Cell Journal (Yakhteh)
Year:2011 | Volume:13 | Issue:2 (50)|Papers Count:9

Apoptosis, a form of programmed cell death that occurs under physiological as well as pathological conditions, is characterized by morphological and biochemical features. While the importance of caspases in apoptosis is established, several noncaspase proteases (Ca 2+-dependent proteases) such as calpain may play a role in the execution of apoptosis. The calpain family consists of two major isoforms, calpain I and calpain II which require mM and mM Ca 2+ concentrations to initiate their activity. An increase in intracellular Ca 2+ level is thought to trigger a cascade of biochemical processes including calpain activation.Once activated, calpains degrade membrane, cytoplasmic and nuclear substrates, leading to the breakdown of cellular architecture and finally apoptosis.The activation of calpain has been implicated in neuronal apoptosis following spinal cord injuries and neurodegenerative diseases. This review focuses on calpain with an emphasis on its key role in the proteolysis of cellular protein substrates following apoptosis.

Objective: This study evaluated in vitro maturation (IVM) of oocytes in the germinal vesicle (GV) stage in stimulated intracytoplasmic sperm injection (ICSI) cycles.Materials and Methods: A total of 26 women, aged 18 -37 years, who were candidates for ICSI at the Fatemeh Zahra Infertility and Health Reproductive Research Center in 2007 were recruited for this study. We used the standard long protocol for ovarian stimulation.Follicles>11 mm were punctured 36-38 hours after administration of 10000 IU human chorionic gonadotrophin (hCG). Immature oocytes were cultured for 24-30 hours. Oocytes that liberated polar bodies were injected by sperm prepared within the previous day. IVM fertilized oocytes were cultured an additional 24-30 hours for cleavage. The rates of maturation, fertilization and cleavage in IVM oocytes were recorded and statistically compared toin vivo matured sibling oocytes.Results: There were 279 collected oocytes (mean±SD: 10.73±6.2), of which 4.08±2.79 were subjected to IVM. An average of 2.73±2.15 GV oocytes (70%) developed to metaphase II (MII). Although the maturation rate significantly differed between the IVM andin vivoMII sibling oocyte groups (p=0.027), the numbers of fertilized oocytes (p=0.795) and cleaved embryos (p=0.529) were not significantly high in thein vivo group. Transfer of IVM embryos occurred in only three cases with one pregnancy that resulted in the delivery of a healthy baby.Conclusion: This study shows that culturing GV oocytes can produce acceptable numbers of four-cell embryos on the transfer day. The developmental competence of oocytes is not significantly different between early stage IVM andin vivo sibling embryos.

Objective: Ocular morbidity is widely observed when radiotherapy includes the orbit.Oxidative stress generated by irradiation is responsible for this complication. In different studies, it has been shown that melatonin has antioxidative properties and a radio protective role. The aim of this study was to evaluate the antioxidant role of melatonin against radiation-induced oxidative injury in rats' lenses after total cranial irradiation.Materials and Methods: Thirty-six adult female Sprague-Dawley rats were divided into six groups. Group I was the control group, group II only received total cranial gamma irradiation of 5 Gy, group III was exposed as the second group but at the dose of 8 Gy, group IV received 30 mg/kg melatonin 30 minutes prior to radiation plus total cranial irradiation of 5 Gy plus 5 mg/kg melatonin daily through intraperitoneal injection for ten days after irradiation, group V was treated similar to the fourth group, i.e. received irradiation plus melatonin, but at the dose of 8 Gy, and group VI only received melatonin (30 mg/kg on the first day and 5 mg/kg on the following days). Ten days after irradiation, all rats were sacrificed and their eyes were enucleated to measure the biochemical parameters i.e.malondialdehyde (MDA) and glutathione (GSH).Results: The levels of MDA in rat lenses increased and the levels of glutathione in lenses decreased after gamma ray irradiation but these parameters were still within normal limits in rats that received melatonin.Conclusion: It could be concluded that melatonin is useful in preventing radiation-induced oxidative injury due to its antioxidative and free radical scavenging properties.

