Invitro Antibacterial Activity of the Prosopis Juliflora Seed Pods on Some Common Pathogens

Introduction: Prosopis juliflora is probably the most widespread species of genus Prosopis and it is a good source of compounds that have been shown to be pharmacologically active. This plant has been used as a traditional treatment for several diseases. Aim: To investigate the in-vitro antibacterial activity of the P. juliflora seed pods from Bushehr, South West of Iran. Materials and Methods: In the present study, the antibacterial activity of P. juliflora seed pods extract was tested against Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli and Pseudomonas aeruginosa. The minimum inhibitory concentration (MIC) of the extract was determined for each test microorganism. Results: P. juliflora seed pods extract exhibited antibacterial activity against all four test organisms. The MIC of the extract was 0.312 mg/ml and 0.078 mg/ml for S. aureus and S. epidermidis, respectively and 1.25 mg/ml for both E.coli and P.aeruginosa. Conclusion: P. juliflora seed pods from Bushehr, South West of Iran could be an appropriate source of antibacterial compounds that makes it a promising candidate for further studies

Characterisation of caprine islet-related genes and enhancement of islet viability and function by heme oxygenase 1 gene

Type 1 diabetes is an important health issue since diabetes is related with serious mortality and morbidity worldwide. One promising approach to treat the disease would be transplantation of the islets which are isolated from the donor pancreas into the patient liver via portal vein. However, a high loss of islets after isolation and transplantation is commonly noted which negatively affects the transplant outcomes. Heme oxygenase 1 (HO-1) is a stress protein induced as a protective mechanism in response to a variety of stimuli. Hence, targeted induction of this protein may be considered as a valuable therapeutic strategy for the protection against inflammatory processes and oxidative tissue damages. The first aim of the current study was to sequence goat HO-1 mRNA. Once the HO-1 mRNA was sequenced, the vector encoding goat HO-1 was constructed and later transfected into the islets. Insulin is the most important hormone of the pancreas, hence, the second objective of the study was aimed to clone and sequence a cDNA encoding an insulin protein in the caprine species. The study was also designed to investigate the transient response of the caprine islets for insulin secretion as well as insulin gene expression after glucose exposure in short period (1 h). The current study also aimed to determine and differentiate the insulin secretion and cell death in association with the size of caprine islets. In this regard, caprine islets were isolated by collagenase digestion and purified by discontinuous Ficoll density gradient centrifugation. Isolated caprine islets were transfected with expression vector containing goat HO-1 gene and cultured for 5 days. The efficacy of GFP transfer to islets was quantified by flow cytometry. Western blots were applied to verify the expression of HO-1 protein. Glucose stimulated insulin release was measured using insulin ELISA assays. Meanwhile, the islet viability was evaluated by the Cell-Titer blue viability assay. Goat HO-1 cDNA was encoding a 288-amino acid protein with a predicted molecular mass of 32.75 kD. The cDNA of goat insulin was successfully amplified using a pair of specifically designed primers. Pairwise comparison showed that goat insulin protein (partially sequenced) was highly similar to insulin protein deposited in public databases, especially cattle preproinsulin with 93% homology. Goat insulin was also similar to human preproinsulin with 72% identity. It was shown that stimulation of insulinproducing cells with high glucose concentrations for only an hour resulted in transient elevation of insulin gene transcription, as verified by measurements of insulin mRNA levels. It was found that isolated caprine islets of large diameter could not be maintained efficiently in culture compared to small islets. After 48 h of culture, small islets showed 2.33% necrosis while a large proportion of necrotic cells was detected in cultured large islets (29.5%). The small and large islets showed an apoptotic death pattern of 5.21 and 7.34%, respectively. Small islets were 92.46% viable, while the viability of the large islets was 63.16%. At 48 h post isolation, under basal conditions, the small islets release 1.39 ± 0.2 ng/IE insulin. With a high glucose concentration, the secreted insulin increased to 2.95 ± 0.33 ng/IE. For the large islet equivalencies, the insulin release under basal conditions and with a high glucose concentration were 0.489 ± 0.2 and 1.01 ± 0.26 ng/IE, respectively. Based on the results, after 2 days of culture, the insulin release with low and high glucose stimulation in large and small islets decreased significantly which clearly indicated that the islet function decreased gradually over time in in vitro culture. In the present study, to ensure the lipidmediated transfection efficiency into the islets, optimization was first done using a reporter protein (Green Fluorescent Protein, GFP). After optimization, transfection and expression of the goat native HO-1 protein in the islet were investigated to determine islet functional outcome after 5 days of cultures in vitro. The insulin stimulation index (SI) in control islets (non-transfected) was 2.02 ± 0.026 while the SI in GFP and HO-1 transfected islets was 1.97 ± 0.026 and 2.07 ± 0.030, respectively. In conclusion, caprine islets could be suitable alternatives for human transplantation with regard to high similarity of the insulin gene to that of pig and human. The study showed that small islets were superior large islets in viability and insulin release under low and high glucose conditions in vitro culture 48 h postisolation. Therefore, protection from core cell death in transplanted islets may improve the success of transplantation by reducing the routine non-functionality of the grafted islets. Transfection of caprine islets with native HO-1 can improve viability and function of the cultured islets which might subsequently increase success of islet transplantation

Size-related assessment on viability and insulin secretion of caprine islets in vitro

