Pathobiology Research
Year:2015 | Volume:17 | Issue:4|Papers Count:8

Objective: Prostate cancer is the fifth most common cancer. In 2012, it was the second leading cause of cancer death for men worldwide. The PI3K/AKT pathway plays an essential role in pathogenesis of prostate cancer; the key role of this pathway in cancer progression makes it an attractive target for prostate cancer therapy. MicroRNAs (miRNAs) that regulate gene expression have a special ability to simultaneously control multiple genes and pathways which make them candidates for therapeutics. This study aims to determine miRNAs which target the PI3K/AKT pathway and evaluate them in prostate cancer cell lines.Methods: In order to determine an effective miRNA for the PI3K/AKT pathway, we assessed six genes from this pathway which have been proposed as drug targets in ten different prediction algorithms. Next, the candidate miRNAs were analyzed in expression profile and pathway analysis databases. Expression of candidate miRNAs in control and prostate cancer cell lines were subsequently evaluated.Results: According to bioinformatics, the miR-29 family could target the most genes from this list. Other bioinformatic estimates confirmed these results. The miR-29 family showed significant downregulation in prostate cancer cell lines LNCAP, PC3 and DU-145 compared to control samples.Conclusion: These results propose the possibility of using the miR-29 family to inhibit the PI3K/AKT pathway in prostate cancer.

Objective: Dendrimers are three-dimensional nanostructures that have numerous applications in medicine, including drug delivery and imaging. Although anionic dendrimer polyethylene glycol–citrate has a high potential to increase solubility of waterinsoluble drugs and drug delivery, its multi-step synthesis procedure is time consuming.In addition, toxic substances such as dichloromethane are used in its synthesis procedure.In this study, we have developed a simple one-step synthesis method using green chemistry.Methods: We examined four different methods to improve the synthesis method of this dendrimer. Products were characterized by FTIR, LC-MS and DLS. Cytotoxicity was assessed by the XTT method.Results: We synthesized a G2 polyethylene glycol–citrate dendrimer in one-step without purifying G1. This process was chosen as a beneficial method for synthesis of the G2 dendrimer. When compared with previous methods, this procedure had higher efficiency and greatly reduced response. This procedure used nontoxic materials. XTT assay results showed that this dendrimer created by green chemistry had no cytotoxicity in Hela and Vero cells up to a concentration of 800 μM.Conclusion: One-step synthesis of anionic polyethylene glycol-citrate G2 dendrimer is a simple, beneficial production method. The dendrimer is biocompatible and can be used as a suitable carrier for drug delivery purposes.

Objective: In order to improve the water solubility and bioavailability of curcumin in cancer therapy, we prepared and tested a novel waterborne cationic polyurethane (PU) as a nano-carrier for curcumin loading (CU-PU). We studied the effect of this prepared nano-drug on melanoma (F10B16) and fibroblasts cells (L929).Methods: Morphology, size and cell internalization ability of the prepared nanoparticles were analyzed by zetasizer, SEM, AFM and fluorescent microscopy, respectively. We anticipated that curcumin was loaded in the hydrophobic core of the PU carrier. Next, the suitable dose and therapeutic effects of CU-PU for both skin cancer and normal cell lines were evaluated by the MTT assay and real-time PCR.Results: The average diameters and polydispersity of the nanoparticles were 62.37 ± 1.7 nm and 0.080 ± 2.1 at 25 C, respectively. The drug encapsulation efficiency was 87 ±0.2%. The morphological analysis confirmed both a spherical shape and good dispersion without remarkable aggregation. The MTT assay results showed that the IC50 at 24 hours was 36.2 mM, whereas it was 25.4 mM at 48 hours. Real-time PCR results indicated that the CU-PU significantly decreased mRNA expressions of VEGF, Bcl-2, MMP-9 and COX-2 genes. An increase in mRNA expression of the BAX gene was also observed.Conclusion: Our result provided acceptable evidence for cell proliferation inhibition and the apoptotic effect of CU-PU on skin cancer cells. There were no adverse effects detected for normal cells.

Objective: Endometriosis is a common disease in which the endometrial stroma and glands grow abnormally outside the uterine cavity. The establishment of animal models can be effective for determining the etiology of endometriosis. In this study we compare two endometriosis models using endometrial fragments and isolated cultured endometrial cells.Methods: We obtained endometrial tissues that were in the proliferative or secretory phases from women who underwent hysteroscopies for benign reasons at Imam Khomeini Hospital (Tehran). Following confirmation of the tissue's normality, we cut the tissues into 2 mm cube pieces. The remainder of endometrial tissues were used for isolation and cultured to the fourth passage. In the first model of endometriosis, the endometrial tissue fragments and in the second model, 2×106 isolated endometrial cells were subcutaneously transplanted into gamma irradiated mice. The mice were kept under controlled, sterile conditions for 20 days. The mice sera were collected before and after transplantation for assessment of 17b estradiol. The ectopic tissues in both models were assessed for morphological staining using hematoxylin and eosin, as well as periodic acid Schiff (PAS) for gland secretion. The gland sections per mm2 were analyzed.Results: At 20 days after tissue transplantation, we observed endometrial-like glands in the subcutaneous tissue of both endometriosis models. The number of gland sections was 57.55 ± 17.18/mm2 for the first model and 271.57 ± 77.98/mm2 for the second model.This result was significantly higher in the second model when compared to the first model. Gland secretion was positive for PAS. The level of 17b estradiol was higher in both models compared to the control group. This level was significantly higher in the second model compared to the first (P≤0.05).Conclusion: The endometriosis model that used cultured endometrial cells showed more efficiency in morphology, gland formation and level of17b estradiol.

