The promotive effect of interleukin 4 with interleukin 2 in the proliferation of tumor-infiltrating lymphocytes from patients with malignant tumor

In adoptive immunotherapy, the number of effector cells is one of the major factors relating to the therapeutic efficacy. We demonstrated that tumor-infiltrating lymphocytes (TILs) were stimulated to proliferate by incubation with interleukin 2 (IL-2) plus interleukin 4 (IL-4). TILs cultured with IL-2 plus IL-4 increased 3.1-fold more than TILs cultured with IL-2 alone. However, IL-4 did not alter the cytotoxic activity of TILs against autologous tumor cells and established tumor cell lines. It is suggested that IL-2 receptor is related to the mechanism of the proliferation of activated TILs cultured by combination with IL-2 and IL-4. Thus, the combination of IL-2 and IL-4 may increase the efficacy of adoptive immunotherapy using activated TILs.     

Comparative Study between Gas Chromatography and Ion-exchange Chromatography on Amino Acid Analysis of Foods

Lactobacillus brevis and Saccharomyces cerevisiae were completely sterilized by the supercritical (SC) CO2 micro-bubble method. Gaseous (G) and liquid (LQ) CO2 were used in a similar manner to compare the sterilizing effect. Among the three treatments, the microorganisms were only effectively sterilized by the SC CO2 treatment at 25 MPa and 35°C.     

Role of calcium-binding sites in calcium-dependent membrane association of annexin A4

Annexin A4 (Anx4) is a cytosolic calcium-binding protein with four repeat domains, each containing one calcium-binding site (CBS). The protein interacts with the phospholipid membrane through the CBS-coordinated calcium ion, although the role of each CBS in the calcium-dependent association is unclear. To determine the role of each CBS, 15 CBS-abolished variants were produced in various combinations by substitution of a calcium-liganding residue on each CBS by Ala. Various mutant combinations produced different influences on calcium-dependent membrane-binding behavior and on the sodium-dependent dissociation of membrane-bound Anx4. Our data suggest the interaction of Anx4 with the lipid membrane consists of strong and weak interactions. CBSs I and IV mediate formation of strong interactions, while CBSs II and III are important for weak interactions. We also suggest Anx4 binds the lipid membrane through CBSs I and IV in the cytoplasmic fluids.     

Effect of a conjugated acridine moiety on the binding and reactivity of Cu(II)[9-acridinylmethyl-1,4,7-triazacyclononane] with DNA

The DNA binding orientation and dynamic behavior of Cu(II) complexes of 1,4,7-triazacyclononane ([9]aneN(3)), 1, and an acridine conjugate, 2, were investigated by DNA fiber EPR (EPR=electron paramagnetic resonance) spectroscopy. Crystal and molecular structure of 2 were determined by X-ray diffraction. It has been shown that 1 binds to DNA in two different modes at room temperature; one species is rapidly rotating and the other is immobilized randomly on the DNA. The introduction of acridine to [9]aneN(3) fixed the [Cu([9]aneN(3))](2+) moiety of 2 in two different environments on the DNA: the g(mid R:mid R:) axis of one species (g( parallel)=2.26) is aligned perpendicularly to the DNA fiber axis whereas that of the other (g( parallel)=2.24) aligns<90 degrees with the DNA fiber axis. The different DNA binding structures of 1 and 2 are reflected also in their different efficiencies of DNA cleavage; 2 was found to be more effective both in oxidative and hydrolytic cleavage reactions.     

Periplasmic secretion of native ovalbumin without signal cleavage in Escherichia coli

In Escherichia coli cells carrying wild-type ovalbumin cDNA, some of the recombinant protein was secreted into the periplasmic space. In contrast, a signal-region mutant form of ovalbumin (deletion, Gly1 to Ala39) was not detected in the periplasm despite being synthesized at the same level as the wild-type protein. Chemical and spectroscopic analyses showed that periplasmic ovalbumin assumes a conformation indistinguishable from that of native egg white ovalbumin. We concluded that a process resembling the secretion of ovalbumin process in the oviduct occurs also in bacteria.     

Physiologically high concentrations of 17beta-estradiol enhance NF-kappaB activity in human T cells

Estrogen has diverse effects on inflammation and immune responses. That pregnancy is associated with remission of some autoimmune diseases and exacerbation of others suggests that physiological fluctuation in estrogen levels could affect the immune responses in humans. However, the molecular basis for these phenomena is poorly understood. We hypothesized that fluctuations of estrogen levels modulate intracellular signaling for immune responses via estrogen receptors (ERs). In reporter assays, 17beta-estradiol (E2) at a physiologically high concentration increased the activity of NF-kappaB in Jurkat cells stimulated by PMA/ionomycin or TNF-alpha. Overexpression and RNA interference experiments suggested that the effects were mediated through ERbeta. Immunoprecipitation assay showed that both ERalpha and ERbeta are directly associated with NF-kappaB in the cell nucleus. Using chromatin immunoprecipitation assay, we confirmed that ERalpha and ERbeta associated with NF-kappaB and steroid hormone coactivators at the promoter region of NF-kappaB regulated gene. Considering that NF-kappaB regulates the expression of various genes essential for cell growth and death, estrogen could regulate the fate of T cells by affecting the activity of NF-kappaB. To determine whether E2 alters the fate of T cells, we investigated E2 actions on T cell apoptosis, a well-known NF-kappaB-mediated phenomenon. E2 increased apoptosis of Jurkat cells and decreased that of human peripheral blood T cells. Our results indicate that E2 at a physiologically high concentration modulates NF-kappaB signaling in human T cells via ERbeta and affects T cell survival, suggesting that these actions may underlie the gender differences in autoimmune diseases.     

Ploidy chimeras induced in haploid sporophytes of <Emphasis Type="Italic">Osmunda claytoniana</Emphasis> and <Emphasis Type="Italic">Osmunda japonica</Emphasis>

Haploid sporophytes of Osmunda claytoniana (2n = x = 22) were apogamously produced from calli when cultivated on a hormone-free medium. Flow cytometric analysis showed that ploidy chimeras were spontaneously produced in a haploid sporophyte of O. claytoniana and those of O. japonica that were obtained in the previous study. In the haploid sporophyte of O. claytoniana, a diploid pinnule and a partially diploid terminal segment were produced in a haploid pinna. In O. japonica, a haploid sporophyte yielded a diploid pinna in a haploid frond, and another haploid sporophyte yielded a diploid pinnule in a haploid pinna. Diploid chimeras were large in size and could be readily distinguished from other haploid parts of the fronds. It is likely that the chimeras were produced clonally from a single diploid cell that established chromosome doubling.