Interleukin-5 induces tumor suppression by peritoneal exudate cells in mice

The antitumor activity of peritoneal exudate cells (PEC) induced by murine interleukin-5 (mIL-5) was examined using Meth-A sarcoma cells transplanted into the peritoneal cavity of mice. Although in vitro treatment of Meth-A sarcoma cells with mIL-5 did not result in inhibition of their growth, treatment of mice intraperitoneally with mIL-5 (1 g/day) from day –5 to +5 (tumor cells were inoculated on day 0) led to a significant increase in survival or even rejection of tumor cells. This antitumor effect depended on the dose of mIL-5. Interestingly, there was identical therapeutic activity when the protocol of days –10 to –1 was used as opposed to –5 to +5. In addition, post-treatment with mIL-5 from day +1 to +10 was ineffective. This suggests that the therapeutic activity of IL-5 is largely prophylactic. Under the former condition, the number of PEC was found to increase over 50-fold when compared to levels in control mice. Moreover, the antitumor effect of mIL-5 was completely abolished by subcutaneous injection of anti-mIL-5 monoclonal antibodies. The treatment of mice injected intraperitoneally with human IL-2 also resulted in an increase in survival. Winn assay experiments using PEC recovered from mIL-5-treated mice (1g/day, from day –10 to –1) revealed that these PEC could mediate antitumor activity against Meth-A sarcoma cells. Furthermore, when the cured mice were re-injected with Meth-A sarcoma cells or syngeneic MOPC 104E cells, they could reject Meth-A sarcoma cells but not MOPC 104E cells, indicating that immune memory had been generated. These results suggest that IL-5 augumented the PEC tumoricidal activity but we have no indication that the tumoricidal activity was mediated through a mIL-5-dependent mechanism.     

Variations in and Inheritance of NS-Glycoprotein in Self-Incompatible Brassica campestris L.

The stigma of Brassica species contain NS-glycoproteins thatexhibit a high degree of structural homology to the S-glycoproteinsof self-incompatibility. Inheritance of and variations in theNS-glycoprotein were studied with reference to self-incompatibility.The detection of NS-glycoproteins was performed by cross-reactionwith an antiserum raised against a purified NS-glycoprotein.In B. campestris, four isoforms of the NS-glycoprotein weredifferentiated by their pI values, but their molecular weightswere identical to one another. The genes for these isoformsof NS-glycoprotein were controlled by alleles at a single locus,tentatively named the NS allele, which was independent of Salleles at both the protein and the DNA level. Segregation ofF₂ plants with respect to the self-incompatibility behaviorof pollen tubes can be explained by the S allele model, butit appears not to be affected by the NS alleles. NS-glycoproteinswere found in all 21 species of Brassica and its allies examinedto date. The pI values of these glycoproteins varied among differentspecies. In addition to the isoforms of the NS alleles, maturestigmas contained other groups of proteins that reacted weaklywith the antiserum against the NS-glycoprotein. (Received July 30, 1991; Accepted February 21, 1992)     

PLD activation in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA

Pertussis toxin-insensitive GTP-binding protein was observed to be involved in prostaglandin F2α(PGF2α)-induced phosphoinositide metabolism in Chinese hamster ovary (CHO) cells transfected with PGF2α receptor cDNA (CHO-PGF2α·R cells) (Ito, S. et al. Biochem. Biophys. Res. Commun. 200: 756, 1994). In the present study, we investigated PGF2α-induced PLD activation in CHO-PGF2α·R cells. PLD activation was examined by measuring the production of [³H]phosphatidylbutanol ([³H]PBut), a specific product of the PLD-catalyzed transphosphatidylation reaction. PGF2α-induced [³H]PBut formation was concentration-dependent with the maximal level obtained at 1 μM PGF2α. The maximal [³H]PBut formation was observed at 2 min after addition of PGF2α. Depletion of extracellular Ca²⁺ with EGTA suppressed PGF2α-induced PLD activation by 50%. PKC inhibitors Ro31–8425 and calphostin C inhibited PGF2α-induced [³H]PBut formation by 50%. PTK inhibitors genistein and herbimycin A failed to inhibit PGF2α-induced PLD activation. A combination of maximal effective concentrations of PGF2α (1 μM) and PMA (100 nM) enhanced PLD activation in an additive manner. Pretreatment of the cells with PMA for 2 h down-regulated PKCα and decreased PGF2α-induced PLD activation. These results suggest that PLD activation by PGF2α is mediated by both PKC-dependent and -independent pathways and that PKCα is involved in the former pathway.     

In vitro neurotoxicity of amyloid β-peptide cross-linked by transglutaminase

Transglutaminase catalyzes the intermolecular cross-linking of peptides between Gln and Lys residues, forming an -(-glutamyl) lysine bond. Amyloid -peptide, a major constituent of the deposits in Alzheimer disease, contains Lys16, Lys28, and Gln15 which may act as substrates of transglutaminase. Transglutaminase treatment of amyloid -peptide (1–28) and amyloid -peptide (1–40) yielded cross-linked oligomers. Transglutaminase-treated A retarded neurite extension of PC12 cells, and rat cultured neurons of hippocampus and septum, brain areas severely affected by Alzheimer disease, and subsequently caused cell death, whereas the transglutaminase-untreated counterparts did not show harmful effects. The transglutaminase-catalyzed oligomers of amyloid -peptide and their neurotoxicity may be involved in two characteristics in Alzheimer disease, neuronal degeneration and formation of the insoluble deposits.Abbreviations: AD – Alzheimer disease, A – amyloid -peptide, DMEM – Dulbecco's modified Eagle's medium, DMEM/F–12–1:1 mixture of DMEM and Ham's F–12 medium, FCS – fetal calf serum, HS – horse serum, PAGE – polyacrylamide gel electrophoresis, MTT – 3-(4,5-dimethylthiazol–2-yl)–2,5-diphenyltetrazolium bromide, NGF – nerve growth factor, TGase – transglutaminase.     

