Expression of ER stress markers (GRP78 and PERK) in experimental nephrotoxicity induced by cisplatin and gentamicin: roles of inflammatory response and oxidative stress.

This study aimed to establish the relationship between two endoplasmic reticulum (ER) stress proteins, glucose-regulated protein 78 (GRP78/BiP) and PKR-like endoplasmic reticulum kinase (PERK), and oxidative stress markers in cisplatin (CIS)-induced and gentamicin (GEN)-induced nephrotoxicity.The study consisted of five groups: control (saline solution only), CIS D2 (2.5 mg/kg for 2 days), CIS D7 (2.5 mg/kg for 7 days), GEN D2 (160 mg/kg for 2 days), and GEN D7 (160 mg/kg for 7 days). All rats were sacrificed 24 h after the last injection for standard clinical chemistry, and ultrastructural and histological evaluation of the kidney.CIS and GEN increased blood urea nitrogen (BUN) and serum creatinine (Cr) levels, as well as total oxidant status (TOS), while decreasing total antioxidant status (TAS) level in CIS D7 and GEN D7 groups. Histopathological and ultrastructural findings were also consistent with renal tubular damage. In addition, expression of markers of renal inflammation (tumor necrosis factor-α (TNF-α) and interleukin 1ß (IL-1ß)) and ER stress markers (GRP78 and PERK) was significantly increased in the kidney tissue of rats treated with CIS and GEN for 7 days.These findings suggest that CIS and GEN administration for 7 days aggravates nephrotoxicity through the enhancement of oxidative stress, inflammation, and ER stress-related markers. As a result, the recommended course of action is to utilize CIS and GEN as an immediate but brief induction therapy, stopping after 3 days and switching to other drugs instead.

A Review on the Design, Synthesis, and Structure-activity Relationships of Benzothiazole Derivatives against Hypoxic Tumors.

There has been a growing body of studies on benzothiazoles and benzothiazole derivatives as strong and effective anti-tumor agents against lung, liver, pancreas, breast, and brain tumors. Due to the highly proliferative nature of the tumor cells, the oxygen levels get lower than that of normal tissues in the tumor microenvironment. This situation is called hypoxia and has been associated with increased ability for carcinogenesis. For the drug design and development strategies, the hypoxic nature of the tumor tissues has been exploited more aggressively. Hypoxia itself acts as a signal initiating system to activate the pathways that eventually lead to the spread of the tumor cells into the different tissues, increases the rate of DNA damage, and eventually ends up with more mutation levels that may increase the drug resistance. As one of the major mediators of hypoxic response, hypoxia-inducible factors (HIFs) have been shown to activate angiogenesis, metastasis, apoptosis resistance, and many other protumorigenic responses in cancer development. In the current review, we will be discussing the design, synthesis, and structureactivity relationships of benzothiazole derivatives against hypoxic tumors such as lung, liver, pancreas, breast, and brain as potential anti-cancer drug candidates. The focus points of the study will be the biology behind carcinogenesis and how hypoxia contributes to the process, recent studies on benzothiazole and its derivatives as anti-cancer agents against hypoxic cancers, conclusions, and future perspectives. We believe that this review will be useful for researchers in the field of drug design during their studies to generate novel benzothiazole-containing hybrids against hypoxic tumors with higher efficacies.

Effects of Bone Marrow and Adipose-Derived Mesenchymal Stem Cells on microRNA Expressions in Acute Alkaline Corneal Burn.

Purpose: The aim of this study was to investigate the microRNA (miRNA) expressions of the corneal tissue after an alkaline burn and to compare the efficiency of adipose- and bone marrow-derived mesenchymal stem cells (MSCs) on expressions. Methods: Thirty-two rats were divided into 4 groups. No intervention was made in the control group. A chemical burn was created by applying 4 µL NaOH soaked in 6 mm filter paper to the right eye of each animal in the other groups. Whereas only subconjunctival 0.1 mL phosphate-buffered saline (PBS) was injected to in the group 1, 2 × 106 adipose- or bone marrow-derived MSC in 0.1 mL PBS was injected subconjunctivally to the animals in the remaining groups (groups 2 and 3, respectively). Tissue samples were collected for miRNA analysis on the third day after the burn. Results: When group 1 was compared with the control group, the expression of 3 of 93 miRNAs increased significantly, whereas the expression of 50 miRNAs decreased significantly. Significant changes in miRNA expressions were observed when group 1 was compared with groups 2 and 3. Although a significant change was observed in the expression of 6 miRNAs in the adipose-derived MSC group, it was found that the expression of 65 miRNAs significantly changed in the bone marrow-derived MSC group. Conclusion: This study shows that there are significant changes in some miRNA expressions after corneal alkaline burn and these changes can be reversed with the subconjunctival injection of MSCs.

