminal plasma. PURPOSE: The purpose was to characterize immunosuppressive proteins in seminal plasma. METHOD: Gel filtration fractions of 100 to >440 kDa were identified that inhibited DNA synthesis and killing activity of interleukin-2 stimulated lymphocytes. RESULTS: The fractions exhibiting immunosuppression also inhibited DNA synthesis in a mink lung cell bioassay commonly used to measure the activity for transforming growth factor β (TGF-β). The negative growth activity was diminished by aTGF-β neutralizing monoclonal antibody. TGF-β was further detected in the active fractions by Western immunoblot. CONCLUSIONS: These results identified TGF-β as an immunosuppressive protein in human seminal plasma and may provide insight into the role of immunosuppression played by seminal plasma, such as in reproduction and neoplasia.     

Tubeimoside-1 inhibits the proliferation and activation of mouse T lymphocytes through signal transduction pathways

Abstract

Tubeimoside-1 (TBMS1) is one of the important components in Bolbostemma paniculatum (Maxim.) Franque. In the study, its immunosuppressive effects on murine T lymphocyte responses were evaluated in vitro and in vivo. The data showed that TBMS1 inhibited ConA-induced T lymphocyte proliferation, decreased the ratio of CD4⁺/CD8⁺, suppressed IL-2, IFN-γ, IL-4 and IL-6 production and mRNA expression, down-regulate activation of NF-κB, NFAT2 and AP-1 signal transduction pathways in vitro. In addition, administration of TBMS1 significantly inhibited T cell-mediated DTH response in vivo. These findings indicated that TBMS1 inhibits the proliferation and activation of T lymphocytes in mice.     

Human interdigitating dendritic cells directly stimulate CD40-activated naive B cells

Human interdigitating dendritic cells (IDC) were isolated from tonsils based on their CD40⁺ lineage-negative expression in situ. Isolated IDC displayed a phenotypic profile similar to that of IDC in tonsils and spleen in situ, characterized by high-level expression of major histocompatibility complex class II, the co-stimulatory molecules B7.1 (CD80) and B7.2 (CD86), expression of the late DC maturation marker CD83, and no expression of CD1a, CD13, or CD33. IDC also showed weak nonspecific esterase staining and had the ability to induce an allogeneic mixed lymphocyte reaction. In this study, we further show that in the presence of surrogate activated T cells in the form of CD40 ligation and IL-2, IDC enhance the proliferation of naive B cells and induce their differentiation into plasma cells producing IgM. Evidence for the anatomical co-localization of naive B cells and IDC in the T cell area together with the data obtained in vitro implies a role for IDC in the initiation of the extrafollicular reaction.     

Human seminal plasma suppresses delayed-type hypersensitivity responses to intravaginally deposited sheep red blood cells and sperm: separation of immunosuppressive factors

Although significant progress has been made in identifying the immunosuppressive factors in seminal plasma (SP) and their possible action in vitro, the potential role of SP in naturally occurring normal immune responses in vivo is less certain. Human SP or its fractions, administered 5 or 10 times at 3-day intervals into the mouse vagina simultaneously with washed human sperm or sheep red blood cells, suppressed the delayed-type hypersensitivity reactions to footpad injection of the antigens. However, SP failed to suppress antibody formation to these antigens in the applied experimental conditions. In an in vitro study, SP suppressed the proliferation of human peripheral blood mononuclear cells to phytohemagglutinin. This suppression of in vitro proliferation by SP was mediated by materials of both low (LMW) and high molecular weight (HMW). Among the HMW materials, a factor with approximately 1,500 kilodaltons, partially purified by DEAE-cellulose chromatography and gel filtration, exhibited the most potent suppressor activity in vitro and in vivo. The suppressive activities of SP or its fractions were not abolished by treatment with both heat (95 degrees C for 10 min) and trypsin (1.0 mg/ml, 37 degrees C for 4 h). These findings indicate that SP could contribute to the development of suppressed cellular immunity to sperm; immunosuppressive actions of SP are mediated by at least two distinct factors in vivo as well as in vitro.     

