Iranian Journal of Basic Medical Sciences
Volume:15 Issue: 2, Mar-Apr 2012

In a recent published paper, Pourshanazari et al found that morphine enhanced angiogenesis in murine matrigel method in sham and two-kidney one-clip (2K-1C) hypertensive animals and they concluded that this effect is via NO pathway (1).Morphine acts both directly and indirectly on angiogenesis, however, the results are contradictory. It seems that the different effect of morphine on angiogenesis process depends on the low or high dose and acute or chronic morphine administration. It is indicated that chronic morphine treatment in wound repair resulted in a significant decrease in the expression of HIF-1α and reduced vascular endothelial growth factor (VEGF) synthesis (2). Morphine increases systemic oxidative stress, impairs mobilization of endothelial progenitor cells (3) and inhibits hypoxia-induced VEGF expression in endothelial cells and cardiac myocytes in ischemic myocardium (4). Morphine also suppresses tumor angiogenesis through a HIF-1alpha/p38MAPK pathway (5). In contrast, it is able to stimulate vascular endothelial cell proliferation in vitro through mitogen-activated protein kinase (MAPK) pathway (6). Topical use of opioids accelerate healing by up-regulating both endothelial and inducible nitric oxide synthase and the vascular endothelial-derived growth factor receptor Flk1 in the wounds (7). Thus, it seems that the effect of morphine on angiogenesis depends on the type of experimental model as well.The authors also concluded that morphine enhanced angiogenesis in sham and 2K-1C animals via NO pathway and they showed that angiogenesis in hypertensive animals is much higher than sham group. Hypertensive subjects have reduced NO bioavailability due to several mechanisms (8) and in that study, morphine increased serum NO concentration in 2K-1C animals which approached normal level. Thus, it seems that the increased serum NO level could not completely explain enhanced angiogenesis in morphine-treated hypertensive animals. On the other hand, there is a relationship between NO and angiotensin II (Ang II). It was shown that high Ang II level decreases NO bioavailability by promoting oxidative stress (9). Thus, morphine probably reduced plasma renin activity and Ang II levels which results in increased NO level. More studies need to clarify the exact role and mechanism of angiogenesis during morphine administration.

Nowadays the most difficult problem in treatment of bacterial infections is the appearance of resistant bacteria to the antimicrobial agents so that the attention is being drawn to other potential targets. In view of the positive findings of phage therapy, many advantages have been mentioned which utilizes phage therapy over chemotherapy and it seems to be a promising agent to replace the antibiotics. This review focuses on an understanding of phages for the treatment of bacterial infectious diseases as a new alternative treatment of infections caused by multiple antibiotic resistant bacteria. Therefore, utilizing bacteriophage may be accounted as an alternative therapy. It is appropriate time to re-evaluate the potential of phage therapy as an effective bactericidaland a promising agent to control multidrug-resistant bacteria.

Objective(s)As there are conflicting findings regarding the clearance-dose and patient characteristics relationships for valproic acid (VPA), this study was conducted to investigate the relationship between patient demographic characteristics, VPA dosage and the drug clearance in adult Iranian patients.Materials and MethodsPatients (N= 47) were either on monotherapy with VPA or were under co-treatment with drugs that have no effect on VPA pharmacokinetic (PK) profile. All of the patients received VPA at therapeutic dose. Steady state trough plasma concentrations of VPA were determined by Fluorescence Polarization Immunoassay (FPIA) and VPA apparent clearance (CL/F) were calculated in each patient.ResultsMean VPA dose and VPA CL/F were 8.93±2.2 mg/kg/day and 0.65±0.55 l/hr respectively. No significant correlations were found between VPA CL/F and patient's age, TBW and VPA dose. VPA CL/F values of male and female patients were compared and no significant difference between these two groups was noted (P> 0.05). Significant correlation between VPA dose and total trough plasma concentration was found (P= 0.001). Mean total VPA plasma concentration was 54.51±23.74 mg/l.ConclusionOur study showed PK of VPA was not affected by age, sex, TBW and VPA dose. However, for detailed results and construction of VPA PK model in Iranian patients, it is necessary to evaluate VPA PK in a larger sample size with different VPA doses, age and TBW ranges.