Evaluation of Anti-Inflammatory and Antiapoptotic Effects of Bone Marrow and Adipose-Derived Mesenchymal Stem Cells in Acute Alkaline Corneal Burn.

Purpose: The aim of the present study is to comparatively evaluate the anti-inflammatory and antiapoptotic effects of bone marrow and adipose-derived mesenchymal stem cells (MSCs) applied subconjunctivally after alkaline corneal burn. Methods: Thirty-two rats were divided into 4 groups and included in the study (n = 8). While no intervention was made in the control group, a chemical burn was created by applying 4 µL of NaOH soaked in 6 mm filter paper to the right eye of each subject in the other groups under general anesthesia. While only subconjunctival 0.1 mL phosphate-buffered saline (PBS) was injected to in the group 1, 2 × 106 adipose or bone marrow-derived MSC in 0.1 mL PBS was applied subconjunctivally to the subjects in the remaining groups (Group 2 and 3, respectively). Tissue samples were collected for histological analysis on the third day after the burn. Tissue samples were evaluated light microscopically and immunohistochemically stained for interleukin-1 beta (IL-1ß), tumor necrosis factor alpha (TNF-α), caspase-3 (Cas-3), and CD68. Results: The IL-1ß and TNF-α staining scores and the number of CD68- and Cas-3-positive stained cells were significantly lower in the groups given bone marrow and adipose-derived MSC compared to the alkaline burn group (P < 0.0001, for all parameters). Epithelial IL-1ß and TNF-α staining scores were significantly lower in the bone marrow-derived MSC group compared to the adipose-derived MSC group (P < 0.0001, for all parameters). Conclusions: The presented study shows that both bone-marrow and adipose-derived MSCs support wound healing in the corneal tissue and strongly suppress the inflammation occured in the tissue.

Expression levels of maternal plasma microRNAs in preeclamptic pregnancies.

The present study aimed to identify the differential expression profiles of microRNAs in the plasma between patients with preeclampsia (PE) and healthy pregnancies using quantitative real-time PCR. The expression profiles of 32 miRNAs in maternal plasma from 31 patients with PE and 32 healthy pregnancies were evaluated. The expression levels of eight miRNAs including miR-210, miR-375, miR-197-3p, miR-132-3p, miR-29a-3p, miR-328, miR-24-3p, and miR-218-5p were significantly upregulated and the expression levels of three miRNAs, including miR-302b-3p, miR-191-5p, and miR-17-5p, were significantly downregulated in patients with preeclampsia when compared to healthy pregnant women. In conclusion, we identified 11 miRNAs that may be potential biomarkers for non-invasive diagnosis and a pivotal role in the prediction of PE. Considering the small cohort of patients, further studies with larger samples from different gestational stages are necessary to confirm our findings.IMPACT STATEMENTWhat is already known on this subject? The alterations in the release pattern of placenta-specific miRNAs detected in maternal serum have been found to be associated with pregnancy-related complications such as preeclampsia (PE).What do the results of this study add? In the present study, the release pattern of seven miRNAs had consistency and two of them had inconsistency with previous researches. Moreover, two novel miRNAs were also defined to demonstrate the interrelationship between PE and miRNAs.What are the implications of these findings for clinical practice and/or future research? The identification of 11 miRNAs that may be potential biomarkers for non-invasive diagnosis and a pivotal role in the prediction of PE. Considering the small cohort of patients, further studies with larger samples from different gestational stages are necessary to confirm our findings.

