Iranian Journal of Basic Medical Sciences
Year:2015 | Volume:18 | Issue:9|Papers Count:15

Pigment epithelium‐derived factor (PEDF) is a glycoprotein that belongs to the family of noninhibitory serpins. The broad spectrum of PEDF biological activity is evident when considering its effects in promoting cell survival and proliferation, as well as its antiangiogenic, antitumor, and antimetastatic properties. Although the structural domains of the PEDF gene that mediate such diverse effects and their mechanisms of action have not been completely elucidated, there is a large body of evidence describing their diverse range of activities; this evidence combined with the regulation of PEDF expression by sex steroids and their receptors have led to the idea that PEDF is not only a diagnostic and prognostic marker for certain diseases such as cancer, but is also a potential therapeutic target. In this manner, this paper aims to generally review the regulation of PEDF expression and PEDF interactions, as well as the findings that relate PEDF to the role of estrogens and estrogen receptors. In addition, this manuscript will review major advances toward potential therapeutic applications of PEDF.

Objective (s): Transforming growth factor‐ b (TGF‐ b) is involved in colorectal cancer (CRC). The SD‐208 acts as an anti‐cancer agent in different malignancies via TGF‐ b signaling. This work aims to show the effect of manipulation of TGF‐Ò signaling on some miRNAs implicated in CRC.Materials and Methods: We investigated the effects of SD‐208 on SW‐48, a colon adenocarcinoma cell line. The cell line was treated with 0.5, 1 and 2 mM concentrations of SD‐208. Then, the xenograft model of colon cancer was established by subcutaneous inoculation of SW‐48 cell line into the nude mice. The animals were treated with SD‐208 for three weeks. A quantitative real‐time PCR was carried out for expression level analysis of selected oncogenic (miR‐21, 31, 20a and 135b) and suppressormiRNAs (let7‐g, miR‐133b, 145 and 200c). Data were analyzed using the 2‐DDCT method through student’s t‐test via the GraphPad Prism software.Results: Our results revealed that SD‐208 could significantly down‐regulate the expression of one key onco‐miRNA, miR‐135b, in either SW‐48 colon cells (P=0.006) or tumors orthotopically implanted in nude mice (P=0.018). Our in silico study also predicted that SD‐208 could modulate the expression of potential downstream tumor suppressor targets of the miR135b.Conclusion: Our data provide novel evidence that anticancer effects of SD‐208 (and likely other TGF‐ b inhibitors) may be owing to their ability to regulate miRNAs expression. Article history:

Objective (s): Sulfur mustard (SM) was used by the Iraqi army against the Iranian troops in the Iran‐Iraq war from 1983–1988. This chemical gas affects different organs including the skin, lungs and the hematopoietic system. Any exposure to SM increases the risk of chromosomal breaking, hyperdiploidy and hypodiploidy. Studies have shown that the risk for acute myeloblastic and lymphoblastic leukemia increases in veterans exposed to SM. FLT3 mutations including ITD and TKD mutations had been observed in some cases of leukemia. Therefore, we aimed to investigate the frequency of FLT3‐ TKD835 mutations in the veterans exposed to SM agent.Materials and Methods: We studied 42 patients who were exposed to SM during the war in Khorasan Razavi province, Mashhad, Iran in 2012. As control group, 30 healthy males were selected from firstdegree relatives of the patients. For assessment of TKD835 mutation, DNA was extracted and RFLPPCR was performed.Results: Analysis of RFLP‐PCR data showed no FLT‐3 TKD mutation in any of the patients. Conclusion: Although contact with SM can increase the risk of malignancy especially hematologic neoplasms, results of the study show that another mechanism of leukemogenesis, other than FLT3‐ TKD mutation, may be the reason for increased risk of leukemia in SM toxicity.

Objective (s): 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.

Objective (s): Anodized treated titanium surfaces, have been proposed as potential surfaces with better cell attachment capacities. We have investigated the adhesion and proliferation properties of H9C2 cardiac myoblasts on anodized treated titanium surface.Materials and Methods: Surface topography and anodized tubules were examined by high‐resolution scanning electron microscopy (SEM). Control and test substrates were inserted to the bottom of 24‐ well tissue culture plates. Culture media including H9C2 cells were loaded on the surface of substrate and control wells at the second passage. Evaluation of cell growth, proliferation, viability and surface cytotoxicity was performed using MTT test. After 48 hr, some samples were inspected by SEM. DAPIstaining was used to count attached cells.Results: MTT results for cells cultured on anodized titanium and unanodized titanium surfaces was equal to 1.56 and 0.55 fold change compared to tissue culture polystyrene (TCPS). The surface had no cytotoxic effects on cells. The average cell attachment to TCPS, unanodized and anodized titanium surface was 2497±40.16, 1250±20.11 and 4859.5±54.173, respectively. Cell adhesion to anodized titanium was showed 1.95 and 3.89 fold increase compared to TCPS and unanodized titanium, respectively (P<0.05).Conclusion: Anodized titanium surfaces can be potentially applied for enhanced recruitment of H9C2 cells. This unique property makes these inexpensive anodized surfaces as a candidate surface for attachment of cardiac cells and consequently for cardiac regeneration purposes.

