Effect of resveratrol on SNARE proteins expression and insulin resistance in skeletal muscle of diabetic rats.

OBJECTIVES: Soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex proteins are involved in membrane trafficking. The expression of isoforms of SNAP-23, syntaxin-4, and VAMP-2 is significantly done in skeletal muscles; they control GLUT4 trafficking. It is believed that type 2 diabetes could be caused by the modifications in the expression of SNARE complex proteins. The purpose of this study was to evaluate the effect of resveratrol on the expression of these proteins in type 2 diabetes. MATERIALS AND METHODS: Forty male Wistar rats were selected. Streptozotocin and nicotinamide were applied for the induction of type 2 diabetes. The animals were divided into five groups. Healthy and diabetic groups were set as control; resveratrol (1, 5, and 10 mg/kg body weight) was applied to treat the three groups of diabetic rats for 30 days. Real-time qRT-PCR was applied to evaluate the expression of SNARE complex proteins. RESULTS: There is a link between diabetes and insulin resistance and up-regulation of SNARE proteins expression. Resveratrol improved hyperglycemia and insulin resistance along with a non-significant reduction in the expression of SNARE proteins. CONCLUSION: Increased expression of SNARE proteins was possibly a compensatory mechanism in response to insulin resistance in the skeletal muscles of diabetic rats. Resveratrol non-significantly reduced the expression of SNARE proteins by enhancing insulin sensitivity, where this effect was dose-dependent. Thus, higher doses of resveratrol and longer intervention periods could probably be more effective. Another molecular mechanism of the anti-diabetic properties of resveratrol was identified with an effect on the expression of SNARE proteins.

Through oxaliplatin resistance induction in colorectal cancer cells, increasing ABCB1 level accompanies decreasing level of miR-302c-5p, miR-3664-5p and miR-129-5p.

Oxaliplatin as a component of (Neo-) adjuvant chemotherapeutic regimens is administered to colorectal cancer patients. Unfortunately, the acquisition of resistance to this drug in nearly 90% of metastatic patients rendered it as an ineffective drug. Therefore, resistance mechanisms to this drug should be elucidated. There are different genes like GSTP1 and ABCB1 which are responsible for oxaliplatin resistance. We hypothesized that miR-129-5p, miR-302c-5p, miR-3664-5p, mir-3714 and miR-513a-3p are targeting ABCB1 gene, while GSTP1 was predicted to be the potential target of miR-3664-5p, mir-3714 and miR-513a-3p. In order to study this hypothesis, resistant colorectal cell lines were generated through intermittent exposure of HCT116, SW480 and HT29 to the increasing doses of oxaliplatin. MTT assays validated this resistance induction. Expression of ABCB1 and GSTP1 in addition to their targeting miRNAs in different cell lines were studied by quantitative real time PCR in the cell lines. Even though in comparison with HCT116 and SW480 cell lines, GSTP1 expression was reduced in resistant cells, ABCB1 expression was upregulated in these cell lines. On the other hand, HT-29 resistant cells showed elevated GSTP1 and unchanged ABCB1 levels. While miR-302c-5p level was downregulated in resistant cell lines, miR-129-5p and miR-3664-5p level showed different pattern of reduction in the resistant SW480 and HCT116 cell lines. GSTP1 level was correlated directly with miR-513a-3p and miR-3664-5p in all SW480 and HCT116 derived cell lines, however in HT-29-OXR1, GSTP1 level was correlated inversely with miR-3664-5p. In conclusion, upregulation of ABCB1 can be considered as the crucial component of poor response to oxaliplatin which is likely controlled by miR-302c-5p.

Genes Encoding GABA-ß and HT1D Receptors in Bipolar I (Manic Phase) Patients.

