Detection of Toxoplasma gondii B1 Gene and IgM in IgG Seropositive Pregnant Women.

BACKGROUND: The onset of acute toxoplasmosis in pregnant women may pose a risk to their growing fetuses. The timely diagnosis of infection in managing the disease and preventing its harmful consequences on the fetus is very important. Therefore, the study was conducted to identify acute toxoplasmosis in the pregnant women by detecting the specific IgM antibody and Toxoplasma gondii B1 gene. METHODS: A total of 653 serum samples of women who attended to Fatemieh Hospital of Hamadan University of Medical Sciences were tested for IgG antibodies against Toxoplasma gondii by enzyme-linked immunosorbent as-say (ELISA). The IgG positive specimens were further examined for IgM by ELISA and polymerase chain reaction (PCR) for B1 gene. In the second phase, change in IgG titers was evaluated on 47 IgG positive samples after two weeks. RESULTS: ELISA data showed that 167 out of 653 and 2 out of 167 samples were positive for IgG (25.6%) and IgM (1.2%), respectively. However, PCR detection showed that 36 cases (21.6%) were positive for the B1 gene. Seven out of 47 IgG positive samples showed an increase in the antibody titer and positive for the B1 gene. The most cases of IgG positives and the B1 gene samples were associated with the third trimester of pregnancy with 49.7% and 14%, respectively, and the most common abundance of the B1 gene was 14.4% in the age group of 26 - 35. The most commonly reported clinical symptoms in the B1 gene-positive women were nausea 15 (41.7%), cough 13 (36.1%), headache 12 (33.3%), and vomiting 11 (30.5%). CONCLUSIONS: Using PCR and the B1 gene in serum samples of pregnant women to detect acute toxoplasmosis is a more appropriate and accurate method than IgM antibody.

Garcinol exhibits anti-proliferative activities by targeting microsomal prostaglandin E synthase-1 in human colon cancer cells.

BACKGROUND: Colorectal cancer is the fourth leading cause of death. Various natural compounds are known to have antitumor properties. Garcinol, a polyisoprenylated benzophenone, has antioxidant and anti-inflammatory properties. In the current study, we investigated the anticancer activity of garcinol on human colorectal adenocarcinoma cell line (HT-29) human colon cancer cells. METHODS: HT-29 cells were treated with various concentrations of garcinol for 24 h. The effect of garcinol on HT-29 cells proliferation was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay; the mRNA expression of microsomal prostaglandin E synthase-1 (mPGES-1), hypoxia-inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF), C-X-C chemokine receptor type 4 (CXCR4), matrix metalloproteinase-2 (MMP-2), and matrix metalloproteinase-9 (MMP-9) were examined by quantitative real-time polymerase chain reaction; apoptosis was detected by proportion of sub-G1 cell; caspase 3 activity and prostaglandin E2 (PGE2) level were determined by enzyme-linked immunosorbent assay and HT-29 cells migration was assessed using scratch test. RESULTS: Garcinol preconditioning markedly decreased the expression of mPGES-1, HIF-1α, VEGF, CXCR4, MMP-2, and MMP-9. The proportion of cells in sub-G1 phase and caspase 3 activity were increased by garcinol treatment whereas the cell proliferation, PGE2 level, and cell migration were decreased in these cells, compared to the control group. CONCLUSION: Our findings suggest that garcinol plays a critical role in elevating apoptosis and inhibiting HT-29 cells proliferation, angiogenesis, and invasion by suppressing the mPGES-1/PGE2/HIF-1α signaling pathways.

All-trans retinoic acid preconditioning enhances proliferation, angiogenesis and migration of mesenchymal stem cell in vitro and enhances wound repair in vivo.

OBJECTIVES: Stem cell therapy is considered to be a suitable alternative in treatment of a number of diseases. However, there are challenges in their clinical application in cell therapy, such as to reduce survival and loss of transplanted stem cells. It seems that chemical and pharmacological preconditioning enhances their therapeutic efficacy. In this study, we investigated effects of all-trans retinoic acid (ATRA) on survival, angiogenesis and migration of mesenchymal stem cells (MSCs) in vitro and in a wound-healing model. MATERIALS AND METHODS: MSCs were treated with a variety of concentrations of ATRA, and mRNA expression of cyclo-oxygenase-2 (COX-2), hypoxia-inducible factor-1 (HIF-1), C-X-C chemokine receptor type 4 (CXCR4), C-C chemokine receptor type 2 (CCR2), vascular endothelial growth factor (VEGF), angiopoietin-2 (Ang-2) and Ang-4 were examined by qRT-PCR. Prostaglandin E2 (PGE2) levels were measured using an ELISA kit and MSC angiogenic potential was evaluated using three-dimensional tube formation assay. Finally, benefit of ATRA-treated MSCs in wound healing was determined with a rat excisional wound model. RESULTS: In ATRA-treated MSCs, expressions of COX-2, HIF-1, CXCR4, CCR2, VEGF, Ang-2 and Ang-4 increased compared to control groups. Overexpression of the related genes was reversed by celecoxib, a selective COX-2 inhibitor. Tube formation and in vivo wound healing of ATRA-treated MSCs were also significantly enhanced compared to untreated MSCs. CONCLUSION: Pre-conditioning of MSCs with ATRA increased efficacy of cell therapy by activation of survival signalling pathways, trophic factors and release of pro-angiogenic molecules.

