1,25-Dihydroxyvitamin D3 enhances NK susceptibility of human melanoma cells via Hsp60-mediated FAS expression.

The active metabolite of vitamin D(3), 1α,25(OH)(2)D(3) , displays anticancer effects by regulating cell cycle and apoptosis in many cancer cells. However, it has not been determined whether 1α,25(OH)(2)D(3) increases the susceptibility of cancer cells to NK cells. Here, we investigated the anticancer effect of 1α,25(OH)(2)D(3) in human melanoma cell lines by investigating enhancement of NK susceptibility and elucidating the mediator of NK cytotoxicity. 1α,25(OH)(2)D(3)-resistant melanoma cells (G-361 and SK-MEL-5) treated with 1α,25(OH)(2)D(3) showed higher susceptibility to NK cells with up-regulation of Fas expression. Furthermore, G-361 cells treated with 1α,25(OH)(2)D(3) showed significantly increased caspase activity. In addition to Fas up-regulation, expression of heat shock protein 60 (Hsp60) was elevated by 1α,25(OH)(2) D(3) . Increased expression of Hsp60 by 1α,25(OH)(2)D(3) was related to not only up-regulation of Fas expression but also to NK susceptibility of G-361 cells. Taken together, our data suggest that 1α,25(OH)(2)D(3) acts as an anticancer agent by increasing expression of Fas on the surface of melanoma cells through Hsp60 induction and strengthens caspase sensitivity to Fas-mediated apoptotic pathway by NK cells. 1α,25(OH)(2)D(3) treatment may therefore have a preventive role in melanoma occurrence or potentiate the anticancer effects of NK-cell immune therapy.

Anaesthetic requirements and stress hormone responses in acute cord-injured patients undergoing surgery of the injured spine.

BACKGROUND AND OBJECTIVE: Neuraxial anaesthesia has been shown to produce a sedative and anaesthetic-sparing effect. The purpose of the present study was to determine the effects of acute spinal cord injury on sevoflurane requirement and stress hormone responses during spinal surgery at the level of the injury. METHODS: Thirty-five patients with traumatic complete spinal cord injury undergoing spinal surgery at the level of the injury were studied. They were grouped into quadriplegics (above C7, n = 20) and paraplegics (below T1, n = 15) according to the level of injury. Patients (n = 35) with spine trauma without neurological impairment undergoing spinal surgery at the respective level served as controls. The bispectral index score was maintained at 40-50 throughout the surgery. Measurements included end-tidal sevoflurane concentrations, mean arterial pressure, heart rate, and plasma concentrations of catecholamines and arginine vasopressin. RESULTS: During the surgery, the mean arterial pressure was significantly lower in both quadriplegics and paraplegics (P < 0.05). The heart rate did not differ significantly in the quadriplegics, but was higher in the paraplegics, compared with their controls. However, end-tidal sevoflurane concentrations and bispectral index score were comparable with controls in both quadriplegics and paraplegics. Throughout the study, the plasma arginine vasopressin concentrations were not altered, although norepinephrine and epinephrine concentrations were lower in the quadriplegics. There were no significant differences in stress hormones between the groups having thoraco-lumbar surgery. CONCLUSION: Spinal cord injury neither alters the anaesthetic requirement regardless of the level of injury during spinal surgery at the level of the injury, nor enhances arginine vasopressin release. However, it blunts catecholamine responses in quadriplegics.

Water-enhanced low-temperature CO oxidation and isotope effects on atomic oxygen-covered Au(111).