INTRODUCTION: According to the cumulative evidence, genes encoding GABA receptors inhibit neurotransmitters in CNS and are intricately involved in the pathogenesis of mood disorders. Based on this hypothesis, these genes may be expressed in bipolar patients. As a result, we evaluated the gene expressions of GABA-ß3 and HT1D receptors to assess their associations with bipolar mood disorder. METHODS: In this study, 22 patients with bipolar I disorder (single manic episode) and 22 healthy individuals were enrolled. All participants were older than 15 years and had referred to Farshchian Hospital, Hamadan, Iran. They were diagnosed based on DSM IV-TR criteria and young mania rating scale in order to determine the severity of mania by a psychiatrist as bipolar Type 1 disorder in manic episode. We evaluated the expression of GABA-ß3 and HT1D receptor genes in peripheral blood mononuclear cells, using real-time RT-PCR analysis. RESULTS: In our study, a reduction in the gene expression of GABA-ß3 and HT1D receptors was observed in peripheral blood mononuclear cells of the patients with bipolar disorders compared to the healthy controls. CONCLUSION: The results of this study supports the hypothesis that the gene expression for serotonin and GABA receptors can be employed in elucidating the pathogenesis of bipolar disorders.

Evaluation of altered expression of miR-9 and miR-106a as an early diagnostic approach in gastric cancer.

BACKGROUND: The role of microRNAs (miRNAs) in cellular processes such as growth, apoptosis, differentiation and proliferation verifies the importance of miRNAs in carcinogenesis. Moreover, levels of miRNAs are dysregulated in cancer cells, so they could be used as novel classes of biomarkers for diagnosing cancer. The oncogenic role of miR-106a and its increased expression have been demonstrated in some cancers. In contrast, there is no consensus for miR-9 expression rate in different cancers. Therefore, this study was done to investigate the role of miR-106a and miR-9 in gastric cancer (GC). METHODS: The current study was performed on 31 GC tissues as case, and 31 healthy adjacent tissues as a control group. Quantitative reverse transcriptase (q-RT) PCR was used for studying the expression rate of both miR-106a and miR-9. RESULTS: The expression rate of both miRNAs in cancerous tissues was significantly higher than healthy adjacent tissues (≈10 folds) (P<0.05). CONCLUSIONS: The results showed that the expression rate of both markers was significantly increased in cancerous tissues. Therefore, they can be suggested as potential biomarkers for cancer diagnosis and prognosis as well as targets for therapy.

Evaluation of miR-22 and miR-20a as diagnostic biomarkers for gastric cancer.

BACKGROUND: Gastric cancer (GC) is the fourth most common cancers and the second reason for cancer-related death around the world, particularly in East Asian countries. Diagnosing GC in its early stages is followed by more successful treatment. Unfortunately, there is no accurate method for GC diagnosis in its early stages. Recently, miRNAs have been investigated in the most cancer researches which have demonstrated that they have been dysregulated in many cancers. METHODS: This case-control study aims to investigate the expression rate of miR-22 and miR-20a in 32 cancerous tissues as well as 32 healthy adjacent tissues. Quantitative reverse transcriptase PCR (q-RT PCR) was used for investigating the expression rate of these miRNAs. RESULTS: The expression rate of miR-20a in cancerous tissues was significantly increased (8.9 times) in comparison with their healthy tissues (P<0.001), while the expression rate of miR-22 in cancerous tissues was significantly decreased (1.9 times) (P<0.05). CONCLUSIONS: The obtained results suggest miR-22 and miR-20a as good diagnostic biomarkers for early detection of GC. However more research is needed to investigate their efficacy.

Evaluation of zinc finger E-box binding homeobox 1 and transforming growth factor-beta2 expression in bladder cancer tissue in comparison with healthy adjacent tissue.

