Pathobiology Research
Year:2018 | Volume:21 | Issue:2|Papers Count:7

Aims Growth factor (GFs) delivery with the certain concentration and release kinetic is one of the main challenges in tissue engineering. The aim of this study was the preparation and characterization of smart poly (N-isopropylacrylamide) nanoparticles containing vascular endothelial growth factor for induction of angiogenesis in human bone marrow-derived mesenchymal stem cells. Materials & Methods In this exprimental study, two different formulations of temperaturesensitive Poly (N-isopropylacrylamide) (PNIPAM) nanoparticles (NPs) were synthesized by free radical polymerization technique. Nanoprecipitation and diffusion methods were used to load the vascular endothelial growth factor (VEGF) in PNIPAM NPs. The effects of released VEGF on the differentiation of human bone marrow stem cells (hBMSCs) into endothelial cells in angiogenic, osteogenic, and 50% angiogenic-osteogenic culture medium were investigated, using flow cytometry and light microscope. Statistical analysis was performed, using the GraphPad Prism 6 software. Findings The nanoprecipitation process caused polymer degradation due to using the organic N, N-Dimethylacetamide solvent. The cumulative VEGF released after 72hours for 70%. A total of 10ng/ml VEGF released from PNIPAM nanoparticles, in 2D culture with cell density of 3×104 hBMSCs, after 7 days, leading to the endothelial differentiation, capillary-like tube formation, and expression of 20% vWF as angiogenic marker. Conclusion The PNIPAM NPs have the potential to load and release the angiogenic GFs for induction of angiogenesis in hBMSCs and in osteogenic medium.

Aims Vaccination with the HBs Ag vaccine formulated in the alum adjuvant prevents hepatitis B infection. Adjuvants increase the efficacy of vaccines. Mycobacterium purified protein derivative (PPD) causes dendritic cell maturation. Considering the role of dendritic cells in the induction of immune responses, the aim of present study was to investigate of application of PPD in the formulation of Commercial Hepatitis B vaccine to improve humoral and cellular immune responses. Materials & Methods In this experimental study, 63 balb/C mice of inbred were divided into 7 groups of 9. The Hepatitis B vaccine with doses of 1 and 10μ g of PPD was injected subcutaneously into HBS-ALUM, HBS-ALUM+PPD1μ g, HBS-ALUM+PPD10μ g and proper control groups, three times with two-week intervals. Total IgG in serum samples and IL-4 and IFN-γ cytokines levels and IFN-γ /IL-4 ratio in spleen cell culture were assessed by ELISA method. Statistical analysis was done using one-way analysis of variance by Graf pad prism 6. 1 software. Findings Using PPD in the formulation of HBs vaccine had no effect on IFN-γ cytokine levels but reduced IL-4 and also increased the IFN-γ /IL-4 ratio compared to the commercial vaccine. Also PPD application in HBs vaccine after the first injection enhanced the humoral response, but after the second injection was ineffective so that after the third injection reduced the antibody response compared to the commercial vaccine. Conclusion PPD in the formulation of hepatitis B vaccine can lead the immune response pattern to the T helper cells 1 (Th1). The first injection enhances the antibody response but after third injection suppressed humoral immunity system.

Aims Botulinum neurotoxins are the strongest known bacterial toxins that cause muscle paralysis due to inhibition of acetylcholine release. Design of inhibitors is still pursued as a major strategy for intracellular inhibition of poisoning caused by these toxins. Investigation of the potential function of design inhibitors, pure poison or catalytic area is essential. The aims of present study were expression and purification of recombinant catalytic domain of botulinum neurotoxin type E (BoNT/E) Type E from a synthetic gene. Materials & Methods In this experimental study, the sequence of the botulinum neurotoxin type E light chain was adopted from GeneBank and codons were optimized according to E. coli BL21 (DE3) codon usage. Other bioinformatics tools were exploited to reach the optimum expression of the gene in the mentioned host. The resultant (gene) was then ordered to synthesize and cloning in pET28a (+) expression vector. The recombinant vector was transferred into E. coli BL21 (DE3) host cells. The expression of the protein was induced by addition of IPTG. The expression conditions were changed to obtain a soluble expression of the protein. Then, the protein was purified by an affinity chromatography, followed by a further purification with amicon filter. SDS-PAGE was used to evaluate expression and purification of the protein and Western blotting was performed to confirm the expressed protein. Findings Codon Adaptation Index of the gene increased to 0. 85. The third predicted structure showed good quality. The thermodynamic analysis of the mRNA structure showed that the predicted structure is stable. The soluble expression was obtained in 18° C and 18h induction by 1 mM IPTG. Protein production with higher more than 90% purity was confirmed. Conclusion Optimization of the protein expression conditions resulted in producing the solution in the culture medium by E. coli BL21 as host.