Objective(s)Cyclooxygenases (COXs) are known to play some roles in physiological mechanisms related to learning and memory. Since sodium salicylate is an inhibitor of COX, we have evaluated the effect of peripheral and intra-hippocampal administration of sodium salicylate on spatial learning and memory in male rats.Materials and MethodsMale rats were studied in two groups; the first group received different intraperitoneal (i.p.) sodium salicylate doses (0, 200, 300, and 400 mg/kg) and the second group received intra-hippocampal doses of the drug (0, 30, 50, and 100 μg/0.5 μl/side). The spatial performance of rats was tested using Morris water maze (MWM) task. The spatial learning and memory parameters were analyzed using ANOVA.ResultsPeripheral and intra-hippocampal administration of sodium salicylate did not lead to a statistically significant change in the mean time (escape latency), and also the distance traveled for finding the hidden platform during the training days, compared with the control group. But at the probe trial, the percentage of time spent in the target quadrant by rats which received the highest doses of drug significantly increased. ConclusionWe found that both peripheral and intra-hippocampal administration of sodium salicylate facilitates the process of spatial memory consolidation in the MWM.

Objective(s)Staphylococcus aureus is a foremost source of numerous nosocomial and community acquired infections. Antibiotic therapy for vancomycin resistant S. aureus (VRSA) can not promise the eradication of infections. Since adhesion is the major route of infections, adhesin based vaccine could suppress S. aureus infections. Fibronectin binding protein A (FnBPA) and clumping factor A (ClfA) are major responsible adhesions involved in S. aureus infections, so they could be candidate vaccine molecules against an extensive range of infections. This project intended to express a new fusion protein construct and analysis of biological activity regarding binding activity.Materials and MethodspfnbA- ClfA construct was transformed to Escherichia coli BL21 (DE3). Transformant E. coli were grown in LB broth and induced with IPTG and cellular extracts were separated on SDS–PAGE. RT-PCR was performed to verify expression. Binding activity of fusion protein was studied using human gingival fibroblast (HGF) cell line. D1-D3 protein from unpublished study was used as control. ResultsThe expected fusion protein fragment showed by SDS-PAGE. RT-PCR verified the existence of mRNA relating to expressed fusion protein. Binding activity of S. aureus decreased after treatment of HGF cells with fusion protein.ConclusionIn total,binding activity of fusion protein was approximately two fold lesser than D1-D3 protein. It is supposed that the fusion protein could not be attached to its ligand easily and would be more accessible to antigen presenting cells and consequentlyprotective antibodies will be produced. This project is pending for in vivo infection study in animal model.

Objective(s)Varicocele is associated with impaired testicular function and male infertility, but the molecular mechanisms by which fertility is affected have not been satisfactorily explained. The aim of our study was to investigate whether or not the polymerase gamma (POLG) polymorphism is associated with Iranian varicocele patients. Materials and MethodsWe determined the POLG CAG repeat length in DNA samples extracted from 40 varicocele patients and 30 control subjects by PCR-denature polyacrylamide gel electrophoresis and sequencing. ResultsThe distribution of the CAG repeat length in varicocele patients showed no notable difference from that in control subjects, but we found a significant statistical inverse correlation between 10/10 and 10/#10 genotypes and varicocele grade. ConclusionThese findings indicate that POLG CAG repeats may affects the varicocele grade, but the mechanisms remain to be elucidated.

Objective(s)Garlic (Allium sativum) is known as a potent spice and a medicine with broad therapeutic properties ranging from antibacterial to anticancer, and anticoagulant. One of the major purified garlic protein components is the 47 kDa protein. In this study, the effect of 47 kDa protein extracted from aged garlic (AGE) was evaluated on mouse dendritic cell (DC) maturation in vitro.Materials and MethodsForty seven kDa protein was purified from AGE by ammonium sulfate precipitation and gel filtration. SDS-PAGE was used to determine the molecular weight and purity of the isolated protein. DCs were purified from spleen of BALB/c mice by Nycodenz centrifugation and their adhesiveness to the plastic dish. The 47 kDa protein isolated from AGE was added to DCs medium during the overnight culture and the expression of DC surface markers was assessed via flowcytometry.ResultsThe 47 kDa protein-treated DCs lowered the expression of DC maturation markers including: CD40, CD86 and MHC-II in comparison with non-treated DCs; (median of 41% versus 47%, 84% versus 91% and 83% versus 90%, respectively) but we observed no statistical difference between the two groups.ConclusionUpon treatment with DCs with 47 kDa protein, DCs down regulated the expression of costimulatory and MHC-II surface molecules, which is similar to tolerogenic DC phenotype. According to the results of the present study, we found that 47 kDa protein purified from AGE can be considered as a potential candidate to generate tolerogenic DCs in vitro.

