Iranian Journal of Basic Medical Sciences
Volume:16 Issue: 2, Feb 2013

Throughout evolution, mammalians have increasingly lost their ability to regenerate structures however rabbits are exceptional since they develop a blastema in their ear wound for regeneration purposes. Blastema consists of a group of undifferentiated cells capable of dividing and differentiating into the ear tissue. The objective of the present study is to isolate, culture expand, and characterize blastema progenitor cells in terms of their in vitro differentiation capacity. Five New Zealand white male rabbits were used in the present study. Using a punching apparatus, a 4-mm hole was created in the animal ears. Following 4 days, the blastema ring which was created in the periphery of primary hole in the ears was removed and cultivated. The cells migrated from the blastema were expanded through 3 successive subcultures and characterized in terms of their potential differentiation, growth characteristics, and culture requirements. The primary cultures tended to be morphologically heterogeneous having spindly-shaped fibroblast-like cells as well as flattened cells. Fibroblast-like cells survived and dominated the cultures. These cells tended to have the osteogenic, chondrogenic, and adipogenic differentiation potentials. They were highly colonogenic and maximum proliferation was achieved when the cells were plated at density of 100 cells/cm2 in a medium which contained 10% fetal bovine serum (FBS). Taken together, blastema tissue-derived stem cells from rabbit ear are of mesenchymal stem cell-like population. Studies similar to this will assist scientist better understanding the nature of blastema tissue formed at rabbit ear to regenerate the wound.

Reflexes that rose from mechanoreceptors in nasal cavities have extensive neuro-regulatory effects on respiratory system. Because of side specific geometry and dual innervations of the nasal mucosa, we investigated the consequences of unilateral nasal stimulations on respiratory mechanics and breathing patterns. Unilateral nasal air-puff stimulation (30 min) in the presence of propranolol (25 mg/kg) and atropine (5 mg/kg) were applied on tracheotomized spontaneously breathing rats. Breathing rate and pattern monitored. Peak inspiratory pressure (PIP) and flow (PIF) were exploited for calculation of resistance, dynamic compliance (Cdyn), and estimation of respiratory system impedance (Zrs). During right-side stimulation, in propranolol (P<0.05) and atropine groups (P<0.01) PIP significantly decreased in comparison to the control group. Alternatively, it significantly increased in left-side and propranolol-left groups (P<0.05) than control group. Mean Cdyn following left-side stimulation and propranolol, revealed significant decrements (P<0.05) than control group. In the case of atropine-right and atropineleft groups, mean Cdyn had significantly decreased in comparison with atropine alone (P<0.05). Airway resistance (R) did not reveal significant difference during nasal stimulations whereas least square approximation revealed a significant side-specific frequency dependent deviation of imaginary part of impedance (X). An inverse correlation was determined for Cdyn versus frequency following right side (R=-0.76) and left side (R=-0.53) stimulations. For the reason that lower airways mechanics changed in a way independentfrom smooth muscle, it may be concluded from our data that unilateral nasal stimulations exert their different controls through higher regulatory centers.

In regard to the high incidence of asthma and the side-effects of the drugs used, finding novel treatments for this disease is necessary. Our previous studies demonstrated the preventive effect of Nigella sativa extract on ovalbumin-induced asthma. In addition, water-soluble substances of N. sativa extract and methanol fraction of this plant were responsible for the relaxant effect of this plant on tracheal chains of guinea pigs. Therefore, for the first time, in the present study, in order to identify main constituents of the methanolic extract, the relaxant effects of five different methanolic fractions (20%, 40%, 60%, 80%, and 100%) of N. sativa on tracheal chains of guinea pigs were examined. The relaxant effects of four cumulative concentrations of each fraction (0.8, 1.2, 1.6, and 2.0 g%) in comparison with saline as negative control and four cumulative concentrations of theophylline (0.2, 0.4, 0.6, and 0.8 mM) were examined by their relaxant effects on precontracted tracheal chains of guinea pig by 60 mM KCl (group 1) and 10 μM methacholine (group 2). In group 1, all concentrations of only theophylline showed significant relaxant effects but all concentrations of these methanolic fractions showed significant contractile effects compared with that of saline (P<0.001 to P<0.05). However, in group 2, all concentrations of theophylline and these methanolic fractions showed significant relaxant effects compared with that of saline (P<0.001 to P<0.05). These results showed a potent relaxant effect of 20% methanolic fractions from N. sativa on tracheal chains of guinea pigs that were higher than that of theophylline at the used concentrations

