Prion protein-dependent regulation of p53-MDM2 crosstalk during endoplasmic reticulum stress and doxorubicin treatments might be essential for cell fate in human breast cancer cell line, MCF-7.

In this study, we investigated the effect of doxorubicin and tunicamycin treatment alone or in combination on MDM-, Cul9-and prion protein (PrP)-mediated subcellular regulation of p53 in the context of apoptosis and autophagy. MTT analysis was performed to determine the cytotoxic effect of the agents. Apoptosis was monitorized by ELISA, flow cytometry and JC-1 assay. Monodansylcadaverine assay was performed for autophagy. Western blotting and immunofluorescence were performed to determine p53, MDM2, CUL9 and PrP levels. Doxorubicin increased p53, MDM2 and CUL9 levels in a dose-dependent manner. Expression of p53 and MDM2 was higher at the 0.25 µM concentration of tunicamycin compared to the control, but it decreased at 0.5 µM and 1 µM concentrations. CUL9 expression was significantly decreased only after treatment of tunicamycin at 0.25 µM. According to its glycosylation status, the upper band of PrP increased only in combination treatment. In combination treatment, p53 expression was higher than control, whereas MDM2 and CUL9 expressions were decreased. Combination treatments may make MCF-7 cells more susceptible to apoptosis rather than autophagy. In conclusion, PrP may be important in determining the fate of cell death through crosstalk between proteins such as p53 and MDM2 under endoplasmic reticulum (ER) stress conditions. Further studies are needed to obtain in-depth information on these potential molecular networks.

Is genetic screening necessary for determining the possibility of venous thromboembolism in cancer patients?

OBJECTIVE: To determine the risk of an association with some genetic polymorphisms involved in venous thromboembolism (VTE) gene variations (FVL, FV H1299R, FII G20210A, MTHFR C677T, MTHFR A1298C, PAI-1 4G/5G, ß-fibrinogen -455 G → A, FXIII Val34Leu and GpIIIa HPA-1a) in cancer patients. SUBJECTS AND METHODS: Among 78 cancer patients, 28 who had proven first episode of VTE were selected as the patient group, with 50 control samples selected from age-, sex- and body mass index-matched healthy volunteers (healthy group). The differences in frequency of genetic polymorphisms were found to be statistically insignificant between these two groups. RESULTS: Logistic regression analysis after adjustment for age, sex, smoking and hypertension showed no difference. The screened mutations of these genes were not significantly associated with VTE risk. CONCLUSION: There is no possible benefit from genetic screening tests regarding VTE in cancer patients.

Electrochemical genosensor based on colloidal gold nanoparticles for the detection of Factor V Leiden mutation using disposable pencil graphite electrodes.

Electrochemical genosensors for the detection of the Factor V Leiden mutation from polymerase chain reaction (PCR) amplicons using the oxidation signal of colloidal gold (Au) is described. A pencil graphite electrode (PGE) modified with target DNA, when hybridized with complementary probes conjugated to Au nanoparticles, responded with the appearance of a Au oxide wave at approximately +1.20 V. Specific probes were immobilized onto the Au nanoparticles in two different modes: (a) Inosine-substituted probes were covalently attached from their amino groups at the 5' end using N-(3-dimethylamino)propyl)-N'-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysulfosuccinimide (NHS) as a coupling agent onto a carboxylate-terminated l-cysteine self-assembled monolayer (SAM) preformed on the Au nanoparticles, and (b) probes with a hexanethiol group at their 5' phosphate end formed a SAM on Au nanoparticles. The genosensor relies on the hybridization of the probes with their complementary targets, which are covalently immobilized at the PGE surface. Au-tagged 23-mer capture probes were challenged with the synthetic 23-mer target, 131-base single-stranded DNA or denatured 256-base polymerase chain reaction (PCR) amplicon. The appearance of the Au oxidation signal shortened the assay time and simplified the detection of the Factor V Leiden mutation from PCR amplified real samples. The discrimination between the homozygous and heterozygous mutations was also established by comparing the peak currents of the Au signals. Numerous factors affecting the hybridization and nonspecific binding events were optimized. The detection limit for the PCR amplicons was found to be as low as 0.78 fmol; thus, it is suitable for point-of-care applications.