Medical Laboratory Journal

Computational Prediction of the Effects of Single Nucleotide Polymorphisms of the Gene Encoding Human Endothelial Nitric Oxide Synthase

Esmaeil Samadian

ABSTRACT

Background and Objective: Genetic variations in the gene encoding endothelial nitric oxide synthase (eNOS) enzyme affect the susceptibility to cardiovascular disease. Identification of the way these changes affect eNOS structure and function in laboratory conditions is difficult and time-consuming. Thus, it seems essential to perform bioinformatics studies prior to laboratory studies to find the variants that are more important. This study aimed to predict the damaging effect of changes in the coding region of eNOS using homology- and structure-based algorithms (SIFT and PolyPhen).

Methods: First, the single nucleotide polymorphisms in the coding region (cSNPs) of the human eNOS gene were extracted from dbSNP. Resulting amino acid changes were reported as primary data required for the study. Then, position and type of amino acid changes along with the complete amino acid sequence were separately entered into the SIFT and PolyPhen tools for analysis.

Results: Of 144 single nucleotide changes, 38 changes by the SIFT, 47 changes by the PolyPhen and 18 amino acid substitutions by both tools were predicted as damaging.

Conclusion: It is predicted that 18 amino acid changes may have damaging phenotypic effects on the structure of the eNOS enzyme that may affect its performance by potentially affecting the enzyme’s various functional regions. Therefore, computational prediction of potentially damaging nsSNPs and prioritizing amino acid changes may be useful for investigating protein performance using targeted re-sequencing and gene mutagenesis experiments.

Effects of Vitamins C and E on Serum Resistin, Hs-CRP, Lipid Profile and Insulin Levels in Patients with Type 2 Diabetes

GH Mohammadzadeh

ABSTRACT

Background and Objective: Vitamins C and E are the two main antioxidants involved in prevention of type 2 diabetes, by reducing oxidative stress. The aim of this study was to evaluate the effects of vitamins C and E supplementation independently, on serum levels of insulin, high sensitivity C-reactive protein (hs-CRP) and resistin in people with type 2 diabetes.

Methods: In this study, 38 patients with type 2 diabetes (17 men and 21 women) received 1,000 mg/day vitamin C, and 40 patients with type 2 diabetes (21 men and 19 women) received 400 IU/day vitamin E orally. Fasting blood glucose and lipid profile were measured using enzymatic method. Hs-CRP was measured by immunoturbidimetric method, and serum insulin and resistin levels were measured by ELISA.

Results: total cholesterol, triglycerides, hs-CRP, insulin and resistin significantly were reduced after vitamin C supplementation (P<0.001). Moreover, the level of total cholesterol (P =0.018), low-density lipoprotein, triglycerides and hs-CRP significantly changed after vitamin E supplementation (P=0.001).

Conclusion: The daily intake of 1,000 mg of vitamin C and 400 IU of vitamin E may be useful in reducing diabetic complications by decreasing serum levels of hs-CRP and lipid profile in people with type 2 diabetes.

Comparison of Epicardial Adipose Tissue Fatty Acid Profile in Cardiovascular Disease Patients Diabetic and Non-Diabetic

Homayun Dolatkhah

ABSTRACT

Background and Objective: The relationship between diabetes mellitus and increased risk of cardiovascular diseases has been demonstrated. The aim of this study was to determine the fatty acid profile of epicardial adipose tissue in diabetic and non-diabetic patients with cardiovascular disease.

Methods: In this study, 28 diabetic and 40 nondiabetic patients were evaluated. The epicardial adipose tissue and blood samples of patients were collected during surgery. Non‐esterified fatty acids and phospholipids were measured by the thin layer chromatography and gas chromatography.

