Role of rs7903146 polymorphism and adropin serum level in patients with diabetes mellitus; a case-control study from Isfahan, Iran.

BACKGROUND: Type-2 diabetes mellitus (T2DM) is the common endocrinopathy which characterised by insulin resistance, insufficient expression or secretion of insulin and decrement of insulin effectiveness. Although T2DM has unknown aetiology, the strongest susceptible gene in this disease is TCF7L2. Adropin peptide may have roles in T2DM pathogenesis due to several roles in glucose tolerance, decrement of insulin resistance, lipid metabolism and energy homoeostasis. AIM: To evaluate the serum level of adropin in T2DM patients and comparing with healthy individuals as well as assessing frequency of rs7903146 genotypes/alleles in patients and control groups. METHODS: We analysed the frequency of rs7903146 genotypes/alleles in 93 patients with T2DM disease and 53 healthy individuals by the method of polymerase chain reaction-restriction fragment length polymorphism analysis. The serum level of adropin was measured by using enzyme-linked immunosorbent assay technique. RESULTS: The mean serum level of adropin was 12.32 ± 2.98 and 9.51 ± 2.73 in patients and control groups, respectively (p value < .001). Also, there were significant difference in frequency of genotypes and alleles of rs7903146 in patients and controls groups (p < .001). The rs7903146T/T and rs7903146C/T genotypes increased risk of T2DM disease (OR: 6.035 and OR: 3.082, respectively). Interestingly, the highest level of adropin was detected in T2DMpatients with rs7903146T/T genotype. CONCLUSION: Our analysis showed higher level of adropin in T2DM patients and increased risk of T2DM with rs7903146T/T and rs7903146C/T genotypes.

Glucosamine attenuates drug resistance in Mitoxantrone-resistance breast cancer cells.

OBJECTIVES: This study was aimed at investigating the cytotoxicity and multi-drug resistance (MDR) reversal effect of Glucosamine (GlcN) on resistant BCRP-overexpressing breast cancer MCF-7/MX cells. METHODS: After confirming the overexpression of BCRP, the cytotoxicity and MDR reversing potential of GlcN on MCF-7/MX mitoxantrone-resistant and MCF-7 sensitive breast cancer cells were assessed via MTT assay. The effects of GlcN on mitoxantrone accumulation were analyzed through flow cytometry. Finally, the expression of BCRP and Epithelial-Mesenchymal Transition (EMT)-related markers following the exposure to GlcN were assessed by real-time RT-PCR. KEY FINDINGS: This study showed that glucosamine had an inhibitory effect on the proliferation of human breast cancer cells. The respective IC50 values for MCF-7/MX cells following exposure to mitoxantrone (MX) in the presence of GlcN (0, 0.5 and 1 mm) for 72 h were 3.61 ± 0.21, 0.598 ± 0.041 and 0.284 ± 0.016 µm, respectively. Furthermore, GlcN reduced the expression of BCRP mRNA without any significant effect on EMT-related markers in breast cancer cells. CONCLUSIONS: These results proposed that glucosamine as a natural sugar could down regulate the BCRP expression and increased MX cytotoxicity in breast cancer cells.

Single-Strand DNA-Like Oligonucleotide Aptamer Against Proprotein Convertase Subtilisin/Kexin 9 Using CE-SELEX: PCSK9 Targeting Selection.

BACKGROUND: Proprotein convertase subtilisin/kexin 9 (PCSK9) serves a key regulatory function in the metabolism of low-density lipoprotein (LDL)-cholesterol (LDL-C) through interaction with the LDL receptor (LDLR) followed by its destruction that results in the elevation of the plasma levels of LDL-C. The aims of the present study were to separate and select a number of single-stranded DNA (ssDNA) aptamers against PCSK9 from a library pool (n > 1012) followed by their characterization. METHODS: The aptamers obtained from the DNA-PCSK9 complexes which presented the highest affinity against PCSK9 were separated and selected using capillary electrophoresis evolution of ligands by exponential enrichment (CE-SELEX). The selected aptamers were amplified and cloned into a T/A vector. The plasmids from the positive clones were extracted and sequenced. The Mfold web server was used to predict the secondary structure of the aptamers. RESULTS: Following three rounds of CE-SELEX, the identified anti-PCSK9 ssDNA aptamers, namely aptamer 1 (AP-1) and aptamer 2 (AP-2), presented half maximal inhibitory concentrations of 325 and 327 nM, lowest dissociation constants of 294 and 323 nM, and most negative Gibbs free energy values of - 9.17 and - 8.28 kcal/mol, respectively. CONCLUSION: The results indicated that the selected aptamers (AP-1 and AP-2) induced potent inhibitory effects against PCSK9. Further in vivo studies demand to find out AP-1 and AP-2 aptamers as suitable candidates, instead of antibodies, for using in therapeutic purposes in patients with hypercholesterolemia and cardiovascular disease.