Objective(s)Erythropoietin has been shown to exert neuroprotective effects in a variety of CNS injury models. Elevation of serum S100β, as a glial damage marker and myelin basic protein (MBP) has been reported to occur in acute carbon monoxide (CO) poisoning. The aim of this study was to evaluate the effect of erythropoietin (EPO) on serum S100β and MBP levels after CO poisoning in rats.Materials and MethodsRats were poisoned with a mixture of 3000 PPM CO in air for 65 min. After exposure, half of the rats received 5000 u/kg EPO and the rest received normal saline. At 3, 6, 12, 24, 48, 72, 144, and 336 hr after exposure samples were taken. Additionally, EPO was administered at three lower doses (625, 1250 and 2500 u/kg). The serum S100β and MBP levels were measured using immunoenzymatic colorimetric assay. Hemoglobin level was alsomeasured.ResultsSerum S100β levels in CO poisoned rats were significantly higher compared to the control group [6 hr (P< 0.01), 12 hr (P< 0. 001), 24 hr (P< 0.001), 48 hr (P< 0.008) and 72 hr (P< 0.008)]. EPO administration could significantly prevent serum S100β elevations after 12 hr (P< 0.008) and 24 hr (P< 0.008) of CO poisoning. Serum MBP levels in CO poisoned rats were not significantly increased in comparison with the control group (P> 0.05). EPO significantly increased the hemoglobin levels.ConclusionEPO could partially prevent neuronal damage. More studies are required to elucidate other aspects of these effects.

Objective(s)Sonodynamic therapy is a physical treatment which utilizes ultrasound waves with an appropriate sensitizer such as protoporphyrin IX (PpIX). The activation of sensitizer depends on cavitation, and therefore, high intensity ultrasound is an important necessity. Beside, high intensity ultrasound can induce side effects on the healthy tissues which have surrounded tumor. The particles in a liquid decrease the ultrasonic intensity threshold needed for onset of cavitation. The non-radiative relaxation time of PpIX in the presence of gold nanoparticles (GNP) is longer than the similar time without GNP. Materials and MethodsThis study was conducted on colon carcinoma tumor in BALB/c mice. The tumors were induced by subcutaneous injection of CT26 cells. Ultrasound irradiation were performed on tumors 24 hr after the injection of PpIX into GNPs. Antitumor effects were estimated by measuring tumor relative volume, doubling time and time being five times of the tumors and by calculating the average survival time of tumor-bearing mice after treatment.ResultsThere is no inhibitory effect in control group. Ultrasound irradiation alone showed a slight antitumor effect which was enhanced by ultrasound plus PpIX (SDT). The synergistic inhibitory effect was significant when ultrasound plus PpIX was conjugated to GNPs. ConclusionOur experiments suggested a significant synergistic effect of ultrasound combined with Au-PpIX that reduced tumor relative volume and increased average animal survival fraction. This effect was obviously stronger than ultrasound alone and synergistic effect of ultrasound combined with PpIX.