Results: Saturated free fatty acids (12: 0) level was higher in diabetic patients compared to nondiabetic patients (P=0.038), while saturated free fatty acids (16: 0) was significantly lower in diabetics (P=0.011). Unsaturated fatty acid (20: 3n-9) was higher in nondiabetics compared to diabetics (P=0.015). There was a significant decrease in level of monounsaturated fatty acids in diabetic patients. The epicardial adipose tissue of diabetics showed a significant increase in free fatty acid (18: 0) and conjugated linoleic acid levels, while there was a significant decrease in the level of free fatty acid (18: 1n-11). Level of epicardial omega-3 free fatty acid (20: 5 n-3) and 22: 6 n -3 was significantly reduced in these patients.

Conclusion: Differences in the serum free fatty acid profile of the two groups may be due to differences in their diet, while changes in the fatty acid composition of epicardial adipose tissue in these two groups could be due to impaired metabolism of fatty acids such as uptake, movement and androgen synthesis as a result of diabetes. These changes increase the risk of developing atherosclerosis in diabetic patients.

Simultaneous Effect of Resistance Training and Portulace Oleraceal Supplementation on Some Biochemical Markers of Bone Dynamics in Women with Type II Diabetes

Parvin Farzanegi

ABSTRACT

Background and Objective: Osteoporosis is one of the complications of diabetes. The aim of this study was to determine the impact of resistance training along with Portulaca oleracea supplementation on OPG and NFκB levels (bone markers) in women with type II diabetes.

Methods: Overall, 28 women with type II diabetes (44 to 60 years old) were randomly and equally assigned into four groups (supplement, training, training-supplement and control). An eight-week resistance training program (three one-hour sessions per week with one repetition maximum) was performed using three types of bands with different resistances, at 40-50% intensity and 50-70% maximum heart rate. P. oleracea seeds were supplemented daily (7.5 grams mixed in yogurt) for eight weeks. The data was analyzed using descriptive statistics, paired t-test and ANOVA.

Results: After 8 weeks, OPG and NFκB levels in the three groups of supplement, training and training-supplement increased and decreased, respectively. The highest change in both variables was observed in the training-supplement group. Moreover, the level of these two variables in the training-supplement group had significant difference with the controls and other groups (P=0.000).

Conclusion: As a non-pharmacological therapeutic approach, the regular resistance training and P. oleracea supplementation can increase bone formation markers and reduce bone resorption in women with type II diabetes.

Detection of Ampc and Extended-Spectrum Beta-Lactamases in Clinical Isolates of Pseudomonas Aeruginosa from Patients with Cystic Fibrosis

Fereshte Eftekhar

ABSTRACT

Background and Objectives: Pseudomonas aeruginosa is the most frequent opportunistic pathogen isolated from the sputum of patients with cystic fibrosis (CF). Resistance to β -lactam antibiotics may arise from over expression of the naturally occurring AmpC cephalosporinases or acquired extended-spectrum β-lactamases (ESBL). The aim of this study was to determine the antibiotic resistance profiles as well as the prevalence of ESBL and AmpC production in clinical isolates of P. aeruginosa from CF patients in Tehran.

Methods: Antibiotic resistance of 50 non-duplicate P. aeruginosa isolates was determined by the disc diffusion method. AmpC β-lactamase production was detected by the antagonism disc test and ESBL production was detected by the phenotypic confirmatory test. The presence of ESBL and AmpC genes was assessed by PCR, followed by sequencing the PCR products.

Results: The antibiotic resistance rates were as follows: 22% to ceftriaxone, 20% to cefotaxime, 10% to imipenem, 8% to carbenicillin and 6% to ticarcillin, 4% each to cefepime, tobramycin, amikacin and aztreonam and 2% to each piperacilin, meropenem and ceftazidime. AmpC production was observed in 47 isolates (94%) and ESBL production was observed in one isolate (2%). PCR results showed that all isolates carried the bla_AmpCβ-lactamase gene. One multidrug-resistant isolate carried both bla_TEM and bla_PER-1 genes.

Conclusion: The results showed that despite the low rate of antibiotic resistance in P. aeruginosa CF isolates,the presence of multiple β-lactamases even in one isolate is alarming and can complicate the already difficult treatment of chronic infections in the lungs of CF patients.

