Exposure to Low-Frequency Radiation Changes the Expression of Nestin, VEGF, BCRP and Apoptosis Markers During Glioma Treatment Strategy: An In Vitro Study.

BACKGROUND: Exposure to physical contamination during chemotherapy, including non-ionizing electromagnetic fields, raises concerns about the widespread sources of exposure to this type of radiation. Glioblastoma multiforme (GBM) is an aggressive central nervous system tumor that is hard to treat due to resistance to drugs such as temozolomide (TMZ). OBJECTIVE: Electromagnetic fields (EMF) and haloperidol (HLP) may have anticancer effects. In this study, we investigated the effects of TMZ, HLP, and EMF on GBM cell lines and analyzed the association between non-ionizing radiation and the risk of change in drug performance. METHODS: Cell viability and reactive oxygen species (ROS) generation were measured by MTT and NBT assay, respectively. Then, the expression levels of breast cancer-resistant protein (BCRP), Bax, Bcl2, Nestin, vascular endothelial growth factor (VEGF) genes, and P53, Bax, and Bcl2 Proteins were evaluated by real-time PCR and western blot. RESULTS: Co-treatment of GBM cells by HLP and TMZ enhanced apoptosis in T-98G and A172 cells by increasing the expression of P53 and Bax and decreasing Bcl-2. Interestingly, exposure of GBM cells to EMF decreased apoptosis in the TMZ+HLP group. CONCLUSION: In conclusion, EMF reduced the synergistic effect of TMZ and HLP. This hypothesis that patients who are treated for brain tumors and suffer from depression should not be exposed to EMF is proposed in the present study. There appears to be an urgent need to reconsider exposure limits for low-frequency magnetic fields, based on experimental and epidemiological research, the relationship between exposure to non-ionizing radiation and adverse human health effects.

Comparing the effects of vitrification, before and after mouse oocyte in vitro maturation on developmental competence, changes in epigenetic regulators and stress oxidative response.

Since the developmental stage of oocyte is a challenging issue in the success of vitrification, this study investigated the effects of vitrification, before and after in vitro maturation, on the survival and maturation rates, developmental competence and the expression levels of genes involved in apoptosis, oxidative stress and epigenetic modifications. Mouse germinal vesicle (GV) oocytes were divided into four groups: fresh in vitro matured oocytes without vitrification (fIVM), in vitro matured oocytes after vitrification (vIVM), in vitro matured oocytes before vitrification (IVMv). In addition, in vivo matured oocytes (MII) were used as control. After oocytes collection, maturation and survival rates as well as the intracellular reactive oxygen species (ROS) level were evaluated. Also, the expression level of various genes was analyzed by qRT-PCR. In addition, following artificial activation (parthenogenesis), the developmental competence of oocytes to the blastocyst stage was evaluated. A significant decrease in maturation rate and survival of vIVM oocytes was observed compared to fIVM and IVMv oocytes. Intracellular ROS levels were significantly increased in both vitrified groups compared to the fIVM group, and no significant difference between vitrified groups. Pro-apoptotic genes; BAX and Bcl2 as well as genes related to oxidative stress response Hsp1a, Hsp1b and SOD1were significantly increased in the vIVM group compared to the IVMv group. Interestingly, epigenetic regulators genes DNMT1, DNMT3a and DNMT3b were highly expressed in IVMv oocytes along with a decrease in the artificial activation rate compared to the vIVM oocytes. Our results indicated that despite observing more negative effects of vitrification before IVM on the survival rate and maturation as well as apoptosis status, less epigenetic changes in vIVM oocytes can make this process a better option in the treatment of infertility than IVM of oocytes followed by vitrification, a hypothesis that needs to be investigation in human oocytes.

Impact of ketamine administration on chronic unpredictable stress-induced rat model of depression during extremely low-frequency electromagnetic field exposure: Behavioral, histological and molecular study.