In our previous study, we reported that capsaicin-induced unmyelinated C-fiber depletion can modulate excitatory and integrative circuits in the somatosensory cortex following experience-dependent plasticity. In this study, we investigated the involvement of the capsaicin-induced acute inactivation of c-fibers on tactile learning in rat. The delayed novel object recognition test was used to assess tactile learning. This procedure consisted of two phases. The first of these (T1) was a training phase during which the animals explored two similar objects. T2, the test phase, occurred 24 hr later, during which the animals explored one novel and one familiar object. In order to induce acute inactivation of the C-fiber pathway, 25–30 μl of a 10% capsaicin was injected subcutaneously into the rat’s upper lip, 6 h prior to T1. Tactile learning was quantified using a discrimination ratio. In T2, the discrimination ratio in capsaicin-treated animals (37.3±3.8%) was lower than that observed in vehicle-treated animals (54.4±5.1%, P<0.05). These findings indicate that the selective inactivation of a peripheral nociceptor subpopulation affects tactile learning.

Spermatogonial Stem Cells (SSCs) maintain spermatogenesis throughout the life of the male. Because of the small number of SSCs in adult, enriching and culturing them is a crucial step prior to differentiation or transplantation. Maintenance of SSCs and transplantation or induction of in vitro spermio-genesis may provide a therapeutic strategy to treat male infertility. This study investigated the enrichment and proliferation of SSCs co-cultured with STO cells in the presence or absence of growth factors. Spermatogonial populations were enriched from the testis of 4-6 week-old male mice by MACS according to the expression of a specific marker, Thy-1. Isolated SSCs were cultured in the presence or absence of growth factors (GDNF, GFRα1 and EGF) on STO or gelatin-coated dishes for a week. Subsequently, the authors evaluated the effects of growth factors and STO on SSCs colonization by alkaline phosphates (AP) activity and flow cytometery of α6 and β1 integrins. SSCs co-cultured with STO cells and growth factors developed colonization and AP activity as well as expression of α6 and β1 integrins (P≤0/05). Our present SSC-STO co-culture provides conditions that may allow efficient maintenance and proliferation of SSCs for the treatment of male infertility.

Breast cancer is the leading cause of cancer-related death in women worldwide. It has been revealed that elevated risk for malignancy may be associated with certain HLA alleles. This study was performed to assess the association of HLA class I alleles with breast cancer in women in Southern Iran. Eighty nine patients included for analyzing the HLA class I alleles frequency using complement dependent cytotoxicity microassay and results were compared to 86 gender-matched healthy volunteers. There were significantly more patients with A24(9) allele than those of healthy individuals (38.2% versus 16.3%) (P-value=0.002). In contrast, HLA-A1 had significantly much less expression in the patient group compared to the controls (P- value=0.04). A24(9) allele appears to be one of the factors increasing an individual’s the susceptibility to breast cancer in our population but further investigation might be required.

