A Comparative Analysis of Photobiomodulation-Mediated Biological Effects of Single Versus Double Irradiation on Dental Pulp Stem Cells: An In Vitro Study.

Objective: In recent years, fractionated irradiation protocols, rather than a simple plan of exposure, have been proposed as a more effective method in the field of tissue regeneration. Thus, this study aimed at a comparative analysis of single versus double irradiation of an 808-nm diode laser, in terms of dental pulp stem cells' (DPSCs) viability and proliferation in vitro. Methods: Subcultured DPSCs were either irradiated, or not (control group), with energy densities of 3, 7, and 12 J·cm-2 in a single- or double-session manner (24 h apart). On 0, 12, 24, 48, and 72 h postirradiation, cell viability and proliferation were evaluated through Trypan Blue and alamarBlue assays, respectively. Results: During the first 48 h postirradiation, the highest rates of DPSC proliferation were assigned to double irradiation at 3 or single exposure to 7 J⋅cm-2, with no cytotoxic effects on cell viability. Inversely, single irradiation at 12, or a double session of exposure to 7 or 12 J⋅cm-2, led to a significant descent in the rates of proliferation and cell viability. Conclusions: Within the limitations of this study, evidence suggests a positive impact on the biological responses of DPSCs following double session of exposure to lower energy densities as well as a single irradiation at a higher energy dosage.

G-CSF augments the neuroprotective effect of conditioned medium of dental pulp stem cells against hypoxic neural injury in SH-SY5Y cells.

Objectives: Dental pulp stem cells (DPSCs) can differentiate into functional neurons and have the potential for cell therapy in neurological diseases. Granulocyte colony-stimulating factor (G-CSF) is a glycoprotein family shown neuroprotective effect in models of nerve damage.we evaluated the protective effects of G-CSF, conditioned media from DPSCs (DPSCs-CM) and conditioned media from transfected DPSCs with plasmid encoding G-CSF (DPSC-CMT) on SH-SY5Y exposed to CoCl2 as a model of hypoxia-induced neural damage. Materials and Methods: SH-SY5Y exposed to CoCl2 were treated with DPSCs-CM, G-CSF, simultaneous combination of DPSCs-CM and G-CSF and finally DPSC-CMT. Cell viability and apoptosis were determined by resazurin (or lactate dehydrogenase (LDH) assay alternatively) and propidium iodide (PI) staining. Western blot analysis was performed to detect changes in apoptotic protein levels. The interleukin-6 and interleukin-10 IL6/IL10 levels were measured with Enzyme-Linked Immunosorbent Assay (ELISA). Results: DPSCs-CM and G-CSF were able to significantly protect SH-SY5Y against neural cell damage caused by CoCl2 according to resazurin and LDH analysis. Also, the percentage of apoptotic cells decreased when SH-SY5Y were treated with DPSCs-CM and G-CSF simultaneously. After transfection of DPSCs with G-CSF plasmid, DPSC-CMT could significantly improve the protection. The amount of ß-catenin, cleaved PARP and caspase-3 were significantly decreased and the expression of survivin was considerably increased when hypoxic SH-SY5Y treated with DPSCs-CM plus G-CSF according to Western blot. Decreased level of IL-6/IL-10, which exposed to CoCl2, after treatment with DPSCs-CM indicated the suppression of inflammatory mediators. Conclusion: Combination therapy of G-CSF and DPSCs-CM improved the protective activity.

Evaluation of the Cytotoxic and Apoptogenic Effects of Glabridin and Its Effect on Cytotoxicity and Apoptosis Induced by Doxorubicin Toward Cancerous Cells.