Evaluation of miRNAs Related with Nuclear Factor Kappa B Pathway in Lipopolysaccharide Induced Acute Respiratory Distress Syndrome.

This study aimed to determine the expression of nuclear factor kappa B (NF-κB) pathway related miRNAs in experimental acute respiratory distress syndrome (ARDS) induced by lipopolysaccharide (LPS) in rats, and to elucidate the underlying molecular mechanism. Twenty four sprague dawley rats were randomly divided into two groups; LPS (n = 12) and control (n = 12). Experimental ARDS was induced by intraperitoneal injection of E. coli LPS in LPS group. Intraperitoneal saline was administered in control group. Serum and lung samples were collected from both groups. Immunohistochemistry staining was performed for interleukin 1ß (IL-1ß), tumor necrosis factor α (TNF-α), CD 68, and caspase-3 in lung samples. Intensity of staining was scored as strong, moderate, weak, and no for evaluation of IL-1ß and TNF-α. In addition, caspase-3 and CD68-positive stained cells were counted in sections. Expressions of 9 miRNAs were determined by quantitative real-time PCR in serum samples. IL-1ß and TNF-α staining scores were significantly higher in the LPS group in comparison with the control group (P = 0.04 and P = 0.02, respectively). In addition, caspase-3 and CD68-positive stained cells were significantly higher in the LPS group (P = 0.02). Expressions of seven miRNAs were significantly changed in the LPS group in comparison with the control group. While six miRNAs (miR-7a-5p, miR-7b, miR-9a-5p, miR-21-5p, miR-29a-3p, and miR-138-5p) were up regulated, only miR-124-3p was down regulated. This study suggests that these miRNAs may have a role in the pathogenesis of ARDS related to NF-κB. However, this relationship needs to be examined in new studies by evaluation of pathways and target genes.

Effects of Bevacizumab, Ranibizumab, and Aflibercept on MicroRNA Expression in a Retinal Pigment Epithelium Cell Culture Model of Oxidative Stress.

PURPOSE: This study aimed to evaluate the effects of bevacizumab, ranibizumab, and aflibercept on the microRNA (miRNA) expression in human retinal pigment epithelium cell (ARPE-19) culture model of oxidative stress. METHODS: Control cells were cultured in the hydrogen peroxide (H2O2)-free medium. In H2O2 group ARPE-19 cells were exposed to 600 µM H2O2 alone for 18 h. In study groups, cells were preincubated with bevacizumab, ranibizumab, and aflibercept (1.25-2.5, 0.5 and 2.0 mg/mL, respectively) for 3 h before H2O2 exposure. Another group of ARPE-19 cells were incubated with drugs for 3 h without H2O2 exposure. Cell viability and vascular endothelial growth factor (VEGF) levels were evaluated by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and enzyme-linked immunosorbent assay. The expression levels of 1,152 miRNAs were determined by quantitative real-time PCR. RESULTS: Incubation with 600 µM H2O2 alone for 18 h decreased cell viability by ∼50%. Cell viability was greater in the anti-VEGF drug groups compared with the H2O2 group, but the differences were not significant (P > 0.05). VEGF levels were significantly lower in the anti-VEGF drug groups compared with the H2O2 group (P < 0.05 for all study groups), with no significant differences between the study groups (P > 0.05). Incubation with anti-VEGF drugs alone had no effect on miRNA expression in ARPE-19 cells. However, preincubation with bevacizumab, ranibizumab, and aflibercept significantly altered the profile of H2O2-modulated miRNA expression. CONCLUSIONS: Preincubation with anti-VEGF drugs can alter the miRNA expression profile in response to H2O2-induced oxidative stress, and these drugs may have epigenetic effects.

Evaluation of microRNA responses in ARPE-19 cells against the oxidative stress.