Water-oxygen interactions and CO oxidation by water on the oxygen-precovered Au(111) surface were studied by using molecular beam scattering techniques, temperature-programmed desorption (TPD), and density functional theory (DFT) calculations. Water thermally desorbs from the clean Au(111) surface with a peak temperature of approximately 155 K; however, on a surface with preadsorbed atomic oxygen, a second water desorption peak appears at approximately 175 K. DFT calculations suggest that hydroxyl formation and recombination are responsible for this higher temperature desorption feature. TPD spectra support this interpretation by showing oxygen scrambling between water and adsorbed oxygen adatoms upon heating the surface. In further support of these experimental findings, DFT calculations indicate rapid diffusion of surface hydroxyl groups at temperatures as low as 75 K. Regarding the oxidation of carbon monoxide, if a C (16)O beam impinges on a Au(111) surface covered with both atomic oxygen ( (16)O) and isotopically labeled water (H 2 (18)O), both C (16)O (16)O and C (16)O (18)O are produced, even at surface temperatures as low as 77 K. Similar experiments performed by impinging a C (16)O beam on a Au(111) surface covered with isotopic oxygen ( (18)O) and deuterated water (D 2 (16)O) also produce both C (16)O (16)O and C (16)O (18)O but less than that produced by using (16)O and H 2 (18)O. These results unambiguously show the direct involvement and promoting role of water in CO oxidation on oxygen-covered Au(111) at low temperatures. On the basis of our experimental results and DFT calculations, we propose that water dissociates to form hydroxyls (OH and OD), and these hydroxyls react with CO to produce CO 2. Differences in water-oxygen interactions and oxygen scrambling were observed between (18)O/H 2 (16)O and (18)O/D 2 (16)O, the latter producing less scrambling. Similar differences were also observed in water reactivity toward CO oxidation, in which less CO 2 was produced with (16)O/D 2 (16)O than with (16)O/H 2 (16)O. These differences are likely due to primary kinetic isotope effects due to the differences in O-H and O-D bond energies.

Familial association and frequency of learning disabilities in ADHD sibling pair families.

In a sample of 235 families with at least two children with Attention-Deficit/ Hyperactivity Disorder (ADHD), the frequency and familial association of learning disabilities (LD) were assessed. Familiality was examined both between sibling pairs and between parents and their children. Two methods for defining LD, a discrepancy-based and a low-achievement model, were employed to examine the occurrence of LD in this sample. The specific types of LD examined included Reading Disability (RD), Math Disability (MD), and Writing Disability (WD). The prevalence rates were highest for RD, followed by WD then MD. The two definitions of LD yielded similar prevalence rates but identified different groups of children with vastly different IQ scores. Strong familial association was demonstrated for RD both between sibling pairs and between parents and children, with weaker association for WD. There was evidence of nonrandom mating for LD among parents, but not for ADHD or for ADHD + LD. Despite the high comorbidity of ADHD and LD among parents, the presence of ADHD in the parents did not predict child LD supporting independent familial factors underlying ADHD and LD.

New Insights into Vitamin D and Autophagy in Inflammatory Bowel Diseases.

Vitamin D exerts an anti-inflammatory effect in both health and disease. The importance of vitamin D in protection against various inflammatory and metabolic diseases, including inflammatory bowel diseases (IBDs), has been discussed. Considerable data indicate a role for vitamin D in the activation of autophagy, an intracellular renewal system that maintains homeostasis by eliminating large protein aggregates and damaged organelles. Recent studies have demonstrated an intricate interplay between autophagy and the regulation of inflammation, suggesting that autophagy-modulating agents could be used to treat IBDs. This review focuses on the role and mechanisms of vitamin D in autophagy and the regulation of intestinal inflammation. Vitamin D shows promise for the prevention and amelioration of pathologic responses in IBD, an effect that is mediated, at least in part, by the induction and modulation of autophagy.

Reactivity of molecularly chemisorbed oxygen on a Au/TiO2 model catalyst.