Aims Breast cancer is the most common cancer among women around the world and in Iran and 22% of all cancers in women is included. About 30% of cases with breast cancers are due to the proliferation or excessive expression of HER-2 protein. Chitosan is an environmentally friendly combination, due to its minimal systemic toxicity of peptide or drug delivery, the application is considered. The aims of this study were to manufactur chitosan nanoparticle containing HER-2 recombinant protein, evaluation of the properties and effect of its oral and injection immunization on spleen lymphocytes. Materials & Methods In this experimental study, pET-28a vector containing HER-2 gene was evaluated using PCR by universal primers. Expression of protein in E. coli was induced with IPTG. The recombinant protein was purified using affinity chromatography and evaluated by Western Blotting analysis. Chitosan nanoparticles containing recombinant protein were prepared by inonic gelation method and their size were characterized by DLS. Mice were immunized with nanoparticles and antibody titers were determined by ELISA. The response of lymphocytes in exposure to nanoparticles was evaluated by MTT assay. One way ANOVA, Duncan test and SPSS 22 software were used for statistical analysis. Findings The protein with a molecular weight of 18 kDa was confirmed. Yield of protein was 12mgL-1. Encapsulation efficiency of recombinant protein in nanoparticles was 70%. The average particle size was 205. 2nm. Immunization of mice induced mucosal and humoral immune response. Conclusion Encapsulation efficiency of recombinant protein in nanoparticles is 70%. The average particle size is 205. 2nm. Immunization of mice induces mucosal and humoral immune response HER-2 recombinant protein, due to the cellular and humoral immune response, is a candidate suitable for cancer diagnostic or therapeutic purposes.

Aims Developing an effective vaccine against Human Immunodeficiency Virus (HIV) is necessary. The aim of this study was the immunological evaluation of HIV-1 VLP harboring MPER-V3 in BALB/c mice model. Materials & Methods In the present experimental research, presenting 40 female mice, which were between 6 and 8 weeks old, were used for immunization with VLP MPER-V3. The mice were divided into 8 groups and in each group, 5 mice were considered. Injections were performed three times at three weeks intervals and subcutaneously in a volume of 100μ l per mouse. Two weeks after last injection, mouse blood samples were collected by retro-orbital bleeding and immune responses were evaluated in serum for levels of total IgG and splenocytes for cytokine assay, using ELISA method. The data analysis was performed, using Mann-Whitney test and one-way analysis of variance. Findings The level of total antibody production was very high in all groups that had VLP alone or with adjuvant immunity, having a significant difference with the control group (p<0. 05). IgG1 was the predominant isotype (Th2-biased response) in groups that had VLP injections alone or with adjuvant. Conclusion VLPs can stimulate the humoral immune system in mice immunized with these particles alone or formulated with adjuvant. Also, the level of production of IL-5 in the presence of the vaccine candidate as VLP increased significantly.

Aims Beta-thalassemia is a blood and genetic disorder. Prevalence of beta-thalassemia trait is high in Iranian population. Therefore, for detection of carriers’ status in premarital screening we need to establish reliable and rapid scanning method. The aim of this study is development a rapid method to detect four common beta globin gene mutations in Iranian population by HRM (High-Resolution Melting Analysis. Materials & Methods In this experimental study, eight samples of beta-thalassemic carriers and illness with IVSI-110 (G>A), IVSII-I (G>A), IVSI-5 (G>C) and CD36/37 (-T) genotypes in Homozygote and heterozygote conditions and one control sample (healthy person) were selected. After blood collection, DNA extraction was performed using salting out method. To ensure, the samples were run binary, and two control samples without DNA were used. Primers were designed with Gene Runner 6. 0. 28, Primer Express v 3. 0. 1 3/1) and Primer3 v. 0. 4. 0 and HRM method was used to determine the above mutations. Findings The best concentration of DNA was obtained at 5ng/μ l. In the difference plot step, the differences between the curves were clearly observed in heterozygous, normal and homozygous conditions. Unexpectedly, in the mutation IVSII-IG>A in the melting curve was seen two peaks in the samples; it was due to the presence of a polymorphism near the desired mutation. Conclusion The HRM method is capable of detecting the four common beta globin gene mutations, and, it can differentiate between heterozygote and homozygous in a single mutation. This method is high throughput, which can analyze 96 samples of DNA in 2hours.

Introduction Myocardium tissue is an electroactive tissue capable of transferring electrical signals, which lead to synchronized beating of heart. Electrical impulses originate from sinoatrial node and spread though myocardium to induce mechanical contraction of cardiomyocytes. As the leading cause of death, worldwide, cardiovascular diseases are often accompanied by disruption of electrical integrity of cardiac tissue and arrhythmia. In many arrhythmias, lack of conduction as well as unidirectional conduction result in insufficient intercellular electrical coupling at gap junctions. Due to limitation of conventional treatment methods such as heart transplantation, pathological and therapeutic researches in cardiac electrical disorders have increased in last few years. The aim of this study was to review the last studies in electrical system of heart and its disorder along with the results and the future of the cardiovascular tissue therapy method based on Conductive biomaterial. Conclusion Electrical integrity is essential for normal functioning of the heart. Among the new methods of treating heart failure and improving the electrical integrity of the disorder caused by these defects, tissue engineering with the use of conductive electrical conductive materials has been widely considered along with other methods. Three main types of conductive materials have been used for tissue engineering application: (1) Gold-based materials (2) Carbon-based materials (3) Conductive polymers.