Evidence for decreased expression of APPL1 associated with reduced insulin and adiponectin receptors expression in PCOS patients.

PURPOSE: To investigate the expression of Adaptor protein containing a PH domain, PTB domain and leucine zipper motif 1 (APPL1), insulin receptor (INSR), adiponectin and adiponectin receptors (adipoR1 and R2) and their possible associations in granulosa cells (GCs) of 22 polycystic ovary syndrome (PCOS) women compared to the 22 non-PCOS controls with normal ovulatory function matched for BMI (body mass index). METHODS: In this study, 44 infertile women aged 18-40 years undergoing in vitro fertilization (IVF) protocol were recruited. After follicular fluid collection, GCs were isolated and then purified with MACS (Micro Beads conjugated to monoclonal anti-human CD45 antibodies). RNA was extracted from GCs and quantitative real-time PCR (qRT-PCR) was performed to assess APPL1 gene expression. RESULTS: Expression of APPL1, insulin receptor and adiponectin system genes was significantly decreased in PCOS group compared to the controls. CONCLUSIONS: Reduction of APPL1, insulin receptor and adiponectin system genes in GCs could be involved in the development of PCOS.

Expression of interleukins 7 & 8 in peripheral blood mononuclear cells from patients with metabolic syndrome: a preliminary study.

BACKGROUND & OBJECTIVES: Metabolic syndrome (MS) is a common but serious public health problem in developed countries. Chronic inflammation plays a key role in MS. Interleukins (IL)-7 and 8 are considered to have proinflammatory effects and may be involved in the pathogenesis of MS. Therefore, the aim of this study was to determine gene expression level of IL-7 and IL-8 in peripheral blood mononuclear cells (PBMCs) of patients with MS compared to healthy control subjects. METHODS: Using real-time RT-PCR, the relative amounts of IL-7 and IL-8 mRNA were determined in PBMCs from 20 female patients with MS and compared with those of 20 healthy control subjects. Biochemical and anthropometric parameters of MS were also assessed. RESULTS: Total cholesterol, triglyceride, and fasting blood sugar were significantly higher in MS patients compared to healthy subjects. There were no significant differences in HDLc and LDLc between the two groups. IL-8 expression in PBMC was significantly decreased in MS versus control subjects (fold of change was 0.395±0.1824), while no difference in the IL-7 expression was detected between them. IL-8 expression had negative correlation with MS components especially with triglyceride and total cholesterol (r=0.5, P<0.001). INTERPRETATION & CONCLUSIONS: In this preliminary study, no detectable differences were found in IL-7 expression and decreased expression of IL-8 in PBMCs of MS patients as compared to those of control subjects. Study on a larger population and investigating the mechanisms involved can reveal more details.

Cholesteryl ester transfer protein (CETP) -629C/A polymorphism and it's effects on the serum lipid levels in metabolic syndrome patients.

Metabolic syndrome is a relatively common disorder with significant morbidity worldwide. Cholesteryl ester transfer protein (CETP) plays a central role in the metabolism of lipoproteins. In this study the effect of -629C/A polymorphism on the concentration of CETP and plasma lipids pattern was elicited in metabolic syndrome patients and control subjects. For this, a sample of 200 patients diagnosed with metabolic syndrome disorder was studied in comparison with 200 healthy controls. This study was performed by using polymerase chain reaction and restriction fragment length polymorphisms. Genotype distribution and allelic frequencies were determined and compared in metabolic syndrome and healthy controls. To determine the relationship between -629C/A polymorphism and lipid levels, lipids and CETP concentration were measured in metabolic syndrome and normal subjects. The results showed a significant difference between two groups in terms of FBS, cholesterol, TG, HDL-C, LDL-C levels as well as BMI, waist circumference, systolic and diastolic blood pressure. The genotype frequencies for this polymorphism differed significantly between metabolic syndrome patients and controls (in control group: CC% 20.5, CA% 76, AA% 3.5 and in patient group: CC% 28.5, CA% 53.5, AA% 18) (p < 0.05) while there was no significant difference in the frequency of the alleles. In the two groups, the levels of the cholesteryl ester transfer protein in AA genotype were lower than other genotypes. In the control group, individuals with AA genotype had the highest levels of LDL-C and TC plasma concentration. Considering the results of this study, it can be concluded that the -629 AA genotype was associated with high cholesterol; high LDL-C and low CETP level, so that it can be related to metabolic syndrome.

Active membrane transport and receptor proteins from bacteria.

A general strategy for the expression of bacterial membrane transport and receptor genes in Escherichia coli is described. Expression is amplified so that the encoded proteins comprise 5-35% of E. coli inner membrane protein. Depending upon their topology, proteins are produced with RGSH6 or a Strep tag at the C-terminus. These enable purification in mg quantities for crystallization and NMR studies. Examples of one nutrient uptake and one multidrug extrusion protein from Helicobacter pylori are described. This strategy is successful for membrane proteins from H. pylori, E. coli, Enterococcus faecalis, Bacillus subtilis, Staphylococcus aureus, Microbacterium liquefaciens, Brucella abortus, Brucella melitensis, Campylobacter jejuni, Neisseria meningitides, Streptomyces coelicolor and Rhodobacter sphaeroides.