PURPOSE: This study aimed to determine microRNA (miRNA) expression profile of human retinal pigment epithelium cell (ARPE-19) against the oxidative stress induced by hydrogen peroxide (H2O2). METHODS: ARPE-19 cells were incubated with different concentrations of H2O2 (200, 600 and 800 µM) for 18 h, and then cell viability, vascular endothelial growth factor levels and total oxidant status were evaluated. Expressions of 1152 miRNA were determined by quantitative real-time PCR in each group. RESULTS: Expressions of 90 miRNA were significantly changed in the ARPE-19 cells incubated with H2O2 compared to control group. However, miR-143-3p was only found to be expressed in groups incubated with H2O2. While 24 miRNA (hsa-miR-200c-3p, miR-192-5p, miR-194-5p, miR-141-3p, miR-658, miR-18 b-5p, miR-486-5p, miR-525-3p, miR-493-3p, miR-518d-3p, miR-29 b-1-5p, miR-675-3p, miR-1238-3p, miR-195-3p, miR-1539, miR-490-5p, miR-3200-5p, miR-1273d, miR-130a-5p, miR-30 b-5p, miR-1247-5p, miR-1910-5p, miR27a-5p and miR-200 b-3p) upregulated due to the increased dose of H2O2, nine miRNA (hsa-miR-96-5p, miR-33a-5p, miR-345-5p, miR-106 b-3p, miR-1285-3p, miR-23 b-5p, miR-27 b-5p, miR-103a-3p and miR-4289) were also found to be downregulated. CONCLUSION: This study suggests that oxidative stress may be an important factor on expression of miRNAs in ARPE-19 cells. These miRNAs may have a role in the pathogenesis of age-related macular degeneration related to oxidative stress. However, this relationship needs to be examined in new studies by evaluation of pathways and target genes.

Protective Effect of Combined Caffeic Acid Phenethyl Ester and Bevacizumab Against Hydrogen Peroxide-Induced Oxidative Stress in Human RPE Cells.

PURPOSE: This study aimed to evaluate the protective effects of caffeic acid phenethyl ester (CAPE) and combined CAPE-bevacizumab against oxidative stress induced by hydrogen peroxide (H2O2) in human retinal pigment epithelium. METHODS: ARPE-19 cells were pretreated with 5, 10, and 30 µM CAPE alone and in combination with bevacizumab for 3 h, then exposed to H2O2 for 16 h. Cell viability was evaluated with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Vascular endothelial growth factor (VEGF) protein levels in the medium were measured using a human VEGF ELISA kit. Total antioxidant status (TAS) and total oxidant status (TOS) were measured in ARPE-19 cells using the test kit from Rel Assay. Expression levels of VEGF, Bax, Bcl-2, cytochrome c, apoptotic protease activating factor-1 (apaf-1), and caspase-3 were determined using reverse transcription polymerase chain reaction. RESULTS: Pretreatment of ARPE-19 cells with 30 µM CAPE and combined CAPE-bevacizumab reduced H2O2 mediated cell death. H2O2-induced oxidative stress increased TOS and VEGF production, which was significantly inhibited by CAPE and the CAPE-bevacizumab combination. VEGF, Bax, cytochrome c, apaf-1, and caspase-3 gene expressions were significantly decreased in cells pretreated with 5, 10, and 30 µM CAPE and combined CAPE-bevacizumab compared to the H2O2 group. In addition, Bcl-2 expression was significantly increased in both the CAPE and CAPE-bevacizumab combination groups compared to the H2O2 group. CONCLUSIONS: CAPE has a protective effect on ARPE-19 cells against oxidative stress, and VEGF protein level and expression can be decreased by incubation with different concentrations of CAPE. These results demonstrate that CAPE suppresses the mitochondria-mediated apoptosis in ARPE-19 cells under oxidative stress. In addition, the use of CAPE in combination with bevacizumab has an additive effect.

Inhibition of Radiation-Induced Oxidative Damage in the Lung Tissue: May Acetylsalicylic Acid Have a Positive Role?

The lung is relatively sensitive to irradiation. It is shown that acetylsalicylic acid (ASA) might reduce oxidative injury and that it has a place in protection from cancer. The aim of this study is to evaluate the potential radioprotective effects of ASA. Whole-body irradiation (6 Gy, single dose) was applied to the rats. Glutathione (GSH), malondialdehyde (MDA), myeloperoxidase (MPO), and nitric oxide (NO) levels in the lung tissue were measured. Control (C), Radiation (R), Radiation + ASA (R + ASA; received irradiation and 25 mg/kg of ASA intraperitoneally (i.p.)), and Radiation + Amifostine (R + WR-2721; received irradiation and 200 mg/kg of WR-2721 i.p.) groups were used. The MPO levels decreased statistically significantly in the group administered ASA. Histopathologically, a radioprotective effect of ASA was more evident in the R + ASA group. ASA is an agent which has not been used as a radioprotector in the clinic yet, and it is worth supporting with more advanced studies.