Purposes: In the present study, we tried for the first time to examine the anti-proliferative and anti-apoptogenic effect of Glabridin (Glab) toward three groups of cancer cells (SKNMC, H1299, and A2780). Furthermore, the possibility of co-administration of Glab with doxorubicin (DOX) to these cells was also examined to find out whether Glab can potentiate the cytotoxic effect of this chemotherapy agent. Methods: Different cellular assays (MTT, caspase-3 activity, MMP, RT-PCR analysis) were carried out on the cancer cells treated with Glab. Results: Cellular toxicity assay revealed that Glab can potentially reduce the viability of these cells with IC50 concentrations up to 10, 12, and 38 µM toward A2780, SKNMC, and H1299 cell lines, respectively. The results of MMP and caspase-3 activity assays, in association with the results corresponding to the BAX and Bcl-2 gene expressions, altogether revealed that Glab can exert apoptogenic effect on these cells. The intrinsic mitochondrial pathway was found to be the main mechanism, in which Glab induced apoptosis toward H1299 cells and SKNMC cells, while the apoptosis mechanism for A2780 cells could be probably through extrinsic pathway. Glab also potentiated the cytotoxic effect of DOX and its accumulation in H1299 cell line. Conclusion: The results of this study revealed the promising cytotoxic role of Glab on different carcinoma cells. These data also suggested that co-chemotherapy method using Glab could be effective for treatment of cancer, but further in-vivo and clinical studies are still needed to assure these results.

The Protective Effect of Different Extracts of Three Artemisia Species against H2O2-Induced Oxidative Stress and Apoptosis in PC12 Neuronal Cells.

BACKGROUND: Oxidative stress causes cell damage and is involved in many neurological diseases. The antioxidant properties of plant materials for the maintenance of health and protecting against different diseases stimulated scientist to investigate different herbs. Different Artemisia species have exhibited antioxidant activity. This study aims to investigate whether different Artemisia species could protect the PC12 cells against oxidative stress mediated by H2O2. METHODS: For this purpose, different extracts of three Artemisia species (Artemisia aucheri, Artemisia turanica, and Artemisia turcomanica) were prepared using petroleum ether, dichloromethane, ethyl acetate, ethanol, and Water: Ethanol mixture (1:1 volume ratio). The protective effect of the prepared extracts against H2O2-induced cytotoxicity and reactive oxygen species production were compared. The effect of treatment of PC12 cells with different extracts on total glutathione (GSH) level, caspase-3 activity, and mitochondrial membrane potential were also compared. RESULTS: The A. aucheri extracts could not rescue the PC12 cells from oxidative stress consequences. The A. turanica and A. turcomanica extracts were found potent in suppressing the toxicity and apoptosis of PC12 cells mediated by H2O2 and significantly antagonized the H2O2-induced GSH depletion. The hydroethanolic and ethyl acetate extracts of A. turanica and the petroleum ether and hydroethanolic extracts of A. turcomanica more efficiently suppressed cytotoxicity and loss of GSH caused by H2O2. CONCLUSION: This study shows the protective effects of Artemisia extracts on PC12 cell line and suggested that these species could be also considered as promising neuroprotective agents in treatment of different neurodegenerative diseases. SUMMARY: Artemisia turanica and Artemisia turcomanica extracts were found to potentially exert neuroprotective effect on PC12 cells. The results exhibited that the cytoprotective potential and anti-apoptotic mechanism of these species is not the same for different extracts, and suggested that based on the type of species and the type of solvents used in extraction, both intrinsic and extrinsic pathways could be included in the anti-apoptotic mechanism of these species. Abbreviations Used: GSH: Glutathion. ROS: Reactive Oxygen Species. GSSG: Glutathione disulfide. DCF-DA:2',7'-Dichlorofluorescin diacetate. FBS: Fetal Bovin Serum. MMP: Mitochondrial Membrane Potential. H-Et: Hydro-ethanolic. DCM: Dichloromethane. PE: Petroleum Ether. Et: Etanolic. EA: Ethyl Acetate.

Bioassay-guided Isolation of Neuroprotective Fatty Acids from Nigella sativa against 1-methyl-4-phenylpyridinium-induced Neurotoxicity.