Glucosamine Reverses P-Glycoprotein-Mediated Multidrug Resistance in the Daunorubicin-Resistant Human Gastric Cancer Cells.

Glucosamine (GlcN) is a natural amino monosaccharide in the human body, and evidence of its anticancer effects is growing. In this study, we aimed to evaluate the effects of GlcN for its cytotoxicity, MDR reversal effects and inhibitory effects on function and expression of P-glycoprotein (P-gp) transporter in the daunorubicin-resistant human gastric cancer cells. Cell viability was measured by MTT assay to evaluate the cytotoxicity and multidrug resistance (MDR) reversal effects of GlcN. The effects of GlcN on function and mRNA expression level of P-gp transporter were assessed by flow cytometry and real-time RT-qPCR, respectively. Our results indicated that GlcN reduced the proliferation of human gastric cancer cell line EPG85-257 and its drug-resistant variant EPG85-257RD in a dose-dependent manner. GlcN (at the concentrations of 0.5 and 1 mM) also enhanced the sensitivity of EPG85-257RDB cells to daunorubicin. The cellular accumulation studies showed that GlcN inhibited efflux activity of P-gp and enhanced the mean fluorescent intensity of Rho123 while ˙it had no effects on P-gp gene expression in these cells. This study suggested that the inhibition of P-gp activity is a novel mechanism of action by which GlcN could reverse MDR in EPG85-257RDB cells.

Glucosamine reverses drug resistance in MRP2 overexpressing ovarian cancer cells.

Glucosamine (GlcN), a natural amino sugar in human body, was reported to exhibit anticancer activity against some tumors. In the present study, we evaluated the cytotoxicity and multi-drug resistance (MDR) reversal activity of GlcN on resistant MRP2-overexpressing ovarian cancer A2780RCIS cells. The cytotoxicity and MDR reversal activity of GlcN on cancer cells were measured by MTT assay. The effects of GlcN on MRP1 and MRP2 mRNA expression and function were evaluated by qRT-PCR and flow cytometry, respectively. The cell migration capacity of ovarian cancer cells were assessed in the presence or absence of GlcN using wound healing migration assay. Furthermore, the effects of GlcN on the mRNA expression of E-cadherin, vimentin and α-smooth muscle actin as Epithelial-Mesenchymal Transition (EMT)-related markers were evaluated by qRT-PCR. Our results indicated that glucosamine reduced the proliferation of human ovarian cancer cell lines (A2780) and its cisplatin resistant variant (A2780RCIS) in a dose-dependent manner. The IC50 values for A2780RCIS cells treated with cisplatin in the presence of different concentrations of GlcN (0, 1, 2 and 3 mM) for 72 h were 44.463 ± 1.603, 35.17 ± 0.025, 22.25 ± 0.018, 17.78 ± 0.012 µM respectively. Also GlcN decreased the expression of MRP1 and MRP2 mRNA in ovarian cancer cells. Our results further demonstrated that although GlcN had no significant effects on the expression of studied EMT-related markers in invasive A2780RCIS cells, it was able to inhibit their migration in vitro. According to these findings, GlcN could effectively enhance cisplatin cytotoxicity in resistant A2780RCIS cells.

An investigation of the association between the level of prolactin in serum and type II diabetes.

As a hormone secreted from the pituitary gland, prolactin (PRL) plays an important role in increasing beta cell proliferation, stimulating the secretion of insulin, preventing the activities of caspases on pathways that cause apoptosis in the Langerhans' islands, and moderating the immune system in regulating the whole body's sensitivity to insulin. Therefore, PRL level changes in type II diabetes and it can be concluded that PRL can play an important role in metabolic disorders of glucose. The present study is carried out in order to investigate the association between serum levels of PRL and type II DM. Blood samples were taken from 64 females affected by type II diabetes and 70 healthy ones, whose PRL level was measured using electrochemiluminescence (ECL) technique. It was a case-control study, and based on the definition dedicated to each group, subjects were assigned to two groups. The patient group included the subjects with type II diabetes while the control group included healthy samples. Data were analyzed using SPSS software (Mann-Whitney test, t-test, and spearman's rho correlation test). According to the results, PRL concentration in the serum of people affected by type II diabetes (5.32 ±â€¯0.36) was significantly (P˂0.05) lower than that of control group (18.38 ±â€¯2.3). The results also showed that in type II diabetes, the level of PRL changes so that the concentration of PRL in the serum of the patients was lower than that of healthy ones. Therefore, PRL concentration in the blood can be related to diabetes.

