Cell Journal (Yakhteh)

76-84

Authors: Mostafa Ebrahimdoost; Mohsen Mohammadi; Narges Obeidi; Seyed Amin Mohammadi; Gholamreza Khamisipour

Objective: Some cationic anti-microbial peptides show a wide range of cytotoxic action versus malignant cells, which may lead to developing a novel group of antitumor medications. In the present study, the anticancer activity of pleurocidin-like peptide WF3 isoform X2 (AMP-WF3), from the Poecilia Mexicana fish, against leukemic cell line Jurkat was evaluated, and the cytotoxicity compared with the effects on normal cells, including peripheral blood mononuclear cells (PBMCs) and human dermal fibroblast (HDF) cells. Materials and Methods: In this experimental study, cells were treated with various dosages of AMP-WF3 for 24 hours. Using methyl thiazole tetrazolium salt reduction (MTT test), the effects of the AMP-WF3 on cell viability and toxicity were evaluated. The impact of this peptide on apoptotic pathways was examined using flow cytometry and Annexin V-PI stains. Additionally, the relative expression of the P53, P21, and BCL-2 genes was evaluated using a real-time polymerase chain reaction. Results: The Jurkat cell line was more susceptible to AMP-WF3 cytotoxicity [half-maximal inhibitory concentration (IC50)=50 μM], while normal cells (PBMCs and HDF) were less susceptible. Flow cytometry verified that the apoptotic activity of AMP-WF3 on Jurkat cells was significantly higher than that of HDF and PBMCs. Peptide-treated Jurkat cells were associated with increased expression of P21, and P53 genes. In contrast, the changes in P21, P53, and BCL-2 genes differed in PBMCs and HDF cells. In HDF cells, simultaneous increase of P21, P53, and BCL-2, and in PBMCs, only the increase of BCL-2 was observed. Conclusion: Our research showed that AMP-WF3 could be developed as a novel treatment agent with minimum side effects for ALL patients.

92-101

Authors: Javad Firouzi; Abbas Hajifathali; Masoumeh Azimi; Neda Parvini; Fatemeh Ghaemi; Niloufar Shayan Asl; Amir Abbas Hedayati Asl; Majid Safa; Marzieh Ebrahimi

Objective: Natural killer (NK) cells are critical immune cells for acute myeloid leukemia (AML) targeting. However, little is known about the relationship between using checkpoint inhibitors and heat shock protein 70 (Hsp70) as NK cell activators to control AML. Therefore, the study aims to find the best formulation of Hsp70, human PD-1 (Programmed cell death protein 1) blocker, and interleukin 15 (IL-15) to activate NK cells against AML. Materials and Methods: In this experimental study, the NK cells were isolated from mononuclear cells (MNCs) by using magnetic activation cell sorting (MACS) and were activated using the different combinations of Hsp70, PD-1 blocker, and IL-15 and then followed by immunophenotyping, functional assays to estimate their killing potential, and evaluation of expression pattern of PRF1, PIK3CB, PD-1, AKT-1, FAS-L, TRAIL, and GER A and B. Results: The expression of PD-1 was significantly (P<0.05) reduced after NK cell activation by the different formulas of IL-15, Hsp70, and PD-1 blocker. The expression of NKG2A in the treated NK cells was reduced particularly in the IL-15 (P<0.01) and IL-15+PD-1 blocker (P<0.05) groups. The addition of Hsp70 increased its expression. The cytotoxic effect of NK cells increased in all groups, especially in IL-15+PD-1 blocker besides increasing interferon-gamma (IFN-γ), Granzymes, and perforin expression (P<0.05). All IL-15+PD-1 blocker group changes were associated with the upregulation of PIK3CB and AKT-1 as key factors of NK cell activation. The presence of Hsp70 reduced IFN-γ releasing, and down-regulation of PIK3CB, AKT-1, Granzymes, and Perforin (P<0.05). Conclusion: We suggested the combination of IL-15 and PD-1 blocker could enhance the killing potential of AMLNK cells. Moreover, Hsp70 in combination with IL-15 and PD-1 blocker interferes activation of AML-NK cells through unknown mechanisms.