Objective: In this study, we investigated the combined effect of 2-Methoxyestradiol (2ME2) and 60Co on the cytogenetic damage of iododeoxyuridine (IUdR) in the spheroid model of U87MG glioblastoma cancer cell lines by alkaline comet assay.Materials and Methods: U87MG cells were cultured as spheroids with diameters of 350mm. As control, the spheroids of one plate were not treated. Other cultures were pretreated with 2ME2 (250mM) for one volume doubling time (1 VDT). After this time, the subsequent treatments were performed according to the following groups: 1. Vehicle (this sample was not treated in the 2nd VDT) 2. Treated with 2ME2 (250mM) for 1 VDT 3. Treated simultaneously with 2ME2 (250mM) and IUdR (1 mM) for 1 VDT 4. Treated with 2ME2 (250mM) for 1 VDT then irradiated with 60Co (2 Gy) 5. Treated simultaneously with 2ME2 (250mM) and IUdR (1 mM) for 1 VDT then irradiated with 60Co (2 Gy) Then the DNA damage was evaluated using the alkaline comet assay method.Results: The results showed that 2ME2 in combination with gamma irradiation of 60Co significantly (p<0.001) increased the DNA damage by IUdR as compared to the control group. Thus the combination of these two agents increased the cytogenetic effects of IUdR in the spheroid culture model of U87MG glioblastoma cell lines.Conclusion: By inhibiting the HIF-1a protein and preventing the G0 phase arrest, 2ME2 causes an increased progression into S phase and increases the IUdR absorption. Then the DNA damage in the spheroid cells increases as the uptake of IUdR is increased.These results suggest that the combined use of 2ME2 and 60Co can increase the radiosensitization effect of IUdR.

Objective: Neuregulin1 (NRG1) gene is among the most promising candidate genes for schizophrenia. This gene is located on 8p22-p12, a region with a reported linkage to schizophrenia.Several studies have reported an association between schizophrenia and the 5' end polymorphisms in this gene. However, some studies have failed to confirm the role ofNRG1 gene in the pathogenesis of schizophrenia. In the current study, we attempt to examine the association of SNP8NRG241930 from theNRG1 gene with schizophrenia in an Iranian population. It is noteworthy that there has been no report on the NRG1 association with schizophrenia in a population from the Middle East region.Materials and Methods: Genomic DNA samples were obtained via isolation from the peripheral blood cells of 95 unrelated subjects with schizophrenia and 95 matched healthy controls from southwest Iran. SNP8NRG241930 was genotyped by PCRRFLP using ScaI as a restriction endonuclease enzyme. Association of the SNP with schizophrenia was examined using the chi-square test. The frequency difference of alleles and genotypes between the two groups were compared. P£0.05 was considered significant.Results: Statistical analysis on the studied polymorphism showed that both case and control groups were in Hardy-Weinberg equilibrium. The frequency of high risk allele (G allele) was 72.6% in patients, while this number was 56.8% in controls. The genotype frequencies in the patient group were as follows: GG (54%), GT (38%) and TT (8%) vs. genotype frequencies in the control group of: GG (26%), GT (63 %) and TT (11%).Conclusion: Considering allele and genotype frequencies, a significant association was observed between schizophrenia and SNP8NRG241930. The current study adds weight to the idea that some functional polymorphisms could exist in the 5′ end of the NRG1 gene which increase susceptibility to schizophrenia. This is the first time that supportive evidence shows an involvement of the NRG1 locus in schizophrenia in an Iranian sample population.

Objective: Global surveillance has shown that drug resistant (DR) tuberculosis (TB) is widespread. Prompt detection of Mycobacterium tuberculosis drug resistance is essential for effective control of TB. The most frequent mutations associated with Isoniazid (INH) resistance in Mycobacterium are substitutions at codons 315 in the katG gene and the mabA-inhA promoter region (-15). This survey evaluated INH resistant-associated mutations in order to determine rapid detection of TB resistance.Materials and Methods: Through a descriptive cross- sectional study total of 96 sputum specimens were digested, examined microscopically for acid-fast bacilli and inoculated into Löwenstein-Jensen slants. Thereafter, the susceptibility and identification tests were performed on culture positive specimens. Subsequently, the strains were subjected to multiplex allele-specific polymerase chain reaction (MAS-PCR) targeting in the codons 315 in the katG gene and the mabA-inhA promoter region. Distinct PCR banding patterns were observed for different mutation profiles.Results: Drug susceptibility testing revealed that out of 96 available isolates, 30 (31.3%) were susceptible, 36 (37.5%) had multi-drug resistance (MDR-TB) and 30 (31.3%) showed mono- drug resistance. In comparison with the culture-based phenotypic drug susceptibility test, the sensitivity and specificity of MAS-PCR assay for drug resistance-related genetic mutations were 76.7% and 71.4%, respectively. The correlation between MAS-PCR and culture-based phenotypic drug susceptibility testing findings was 99.4%.Conclusion: The profile of the isolates suggests a significant number of different DR strains with a high frequency of mutations at codon 315 of the katG gene. MAS-PCR provides a rapid, simple and cost-effective method for detecting MDR-TB.