OBJECTIVE: Parkinson's disease, a slowly progressive neurological disease, is associated with degeneration of the basal ganglia of the brain and a deficiency of the neurotransmitter dopamine. The main aspects of researches are the protection of normal neurons against degeneration. Fatty acids (FAs), the key structural elements of dietary lipids, are carboxylic straight chains and notable parameters in nutritional and industrial usefulness of a plant. MATERIALS AND METHODS: Black cumin, a popular anti-inflammatory and antioxidant food seasoning, contains nonpolar constituents such as FAs which were extracted using hexane. Different fractions and subfractions were apt to cytoprotection against apoptosis and inflammation induced by 1-methyl-4-phenylpyridinium (MPP+) in rat pheochromocytoma cell line (PC12) as a neural cell death model. The experiment consisted of examination of cell viability assessment, mitochondrial membrane potential (MMP), caspase-3 and -9 activity, and measurement of cyclooxygenase (COX) activity. RESULTS: MPP+ induced neurotoxicity in PC12 cells. Pretreatment with subfractions containing FA mixtures attenuated MPP+-mediated apoptosis partially dependent on the inhibition of caspase-3 and -9 activity and increasing the MMP. A mixture of linoleic acid, oleic acid, and palmitic acid also decreased the COX activity induced by MPP+ in PC12 cells. CONCLUSION: Our observation indicated that subtoxic concentration of FA from Nigella sativa may exert cytoprotective effects through their anti-apoptotic and anti-inflammation actions and could be regarded as a dietary supplement. SUMMARY: MPP+ induced neurotoxicity in PC12 cellsNigella sativa contains bioactive fatty acidsPretreatment with fatty acids attenuated MPP+ mediated apoptosis through inhibition of caspase 3 and 9 activityA mixture of linoleic acid, oleic acid, and palmitic acid decreased the COX activity induced by MPP+ in PC12 cellsDue to cytoprotective, anti apoptotic and anti inflammation actions of N. sativa, it could be regarded as a dietary supplement. Abbreviations used: ANOVA: Analysis of variance; Ca: Calcium; CDCl3: Chloroform; COX: Cyclooxygenase; DMSO: Dimethyl sulfoxide; EA: Elidic acid; EDTA: Ethylene diamine tetraacetic acid; ELISA: Enzyme Linked Immunosorbent Assay; ESI-MS: Electron spray mass spectroscopy; FAs: Fatty acids; FBS: Fetal bovine serum; GC: Gas chromatography; 1HNMR: Hydrogen nuclear magnetic resonance; LA: Linoleic acid; MPP+: 1-Methyl-4-phenylpyridinium; MPTP: 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine; MTT: 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide; N. sativa: Nigella sativa; OA: Oleic acid; PA: Palmitic acid; PBS: Phosphate buffer saline; PC12: Rat pheochromocytoma cell line; PD: Parkinson's disease; PDA: Photo diode array detector; PGE2: Prostaglandin E2; TLC: Thin layer chromatography; TMPD: N,N,N',N'-tetramethyl-p-phenylenediamine; USA: United states of America.

Protective effect of bioactive compounds from Echinophora cinerea against cisplatin-induced oxidative stress and apoptosis in the PC12 cell line.

OBJECTIVES: The present study aims to evaluate the protective effect of the compounds isolated from Echinophora cinerea (E. cinerea) against oxidative stress and apoptosis induced by cisplatin (CIS) in PC12 cells. MATERIALS AND METHODS: Six compounds were isolated as quercetrin-3-O-ß-D-glucopyranoside (QUE), osthol (OST), verbenone-5-O-ß-D-glycopyranoside (VER), Isoimperatorin (ISO), kaempferol-3-O-ß-D-glucopyranoside (KAM), and echinophorin B (ECH). For this study, we used MTT reduction assay for detection of protective effects of isolated compounds on CIS-induced cytotoxicity in PC12 cells. The effects of isolated compounds against apoptosis induced by CIS were investigated through the measurement of mitochondrial membrane potential (MMP), Bax and Bcl2 mRNA expression, and caspase-3 activation. We also assessed oxidative stress by measuring reactive oxygen species (ROS) generation with 2', 7'-dichlorofluorescein diacetate (DCFH-DA). RESULTS: Treatment of cells with QUE and OST before exposure to the CIS increased cell viability, i.e., these compounds protected the cells against CIS -induced cytotoxicity. In addition, pre-treatment with QUE and OST decreased CIS-induced apoptosis through up-regulation of Bcl-2, inhibition of caspase-3 activity, and mitochondrial membrane potential (MMP) increase. OST decreased ROS generation induced by CIS, as well. CONCLUSION: Our in vitro experiment showed that QUE and OST are apoptotic inhibitors that effectively block CIS-induced neurotoxicity predicting their therapeutic potential in the prevention of chemotherapy-induced neurotoxicity.