It has been reported that ischemic postconditioning, conducted by a series of brief occlusion and release of the bilateral common carotid arteries, confers neuroprotection in permanent or transient models of stroke. However, consequences of postconditioning on embolic stroke have not yet been investigated. In the present study, rats were subjected to embolic stroke (n=30) or sham stroke (n=5). Stroke animals were divided into control (n=10) or three different patterns of postconditioning treatments (n=20). In the first pattern of postconditioning (PC10, n=10), the common carotid arteries (CCA) were occluded and reopened 10 and 30 sec, respectively for 5 cycles. Both occluding and releasing times in pattern 2 (PC30, n=5) and 3 (PC60, N=5) of postconditionings, were five cycles of 30 or 60 sec, respectively.Postconditioning was induced at 30 min following the stroke. Subsequently,cerebral blood flow (CBF) was measured from 5 min before to 60 min following to stroke induction. Infarct size, brain edema and neurological deficits and reactive oxygen species (ROS) level was measured two days later. While PC10 (P<0.001), PC30 and PC60 (P<0.05) significantly decreased infarct volume, only PC10 decreased brain edema and neurological deficits (P<0.05). Correspondingly, PC10 prevented the hyperemia of brain at 35, 40, 50 and 60 min after the embolic stroke (P<0.005). No significant difference in ROS level was observed between PC10 and control group. Ischemic postconditioning reduces infarct volume and brain edema, decreases hyperemia following to injury and improves neurological functions after the embolic model of stroke.

Polyethylenimine (PEI) is a potent non-viral gene delivery carrier. PEI surface charge plays a major role in its condensation ability which in turn is a necessary requirement for high transfection efficiency. As the PEI charge density is dependent on pH, the effect of pH changing PEI on PEI condensation ability, transfection efficiency, and cytotoxicity was investigated. In order to compare the different values of pH, 25 kDa PEI solutions were prepared at the pH values of 5, 7 and 10, afterward complexed with two plasmids encoding reporter genes of luciferase and enhanced green fluorescence protein to evaluate the capability of polymers in plasmid delivery at two incubation time (4 versus 24 hr). The condensation ability of PEI in acidic, neutral and basic environments was similar. At low pH value, the polyplex had negative surface charge whereas at a higher pH value, the surface charge increased and became positive. The higher transfection efficiency and lower cytotoxicity were achieved by the polyplexes prepared at the pH value of 5 with longer incubation time of 24 hr. The results showed the impact of pH value of PEI on biological and biophysical properties of polyplexes. Although the proton sponge effect can justify several properties of PEI, these results suggest that the proton sponge hypothesis may not be applicable for polymers with low buffering capacity at the pH values around 5.

The aim of this study was to investigate ascorbic acid and garlic protective effects on lead-induced neurotoxicity during rat hippocampus development. 90 pregnant wistar rats were divided randomly into nine groups: 1- Animals received leaded water (L). 2- Rats received leaded water and ascorbic acid (L+AA). 3- Animals received leaded water and garlic juice (L+G). 4-Animals received leaded water, ascorbic acid and garlic juice (L+G+AA). 5- Rats treated with ascorbic acid (AA). 6- Rats treated with garlic juice (G). 7- Rats treated with ascorbic acid and garlic juice (AA+G). 8- Rats treated with tap water plus 0.4 ml/l normal hydrogen chloride (HCl) and 0.5 mg/l Glucose (Sham). 9- Normal group (N). Leaded water (1500 ppm), garlic juice (1 ml/100g/day, gavage) and ascorbic acid (500 mg/kg/day, IP) were used. Finally, blood lead levels (BLL) were measured in both rats and their offspring. The rat offspring brain sections were stained using Toluidine Blue and photographed. Dark neurons (DNs) were counted to compare all groups. BLL significantly increased in L group compared to control and sham groups and decreased in L+G and L+AA groups in comparison to the L group (P<0.05). the number of DNs in the CA1, CA3, and DG of rat offspring hippocampus significantly increased in L group in comparison to control and sham groups (P<0.05) and decreased in L+G and L+AA groups compared to L group (P<0.05). Garlic juice and ascorbic acid administration during pregnancy and lactation may protect lead-induced neural damage in rat offspring hippocampus.

