Iranian Biomedical Journal
Volume:17 Issue: 3, Jul 2013

Spinal cord has a limited capacity to repair; therefore، medical interventions are necessary for treatment of injuries. Transplantation of Schwann cells has shown a great promising result for spinal cord injury (SCI). However، harvesting Schwann cell has been limited due to donor morbidity and limited expansion capacity. Furthermore، accessible sources such as bone marrow stem cells have drawn attentions to themselves. Therefore، this study was designed to evaluate the effect of bone marrow-derived Schwann cell on functional recovery in adult rats after injury. Mesenchymal stem cells were cultured from adult rats’ bone marrow and induced into Schwann cells in vitro. Differentiation was confirmed by immunocytochemistry and RT-PCR. Next، Schwann cells were seeded into collagen scaffolds and engrafted in 3 mm lateral hemisection defects. For 8 weeks، motor and sensory improvements were assessed by open field locomotor scale، narrow beam، and tail flick tests. Afterwards، lesioned spinal cord was evaluated by conventional histology and immunohistochemistry. In vitro observations showed that differentiated cells had Schwann cell morphology and markers. In this study، we had four groups (n = 10 each): laminectomy، control، scaffold and scaffold + Schwann cells. Locomotor and sensory scores of cell grafted group were significantly better than control and scaffold groups. In histology، axonal regeneration and remyelination were better than control and scaffold groups. This study demonstrates that bone marrow-derived Schwann cells can be considered as a cell source for Schwann cells in SCI treatment.

The mitochondria are an important source of adenosine triphosphate (ATP) production in pre-implantation embryo. Therefore، the objective of this study was to investigate the effect of vitrification and in vitro culture of mouse embryos on their mitochondrial distribution and ATP content. The embryos at 2-PN، 4-cell and blastocyst stages were collected from the oviduct of stimulated pregnant mice and uterine horns. Then، the embryos were vitrified with the cryotop method using ethylene glycol and dimethylsulphoxide. After evaluating the survival rates of vitrified embryos، their development to hatching stages were assessed. The ATP content of collected in vivo and in vitro embryos at different stages was measured by luciferin-luciferase bioluminescence assay. The distribution of mitochondria was studied using Mito-tracker green staining under a fluorescent microscope. The survival rates of vitrified embryos at 2-PN، 4-cell and early blastocyst stages were 84. 3، 87. 87 and 89. 89%، respectively. The hatching rates in previous developmental stages in vitrified group were 57. 44، 66. 73 and 70. 89% and in non-vitrified group were 66. 32، 73. 25 and 75. 89%، respectively (P>0. 05) The ATP content of in vivo or in vitro collected embryos was not significantly different in both vitrified and non-vitrified groups (P>0. 05). Mitochondrial distribution of vitrified and non-vitrified 2-PN embryos was similar، but some clampings or large aggregation of mitochondria within the vitrified 4-cell embryos was prominent. Vitrification method did not affect the mouse embryo ATP content. Also، the cellular stress was not induced by this procedure and the safety of vitrification was shown.

Infectious by Pseudomonas aeruginosa has spread worldwide and metallo-beta-lactamases (MBL) are being reported with increasing frequency. The aim of this study was to investigate the antibiotic susceptibility and distribution of blaVIM and blaIMP genes in P. aeruginosa isolates from Zanjan Province of Iran. A total of 70 P. aeruginosa isolates were identified from patients admitted at intensive care units. The antimicrobial susceptibility was tested by disk diffusion (Kirby-Bauer) method and for production of MBL using double-disk synergy test (DDST). After DNA extraction، the presence of blaVIM and blaIMP genes and class 1 integron were detected by PCR. Most of the isolates were resistant to meropenem، cefotaxime and imipenem (IPM). Also، 44/70 (62. 85%) IPM resistant isolates were confirmed by DDST. Of the 44 clinical isolates، 41 (93%) isolates showed MIC≥4 µg/ml for IPM. Based on the DDST results، 36 (87. 8%) were confirmed to be MBL producers. PCR amplification showed that 23/41 (56%) carried blaVIM and 10/41 (24. 3%) possessed blaIMP gene. Also، 31/44 (70. 5%) isolates contained class 1 integron gene. Our results highlight that the genes for Verona integron-encoded metallo-β-lactamase، IPM β-lactamases and class 1 integrons were predominantly present among the IPM-resistant P. aeruginosa tested in our province and also the frequency of blaVIM type is higher than blaIMP. This is the first report of P. aeruginosa strains producing blaIMP with high frequency from Zanjan province of Iran.

Ritalin has high tendency to be abused. It has been the main indication to control attention deficit hyperactivity disorder. The college students may seek for it to improve their memory، decrease the need for sleep (especially during exams)، which at least partially، can be related to serotonergic system. Therefore، it seems worthy to evaluate the effect of Ritalin intake on mature brain. There are many studies on Ritalin effect on developing brain، but only few studies on adults are available. This study was undertaken to find Ritalin effect on serotonin transporter (SERT) density in medial frontal cortex (MFC) of mature rat. Thirty male Wistar rats were used in the study. Rats were assigned into five groups (n = 6 per group): one control، two Ritalin and two vehicle groups. Twelve rats received Ritalin (20 mg/kg/twice a day) orally for eleven continuous days. After one week of withdrawal and another two weeks of rest، in order to evaluate short-term effects of Ritalin، six rats were sacrificed. Another six rats were studied to detect the long-term effects of Ritalin; therefore، they were sacrificed 12 weeks after the previous group. The immunohistochemistry was performed to evaluate the results. Immunohistochemistry studies showed a higher density of SERT in both 2 and 12 weeks after withdrawal from Ritalin intake in MFC of rat and there was no significant difference between these two groups. Our findings demonstrated both short- and long-term effects of Ritalin on frontal serotonergic system after withdrawal period.