Synthesis and Biological Evaluation of N-(5-(pyridin-2-yl)-1,3,4-thiadiazol-2-yl)benzamide Derivatives as Lipoxygenase Inhibitor with Potential Anticancer Activity.

In the recent years, the role of LOX enzymes in the origin of neoplastic diseases such as colorectal, skin, pancreatic and renal cancers has been confirmed. A new series of 1,3,4-thiadiazole derivatives bearing 2-pyridyl moiety was synthesized and the cytotoxicity of the members of this series was assessed using MTT protocol. Enzyme inhibitory activity of the prepared compounds was also tested against 15-lipoxygenase-1 as a novel target for the discovery of anticancer drugs. PC3, HT29 and SKNMC cell lines were utilized and the obtained results were compared with doxorubicin. Overall, nitro containing derivatives exerted a higher cytotoxic activity against PC3 cell line and methoxylated derivatives showed an acceptable activity against SKNMC cell line. Methoxylated derivatives were also the most potent enzyme inhibitors especially at position ortho of the phenyl residue.

The Role of Plasma Creatine Phosphokinase (CPK) Level in Prediction of Response to Methotrexate for Ectopic Pregnancy.

OBJECTIVE: To evaluate the plasma creatine phosphokinase (CPK) level after injection of methotrexate (MTX) as a predictor of treatment success in ectopic pregnancy (EP). MATERIALS AND METHODS: One hundred women treated with single dose of methotrexate for ectopic pregnancy were evaluated in a prospective study, for CPK and ß-subunit of human chorionic gonadotropin (ß-hCG) levels. They received intramuscular MTX at a dose of 50 mg/m2. The day of injection was considered as day 1 (D1). CPK level on the day of Methotrexate injection was compared between success group who were treated by a single MTX injection, and the unsuccessful groupwho were treated by two or three MTX injections or by surgery. RESULTS: The success rate of single dose of MTX injection was 78 (78%). The mean of CPK was higher in success group than unsuccessful group. (86 ± 10.7 vs. 73 ± 11.8), the difference was significant (p = 0.04). The mean of ß-hCG was significantly lower in treatment success group than unsuccessful group (1421.3 ± 443.6 vs. 1925.6 ± 185.4, p = 0.01). CONCLUSION: The success of single dose of MTX treatment in ectopic pregnancy may be predicted by CPK levels and the higher levels of CPK may be useful for detecting of patients with successful response to MTX.

Protective effects of fractions from Artemisia biennis hydro-ethanolic extract against doxorubicin-induced oxidative stress and apoptosis in PC12 cells.

OBJECTIVES: This study was designed to indicate whether different fractions from Artemisia biennis hydroethanolic extract could provide cytoprotection against oxidative stress and apoptosis induced by doxorubicin (DOX) in rat pheochromocytoma cell line (PC12). MATERIAL AND METHODS: Cell viability was determined by MTT assay. Also, activation of caspase-3 and superoxide dismutase were evaluated by spectrophotometry. Detection of reactive oxygen species (ROS) and measurement of mitochondrial membrane potential (MMP) were performed by flowcytometry. RESULTS: Treatment of PC12 cells with DOX reduced viability dose dependently. For evaluation of the effect of fractions (A-G) on DOX-induced cytotoxicity, PC12 cells were pretreated for 24 hr with the A. biennis fractions and then cells were treated with DOX. The fractions C and D increased PC12 cells viability significantly compared to DOX treated cells. Moreover, pretreatment with fractions C and D for 24 hr attenuated DOX-mediated apoptosis and the anti-apoptotic action of A. biennis fractions was partially dependent on inhibition of caspase 3 activity and also increasing the mitochondrial membrane potential (MMP). Selected A. biennis fractions also suppressed the generation of ROS and increased superoxide dismutase (SOD) activity. CONCLUSION: Taken together our observation indicated that subtoxic concentration of aforementioned fractions of A. biennis hydroetanolic extract has protective effect against apoptosis induced by DOX in PC12 cell. The results highlighted that fractions C and D may exert cytoprotective effects through their antioxidant actions.