OBJECTIVES: In the study, we examined the effects of ketamine and extremely low-frequency electromagnetic fields (ELF-EMF) on depression-like behavior, learning and memory, expression of GFAP, caspase-3, p53, BDNF, and NMDA receptor in animals subjected to chronic unpredictable stress (CUS). METHODS: After applying 21 days of chronic unpredictable stress, male rats received intraperitoneal (IP) of ketamine (5 mg/kg) and then were exposed to ELF-EMF (10-Hz, 10-mT exposure conditions) for 3 days (3 h per day) and behavioral assessments were performed 24 h after the treatments. Instantly after the last behavioral test, the brain was extracted for Nissl staining, immunohistochemistry, and real-time PCR analyses. Immunohistochemistry (IHC) was conducted to assess the effect of ketamine and ELF-EMF on the expression of astrocyte marker (glial fibrillary acidic protein, GFAP) in the CA1 area of the hippocampus and medial prefrontal cortex (mPFC). Also, real-time PCR analyses were used to investigate the impacts of the combination of ketamine and ELF-EMF on the expression of caspase3, p53, BDNF, and NMDA receptors in the hippocampus in rats submitted to the CUS procedure. Results were considered statistically significant when p < .05. RESULTS: Our results revealed that the combination of ketamine and ELF-EMF increased depression-like behavior, increased degenerated neurons and decreased the number of GFAP (+) cells in the CA1 area and mPFC, incremented the expression of caspase-3, and reduced the expression of BDNF in the hippocampus but showed no effect on the expression of p53 and NMDA-R. CONCLUSIONS: These results reveal that combining ketamine and ELF-EMF has adverse effects on animals under chronic unpredictable stress (CUS).

Pulsed and Discontinuous Electromagnetic Field Exposure Decreases Temozolomide Resistance in Glioblastoma by Modulating the Expression of O6-Methylguanine-DNA Methyltransferase, Cyclin-D1, and p53.

Background: Glioblastoma is a malignant and very aggressive brain tumor with a poor prognosis. Despite having chemotherapy concomitant with surgery and/or radiation therapy, the median survival of glioblastoma-affected people is less than 1 year. Temozolomide (TMZ) is a chemotherapeutic used as a first line treatment of glioblastoma. Several studies have reported that resistance to TMZ due to overexpression of O6-methylguanine-DNA methyltransferase (MGMT) is the main reason for treatment failure. Several studies described that pulsed-electromagnetic field (EMF) exposure could induce cell death and influence gene expression. Materials and Methods: In this study the authors assessed the effects of EMF (50 Hz, 70 G) on cytotoxicity, cell migration, gene expression, and protein levels in TMZ-treated T98 and A172 cell lines. Results: In this study, the authors show that treatment with a combination of TMZ and EMF enhanced cell death and decreased the migration potential of T98 and A172 cells. The authors also observed overexpression of the p53 gene and downregulation of cyclin-D1 protein in comparison to controls. In addition, T98 cells expressed the MGMT protein following treatment, while the A172 cells did not express MGMT. Conclusion: Their data indicate that EMF exposure improved the cytotoxicity of TMZ on T98 and A172 cells and could partially affect resistance to TMZ in T98 cells.

The prophylactic effect of date palm (Phoenix dactylifera L.) fruit extract on testicular toxicity induced by formaldehyde: An experimental study.

BACKGROUND: Formaldehyde (FA) is one of the most widely used materials in industries and in sciences. Prolonged contact with FA might have harmful effects on fertility due to the increase in the reactive oxygen species level. On the other hand, date palm (Phoenix Dactilifera L.) fruit extract (DPFE) contains a high concentration of natural antioxidants that could scavenge free radicals. Objective: The aim was to investigate the prophylactic effects of DPFE, with strong antioxidant properties, on FA-induced testicular toxicity in male mice. MATERIALS AND METHODS: Thirty-two adult NMRI male mice with a weight range of 25-35 gr (9-10 wk old) were randomly divided into four groups: control group (distilled water, orally for 35 days), FA group (FA; 0.25 mg/kg intraperitoneally (i.p.) for 20 days), treatment group (Date (DT) + FA; DPFE, 4 mg/kg for 35 days followed by FA administration, 0.25 mg/kg, i.p., for 20 days), date fruit extract group (DT; DPFE, 4 mg/kg, orally for 35 days). After this, blood was collected and left epididymis and testis tissues were isolated to evaluate the sperm parameters and histological examination, respectively. RESULTS: The FA administration increased the sperm morphological anomalies and reduced the sperm count, viability and motility, and also testosterone compared to the control group (p ≤ 0.001). In addition, histological studies of the testes showed that FA causes changes in the testis seminiferous tubules such as destruction of germinal epithelium and vacuolization of the tubules. The DPFE consumption before FA administration could partially ameliorate the reduced testosterone, sperm, and testicular parameters due to FA. CONCLUSION: The DPFE use might have discount effects on FA-induced testicular toxicity.

Comparison of detection percentage and morphology of myocardial bridge between conventional coronary angiography and coronary CT angiography.