Selection and Characterization of Single-Stranded DNA Aptamers Binding Human B-Cell Surface Protein CD20 by Cell-SELEX.

The B-lymphocyte antigen (CD20) is a suitable target for single-stranded (ss) nucleic acid oligomer (aptamers). The aim of study was selection and characterization of a ssDNA aptamer against CD20 using Cell-Systematic Evolution of Ligands by Exponential Enrichment (Cell-SELEX). The cDNA clone of CD20 (pcDNA-CD20) was transfected to human embryonic kidney (HEK293T) cells. Ten rounds of Cell-SELEX was performed on recombinant HEK-CD20 cells. The final eluted ssDNA pool was amplified and ligated in T/A vector for cloning. The plasmids of positive clones were extracted, sequenced and the secondary structures of the aptamers predicted using DNAMAN® software. The sequencing results revealed 10 different types; three of them had the highest thermodynamic stability, named AP-1, AP-2 and AP-3. The AP-1 aptamer was the most thermodynamically stable one (ΔGAP-1 = -10.87 kcal/mol) with the highest binding affinity to CD20 (96.91 ± 4.5 nM). Since, the CD20 is a suitable target for recognition of B-Cell. The selected aptamers could be comparable to antibodies with many advantages. The AP-1, AP-2 and AP-3 could be candidate instead of antibodies for diagnostic and therapeutic applications in immune deficiency, autoimmune diseases, leukemia and lymphoma.

Transcription factor 7-like 2 polymorphism and context-specific risk of metabolic syndrome, type 2 diabetes, and dyslipidemia.

BACKGROUND: The transcription factor 7-like 2 gene (TCF7L2) is an element of the Wnt signaling pathway. There is lack of evidence if TCF7L2 has a functional role in lipid metabolism and regulation of the components constitutes the metabolic syndrome (MetSyn). The aims of this study were to evaluate whether the risk allele of TCF7L2 gene polymorphism is associated with dyslipidemia and MetSyn. MATERIALS AND METHODS: The MetSyn subjects were participated only based on the National Cholesterol Education Program - Third Adult Treatment Panel criteria. In this case-control study, the DNA from MetSyn patients without (n = 90) and with type 2 diabetes (T2D) (n = 94) were genotyped. RESULTS: The results show that the genotype-phenotype for CC, CT/TT of TCF7L2 gene polymorphism correlated with body mass index and waist circumference in MetSyn and MetSyn + T2D subjects (r = -0.949 and r = -0.963, respectively). The subjects that only possess MetSyn but are not diabetics show the 2 h postprandial glucose and fasting blood glucose, glycated hemoglobin significantly lower (P < 0.05) than those subjects have both abnormality. The level of triglyceride in CT/TT carriers in MetSyn was higher than CC carriers (P = 0.025). A comparison with the controls subjects, the frequencies of the T allele in the groups of MetSyn (46.66%) and MetSyn + T2D (47.34%) show significantly different (P < 0.05). The odds ratios for T allele in (MetSyn)/(normal), (MetSyn + T2D)/(normal), and in (MetSyn + T2D)/(MetSyn) were 3.59 (95% confidence interval [CI], 1.33-9.67, P = 0.0093), 3.76 (95% CI, 1.40-10.07, P = 0.0068), and 1.08 (95% CI: 0.55- 2.11, P = 0.834), respectively. CONCLUSION: The results revealed the important insights essential for the role of TCF7L2 that the T allele of TCF7L2 plays a significant role in the susceptibility to dyslipidemia, MetSyn, and T2D.

Synthesis and evaluation of the complex-forming ability of hydroxypyranones and hydroxypyridinones with Ni (II) as possible inhibitors for urease enzyme in Helicobacter pylori.

