Juxtacrine function of interleukin-15/interleukin-15 receptor system in tumour derived human B-cell lines.

Interleukin-15 (IL-15) is a cytokine that induces proliferation and promotes cell survival of human T, B and NK cells. IL-15 and interleukin-2 (IL-2) exhibit a similar spectrum of immune effects and share the IL-2 receptor (IL-2R) subunits IL-2Rbeta and IL-2Rgamma(c) for signalling in haematopoietic cells. Furthermore, each cytokine has a private alpha receptor, namely IL-2Ralpha for IL-2 and IL-15Ralpha for IL-15, that functions in ligand binding. Using reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA) methods, the expression and secretion of IL-15 and IL-15Ralpha in tumour-derived B-cell lines were studied. The results as presented in this study identify that IL-15 mRNA is predominantly expressed in EBV positive (EBV(+)) B-cell lines, although IL-15Ralpha is ubiquitously and constitutively expressed in all these B-cell lines. Although no detectable levels of IL-15 protein secretion were observed in any of these cell lines, we were able to detect membrane-bound expression of IL-15 protein by FACS analysis in some cell lines. These data imply that the IL-15/IL-15R system requires complex regulatory mechanisms for protein secretion. Taken together, we speculate that these results suggest a juxtacrine, intracrine function for IL-15/IL-15R.

Induced mitogenic activity in AML-12 mouse hepatocytes exposed to low-dose ultra-wideband electromagnetic radiation.

Ultra-wideband (UWB) technology has increased with the use of various civilian and military applications. In the present study, we hypothesized that low-dose UWB electromagnetic radiation (UWBR) could elicit a mitogenic effect in AML-12 mouse hepatocytes, in vitro. To test this hypothesis, we exposed AML-12 mouse hepatocytes, to UWBR in a specially constructed gigahertz transverse electromagnetic mode (GTEM) cell. Cells were exposed to UWBR for 2 h at a temperature of 23 degrees C, a pulse width of 10 ns, a repetition rate of 1 kHz, and field strength of 5-20 kV/m. UWB pulses were triggered by an external pulse generator for UWBR exposure but were not triggered for the sham exposure. We performed an MTT Assay to assess cell viability for UWBR-treated and sham-exposed hepatocytes. Data from viability studies indicated a time-related increase in hepatocytes at time intervals from 8-24 h post exposure. UWBR exerted a statistically significant (p < 0.05) dose-dependent response in cell viability in both serum-treated and serum free medium (SFM) -treated hepatocytes. Western blot analysis of hepatocyte lysates demonstrated that cyclin A protein was induced in hepatocytes, suggesting that increased MTT activity after UWBR exposure was due to cell proliferation. This study indicates that UWBR has a mitogenic effect on AML-12 mouse hepatocytes and implicates a possible role for UWBR in hepatocarcinoma.

Arsenic trioxide-induced transcriptional activation of stress genes and expression of related proteins in human liver carcinoma cells (HepG2).

Arsenic is a naturally occurring element, but anthropogenic activities can lead to a substantial contamination of the environment. Exposure to arsenic has been associated with a significant number of adverse health effects in humans including: cardiovascular disease, diabetes, hearing loss, developmental abnormalities, anemia, neurologic and neurobehavioral disorder, leukopenia, eosinophilia, fibrosis of the liver and the kidney and various neoplasms. However, the cellular and molecular events associated with arsenic toxicity are poorly understood. Also, the precise mechanisms by which arsenic acts as a carcinogen in humans remain to be elucidated. In the present study, we used human liver carcinoma (HepG2) cells as a model to study the molecular mechanisms of arsenic-induced toxicity and carcinogenesis. We hypothesized that arsenic-induced expression of stress genes and related proteins may play a role in the cellular and molecular events leading to toxicity and tumorigenesis in liver cells. To test this hypothesis, we performed the MTT-assay for cell viability, the CAT-Tox (L) assay for gene induction, and the Western Blot analysis to assess the expression of cellular proteins including c-fos, HMTIIA, HSP70 and p53. Data obtained from the MTT assay indicated a strong dose-response relationship with respect to arsenic trioxide toxicity. Upon 48 hr of exposure, the chemical dose required to cause 50% reduction in cell viability (LD50) was computed to be 8.55 +/- 0.58 microg/ml. The CAT-Tox (L) assay showed statistically significant inductions (p<0.05) of c-fos, HMTIIA, and HSP70. Western blot analysis also demonstrated a dose-response relationship with regard to expression of specific cellular proteins. The p53 protein was expressed in arsenic trioxide-treated cells, however, the densitometric analysis did not show any significant differences (p<0.05) between treated and control cells. The lack of a significant induction of p53 may be due to the potential mitogenic effect of arsenic at low levels of arsenic exposure.

Neurotransmitter and neuropeptide modulation of high affinity choline uptake in Limulus brain.

The role of neurotransmitters in the modulation of the sodium-dependent high affinity choline uptake system (HAChUS) of the horseshoe crab, Limulus polyphemus has been investigated utilizing a tissue slice preparation. Choline uptake was significantly decreased by carbachol but unaffected by atropine and d-tubocurarine. The muscarinic agonist oxotremorine decreased choline uptake by 30.4% while the muscarinic antagonist, pirenzepine, increased uptake by 29.6%. Applied in combination, pirenzepine and oxotremorine abolished their individual effects resulting in control values for choline uptake. The non-cholinergic transmitters octopamine and serotonin significantly enhanced choline uptake. The neuropeptide proctolin elicited a 20% increase in choline transport whereas Phe-Met-Arg-Phe (FMRF) amide was without effect. This study demonstrates that neurotransmitters and neuropeptides modulate the HAChUS, possibly through specific receptor-mediated second messenger systems.

Differential expression and subcellular localization of protein kinase C isoforms in neuronal and cardiac tissues of Limulus polyphemus.

The presence and subcellular distribution of specific protein kinase C (PKC) isoforms in Limulus neuronal and cardiac tissue extracts were determined using polyclonal antibodies to individual PKC isoforms. Western blot analysis revealed that the brain, abdominal ganglia, cardiac ganglia and heart all contained PKC's alpha, beta, gamma, epsilon and zeta. PKC delta was detected in all neuronal tissues. PKC theta was found only in the cardiac ganglia and heart. PKC's gamma and delta were predominantly localized in membrane fractions while the alpha, beta, epsilon, zeta and theta isoforms were found in both fractions. Antibodies to PKC's epsilon and zeta detected relatively high molecular weight bands in membrane fractions and lower molecular weight bands in the cytosolic fractions. The results demonstrate that Limulus neuronal and cardiac tissues differentially express the conventional, novel and atypical isoforms of protein kinase C.