Advanced Biomedical Research
Volume:4 Issue: 2, Feb 2014

There are several methods commonly practicing for deoxyribonucleic acid (DNA) purification from agarose gel. In most laboratories, especially in developing countries, present methods for recovering of DNA fragments from the gel are mostly involved organic solvents. However, manual purification using organic solvents are toxic, labor intensive, time consuming and prone to contamination owing to several handling steps. The above mentioned burdens as well as cost and long time to import them, especially in developing countries, prompted us to design and develop a chamber system for rapid, non-toxic, costeffective and user friendly device for polymerase chain reaction (PCR) products purification from agarose gel.

The device was made from plexiglass plates. After amplification of two fragments of 250 and 850 bp, PCR products were electrophoresed. Subsequently, the desired bands were excised and purified with three

HiPer Mini chamber, phenol extraction method and spin column procedure. To assess the suitability of the purified DNAs, restriction digestion was applied.

Results showed that the yield of recovered DNA in our method was above 95%, whereas the yields obtained with conventional phenol extraction and spin column methods were around 60%.

In conclusion, the current method for DNA elution is quick, inexpensive and robust and it does not require the use of toxic organic solvents. In addition, the purified DNA was well has suited for further manipulations such as restriction digestion, ligation, cloning, sequencing and hybridization.

Mammography as a non invasive method has been suggested to be helpful in predicting coronary artery disease. This study aimed to investigate whether presence and severity of breast artery calcification (BAC) on mammograms is associated with computed tomography coronary angiography (CTCA) finding such as coronary artery calcium (CAC) score and the severity of coronary artery stenosis.

This cross-sectional study was performed on 150 women aged >40 years who were referred for CTCA. Women who had undergone screening mammography during the first year after CTCA entered the study. CAC score was determined and the severity of coronary artery stenosis was classified into normal, non-significant stenosis, or significant stenosis. Based on the severity of BAC, patients were also grouped into normal, mild, moderate, or severe groups. Then, the correlation between BAC severity and CAC score was determined. Patients with different BAC severity were also compared regarding the relative frequency of different grades of coronary artery stenosis.

Mean age of subjects with BAC (n: 35) was significantly higher than patients without BAC (n: 115) (68.03 ± 6.16 versus 54.36 ± 7.63 years, P < 0.0001). Although the relative frequency of different grades of coronary artery stenosis was significantly higher in women with BAC (P < 0.0001), after controlling for age, there was no significant difference between patients with different severity of BAC in the mean of CAC score (P: 0.09). In addition, the correlation between BAC severity and CAC score was not statistically significant (R: 0.09, P: 0.26).

We concluded that presence and severity of BAC have no significant correlation with CAC score on CTCA.

Different risk factors have been suggested for ischemic stroke in young adults. In a group of these patients despite of extensive diagnostic work-up, the primary cause remains unknown. Coagulation tendency is accounted as a possible cause in these patients. Previous studies on factor V Leiden (FVL) as the main cause of inherited thrombophilia for clarifying the role of FVL in stroke have resulted in controversial findings. The current study investigates the role of this factor in ischemic stroke among Iranians.

This case-control study was performed between September 2007 and December 2008 in Isfahan, Iran. The case group comprised of 22 patients of which 15 were males and 7 were females with age range of ≤50 years, diagnosed as ischemic stroke without classic risk factors and the control group consisted of 54 healthy young adults. After filling consent form, venous blood samples were obtained and sent to the laboratory for genetic examination.

No FVL mutation was found in the case group. There was one carrier of the mutation as heterozygous in the control group (relative frequency = 1.85%).

Based on our study, FVL might not be considered as an independent risk factor for ischemic stroke in Iranian individuals who are not suffering from other risk factors of ischemic stroke.

Refractory epilepsy is a significant problem in clinical practice. Sometimes, multiple antiepileptic drugs are required to control the attacks. To avoid various complications ensuring from these drugs, new methods of treatment such as vagus nerve stimulation (VNS) have been recommended. Trigeminal nerve stimulation (TNS) is a new method under evaluation. The purpose of this paper is to determine whether this method is effective or not.

Percutaneous simulation of supraorbital branches of the trigeminal nerve by an electrical device was planned in 18 patients over a six-month period. Participants who fulfilled the research criteria were selected randomly from epileptic patients referred to the clinic. (November 2011-December 2012). T-test was used for data analysis.

Only eight of 18 patients stayed in the study during all 6 months. A 47.9% reduction in daily seizure frequency was seen in this group (P = 0.022). Other subjects left the study earlier. In this group, seizure frequency increased by 10.6% (P = 0.82).