110-117

Authors: Leila Ahmadi; Nahid Eskandari; Mustafa Ghanadian; Mahshid Rahmati; Neda Kasiri; Masoud Etamadifar; Mohadeseh Toghyani; Fereshteh Alsahebfosoul

Objective: The function of Th17 cells in the neuroinflammatory process in multiple sclerosis (MS) has been previously clarified. It has been suggested that Quercetin can influence MS due to a variety of anti-inflammatory effects. The present study aimed to examine in vitro immunomodulatory aspects of Quercetin Penta Acetate as a modified compound on Th17 cells of MS patients and also to compare its effects with Quercetin. Materials and Methods: In this experimental study, peripheral blood mononuclear cell (PBMCs) were isolated and stained with CFSE then, half-maximal inhibitory concentration (IC50) values were determined using different doses and times for Quercetin Penta Acetate, and Methyl Prednisolone Acetate. Th17 cell proliferation was analyzed by flow cytometry and the expression levels of IL-17 and RORc genes were assessed by real-time polymerase chain reaction (PCR) method. Results: The results showed that IL-17A gene expression was inhibited by Quercetin Penta Acetate (P=0.0081), but Quercetin Penta Acetate did not have a significant inhibitory effect on Th17 cells proliferation (P= 0.59) and RORc gene expression (P=0.1), compared to Quercetin. Conclusion: Taken together, our results showed some immunomodulatory aspects of Quercetin Penta Acetate on Th17 cells are more effective than Quercetin and it could be considered in the treatment of MS.

126-134

Authors: Mohammad Danesh-Doust; Rasoul Irajirad; Fereshteh Vaziri Nezamdoust; Sara Khademi; Alireza Montazerabadi

Objective: Increasing research has been focused on the development of various nanocomplexes as targeted contrast media in diagnostic modalities, mainly in computed tomography (CT) scan imaging. Herein, we report a new method that uses Triptorelin [a luteinizing hormone-releasing hormone (LHRH) agonist]-targeted gold nanoparticles (AuNPs) via alginate for early detection of cancer by molecular CT imaging. Materials and Methods: In the experimental study, the formed multifunctional AuNPs coated with alginate conjugated with Triptorelin peptide (Triptorelin-Alginate-AuNPs) were synthesized and characterized via different techniques, including transmission electron microscopy (TEM), dynamic light scattering (DLS), and fourier transform infrared (FTIR) spectroscopy. The MTT assay was applied to calculate the toxicity of the NPs. Results: The results indicated that the formed Triptorelin-Alginate-AuNPs with an Au core size of ~18 nm are noncytotoxic at 127-, 254-, 381- and 508-mM concentrations and revealed significant improvement in the attenuation of X-rays intensity and contrast to noise ratio (CNR), compared with non-targeted cells at the highest energies (90, 120, 140 kVp). At 90 kVp, compared to non-targeted cells, targeted cells (Triptorelin-Alginate-AuNPs) enable 1.58, 1.69, 3.7 and 3.43 times greater contrast at a concentration of 127 mM, 254 mM, 381 mM, and 508 mM, respectively. Conclusion: These results suggest that the developed Triptorelin-Alginate-AuNPs may be considered an effective contrast agent for molecular CT imaging of gonadotropin-releasing hormone (GnRH) receptor-expressing cancer cells.

