Iranian Biomedical Journal
Volume:17 Issue: 1, Jan 2013

Infections due to Pseudomonas aeruginosa are among the leading causes of morbidity and mortality in patients who suffer from impaired immune responses and chronic diseases such as cystic fibrosis. At present، aggressive antibiotic therapy is the only choice for management of P. aeruginosa infections، but emergence of highly resistant strains necessitated the development of novel alternative therapeutics including an effective vaccine. Several P. aeruginosa antigens have been tested for vaccine development، including lipopolysaccharide alone، polysaccharides alginate، extracellular proteins، exotoxin A (exo A) and killed whole cell. However، none of them are currently available clinically. In this research، recombinant exoA-flagellin (fliC) fusion protein as a cocktail antigen was expressed، purified and its antigenic characteristics were evaluated. Expression of recombinant fusion protein by E. coli using pET22b vector resulted in production of exoA-fliC fusion protein in high concentration. Based on Western-blotting results، recombinant fusion protein showed a good antigenic interaction with sera from patients with various P. aeruginosa infections. These results suggested that recombinant exoA-fliC fusion protein can be produced in the laboratory، and tested as a candidate vaccine in P. aeruginosa infections.

Bone marrow stromal stem cells (BMSC) are appropriate source of multipotent stem cells that are ideally suited for use in various cell-based therapies. It can be differentiated into neuronal-like cells under appropriate conditions. This study examined the effectiveness of co-stimulation of creatine and retinoic acid in increasing the differentiation of BMSC into GABAergic neuron-like cells (GNLC).

BMSC isolated from the femurs and tibias of adult rats were cultured in DMEM/F12 medium supplemented with 10% FBS، pre-induced using β-mercaptoethanol (βME) and induced using retinoic acid (RA) and creatine. Immunostaining of neurofilament 200 kDa، neurofilament 160 kDa، nestin، fibronectin، Gamma-amino butyric acid (GABA) and glutamic acid decarboxylase (GAD) 65/67 were used to evaluate the transdifferentiation of BMSC into GNLC and to evaluate the effectiveness of pre-induction and induction assays. The expression of genes that encode fibronectin، octamer-binding transcription factor 4 (Oct-4)، GAD 65/67 and the vesicular GABA transporter was examined in BMSC and GNLC by using RT-PCR assays during transdifferentiation of BMSC into GLNC.

Co-stimulation with RA and creatine during the induction stage doubled the rates of GABAergic differentiation compared with induction using creatine alone، resulting in a 71. 6% yield for GLNC. RT-PCR showed no expression of Oct-4 and fibronectin after the induction stage.

The results of this study showed that the application of βME، RA، and creatine induced the transdifferentiation of BMSC into GLNC.

Efficient screening for detection of colorectal cancer (CRC) at earlier stages reduces its mortality. The purpose of this study was to investigate expression of carcinoembryonic antigen (CEA) and human telomerase reverse transcriptase (hTERT) mRNA in peripheral blood of CRC patients and to present strategies for early detection screen test.

Twenty seven patients in non-metastatic stage and 27 healthy individuals were studied. Expression of CEA، hTERT mRNA and 18srRNA (18s subunit of ribosomal RNA، as reference gene) were determined based on real-time RT-PCR on 3 µg of total RNA from blood in 3 separate vials (1 µg per vial).

Positive expression rate of CEA mRNA (78%) and hTERT mRNA (81%) were higher in patient group (P<0. 001). These rates were meaningfully higher than the results of individual vials containing only 1 µg of total RNA. Difference between Ct values of markers with 18srRNA (ΔCt) was higher in healthy group than patient one. Therefore، a ΔCt cut-off value was determined for distinguishing between true- and false-positive results. Concurrent expression of both markers was found in 67% of the patients، which was higher than healthy cases (11%). Combination of concurrent marker expression with cut-off point strategy increased specificity to 100%.

These results showed that concurrent evaluation of marker expression and performing the test on 3 µg of samples in 3 separate vials may increase specificity and sensitivity of real-time RT-PCR for early detection of non-metastatic CRC. However، more investigations with larger numbers of samples are needed to verify these results.

Oxidative modification of low-density lipoprotein (LDL) appears to be an early step in the pathogenesis of atherosclerosis. Meanwhile، myeloperoxidase (MPO) -catalyzed reaction is one of the potent pathway for LDL oxidation in vivo. The aim of this study was to evaluate in vitro antioxidant effects of vitamins C and E on LDL oxidation mediated by MPO. MPO was isolated from fresh plasma by sequential centrifugation using density ultracentrifugation. It was incubated with LDL and the LDL oxidation level was determined spectrophotometrically by measuring conjugated diene absorbance at 234 nm. Furthermore، vitamin C (50-200 mM) and vitamin E (10-40 mM) were added and the LDL oxidation level was determined. The purity index of MPO and its enzymatic activity were 0. 69 and 1127 U/mg protein، respectively. It was demonstrated that vitamin C in vitro inhibited LDL oxidation mediated by MPO; however، vitamin E was unable to act in the same way. The protection by vitamin C was concentration dependent and maximum protective effect of vitamin C was observed at 150 mM، where about 64% of the LDL oxidation was inhibited. Vitamin C increased lag time of LDL oxidation mediated by MPO up to 2. 4 times. It can be concluded from our results that vitamin C is able to improve LDL resistance to oxidative modification in vitro. In addition، vitamin C might be effective in LDL oxidation mediated by MPO in vivo، resulting in reduction of atherosclerosis process rate.