CtxB (Cholera toxin B subunit) contributes to a vaccine's efficacy by stimulating production of the anti-CtxB antibody. Various attempts have been made to increase production of this antibody. Chitosan is a mucoadhesive polysaccharide that has tremendous potential for oral vaccine delivery in terms of its exclusive features that include biocompatibility, biodegradability, high charge density and non-toxicity. We investigated the potential for chitosan nanofibers and nanocapsules as novel carrier systems for the oral delivery of CtxB.Methods: Antigen-containing chitosan nanofibers were prepared by electrospinning a chitosan/AcOH solution. Encapsulation of the antigen inside the chitosan nanofibers was confirmed through infrared spectroscopy analysis (FTIR). Guinea pigs were immunized with free antigen and CtxB antigen or antigen alone by direct administration of antigencontaining chitosan nanofibers into the buccal cavity. Serum immunoglobulin G (IgG) and intestinal immunoglobulin A (IgA) antibody responses were determined Results: The results indicated that antigen in the chitosan nanofibers or nanocapsules elicited very high IgA and IgG responses. No detectable IgA and IgG responses were obtained after oral immunization with CtxB. The results of the antibody titer were analyzed using the ANOVA and LSD tests.Conclusion: CtxB inside the nanofiber increased antibody production when administered orally. This system might be used for delivery of other antigens.

Objective: More similarity to in vivo may help to increase the proliferation and differentiation of cells at in vitro culture. The present study has investigated the effect of a dynamic culture medium and nano hydroxyapatite (nHA) presence on proliferation and differentiation of mesenchymal stem cells (MSCs) to bone cells using electrospun polycaprolactone (PCL) scaffolds.Methods: We prepared PCL and PCL-nHA scaffolds by electrospinning. After static culturing of the scaffolds with MSCs, the scaffolds, were divided into two groups of static and dynamic cultures. The dynamic culture scaffolds were placed on a shaker. Cell proliferation and differentiation at days 3, 7 and 14 were investigated by MTT, and the calcium and alkaline phosphatase assays.Results: The obtained results from the MTT assay on day 14 showed an increase of 1.1 times in cell proliferation in the dynamic culture compared to the static culture. During this period, the calcium content produced by cells in the dynamic culture at day 14 were 1.23 times higher for PCL scaffolds and 1.46 times higher for PCL-nHA scaffolds compared to the static culture. Alkaline phosphatase levels for the dynamic state PCL scaffold were 1.24 times more and for PCL-nHA scaffolds were 1.28 times more compared to the static culture at day 14.Conclusion: The obtained results from dynamic culture, showed higher proliferation and differentiation of stem cells to bone for both PCL and PCL-nHA scaffolds compared to the static culture. The amount of cell proliferation and differentiation in the scaffolds that contained nHA was more than scaffolds that did not have nHA.

Objective: Acetaminophen (APAP) overdose causes acute liver injuries. Studies show that stem cell factor (SCF) and its receptor, c-Kit, enhance liver recovery from APAPinduced injuries in mice. In this study we explore the effect of SCF on activity of glutathione S-transferase (GSTs) enzymes which are considered to be important in APAP metabolism.Methods: We divided 45 Balb/c mice into three groups. Within each group there were three sub-groups of five mice per subgroup. The groups included: 1. APAP (300 mg/kg B.W., i.p.); 2. SCF (40 μg/kg B.W., i.p.) given.30 minutes after APAP (300 mg/kg B.W., i.p.), and 3.control mice treated with normal saline. The mice were sacrificed at 1, 12 and 24 hours, respectively. Hepatotoxicity was evaluated in the 24 hour group by histopathology and assessment of biochemical serum markers (ALT and AST). We assessed the levels of SCF receptor (c-Kit) protein and GST enzyme activities in the liver tissues.Results: Hepatotoxicity was induced by APAP (300 mg/kg, B.W) as evident by both histopathological observations and a significant (p<0.05) increase in serum ALT and AST levels, which were reversed by SCF administered post-APAP. SCF administration after APAP administration significantly increased GSTs enzyme activity levels by 24 hours, however it led to a significant decrease in c-Kit protein level compared to the control and APAP groups.Conclusion: Our data suggest that SCF binding to its receptor (c-Kit) on liver cells may attenuate APAP-induced liver injuries by increasing GST activities in the livers of mice.

Objective: The incidence of breast cancer is approximately one million which makes this cancer one of the most common among women worldwide. Breast cancer comprises 7% of the total death rate caused by cancers. Several strategies that use tumor-associated antigen (TAA) vaccination and early detection of breast cancer are clinically being developed. Breast cancer is caused by increased over expression of certain genes. HER-2 is a tyrosine kinase receptor in the epidermal growth factor family. The role of HER-2 in breast cancer has been extensively studied. HER-2 is found in 25%-30% of breast cancer patients. Herceptin, a human antibody, is used as a therapeutic target for HER-2. The purpose of this study is to produce recombinant protein HER-2 for early detection of breast cancer cells.Methods: We used specific primers to amplify the HER-2 gene. The amplified gene was cloned into pET28a as an expression vector. Cloning was confirmed by restriction analysis and sequencing. Expression was induced using IPTG and the recombinant protein was analyzed by SDS-PAGE.Results: Cloning of the HER-2 gene was confirmed by enzyme digestion and sequencing.The gene was expressed in E.coli BL21 DE3. The pET-28a vector which contained the HER-2 gene showed a high level of expression. The recombinant protein was confirmed by Western blot analysis.Conclusions: A portion of the HER-2 gene was expressed as a recombinant in E.coli.This could be a good diagnostic test for breast cancer.