Is mycophenolate mofetil an alternative agent to corticosteroids in traumatic nerve paralysis?

OBJECTIVE: The effects of an immunosuppressive agent, mycophenolate mofetil (MM), were investigated and compared with those of methylprednisolone (MP) and dexamethasone (DXM) on the traumatic nerve function. STUDY DESIGN: This is a randomized controlled animal study. MATERIALS AND METHODS: This experimental study was performed on 84 male Wistar albino rats. The rats were assigned to 12 groups each consisting of 7 animals. The groups were formed according to application of normal-dose DXM (group 1A-B), high-dose MP (group 2A-B), normal-dose MP (group 3A-B), MM (group 4A-B), and MM with high-dose MP combination therapies (group VA-B). Right sciatic nerve dissection was performed, and compound muscle action potential thresholds were recorded. The nerve was traumatized with the compression of a Jeweller forceps for 20 seconds. Posttraumatic thresholds were also recorded. The compound muscle action potential thresholds were recorded in the first and fourth weeks for the assigned groups. Then, the nerve was transected and prepared for electron microscopic and histopathologic examinations. Nitric oxide and malondialdehyde assessments were performed on both tissue and blood samples. RESULTS: Only the MM and MP+MM groups had satisfactory electron microscopic findings and were about to reach the tissue characteristics of the control animals. Despite the electrophysiologic recovery, the DXM group was found to have poor electron microscopic scoring. CONCLUSIONS: Mycophenolate mofetil has been found to be beneficial in the treatment of traumatic nerve paralysis. Although a complementary investigation is needed, this immunosuppressive agent may be an alternative to corticosteroids for the selected cases where steroid therapy is contraindicated.

Evaluation of circulating miRNAs in wet age-related macular degeneration.

PURPOSE: In the present study, we aimed to investigate the changes in plasma miRNA in patients with wet age-related macular degeneration. METHODS: The expression profiles of 384 miRNAs in plasma from 33 patients (22 male, 11 female) who were diagnosed with wet age-related macular degeneration with fundus examination, fundus fluorescein angiography, and optical coherence tomography and 31 controls (17 male, 14 female) were evaluated using high-throughput quantitative real-time PCR. RESULTS: Our results demonstrated that the expression level of five miRNAs (miR-17-5p, miR-20a-5p, miR-24-3p, miR-106a-5p, and miR-223-3p) was significantly upregulated in patients with age-related macular degeneration when compared to the control group (p<0.05). The expression level of 11 miRNAs (miR-21-5p, miR-25-3p, miR-140-3p, miR-146b-5p, miR-192-5p, miR-335-5p, miR-342-3p, miR-374a-5p, miR-410, miR-574-3p, and miR-660-5p) was significantly downregulated in patients (p<0.05). In addition, ten miRNAs (miR-26b-5p, miR-27b-3p, miR-29a-3p, miR-139-3p, miR-212-3p, miR-324-3p, miR-324-5p, miR-532-3p, miR-744-5p, and miR-Let-7c) were expressed only in the patient group. CONCLUSIONS: Our results suggest that plasma miRNA levels may change in wet age-related macular degeneration. These molecules may have an important therapeutic target in patients who are unresponsive to antivascular endothelial growth factor therapy. However, further studies must be conducted for possible effects of miRNAs in vascular disorders of eye such as age-related macular degeneration.

The G1057D polymorphism of insulin receptor substrate-2 associated with gestational diabetes mellitus.