Objective (s): The aim of the study was to investigate the relationship between insulin resistance (IR) and leptin (LEP) gene polymorphisms in peritoneal dialysis (PD) patients.Materials and Methods: From July 1, 2011 to August 1, 2011, patients who received chronic PD were chosen and divided into three groups (DM, high HOMR‐IR, and low HOMR‐IR). Two PCR products of LEP were sequenced and aligned and the distribution of polymorphisms was analyzed using χ2 analysis. In addition, serum leptin level, PD conditions, and biochemical parameters according to different genotype of G‐2548A and A19G were statistically analyzed (P‐value<0.05). The relationship between LEP gene polymorphisms and prognosis was explored.Results: Totally 157 patients with average age of 55±15 years old were chosen. Distribution of genotype frequencies was complied with Hardy‐Weinberg equilibrium. Leptin level and BMI (body mass index) of the GG genotype of G‐2548A were higher than that of GA or AA. The fasting glucose, cholesterol, etc. of AA genotype were lower, and the nPCR was higher than the two other genotypes. Serum leptin level and BMI of AA genotype of A19G was higher than GA and GG genotypes; meanwhile, fasting blood glucose of that genotypes was the highest. In addition, survival rate of AA group of A19G was very low.Conclusion: The G‐2548A and A19G polymorphisms were correlated with serum leptin level and IR. Leptin A19G polymorphism may be prognostic for PD patients. This study may facilitate early intervention for IR in PD patients.

Objective (s): Various methods for repairing bone defects are presented. Cell therapy is one of these methods. Bone marrow stromal cells (BMSCs) seem to be suitable for this purpose. On the other hand, lots of biomaterials are used to improve and repair the defect in the body, so in this study we tried to produce a similar structure to the bone by the chitosan and hydroxyapatite.Materials and Methods: In this study, the solution of chitosan‐nanohydroxyapatite‐polyethylene oxide (PEO) Nanofibers was produced by electrospinning method, and then the BMSCs were cultured on this solution. A piece of chitosan‐nanohydroxyapatite Nanofibers with BMSCs was placed in a hole with the diameter of 1 mm at the distal epiphysis of the rat femur. Then the biomechanical and radiographic studies were performed.Results: Biomechanical testing results showed that bone strength was significantly higher in the Nanofiber/BMSCs group in comparison with control group. Also the bone strength in nanofiber/BMSCs group was significant, but in nanofiber group was nearly significant. Radiographic studies also showed that the average amount of callus formation (radio opacity) in nanofiber and control group was not significantly different. The callus formation in nanofiber/BMSCs group was increased compared to the control group, and it was not significant in the nanofiber group.Conclusion: Since chitosan‐nanohydroxyapatite nanofibers with BMSCs increases the rate of bone repair, the obtained cell‐nanoscaffold shell can be used in tissue engineering and cell therapy, especially for bone defects.

Objective (s): The neuropeptide calcitonin gene‐related peptide (CGRP) has long been postulated to play an integral role in the pathophysiology of migraine. Earlier studies showed that CGRP can stimulate the synthesis and release of nitric oxide (NO) and cytokines from trigeminal ganglion glial cells. The purpose of this study was to determine the effect of 17b‐estradiol in regulation of CGRP expression, inducible nitric oxide synthase (iNOS) activity, and NO and interleukin‐1beta (IL‐1 b) release in cultured peripheral blood mononuclear cells (PBMCs) from patients with pure menstrual migraine and healthy individuals.Materials and Methods: This study was performed on twelve patients with pure menstrual migraine and twelve age‐and sex‐matched healthy individuals. PBMCs treated with 17 b ‐estradiol for 24 hr at physiological and pharmacological doses. Gene expression was evaluated by real time‐PCR. CGRP and IL‐1 b proteins in culture supernatant were determined by ELISA method. Activity of iNOS in PBMCs and total nitrite in the culture supernatant were measured by colorimetric assays.Results: Treatment with 17 b ‐estradiol had a biphasic effect on expression of CGRP. We found that 17 b ‐estradiol treatment at pharmacological dose significantly increases mRNA expression of CGRP in both groups (P<0.001), whereas at physiological dose it could significantly decrease CGRP mRNA expression (P<0.001), CGRP protein levels, IL‐1 b release, NO production and iNOS activity only in patient groups (P<0.05).Conclusion: Collectively, it appears that 17 b ‐estradiol can exert protective effect on decrease of inflammation in migraine via decrease in levels of CGRP, IL‐1 b and iNOS activity; however, more studies are necessary in this regard.

