Effect of TRAIL in combination with DDP on the expression of MDR1 gene in gastric cancer cells.

INTRODUCTION: Gastric cancer is one of the most common malignant tumor, and gastric cancer is the second most common cause of cancer mortality worldwide. Although chemotherapy is one of the most important treatment options for gastric cancer, and could improve the overall survival rate and quality of live, one significant reason for its failure is multidrug resistance (MDR). AIM: To study the effect of tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) combined with chemotherapeutic drug cisplatin (DDP) on the expression of multidrug resistance gene 1 (MDR1) in the gastric cancer cell line SGC-7901/VCR. MATERIAL AND METHODS: SGC-7901/VCR cells were cultured with DDP and TRAIL in various concentrations. The apoptosis rate was separately measured by a flow cytometer in DDP (sub-toxic dose) alone, TRAIL (200 µg/l) alone and in a combination of the two. Expression levels of MDR1 mRNA and P-glycoprotein (P-gp) were detected by RT-PCR and ELISA analysis, respectively. RESULTS: The apoptosis rate in the combination group was significantly higher than that in the other groups (p < 0.05). According to the results of RT-PCR and ELISA, the expressions of MDR1 mRNA and P-gp in the combination group were statistically significant different compared with other groups (p < 0.05). CONCLUSIONS: The combination of TRAIL with DDP could reverse MDR phenotype in gastric cancer cell line SGC7901/VCR. The mechanism may be involved in the down-regulation of MDR1 mRNA and P-gp, which may play an essential role in overcoming the chemotherapeutic resistance of gastric cancer cells. This study indicates that a combination of chemotherapy and TRAIL may be an effective strategy to treat MDR gastric cancer.

The effect of TRAIL on the expression of multidrug resistant genes MDR1, LRP and GST-π in drug-resistant gastric cancer cell SGC7901/VCR.

BACKGROUND/AIMS: To investigate the effect of tumor necrosis factor related apoptosis inducing ligand(TRAIL) on the expression of multidrug resistant genes MDR1, LRP and GST-n in drug-resistant gastric cancer cell strain SGC7901/VCR, and discuss a potential mechanism that reverses the multidrug resistance of gastric cancer with TRAIL as the target point. METHODOLOGY: SGC7901/VCR cell strain was treated over 48 h with TRAIL (50, 100, 200 and 400ig/L, respectively). The expression ofMDR1, LRP, GST-r mRNA in different groups of gastric cell strains was tested by RT-PCR and the expression of P-gp, LRP and GST-n by ELISA. RESULTS: Under the action of TRAIL of different concentrations, different degrees of inhibition were observed in the expression of the mRNA and protein, and the difference from the reference group was statistically significant(p<0.01). Except for the insignificant inhibition degree of mRNA and protein in MDR1, LRP and GST-nr as compared between the 400lg/L and the 2001ig/L group(p>0.05), the differences between other groups were all statistically significant (p<0.05). CONCLUSIONS: As preliminarily estimated from the results of the study, TRAILis negatively correlated with drug-resistant genes. It is possible that TRAIL increases the apoptosis and growth inhibition of chemotherapy drug tumor cells by reducing the expression of drug-resistant genes MDR1, LRP and GST-Tt, thereby participating in the reversion of the multidrug resistance of gastric cancer

Reducing ischaemia/reperfusion injury through delta-opioid-regulated intrinsic cardiac adrenergic cells: adrenopeptidergic co-signalling.

AIMS: The purpose of this study was to determine whether intrinsic cardiac adrenergic (ICA) cells release calcitonin gene-related peptide (CGRP), exerting synergistic adrenopeptidergic cardioprotection. METHODS AND RESULTS: In situ hybridization coupled with immunostaining demonstrated that ICA cells exclusively expressed CGRP mRNA and co-expressed CGRP and delta-opioid receptor in human and rat left ventricular (LV) myocardium. Radioimmunoassay detected constitutive CGRP release from ICA cells in human and rat hearts. The delta-opioid agonist [D-Pen(25)]-enkephalin (DPDPE) increased CGRP release from ICA cells in denervated rat heart. In an ischaemia/reperfusion rat model, pre-ischaemic treatment with DPDPE reduced infarct size (IS) by 51 +/- 16% (P < 0.01). Co-infusion of beta(2)-adrenergic receptor (beta(2)-AR) and CGRP receptor (CGRP-R) antagonists increased IS by 62 +/- 23% (P < 0.01) compared with saline and abolished DPDPE-initiated IS reduction. Pre-treatment of ICA cell-myocyte co-culture with the beta(2)-AR/CGRP-R antagonists increased myocyte death rate by 24 +/- 4% (P < 0.01) and abolished DPDPE-initiated myocyte protection against hypoxia/reoxygenation (re-O(2)). In the ICA cell-depleted myocyte culture, DPDPE did not confer myocyte protection. Supplementing ICA cell-depleted myocyte culture with beta(2)-AR/CGRP-R agonists reduced hypoxia/re-O(2)-induced myocyte death by 24 +/- 5% (P < 0.01), simulating endogenous neurohormonal effects of ICA cells. Western blot analysis showed that DPDPE markedly increased phosphorylated myocardial Akt levels. This effect was abolished in the presence of beta(2)-AR/CGRP-R blockade. Terminal dUTP nick-end labelling staining analysis of the LV infarct zone demonstrated that DPDPE reduced myocyte apoptosis by 58 +/- 19% (P < 0.05), an effect that was eliminated in the presence of beta(2)-AR/CGRP-R blockade. Finally, echocardiography showed that DPDPE increased LV contractility in a manner dependent on beta-AR/CGRP-R stimulation. CONCLUSION: ICA cells constitute a delta-opioid-regulated adrenopeptidergic paracrine system conferring robust cardioprotection through beta(2)-AR/CGRP-R co-signalling, resulting in the activation of an anti-apoptotic pathway during ischaemia/reperfusion.