The complex-forming ability of 2-methyl-3-hydroxypyran-4-one (1a), 2-ethyl-3-hydroxypyran-4-one (1b), 1,2-dimethyl-3-hydroxypyridin-4-one (4a) and 1-ethyl-2-methyl-3-hydroxypyridin-4-one (4b) with nickel(Ni(II)) were characterized by infrared, ultraviolet, proton nuclear magnetic resonance spectroscopy and melting point. The mole-ratio of nickel:ligands was analyzed by atomic-absorption-spectrometry. The partition-coefficients (KOW) of the compounds were also determined. The binding of ligands with Ni(II) are through deprotonated hydroxyl group (-O(-), disapeared at 3259 cm(-1)) and ioan-pairs of carbonyl group (=CO(.), shifted from 1650 to 1510-1515 cm(-1)). The characterization of complex geometry for bis-(2-methyl-3-hydroxypyranonato)Ni(II) (5a) and bis-(2-ethyl-3-hydroxypyranonato)Ni(II) (5b) predicted to be square-planer while for bis-(1,2-dimethyl-3-hydroxypyridinonato)Ni(II) (5c) and bis-(1-ethyl-2-methyl-3-hydroxypyridinonato)Ni(II) (5d) distorted to tetrahedral-geometry. Inhibitors of Helicobacter pylori urease are nickel chelators. The compounds 1a, 4a and 4b are likely suitable ligands with complex forming-ability to make complexes of 5a, 5c and 5d with nickel. The KOW values show the compound 5c with low partition-coefficient is more suitable ligand with lower penetration from GI lumen. Future studies demand to find out the biological activity of developed compounds on H. pylori.

Preliminary study on patients located at the Kashani/Isfahan Hospital with multiple sclerosis between the years 2011 and 2013.

BACKGROUND: Multiple sclerosis (MS) is a multifactorial disease that could result from demyelination of the myelin sheath. The aim of this study is to investigate the demographic features and rank the immunomodulating drugs in patients with MS. MATERIALS AND METHODS: This study was conducted in the MS clinic of the Isfahan Kashani Hospital, from 22 May, 2011 to 18 March, 2013. The data analyses (n = 1067) were divided into two periods: (1) 2011/05/22 to 2012/03/18 denoted as P1 and (2) 2012/04/02 to 2013/03/18 denoted as P2. RESULTS: Most of drugs prescribed within the population studied were: Avonex, Betaferone, and Rebif. There was an increase in the number of female (n = 811) and male subjects (n = 256). During P1/P2 there was an increase from 460 to 607 in the total number of patients, respectively. The number of patients who attended the MS clinic once was 250 (P1) versus 430 (P2), and those more than four times was 71 (P1) versus 59 (P2) correspondingly. CONCLUSION: The number of females increased from 2011 to 2013. Because of dissimilar ingredients additive of different pharmaceutical companies, it could be suggested that pharmacotherapy strategies, especially in Iranian population of MS with first-line treatment using Avonex, Betaferone and Rebif, more spotlighted on inter- and intra-individual variability based on clinical pharmacokinetics parameters.

Quantitative analysis of the nutritional components in leaves and seeds of the Persian Moringa peregrina (Forssk.) Fiori.

BACKGROUND: Moringa peregrina (M. peregrina) is an important tropical tree recognized for its nutritional and medicinal properties. The objective of this study was to investigate the nutritional component in the leaves and seeds of the Persian M. peregrina (Forssk.) Fiori. MATERIALS AND METHODS: The M. peregrina leaves and seeds of wild cultivated trees were collected from the areas of arid environment located in the South-East of the Iran. The leaves and seeds of M. peregrina were dried and grounded to a fine powder and kept in dark for the day of experiment. The acidic digested leaves and seeds were analyzed for Vitamins C and A, calcium, and potassium using atomic adsorption and flame emission spectrophotometer. RESULTS: The analytical data revealed that the leaves and seeds of the Persian M. peregrina (Forssk.) Fiori contain sufficient amounts of Vitamin C: 83 ± 0.5 and 14 ± 0.6 mg/100 g/DW; and Vitamin A: 6.8 ± 0.7 and 24.8 ± 0.7 mg/100 g/DW, respectively. The elemental analysis in the leaves and seeds showed that the calcium content are 764.8 ± 1.6 and 1164.8 ± 43.4 mg/100 g/DW and for potassium content are 900.2 ± 14 and 572 ± 10 mg/100 g/DW, respectively. CONCLUSIONS: The nutritional characteristics of the Persian M. peregrina (Forssk.) Fiori, investigated in this study revealed that, daily use of leaves and seeds of this plant could significantly provide the recommended dietary allowance for the Vitamins C and A, and minerals, such as calcium and potassium.