1-10

Authors: Marjan Asadi; Mohammad Ali Gholampour; Farzad Kompani; Shaban Alizadeh

Objective: Long non-coding RNA (lncRNA) H19 has essential roles in growth, migration, invasion, and metastasis of most cancers. H19 dysregulation is present in a large number of solid tumors and leukemia. However, the expression level of H19 in acute lymphoblastic leukemia (ALL) has not been elucidated yet. The current study aimed to explore H19 expression in ALL patients and cell lines. Materials and Methods: This experimental study was conducted in bone marrow (BM) samples collected from 25 patients with newly diagnosed ALL. In addition, we cultured the RPMI-8402, Jurkat, Ramos, and Daudi cell lines and assessed the effects of internal (hypoxia) and external (chemotherapy medications L-asparaginase [ASP] and vincristine [VCR]) factors on h19 expression. The expressions of H19, P53, c-Myc, HIF-1α and β-actin were performed using quantitative real-time polymerase chain reaction (qRT-PCR) method. Results: There was significantly increased H19 expression in the B-cell ALL (B-ALL, P<0.05), T-cell ALL (T-ALL, P<0.01) patients and the cell lines. This upregulation was governed by the P53, HIF-1α, and c-Myc transcription factors. We observed that increased c-Myc expression induced H19 expression; however, P53 adversely affected H19 expression. In addition, the results indicated that chemotherapy changed the gene expression pattern. There was a considerable decrease in H19 expression after exposure to chemotherapy medications; nonetheless, hypoxia induced H19 expression through P53 downregulation. Conclusion: Our findings suggest that H19 may have an important role in pathogenesis in ALL and may act as a promising and potential therapeutic target.

17-24

Authors: Shima Abbasihormozi; Azam Kouhkan; Abdolhossein Shaverdi; Mohammad Ali Sadighi Gilani; Vahab Babapour; Amir Niasari Naslji; Vahid Akbarinehad; AliReza Alizadeh

Objective: Although the role of obesity and diabetes mellitus (DM) in male infertility is well established, little information about the underlying cellular mechanisms in infertility is available. In this sense, nuclear factor kappa-B (NF-kB) has been recognized as an important regulator in obesity and DM; However, its function in the pathogenesis of male infertility has never been studied in obese or men who suffer from diabetes. Therefore, the main goal of current research is assessing NF-kB existence and activity in ejaculated human spermatozoa considering the obesity and diabetics condition of males. Materials and Methods: In an experimental study, the ELISA technique was applied to analyze NF-kB levels in sperm of four experimental groups: non-obese none-diabetic men (body mass index (BMI) <25 kg/m2; control group; n=30), obese non-diabetic men (BMI >30 kg/m2; OB group; n=30), non-obese diabetic men (BMI <25 kg/m2; DM group; n=30), and obese diabetic men (BMI >30 kg/m2; OB-DM group; n=30) who were presented to Royan Institute Infertility Center. In addition, protein localization was shown by Immunocytofluorescent assay. Sperm features were also evaluated using CASA. Results: The diabetic men were older than non-diabetic men regardless of obesity status (P=0.0002). Sperm progressive motility was affected by obesity (P=0.035) and type A sperm progressive motility was affected by DM (P=0.034). The concentration of sperm (P=0.013), motility (P=0.025) and morphology (P<0.0001) were altered by obesity × diabetes interaction effects. The NF-kB activity was negatively influenced by the main impact of diabetics (P=0.019). Obesity did not affect (P=0.248) NF-kB activity. Uniquely, NF-kB localized to the midpiece of sperm and post-acrosomal areas. Conclusion: The current study indicated a lower concentration of NF-kB in diabetic men, no effect of obesity on NF-kB was observed yet. Additionally, we revealed the main obesity and diabetes effects, and their interaction effect adversely influenced sperm characteristics.