Loss of cell-adhesion molecule complexes in solid pseudopapillary tumor of pancreas.

Solid pseudopapillary tumor of pancreas (SPT) is a rare neoplasm that occurs most often in young females with the two distinct features, the 'solid-cystic' gross appearance, and the 'solid-pseudopapillary' microscopic pattern. It has been reported that almost all SPT tumors contain a mutation in the beta-catenin gene; however, the histogenetic origin of this tumor remains largely a mystery. E-cadherin is a cell adhesion molecule that links to catenins to form cell adhesion junctions, which is associated with the cytoskeleton formation. In this study, we examined the expression of E-cadherin and beta-catenin from SPT in an attempt to determine the molecular basis for the unusual morphology of this tumor. Nine cases of SPT were retrieved from Surgical Pathologic Archives of three institutions, including one male and eight females. H&E slides of each case were reviewed to confirm the diagnosis. The beta-catenin gene was sequenced in one case. E-cadherin and beta-catenin immunostains, were performed on all nine cases. Sequencing analysis on one case showed a point mutation of the beta-catenin gene, confirming previous findings that almost all SPT tumors contain mutation in the beta-catenin gene. Immunostains showed that, in both solid and pseudopapillary areas, all the tumor cells lost expression of E-cadherin, and beta-catenin nuclear expression was observed in all cases. Our findings suggest that loss of cytoplasmic beta-catenin protein in the cell adhesion complex due to beta-catenin gene mutation, results in instability of the complex, loss of E-cadherin in cell membrane, and eventually dissociation of the tumor cells to form the pseudopapillary pattern.

Progesterone-induced sphingosine kinase-1 expression in the rat uterus during pregnancy and signaling consequences.

Sphingosine 1-phosphate (Sph-1-P), a product of sphingomyelin metabolism, can act via a family of cognate G protein-coupled receptors or as an intracellular second messenger for agonists acting through their membrane receptors. In view of the general growth promoting and developmental effects of Sph-1-P on target cells, we hypothesized that it plays a role in adaptation of the uterus to pregnancy. We analyzed its potential role and that of the related lysophospholipid lysophosphatidic acid in the pregnant rat uterus by examining changes in mRNA levels of cognate receptors and enzymes involved in their turnover. Of these, only sphingosine kinase-1 (SphK1) was markedly changed ( approximately 30-fold increase), being localized in the glandular epithelium, vasculature, and the myometrium. Uterine SphK1 mRNA and protein levels paralleled those of serum progesterone, and treatment with progesterone or an antagonist elevated or reduced SphK1 mRNA expression, respectively. Progesterone also increased SphK1 mRNA steady-state levels in a rat myometrial/leiomyoma cell line (ELT3). Overexpressing human SphK1 in these cells resulted in increased levels of the cell cycle regulator cyclin D1 and increased myosin light-chain phosphorylation. Ectopic expression of SphK1 also resulted in increased proliferation rates, possibly in conjunction with increased cyclin D1 expression. These studies suggest that the uterine expression of SphK1 mediates processes involved in growth and differentiation of uterine tissues during pregnancy.

Estrogenic regulation of host immunity against an estrogen receptor-negative human breast cancer.

PURPOSE: The risk of developing breast cancer is positively correlated with exposure to increased levels of estrogen and/or an increased duration of estrogen exposure. Many different mechanisms have been proposed to explain the association of estrogens with breast cancer risk; however, the well-documented immune modulatory properties of estrogen have received little attention. In part, this is due to a lack of suitable models for studying this relationship. EXPERIMENTAL DESIGN: We have developed an animal model using estrogen receptor (ER)-negative human breast cancer cell line, MDA-MB-468, xenografted into severe combined immunodeficient (SCID) mice. We also generated the ER-alpha knockout (ER-alphaKO) mice on the SCID background and then tested the ability of 17beta-estradiol to stimulate growth of xenografted ER-negative human breast cancer tumors in wild-type and ER-alphaKO SCID mice. We quantified vascularization of tumors, macrophage recruitment to the tumor site by immunocytochemistry, and inflammatory cytokine production. RESULTS: We show that estrogen treatment of C57BL/6/SCID mice promotes the growth of xenografted ER-negative tumors in wild-type mice and this estrogen-induced tumor growth is abrogated in ER-alphaKO mice. Tumor neovascularization of estrogen-treated mice was unchanged versus control; however, estrogen treatment of the C57BL/6/SCID host suppressed macrophage recruitment to and inflammatory cytokine production at the tumor site. CONCLUSIONS: These data are consistent with estrogen modulation of the inflammatory response as a contributing factor in estrogen-stimulated growth of an ER-negative tumor. This effect on the host innate immune response was mediated by ER-alpha.