We present results of an investigation into the reactivity of molecularly chemisorbed oxygen with CO on a Au/TiO2 model catalyst at 77 K. We previously discovered that exposing the model catalyst sample to a radio-frequency-generated plasma jet of oxygen results in co-population of both atomically and molecularly chemisorbed oxygen species on the sample. We tested the reactivity of the molecularly chemisorbed oxygen by comparing the CO2 produced from a sample populated with both species to the CO2 produced from a sample that has been cleared of molecularly chemisorbed oxygen employing collision-induced desorption. Samples that are populated with both species consistently result in greater CO2 produced than samples with only atomic oxygen. We interpret this result to indicate that molecularly chemisorbed oxygen on the sample can directly participate in the CO oxidation reaction. The reactivity of molecularly chemisorbed oxygen has been investigated for five different gold coverages (0.5, 0.75, 1, 1.25, and 2 ML), and we observe that there is a greater fractional difference in the CO2 produced (difference between sample populated with both molecularly and atomically adsorbed oxygen and sample populated solely with atomically adsorbed oxygen) for the 1 ML Au coverage than for the other coverages for equivalent oxygen plasma-jet exposures. However, it is not possible to unambiguously conclude that this observation is directly related to a particle size effect on the chemistry since the absolute O(2,a) and O(a) content on the various surfaces is different for all the coverages studied because of the plasma-jet technique that we employed for populating the surfaces with oxygen. Unfortunately, this precludes a direct comparison of the reactivity of molecular oxygen in the carbon monoxide oxidation reaction as a function of gold coverage and hence particle size.

Ballistocardiogram artifact removal from EEG signals using adaptive filtering of EOG signals.

We estimated ballistocardiogram (BCG) components in EEG signals recorded inside an MRI magnet using the electro-oculogram (EOG) signals recorded simultaneously with the EEG signals. Since the EOG signals are measured near the EEG measuring points, it is thought that the BCG components in the EOG signals resemble the BCG components in the EEG signals. To estimate the BCG components in the EEG signals, we applied the Kalman filter to the EOG and EEG signals recorded inside a 3.0 T MRI magnet. After removing the estimated BCG components from the EEG signals, we extracted the visual-evoked potentials (VEPs) from the BCG-removed EEG signals. To validate the efficacy of Kalman filtering in the BCG artifact removal, we have compared three types of VEPs of eight healthy subjects: one extracted from the raw EEG signals measured outside the magnet and the others extracted from the BCG-removed EEG signals measured inside the magnet. The BCG artifacts have been removed with Kalman filtering as well as with the conventional BCG template subtraction method for the sake of comparison. No significant difference in waveforms, latencies and amplitudes has been found between the two types of VEPs extracted from the two kinds of BCG-removed EEG signals.

Immunity to squamous carcinoma in mice immunized with dendritic cells transfected with genomic DNA from squamous carcinoma cells.

Immunotherapy of squamous cell carcinoma (SCC) at an early stage of the disease increases the likelihood of success. We report a new vaccination strategy designed to prepare SCC vaccines from microgram amounts of tumor tissue, enabling the treatment of patients with minimal residual disease. The vaccine was prepared by transfer of sheared genomic DNA-fragments (25 kb) from KLN205 cells, an SCC cell line of DBA/2 mouse origin, into syngeneic bone marrow-derived mature dendritic cells (DCs). More than 90% of the transfected DCs took up DNA from the neoplasm and transferred genes were expressed as protein. The DCs expressed CD11c, CD11b, and the costimulatory molecules CD40, CD80 and CD86, characteristic of mature DCs. Syngeneic DBA/2J mice, highly susceptible to the growth of KLN205 cells, were injected intravenously (i.v.) with the transfected DCs, followed by a subcutaneous (s.c.) injection of the tumor cells. The strong immunogenic properties of the transfected cells were indicated by the finding that the survival of the tumor-bearing mice was prolonged (P<.001), relative to that of mice in various control groups. Enzyme-linked immuno spot (ELISPOT IFN-gamma) assays revealed the activation of cell-mediated immunity directed toward the SCC in mice immunized with the transfected DCs. Two independent in vitro cytotoxicity assays indicated the presence of robust cell-mediated immunity directed toward the SCC in mice immunized with the transfected cells.

Formation of molecularly chemisorbed oxygen on TiO2-supported gold nanoclusters and Au(111) from exposure to an oxygen plasma jet.