PURPOSE: The fifth most common cancer is allocated to bladder cancer (BC) worldwide. Understanding the molecular mechanisms of BC invasion and metastasis to identify target therapeutic strategies will improve disease survival. So the aim of this study was to measure expression rate of zinc finger E-box binding homeobox 1 (ZEB1) and transforming growth factor-beta2 (TGF-ß2) mRNA in tissue samples of patients with BC and its healthy adjacent tissue samples and their association with muscle invasion, size and grade of the tumor. MATERIALS AND METHODS: Tissue samples were collected from 35 newly diagnosed untreated patients with BC from 2013 to 2014. Total RNA was extracted from about 50-mg tissue samples using TRIzol reagent. TAKARA SYBR Premix EX Tag II was applied to determine the rate of mRNA expression by real-time polymerase chain reaction (PCR). To obtain final validation, PCR product of ZEB1 and TGF-ß2 were sequenced. STATA 11 software was used to analyze the data. RESULTS: The expression level of ZEB1 in tumor samples was significantly more than of in healthy adjacent tissue samples. Up-regulation of TGF-ß2 showed a strong association with muscle invasion (p=0.017). There was also demonstrated a relationship between over expression of ZEB1 with the tumor size (p=0.050). CONCLUSIONS: It looks ZEB1 and TGF-ß2 had a role in BC patients. In this study ZEB1 expression was higher in BC tissues than that of in healthy control tissues. There was demonstrated a markedly association between overexpression of TGF-ß2 and muscle invasion. Therefore, they are supposed to be candidate as potential biomarkers for early detection and progression of BC.

Effect of resveratrol on resistin and apelin gene expressions in adipose tissue of diabetic rats.

BACKGROUND/AIM: Adipose tissue plays a major role in glucose homeostasis. Dietary antioxidants such as resveratrol (RSV) may offer some protection against the early stage of diabetes mellitus and the development of complications. The present study investigated the effects of RSV on biochemical parameters and resistin and apelin gene expressions in adipose tissue of rats with type 2 diabetes. MATERIALS AND METHODS: Diabetes was induced using a single dose of streptozotocin + nicotinamide. Three groups of diabetic rats were treated with different doses of RSV (1, 5, and 10 mg/kg body weight per day). Oxidative status, serum biochemical parameters, insulin, and HOMA index were measured. Finally, resistin and apelin gene expressions were determined in rats' adipose tissue using RT- PCR (qRT-PCR). RESULTS: A significant reduction in serum glucose level was observed in rats treated with 5 and 10 mg/kg per day RSV compared with the diabetic control. Resistin expression in adipose tissue was reduced in RSV-treated groups, while no significant changes were observed in apelin expression. CONCLUSION: The results showed that RSV improves insulin sensitivity due to a simultaneous decrease in blood glucose and an increase in insulin. We concluded that RSV has potential hypoglycemic effect, probably by increasing insulin levels and changing the expression of resistin.

A Pilot Study of CK19, CK20 and GCC mRNA in the Peripheral Blood as a Colorectal Cancer Biomarker Panel.

Colorectal cancer remains one of the major cancer- related deaths despite progress in the treatment during past decades. Detection of disease at earlier stages reduces its mortality. The aim of current study was to investigate expression of Cytokeratin 19 (CK19), Cytokeratin 20 (CK20) and Guanylyl Cyclase C (GCC) mRNA in peripheral blood of non- metastatic colorectal cancer patients which may result into introducing of an early detection test. 25 patients with colorectal cancer and 25 healthy controls were recruited. Blood was obtained from all individuals. Expression of CK19 and CK20 and GCC mRNA and 18SrRNA (as reference gene) were determined based on real- time RT-PCR on total RNA from blood. CK19, CK20 and GCC expression had been detected in 68%, 76% & 52% of patient group, respectively, which was higher than healthy group, with 8%, 32% and 0% expression, respectively (p<0.05). CK20 was over-expressed 8- fold more in patients compared to controls. Similar result was found for CK19 with 4- fold over- expression. Sensitivity and specificity of combination of markers were 88% and 68%, respectively. Current data suggest that the detection of CK20 & CK19 as relative sensitive markers may become a valuable tool for primary diagnosis of colorectal cancer in early stages. GCC could be considered as a specific tumor marker for detection of colorectal cancer. Higher expression of these markers in patients may be considered as a relative good tool for the diagnosis of disease in non- metastatic stages.

