Molecular Detection of Campylobacter Species: Comparision of 16SrRNA with slyD,cadF, rpoA,and dnaJ Sequencing

Background: Campylobacter spp. are the main cause of human gastroenteritis. The 16SrRNA sequencing is one of fast molecualr method to detect this fastidious. In this study, we compared the sequencing of 16srRNA genewith four housekeeping genestodetect Campylobacter spp. in patients with diarrhea and healthy people.Methods:60 samples of Campylobacter DNA extracted from stool samples of 30 patients with diarrhea and 30 healthy people were used. In order to detect Campylobacter, we designed primers for proliferation of 16SrRNA, cadF, dnaJ, slyD , and rpoA genes using Primer 3, Mega 4.0 and Blast software. Then the PCR products were sequenced using ABI system.Results:The sequencing showed concordance of PCR-products with deposited sequences in the Gene Bank. Among diarrhea patients, 53.3% of samples were significantly (p< 0.05) positive for slyD and cadF genes and 50% of samples were positive using 16SrRNA, rpoA, and dnaJ genes by PCR assay. The average of sensitivity and specificity were found 53.33% and 83.33%, respectively.Conclusion:Due to various copies of repeated sequences of 16SrRNA gene, analyzing its amplicons on electrophoresis may be more difficult than the slyD and cadF genes. According to our results, among the 5 studied genes; the highest detection rate was related to slyD and cadF genes. Although, dnaJ and rpoA genes,instead of 16SrRNA gene, can be considered as appropriate genes for molecular detection of Campylobacter bacteria.     

Endothelial cells' biophysical,biochemical, and chromosomal aberrancies in high‐glucose condition within the diabetic range

To date, many studies have been conducted to find out the underlying mechanisms of hyperglycemia‐induced complications in diabetes mellitus, attributed to the cellular pathologies of different cells—especially endothelial cells. However, there are still many ambiguities and unresolved issues to be clarified. Here, we investigated the alteration in biophysical and biochemical properties in human umbilical vein endothelial cells exposed to a high‐glucose concentration (30mM), comparable to glucose content in type 2 diabetes mellitus, over a course of 120 hours. In addition to a reduction in the rate of cell viability and induction of oxidative stress orchestrated by the high‐glucose condition, the dynamic of the fatty acid profile—including polyunsaturated, monounsaturated, and saturated fatty acids—was also altered in favor of saturated fatty acids. Genetic imbalances were also detected at chromosomal level in the cells exposed to the abnormal concentration of glucose after 120 hours. Moreover, the number of tip cells (CD31⁺/CD34⁺) and in vitro tubulogenesis capability negatively diminished in comparison to parallel control groups. We found that diabetic hyperglycemia was associated with a decrease in the cell‐cell tight junction and upregulation in vascular endothelial cadherin and zonula occludens (ZO)‐1 molecules after 72 and 120 hours of exposure to the abnormal glucose concentration, which resulted in a profound reduction in transendothelial electrical resistance. The surface plasmon resonance analysis of the human umbilical vein endothelial cells immobilized on gold‐coated sensor chips confirmed the loosening of the cell to cell intercellular junction as well as stable attachment of each cell to the basal surface. Our findings highlighted the disturbing effects of a diabetic hyperglycemia on either biochemical or biophysical properties of endothelial cells.     

Functional in vitro characterization of small extracellular vesicles isolated from menstrual blood-derived mesenchymal stem/stromal cells

Menstrual blood is a rich source of mesenchymal stem/stromal cells (MenSCs) with a diverse potential to differentiate into various cell types. Similar to other cells, MenSCs produce extracellular vesicles from which, small extracellular vesicles have attracted much interest due to their therapeutic and regenerative capacities. Here using in vitro approaches, several properties of MenSC-derived small extracellular vesicles (MEX) have been investigated. HUVEC angiogenesis assay was used to evaluate the proangiogenic function of MEX. The immune regulatory property of MEX was assessed using a T cell proliferation assay. Proliferation, migration, and gene expression of primary fibroblasts were selected to determine the scar-related activity of MEX. Finally, the anti-cancer effect of MEX on the proliferation of cancerous cell lines was tested. Our results demonstrated that the small extracellular vesicles isolated from MenSCs have proangiogenic and immune-suppressive abilities. Moreover, these vesicles performed as an anti-proliferative agent for cancerous cell lines. MEX was also able to reduce the migration of primary fibroblasts. In summary, MEX has shown promising in vitro characteristics for regenerative applications. They may offer a great cell-free strategy with therapeutic potential for a diverse range of diseases. For future therapeutic applications and further clinical translation, more studies are needed to elucidate the involved mechanisms.     

Recognition of (Sesc) for Easy Identification of Staphylococcus Epidermidis and Molecular and Phenotypic Study of Β-Lactam Resistance in Staphylococcus Epidermidis Isolates in Isfahan

Background:Not only is it crucial to rapidly detect Staphylococcus epidermidis (S. epidermidis) isolates from a broad range of bacteria, but recognizing resistance agents can greatly improve current diagnostic and therapeutic strategies.Methods:The current cross-sectional study investigated 120 clinical isolates from a nosocomial S. epidermidis infection. The isolates were identified using common biochemical tests, and specific S. epidermidis surface protein C (SesC) primers were used to confirm the presence of S. epidermidis. PCR and special primers were used to detect the β-lactamase gene (blaZ). Methicillin resistance was measured using the agar screening method and antibiotic susceptibility was measured by disk diffusion. Results:100 samples were characterized as S. epidermidis using a phenotypic and genotypic methods. From the 100 specimens examined, 80% contained blaZ. According to agar screening, 60% of isolates were methicillin-resistant. S. epidermidis isolates demonstrated the highest resistance to penicillin (93%) and the highest sensitivity to cefazolin (39%).Conclusion:The increased resistance to β-lactam antibiotics in S. epidermidis isolates is alarming, and certain precautions should be taken by healthcare systems to continuously monitor the antimicrobial pattern of S. epidermidis, so that an appropriate drug treatment can be established.Key Words: Antibiotic resistance, β-lactam, Staphylococcus epidermidis     