Laboratory Diagnosis of Nosocomial Bacterial Infections Using Standard Methods

Saeid Abediankenari (Phd)

Background and objective: Implementation of standard methods for accurate detection of bacteria, correct antibiotic susceptibility testing and effective treatment of bacterial infections play important roles in development of public health and prevention of drug resistance. This study aimed to detect bacteria using standard methods and compare the results with the results obtained in teaching hospitals’ laboratories.

Methods: Positive culture plates containing bacteria isolated from patients in hospital laboratories in city of Sari were transferred to microbiology laboratory of Faculty of Medicine at Mazandaran University of Medical Sciences, after determining the genus and species of bacteria and antibiotic susceptibility testing of the isolates. The samples were re-examined based on standard protocols, and antibiotic susceptibility testing was done using the Kirby-Bauer method.

Results: Of 101 patients, 20% of bacteria and 22.5% of antibiotic sensitivity results reported by the hospital laboratories were incorrect. There were significant differences between the two study groups in terms of bacterial species detection and sensitivity to some drugs (P<0.05).

Conclusion: In the present study, lack of implementation of internal quality control programs in some hospital laboratories and lack of proper monitoring by regulatory authorities in different departments of the hospital have caused 20% false-detection results in hospital reports. Inconsistency in results of laboratories, false antibiograms and subsequent false laboratory reports cause drug resistance in some patients. This indicates the necessity of continuous training in the field of Microbiology and implementation of standard protocols and methods for detection of bacterial species and antibiotic susceptibility testing.

Detection of Herpes Simplex Virus Infection in Cerebrospinal Fluid

Mohammad Taghi Kardi

ABSTRACT

Background and Objective: Herpes simplex encephalitis is a life-threatening consequence of the central nervous system (CNS) infection with Herpes simplex virus (HSV). Although it is a rare disease, mortality rates reach 70% in the absence of therapy and only a minority of individuals can return to normal function. The aim of this study was to determine possible correlation between HSV infection and the incidence of encephalitis in patients with neurological signs.

Methods: Overall, 152 CSF samples were tested from patients with neurological signs referred to Mahdieh Clinical Laboratory in Isfahan from 2010 to 2013. After cerebrospinal fluid (CSF) collection, DNA was extracted and real-time polymerase chain reaction (PCR) was performed for HSV detection.

Results: Of 152 patients tested, 50 were diagnosed with encephalitis. HSV DNA was present in the CSF of 13 patients with encephalitis. HSV was significantly higher (p< 0.05) in patients with encephalitis, which shows the significance of infection as an etiological factor of this disease. About 60% of the encephalitis cases were in age range of 1-24 months.

Conclusion: According to the findings of the present study, Cesarean section is recommended for HSV-positive mothers. A routine real-time PCR test is suggested for HSV detection in patients with encephalitis to avoid unnecessary antiviral treatments.

Echophytochemical, Antioxidant and Ethnopharmacological Properties of Stachys inflata Benth.Extract from Chahar Bagh Mountain

Amir Mohammadi

ABSTRACT

BACKGROUND AND OBJECTIVE: Stachys inflata Benth. is used as an anti-inflammatory and antiseptic agent in traditional medicine in most mountain villages of Golestan province. Therefore, this study aimed to investigate the antioxidant, ethnopharmacological and phytochemical properties of extract from different parts of S. inflata, collected from Chahar Bagh Mountain.

METHODS: Flowering branches and root of the plant were collected from Chahar Bagh Mountain (2100 m) in July 2013. At the same time, the most important information about traditional uses of the plant (ethnopharmacology) was recorded by questioning local people. Phytochemical evaluation (total phenolic, flavonoid and anthocyanins content) of ethanolic extract of plant organs was done using spectrophotometry and folin-ciocalteu. The antioxidant activity of the extract was evaluated by DPPH test. P ≤0.05 was considered as statistically significant.