Objective(s): Acrylamide (ACR) has many applications in different industries. ACR damages the central and the peripheral nervous system in human and animals. Importance of ACR‐induced neurotoxicity encouraged researchers to find both different mechanisms involved in ACR neurotoxicity and potent neuroprotective agents. Therefore, this study was designed to investigate the protective effect of crocin, an active constituent ofCrocus sativus L. (saffron) on ACR‐induced neurotoxicity in Wistar rats.Materials and Methods: Animals were treated with ACR (50 mg/kg, IP) 11 days for induction neurotoxicity. Crocin (12.5, 25 and 50 mg/kg, IP) were used during treatment with ACR. At the end of treatment, gait score examination was performed. Then, rats were sacrificed and the severity of damage in brain tissue was determined using pathological tests. The level of malondialdehyde (MDA) and glutathione (GSH) content were evaluated in cerebral cortex and cerebellum to determine the role of oxidative stress in this model.Results: Exposure to ACR induced severe gait abnormalities and pathological changes, but administration of crocin markedly improved behavioral index and histopathological damages. The elevation of lipid peroxidation parallel with reduction of GSH level was observed in cerebral cortex and cerebellum following exposure to ACR. Treatment with crocin markedly decreased MDA level, while elevated GSH content in nervous system as compared to ACR‐treated animals.Conclusion: The administration of crocin markedly improved behavioral and histopathological damages in Wistar rats exposed to ACR. Reduction of oxidative stress can be considered as an important mechanism of neuroprotective effects of crocin against ACR‐induced toxicity.

Objective(s): Rapid and accurate detection of Brucella abortus and Brucella melitensis from clinical samples is so important because antibiotic treatment has major side effects. This study reveals a new method in detection of clinical samples of brucellosis using real‐time PCR and high‐resolution melt (HRM) curve analysis.Materials and Methods: 160 brucellosis suspicious samples with more than 1/80 serum antibody titers were collected and the results were compared with the RFLP method. In order to amplify the sequences for HRM analysis, vdcc, int‐hyp and glk and for RFLP, omp2a and omp2b with PstI and Hinf1 restriction endonuclease were used. At last, the accuracy and specificity of the two methods were compared with each other.Results: Out of these 160 samples, multiplex real time PCR showed 108 positive samples (67.5%), including 56%B. melitensis and 44% B. abortus; whereas in PCR‐RFLP 52 out of 160 samples were positive, where recognition of two species were accordant with HRM analysis, separation was based on the size of the amplified fragment. Using the designed primers and performing the assay, we confirmed this method to be much faster and have lower cost with more than 99% accuracy compared to methods such as RFLP.Conclusion: The present study showed that this technique, which scans gene segments and creates an analysis pattern for detection of clinical samples, is useful and more dominant compared with PCR‐RFLP.Thus, this method can be used for brucellosis detection, and clinical and epidemiological research.

Objective (s): Cutaneous Leishmaniasis is a common and endemic disease in Khorasan province in North‐East of Iran. The pentavalant antimony (Sb V) is the mainstay of treatment that has many side effects and resistance to the drug has been reported. The microbicidal effect of ozone was proven in different microorganisms. Since there is no study in this respect and to achieve a low cost and effective treatment, we decided to evaluate the efficacy of ozone against promastigotes ofLeishmania major, in vitro.Materials and Methods: Ozonated olive oil was prepared after production of ozone by bubbling ozoneoxygen gas produced by ozone generator through olive oil until it solidified. Promastigotes of L. major were cultivated in two phasic media. After calculation of the number of promastigotes, they were incubated with ozonated olive oil (0, 0.626, 0.938, 1.25, 2.5, 5, 10 mcg/ml) at 28oc for 24 hr. Parasites survival percentage was evaluated using MTS and microscopic assay, and then compared with Glucantime and non‐ozonated olive oil.Results: According to the results, there were significant differences in parasites survival percentage between ozonated olive oil and non‐ozonated olive oil, at similar concentrations (P<0.001). Ozonated olive oil was more effective than Glucantime. According to MTS results, Glucantime and ozonated olive oil gel concentrations that are required to inhibit the growth of L. major promastigotes by 50% (IC50), were 165 and 0.002 mg/ml, respectively.Conclusion: Ozonated olive oil has in vitro activity against the promastigotes of L. major and this effect is dose dependent.