We present results of an investigation into the low-temperature formation of molecularly chemisorbed oxygen on a Au/TiO(2) model catalyst and on a Au(111) single crystal during exposure to a plasma jet of oxygen. Through the use of collision-induced desorption measurements and isotopic mixing experiments we show evidence suggesting that at least some of the molecular oxygen is formed as a result of recombination of oxygen atoms on the samples during the plasma exposure. Of course, adsorption of excited molecular oxygen directly from the gas phase may also take place. We also present evidence showing that the adsorption of oxygen atoms on the surface assists in the molecular chemisorption of oxygen on the Au/TiO(2) model catalyst samples. Thus, oxygen molecules impinging on the samples during plasma-jet exposures (plasma jet has approximately 40% dissociation fraction) could have an enhanced probability of adsorption due to simultaneous oxygen atom adsorption.

Induction of human leukemia HL-60 cell differentiation via a PKC/ERK pathway by helenalin, a pseudoguainolide sesquiterpene lactone.

Helenalin, a cell-permeable pseudoguainolide sesquiterpene lactone, is a potent anti-inflammatory agent that inhibits nuclear factor-kappa B (NF-kappa B) DNA binding activity. In this report, we investigated the effect of helenalin on cellular differentiation in the human promyelocytic leukemia HL-60 cell culture system. Helenalin by itself markedly induced HL-60 cell differentiation in a concentration-dependent manner. Cytofluorometric analysis and cell morphologic studies indicated that helenalin induced cell differentiation predominantly into granulocytes. Protein kinase C (PKC) and extracellular signal-regulated kinase (ERK) inhibitors significantly inhibited HL-60 cell differentiation induced by helenalin, while p38 mitogen-activated protein kinase (MAPK) inhibitors did not. Moreover, helenalin enhanced PKC activity and protein level of PKC beta I and PKC beta II isoforms, and also increased the level of pERK in a concentration-dependent manner. In addition, the enhanced levels of cell differentiation closely correlated with the decreased levels of NF-kappa B binding activity by helenalin. These results indicate that PKC, ERK, and NF-kappa B may be involved in HL-60 cell differentiation induced by helenalin.

Potentiation of 1,25-dihydroxyvitamin D(3)-induced differentiation of human promyelocytic leukemia cells into monocytes by costunolide, a germacranolide sesquiterpene lactone.

Costunolide, a germacranolide sesquiterpene lactone that exists in several medicinal plants, is known to be a possible anti-cancer and chemopreventive agent for tumorigenesis. In this report, we investigated the effect of costunolide on cellular differentiation in the human promyelocytic leukemia HL-60 cell culture system. Costunolide markedly increased the degree of HL-60 leukemia cell differentiation when simultaneously combined with 5nM 1,25-dihydroxyvitamin D(3) (1,25-(OH)(2)D(3)). Costunolide by itself had very weak effects on the differentiation of HL-60 cells. Cytofluorometric analysis and cell morphologic studies indicated that costunolide potentiated 1,25-(OH)(2)D(3)-induced cell differentiation predominantly into monocytes. Inhibitors for PKC, PI3-K, and ERK markedly inhibited HL-60 cell differentiation induced by costunolide in combination with 1,25-(OH)(2)D(3). In addition, pretreatment of HL-60 cells with costunolide before the 1,25-(OH)(2)D(3) addition also potentiated cell differentiation in a concentration- and time-dependent manner, and the enhanced levels of cell differentiation closely correlated with the inhibitory levels of NF-kappaB-binding activity by costunolide. These results indicate that PKC, PI3-K, ERK and NF-kappaB may be involved in 1,25-(OH)(2)D(3)-mediated cell differentiation enhanced by costunolide.

Prevalence of radiologic superior canal dehiscence in normal ears and ears with chronic otitis media.