Objective: Mitochondrial DNA (mtDNA) is a useful tool for population studies, identification of humans and forensic DNA studies. The existence of several hundreds copies of mtDNA per cell permit its extraction from minute or degraded samples. In addition, the level of polymorphism in the hypervariable (HV) region is high enough to permit its use in human identity testing. However, the presence of several heteroplasmy might lead to ambiguous results.Materials and Methods: This study was an experiental study. This study evaluated heteroplasmy in the HV region of mtDNA in blood samples of 30 Iranians who belonged to ten unrelated families from three sequential generations (grandmother, mother and daughter).Results: There were no heteroplasmic substitutions in the HV1 region, but analysis of HV2 showed heteroplasmic substitutions in two out ten families. In the first family the grandmother showed heteroplasmy (T/C) in nucleotide positions 146 and 151, however it was not detected in the mother and daughter. In second family, a triple heteroplasmy (T/C) was detected in the daughter in nucleotide positions 146, 151 and 295, but these heteroplasmic substitutions were not obvious in the grandmother and mother.Conclusion: Heteroplasmy in mtDNA is not a rare phenomenon and probably exists in everyone, but a triple heteroplasmy in one family member is a novel finding. Our results demonstrate that one or two sequence differences between samples in mtDNA do not warrant exclusion. In our study, the average nucleotide difference between unrelated persons in the HV2 region was 2.8 nucleotides, whereas there was a triple heteroplasmy in one person which was not obvious in her family.

Objective: Azadirachta indica (Neem) has been used traditionally for many centuries.Some impressive therapeutic qualities have been discovered. However, the therapeutic effect of neem leaf extract in 4T1 breast cancer has not been documented. The purpose of the present study is to investigate the therapeutic effect of ethanolic Neem leaf extract in anin vivo 4T1 breast cancer model in mice.Materials and Methods: A total of 84 female BALB/c mice were divided randomly into 7 groups (3 non-cancerous groups and 4 cancerous groups) consisting of 12 mice per group. The 3 non-cancerous groups were normal mice treated with 0.5% of Tween 20 in phosphate buffer saline (PBS) (NC), 250 mg/kg Neem (N250) or 500 mg/kg Neem (N500).The 4 cancerous groups were; cancer controls treated with 0.5% of Tween 20 in PBS (CC), and cancerous mice treated with 0.5μg/mL tamoxifen citrate (CT), 250 mg/kg Neem leaf extract (CN 250) or 500 mg/kg Neem leaf extract (CN 500). Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays were used to evaluate apoptosis (cell death) in the breast cancer tissues. SPSS software, version 14 was used for statistical analysis. Statistical significance was defined as p<0.05. Non parametric analysis of variance (ANOVA) was performed with the Kruskal Wallis test for the TUNEL assays.Parametric data among the groups was compared using ANOVA.Results: TUNEL assays showed that the CN 250 and CN 500 groups had a higher incidence of apoptosis compared with the cancer controls.Conclusion: The findings showed that neem leaf extract induces apoptosis in 4T1 breast cancer BALB/c mice.

Objective: Cartilage mass produced from mesenchymal stem cell (MSC) differentiation would be a suitable candidate for use in regenerative medicine. Since the proper function of cartilage tissue is largely dependent on matrix glycosaminoglycan (GAG) contents, the objective of this study was to investigate the enhancing effect of two GSK3 inhibitors on the GAG content of cartilage produced by human marrow MSCs in vitro chondrogenesis.Materials and Methods: MSCs that were used in this experimental study were derived from human marrow aspirates and confirmed using standard assays. Optimal concentrations of Lithium chloride and SB216763 were determined based on the yield of viable cell numbers in MSC cultures treated with varying concentrations of either Lithium chloride or SB216763. Passaged-3 MSCs were then centrifuged into small aggregates and provided with a chondrogenic medium supplemented with either lithium or SB216763 reagent at the optimal concentration determined in the previous experiment. Three weeks after, GAG contents of the culture were quantified and compared to each other and the control.Results: According to our data, the cultures treated with 5 mM Lithium and 1 mM SB216763 tended to have comparatively more viable cells; therefore these concentrations were used in the differentiation experiments. The addition of either SB216763 or lithium to chondrogenic cultures appeared to significantly enhance cartilage matrix production. In SB216763 and Lithium-treated cultures average GAG concentrations were 6.17±0.7 and 6.12±1.1mg/ml compared to 2.00±0.3 mg/ml in the control (p<0.05).Conclusion: Using SB216763 and Lithium as supplements in human marrow MSC chondrogenic culture can lead to the production of cartilage mass high in GAG content.