Our objective was to evaluate the effects of a triple antioxidant combination [α-tocopherol (AT), ascorbic acid (AA) and α-lipoic acid (LA); AT+AA+LA] on the cholesterol and glutathione levels, and the fatty acid composition of liver and muscle tissues in diabetic rats. Forty-three Wistar rats were randomly divided into five groups. The first group was used as a control. The second, third and fourth groups received STZ (45 mg/kg) in citrate buffer. The fourth and fifth groups were injected with intraperitoneal (IP) 50 mg/kg DL-AT and 50 mg /kg DL-LA four times per week and received watersoluble vitamin C (50 mg/kg) in their drinking water for a period of six weeks. Liver cholesterol levels in the AT+AA+LA group were lower than the control (P<0.05). Glutathione level was lower in D-2 (P<0.05) and were higher in D+AT+AA+LA and AT+AA+LA groups than the control groups (P≤ 0.05). The muscle cholesterol levels in the D-1 and D+AT+AA+LA groups were higher than the control group (P≤ 0.05). The levels of oleic acid were higher in the D-1 group and lower in the D-2 group (P<0.001). The arachidonic acid level in the D-1 and D-2 groups were lower (P<0.05), and higher in the D+AT+AA+LA group. Our results revealed that glutathione levels and the Stearoyl CoA Desaturase enzyme products in liver tissues of diabetic and non-diabetic rats were increased by triple antioxidant mixture.

Klebsiella infections are caused mainly by K. pneumoniae and K. oxytoca. In the last two decades, a new type of invasive Klebsiella pneumoniae which contains mucoviscosity-associated gene (magA) has emerged. The aim of this study was to investigate the prevalence of magA gene and to detect antimicrobial susceptibility patterns of Klebsiella spp. isolated from clinical samples. Klebsiella isolates were collected from patients admitted to referral hospitals of Hamadan, Iran, during a 12-month period from 2007 to 2008. The samples were analyzed by conventional microbiological methods and polymerase chain reaction (PCR). The hypermucoviscosity (HV) phenotype of Klebsiella isolates was characterized by formation of viscous strings >5 mm as a positive test. The susceptibility of isolates to routine antibiotics was assessed by agar disk diffusion method. Out of 105 Klebsiella isolates, 96.2% was identified as K. pneumoniae and 3.8% as K. oxytoca by PCR. magA gene was detected in 4 (3.8%) isolates of K. pneumoniae. The isolates of K. oxytoca contained no magA gene. From 4 isolates with positive magA gene, two of them were HV+ and two were HV- phenotype. Overall, sixty-four isolates (60.95%) of K. pneumoniae showed an HV positive phenotype and all isolates of K. oxytoca were HV-phenotype. The most effective antibiotics against the isolates were tobramycin (79.05%), ceftazidime (79.05%), ceftizoxime (78.09%), ciprofloxacin (76.19%), ceftriaxone (76.24%) and amikacin (74.29%). The results suggest that there is also magA associated serotype of the K. pneumoniae in this region. In addition, the presence of HV+ phenotype may not be associated with magA.

Radiation effect induced in nonirradiated cells which are adjacent or far from irradiated cells is termed radiation-induced bystander effect (RIBE). Published data on dose-response relationship of RIBE is controversial. In the present study the role of targeted and bystander cells in RIBE dose-response relationship of two cell lines have been investigated. Two cell lines (QU-DB and MRC5) which had previously exhibited different dose-response relationship were selected. In the previous study the two cell lines received medium from autologous irradiated cells and the results showed that the magnitude of damages induced in QU-DB cells was dependent on dose unlike MRC5 cells. In the present study, the same cells irradiated with 0.5, 2 and 4 Gy gamma rays and their conditioned media were transferred to nonautologous bystander cells; such that the bystander effects due to cross-interaction between them were studied. Micronucleus assay was performed to measure the magnitude of damages induced in bystander cells (RIBE level). QU-DB cells exhibited a dose-dependent response. RIBE level in MRC5 cells which received medium from 0.5 and 2 Gy QU-DB irradiated cells was not statistically different, but surprisingly when they received medium from 4Gy irradiated QU-DB cells, RIBE was abrogated. Results pertaining to QU-DB and MRC5 cells indicated that both target and bystander cells determined the outcome. Triggering the bystander effect depended on the radiation dose and the target cell-type, but when RIBE was triggered, dose-response relationship was predominantly determined by the bystander cell type.