OBJECTIVES/HYPOTHESIS: Although labyrinth fistulae are caused mostly by cholesteatoma, they can occur in long-standing chronic otitis media (COM) without cholesteatoma. We aimed to compare the prevalence of radiologic SCD on computed tomography (CT) between normal ears and contralateral COM ears in patients with unilateral COM and to assess the prevalence of superior canal dehiscence (SCD) according to the age. STUDY DESIGN: Case series with comparison performed at a tertiary care academic referral center. METHODS: We retrospectively reviewed consecutive temporal bone CT scans of 759 patients with unilateral COM between 2009 and 2011. The mean (± standard deviation) age was 48 years (±14 years). Images were independently evaluated by two otologists, and the bone overlying the superior canal was characterized as normal, suspicious, or definite SCD. RESULTS: The prevalence (3.4%) of definite SCD in COM ears was significantly higher than that (0.3%) in normal ears. The prevalence (6.6%) of suspicious or definite SCD in COM ears was also higher than that (1.2%) in normal ears. There was no correlation between the prevalence of SCD and age in either normal or COM ears. All of the normal ears with suspicious or definite SCD also showed contralateral suspicious or definite SCD (bilateral involvement). CONCLUSIONS: Our present findings suggest that the COM is related to the presence of SCD. The roof of the temporal bone may become thin by the failure of postnatal bone development and susceptible to chronic brain pulsation and pressure exerted by the temporal lobe in COM ears.

Gene transfer of AIMP1 and B7.1 into epitope-loaded, fibroblasts induces tumor-specific CTL immunity, and prolongs the survival period of tumor-bearing mice.

T helper type 1 (Th1) cell-mediated immune responses play various roles in cellular immunity, including inducing cytotoxic T lymphocytes (CTLs) and they have been shown to be crucial in cancer immunotherapy. Previously, we found that aminoacyl-tRNA synthetase-interacting multifunctional protein 1 (AIMP1) stimulated antigen-presenting cells to secrete IL-12, leading to enhanced Th1 cell responses. In this study, as a way of enhancing antigen-specific Th1 responses, mouse fibroblasts (H-2(b)) were genetically modified to express an AIMP1 and a costimulatory B7.1 (Fb/AIMP1/B7.1). Fb/AIMP1/B7.1 cells were then loaded with an ovalbumin epitope as a model antigen (Fb/AIMP1/B7.1/OVA), and tested to determine if they induced OVA-specific CTLs in C57BL/6 mice (H-2(b)). Immunization with Fb/AIMP1/B7.1/OVA cells induced strong cytotoxic activities against OVA-expressing EG7 tumor cells, but not against other H-2(b) tumor cells. The levels of the cytotoxic response in the immunized mice with Fb/AIMP1/B7.1/OVA cells were significantly higher than the responses in mice immunized with other cell constructs. CD8(+) T cells were a major cell-type of OVA-specific antitumor immunity induced by Fb/AIMP1/B7.1/OVA cells. Furthermore, treatment with Fb/AIMP1/B7.1/OVA cells significantly prolonged the survival period of EG7 tumor-bearing mice. These results indicate that AIMP1-secreting, epitope-loaded fibroblasts efficiently induce antigen-specific CTL responses in mice.

Anti-CD3 single-chain Fv/interleukin-18 fusion DNA induces anti-mycobacterial resistance via efficient interferon-gamma production in BALB/c mice infected with Mycobacterium avium.

T helper type 1 (Th1) cell-mediated immune response performs a critical role in the induction of host defenses against a variety of intracellular pathogens. We recently demonstrated that the linkage between interleukin (IL)-18 and anti-CD3sFv genes confined the effect of IL-18 to the CD3+ T cells, resulting in the efficient induction of Thl immune responses. In this study, we attempted to determine whether the anti-CD3 single-chain Fv/IL-18 fusion DNA (pAnti-CD3sFv/IL-18) could stimulate resistance to Mycobacterium avium infection in genetically susceptible BALB/c mice. Immunization with pAnti-CD3sFv/IL-18 DNA during intranasal infection with M. avium resulted in a significant reduction in the bacterial load in the lung throughout the entire 10-week observation period. Immunization with pAnti-CD3sFv/IL-18 DNA induced and maintained significantly higher levels of cytotoxic activity and nitric oxide production by the lung cells, as compared with the injection of pAnti-CD3sFv DNA, or of a simple mixture of pAnti-CD3sFv DNA and pIL-18 DNA. Furthermore, the lung cells of the mice injected with pAnti-CD3sFv/IL-18 DNA exhibited persistent production of IFN-gamma. Treatment with anti-IFN-gamma antibody resulted in an abrogation of anti-mycobacterial effects in mice injected with pAnti-CD3sFv/IL-18 DNA. These results indicate that vaccination with the pAnti-CD3sFv/IL-18 fusion DNA induces significant resistance to M. avium infection, via the production of IFN-gamma in CD4+ T cells.