Introduction: Myocardial bridge (MB) is a congenital anomaly in which a segment of a coronary artery is surrounded by myocardium. In our study, we want to use conventional coronary angiography (CCA) to describe morphologic characteristics of MB (unidentified or identified) in the patients with documented evidence of MB in coronary computed tomography angiography (CCTA). Methods: The present study was designed as cross-sectional and was conducted on 47 patients with documented evidence of MB in CCTA, who were referred to Nemazee and Faghihi hospitals for performing coronary angiography during a one year period. We compared the morphologic characteristics of tunneled segments, which were missed at CCA (unidentified), and the tunneled segments which were identified with CCA. Results: In sum, MB was found in 16 (34%) patients at CCA (identified), and it was not found in 31 (66%) patients (unidentified) based on compression sign. No significant correlation was found between the percentage of systolic compression and the length and depth of the tunneled segment in identified group (r=0.73, P = 0.18; r=1.09, P = 0.15; respectively). Degree of atherosclerotic plaque formation (diameter stenosis, percentage) (mean, 0.25 (25%) ±0.29; range, 0-0.98) of the tunneled segments in unidentified group was significantly more than the same degree (mean, 0.07 (7%) ±0.13; range, 0-0.41) of the identified group (P = 0.03). The measurement of the trapezoid area under the tunneled segment with this formula [(MB length+ intramyocardial segment) ×depth/2] had significant relation with systolic compression (r=0.304, P = 0.03) and defined the cut-off value of 250 mm2 as the value of significant difference in detecting myocardial bridging with CCA. Conclusion: Our results showed that in most of identified MBs in CCA the tunneled segment area was equal and more than 250 mm2. In addition, the degree of atherosclerotic plaque of the tunneled segments at CCA was significantly more in unidentified group.

The Effects of Olive Leaf Extract on The Testis, Sperm Quality and Testicular Germ Cell Apoptosis in Male Rats Exposed to Busulfan.

BACKGROUND: Busulfan (BU) has a destructive effect on the male reproductive system. The goal of this study was to assess the effects of olive leaf extract (OLE) as a source of antioxidants and phenolic compounds, on BU-induced damages in rat testes. MATERIALS AND METHODS: In this experimental study, 40 male Wistar rats were randomly divided into 5 groups. The control group (CTL) received a single intraperitoneal (i.p.) injection of dimethyl sulfoxide (DMSO), followed by oral administration of distilled water for 5 weeks. In BU group, BU (10 mg/kg) was administrated i.p. once. In cotreatment groups, first, received BU (10 mg/kg, a single i.p. injection) then, OLE was administrated orally at different doses of 250 mg/kg (BU+OLE 250), 500 mg/kg (BU+OLE 500) and 750 mg/kg (BU+OLE 750), for 5 weeks. Next, blood and sperm samples were collected. The left testis was removed to investigate testicular parameters and apoptosis by using H and E and TUNEL staining, respectively. All data were analyzed by SPSS software and a P<0.05 was considered significant. RESULTS: There was a significant decline in sperm viability (P=0.017), number of primary spermatocyte (PS) (P=0.001) and Leydig cells (P=0.023) in the BU group versus the CTL group. OLE at three doses could repair these defects versus BU group. Increases in apoptotic spermatogonia cells (SG) due to BU were significantly reduced by OLE 250 and 500 mg/kg (P<0.01). A reduction in germinal epithelium height and an increase in apoptotic SG were observed in BU+OLE 750 group vs. other groups (P<0.01) and alkaline phosphatase (ALP) was at the highest level, also Aspartate aminotransferase (AST) increased markedly vs. CTL (P=0.024). CONCLUSION: Oral administration of OLE at the doses of 250 and 500 mg/kg could be helpful in ameliorating BUinduced toxicity in rat testes, while OLE 750 mg/kg not only did not cause positive effects, but also could exacerbate the harmful effects.

Induction of cross-tolerance between protective effect of morphine and nicotine in 6-hydroxydopamine-induce neurotoxicity in SH-SY5Y human dopaminergic neuroblastoma cells.