An optimized Protocol for Human M2 Macrophages using M‐CSF and IL‐4/IL‐10/TGF‐β Yields a Dominant Immunosuppressive Phenotype

Monocytes are highly abundant circulatory effector cells and play a vital role in driving or resolving inflammatory processes depending on their activation phenotype. We investigated and compared a panel of polarization protocols of blood‐derived monocytes to achieve a stable, optimal and effective regimen for in vitro induction of immunosuppressive human macrophages, evaluating their surface receptor expression, cytokine profile, scavenging function and ability to suppress T‐cell proliferation. Importantly, we assessed the effect of copolarization or secondary pro‐inflammatory stimulation of a primary anti‐inflammatory activation phenotype. A combination of IL‐4/IL‐10/TGF‐β yielded a relatively stable and dominant immunosuppressive phenotype characterized by higher IL‐10 production and down‐regulated TNF‐α, IL‐6, CD86, CD274 and MHC II expression. Functionally, IL‐4/IL‐10/TGF‐β‐stimulated macrophages (M2) had a potent deactivating effect on a subsequent pro‐inflammatory LPS/IFNγ‐activated macrophage (M1) stimulation and significantly suppressed T‐cell proliferation. Monocytes derived from patients with chronic inflammatory diseases could be induced to be anti‐inflammatory using this protocol. Pre‐differentiation with GM‐CSF or M‐CSF was further demonstrated to enhance final M1/M2 activation status. Our findings indicate a robust polarization protocol for generation of specific immunosuppressive human monocyte‐derived macrophages.     

Fentanyl Suppresses the Survival of CD4+ T Cells Isolated from Human Umbilical Cord Blood through Inhibition of IKKs‐mediated NF‐κB Activation

The aim of this study was to investigate the effects and the underlying mechanisms of fentanyl anaesthetic on T lymphocytes isolated from human umbilical cord blood in vitro. The percentages of CD4⁺, CD8⁺ and regulatory T (Treg) cells in human umbilical cord blood mononuclear cells (UBMC) treated with fentanyl in vitro were analysed by flow cytometry. The levels of cytokines IFN‐γ, IL‐2, IL‐4 and IL‐17 secreted by activated CD4⁺ T cells were measured by ELISA assays. Expressions of MAPK and NF‐κB signalling pathway proteins were determined by Western blotting. Effects of fentanyl on IKK and p65 expression promoter activities were analysed by luciferase assay. Fentanyl decreased the percentages and amounts of CD4⁺, CD8⁺ and Foxp3⁺Treg T lymphocyte subsets in UBMCs in a dose‐dependent manner. Fentanyl inhibited the proliferation and induced apoptosis of activated CD4⁺ T cells dose dependently. Fentanyl could not reverse the increase of cell proliferation in activated groups to be equivalent with those in inactivated group. Secretions of IFN‐γ, IL‐2 and IL‐4 cytokines were significantly decreased by moderate to high dose of fentanyl compared with controls. No significant differences were observed in protein expressions of MAPK pathway. In addition, fentanyl suppressed the IKKs‐mediated activation of NF‐κB. This study demonstrates that fentanyl exerts immunosuppressive effects on T lymphocytes obtained from UBMCs. Thus, the clinical application of fentanyl would not only relieve pain caused by surgery but regulate immune responses post‐operation possibly through inhibition of IKKs‐mediated NF‐κB activation.     