The investigation of correlation between Iminoral concentration and neurotoxic levels after kidney transplantation.

BACKGROUND: Neurotoxicity side effects related to cyclosporine kinetics could lead to dysfunction of kidney graft and patient outcome after transplantation. The aim of this study was evidence-based pharmacotherapy of kidney transplant recipients and to investigate neurotoxic levels of Iminoral. MATERIALS AND METHODS: The results of 2239 cyclosporine trough levels obtained from 743 patients were studied. Seventy-five adult kidney recipients who received Iminoral were studied for neurotoxicity symptoms. Demographic, clinical, hematology and biochemical data were recorded in d-base and analyzed using SPSS application for windows. RESULTS: The mean value related to cyclosporine C0 was 246.3 µg/l. In the 48% the signs of neurotoxicity such as tremor and headache were noted, but only in 9% the levels of cyclosporine C0 were >400 µg/l. Further studies on 75 patients showed that the incidence of neurotoxic side effects were as follows: Tremor in 35, headache in 24 and anxiety in 34 recipients of kidney. The prescribed drug regimens from the day of transplant in most patients were based on mycophenolic acid or cellcept, pulse therapy using methylprednisolone (daily from kidney transplant up to 3 days after transplant), cyclosporine or Iminoral plus other drugs related to each individual. Administrations of ganciclovir, thymoglobulin, clotrimazol and prednisolone were also distinguished with immunosuppressant-based therapy simultaneously. CONCLUSION: Evidence-based study related to pharmacotherapy of Iminoral showed that clinical presentation related to neurotoxic side effects such as tremor, headache and anxiety might be due to many factors such as polypharmacy. Planning immunosuppression to individual patients based on programmed therapeutic Iminoral monitoring, avoiding polypharmacy in terms of removal or drug minimization and focusing on first week after transplant seem to be a realistic option.

Association of a common variant in TCF7L2 gene with type 2 diabetes mellitus in a Persian population.

Diabetes is one of the most common and challenging health problems. Studies in several nations show that polymorphisms within the transcription factor 7-like 2 genes could be associated with type 2 diabetes (T2D). Therefore, a case-control study was conducted to find the association between SNP rs7903146 and T2D in our population. The study consists of 110 patients referring to clinic and 80 healthy controls randomly selected based on WHO guideline. DNA was extracted from blood and genotyped by PCR-RFLP with specific primers to amplify a fragment for restriction enzyme (RsaI). A chi-square test was calculated to compare the proportions of genotypes or alleles. Using a logistic regression model, the odds ratio for risk of developing T2D was calculated with and without adjustment for age, sex, and BMI. The frequency of the T allele of rs7903146 (C/T) polymorphism was significantly higher in diabetic subjects (47.3%) compared to that in normal subjects (34.4%). Logistic regression analysis of the rs7903146 polymorphism showed that the odds ratio was 3.71(95% CI: 1.43-9.56; P: 0.008) for the TT genotype and 1.26 (95% CI: 0.67-2.39; P: 0.516) for the CT genotype when compared with the CC genotype. Odds ratio adjusted for age, sex, and BMI have shown similar results. The results show that rs7903146 of TCF7L2 gene is an important susceptibility gene for T2D mellitus in the province of Isfahan, Iran. Our results support the recent findings that rs7903146 of TCF7L2 gene is an important genetic risk factor for the development of T2D in multiple ethnic groups.

Gene delivery to brain cells with apoprotein E derived peptide conjugated to polylysine (apoEdp-PLL).

A promising strategy to carry genetic material to brain cells either in vitro or in vivo is using the LDL receptor (LDLr) on blood-brain barrier. LDLr naturally help to low density lipoproteins (LDL(S)) transporting across the BBB by endocytosis. Here we present the idea of using the LDLr-mediated pathway for transporting genetic material to brain cells. A tandem dimer Sequence of apoprotein-E (apoE) (141-150) conjugated to polylysine sequence was used as a novel DNA Delivery vector for transfecting of brain cells either in vitro or in vivo. DNA condensation occurs with this vector because electrostatic interaction between DNA and polylysine. The vector favors to protection of DNA from enzymatic degradation and also helps to DNA carrying in blood stream to reach BBB and transport it to brain cells and eventually help DNA expression in target cells. These results suggest a novel gene delivery vector for gene therapy of brain disease.