Cytokine single nucleotide polymorphisms in patients’ with gallstone: dose TGF-β gene variants affect gallstone formation?

Gallstone is a common biliary disorder with several risk factors. Immune responses and inflammatory cytokines are important in this disease; as a result, some cytokines can be detected in bile fluid. In this research, cytokine gene polymorphisms were studied, and their effects on gallstone formation were evaluated. On 158 gallstone patients and 254 normal subjects, by PCR- RFLP method, IL-4-C590T polymorphism and by ARMS-PCR method, IFN-γ T+874A, TNF-α-A308G, IL-6 G-174C and TGF-β T+869C variants were studied. Pathologic evaluations were done on surgical specimens. There were no significant differences in distribution of evaluated polymorphisms between patient group and normal control group (P > 0.05), except TGF-β +869T allele (P = 0.04, OR = 1.23, 95 % CI = 1–1.79) which was higher in patients with gallstone. Although the pro-inflammatory cytokines such as TNF-α and IL-6 may promote gallstone formation, in this study no significant correlation between TNF-α and IL-6 polymorphisms and gallstone formation was seen. It is taught that TGF-β may affect gallbladder cells to promote gallstone formation and higher producer TGF-β +869T allele can be a risk factor of gallstone disease, so further studies would be more elucidative.     

Interconnections between apoptotic,autophagic and necrotic pathways: implications for cancer therapy development

The rapid accumulation of knowledge on apoptosis regulation in the 1990s was followed by the development of several experimental anticancer‐ and anti‐ischaemia (stroke or myocardial infarction) drugs. Activation of apoptotic pathways or the removal of cellular apoptotic inhibitors has been suggested to aid cancer therapy and the inhibition of apoptosis was thought to limit ischaemia‐induced damage. However, initial clinical studies on apoptosis‐modulating drugs led to unexpected results in different clinical conditions and this may have been due to co‐effects on non‐apoptotic interconnected cell death mechanisms and the ‘yin‐yang’ role of autophagy in survival versus cell death. In this review, we extend the analysis of cell death beyond apoptosis. Upon introduction of molecular pathways governing autophagy and necrosis (also called necroptosis or programmed necrosis), we focus on the interconnected character of cell death signals and on the shared cell death processes involving mitochondria (e.g. mitophagy and mitoptosis) and molecular signals playing prominent roles in multiple pathways (e.g. Bcl2‐family members and p53). We also briefly highlight stress‐induced cell senescence that plays a role not only in organismal ageing but also offers the development of novel anticancer strategies. Finally, we briefly illustrate the interconnected character of cell death forms in clinical settings while discussing irradiation‐induced mitotic catastrophe. The signalling pathways are discussed in their relation to cancer biology and treatment approaches.     

Cytotoxic activity of some marine brown algae against cancer cell lines

The aim of this study was to investigate the in vitro cytotoxic activity of total extract of MeOH (70%) and partition fractions of hexan, chloroform (CHCL3), ethylacetate (EtOAc) and MeOH-H2O of brown algae species (Sargassum swartzii, Cystoseira myrica, Colpomenia sinuosa) found in the Persian Gulf against in different cell lines including HT-29, Caco-2, T47D, MDA-MB468 and NIH 3T3 cell lines by MTT and AnnexinV-PI assay. The hexan fraction of S. swartzii and C. myrica showed selective cytotoxicity against proliferation of Caco-2 cells (IC50 < 100 μg/ml) T47D cell line (IC50<100 μg/ml), respectively. S. swartzii and C. myrica were also observed for increasing apoptosis in Caco-2 and T47D cells. Total extract and fractions of C. sinuosa did not show any significant cytotoxicity against the studied cell lines. MDA-MB468 cells were more sensitive to C. myrica than was T47D (IC50 99.9 ± 8.11 vs. 56.50' ± 0.88). This reflects an estrogen receptor independent mechanism for cytotoxicity of the extract. The IC50 of the hexan fraction of C. myrica on T47D parent cells was lower than it was on T47D-TR cells (IC50 99.9 ± 8.11 vs. 143.15 ± 7.80). This finding suggests a role for the MDR-1 in the development of possible future tolerance to the extract.     

The Thyroid Receptor Modulator KB3495 Reduces Atherosclerosis Independently of Total Cholesterol in the Circulation in ApoE Deficient Mice

Background

Thyroid hormones (TH) regulate cholesterol metabolism but their use as lipid-lowering drugs is restricted due to negative cardiac effects. TH mimetic compounds modulating TH receptor β (THRβ) have been designed as potential drugs, reducing serum cholesterol levels while avoiding apparent deleterious cardiac effects.

Objective

Using ApoE deficient mice, we examined whether KB3495, a TH mimetic compound, reduces atherosclerosis and if there is a synergistic effect with atorvastatin. The effect of KB3495 was investigated after 10 and 25 weeks.

Results

KB3495 treatment reduced atherosclerotic plaque formation in aorta and decreased the cholesteryl ester (CE) content by 57%. Treatment with KB3495 was also associated with a reduction of macrophage content in the atherosclerotic plaques and reduced serum levels of IL-1β, TNFalpha, IL-6, Interferon γ, MCP-1 and M-CSF. Serum lipoprotein analysis showed no change in total cholesterol levels in ApoB-containing lipoproteins. KB3495 alone increased fecal BA excretion by 90%. The excretion of neutral sterols increased in all groups, with the largest increase in the combination group (350%). After 25 weeks, the animals treated with KB3495 showed 50% lower CE levels in the skin and even further reductions were observed in the combination group where the CE levels were reduced by almost 95% as compared to controls.

