Possible synergistic effect of apoE and LRP1 genotypes on metabolic syndrome development in Serbian patients.

The modern way of life contributes to the higher frequency of a complex state medically called metabolic syndrome (MetS), which is an inevitable consequence of several most common diseases of modern civilization. Patients with MetS have three times higher risk of experiencing a heart attack or a stroke and twice higher possibility to die from them. Serbia holds the infamous third place in Europe in mortality from heart disease, just behind Russia and Ukraine. The study explores the correlation of every combination of genotypes of apoE (apolipoprotein E) and LRP1 (low density receptor- related protein 1) genes with presence of MetS, and the connection with each anthropometric and biochemical parameter in both tested groups. Study demonstrates the impact of genotype combinations on the emergence and development of the MetS in Serbia. 63 patients and 30 controls were included in the study, aged from 19 to 65. Each person genotype was determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) profile. Odds ratio (OR) values showed that the presence of apoE e3e4/LRP1 CC genotype combination of genotypes in patients multiplies the chance (7.6 times) for the occurrence of the MetS in comparison to the presence of other genotype combinations. Determining the genetic basis of MetS is one of the necessary steps in the prevention of disease, saving the cost of treatment, and in the design of targeted therapies.

The LRP1 Gene Polymorphism is associated with Increased Risk of Metabolic Syndrome Prevalence in the Serbian Population.

The determination of genetic background in metabolic syndrome (MetS) represents one of the necessary steps to prevent the disorder, thus reducing the cost of medical treatments and helping to design targeted therapy. The study explores the association between individual alleles of the LRP1 gene and the diagnosis of MetS to find correlation between the low-density lipoprotein receptor-related (LRP1) gene polymorphism and each individual anthropometric and biochemical parameter. The study included 93 males and females, aged from 19 to 65, divided into two groups. The genotype of each person was determined from the restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR) profile. Results indicated the association of the T allele form of exon 3 LRP1 gene with development and progression of MetS that further pointed out its negative impact on tested anthropometric and biochemical parameters. The presence of the T allele in patients multiplies the chance of occurrence of deviations from the reference values of body mass index (BMI), (4.24-fold) and low-density lipoprotein (LDL) (20.26-fold) compared to C allele carriers. The results showed that T allele presence multiplies the chance (4.76 fold) for the occurrence of MetS in comparison to C allele carriers. Correlation found that the T allele of the LRP1 gene with MetS determinants is not negligible, therefore, the T allele may be considered as a risk factor for MetS development.

Expression analysis of genes involved in apoptosis, proliferation and endoplasmic reticulum stress in ionomycin/PMA treated Jurkat cells.

PURPOSE: Activation of T cells by direct stimulation with phorbol 12-myristate 13-acetate (PMA) and ionomycin (Io) results in numerous downstream signals that activate pathways enabling T cells to proliferate and produce cytokines. Inducible T cell activation is regulated predominantly at the transcriptional level. Therefore, we were interested to analyze the transcriptional activity of the 19 genes involved in the regulation of several important cellular processes. METHODS: Quantitative real-time (RT) PCR analysis was performed using mRNA-specific primers and SybrGreen for relative mRNA expression levels of all the examined genes. RESULTS: Our results showed c-kit expression in Jurkat cells, further confirmed by sequencing of c-kit mRNAspecific PCR product. The expected increased expression of interleukin (IL)-2 mRNA, together with moderate Ki-67 upregulation, indicate the proliferation of PMA/Io treated Jurkat cells. Significant upregulation of nuclear factor (NF)-κB, JNK and the prosurvival Bcl-2 was followed by activation of only one protein kinase RNA-like endoplasmic reticulum kinase (PERK) out of 3 main endoplasmic reticulum (ER) stress subpathways (ATF6 and spliced XBP were downregulated). NF-κB and JNK activation, as well as ERK downregulation were reactive oxygen species (ROS)-independent, shown by the lack of activation of antioxidative enzymes (SOD, NOS, GSTP1, gGCS and GR). C-kit was downregulated in the absence of exogenous SCF (c-kit ligand). CONCLUSION: Based on these data it is concluded that the PMA/Io treatment of Jurkat cells induced increased expression of IL-2, followed by upregulation of prosurvival genes belonging to the Bcl-2 family. Neither c-kit nor the antioxidative system were activated, excluding their role in Jurkat T-cell activation in the absence of exogenous c-kit ligand SCF.

Ethical and legal aspects of oncogenomics.

The discovery of the genes and cellular pathways that play fundamental roles in several diseases, and the understanding of many diseases at a molecular level due to the advances in the field of genomics, have revolutionized the diagnosis, therapy and prevention of human diseases. Application of genetic testing in numerous medical fields, including pharmacogenomics and oncogenomics, raised numerous ethical questions and introduced legal instruments that are aimed at ensuring the appropriate protection of human research participants. For the effective development of human genomics and translation of novel, validated biomarkers into potentially useful clinical applications in personalized medicine, there is a need for clear ethical standards and principles in all phases of clinical research.