Water activated by atomic oxygen on Au(111) to oxidize CO at low temperatures.

The Au(111) surface was populated with atomic oxygen [16O] followed by oxygen-labeled water [H218O] at surface temperatures as low as 77 K. When a CO beam was impinged on this surface, both [C16O16O] and [C16O18O] were produced. The results strongly suggest the direct involvement and promoting role of water in CO oxidation on oxygen covered Au(111) at low temperatures.

Prolongation of the survival of breast cancer-bearing mice immunized with GM-CSF-secreting syngeneic/allogeneic fibroblasts transfected with a cDNA expression library from breast cancer cells.

Breast cancer cells, like other types of neoplastic cells, form weakly immunogenic tumor-associated antigens. The antigenic properties of the tumor-associated antigens can be enhanced if they are expressed by highly immunogenic cells. In this study, a cancer vaccine was prepared by transfer of a cDNA expression library from SB5b breast carcinoma into mouse fibroblast cells of C3H/He mouse origin (H-2(k)), that had been previously modified to secrete GM-CSF and to express allogeneic class I-determinants (H-2(b)). The transfected syngeneic/allogeneic fibroblasts secreting GM-CSF were used as a vaccine in C3H/He mice. Robust cell-mediated immunity toward the breast cancer cells was generated in mice immunized with the cDNA-based vaccine. The immunity, mediated predominantly by CD8(+) T lymphocytes, was directed toward the breast cancer cells, but not against either of two other non-cross-reactive neoplasms of C3H/He mice. The immunity was sufficient to prolong the survival of mice with established breast cancer. Among other advantages, preparation of the vaccine by cDNA-transfer into a fibroblast cell line enabled the recipient cells to be modified in advance of DNA-transfer to augment their immunogenic properties. As the transferred DNA is replicated as the transfected cells divide, the vaccine could be prepared from microgram quantities of tumor tissue.

Selective catalytic oxidation of ammonia to nitrogen on atomic oxygen precovered Au(111).

We demonstrate ammonia oxidation promoted by an atomic oxygen precovered Au(111) surface. The selectivity of the catalytic oxidation of ammonia to NO or N2 on Au(111) is tunable by the atomic oxygen coverage. We propose that N2 and NO are produced via the recombination reactions of Nad + Nad and Nad + Oad.

Formononetin, a phyto-oestrogen, and its metabolites up-regulate interleukin-4 production in activated T cells via increased AP-1 DNA binding activity.

Phyto-oestrogens are polyphenolic non-steroidal plant compounds with oestrogen-like biological activity. Phyto-oestrogens have many biological effects including oestrogen agonist/antagonist properties. However, the effect of phyto-oestrogens on allergic responses remains unclear. In this study we investigated whether formononetin, a phyto-oestrogen, and its metabolites, daidzein and equol, affect production of interleukin-4 (IL-4), a pro-inflammatory cytokine closely associated with allergic immune response, in primary CD4+ T cells and EL4 T lymphoma cells. Formononetin, daidzein and equol significantly enhanced IL-4 production from both CD4+ T cells and EL4 cells in a dose-dependent manner. Formononetin, daidzein and equol also enhanced IL-4 gene promoter activity in EL4 cells transiently transfected with IL-4 gene promoter constructs, but this effect was impaired in EL4 cells transfected with an IL-4 promoter construct deleted of P4 site carrying nuclear factor of activated T cells (NF-AT) and activator protein-1 (AP-1) binding sites. In addition, formononetin, daidzein and equol increased AP-1 DNA binding activities while did not affect NF-AT DNA binding activities. The enhancing effects on IL-4 production and AP-1 DNA binding activities were abrogated by specific inhibitors for phosphatidylinositol-3-kinase (PI3K), protein kinase C (PKC) and p38 mitogen-activated protein kinase (MAPK), indicating that formononetin, daidzein and equol might enhance IL-4 production by increased activation of AP-1 through the PI3-K/PKC/p38 MAPK signalling pathway. These results suggest that phyto-oestrogens and some of their metabolites may increase allergic responses via the enhancement of IL-4 production in T cells.