Conclusion

KB3495 treatment reduced atherosclerosis independently of total cholesterol levels in ApoB-containing lipoproteins likely by stimulation of sterol excretion from the body and by inhibition of the inflammatory response.     

Hepatic ACAT2 Knock Down Increases ABCA1 and Modifies HDL Metabolism in Mice

Objectives

ACAT2 is the exclusive cholesterol-esterifying enzyme in hepatocytes and enterocytes. Hepatic ABCA1 transfers unesterified cholesterol (UC) to apoAI, thus generating HDL. By changing the hepatic UC pool available for ABCA1, ACAT2 may affect HDL metabolism. The aim of this study was to reveal whether hepatic ACAT2 influences HDL metabolism.

Design

WT and LXRα/β double knockout (DOKO) mice were fed a western-type diet for 8 weeks. Animals were i.p. injected with an antisense oligonucleotide targeted to hepatic ACAT2 (ASO6), or with an ASO control. Injections started 4 weeks after, or concomitantly with, the beginning of the diet.

Results

ASO6 reduced liver cholesteryl esters, while not inducing UC accumulation. ASO6 increased hepatic ABCA1 protein independently of the diet conditions. ASO6 affected HDL lipids (increased UC) only in DOKO, while it increased apoE-containing HDL in both genotypes. In WT mice ASO6 led to the appearance of large HDL enriched in apoAI and apoE.

Conclusions

The use of ASO6 revealed a new pathway by which the liver may contribute to HDL metabolism in mice. ACAT2 seems to be a hepatic player affecting the cholesterol fluxes fated to VLDL or to HDL, the latter via up-regulation of ABCA1.     

2-Methoxyestradiol-induced apoptosis in prostate cancer cells requires Smad7

Prostate cancer is the second most common cause of death related to cancer in Western society. 2-Methoxyestradiol (2-ME), an endogenous metabolite of estradiol-17beta, inhibits tumor angiogenesis while also exerting potent cytotoxic effects on various cancer cells. 2-ME has been shown to activate the p38 MAPK and JNK pathways and to induce apoptosis in cells, although the underlying molecular mechanisms for this are unknown. Here we report that the expression of Smad7, an adaptor molecule required to activate p38 MAPK in the transforming growth factor beta signaling pathway, is also required for 2-ME-induced p38 activation and apoptosis in human prostate cancer cells (PC-3U). PC-3U/AS-S7 cells stably transfected with an antisense Smad7 construct, or PC-3U cells transiently transfected with short interfering RNA for Smad7, were protected against 2-ME-induced apoptosis. 2-ME-induced apoptosis was found to involve p38 MAPK and JNK, because simultaneous treatments with 2-ME and a specific p38 inhibitor (SB203580) or an inhibitor of JNK (L-JNK1) prevented 2-ME-induced apoptosis. Most interestingly, Smad7 was shown by both antisense and short interfering RNA techniques to affect levels of beta-catenin, which has been implicated previously in the regulation of apoptosis. Moreover, Smad7 was found to be important for the basal expression of Bim, a pro-apoptotic Bcl-2 family member, and for 2-ME-induced expression of Bim. These results suggest that expression of Smad7 is crucial for 2-ME-induced apoptosis in human prostate cancer cells.     

Synthesis and biological evaluation of novel carbon-11-labelled analogues of citalopram as potential radioligands for the serotonin transporter

Three serotonin reuptake inhibitors where the 5-cyano group in citalopram [1-(3-dimethylamino-propyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile (1)] was replaced with a methyl, acetyl and piperidinyl carbonyl group, respectively, were synthesized. In a Stille reaction applying [(11)C]methyl iodide the labelled compound [5-methyl-(11)C][3-[1-(4-fluorophenyl)-5-methyl-1,3-dihydroisobenzofuran-1-yl]-propyl]-dimethylamine ([(11)C]-2) was synthesized in 60-90% radiochemical yield. [5-carbonyl-(11)C][1-[1-(3-dimethylaminopropyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-yl]-1-piperidin-1-yl-methanone] ([(11)C]-3) was synthesized in 62% radiochemical yield in a palladium mediated cross-coupling reaction utilizing [(11)C]carbon monoxide. The specific activity of [(11)C]-2 was highly dependent on whether the corresponding trimethyltin or tributyltin precursor was applied. In ex vivo rodent studies compound [(11)C]-2 exhibited a good blood-brain barrier (BBB) penetration whereas [(11)C]-3 did not. The brain distribution of [(11)C]-2 was investigated in a non-human primate using PET. There was a rapid uptake of radioactivity into the brain. Accumulation of the radiotracer was in agreement with the known distribution of serotonin transporters. The maximal thalamus to cerebellum ratio of 1.3 was reached after 85 min and the specific binding was partly blocked after pre-treatment with citalopram. Thus, [(11)C]-2 does not exhibit appropriate properties as radioligand for visualization of the serotonin transporter in vivo.     

CD40 expression in Wehi-164 cell line

CD40-CD154 interaction is an important process for cellular and humoral immunity regulation and can be effective in the body’s defense against tumors. In the present study, we evaluated the expression of CD40 in Wehi-164 cell line. CD40 expressions on the cell surface and in the cytoplasm were assessed by flow cytometry and intracellular staining assay, respectively. Also, the mRNA expression was identified by real time-PCR. The obtained results showed the high mRNA and cytoplasmic protein expression of CD40 but no surface expression. These results suggest that the Wehi-164 cell line down regulates expression of CD40 on the surface for evasion of immune system.