The successful isolation, purification, and culture of caprine islets has recently been reported. The present study shows arange of size distribution in caprine islet diameter from 50 to 250 μm, in which 80% of the total islet yield was comprised of small islets. Caprine islets were isolated and purified. Islets were handpicked and the diameter of the islets was recorded using light microscopy. Viablility of the islets was analyzed by confocal microscopy. Insulin secretion assay was carried out and analyzed by ELISA. When tested at 48 h after isolation, these small islets were 29.3% more viable compared to the large-sized islets. Large islets showed a high ratio (P < 0.01) of central core necrosis (29.5% ± 1.92) whilst no significant core death was observed in small islets (2.33% ± 0.59). The annexin assay demonstrated 5.21% ± 0.97 and 7.34% ± 0.78 apoptotic death for small and large islets, respectively. During static incubation, small islets released 2.89-fold (1.39 ± 0.2 ng/IE) higher insulin level under low glucose induction (3.3 mm) and simultaneously 2.92-fold (2.95 ± 0.33 ng/IE) more insulin under high glucose condition (16.7 mm) in comparison to large islets at the same islet equivalents (P < 0.05). The present findings evidenced the superior quality of smaller caprine islets compared to larger ones under an optimized basal maintenance condition. As it is equally important to preserve the quality of larger caprine islets, this work warrants further investigation on special culture conditions to support these islets

Emerging Role of Enhancer RNAs as Potential Diagnostic and Prognostic Biomarkers in Cancer

Enhancers are distal cis-acting elements that are commonly recognized to regulate gene expression via cooperation with promoters. Along with regulating gene expression, enhancers can be transcribed and generate a class of non-coding RNAs called enhancer RNAs (eRNAs). The current discovery of abundant tissue-specific transcription of enhancers in various diseases such as cancers raises questions about the potential role of eRNAs in disease diagnosis and therapy. This review aimed to demonstrate the current understanding of eRNAs in cancer research with a focus on the potential roles of eRNAs as prognostic and diagnostic biomarkers in cancers

Heterotypic tumor spheroids: a platform for nanomedicine evaluation

Abstract Nanomedicine has emerged as a promising therapeutic approach, but its translation to the clinic has been hindered by the lack of cellular models to anticipate how tumor cells will respond to therapy. Three-dimensional (3D) cell culture models are thought to more accurately recapitulate key features of primary tumors than two-dimensional (2D) cultures. Heterotypic 3D tumor spheroids, composed of multiple cell types, have become more popular than homotypic spheroids, which consist of a single cell type, as a superior model for mimicking in vivo tumor heterogeneity and physiology. The stromal interactions demonstrated in heterotypic 3D tumor spheroids can affect various aspects, including response to therapy, cancer progression, nanomedicine penetration, and drug resistance. Accordingly, to design more effective anticancer nanomedicinal therapeutics, not only tumor cells but also stromal cells (e.g., fibroblasts and immune cells) should be considered to create a more physiologically relevant in vivo microenvironment. This review aims to demonstrate current knowledge of heterotypic 3D tumor spheroids in cancer research, to illustrate current advances in utilizing these tumor models as a novel and versatile platform for in vitro evaluation of nanomedicine-based therapeutics in cancer research, and to discuss challenges, guidelines, and future directions in this field. Graphical Abstrac

In vitro assessment of Pediococcus acidilactici Kp10 for its potential use in the food industry

Background: Selection of a microbial strain for the incorporation into food products requires in vitro and in vivo evaluations. A bacteriocin-producing lactic acid bacterium (LAB), Pediococcus acidilactici Kp10, isolated from a traditional dried curd was assessed in vitro for its beneficial properties as a potential probiotic and starter culture. The inhibitory spectra of the bacterial strain against different gram-positive and gram-negative bacteria, its cell surface hydrophobicity and resistance to phenol, its haemolytic, amylolytic and proteolytic activities, ability to produce acid and coagulate milk together with its enzymatic characteristics and adhesion property were all evaluated in vitro. Results: P. acidilactici Kp10 was moderately tolerant to phenol and adhere to mammalian epithelial cells (Vero cells and ileal mucosal epithelium). The bacterium also exhibited antimicrobial activity against several gram-positive and gram-negative food-spoilage and food-borne pathogens such as Listeria monocytgenes ATCC 15313, Salmonella enterica ATCC 13311, Shigella sonnei ATCC 9290, Klebsiella oxytoca ATCC 13182, Enterobacter cloaca ATCC 35030 and Streptococcus pyogenes ATCC 12378. The absence of haemolytic activity and proteinase (trypsin) and the presence of a strong peptidase (leucine-arylamidase) and esterase-lipase (C4 and C8) were observed in this LAB strain. P. acidilactici Kp10 also produced acid, coagulated milk and has demonstrated proteolytic and amylolactic activities. Conclusion: The properties exhibited by P. acidilactici Kp10 suggested its potential application as probiotic and starter culture in the food industry

Primary recovery of a bacteriocin-like inhibitory substance derived from Pediococcus acidilactici Kp10 by an aqueous two-phase system

A polymer–salt aqueous two-phase system (ATPS) consisting of polyethylene–glycol (PEG) with sodium citrate was developed for direct recovery of a bacteriocin-like inhibitory substance (BLIS) from a culture of Pediococcus acidilactici Kp10. The influences of phase composition, tie-line length (TLL), volume ratio (VR), crude sample loading, pH and sodium chloride (NaCl) on the partition behaviour of BLIS was investigated. Under optimum conditions of ATPS, the purification of BLIS was achieved at 26.5% PEG (8000)/11% sodium citrate with a TLL of 46.38% (w/w), VR of 1.8, and 1.8% crude load at pH 7 without the presence of NaCl. BLIS from P. acidilactici Kp10 was successfully purified by the ATPS up to 8.43-fold with a yield of 81.18%. Given that the operation of ATPS is simple, environmentally friendly and cost-effective, as it requires only salts and PEG, it may have potential for industrial applications in the recovery of BLIS from fermentation broth