OBJECTIVE: The Gly1057D polymorphism in the insulin receptor substrate-2 (IRS-2) gene has been reported to be associated with insulin resistance, obesity and type 2 diabetes; little is known about its possible association with gestational diabetes mellitus (GDM). To investigate this association we determined the distribution of its genotypes and frequency of alleles in GDM patients. MATERIALS AND METHODS: The study population consisted of 94 subjects; among them were 44 patients with GDM and 50 healthy controls without diabetes. Genomic DNA was extracted from the leukocyte by high pure polymerase chain reaction (PCR) template preparation kit. Genetic polymorphism of IRS-2 G1057D was detected by using PCR-based restriction fragment-length polymorphism (RFLP). RESULTS: For IRS-2 G1057D polymorphism, there was no significant difference in genotype distribution between GDM patients and controls. The risk for GDM was 2.97 times higher (95% CI: 0.89-9.93, p = 0.076) in the individuals with the IRS-2 DD genotype compared to the GG genotype. Also individuals with the IRS-2 D allele had a significantly higher risk of GDM compared with individuals with the IRS-2 G allele, with a relative risk of 1.86 (95% CI: 1.02-3.37, p = 0.042) for cases compared with population controls. CONCLUSION: These results suggest that IRS-2 1057D allele may be associated with GDM.

Contribution of RhoA/Rho-kinase/MEK1/ERK1/2/iNOS pathway to ischemia/reperfusion-induced oxidative/nitrosative stress and inflammation leading to distant and target organ injury in rats.

The small G protein RhoA and its downstream effector Rho-kinase play an important role in various physiopathological processes including ischemia/reperfusion (I/R) injury. Reactive oxygen and nitrogen species produced by iNOS and NADPH oxidase are important mediators of inflammation and organ injury following an initial localized I/R event. The aim of this study was to determine whether RhoA/Rho-kinase signaling pathway increases the expression and activity of MEK1, ERK1/2, iNOS, gp91(phox), and p47(phox), and peroxynitrite formation which result in oxidative/nitrosative stress and inflammation leading to hindlimb I/R-induced injury in kidney as a distant organ and gastrocnemius muscle as a target organ. I/R-induced distant and target organ injury was performed by using the rat hindlimb tourniquet model. I/R caused an increase in the expression and/or activity of RhoA, MEK1, ERK1/2, iNOS, gp91(phox), p47(phox), and 3-nitrotyrosine and nitrotyrosine levels in the tissues. Although Rho-kinase activity was increased by I/R in the kidney, its activity was decreased in the muscle. Serum and tissue MDA levels and MPO activity were increased following I/R. I/R also caused an increase in SOD and catalase activities associated with decreased GSH levels in the tissues. Y-27632, a selective Rho-kinase inhibitor, (100µg/kg, i.p.; 1h before reperfusion) prevented the I/R-induced changes except Rho-kinase activity in the muscle. These results suggest that activation of RhoA/Rho-kinase/MEK1/ERK1/2/iNOS pathway associated with oxidative/nitrosative stress and inflammation contributes to hindlimb I/R-induced distant organ injury in rats. It also seems that hindlimb I/R induces target organ injury via upregulation of RhoA/MEK1/ERK1/2/iNOS pathway associated with decreased Rho-kinase activity.

Intravitreal bevacizumab effects on VEGF levels in distant organs: an experimental study.

PURPOSE: The aim of this study was to determine the effects of single-dose intravitreal bevacizumab on the levels of vascular endothelial growth factor (VEGF) in serum and distant organs. METHODS: Adult New Zealand albino rabbits (n = 40) were divided into experimental and control groups. Experimental rabbits received a single 0.05 ml intravitreal injection of 1.25 mg bevacizumab (Avastin) into the right eye, and control rabbits (n = 8) received no injection. Following injection, group 1 rabbits (n = 8) were sacrificed on day 1, group 2 rabbits (n = 8) on day 7, group 3 rabbits (n = 8) on day 14, and group 4 rabbits (n = 8) on day 28; control rabbits were sacrificed on day 28. After sacrifice, samples of brain, heart, liver, kidney and blood were collected. Levels of VEGF in serum and tissue were measured using enzyme-linked immunosorbent assay. The presence of bevacizumab was evaluated by immunofluorescence staining in tissues. RESULTS: Positive bevacizumab immunoreactivity was observed in brain, heart and kidney. Serum VEGF levels significantly decreased in groups 3 and 4 compared with controls (p < 0.05). Liver VEGF levels significantly decreased in group 3 compared with controls (p < 0.05). CONCLUSIONS: Intravitreal bevacizumab not only may escape from the blood-retinal barrier and enter the general circulation, but also may be disseminated to distant organs. Our study demonstrates that a single dose of intravitreally injected bevacizumab decreases VEGF levels in serum and liver.

