Avicenna Journal of Medical Biotechnology
Volume:7 Issue: 1, Jan-Mar 2015

Prostate Specific Antigen (PSA) is an important laboratory marker for diagnosis of prostatic cancer. Thus, development of diagnostic tools specific for PSAplays an important role in screening, monitoring and early diagnosis of prostate cancer.In this paper, the production and characterization of a panel of murine monoclonalantibodies (mAbs) against PSA have been presented. Balb/c mice were immunized with PSA, which was purified from seminal plasma. Splenocytes of hyperimmunized mice were extracted and fused with Sp2/0 cells. By adding selective HAT medium, hybridoma cells were established and positiveclones were selected by ELISA after four times of cloning. The isotypes of producedmAbs were determined by ELISA and then purified from ascitic fluids using Hi-Trapprotein G column. The reactivities of the mAbs were examined with the purified PSAand seminal plasma by ELISA and western blot techniques. Furthermore, thereactivities of the mAbs were assessed in Prostate Cancer (PCa), Benign Prostatic Hyperplasia (BPH) and brain cancer tissues by Immunohistochemistry (IHC). Five anti-PSA mAbs (clones: 2G2-B2, 2F9-F4, 2D6-E8, IgG1/К) and clones (2C8-E9, 2G3-E2, IgG2a/К) were produced and characterized. All mAbs, except 2F9- F4 detected the expression of PSA in PCa and BPH tissues and none of them reactedwith PSA in brain cancer tissue in IHC. Besides, all mAbs could detect a protein bandaround 33 kDa in human seminal plasma in western blot. These mAbs can specifically recognize PSA and may serve as a component of PSA diagnostic kit in various biological fluids.

Prostate cancer is one of the most widespread cancers in men and is fundamentally a genetic disease. Identifying regulators in cancer using novel systems biology approaches will potentially lead to new insight into this disease. It was sought to address this by inferring gene regulatory networks (GRNs). Moreover, dynamicalanalysis of GRNs can explain how regulators change among different conditions, suchas cancer subtypes. In our approach, independent gene regulatory networks from each prostate state were reconstructed using one of the current state-of-art reverse engineering approaches. Next, crucial genes involved in this cancer were highlighted by analyzing each network individually and also in comparison with each other. In this paper, a novel network-based approach was introduced to find critical transcription factors involved in prostate cancer. The results led to detection of 38 essential transcription factors based on hub type variation. Additionally, experimentalevidence was found for 29 of them as well as 9 new transcription factors. The results showed that dynamical analysis of biological networks may provide useful information to gain better understanding of the cell.

Emergence of drug resistance has brought major problems in chemotherapy. Using nutrients in combination with chemotherapy could be beneficial forimprovement of sensitivity of tumors to drug resistance. Soybean-derived isoflavones have been suggested as chemopreventive agents for certain types of cancer, particularlybreast cancer. In this study, the synergistic effects of soy isoflavone extract in combination with docetaxel in murine 4T1 breast tumor model were investigated. In this study, mice were divided into 4 groups (15 mice per group) of control, the dietary Soy Isoflavone Extract (SIE, 100 mg/kg diet), the Docetaxel (DOCE, 10 mg/kg) injection and the combination of dietary soy isoflavone extract and intravenousdocetaxel injection (DOCE+SIE). After 3 injections of docetaxel (once a week), 7 mice were sacrificed to analyze MKI67 gene and protein expressions and the rest weremonitored for diet consumption, tumor growth and survival rates. In DOCE+SIE group, diet consumption was significantly higher than DOCE group. While lifespan showed a trend towards improvement in DOCE+SIE group, nosignificant difference was observed among the 4 studied groups. Tumor volume was not significantly affected in treated groups. A lower but not significant MKI67 protein expression was detected in western blot in DOCE+SIE group. The mRNA expression was not significantly different among groups. The results suggest that the combination of soy isoflavone as an adjunct to docetaxel chemotherapy can be effective in improving diet consumption in breast cancer.