PURPOSE: Parkinson's disease is a progressive neurodegenerative disease characterized by progressive and selective death of dopaminergic neurons. It has been reported that nicotine and morphine have protective roles during neuronal damage in Parkinson's disease. In addition, the induction of cross-tolerance between their biological effects has been shown in numerous reports. METHODS: Here, we investigated the effects of nicotine and morphine on 6-OHDA-induced neurotoxicity in human neuroblastoma SH-SY5Y cell line as an in vitro model of Parkinson's disease. Cell damage was induced by 150 µM 6-OHDA and the cells viability was examined by MTT assay. Intracellular reactive oxygen species, calcium level, and mitochondrial membrane potential were determined by fluorescence spectrophotometer method. Biochemical markers of apoptosis were also evaluated by immunoblotting. RESULT: The data showed that morphine and nicotine prevent 6-OHDA- induced cell damage and apoptosis. However, the protective effects of nicotine were not observed in chronic morphine-pretreated cells. Morphine had no protective effects in chronic nicotine-incubated cells. CONCLUSION: A cross-tolerance between protective effects of morphine and nicotine was occurred in 6-OHDA-induced SH-SY5Y cell toxicity.

Assessment of mouse oocytes ultrastructure following vitrification before and after in vitro maturation / Evaluación de la ultraestructura de los ovocitos de ratón después de la vitrificación antes y después de la maduración in vitro

SUMMARY: Vitrification is a physical process in which the concentrated cryoprotectant solution after exposure to extreme cold without ice crystal formation in living cells to be converted glassing state. In this study, maturation rate and ultrastructure of mouse oocytes followed by vitrification before or after in-virto maturation (IVM) were evaluated. A total of 373 germinal vesicle oocytes were obtained from ovaries and divided into three fresh IVM, IVM vitrified, vitrified IVM groups. Ten metaphase II oocytes were obtained from uterine tubes and considered as the control group. Oocytes in vitrified groups were vitrified by Cryotop using vitrification medium and kept in liquid nitrogen. The maturation media was a-MEM supplemented with rFSH + hCG. After 24-48 h of incubation, the oocytes were investigated for nuclear maturation and ultrastructural changes using transmission electron microscopy (TEM). The oocyte maturation rate in vIVM group was significantly lower than IVMv group, when the two groups were compared with vIVM had the highest maturity. The evaluation ultrastructure of the four groups showed that the number of cortical granules, microvilli and mitochondria-SER aggregates in vIVM group were lowest and the highest amongst the number of vacuoles. Zona pellucida was darker than the control group in two freeze groups vIVM and IVMv. Most similar groups to the control group were group vIVM, Group IVMv and ultimately vIVM group, respectively. According to the results, IVM procedure is more efficient when it is performed before oocyte vitrification.

RESUMEN: La vitrificación es un proceso físico en el que la solución concentrada de crioprotectores, después de la exposición al frío extremo sin formación de cristales de hielo en las células vivas, se convierte en estado de cristal. En este estudio, se evaluaron la velocidad de maduración y la ultraestructura de los ovocitos de ratón seguidos por la vitrificación antes o después de la maduración in vitro (IVM). Se obtuvieron un total de 373 ovocitos, de vesículas germinales de ovarios, y se dividieron en tres grupos de IVM vitrificados, IVM e IVM frescos. Diez ovocitos metafase II se obtuvieron a partir de tubas uterinas y se consideraron como el grupo de control. Los ovocitos en grupos vitrificados fueron vitrificados por Cryotop usando medio de vitrificación y mantenidos en nitrógeno líquido. El medio de maduración fue a-MEM suplementado con rFSH + hCG. Después de 24-48 h de incubación, fueron observados en los ovocitos la maduración nuclear y cambios ultraestructurales utilizando microscopía electrónica de transmisión (MET). La tasa de maduración de los ovocitos en el grupo vIVM fue significativamente más baja que en el grupo IVM v, cuando los dos grupos se compararon con los que tenían la mayor madurez. La evaluación de la ultraestructura de los cuatro grupos mostró que el número de gránulos corticales, microvellosidades y acúmulos de mitocondrias-SER en el grupo vIVM fue el más bajo y el más alto entre el número de vacuolas. La zona pelúcida fue más oscura en dos grupos de congelación vIVM e IVMv, que en el grupo control. La mayoría de los grupos, similares al grupo de control, fueron los grupos vIVM, IVMv y,finalmente, el grupo vIVM, respectivamente. De acuerdo con los resultados, el procedimiento de IVM es más eficiente cuando se realiza antes de la vitrificación de ovocitos.

Increased risk of coronary perforation during percutaneous intervention of myocardial bridge: What histopathology says.