Mesenchymal Stem Cells Overexpressing Interleukin‐35 Propagate Immunosuppressive Effects in Mice

To explore generation of interleukin (IL)‐35‐expressing mouse adipocyte‐derived mesenchymal stem cells (Ad‐MSCs) using lentiviral vector and their potential immunosuppressive effects in mice. Ad‐MSCs were isolated and cultured in vitro and transfected with a lentivirus vector for overexpression of the therapeutic murine IL‐35 gene. IL‐35 expression in transfected MSCs (IL‐35‐MSCs) was quantified by enzyme‐linked immunosorbent assay (ELISA). The lymphocytes subsets after one‐way mixed lymphocyte culture and in vivo intravenous transplantation were analysed by flow cytometry to evaluate the immunosuppressive effects of IL‐35‐MSCs. ELISA was performed to examine IL‐10, IL‐17A and IL‐35 expression in lymphocyte culture. Mouse Ad‐MSCs were isolated and cultured. IL‐35 was expressed in the MSC supernatant and serum after IL‐35 transduction into Ad‐MSCs by lentiviral vector transfection in vitro and in vivo. The percentage of CD4⁺ CD25⁺ T regulatory (Treg) cells in mice treated with IL‐35‐MSCs significantly increased. IL‐35‐MSCs upregulated the CD4⁺ CD25⁺ Treg cells in the allogeneic mixed lymphocyte reaction system, and lowered the percentage of CD4⁺ T cells compared with the other two control groups (P < 0.01). IL‐17A expression significantly decreased and IL‐10 expression significantly increased in IL‐35‐MSCs and MSCs when compared by ELISA to the control groups (P < 0.01). IL‐35‐transduced Ad‐MSCs in vivo can enhance proliferation of CD4⁺ CD25⁺ Treg cells and suppress the function of effector T cells such as T helper (Th) 1, Th2 and Th17 cells and may reduce the development of allograft rejection. Our data suggest that transduced Ad‐MSCs overexpressing IL‐35 may provide a useful approach for basic research on cell‐based immunotolerance therapy for inducing transplantation tolerance.     

Asthma and urticaria induced by seminal plasma in a woman with IgE antibody and T-lymphocyte responsiveness to a seminal plasma antigen

A patient with asthma urticaria and angioedema induced by allergy to seminal plasma was examined at intervals for 10 years. Before treatment her anaphylactic susceptibility to seminal plasma was manifested by very strong prick-test responses, IgE antibody to an allergenic fraction of seminal plasma determined by RAST, and by antigen-induced histamine release from her blood leucocytes. The skin test and in vitro lymphocyte tests indicated concomitant delayed hypersensitivity to the same allergen. The patient's lymphocytes treated with seminal plasma allergen fraction showed much increased incorporation of thymidine, and also synthesis of a product (NIF) that inhibited migration of neutrophils from a normal donor. The allergen fraction of seminal plasma had about five components in the range of 20000–40000 daltons molecular weight; the major fraction binding IgE appeared to be a glycoprotein. The patient was successfully desensitized by injections of her husban?s seminal plasma. Desensitization was not associated with persistent amounts of antigen-specific IgG antibodies.     

C. elegans Rassf homolog,rasf‐1, is functionally associated with rab‐39 Rab GTPase in oxidative stress response

The Ras association domain family (Rassf) is one of the Ras effectors, which can bind to several GTP‐charged Ras‐like GTPases. The Rassf proteins are widely conserved beyond species from nematode to human. To explore the novel functions of Rassf proteins, we took advantage of nematode C. elegans as a model animal with only one Rassf homolog, T24F1.3 (rasf‐1). The rasf‐1‐mutant as well as rasf‐1‐knockdown animals were found to be more sensitive to oxidative stress of arsenite than in wild type, indicating that rasf‐1 is involved in oxidative stress response. We next screened for proteins that interact with RASF‐1 by the yeast two‐hybrid system and identified RAB‐39 Rab GTPase as an interacting partner of RASF‐1. We not only confirmed specific binding between these molecules but also demonstrated that RASF‐1 binds to GTP‐bound form but not GDP‐bound form of RAB‐39. Importantly, rab‐39 mutant animals were also sensitive to oxidative stress, which was dependent on rasf‐1 according to the epistasis analysis. Moreover, Rassf1 and Rab39, mammalian homologs of rasf‐1 and rab‐39, respectively, were shown to interact with each other in vitro. These results indicate that the RASF‐1 functionally interacts with RAB‐39 and that the interaction between their homologs is conserved in mammals.