Metabolic derangements in type 2 diabetes mellitus (T2DM) are likely to affect skeletal muscle contractile functions adversely. Levo-carnitine improves muscle contractile functions in healthy humans and rats and corrects metabolic derangements in T2DM. Therefore، it is likely to improve muscle contractile functions in T2DM as well. This study was designed to determine the effect of levo-carnitine on serum levo-carnitine levels، oxidative stress and contractile parameters of fast muscle in T2DM. Ninety Sprague-Dawley rats were randomly divided into three equal groups. Healthy rats served as the controls، while T2DM was induced in diabetic and carnitine groups. The carnitine group was administered levo-carnitine 200 mg/kg/day intraperitoneally for 6 days. At 28th day، extensor digitorum longus muscles were removed and their functions were assessed using iWorx data acquisition unit (AHK/214). Blood obtained by intra-cardiac sampling at 28th day was used for estimation of serum malondialdehyde (MDA) and levo-carnitine levels. Maximum isometric twitch tension، time-to-peak twitch tension and time-to-relax to 50% of the peak twitch tension were not significantly different amongst the groups. Carnitine group showed significant improvement in maximum fused tetanic tension، maximum fused tetanic tension after fatigue protocol and recovery from fatigue after 5 minutes of rest period compared to the diabetic group. Serum MDA levels were reduced، while serum levo-carnitine levels were elevated significantly in carnitine group as compared to the diabetic group. Levo-carnitine supplementation increases serum levo-carnitine levels which decreases oxidative stress. This action improves contractile force but delays fatigue in fast muscles of diabetic rats.

Helicobacter pylori، which is associated with many upper gastrointestinal diseases، is found in half of the population of the world. Several special stains and immunohistochemistry stain for H. pylori are available. The need for and usefulness of immunohistochemical (IHC) technique has been debated for years. Toluidine blue is a simple stain for microbiological studies and is easily available in laboratories. Therefore، this study was conducted to compare hematoxylin and eosin (H&E)، Giemsa and toluidine blue staining with immunehistochemistry for detection of H. pylori in patients with gastritis and also to correlate the results of these staining methods with pathological grading.

We reviewed 54 consecutive gastric biopsy specimens stained by H&E and Giemsa as well as by toluidine blue and immunohistochemistry stains for H. pylori.

H. pylori was positively identified by IHC in 43 (79. 63%) patients، while positive samples were found in 18 (33. 33%)، 24 (44. 44%) and 33 (61. 11%) patients using H&E، Giemsa and toluidine blue staining methods. Our results showed that classical histological staining methods are not sensitive enough to identify low numbers or coccoid forms of organism، while toluidine blue and immunohistochemistry play an important role in detection of H. pylori infection.

Toluidine blue has been proved to be much more reliable than H&E and Giemsa in detection of H. pylori. In addition، in post treatment biopsies and in biopsies with unexplained chronic active gastritis without histological evidence of H. pylori should have immunohistochemistry done to detect possible low density or coccoid form of organisms.

Through its membrane and intracellular receptors، vitamin D regulates many vital functions in the body including its well known actions on musculoskeletal system. Growing body of evidences demonstrate that vitamin D undergoes some of behavioral aspects of neurocognition. The present study was designed to evaluate the effect of food regimens without vitamin D or with a supplement of 1،25 (OH) 2D3 on spatial performance of adult rats. The animals were trained in the Morris water maze to find a hidden platform. The time spent and the distance travelled to find the platform، speed of navigation and the percentage of unsuccessful trials were considered for assessment of the task learning. Our findings indicated that the vitamin D-deprived rats had a significant lower performance compared to both the controls and the animals receiving 1،25 (OH) 2D3 supplementation. Concerning the unsuccessful trials، lack of vitamin D resulted in the highest failures in the maze navigation. The regimen with additional 1،25 (OH) 2D3 did not considerably influence learning of the maze task. We concluded that while vitamin D deficiency deteriorates the spatial task learning، the 1،25 (OH) 2D3 supplementation did not effectively underlie the maze performance.

Although effects of trace elements on secretion of sex steroids and insulin have been studied، the effects of these hormones on serum level of trace elements have been rarely investigated. The aim of the present study was to evaluate the effect of testosterone and finasteride administration and castration on serum levels of testosterone، insulin، zinc and chromium. Male adult rats (n = 32) were divided into 4 groups (n = 8). Group 1، control; Group 2، castration، castration was done at the first day of the study; Group 3، finasteride (20 mg/kg/day، dissolved in drinking water) and Group 4، testosterone (5 mg/kg/day، i. p.). At the end of the period of the study (35 days)، serum testosterone، insulin، zinc and chromium levels were determined in the blood samples collected directly from the right atrium of the heart of the animals. The data indicated that the serum levels of testosterone، insulin and zinc were significantly increased (P<0. 01) in testosterone-administrated and finasteride groups، but the level of chromium was decreased in both groups (P<0. 01). Castrated group had the lowest serum levels of testosterone، insulin and zinc (P<0. 05). Also، the levels of serum chromium in this group were increased. The study demonstrates that testosterone and finasteride increases insulin and zinc levels and decreases chromium levels in the serum of male adult rats. According to these data، it seems that testosterone may affect glucose cycle through effect on serum insulin levels and trace elements such as zinc and chromium.