Liver Mitochondrial DNA Copy Number and Deletion Levels May Contribute to Nonalcoholic Fatty Liver Disease Susceptibility.

BACKGROUND: There is growing evidence that deficiencies observed in the mitochondrial DNA (mtDNA) functions could play an important role in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). We hypothesized that genetic variations in mtDNA could affect the mitochondrial function and contribute to the NAFLD susceptibility. OBJECTIVES: In this study, the possible association of the mtDNA copy number and 4,977-bp deletion levels with NAFLD susceptibility in a sample of Iranian population was evaluated. METHODS: This case-control study included 43 NAFLD patients and 20 control subjects. Genomic DNA was extracted from fresh liver tissue samples by using a DNA isolation kit. The mtDNA copy number and mtDNA deletion levels were measured by quantitative real-time PCR and multiplex PCR. RESULTS: The relative expression of mtDNA copy number was 3.7 fold higher in NAFLD patients than healthy controls (P < 0.0001). The results remained significant after adjustment for age, BMI, and gender (P = 0.02). In addition, the mtDNA copy number was 4.3 (P < 0.0001) and 3.2-fold (P < 0.0001) higher in nonalcoholic fatty liver (NAFL) and non-alcoholic steatohepatitis (NASH) patients than healthy controls, respectively. Finally, the results showed that the 4,977-bp deletion is not detected in any of liver tissue samples obtained from the 20 control subjects whereas 8 out of 43 NAFLD patients (18.6%) showed the 4,977 -bp deletion in their liver tissues (P = 0.039). CONCLUSIONS: This study indicated an association between mtDNA content in the liver tissue and NAFLD susceptibility that may be a consequence of compensatory response to the cumulative exposures to oxidative damage.

Expression Analysis of mir-21 and mir-221 in Cancerous Tissues from Iranian Patients with Gastric Cancer.

BACKGROUND: Early detection is a key to survival for gastric cancer. Molecular markers such as miRNA (microRNA) can have great importance in the early diagnosis of gastric cancer. Expression of miR-21 and miR-221 are deregulated in many types of human cancers. This study aimed to investigate the differences in miRNA expression patterns within the Iranian population. METHODS: Total RNA was extracted from gastric cancer tissues and adjacent non-cancerous tissues from 32 patients. Expression levels of miR-21 and miR-221 were detected by Real time RT-PCR using a specific primer, with 5s rRNA as the internal reference gene. RESULTS: Our data showed that the expression levels of miR-21 and miR-221 in gastric cancer samples were significantly higher than in paired non-cancerous samples (P value less than 0.05). The receiver operating characteristic (ROC) analyses yielded the area under the curve (AUC) values of 80.30 for miR-21 and 93.30 for miR-221, and combined ROC analysis revealed the highest AUC value of 96.90 in discriminating GC patients from healthy controls. CONCLUSION: It seems that miR-21 and miR-221 expression pattern in Iranian patients with gastric cancer are similar to any other population. Considering the increased expression level of two miRNAs in cancerous tissue compared to normal tissue as well as the area under ROC curve, miR-21 and miR-221 can be used for early detection of gastric cancer.

Resveratrol-Dependent Down-regulation of Receptor for Advanced Glycation End-products and Oxidative Stress in Kidney of Rats With Diabetes.

BACKGROUND: Millions of people in the world have diabetes mellitus and its prevalence is growing. Oxidative stress, advanced glycation end-products (AGEs) and receptor for advanced glycation end-products (RAGE) play key role in the pathogenesis of diabetes. New and safe strategies of remedy are needed for this disease. OBJECTIVES: We hypothesized that resveratrol may exert a renal protective effect on diabetic rats. MATERIALS AND METHODS: Male rats with diabetes were treated with or without resveratrol as 1, 5, 10 mg/kg of body weight for 30 days. The total AGEs and malondialdehyde levels in kidney tissues were determined by spectrofluorimetric method and the insulin level was assayed using ELISA. The total antioxidant capacity contents in kidney and the glucose in plasma were measured by a colorimetric assay. The expression of RAGE was assayed in kidneys of all animals using quantitative PCR. RESULTS: In resveratrol-treated rats with diabetes, malondialdehyde levels, plasma glucose and expression of RAGE were significantly reduced compared with the untreated group. Moreover, the total antioxidant and insulin levels significantly increased in treated rats. There was no significant difference in the AGEs contents among all the groups. CONCLUSIONS: These results revealed that resveratrol has beneficial effects on kidney by extenuating the oxidative stress and down-regulation of RAGE expression.