Differential expression of microRNAs in plasma of patients with laryngeal squamous cell carcinoma: potential early-detection markers for laryngeal squamous cell carcinoma.

PURPOSE: Altered microRNA (miRNA) expression has been found in many cancers, including lung, breast, prostate, bladder and colorectal cancer. Many recent studies have demonstrated that aberrant plasma miRNAs were also found in various types of cancers. However, the alteration in plasma miRNA expressions in laryngeal squamous cell carcinoma (LSCC) remains unclear. The present study aimed to investigate the alterations in plasma miRNAs in LSCC. METHODS: In the present study, the expression profiles of 738 miRNAs in plasma from 20 patients and 44 healthy subjects were evaluated using high-throughput real-time quantitative polymerase chain reaction. RESULTS: Our results demonstrated that expression levels of 17 miRNAs were significantly upregulated in patients with LSCCs when compared to control group (p < 0.05). Expression levels of nine miRNAs were found significantly downregulated in LSCC patients (p < 0.05). In addition, 17 miRNAs were expressed only in LSCC group, and five of these miRNAs (miR-331-3p, 603, 1303, 660-5p and 212-3p) are LSCC specific and never seen before in plasma of any human subject. CONCLUSION: In conclusion, our study suggests that detecting these LSCC-specific miRNAs in plasma might serve as novel noninvasive biomarkers for LSCC.

Determination of plasma microRNA for early detection of gastric cancer.

Gastric cancer is the fourth most prevalent malignancy worldwide and remains the second most common cause of cancer-related death globally. Understanding the molecular structure of gastric carcinogenesis might identify new diagnostic and therapeutic strategies for this disease. Thus, early detection of gastric cancer is a key measure to reduce the mortality and improve the prognosis of gastric cancer. There have recently been several reports that microRNAs (miRNAs) circulate in highly stable, cell-free forms in blood. Because serum and plasma miRNAs are relatively easy to access, circulating miRNAs also have great potential to serve as non-invasive biomarkers. Although a number of miRNAs associated with gastric cancer have been identified, the underlying mechanism of these miRNAs in tumorigenesis and tumor progression remains to be investigated. The purpose of this study is to identify the potential of serum miRNAs as biomarkers for early detection of gastric cancer patients. RNA was isolated using the High Pure miRNA Isolation Kit (Roche) following the manufacturer's protocol. cDNA and preamplification protocols were obtained from the isolated plasma miRNAs. The BioMark™ 96.96 Dynamic Array (Fluidigm Corporation) for real-time qPCR was used to simultaneously quantite the expression of 740 miRNAs. All statistical analyses were performed using the Biogazelle's qbase PLUS 2.0 software. In this study, among 740 miRNAs that we analyzed only miR-195-5p was significantly (p < 0.05, fold changes = 13, 3) down-regulated in gastric cancer patients compared with control. We demonstrated that miR-195-5p is a novel tumor suppressor miRNA and may contribute to gastric carcinogenesis. The miRNA expression profile described in this study should contribute to future studies on the role of miRNAs in gastric cancer.

Effects of rosiglitazone with insulin combination therapy on oxidative stress and lipid profile in left ventricular muscles of diabetic rats.