Bone marrow stromal cells (BMSC) have been successfully employed for movement deficit recovery in spinal cord injury (SCI) rat models. One of the unsettled problems in cell transplantation is the relative high proportion of cell death، specifically after neural differentiation. According to our previous studies، p75 receptor، known as the death receptor، is only expressed in BMSC in a time window of 6-12 hours following neural induction. Moreover، we have recently reported a decreased level of apoptosis in p75-suppressed BMSC in vitro. Therefore، our objective in this research was to explore the functional effects of transplanting p75: siRNA expressing BMSC in SCI rats.

Laminectomy was performed at L1 vertebra level to expose spinal cord for contusion using weight-drop method. PBS-treated SCI rats (group one) were used as negative controls، in which cavitations were observed 10 weeks after SCI. pRNA-U6. 1/Hygro- (group two، as a mock) and pRNA-U6. 1/Hygro-p75 shRNA- (group three) transfected BMSC were labeled with a fluorescent dye، CM-DiI، and grafted into the lesion site 7 days after surgery. The Basso-Beattie-Bresnehan locomotor rating scale was performed weekly for 10 weeks.

There was a significant difference (P≤0. 05) between all groups of treated rats regarding functional recovery. Specifically، the discrepancy among p75 siRNA and mock-transfected BMSC was statistically significant. P75 siRNA BMSC also revealed a higher level of in vivo survival compared to the mock BMSC.

Our data suggest that genetically modified BMSC that express p75: siRNA could be a more suitable source of cells for treatment of SCI.

Endothelial progenitor colony forming unit-endothelial cells (CFU-EC) were first believed to be the progenitors of endothelial cells، named endothelial progenitor cells. Further studies revealed that they are monocytes regulating vasculogenesis. The main hindrance of these cells for therapeutic purposes is their low frequency and limited replicative potentials. This study was undertaken to determine telomerase activity and alternative splicing variants in CFU-EC as a potential cause of limited replicative capacity in these cells. CFU-EC were isolated from peripheral blood using a standard cell culture assay. Colonies were detached mechanically and alternative splicing variant mRNA were evaluated using real-time PCR. Telomerase enzyme activity was assessed using telomerase repeat amplification protocol. The same procedures were done on the cancer cell line Calu6 as the positive control. The cultured peripheral blood mononuclear cells formed colonies with spindle-shaped monocytic cells sprouted from the clusters. These morphological characteristics fulfill the definition of CFU-EC. Telomere length amplification protocol assay revealed no telomerase activity and real-time PCR showed no expression of telomerase enzyme mRNA in CFU-EC. Both parameters were significantly higher in the cancer cell line Calu6 taken as the positive control. The absence of telomerase activity in the CFU-EC is a result of pre-transcriptional regulation of gene expression rather than other mechanisms for controlling telomerase activity such as post-transcriptional modifications. This finding can explain the limited proliferative activity of CFU-EC cells. We propose that absence of telomerase activity in CFU-EC can be attributable to their more mature monocytic nature that needs further investigations.

It is well known that the development of brain oxidative stress is one of the most serious complications of arterial hypertension that evokes brain tissue damage. The aim of this study was to examine the effects of atorvastatin treatment (20 mg/kg/day)، as an antioxidant، to prevent the brain tissue oxidative stress in the hypertensive (HTN) rats. Experiments were performed in four groups of rats (n = 5 each group): sham، sham-treated، HTN and HTN treated. Rats were made HTN by aortic constriction above the renal arteries. After 30 days، rats were slaughtered under deep anesthesia to remove brain hemispheres. After tissue homogenization، enzyme activities of superoxide dismutase (SOD) and catalase (CAT)، as well as glutathione (GSH) content and malondialdehyde (MDA) level were determined by biochemical methods. In HTN rats، arterial blood pressure was increased about 40% and brain enzyme activities of SOD and CAT were significantly decreased compared with sham group. Induction of hypertension significantly decreased GSH content and increased MDA level of brain tissue. Treatment with atorvastatin enhanced the activity of SOD and prevented from GSH decrement during hypertension. Based on the findings of this study، treatment with atorvastatin might have saved the brain tissue of HTN rats from hypertension-induced oxidative stress.

Human granulocyte colony-stimulating factor (hG-CSF) can serve as valuable biopharmaceutical for research and treatment of the human blood cancer. Transplastomic plants have been emerged as a new and high potential candidate for production of recombinant biopharmaceutical proteins in comparison with transgenic plants due to extremely high level expression، biosafety and many other advantages. hG-CSF gene was cloned into pCL vector between prrn16S promoter and TpsbA terminator. The recombinant vector was coated on nanogold particles and transformed to lettuce chloroplasts through biolistic method. Callogenesis and regeneration of cotyledonary explants were obtained by Murashige and Skoog media containing 6-benzylaminopurine and 1-naphthaleneacetic acid hormones. The presence of hG-CSF gene in plastome was studied with four specific PCR primers and expression by Western immunoblotting. hG-CSF gene cloning was confirmed by digestion and sequencing. Transplastomic lettuce lines were regenerated and subjected to molecular analysis. The presence of hG-CSF in plastome was confirmed by PCR using specific primers designed from the plastid genome. Western immunoblotting of extracted protein from transplastomic plants showed a 20-kDa band، which verified the expression of recombinant protein in lettuce chloroplasts. This study is the first report that successfully express hG-CSF gene in lettuce chloroplast. The lettuce plastome can provide a cheap and safe expression platform for producing valuable biopharmaceuticals for research and treatment.