RESULTS: The amount of chemical compounds in the extract of flowering branches and root extract was significantly different. The total phenolic (129.96 ± 5.6 mgGAE/g), flavonoid (29.62 ± 1.4 mgQUE/g) and anthocyanin (0.021 ± 0.001 µg/g) content in the extract of aerial parts of the plant was approximately 1.5 to 3 times higher than those in the root. Due to higher production of active compounds, the antioxidant activity of the aerial parts’ extract showed a greater potential in free radical scavenging (IC₅₀= 76.33 ± 4.2 µg/ml) compared to the root extract.

CONCLUSION: Phytochemical findings and antioxidant activity of the extract of aerial parts of the plant in free radical scavenging, confirm the traditional applications of this plant as analgesic, anti-inflammatory and antiseptic agent in treatment of rheumatism, wounds, burns and diarrhea. It is recommended that further evaluation of the plant’s traditional applications be conducted in vivo and in vitro.

Invitro Anti-Bacterial and Anti-Oxidative Activity of GlycyrrhizaGlabra L. from North of Golestan Province

Ali Varasteh Moradi

ABSTRACT

Background and Objective: Glycyrrhizaglabra L. is one of the most widely used medicinal herbs in Golestan province that is known for its anti-inflammatory, carminative, antiviral, anti-infection and anti-ulcer properties in Iranian traditional medicine. This study aimed to assess the anti-bacterial and anti-oxidative activity of G. glabrafrom the Golestan province.

Methods: The rip root of the plant was collected in autumn 2013. The ethanolic extract of the plant was prepared by maceration method. The anti-oxidative property of the plant was assessed by total antioxidant capacity (TAC), reducing power (RP) and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capacity assays. The anti-bacterial activity was assessed using agar-well diffusion method and the minimum inhibitory concentration (MIC) assay.

Results: The ethanolic extract of G. glabrahad relatively high anti-oxidative activity with IC50 value of 130 µg/ml, especially in the DPPH method. The extract also exhibited high anti-bacterial activity against the following Gram-positive bacteria: Staphylococcus aureus (21.1±0.7 mm), Staphylococcus epidermidis (19.6±0.2 mm), Bacillus subtilus (19.3±0.6 mm), followed by Escherichia coli (12.1±0.8 mm), Enterococcus faecalis (13.2±0.1 mm) and Kelebsiellapneumoniae (11.5±0.4 mm) with MIC values in the range of 31 - 132 mg. mL^-1.

Conclusion: According to results, the root extract of G.glabrais a good source of antioxidant compounds with suitable anti-bacterial activity, which can be used as natural anti-infection and anti-inflammatory agent for treatment of many diseases.

Investigation of Composition and Antimicrobial Properties of Lavandula stoechas Essential Oil Using Disk Diffusion and Broth Microdilution

Jila Asghari

ABSTRACT

Background and Objective: Lavandula stoechas is a species of native and permanent plants in Golestan province that belongs to the family Lamiaceae. L. stoechas has been used in traditional medicine for treatment of various diseases. The aim of this study was to extract essential oil using steam distillation method from the flowers of L. stoechas collected from Jahan-nama region in the Golestan province, and evaluate its antibacterial activity.

Methods: Steam distillation (Clevenger) and GC-MS system were used to separate volatile oils and identify the essential oil components, respectively. Two methods of disk diffusion and broth micro dilution were used to evaluate the antimicrobial effect of L. stoechas essential oil. Six bacterial species including Staphylococcus aureus, Bacillus sp., Enterococcus faecalis, Salmonella enteritidis, Escherichia coli and Pseudomonas aeruginosa were tested.

Results: The essential oil yield was 0.28%. The main components were camphor (71.86%), 1, 8-cineole (4.08%), linalool (3.77%) and borneol (3.19%). The essential oil showed no inhibitory effect on P. aeruginosa and E. faecalis, while it had different inhibitory effects on other bacteria. S. aureus and Bacillus sp. showed the highest sensitivity with inhibition zone diameter of 32 and 29 mm, respectively.

Conclusion: The results of this study showed that the essential oil of L. stoechas has high inhibitory and antimicrobial activity particularly against Gram-positive bacteria, which may be due to the presence of 71.86% camphor in its composition.