Peroxidation of lipids and growth inhibition induced by UV-B irradiation

Cotyledons excised from dark-grown seedlings of cucumber (Cucumis sativus L.) were cultured in vitro under UV radiation at different wavelengths, obtained by passage of light through cut-off filters with different transmittance properties. Growth and the synthesis of chlorophyll (Chl) in cotyledons were inhibited and malondialdehyde was accumulated upon irradiation at wavelengths below 320 nm. Exogenous application of scavengers of free radicals reversed the growth inhibition induced by UV-B. Measurement of the fluorescence of Chl a suggested that electron transfer in photosystems was affected by UV-B irradiation. On the basis of these results, the involvement is postulated of active species of oxygen in damages to thylakoid membranes and the growth inhibition that are induced by UV-B irradiation.Abbreviations Chl chlorophyll - F_m maximal fluorescence (dark) - F_m maximal fluorescence (light) - F_v variable fluorescence (dark) - F_v variable fluorescence (light) - MDA malondialdehyde - O₂ Superoxide radical - PS photosystem - q_N non-photochemical quenching of fluorescence - q_P photochemical quenching of fluorescence - UV-B_BE biologically effective UV-B radiation - WL(T = 0.5) wavelength at which 50% transmittance occurs     

Occurrence and induction of spermidine-N1-acetyltransferase in Escherichia coli

Spermidine acetylase activity was detected in extracts prepared from $\text{Escherichia coli}$ and there was a marked increase in activity over the early period of growth. This increase reached a maximum 3 h after inoculation and was followed by an increase in ornithine decarboxylase activity. The acetylase was also able to use spermine as a substrate, but not putrescine. With spermidine and acetyl-CoA as substrate, the product formed was exclusively N¹-acetyl-spermidine. This is the first evidence for the occurrence in bacteria of spermidine-N¹-acetyltransferase, an enzyme which has previously been described in mammalian cells. These results suggest that acetylation of spermidine may be involved in the growth of $\text{Escherichia coli}$ and in the regulation of its polyamine content.     

Prostaglandin E2 Activates Ca2+ Channels in Bovine Adrenal Chromaffin Cells

We have demonstrated that prostaglandin E2 (PGE2) treatment of bovine adrenal chromaffin cells results in a sustained elevation of intracellular Ca2+ concentration ([Ca2+]i) in these cells. Because the continued elevation of [Ca2+]i was dependent on extracellular Ca2+ concentration, it can be assumed that the PGE2-induced [Ca2+]i increase is due, at least in part, to an opening of membrane Ca2+ channels. In this study, we used electrophysiological methods to examine the mechanism of the PGE2-induced [Ca2+]i increase directly. Puff application of PGE2 to the external medium resulted in a prolonged depolarization in about half of the chromaffin cells examined. In whole-cell voltage-clamp recordings, an increase in inward current was observed over a 6-7 min period following bath application of PGE2 (greater than or equal to 10 microM), even in the absence of external Na+. This inward current was abolished when the recordings were made with the cells in a Ca2(+)-free medium, but it was not inhibited by Mn2+, a blocker of voltage-dependent Ca2+ channels. In cell-attached patch-clamp configuration, PGE2 produced an increase in the opening frequency of inward currents. The reversal potential of the PGE2-induced currents was about +40 mV, which is close to the reversal potential of the Ca2+ channel. The opening frequency was not affected by membrane potential changes. In inside-out patch-clamp configuration, inositol 1,4,5-trisphosphate (2 microM) added to the cytoplasmic side activated the Ca2(+)-channel currents, but PGE2 was ineffective when applied to the cytoplasmic side. These results suggest that PGE2 activates voltage-independent Ca2+ channels in chromaffin cells through a diffusible second messenger, possibly inositol 1,4,5-trisphosphate.     

Effect of glucose on protein phosphorylation in rat pancreatic islets

Isolated rat pancreatic islets, incubated in the presence of extracellular ³²Pi to steady state ³²P incorporation into cellular phosphopeptides, were exposed to glucose for 10 min. Glucose (16.7 mM) significantly stimulated the phosphorylation of six phosphoproteins with molecular weights of 15,000, 35,000, 49,000, 64,000, 93,000 and 138,000. Mannoheptulose (16.7 mM) markedly inhibited glucose-stimulated phosphorylation of these six phosphoproteins. This protein phosphorylation might be important in mediating glucose-stimulated insulin release.     

Characterization of O-trimethylsilyl derivatives of d-glucose,d-galactose,and d-mannose by gas-liquid chromatography-chemical-ionization mass spectrometry with ammonia as reagent gas

The per(trimethylsilyl) ethers of d-glucose, d-galactose, and d-mannose were analyzed by g.l.c.-c.i.m.s. with ammonia as the reagent gas. C.i.m.s. gave simple fragmentation and fragment ions of high intensity in the high-mass range where the QM⁺ ion is also detected. The β-d anomers gave ions at m/e 558 showing intensities 3–12 times those of the α-d anomers. The epimers could be distinguished by differences in the intensities of the ions and by the observation that d-glucose gave a base peak at m/e 198, d-galactose at m/e 468, and d-mannose at m/e 204. The pyranose and furanose structures could be distinguished by comparing the ion intensities at m/e 198, m/e 271, m/e 361, m/e 396, and m/e 451. A similar analysis was also performed with 2-methylpropane as the reagent gas.