PURPOSE: In this study, we tested the hypothesis that rosiglitazone (RSG) with insulin is able to quench oxidative stress initiated by high glucose through prevention of NAD(P)H oxidase activation. METHODS AND MATERIALS: Male albino Wistar rats were randomly divided into an untreated control group (C), a diabetic group (D) that was treated with a single intraperitoneal injection of streptozotocin (45 mg kg(-1)), and rosiglitazone group that was treated with RSG twice daily by gavage and insulin once daily by subcutaneous injection (group B). HbA1c and blood glucose levels in the circulation and malondialdehyde and 3-nitrotyrosine levels in left ventricular muscle were measured. RESULT: Treatment of D rats with group B resulted in a time-dependent decrease in blood glucose. We found that the lipid profile and HbA1c levels in group B reached the control group D rat values at the end of the treatment period. There was an increase in 3-nitrotyrosine levels in group D compared to group C. Malondialdehyde and 3-nitrotyrosine levels were found to be decreased in group B compared to group D (P < 0.05). CONCLUSION: Our data suggests that the treatment of diabetic rats with group B for 8 weeks may decrease the oxidative/nitrosative stress in left ventricular tissue of rats. Thus, in diabetes-related vascular diseases, group B treatment may be cardioprotective.

Serum levels and H/L gene polymorphism of mannose-binding lectin in primary open angle glaucoma.

PURPOSE: To analyze the serum levels and H/L gene polymorphisms of mannose-binding lectin-2 (MBL-2) in primary open angle glaucoma (POAG) cases and control subjects to investigate whether MBL-2 has a possible role in the development and pathogenesis of POAG. MATERIALS AND METHODS: In 45 POAG cases and age and sex-matched 45 healthy controls, Elisa Kit was used to determine serum levels of MBL-2. The genomic DNA of patient and control groups was extracted from whole blood using High Pure PCR template preparation kit. Genotyping of MBL-2 polymorphisms were detected by using a MBL-2 mutation detection kit in real-time PCR. Chi-square or Fisher's Exact Tests were used to evaluate the distribution of MBL-2 H/L genotypes among patients and control subjects. Associations between the H/L genotype and POAG risk were analyzed by using binary logistic regression. The serum MBL-2 levels of both groups were compared with Independent Sample t-test. RESULTS: Mean MBL-2 serum levels in the patient group (21.30 ± 4.97 µg/mL) was significantly higher than the control group (17.48 ± 3.66 µg/mL), (p < 0.001). The distribution of alleles in the patient group was 28.9% for LL, 44.4% for HL, 26.7% for HH and in controls was 33.3% for LL, 37.8% for HL, 28.3% for HH. According to genotype ratios, the two groups were not different from each other. CONCLUSIONS: Our findings may suggest an association between high serum MBL-2 levels and POAG, but H/L gene polymorphism of MBL-2 seems not to be associated with POAG.

The distribution of circulating microRNA and their relation to coronary disease.

BACKGROUND: MicroRNAs (miRNAs) are small RNAs that regulate gene expression by suppressing protein translation and may influence RNA expression. MicroRNAs are detected in extracellular locations such as plasma; however, the extent of miRNA expression in plasma its relation to cardiovascular disease is not clear and many clinical studies have utilized array-based platforms with poor reproducibility. METHODS AND RESULTS: Initially, to define distribution of miRNA in human blood; whole blood, platelets, mononuclear cells, plasma, and serum from 5 normal individuals were screened for 852 miRNAs using high-throughput micro-fluidic quantitative RT-PCR (qRT-PCR). In total; 609, 448, 658, 147, and 178 miRNAs were found to be expressed in moderate to high levels in whole blood, platelets, mononuclear cells, plasma, and serum, respectively, with some miRNAs uniquely expressed. To determine the cardiovascular relevance of blood miRNA expression, plasma miRNA (n=852) levels were measured in 83 patients presenting for cardiac catheterization. Eight plasma miRNAs were found to have over 2-fold increased expression in patients with significant coronary disease (≥70% stenosis) as compared to those with minimal coronary disease (less than 70% stenosis) or normal coronary arteries. Expression of miR-494, miR-490-3p, and miR-769-3p were found to have significantly different levels of expression. Using a multivariable regression model including cardiovascular risk factors and medications, hsa-miR-769-3p was found to be significantly correlated with the presence of significant coronary atherosclerosis. CONCLUSIONS: This study utilized a superior high-throughput qRT-PCR based method and found that miRNAs are found to be widely expressed in human blood with differences expressed between cellular and extracellular fractions. Importantly, specific miRNAs from circulating plasma are associated with the presence of significant coronary disease.