Objective (s): Occlusal trauma is one of the most common forms of oral biting dysfunction. Long‐term occlusal trauma could weaken the stomatognathic system; especially damage one’s masticatory muscle. Through using the rat model, this study investigated the trophic effect of ciliary neurotrophic factor (CNTF) on injured masseter muscle.Materials and Methods: Male Wistar rats (n=36) were randomly divided into five experimental groups and one control group (6 rats per group). Animals in the experimental group were cemented modified crowns on their mandibular first molars to artificially induce occlusal trauma in 1, 3, 7, 14, and 28 days. Control group was sham‐treated with forced mouth‐opening for about 5 min, while no crowns were placed. After 28 days of treatment, all rats were euthanized and their masseter muscle was collected. Through immunofluorescence and real‐time quantitative PCR, the expression of desmin, CNTF, and CNTFRa was investigated in rat masseter muscle. The microstructure of masseter muscle was observed by transmission electron microscope.Results: The expression of desmin showed a time‐dependent decrease on traumatic and non‐traumatic sides masseter, until reached the nadir at the 14th day, then restored to its normal level at the 28th day; however, the expression of CNTF and CNTFRa on the traumatic and non‐traumatic sides increased from day 7, reached the peak at the 14th day, and returned to normal level on the 28th day.Conclusion: CNTF, as an important neurotrophic factor, was tightly associated to the restoring of rat injured masseter muscle, which provides new target and treatment method for clinical application.

Objective(s): Tadalafil can restore endothelial function and treat atherosclerosis. However, the effect of tadalafil on transplant arteriosclerosis remains unclear. In this study, we explore the effects of tadalafil on allograft vasculopathy.Materials and Methods: Male Brow‐Norway rats supplied aorta grafts for Male Lewis rats. All recipients were divided into 3 groups: saline as placebo (control) treated group, low dose tadalafil (0.5 mg/kg/day) treated group, and high dose tadalafil (1.0 mg/kg/day) treated group. Eight weeks after transplantation, the grafts were harvested at and analyzed by histological and Western blot analysis. An enzyme‐linked immunosorbent assay (ELISA) was used for measure of plasma cyclic guanylate monophosphate (cGMP).Results: the treatment with tadalafil significantly alleviated the neointimal thickness of aortas compared with the control group (P<0.05). Tadalafil also remarkably enhanced the production of cGMP in plasma and expression of cGMP‐dependent kinase I (PKG‐I) and RhoA compared with control group (P<0.05).Conclusion: These results showed that tadalafil can attenuate graft arteriosclerosis by cGMP ‐PKGI pathway.

Objective(s): Growing evidence indicates that extract of Ginkgo biloba (EGb) attenuates hippocampaldependent memory deficit in aged individuals; however, very little is known about the effect of EGb on the structure of hippocampus. Therefore we examined the EGb‐induced morphological changes of the Cornu Ammonis (CA) region in aged rats.Materials and Methods: Sixteen aged male Wistar rats, 24 months old, were randomly divided into experimental and control groups. Experimental group was orally administered EGb (100 mg/kg/d for 8 weeks), and the control group received a similar volume of water. The volume estimation of CA hippocampal field was done by Cavalieri principle and a quantitative Golgi study was also used for analysis of dendritic arborizations of CA3 and CA1 pyramidal cells.Results: Results revealed that EGb‐treated aged rats had greater volumes than control animals in the layers of pyramidal and radiatum lacunosum moleculare in both CA3 and CA1 subfields. The neurons of CA3 and CA1 in experimental rats had more dendritic segments and larger total dendritic length compared to the control. The results also showed that the aged rats treated by EGb had more numerical branching density in the apical dendrites of CA3 and CA1 pyramidal cells.Conclusion: The results of the present study show that long‐term administration of EGb could produce morphometrical changes in hippocampal pyramidal cells in aged rats. Results also provide a neuroanatomical basis for memory improvement due to chronic treatment with EGb.

Objective (s): There are supportive evidences about the possible role of latent Toxoplasma. Gondii infections on the behavior and neurologic functions, such as increased dopamine levels in the brain. The aim of this study was to examine anti‐toxoplasma activity of aripiprazole that is an atypical antipsychotic drug in mice.Materials and Methods: Mice were randomly divided into four groups, including; control, vehicle, aripiprazole 10 mg/kg, and aripiprazole 20 mg/kg. The mice were inoculated intraperitoneally with mice brain suspension containing tissue cysts. At the end of second month, the number of cysts was counted in smears prepared from brain homogenate by optical microscope.Results: There was no significant difference between mean logarithms of brain cyst numbers of aripiprazole groups compared with control.Conclusion: Results indicate that in aripiprazole groups, the brain cystogenesis was not decrease. Further study needs to investigate the role of anti‐psychotic drugs on T. gondii.