Pleiotropic functions of bile acids mediated by the farnesoid X receptor.

In addition to their well-established role in the digestion and absorption of dietary lipids, bile acids (BAs) are recognized as signalling molecules in a wide range of metabolic processes. Bile acids regulate their own metabolism and enterohepatic circulation by activating the farnesoid X receptor (FXR). BAs have been shown to affect lipid metabolism, to decrease levels of circulating triglycerides, improve hyperglycemia and insulin signalling, directly act on the arterial wall and protect hepatocytes against cholestatic liver injury. Given that BAs are an integrated part of the complex metabolic network regulated by FXR, acting as a major underlying pathway, specific therapeutic targeting of this nuclear receptor represents an attractive therapeutic approach for a wide range of disorders. During a phase II clinical trial, the administration of a semisynthetic BA derivative 6-ethyl-chenodeoxycholic acid (6-ECDCA) to patients with diabetes, non-alcoholic fatty liver disease (NAFLD) and primary biliary cirrhosis (PBC), led to encouraging results, despite side effects being observed in pre-clinical studies. Chemical manipulations of the side chain and the steroid nucleus of BAs could lead to the discovery of novel semisynthetic BA derivatives that are more specific and selective FXR activators.

Expression analysis of genes involved in epigenetic regulation and apoptosis in human malignant haematopoietic cell lines treated with 5-azacytidine.

PURPOSE: The aim of this study was to investigate the modulation of the expression status of 10 different genes involved in epigenetic regulation and apoptosis by the DNA methyltransferase (DNMT) inhibitor 5-azacytidine (5-Aza), as markers of response to treatment, in two different human malignant haematopoietic cell lines. METHODS: In our analysis we used the SybrGreen technology and gene-specific primers for the qRT-PCR analysis of 10 genes, in cDNA of PC-MDS and K562 cell lines, treated by 1 micromole of 5-Aza for 24h. RESULTS: DNMT1 and DNMT3A showed statistically significant decrease of expression in 5-Aza-treated PC-MDS cells, whereas DNMT3B showed significantly decreased expression in 5-Aza-treated K562 cells. The members of the Bcl- 2 family of apoptosis-regulating genes Bcl-2 and Bax showed statistically significant differences in expression, in comparison with non-treated PC-MDS cells. Our most interesting result was the significant upregulation (re-expression) of p15, in 5-Aza-treated PC-MDS cells. CONCLUSION: The re-expression of p15 in PC-MDS cell line evaluated by qRT-PCR makes this novel cell line a suitable model for the studies of pharmacologic demethylation as a plausible mechanism resulting in hematologic response in myelodysplastic syndrome (MDS).

Quantitative real time-polymerase chain reaction method in Bcr-Abl translocation diagnostics.

PURPOSE: The quantitative real-time polymerase chain reaction (qRT-PCR) is used in the detection of molecular events involved in leukemogenesis, such as the Bcr-Abl gene translocation, the most important factor in the pathogenesis of chronic myeloid leukaemia (CML). The main aim of our study was to test the reproducibility, specificity and sensitivity of the qRT-PCR in the detection of Bcr-Abl gene translocation. METHODS: In complementary (c)DNA, isolated from K562 Bcr-Abl positive cell line, we performed qRT-PCR analysis with Bcr-Abl specific primers. For qRT-PCR analysis, we used serial dilutions of the newly synthesized cDNA in order to establish the detection threshold of this method. RESULTS: Using the specific primers for the Bcr-Abl translocation, we obtained the specific translocation product in cDNA sample of K562 human erythroid leukemia cell line. qRT- PCR showed significant sensitivity with the detection threshold for the Bcr-Abl fluorescent signal, which enabled the precise detection that was accurate within a 10-fold dilution range, and a dynamic range of 5 orders of magnitude. CONCLUSION: The results of our study showed that the application of the qRT-PCR is the optimal method for the detection of Bcr-Abl gene translocation, characterized by high reproducibility, specificity and sensitivity.

Evidence for interaction between the TCO and NMTC1 loci in familial non-medullary thyroid cancer.