Evaluation of miR-141, miR-200c, miR-30b Expression and Clinicopathological Features of Bladder Cancer.

Bladder cancer (BC) ranks the second most common genitourinary tract malignant tumor with high mortality and 70% recurrence rate worldwide. MiRNAs expression has noticeable role in bladder tumorigenesis. The purpose of this study was to assess miR-200c, miR-30b and miR-141 in tissue samples of patients with BC and healthy adjacent tissue samples and their association with muscle invasion, grade and the size of the tumor. Transurethral resection tissue samples were collected from thirty- five newly diagnosed untreated patients with BC from 2013 to 2014. The control group consisted of adjacent normal urothelium. All samples, observed by two pathologists, were diagnosed transitional cell carcinomas (TCC) with the proportion of tumor cells greater than 80%. Total RNA including miRNAs was extracted from about 50 mg tissue samples by applying TRIzol reagent. 2((-ΔΔ CT)) method was used to calculate relative quantification of miRNA expression. Two of 35 patients were females and the other 33 were males. Invasion to bladder muscle was observed in 13 (37%) cases. MiR-141, miR-200-c and miR30-b were up-regulated in 91%, 79% and 64% of malignant tissues, respectively. Down-regulation of miR-141 had a strong association with muscle invasion (P= 0.017). Significant inverse correlation between grading and miRNA-141 level was observed (P= 0.043).

Association between tissue miR-141, miR-200c and miR-30b and bladder cancer: a matched case-control study.

PURPOSE: To evaluate the expression of microRNAs in tissue samples from patients with bladder cancer and to compare it with healthy adjacent tissue samples as controls. MATERIALS AND METHODS: Thirty five tissue samples from patients with newly diagnosed untreated bladder transitional cell carcinoma and 35 adjacent normal urothelium were collected during 2013 to 2014. TRIzol reagent was used to isolate total RNA including microRNAs. RNA concentration and purity were determined using a nanodrop spectrophotometer. Also 1% agarose gel electrophoresis was used to assess integrity of RNA. Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) method was performed using the PARSGENOME microRNA RT-PCR system. Data was analyzed by STATA 11. RESULTS: A couple of patients were female the remainder were male. Mean age of patients were 71.06 ± 11.43 years. The expression level of miR-30b, miR-141 and miR-200c in case group were significantly higher than that of control normal tissue samples. miR-141 had higher expression rate in malignant tissue than two other miRNAs (P < .001). CONCLUSION: There was a more expression rate of miR-200c, miR-141 and miR-30b in bladder cancer tissues than healthy adjacent control tissues. Further studies are needed to draw final conclusion.

Expression of two basic mRNA biomarkers in peripheral blood of patients with non-small cell lung cancer detected by real-time rt-PCR, individually and simultaneously.

INTRODUCTION: Although extensive research has been conducted on lung cancer markers, a singular clinically applicable marker has not been found yet. The objective of this study was to evaluate the sensitivity and the specificity of carcinoembryonic antigen (CEA) mRNA and lung-specific X protein (LUNX) mRNA biomarkers in peripheral blood to detect lung cancer individually and simultaneously. METHODS: Thirty patients affected by lung cancer and 30 healthy individuals were studied in this research. Three vials of cDNA were made from each sample after taking peripheral blood samples and extracting total RNA. Each sample was examined by the real-time RT-PCR technique. The result from each vial was then compared with the sensitivity of overall marker. RESULTS: The CEA mRNA was positive in 24 out of 30 lung cancer patients. Hence, its sensitivity was determined at 80%, differing significantly from that observed in healthy individuals, where 11 positive cases were seen. The overall sensitivity of this marker was significantly associated with positivity in vials 2 and 3 but not in vial 1. The LUNX mRNA was positive in 21 out of 30 patients, indicating 70% sensitivity. This finding significantly differed from that in healthy individuals. The overall sensitivity of this marker was significantly associated with positivity in vials 1 and 3, but not in vial 2. In 93.3% of the patients, at least one positive marker was observed. CONCLUSION: The mentioned mRNA could be suggested as sensitive and specific markers in peripheral blood for primary diagnosis of lung cancer.