Objective(s)There is well documented evidence for the increase in widespread use of complementary and alternative medicine in the treatment of physical and psychiatric symptoms and disorders within the populations. In the present study, we investigated the influence of Vitex agnus-castus (vitex) on anxiety-like behaviors of rats. Materials and MethodsElevated plus maze which is one of the methods used for testing anxiety is used in our present study. Rats were orally administrated with vitex for two week. The anxiety test was carried out after two weeks of oral administration of vitex. For evaluating interaction of vitex and serotonergic systems, rats were anaesthetized with ketamine and special cannulas were inserted stereotaxically into the third ventricle (TV) of brain. After 1 week recovery, the effects of serotonegic agents on anxiety were studied.ResultsOral administration of vitex (100, 200, 300 mg/kg) for two weeks induced an anxiogenic-like effect which was shown through specific decreases in the percentages of open arm time (OAT %) and open arm entries (OAE %). Intra-TV infusion of 5HT1A receptor agonist, 8-OH-DPAT (5, 10 and 25 ng/rat) increased OAT% and OAE%, indicating anxiolytic–like behavior. However, injection of 5HT1A receptor antagonist NAN190 (0.25, 0.5 and 1 µg/rat) produced anxiogenic-like behavior. The most effective dose of 8-OH-DPAT (10 ng/rat), when co-administered with vitex (100, 200, 300 mg/kg), attenuated the anxiogenic-like effects of vitex significantly. Injection of the less effective dose of NAN190 (0.5 µg/rat), in combination with vitex (100, 200, 300 mg/kg), potentiate anxiogenic effects of vitex.ConclusionsThese results illustrate that 5HT1A receptor is involved in the anxiogenic effects of vitex.

Objective(s)In the last few years, serum procalcitonin has been proposed as an early marker of infections in neonates, with varying results. In this study, we aimed to investigate the value of procalcitonin, and C- reactive protein in establishing the diagnosis of neonatal sepsis.Materials and MethodsBlood samples were collected at admission from 69 neonates with suspected infection (admitted to the Neonatal Intensive Care Units at Alzahra and Dr Beheshti Hospital in Isfahan and Fatema-Zahra in Najafabad from May 2005 to April 2006). Patients were categorized in different groups according to clinical symptoms of sepsis, bacteriological and laboratory results. Group I consisted of 20 newborns with positive blood cultures and other biological tests which suggested infection. Group II consisted of 49 neonates with negative blood cultures but had two or three of clinical signs of sepsis. The control group included 18 healthy neonates with physiological hyperbilirubinemia and no clinical and biological data of infection, referred to the hospital for bilirubin determination. Procalcitonin and C-reactive protein (CRP) were determined by immunoluminometric assay and nephlometry method respectively.ResultsMean levels of procalcitonin and CRP in septic neonates (group I) were significantly higher than the other two groups (P< 0.005). Sensitivity, specificity, positive predictive value and negative predictive value were determined for all markers and compared with each other.ConclusionWe conclude that procalcitonin is a better marker than CRP in the diagnosis of neonatal sepsis.

Objective(s)Staphylococcus aureus is a common cause of human infection, and emergence of vancomycin-resistanceS. aureus is a great concern for treatment of methicillin-resistant S. aureus,(MRSA) in recent years (MRSA). Here, we report the isolation of high-level VRSA.Materials and MethodsS. aureus was isolated from foot ulcer of a diabetic woman in Tehran, Iran. Antibiotic susceptibility was determined according to CLSI guidelines. VanA gene cluster PCR was carried out and PCR amplicon of vanA was sequenced.ResultsS. aureus had high-level vancomycin-resistant (MIC 512 ≥ µg/ml). Patient's history revealed that VRSA isolate was acquired through community transmission. Only vanA, vanR and vanS genes were amplified in our isolate. Sequencing revealed that the vanA sequence had high similarity to the vanA sequence of Tn1546.ConclusionAlthough VRSA infection continues to be rare, isolation of community–acquired VRSA is a significant issue and it needs the efforts of public health authorities.

Objective(s)Annual incidence of cutaneous leishmaniasis is increasingly growing and development of the alternative drugs against it is a major concern. Artemisia genus is a traditional medicinal plant in Iran. The aim of this study was to examine the leishmanicidal activity of various Iranian Artemisia species extracts. Materials and MethodsDifferent extracts were gathered from eleven Iranian Artemisia species. Their leishmanicidal activities against the growth of Leishmania major (L. major)promastigotes were examined as the half maximal inhibitory concentration(IC50) using MTT assay. ResultsObtained results showed that ethanol extracts especially those taken from A. ciniformis, A. santolina and A. kulbadica have the strongest effects. ConclusionLooking for the effective leishmanicidal agents from natural resources in Iran, we found that the ethanol extract of collected Artemisia species had significant effect on in vitro leishmanicidal activity and may be suitable candidates in the treatment of leishmaniasis.