A phenetic analysis on the genus Lolium (Poaceae) in Iran

In this study, 68 specimens of the genus Lolium were scored for 27 characters, comprising 12 vegetative, 12 inflorescence and 3 seed characters. The aim of the study was to investigate the species relationships within the genus in Iran. The data were analysed using principal components analysis (PCA) and cluster analysis. Both analyses separated L. perenne, L. persicum, L. temulentum, L. multiflorum from each other. According to previous authors, L. rigidum and L. loliaceum show little difference from each other and are not separable to a species level. In this study these two species separated clearly from the other species, but were most closely related to L. perenne following cluster analysis and to L. multiflorum following PCA. L. persicum was found to be the most distinct species within the genus Lolium in Iran.     

Mesenchymal stem cells engineered by modified polyethylenimine polymer for targeted cancer gene therapy,in vitro and in vivo

Cell-based delivery system is a promising strategy to protect therapeutic agents from the immune system and provide targeted delivery. Mesenchymal stem cells (MSCs) have recently been introduced as an encouraging vehicle in cell-based gene therapy due to their unique features including tumor-tropic property and migratory ability. However, gene transfer into MSCs is limited due to low efficiency and cytotoxicity of carriers. In this study, we designed a novel delivery system based on polyethylenimine (PEI₂₅) to improve these features of carrier and transfect plasmid encoding TRAIL to MSCs. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a death ligand of TNF family with selective effect on cancerous cells. Then, death induction and migration ability of TRAIL-expressing MSCs was studied in melanoma cells. The effect of engineered-MSCs as an antitumor vehicle was also investigated in mice bearing melanoma cells. Our findings indicated that heterocyclic amine derivative of PEI₂₅ showed significant improvement in MSCs viability determined by MTT assay and gene expression using fluorescent microscopy, flow cytometry, and Western blot analysis. We observed that engineered-MSCs could migrate toward and induce cell death in B16F0 cells in vitro. The single administration of TRAIL-expressing MSCs could delay tumor appearance and efficiently reduce tumor weights. Hematoxylin and eosin staining of tumor sections revealed extensive neoplastic cells necrosis. Furthermore, engineered-MSCs could migrate and localize to tumors sites within 5 days. Our results indicated that MSCs engineered by modified-PEI/TRAIL complexes could be considered as a promising cellular vehicle for targeted tumor suppression.     

Anti-hyperglycemic effects of Eryngium billardierei F. Delaroche extract on insulin-resistance HepG2 cells in vitro

Background

Insulin resistance as a major problem is associated with type 2 diabetes mellitus. This study investigated the effect of Eryngium billardierei on insulin-resistance induced HepG2 cells.

Methods and results

MTT method was used to evaluate the viability of HepG2 cells treated with various doses of E. billardierei extract. An insulin-resistance model was established in HepG2 cells. Next, MTT assay and Acridine orange staining were performed to investigate the viability of cells in the vicinity of different concentrations of insulin, pioglitazone, and E. billardierei extract in an insulin-resistance media. The glucose uptake test was performed to select the optimal insulin concentration. Expression levels of IR, G6Pase, and PEPCK genes were assessed by real-time RT-PCR. According to obtained data, E. billardierei at concentrations of 0.5 and 1 mg/mL show no toxicity on cells. Furthermore, based on MTT assay and glucose uptake test 10^?5 mol/L insulin was chosen as the model group to induce insulin-resistance in HepG2 cells for gene expression analysis. Finally, 1 mg/mL E. billardierei not only induced no cytotoxicity but also showed an increase in the expression of IR as well as a reduction in G6Pase and PEPCK level compared to the control and model groups.

Conclusions

The obtained data indicated that 1 mg/mL E. billardierei might have an anti-insulin resistance effect on insulin-resistance HepG2 cells in vitro and could be a promising candidate with anti-hyperglycemic properties for diabetes treatments.

Graphical abstract

     

Yeast cells for encapsulation of bioactive compounds in food products: A review

Nowadays bioactive compounds have gained great attention in food and drug industries owing to their health aspects as well as antimicrobial and antioxidant attributes. Nevertheless, their bioavailability, bioactivity, and stability can be affected in different conditions and during storage. In addition, some bioactive compounds have undesirable flavor that restrict their application especially at high dosage in food products. Therefore, food industry needs to find novel techniques to overcome these problems. Microencapsulation is a technique, which can fulfill the mentioned requirements. Also, there are many wall materials for use in encapsulation procedure such as proteins, carbohydrates, lipids, and various kinds of polymers. The utilization of food-grade and safe carriers have attracted great interest for encapsulation of food ingredients. Yeast cells are known as a novel carrier for microencapsulation of bioactive compounds with benefits such as controlled release, protection of core substances without a significant effect on sensory properties of food products. Saccharomyces cerevisiae was abundantly used as a suitable carrier for food ingredients. Whole cells as well as cell particles like cell wall and plasma membrane can act as a wall material in encapsulation process. Compared to other wall materials, yeast cells are biodegradable, have better protection for bioactive compounds and the process of microencapsulation by them is relatively simple. The encapsulation efficiency can be improved by applying some pretreatments of yeast cells. In this article, the potential application of yeast cells as an encapsulating material for encapsulation of bioactive compounds is reviewed.