The mechanism of the antiepileptic effects of TNS is not yet clear. In animal studies, it is suggested that the trigeminal nucleus and its projection to nucleus tractus solitarius (NTS) and the locus ceruleus, are involved in seizure modulation.Although in comparison with seizure frequency prior to the study there was significant seizure reduction, according to the usual criteria for VNS i.e. 50% seizure frequency reduction, the effect of TNS per se may not yet be adequate for treatment of seizures. Trigeminal nerve stimulation may be an effective “adjuvant” method for treatment of intractable seizure.

For the past 30 years, bacterial L-asparaginases have been used as therapeutic agents in the treatment of acute childhood lymphoblastic leukemia. It is found in a variety of organisms such as microbes, plants and mammals. Their intrinsic low-rate glutaminase activity, however, causes serious side-effects, including neurotoxicity, hepatitis, coagulopathy and other dysfunctions. Erwinia carotovora asparaginase shows decreased glutaminase activity, so it is believed to have fewer side-effects in leukemia therapy. Our aim was to clone, express, purify and characterize E. carotovora asparaginase.

L-asparaginase from E. carotovora NCYC 1526 (ErA) was cloned and expressed in Escherichia coli strain BL21 (DE3). The enzyme was purified to homogeneity by affinity chromatography. Various conditions were tested to maximize the production of recombinant asparaginase in E. coli.

A new L. asparaginase from E. carotovora NCYC 1526 (ErA) was successfully cloned, expressed and purified in E. coli BL21 (DE3). The specific activity of the enzyme was 430 IU/mg.

The results of the present work form the basis for a new engineered form of ErA for future therapeutic use, which could be extended with crystallographic studies.

Arterial baroreflex (ABR) is an important factor in preventing of blood pressure fluctuations that determined by baroreflex sensitivity (BRS). Estrogen is an ovarian hormone that has influence on ABR. The mechanism of this effect of estrogen unknown and may be mediated by β-adrenergic receptor of nucleus tractus solitarius (NTS), an important area in regulation of baroreflex. Therefore, in this study changing of BRS by estrogen after blockade β-adrenergic receptor of NTS in ovariectomized rats (Ovx) and Ovx treated with estrogen (Est) was examined.

After ovariectomy, all female rats divided to Ovx and Ovx + Est groups and two series of experiments were performed. In the first experiment, phenylephrine was [intravenously, IV] injected in both the Ovx and Ovx + Est groups, and mean arterial pressure (MAP), heart rate (HR), and BRS were evaluated (n = 8 for each group). In the second experiment, each of Ovx and Ovx + Est groups divided into saline and propranolol (pro) groups, saline and pro stereotaxically were microinjected into NTS, respectively. Further, phenylephrine (IV) was injected in all groups and BRS was evaluated.

BRS significantly increased in estrogen-treated groups (Ovx + Est) compared to Ovx groups (P < 0.01). The blockade β-adrenergic receptor of NTS by pro did not significantly changed BRS in both Ovx and Ovx + Est groups.

We concluded that there aren’t any intraction between estrogen and β-adrenergic receptor of NTS in BRS.

A retrospective study has shown lesser days of hospital stay in patients with increased levels of intra-operative end-tidal carbon dioxide (ETCO2 ). It is probable that hypercapnia may exert its beneficial effects on patients’ outcome through optimization of global hemodynamic and tissue oxygenation, leading to a lower rate of post-operative complications. This study was designed to test the hypothesis that higher values of intra-operative ETCO2 decrease the rate of post-operative complications.

In this randomized, double-blind clinical trial, 78 adult patients scheduled for percutaneous nephrolithotomy (PCNL) were prospectively enrolled and randomly divided into three groups. ETCO2 was set and maintained throughout the procedure at 31-33, 37-39 and 43-45 mmHg in the hypocapnia, normocapnia and hypercapnia groups, respectively. The rates of post-operative complications were compared among the three groups.

Seventy-five patients completed the study (52 male and 23 female). Ten (38.5%), four (16%) and two (8.3%) patients developed post-operative vomiting in the hypocapnia, normocapnia and hypercapnia groups, respectively (P = 0.025). The nausea score was significantly lower in the hypercapnic group compared with the other groups (3.9 ± 1.8, 3.2 ± 2.1 and 1.3 ± 1.8 in the hypocapnia, normocapnia and hypercapnia groups, respectively; P = 0.000). Time to return of spontaneous respiration and awakening were significantly decreased in the hypercapnia group compared with the other groups (P < 0.01).

Mild intra-operative hypercapnia has a protecting effect against the development of postoperative nausea and vomiting and decreases the duration of emergence and recovery from general anesthesia.

