Gene, Cell and Tissue
Volume:6 Issue: 1, Jan 2019

There are few methods for cell labeling in basic studies of cell-migration, tissue engineering, and cell therapy. Recent progress in cell trackers has led to an increasing number of clinical trials involving the treatment of various diseases based on cell transplantation. In this study, we investigated the effects of different concentrations of DiI as a cell labeling dye on the viability and proliferation of mouse embryonic fibroblasts (MEFs). MEFs were divided into two groups of control and experimental (treated with Dil dye). We treated MEFs with three concentrations of the dye and after 1, 5, 7 and 14 days, these cells were analyzed by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and 4’, 6-diamidino-2-phenylindole (DAPI) staining. Also, the stability of the dye in treated MEFs was examined by immunofluorescence. Our results showed that cell viability was significantly different between the groups that received Dil at the concentrations of 1 and 2 µg/mL and the control group. However, cell viability was not significantly different between the group that received Dil (0.5 µg/mL) and the control group. Therefore, Dil at concentrations higher than 0.5 µg/mL was toxic for MEFs. We suggest that Dil dye, which is concentration-dependent, can be used as a safe and stable fluorescent dye for cell labeling both in-vivo and in-vitro.

Phenylketonuria is an inborn metabolic disorder inherited in an autosomal recessive pattern. The detection of pathogenic variations improves the power of at-risk carrier and prenatal detection. We previously found Q375R a novel phenylalanine hydroxylase variation in phenylketonuria patients from the south-west of Iran. Here, we aimed to evaluate the rate of the pathogenicity of this novel variant and three other intron variants (IVS9 + 32insA, IVS11 + 163delC, and IVS12 + 30C>T). The pathogenicity and some structural features of Q375R were analyzed using bioinformatics tools including SIFT, PolyPhen, Mutpred, MutationTaster, nSSNP Analyzer, SNP effect, 3DLigandSite, GeneSplicer, Human Splicing Finder, MaxEntScan, and FSPLICE. According to the SIFT, PolyPhen, Mutpred, and MutationTaster reports, Q375R could be disease-causing. SNAP predicted Q375R as an intermediate damaging variation and nSSNP Analyzer predicted this variation to be neutral. I-Mutant3.0, FoldX, and Mustab showed a decrease in phenylalanine hydroxylase stability upon Q375R alteration. 3DLigandSite predicted that phenylalanine hydroxylase binding sites vary in mutant and wild-type proteins. Q375R could be considered an effective factor in the structure and function of phenylalanine hydroxylase. This may be useful in clinical detection of phenylketonuria in Iranian patients and their at-risk relatives. However, we need to do complementary in vitro and in vivo functional assessments for the evaluation and validation of the effects of this variation on phenylalanine hydroxylase function and structure.

Exposure to cadmium can cause cardiovascular disorders, including possible risks of angiogenesis abnormalities associated with Hypoxia Inducible Factor (HIF), in pregnancy period. This study aimed to investigate the effects of resistance and endurance training on HIF-1α and vascular endothelial growth factor (VEGF) levels in heart tissue of pregnant rats with cadmium toxicity. In this study, 48 female rats were selected and divided into 6 groups of 8 rats, including (1) cadmium, (2) resistance training, (3) resistance training with cadmium consumption, (4) endurance training, (5) endurance training with cadmium consumption, and (6) control. Groups 2, 3, 4, and 5 performed three weeks of resistance and endurance training, five sessions per week. Also, groups 1, 3, and 5 received cadmium as cadmium chloride dissolved in water as 400 mg per liter through drinking water. To analyze the data, Kolmogorov-Smirnov, One-way ANOVA, and Tukey’s post hoc tests were used and P ≤ 0.05 was considered statistically significant. Cadmium had a significant effect on the increase of HIF1-α and VEGF (P = 0.001); resistance training had a significant effect on the increase of HIF1-α (P = 0.03), resistance training (P = 0.02) and endurance training (P = 0.001) had a significant effect on the increase of VEGF. Also, resistance training with cadmium consumption as well as endurance training with cadmium consumption significantly decreased HIF1-α (P = 0.001) and VEGF (P = 0.001), compared with cadmium consumption. Cadmium appears to lead to an abnormal increase in HIF-1α and VEGF in the heart tissues of pregnant rats, which can stimulate abnormal angiogenesis of tumors; however, resistance and endurance training can reduce the severity of cadmium effects on abnormal angiogenesis of the heart tissue of pregnant rats.

Uterine leiomyoma (ULM) is one of the most common medical conditions where the molecular pathogenesis in women in still unknown. The aim of the present investigation was to demonstrate the association between catechol-methyltransferase (COMT) (rs4680 G>A) gene polymorphism with the risk of ULM. In the present case-control study we included 200 peripheral blood samples of women, which consisted of 100 women with ULM and 100 women who were healthy. The genotype frequencies were assessed using polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) method. The genotype frequencies of COMT (rs4680) gene polymorphism AA + GG vs. GA revealed a significant association with the risk of ULM (OR = 2.453; CI = 1.274 - 4.723, P = 0.006). The findings suggested that the COMT (rs4680) gene polymorphism may be contributed as a predisposing risk factor to ULM.

Pseudomonas aeruginosa is a human and plant pathogen. The aim of this study was to investigate the antibacterial effects of ethanol and ethyl acetate extracts of Myrtus communis against antibiotic-resistant P. aeruginosa isolates. The plant was collected from the plains of Kerman province, and its ethanol and ethyl acetate extracts were prepared using a rotary machine. Pseudomonas aeruginosa strains were isolated from urine and blood samples of patients in Zabol, Iran. Antibiotic resistance pattern was determined by the agar diffusion method. Finally, the minimum inhibitory concentration (MIC) and minimum trace concentration (MTC) were determined by the microdilution method. The antibiotic-resistance patterns of the standard and clinical strains showed that P. aeruginosa was resistant to all antibiotics at the following rates: Azithromycin (25%), ampicillin (12.5%), gentamycin (0%), amoxi-clav (12.5%), cefazolin (12%), and amikacin (12.5%). The study of the effect of ethanol extract on clinical and standard P. aeruginosa strains showed that the MIC of the ethanolic extract against the standard strain of P. aeruginosa was 25 ppm. The results of this study showed good antimicrobial effects of the plant extracts against antibiotic-resistant P. aeruginosa, which can be used to treat Pseudomonas infections.