An intradermal environment promotes a protective type-1 response against lethal systemic monocytotropic ehrlichial infection.

Immune responses against monocytotropic ehrlichiosis during infection with a strain of Ehrlichia from Ixodes ovatus (IOE) were evaluated using a model that closely reproduces the pathology and immunity associated with tick-transmitted human monocytotropic ehrlichiosis. C57BL/6 mice were inoculated intradermally or intraperitoneally with high-dose highly virulent IOE or intraperitoneally with mildly virulent Ehrlichia muris. Intradermal (i.d.) infection with IOE established mild, self-limited disease associated with minimal hepatic apoptosis, and all mice survived past 30 days. Intraperitoneal (i.p.) infection with IOE resulted in acute, severe toxic shock-like syndrome and severe multifocal hepatic apoptosis and necrosis, and all mice succumbed to disease. Compared to i.p. infection with IOE, intradermally infected mice had a 100- to 1,000-fold lower bacterial load in the spleen with limited dissemination. Compared to mice infected intraperitoneally with IOE, i.d. infection stimulated a stronger protective type-1 cell-mediated response on day 7 of infection, characterized by increased percentages of both CD4+ and CD8+ splenic T cells, generation of a greater number of IOE-specific, gamma interferon-producing CD4+ Th1 cells, and higher levels of tumor necrosis factor (TNF-alpha) in the spleen but lower concentrations of serum TNF-alpha and interleukin-10. These data suggest that under the conditions of natural route of challenge (i.e., i.d. inoculation), the immune response has the capacity to confer complete protection against monocytotropic ehrlichiosis, which is associated with a strong cell-mediated type-1 response and decreased systemic production of pro- and anti-inflammatory cytokines.

Hormonal regulation of catechol-O-methyl transferase activity in women with uterine leiomyomas.

Catechol-O-methyl transferase (COMT) expression is higher in leiomyomas compared with paired normal myometrium. The expression of COMT in leiomyoma cells is hormonally regulated-estrogen down-regulates, whereas P and dexamethasone up-regulate, COMT expression.

p75(NGFR) immunostaining for the detection of perineural invasion by cutaneous squamous cell carcinoma.

BACKGROUND: Perineural invasion (PNI) in cutaneous squamous cell carcinoma (CSCC) may portend a poor prognosis for patients. p75NGFR (nerve growth factor receptor) is part of a membrane receptor complex that binds nerve growth factor. Its use for detecting PNI in CSCC in comparison with S-100 immunohistochemical staining has not been explored. OBJECTIVE: To determine whether detection of PNI may be improved by staining with p75NGFR compared with hematoxylin and eosin (H&E) and S-100. METHODS: Thirty-four cases of CSCC were retrospectively evaluated for the presence of PNI using standard H&E, as well as S-100 and p75NGFR immunohistochemical stains. Staining intensity was correlated to the presence or absence of PNI and tumor differentiation. RESULTS: The results showed a positive correlation between staining intensity and the presence of PNI detected by p75NGFR (p=.04). Using p75NGFR allowed for the detection of seven cases of PNI not detected by H&E alone. Five of these cases were detected by S-100, with two cases seen by p75NGFR only. Six cases of PNI were detected using S-100 not seen on H&E, with one case also not seen using p75NGFR. CONCLUSION: p75NGFR immunostaining increased detection of PNI compared with H&E. p75NGFR could serve as an alternative to S-100 in the detection of PNI or as part of an immunostaining panel for PNI detection.

Effect of chronic hyperghrelinemia on ingestive action of ghrelin.

The stomach hormone ghrelin is the endogenous ligand for the growth hormone secretagogue receptor (GHS-R). Systemic administration of ghrelin will cause elevations in growth hormone (GH) secretion, food intake, adiposity, and body growth. Ghrelin also affects insulin secretion, gastric acid secretion, and gastric motility. Several reports indicate that repeated or continuous activation of GHS-R by exogenous GHSs or ghrelin results in a diminished GH secretory response. The purpose of this study was to examine the extent to which the acute stimulation of food intake by exogenous ghrelin is altered by chronic hyperghrelinemia in transgenic mice that overexpress the human ghrelin gene. The present findings show that the orexigenic action of exogenous ghrelin is not diminished by a chronic hyperghrelinemia and indicate that the food ingestive pathway of the GHS-R is not susceptible to desensitization. In contrast, the epididymal fat pad growth response, like the GH response, to exogenous ghrelin is blunted in ghrelin transgenic mice with chronic hyperghrelinemia.