Simultaneous analysis of multidrug resistance 1(MDR1) C3435T, G2677T/A, and C1236T genotypes in Hamadan City population, West of Iran.

BACKGROUND: One of the limitations in the treatment of common diseases such as cancer chemotherapy is development of multidrug resistance (MDR). Polymorphisms could alter the expression level of MDR1 gene, which plays an important role in MDR. In this research, the frequency of C3435T, C1236T, and G2677T/A polymorphisms of MDR1 gene was investigated in a large group of population from Hamadan city to provide a sample data resource. METHODS: Peripheral blood (2 ml) was taken, and DNA extraction was carried out. Multiplexed mutagenically separated PCR, which was followed by polyacrylamide gel electrophoresis and silver staining, was applied to detect the mentioned polymorphisms in 935 individuals. Sequencing performed for confirmation of gel electrophoresis resulted in 10 random cases. In total, alleles and genotypes of 933 persons (776 women and 157 men) were determined. RESULTS: The most frequent alleles of the polymorphisms were: 3435T, C1236, and G2677. The most frequent genotypes were: 3435C/T, 1236C/T, and 2677G/A, and their concurrent presence was also found as the most frequent simultaneous genotypes. There was not any meaningful difference among the prevalence of these genotypes in groups of men and women. CONCLUSION: Our results were close to those of other studies performed in Iran and compared to the other ethnic groups, which showed more similarity to Asian peoples than Europeans. As an aspect of personalized medicine, it could be used by chemotherapists to improve the routine methods of cancer treatment.

Cholesterol Transporters ABCA1 and ABCG1 Gene Expression in Peripheral Blood Mononuclear Cells in Patients with Metabolic Syndrome.

ABCA1 and ABCG1 genes encode the cholesterol transporter proteins that play a key role in cholesterol and phospholipids homeostasis. This study was aimed at evaluating and comparing ABCA1 and ABCG1 genes expression in metabolic syndrome patients and healthy individuals. This case-control study was performed on 36 patients with metabolic syndrome and the same number of healthy individuals in Hamadan (west of Iran) during 2013-2014. Total RNA was extracted from mononuclear cells and purified using RNeasy Mini Kit column. The expression of ABCA1 and ABCG1 genes was performed by qRT-PCR. Lipid profile and fasting blood glucose were measured using colorimetric procedures. ABCG1 expression in metabolic syndrome patients was significantly lower (about 75%) compared to that of control group, while for ABCA1 expression, there was no significant difference between the two studied groups. Comparison of other parameters such as HDL-C, FBS, BMI, waist circumference, and systolic and diastolic blood pressure between metabolic syndrome patients and healthy individuals showed significant differences (P < 0.05). Decrease in ABCG1 expression in metabolic syndrome patients compared to healthy individuals suggests that hyperglycemia, related metabolites, and hyperlipidemia over the transporter capacity resulted in decreased expression of ABCG1. Absence of a significant change in ABCA1 gene expression between two groups can indicate a different regulation mechanism for ABCA1 expression.

Lipid Lowering Effects of Hydroalcoholic Extract of Anethum graveolens L. and Dill Tablet in High Cholesterol Fed Hamsters.