Introduction: Myocardial bridge (MB) is a segment of a major epicardial coronary artery that goes intramurally under a bridge of overlying myocardium. Complications have been reported during or after stent implantation particularly coronary perforation. The aim of this study was to determine histological differences between proximal left anterior descending artery (LAD) and the tunneled segment that may have a possible role in increased risk of coronary artery perforation during percutaneous coronary intervention. Methods: Twenty specimens of MB were obtained from dissection of 45 cadavers. Sections were stained using hematoxylin and eosin (H&E), and trichrome methods. The proximal section and the tunneled artery were compared with a normal sample in terms of the characteristics of a muscle artery. Results: The findings of this study showed an MB prevalence of 51%, as 23 out of the 45 examined cadavers were discovered to be afflicted by the MB. The intima layer in the suffering artery had gone through significant hypertrophy, while it had remained thin in the tunneled artery section. The epithelial cells under the bridge were spindle-shaped, while they were polygonal in the proximal section. In the myocardium the nuclei of the muscle fibers in the MB section were smaller than the normal section. Adventitial layer was almost normal. Conclusion: The histopathological differences between MB and proximal part of vessel combined with small vessel diameter in the tunneled segment can explain the high incidence of the LAD rupture and perforation in the section under the bridge.

Anthropometric Analysis of Cephalofacial Dimensions in Kerman, Iran.

The human body dimensions are affected by ecological, biological, geographical, racial, sex, and age factors. Craniofacial measurements can be considered to be one of the important tools for determination of the morphological characteristics of the head and face. In this study, which was conducted on Persian adolescents living in Kerman/Iran, different forms of head and face were determined for using in various aspects of medicine. The study was conducted on 732 participants including 366 males and 366 females in the age of 18-20-year-old. In addition to the height and weight of the participants, cephalofacial sizes of them were measured and then cephalic, facial, and brain indices were calculated. Among the cephalofacial sizes, cranial length and breadth, cranial circumference, prosopic length and prosopic breadth were significantly greater in males compared to females (P<0.005). Also, volume and weight of brain were significantly greater in male comparing to female participants (P<0.005). The predominant type of head was meso-cephal, and the predominant type of face was meso-prosopic in both sexes.

Mesenchymal Stem Cell-Conditioned Medium Modulates Apoptotic and Stress-Related Gene Expression, Ameliorates Maturation and Allows for the Development of Immature Human Oocytes after Artificial Activation.

The aim of the present study was to determine whether mesenchymal stem cell-conditioned medium (MSC-CM) modulates apoptotic and stress-related gene expression, and ameliorates maturation and developmental potential of immature human oocytes after artificial activation. A total of 247 surplus immature germinal vesicle (GV) oocytes obtained from infertile women were allocated into two in vitro maturation (IVM) groups: 1: GV oocytes (n = 116) matured in vitro (fIVM), and 2: GV oocytes (n = 131) that were vitrified, then in vitro matured (vIVM). Also, two maturation media were used: Alpha-minimum essential medium (α-MEM) and human umbilical cord-derived MSCs (hUCM). After 36 h of incubation, the IVM oocytes were examined for nuclear maturation. In IVM-matured oocytes, cytoplasmic maturation was evaluated after artificial activation through Ionomycin. Moreover, the quantitative expressions of B-cell CLL/lymphoma 2 (BCL2), BCL2-associated X protein (BAX), superoxide dismutase (SOD), and Heat shock proteins (HSP70) in matured oocytes were assessed by quantitative Real-time polymerase chain reaction (qRT-PCR) and compared with fresh and vitrified in vivo matured oocytes, which were used as fIVM and vIVM controls, respectively. The highest maturation rate was found in hUCM in fIVM, and the lowest maturation rate was found using α-MEM in vIVM (85.18% and 71.42%, respectively). The cleavage rate in fIVM was higher than that in vIVM (83.4% vs. 72.0%). In addition, the cleavage rate in α-MEM was lower than that in the hUCM (66.0% vs. 89.4%). Furthermore, the difference between parthenote embryo arrested in 4-8 cells (p < 0.04) and the quality of embryo arrested in 8-cell (p < 0.007) were significant. The developmental stages of parthenote embryos in hUCM versus α-MEM were as follows: 2-4 cell (89.45% vs. 66.00%, respectively), 4-8 cell (44.31% vs. 29.11%, respectively), morula (12.27% vs. 2.63%, respectively), and blastocysts (2.5% vs. 0%, respectively). The messenger RNA (mRNA) expression levels of BCL2, BAX and SOD were significantly different (p < 0.05) between the matured IVM oocytes. Overall, hUCM showed potential efficacy in terms of ameliorating oocyte maturation and in promoting the development and mRNA expression of BAX, BCL2, and SOD.

Melatonin treatment reduces astrogliosis and apoptosis in rats with traumatic brain injury.