A semipolar fraction of petroleum ether extract of Artemisia aucheri induces apoptosis and enhances the apoptotic response to doxorubicin in human neuroblastoma SKNMC cell line.

Artemisia is an important genus of Iranian flora whose potent anti-proliferative effect has been demonstrated previously on human cancerous cell lines. In the current study, further fractionation was carried out on the petroleum ether extract of A. aucheri and their cytotoxic effects were evaluated on three human cancer cell lines. Cell viability was determined by 3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide assay. Real time polymerase chain reaction (RT-PCR) was used to evaluate the expression of apoptotic related genes. Activation of caspases and detection of intracellular doxorubicin (DOX) accumulation were evaluated using a spectrophotometer. Mitochondrial membrane potential (MMP) was measured using flow cytometry. The fraction NO-7 (F7) of petroleum ether extract showed the highest anti-proliferative effect, especially against SKNMC cells. Therefore, we focused on a description of the cytotoxic mechanism of the most potent fraction on SKNMC cells. The results indicated that F7 was able to induce apoptosis through MMP disruption, activation of caspases and increament of proapoptotic genes Bax and Smac/DIABLO. Moreover, our observation indicated that F7 is able to increase the cytotoxicity of DOX in SKNMC cells. The combination of F7+DOX significantly increased the intracellular accumulation of DOX. These results indicated that F7 induces apoptosis in SKNMC cells. Moreover, it might enhance the antitumor activity of DOX, through modulating the activity of multidrug resistant cancer cells and inducing apoptosis.

Phthalimide analogs as probable 15-lipoxygenase-1 inhibitors: synthesis, biological evaluation and docking studies.

BACKGROUND: Recent studies have been explained the role of lipoxygenases (LOX) in the origin of cancer. Among the lipoxygenases, the 5-LOX, 12-LOX and 15-LOX are more important in the cause of neoplastic disorders. In the present investigation, a new series of anticancer agents with 1,3,4-thiadiazole and phthalimide substructures were synthesized and their in vitro cytotoxicity was evaluated by MTT assay. Moreover, enzyme inhibitory potency was also assessed by enzymatic protocol towards 15-LOX-1. Molecular docking was performed to explore in silico binding mode of the target compounds. RESULTS: Tested compounds showed a better cytotoxic activity against HT29 cell line (colorectal cancer) in comparison with other cell lines (PC3: prostate carcinoma; SKNMC: neuroblastoma). Unfortunately, all of the tested derivatives rendered lower inhibitory potency than quercetin towards 15-LOX-1. Four hydrogen bonds were detected in docking studies for compound 4d as the most potent derivative in enzymatic assay. CONCLUSIONS: The biological results of reported compounds in this research were not so satisfactory. But, further structural modifications are necessary to improve the bioactivity of these derivatives.

Histopathologic and sonographic analysis of laparoscopic removal ovarian nonendometriotic cyst: the evaluating effects on ovarian reserve.

Currently, laparoscopic cystectomy is the first-line therapy for ovarian benign cysts that are resistant to current therapies. There are different studies that point to ovarian reserve damage due to laparoscopic cystectomy. In this study, we evaluate the ovarian damage following laparoscopic cystectomy for non-endometriosis cysts using ultrasound and pathology findings. This is a prospective cohort study conducted between 7 rd month of 2011 and 10th month of 2012 in Women hospital affiliated to Tehran university of medical sciences.45 non-endometriosis cysts (17 teratoma,7 mucinous, 10 simple serous and 11 simple cysts) underwent laparoscopic cystectomy with stripping technique. Amount of excised parenchyma, number of lost oocytes and cyst wall fibrosis thickness were histopathologically studied. Before and 3 months after surgery antral follicle count was evaluated by ultrasound. AFC after cystectomy for teratoma and simple serous was significantly reduced P<0.05. By larger teratomas and more parenchyma inadvertently removed during their excision (1.64, 0.255) reduced AFC was seen and in simple serous cysts with more removed parenchyma amount (1.5) reduced AFC occurred. In our study simple cysts excision led to a loss in AFC that was not associated with any other cyst parameters. Mucinous cysts resection led to no specific ovarian reserve damage. Laparoscopic cystectomy for non-endometriosis leads to reduced ovarian reserve.