BACKGROUND: Familial non-medullary thyroid cancer (fNMTC) is a complex genetic disorder that is more aggressive than its sporadic counterpart. Thus far, three genetic loci have been implicated in susceptibility to fNMTC by linkage analysis. METHODS: We used linkage analysis to test the significance of two of the known susceptibility loci for fNMTC, TCO on 19p13 and NMTC1 on 2q21 in 10 fNMTC families, nine of which present with cell oxyphilia, a rare histological phenotype associated with TCO. Furthermore, we used two-locus linkage analysis to examine the possibility that the TCO and NMTC1 loci interact to increase the risk of NMTC. RESULTS: The 10 families provided evidence for linkage at both TCO and NMTC, with LOD scores of 1.56 and 2.85, respectively. Two-locus linkage analysis, using a multiplicative risk model for the development of NMTC, achieved a maximum LOD of 3.92, with an LOD of 4.51 when assuming 70% of families were linked, indicating that the segregation in these families is consistent with an interaction model. Most of this evidence came from a large Tyrolean family that singularly achieved a two-locus LOD of 3.21. CONCLUSIONS: These results provide further evidence that susceptibility genes for fNMTC exist at 19p13 and 2q21, and furthermore, raise the possibility that in a subset of fNMTC pedigrees, these loci interact resulting in significantly increased risk of NMTC for patients that carry both susceptibility loci.

[The effect of low-frequency electromagnetic fields on the neuroendocrine system]. / Uticaj niskofrekventnih elektromagnetskih polja na neuroendokrini sistem.

This paper presents literature data about effects of low-intensity variable electromagnetic fields on the neuroendocrine system of experimental animals. We mostly paid attention to electromagnetic fields frequently found in our environment, in technological processes, even in our everyday life. This study shows that the regulatory systems (nervous and endocrine) are extremely sensitive to effects of electromagnetic fields. In regard to structures of the central nervous system hypothalamus shows particularly high sensitivity whereas we can consider a hypothesis that effects of this physical factor may be expected in other systems too. It has been emphasized that the effects of electromagnetic fields on regulatory mechanisms may be connected with primary disturbances on the cellular and subcellular (mitochondrial) level.

Nonreciprocal optical device based on second-harmonic generation.

We propose a novel device that exhibits nonreciprocity in optical reflection or transmission. The device is formed by two frequency-doubling crystals and a suitable dichroic mirror or a filter. The nonreciprocity may exceed 100 and be accompanied by nonlinear reflection or transmission. Applications in unidirectional and mode-locked lasers are discussed.

Mode locking of a Nd:YAlO(3) laser at the 1.34-microm transition by a second-harmonic nonlinear mirror.

A second-harmonic nonlinear mirror formed by a LiIO(3) frequency doubler and a dichroic output mirror is employed to mode lock the 1.34-microm transition of a pulsed Nd:YAlO(3) laser. Pulses of 15-ps duration are reliably generated simultaneously at 1.34 and 0.67 microm.

Frequency-domain analysis of the mode-locking process in a laser with a second-harmonic nonlinear mirror.

We present a computer simulation of the mode-locking process in a pulsed Nd:YAG laser with a nonlinear mirror based on second-harmonic generation. For the first time to our knowledge, a frequency-domain description of the passive mode-locking process is presented, which provides complete information for the amplitude and phase evolution of the individual modes. The mode locking is described as being due to the interaction of the longitudinal modes in the nonlinear mirror through second-harmonic and sum- and difference-frequency generation.

Frequency-doubling mode locker: the influence of group-velocity mismatch.

The influence of the group-velocity mismatch between the fundamental and the second harmonic in the frequency-doubling nonlinear mirror is investigated theoretically. The nonstationary analysis provides an estimation for the maximum pulse-shortening capabilities of the device when it is used as a passive mode locker. The pulse-shape deformation is also investigated.

Use of a vacuum-planar photodiode to drive an electro-optic Q switch directly.

A vacuum photodiode was used to drive a Pockels cell directly in an optical-feedback arrangement. This technique was used to achieve Q switching and a single-longitudinal-mode operation in a flash-lamp-pumped Nd:YAG laser. Synchronization within 2 ns with an external short-pulse laser source was demonstrated.

Methods of passive and active mode locking using intracavity optical frequency mixing.

The mode-locking technique based on intracavity second-harmonic generation is extended to sum- and difference-frequency generation. Simultaneous passive mode locking of two different types of laser in a combined cavity is thus possible. By using the same principle, a laser may be actively mode locked by injection of a pulse train from another mode-locked laser.

Mode locking by a frequency-doubling crystal: generation of transform-limited ultrashort light pulses.

Transform-limited ultrashort green and infrared light pulses have been generated from a Nd:YAG laser mode locked by an intracavity second-harmonic crystal in combination with a dichroic output coupler. The mode-locking process is based on substantial reconversion of the second-harmonic radiation back into fundamental radiation after reflection by the dichroic mirror. With this technique light pulses as short as 45 psec have been generated.

Pulse shortening by a nonlinear mirror mode locker.

A device, composed of a nonlinear crystal and a dichroic mirror, features intensity-dependent reflection and may be used as a passive mode locker. The pulse shortening of a Gaussian light pulse due to reflection by this nonlinear mirror has been calculated and compared to that of a saturable absorber used for mode locking.