Parathyroid hormone-related protein ameliorates death receptor-mediated apoptosis in lung cancer cells

Parathyroid hormone-related protein (PTHrP) is expressed in more advanced, aggressive tumors and may play an active role in cancer progression. This study investigated the effects of PTHrP on apoptosis after UV irradiation, Fas ligation, or staurosporine treatment in BEN human squamous lung carcinoma cells. Cells at 70% confluency were treated for 24 h with 100 nM PTHrP-(1-34), PTHrP-(38-64), PTHrP-(67-86), PTHrP-(107-139), or PTHrP-(140-173) in media with serum, exposed for 30 min to UV-B radiation (0.9 mJ/cm²), and maintained for another 24 h. Caspase-3, caspase-8, and caspase-9 activities increased fivefold. Pretreatment with PTHrP-(1-34) and PTHrP-(140-173) ameliorated apoptosis after UV irradiation, as indicated by reduced caspase activities, increased cell protein, decreased nuclear condensation, and increased clonal survival. Other peptides had no effect on measures of apoptosis. PTHrP-(140-173) also reduced caspase activities after Fas ligation by activating antibody, but neither peptide had effects on caspase-3 or caspase-9 activity after 1 µM staurosporine. These data indicate that PTHrP-(1-34) and PTHrP-(140-173) protect against death receptor-induced apoptosis in BEN lung cancer cells but are ineffective against mitochondrial pathways. PTHrP contributes to lung cancer cell survival in culture and could promote cancer progression in vivo. The mechanism for the protective effect against apoptosis remains to be determined. caspases; cell surface receptors; growth substances     

The role of calcium in oligogalacturonide-activated signalling in soybean cells

Alpha-1,4-Linked oligogalacturonides (OGs) are pectic fragments of the plant cell wall that are perceived by the plant cell as signalling molecules. Using cytosolic aequorin-expressing soybean (Glycine max L.) cells, we have analysed cytosolic Ca(2+) changes and the oxidative burst induced by OGs with different degrees of polymerization. Our results provide evidence that different OGs are sensed through transient elevations of cytosolic Ca(2+) that show different kinetics. Specificity of the Ca(2+) signature relies also on the precise structural characteristics of the OG molecules, such as the methylesterification of galacturonic acid residues and the steric conformation. Inhibition of the OG-induced Ca(2+) transient also blocks the oxidative burst, indicating that the cytosolic Ca(2+) increase is one of the earliest steps in OG-activated signalling. However, a phosphorylation event seems to precede the Ca(2+) rise, because the Ca(2+) transient could be abolished by the protein kinase inhibitor 4,5,6,7-tetrabromobenzotriazole (TBB). A pharmacological approach with different antagonists that interfere with the induction of the cytosolic Ca(2+) rise indicates that both extracellular Ca(2+) influx and intracellular Ca(2+) release participate in transducing the OG signal. Treatment of cells with OGs establishes a refractory state, which impairs the ability of the cell to respond to a second stimulus with the same elicitor for up to 16 h. This desensitization period could be prolonged with the phosphatase inhibitor okadaic acid, and eliminated with the protein kinase inhibitor Ro 31-8220, suggesting that phosphorylation events may be involved in the establishment of the cell refractory state.     

Effect of Pomphorhynchus laevis (Acanthocephala) on putative neuromodulators in the intestine of naturally infected Salmo trutta

Immunohistochemical and pathological studies were carried out on the digestive tract of parasitized and uninfected specimens of Salmo trutta (L.). A total of 124 brown trout were collected on several occasions from 3 tributaries of the Brenta River, northern Italy. Twenty-eight individuals of S. trutta (22.6%) were parasitized with Pomphorhynchus laevis (Miller, 1776). The occurrence of P. laevis in the trout gut significantly increased the number of endocrine cells immunoreactive to calcitonin gene-related peptide (CGRP), beta-endorphin, met-enkephalin, neuropeptide Y (NPY) and Substance P (SP) antisera. Moreover, bombesin-, cholecistokinin-8- (CCK-8), leu-enkephalin- and serotonin- (5-HT)-like immunoreactive cells were less numerous in the intestine of the parasitized brown trout. A strong positive immunoreactivity was observed in nerve fibres and neurones of the myenteric plexus of the parasitized fish; the antisera involved in this positive reactivity were bombesin, met-enkephalin, SP and vasoactive intestinal peptide (VIP). More neurones immunoreactive to anti-CGRP and anti-5-HT sera were noted in the myenteric plexus and in the inner layer of the tunica muscularis of the infected fish. Most of the above-mentioned neuromodulators are known to control gut motility, digestive/absorptive processes, as well as the immune response. The changes induced by parasites in the neuroendocrine system of the brown trout are discussed.     

Interleukin-18 enhances HIV-1 production in a human chronically-infected T cell line (H9-V)

Interleukin-18 (IL-18) is a recently identified immunoregulatory cytokine expressed by activated macrophages, that induces production of interferon-gamma (IFN-gamma) and Th-1 development. Recently some investigators reported controversial in vitro data on IL-18 stimulation of HIV-1 replication in several cell lines. In the present study the effect of IL-18 on HIV replication in a human chronically HIV-1-infected lymphocytic T cell line (H9-V) was investigated. HIV-1 replication was determined by an immunoassay method in order to evaluate the content of p24 antigen in the cell culture supernatants. Stimulation of H9-V cells with IL-18 resulted in increased production of p24, especially at concentrations of 0.01 microg ml(-1) and 0.10 microg ml(-1). Moreover a significant and persistent IL-18 stimulation of HIV-1 replication was observed at a concentration of 0.01 microg ml(-1) during a 7-day period. Pre-treatment of IL-18 with a specific neutralizing monoclonal antibody significantly reduced HIV-1 replication. These experiments show that IL-18 promotes the increase of HIV-1 replication in human chronically-infected lymphocytic T cells and confirm the role of IL-18 as a proimflammatory cytokine in stimulating and maintaining HIV-1 replication during the course of the disease. In a successive set of experiments, since one of the main activities of IL-18 is the induction of IFN-gamma, we evaluated the effect of this biological modifier on H9-V cells. In particular, IFN-gamma shows a significant effect on cell replication and on reduction of CD4 and CD71 surface expression.     

Pathogenesis of the deafness-associated A1555G mitochondrial DNA mutation

The pathogenic mechanisms of the A1555G mitochondrial DNA mutation in the 12S rRNA gene, associated with maternally inherited sensorineural deafness, are largely unknown. Previous studies have suggested an involvement of nuclear factor(s). To address this issue cybrids were generated by fusing osteosarcoma cells devoid of mtDNA with enucleated fibroblasts from two genetically unrelated patients. Furthermore, to determine the contribution, if any, of the mitochondrial and nuclear genomes, separately or in combination, in the expression of the disease phenotype, transmitochondrial fibroblasts were constructed using control and patient's fibroblasts as nuclear donors and homoplasmic mutant or wild-type cybrids as mitochondrial donors. Detailed analysis of mutant and wild-type cybrids from both patients and transmitochondrial fibroblast clones did not reveal any respiratory chain dysfunction suggesting that, if nuclear factors do indeed act as modifier agents, they may be tissue-specific. However, in the presence of high concentrations of neomycin or paromomycin, but not of streptomycin, mutant cells exhibit a decrease in the growth rate, when compared to wild-type cells. The decrease did not correlate with the rate of synthesis or stability of mitochondrial DNA-encoded subunits or respiratory chain activity. Further studies are required to determine the underlying biochemical defect.