Exposure to 4-tert-octylphenol, an environmentally persistent alkylphenol, enhances interleukin-4 production in T cells via NF-AT activation.

4-tert-Octylphenol (OP) is a representative endocrine disruptor that may have adverse effects on human health. The influence of this compound on allergic immune responses remains unclear. In this study, we have examined the effects of OP on production of interleukin-4 (IL-4), a pro-inflammatory cytokine closely associated with allergic immune responses. OP significantly enhanced IL-4 production in antigen-primed T cells in a dose-dependent manner. Treatment with OP in vivo resulted in significant increase of IL-4 production in T cells and of IgE levels in sera of antigen-primed mice. Furthermore, OP enhanced the activation of IL-4 gene promoter in EL4 T cells transiently transfected with IL-4 promoter/reporter constructs, and the enhancing effect mapped to a region in the IL-4 promoter containing binding sites for nuclear factor of activated T cell (NF-AT). Activation of T cells by phorbol-12-myristate-13-acetate (PMA) resulted in markedly enhanced binding activities to the NF-AT site, which significantly increased upon addition of OP, indicating that the transcription factor NF-AT was involved in the enhancing effect of OP on IL-4 production. The enhancement of IL-4 production by OP was blocked by FK506, a calcineurin inhibitor, but not by the estrogen receptor (ER) antagonist ICI 182780. FK506 inhibited the NF-AT-DNA binding activity and IL-4 gene promoter activity enhanced by OP in a dose-dependent manner. These findings demonstrate that OP enhances IL-4 production in T cells via the stimulation of calcineurin-dependent NF-AT activation.

Generation of tumor-specific cytotoxic T lymphocyte and prolongation of the survival of tumor-bearing mice using interleukin-18-secreting fibroblasts loaded with an epitope peptide.

There is currently much interest in generating cytotoxic T lymphocyte (CTL) responses against tumor antigens as a therapy for cancer. In this study mouse fibroblasts (H-2(b)) were genetically modified to express a costimulatory B7.1 and a mature interleukin (IL)-18, and then loaded with an ovalbumin (OVA) epitope (SIINFEKL, H-2K(b) restricted) as a model antigen, and tested for the induction of OVA-specific CTLs in C57BL/6 mice (H-2(b)). The genetically modified fibroblasts lacking either IL-18 or B7.1 were also constructed. Immunization with the IL-18/B7.1-transfected fibroblasts induced strong cytotoxic activities against OVA-expressing EL4 (EG7) tumor cells, but not against other H-2(b) tumor cells such as EL4, C1498, and B16F1 cells. The magnitude of the cytotoxic response in mice with the IL-18/B7.1-transfected fibroblasts was significantly higher than the response in mice immunized with any other cell constructs. CD8(+) T cells with OVA-specific cytotoxic activities were predominant in mice immunized with the IL-18/B7.1-transfected fibroblasts. Furthermore, treatment with the IL-18/B7.1-transfected fibroblasts significantly prolonged the survival period of EG7 tumor-bearing mice. Anti-tumor CTL immunity by the IL-18/B7.1-transfected fibroblasts could be induced without the help of host antigen-presenting cells (APCs) and NK1.1(+) cells, whereas partially decreased by the depletion of CD4(+) T cells at the inductive stage. These results support the ability of IL-18/B7.1 gene transfer to enhance the antigen-presenting capacity of fibroblasts for inducing antigen-specific CTL response.