Performance of pregnant women on folic acid intake.

The cause of neural tube defects (NTDs) is multifactorial and in this case folic acid has an important role. Since the neural tube is closed during 21-28 days of pregnancy, most of women are not informed about their pregnancy at this time, and as a result the golden time of folic acid consumption is missed. The aim of this study was evaluating the performance of pregnant women attending to Tehran Women's Hospital in regard to folic acid intake during pre-conceptional period between 2011 and 2012. This cross-sectional study was conducted in 370 pregnant women attending the prenatal clinic of a hospital affiliate to Tehran University of Medical Sciences between 2011 and 2012. Data were collected through interview using a questionnaire. Although 70% of the pregnancies were planned, but 70.5% of pregnant women had not taken folic acid before conception or in necessary time. There was found a significant relationship between level of education, history of abnormalities in children and the number of abortions and taking folic acid before pregnancy (P=0.005, P=0.000 and P=0.000, respectively).

Pregnancy outcome using general anesthesia versus spinal anesthesia for in vitro fertilization.

BACKGROUND: There is a considerable rate of fertility failure and this causes a great burden of untoward effects for patients. Usually a considerable number of these patients undergo anesthesia for their treatment. OBJECTIVES: This study was designed to compare the effects of general and spinal anesthesia on these patients. PATIENTS AND METHODS: In a randomized clinical trial, after taking informed written consent from the patients, 200 patients entered the study; 100 in each. During a 2 year period, women aged 20 to 40 years entered the study (one group receiving spinal anesthesia and the other, receiving general anesthesia). Ovum retrieval protocols were the same. Nonparametric and parametric analyses were used for data analysis. P value less than 0.05 was considered significant. RESULTS: There was no difference between the two groups regarding demographic variables. 15 of 100 patients (15%) in the general anesthesia group and 27 of 100 patients (27%) in the spinal anesthesia group had successful pregnancy after IVF; so, spinal anesthesia increased significantly the chance of IVF success (P value < 0.001; Chi Square). CONCLUSIONS: The results of this study demonstrated that spinal anesthesia increased the chance of fertilization success.

Extracts of Artemisia ciniformis Protect Cytotoxicity Induced by Hydrogen Peroxide in H9c2 Cardiac Muscle Cells through the Inhibition of Reactive Oxygen Species.

Objective. Artemisia ciniformis (Asteraceae) and A. biennis are two of 34 Artemisia species growing naturally in Iran. In this study we investigated whether different extracts of A. ciniformis and A. biennis have protective effect against hydrogen peroxide-induced cytotoxicity in rat cardiomyoblast cells (H9c2). Method. The dried and ground aerial parts of these two species were extracted successively using petroleum ether (40-60), dichloromethane, ethyl acetate (EA), ethanol (EtOH) and ethanol : water (1 : 1) by maceration method. To evaluate whether different extracts of A. ciniformis and A. biennis protect cardiomyoblast H9c2 cells from H2O2 cytotoxicity, we examined the direct cytotoxic effect of H2O2 on H9c2 cells in the presence and absence of different extracts. After then, cell viability was measured by MTT assay. Results. H2O2 induced cytotoxicity in a concentration dependent manner. The IC50 value was 62.5 µ M for 24 h exposure. However, pretreatment of cells with various concentrations of EA, EtOH, and EtOH/wt extract of A. ciniformis protected cells from H2O2-induced cytotoxicity. Moreover, pretreatment with EA, EtOH and EtOH/wt extracts of A. ciniformis lead to a decrease in the reactive oxygen species (ROS) generation. Taken together our observation indicated that nontoxic concentration of different extracts of A. ciniformis has protective effect on H2O2-induced cytotoxicity in H9c2 cells.