OBJECTIVES: Melatonin is known as an anti-inflammatory agent, and it has been proven to exert neuroprotection through inhibition of cell death (apoptosis) in several models of brain injury. Secondary injury following the primary traumatic brain injury (TBI) results in glial cells activation, especially astrocytes. In fact, astrocyte activation causes the production of pro-inflammatory cytokines that may lead to secondary injury. Since most TBI research studies have focused on injured neurons and paid little attention to glial cells, the aim of current study was to investigate the effects of melatonin against astrocytes activation (astrogliosis), as well as inhibition of apoptosis in brain tissue of male rats after TBI. MATERIALS AND METHODS: The animals were randomly allocated into five groups: sham group, TBI+ vehicle group (1% ethanol in saline) and TBI+ melatonin groups (5 mg/kg, 10 mg/kg and 20 mg/kg). All rats were intubated and then exposed to diffuse TBI, except for the sham group. Immunohistochemical methods were conducted using glial fibrillary acidic protein (GFAP) marker and TUNEL assay to evaluate astrocyte reactivity and cell death, respectively. RESULTS: The results showed that based on the number of GFAP positive astrocytes in brain cortex, astrogliosis was reduced significantly (P<0.05) in melatonin- treated groups (no dose dependent) compared to the vehicle group. Furthermore, based on TUNEL results, melatonin treatment considerably reduced the number of apoptotic cells (P<0.05). CONCLUSION: In total, the present findings suggest that melatonin treatment following TBI diminishes astrocyte reactivity and neuronal cells apoptosis in brain cortex in the rat model.

Evaluation of neurogenic potential of human umbilical cord mesenchymal cells; a time- and concentration-dependent manner.

BACKGROUND: Retinoic acid as one of the most important regulators for cell differentiation was examined in this study for differentiation of human umbilical mesenchymal cells (hUCM). METHODS: After isolation, hUCM were evaluated for mesenchymal stem cell properties by flow cytometry and alkaline phosphatase assay. Also, doubling time of the cells and their differentiation potential into adipogenic and osteogenic cells were tested. hUCM were then cultured with different concentrations of retinoic acid, and on days 1, 7, and 12, the percentage of differentiated cells was determined by immunostaining for nestin, anti-microtubule associated protein 2 (MAP2), glutamic acid decarboxylase (GAD), and gamma-aminobutyric acid (GABA) markers. RESULTS: The isolated cells were negative for the hematopoietic markers and positive for the mesenchymal markers. They showed the population doubling time 60 ± 3 hours and differentiated into osteogenic and adipogenic cells. A descending trend in nestin and an ascending trend in MAP2, GAD, and GABA expression were observed from the first day until the last day between different concentrations of retinoic acid. CONCLUSION: hUCM cells may have the potential to differentiate into neural cells in the presence of different incubation period and concentration of retinoic acid.

Morphine protects SH-SY5Y human neuroblastoma cells against 6-hydroxydopamine-induced cell damage: involvement of anti-oxidant, calcium blocking, and anti-apoptotic properties.

Parkinson's disease is a neurodegenerative disorder characterized by progressive and selective death of dopaminergic neurons. Understanding the neuroprotective effects of chemical reagents has attracted increasing attention. The µ opioid agonist morphine exerts both toxic and protective effects. However, until recently, the neuroprotective role of morphine against 6-hydroxydopamine (6-OHDA)-induced cell death has not been studied. Here, we investigated the effects of morphine on 6-OHDA-induced neurotoxicity in human neuroblastoma SH-SY5Y cell line as an in vitro model of Parkinson's disease. Cell damage was induced by 150 µM 6-OHDA, and the cells' viability was examined by MTT assay. Intracellular calcium, reactive oxygen species (ROS), and mitochondrial membrane potential were determined by the fluorescence spectrophotometry method. Fragmented DNA and biochemical markers of apoptosis were also determined by gel electrophoresis and immunoblotting, respectively. The data showed that 6-OHDA caused a loss of cell viability and mitochondrial membrane potential. In addition, intracellular ROS and calcium levels, activated caspase-3, Bax:Bcl-2 ratio, cytochrome c release, as well as DNA fragmentation were significantly increased in 6-OHDA-treated cells. Incubation of SH-SY5Y cells with morphine (100 µM) elicited a protective effect and reduced biochemical markers of cell damage and death. These results suggest that morphine has neuroprotective effects against 6-OHDA-induced neurotoxicity, and such effects are accompanied by its anti-oxidant, calcium blocking, and anti-apoptotic properties.