Diabetes Mellitus (DM), simply known as diabetes, refers to a group of metabolic diseases in which there are high blood sugar levels over a prolonged period. In this study, the feasibility and safety of intravenous transplantation of Very SmallEmbryonic Like stem cells (VSELs) were investigated for diabetes repair, and finallythe migration and distribution of these cells in hosts were observed. Mouse bone marrow VSELs were isolated by Fluorescent Activating Cell Sorting (FACS) method by using fluorescent antibodies against CD45, CXCR4 and Sca1markers. Sorted cells were analyzed for expression of oct4 and SSEA1 markers with immunocytochemistry staining method. To determine multilineage differentiation, sorted cells were differentiated to Schwann, osteocyte and beta cells. Ten days after the establishment of a mouse model of pancreas necrosis, DiI-labeled VSELs were injected into these mice via tail vein. Pancreases were harvested 4 weeks after transplantation and the sections of these tissues were observed under fluorescent microscope. It was proved that CD45-, CXCR4+, and Sca1+ sorted cells express oct4 and SSEA1. Our results revealed that intravenously implanted VSELs could migrate intothe pancreas of hosts and survive in the diabetic pancreas. In treated groups, blood glucose decreased significantly for at least two month and the weights of mice increasedgradually. This study provides a strategy for using VSELs for curing diabetes and other regenerative diseases, and the strategy is considered an alternative for otherstem cell types.

Development of tissue engineering and regenerative medicine has led to designing scaffolds and their modification to provide a better microenvironment which mimics the natural niche of the cells. Gelatin surface modification was appliedto improve scaffold flexibility and cytocompatibility. PLLA/PCL aligned fibrous scaffold was fabricated using electrospinningmethod. ADSCs were seeded after O2 plasma treatment and gelatin coating of the scaffolds. The morphological and mechanical properties of blends were assessed byScanning Electron Microscopy (SEM), tensile test and ATR-FTIR. The cells proliferationwas evaluated by MTT assay. Based on the results, it is supposed that gelatin coating is a brilliant method of surface modification which significantly increases the mechanical properties of scaffold without any changes on the construction or on the direction of nanofibers whichconducts cell’s elongation. MTT analysis exhibited that ADSCs attachment, viabilityand proliferation significantly (p<0.05) increased after gelatin treatment. Gelatin surface modification is a highly beneficial method to improvecytocompatibility, flexibility and mechanical features of the scaffolds which doesn’t affectthe nanofibers construction. Proliferation of Adipose Derived Stem Cells (ADSCs) as a remarkable source of stem cells was investigated for the first time on PLLA/PCL hybrid scaffold.

CD19 is a pan B cell marker that is recognized as an attractive target for antibody-based therapy of B-cell disorders including autoimmune disease and hematological malignancies. The object of this study was to stably express the human CD19 antigen in the murine NIH-3T3 cell line aimed to be used as an immunogen in our future study. Total RNA was extracted from Raji cells in which high expression of CD19 was confirmed by flow cytometry. Synthesized cDNA was used for CD19 gene amplification by conventional PCR method using Pfu DNA polymerase. PCR product was ligated to pGEM-T Easy vector and ligation mixture was transformed to DH5α competentbacteria. After blue/white selection, one positive white colony was subjected to plasmid extraction and direct sequencing. Then, CD19 cDNA was sub-cloned into pCMV6-Neo expression vector by double digestion using KpnI and HindIII enzymes. NIH-3T3 mouse fibroblast cell line was subsequently transfected by the construct using Jet-PEI transfection reagent. After 48 hours, surface expression of CD19 was confirmed by flow cytometry and stably transfected cells were selected by G418 antibiotic. Amplification of CD19 cDNA gave rise to 1701 bp amplicon confirmed byalignment to reference sequence in NCBI database. Flow cytometric analysis showed successful transient and stable expression of CD19 on NIH-3T3 cells (29 and 93%, respectively). Stable cell surface expression of human CD19 antigen in a murine NIH- 3T3 cell line may develop a proper immunogene which raises specific anti-CD19 antibody production in the mice immunized sera.

The objective of this study is to establish Raman signatures from pure cultures of different Candida species using Raman Spectroscopy (RS) and use thesesignatures for rapid identification of unknown Candida species. Pure cultures of five Candida species were evaluated using RS to build a limited signature library. ‘Raman Processing’ (RP) software was used for PrincipalComponent Analysis (PCA) and Differential Functional Analysis (DFA). Eleven principal components described at least 95% variance in the spectra. Raman signatures from these known Candida species were able to identify the species of unknown Candida cultures with 100% accuracy. Raman spectroscopy can improve early identification of Candida species and may facilitate early optimal antifungal therapy.