Objective. This study was aimed to determine the effect of Anethum graveolens extract and Anethum graveolens (dill) tablet on lipid profile, liver enzymes, and gene expression and enzymatic activity of HMG-CoA reductase in high cholesterol fed hamsters. Materials and Methods. Golden Syrian male hamsters (130 ± 10 g) were randomly divided into 6 groups (n = 6) and received daily the following: group 1 received chow + 2% cholesterol + 0.5% cholic acid (HCD), groups 2 and 3 received HCD diet plus 100 and 200 mg/kg hydroalcoholic extract of dill, respectively, and groups 4 and 5 received HCD diet plus 100 and 200 mg/kg dill tablet, respectively. Group 6 received only chow. After 1 month feeding serum biochemical factors were determined. HMG-CoA reductase mRNA level was measured (real-time PCR) and its activity was determined spectrophotometrically. Results. Compared with hypercholesterolemic group 1, lipid profile, blood glucose, and liver enzymes significantly decreased in all dill tablet or dill extract treated groups (p < 0.05). The changes in HMG-CoA reductase gene expression level and enzyme activity significantly reduced in animals that received 200 mg/kg of extract or tablet. Conclusion. Dill extract and dill tablet showed potential hypocholesterolemic properties in hamsters by inhibition of HMG-CoA reductase activity.

Effect of myomectomy on endometrial glutathione peroxidase 3 (GPx3) and glycodelin mRNA expression at the time of the implantation window.

BACKGROUND: In fertile women, glycodelin and glutathione peroxidase 3 (GPx3) genes expression rises during the luteal phase, with a peak occurring during the implantation window. The expression of these genes decreases in women with myomas. To determine whether myomectomy would reverse glycodelin and GPx3 expression, we evaluated the transcript levels of these genes in the endometrium of patients before and after myomectomy. METHODS: Expression of glycodelin and GPx3 genes were examined prospectively during the midluteal phase in the endometrium obtained from infertile women with myoma (n = 12) before and three months after myomectomy. Endometrial expression of these genes was evaluated using quantitative real-time RT-PCR. RESULTS: Endometrial glycodelin mRNA expression levels (normalized to 18S rRNA expression) were increased significantly in endometrium of patients after myomectomy (P = 0.02). GPx3 mRNA expression was increased insignificantly after myomectomy (P = 0.43). CONCLUSION: The results showed that myomectomy increased endometrial glycodelin (significantly) and GPx3 (not significantly) gene expression after 3 months. Study at different times and detecting expression of these genes can reveal more details.

Improved real-time rt-PCR assays of two colorectal cancer peripheral blood mRNA biomarkers: a pilot study.

BACKGROUND: Efficient screening for detection of colorectal cancer (CRC) at earlier stages reduces its mortality. The purpose of this study was to investigate expression of carcinoembryonic antigen (CEA) and human telomerase reverse transcriptase (hTERT) mRNA in peripheral blood of CRC patients and to present strategies for early detection screen test. METHODS: Twenty seven patients in non-metastatic stage and 27 healthy individuals were studied. Expression of CEA, hTERT mRNA and 18srRNA (18s subunit of ribosomal RNA, as reference gene) were determined based on real-time RT-PCR on 3 µg of total RNA from blood in 3 separate vials (1 µg per vial). RESULTS: Positive expression rate of CEA mRNA (78%) and hTERT mRNA (81%) were higher in patient group (P<0.001). These rates were meaningfully higher than the results of individual vials containing only 1 µg of total RNA. Difference between Ct values of markers with 18srRNA ΔCt) was higher in healthy group than patient one. Therefore, a ΔCt cut-off value was determined for distinguishing between true- and false-positive results. Concurrent expression of both markers was found in 67% of the patients, which was higher than healthy cases (11%). Combination of concurrent marker expression with cut-off point strategy increased specificity to 100%. CONCLUSION: These results showed that concurrent evaluation of marker expression and performing the test on 3 µg of samples in 3 separate vials may increase specificity and sensitivity of real-time RT-PCR for early detection of non-metastatic CRC. However, more investigations with larger numbers of samples are needed to verify these results.