Real-time polymerase chain reaction (PCR) is based on the revolutionary method of PCR. This technique is the result of PCR enormous sensitivity and real-time monitoring combination. In quantitative gene expression analysis, two methods have more popularity, SYBR Green and TaqMan, SYBR Green is relatively cost benefit and easy to use and technically based on binding the fluorescent dye to doublestranded deoxyribonucleic acid (dsDNA) where TaqMan method has more expensive and based on dual labeled oligonucleotide and exonuclease activity of Taq polymerase enzyme. Specificity is the most important concern with the usage of any non-specific dsDNA-binding Dyes such as SYBR Green whiles more specificity showed by labeled oligonucleotide method such as TaqMan. In this study, we compared two common RT PCR methods, TaqMan and SYBR Green in measurement gene expression profile of adenosine receptors.

Gene expression profiles of A1, A2A, A2B and A3 Adenosine receptors were analyzed by optimized TaqMan and SYBR Green quantitative RT PCR in breast cancer tissues. Primary expression data was normalizing by B. actin reference gene.

Efficiencies were calculated more than 95% for TaqMan and SYBR Green methods in all genes. The correlations between means of normalized data of each gene in two methods were positive and significant (P < 0.05).

Data analysis showed that with the use of high performance primer and by use proper protocols and material we can make precise data by SYBR Green as TaqMan method. In other word by optimization of SYBR Green method, its performance and quality could be comparable to TaqMan method.

Recently, higher efficacy of the combination of long-acting beta2-adrenoceptor agonist and inhaled corticosteroids on controlling asthma symptoms has been hypothesized. This study aimed to examine the clinical effects of the combination of Budesonide with formoterol (Symbicort) and Budesonide (Pulmicort) alone in persistent asthma.

In a randomized double-blinded clinical trial, 76 patients with definite diagnosis of moderate-to-severe asthma were randomized to receive Pulmicort 180 mcg/inhalation two puffs twice daily, or receive Symbicort 80/4.5 mg/inhalation two puffs twice daily, or receive Symbicort 160/4.5 mg/inhalation two puffs twice daily for 3 months. All participants were initially evaluated by spirometry for assessing respiratory parameters and also the level of asthma control was assessed by Asthma Control Test (ACT).

More significant improvement in spirometry parameters, including forced expiratory volume in 1 second (FEV1), forced vital capacity (FVC), FEV1/FVC ratio, as well as in peak expiratory flow (PEF) in both groups of Symbicort with the regimens 80/4.5 mg/inhalation or 160/4.5 mg/inhalation 2 puffs twice daily compared with Pulmicort group, ACT score was significantly improved in Symbicort group with the regimens 160/4.5 mg/inhalation compared with both Symbicort groups with lower dosage and Pulmicort group . Response to treatment in PEF parameter and also in ACT level was significantly more in those who received Symbicort with the regimens 160/4.5 mg/inhalation compared with other two interventional groups adjusted for gender and age.

Symbicort with the regimens 160/4.5 mg/inhalation has higher efficacy in reducing asthma symptom and improving its control compared with low doses of this drug and with Pulmicort.

Helichrysum oligocephalum DC. from Asteraceae family is an endemic plant growing wild in Iran. This study was carried out to investigate the effect of H. oligocephalum hydroalcoholic extract (HOHE) on ulcerative colitis (UC) induced by acetic acid (AA) in rats.

Rats were grouped (n = 6) and fasted for 24 h before colitis induction. Treatments were started 2 h before the induction of colitis and continued for two consecutive days with different doses of HOHE (100, 200, and 400 mg/kg) orally (p.o.) and intraperitoneally (i.p.). The colon tissue was removed and tissue damages were scored after macroscopic and histopathologic assessments.

Among the examined doses of HOHE, 100 mg/kg was the most effective dose that reduced the extent of UC lesions and resulted in significant alleviation. Weight/length ratio as an index of tissue inflammation and extravasation was also diminished in the treatment group administered HOHE at a dose of 100 mg/kg, and the results showed correlation with macroscopic and histopathologic evaluations. These data suggest that HOHE (100 mg/kg) administered either p.o. or i.p. was effective in diminishing inflammation and ulcer indices in this murine model of acute colitis in a non–dose-related manner.

H. oligocephalum could be considered as a suitable anticolitis alternative; however, further studies are needed to support this hypothesis for clinical setting.

Leiomyoma is benign mesenchymal tumor, that frequently occur in uterus but it rarely happens in ovary. Ovarian leiomyomas are seen concomitantly with uterine leiomyoma in about 78% of cases. They often discover incidentally but their most clinical manifestations are abdominal pain and palpable mass. Herein, we reported a large ovarian leiomyoma in a 22-year-old woman with abdominal pain and palpable mass at lower abdominal region. Computed tomography scan revealed large adnexal mass. Microscopic appearance was typical for leiomyoma but because its rarity in ovary the immunohistochemical staining was done. Major differential diagnostic considerations for this tumor in ovary are fibroma/thecoma, sclerosing stromal tumor, and leiomyosarcoma. The immunohistochemical staining with desmin, inhibin, and α-smooth muscle actin are helpful to rule out this differential diagnosis.