35-44

Authors: Bahram Jambar Nooshin; Tahereh Tayebi; Amirhesam Babajani; Mohammad Mehdi Khani; Hassan Niknejad

Objective: Organ transplantation is the last therapeutic choice for end-stage liver failure, which is limited by the lack of sufficient donors. Decellularized liver can be used as a suitable matrix for liver tissue engineering with clinical application potential. Optimizing the decellularization procedure would obtain a biological matrix with completely removed cellular components and preserved 3-dimensional structure. This study aimed to evaluate the decellularization efficacy through three anatomical routes. Materials and Methods: In this experimental study, rat liver decellularization was performed through biliary duct (BD), portal vein (PV), and hepatic vein (HV); using chemical detergents and enzymes. The decellularization efficacy was evaluated by measurement of DNA content, extracellular matrix (ECM) total proteins, and glycosaminoglycans (GAGs). ECM preservation was examined by histological and immunohistochemical (IHC) staining and scanning electron microscopy (SEM). Scaffold biocompatibility was tested by the MTT assay for HepG2 and HUVEC cell lines. Results: Decellularization through HV and PV resulted in a transparent scaffold by complete cell removal, while the BD route produced an opaque scaffold with incomplete decellularization. H&E staining confirmed these results. Maximum DNA loss was obtained using 1% and 0.5% sodium dodecyl sulfate (SDS) in the PV and HV groups and the DNA content decreased faster in the HV group. At the final stages, the proteins excreted in the HV and PV groups were significantly less than the BD group. The GAGs level was diminished after decellularization, especially in the PV and HV groups. In the HV and PV groups the collagen amount was significantly more than the BD group. The IHC and SEM images showed that the ECM structure was preserved and cellular components were entirely removed. MTT assay showed the biocompatibility of the decellularized scaffold. Conclusion: The results revealed that the HV is a more suitable route for liver decellularization than the PV and BD.

51-61

Authors: Sara Momeni; Ahmad Shanei; Ameneh Sazgarnia; Neda Attaran; Seyed Amir Aledavood

Objective: The multimodality treatment of cancer provides a secure and effective approach to improve the outcome of treatments. Cold atmospheric plasma (CAP) has got attention because of selectively target and kills cancer cells. Likewise, gold nanoparticles (GNP) have been introduced as a radiosensitizer and drug delivery with high efficacy and low toxicity in cancer treatment. Conjugating GNP with indocyanine green (ICG) can develop a multifunctional drug to enhance radio and photosensitivity. The purpose of this study is to evaluate the anticancer effects of GNP@ICG in radiotherapy (RT) and CAP on DFW melanoma cancer and HFF fibroblast normal cell lines. Materials and Methods: In this experimental study, the cells were irradiated to RT and CAP, alone and in combination with or without GNP@ICG at various time sequences between RT and CAP. Apoptosis Annexin V/PI, MTT, and colony formation assays evaluated the therapeutic effect. Finally, the index of synergism was calculated to compare the results. Results: Most crucially, the cell viability assay showed that RT was less toxic to tumors and normal cells, but CAP showed a significant anti-tumor effect on melanoma cells with selective toxicity. In addition, cold plasma sensitized melanoma cells to radiotherapy so increasing treatment efficiency. This effect is enhanced with GNP@ICG. In comparison to RT alone, the data showed that combination treatment greatly decreased monolayer cell colonization and boosted apoptotic induction. Conclusion: The results provide new insights into the development of better approaches in radiotherapy of melanoma cells assisted plasma and nanomedicine.

73-75

Authors: Sepand Tehrani Fateh; Abbas Behgozin; Farshid Yekani; Lobat Geranpayeh; Asieh Olfatbakhsh; Shiva Moghaddam; Ramin Sarami Forroshani; Amir Salehi-Najafabadi; Faezeh Shekari

Considering HER2 as one of the well-known biomarkers in the cancer field, and published articles regarding serum levels of HER2, in this paper we tried to highlight the issue that most studies don’t stratify the HER-2 concentration of individuals in terms of gender. In this brief survey, healthy individuals with no prior non-communicable diseases were categorized as males (n=34) and females (n=43), and all samples were evaluated for plasma HER-2 levels at once. Surprisingly, the plasma level of HER-2 of healthy male individuals (mean= 2.28 ± 0.21 ng/mL) was significantly (P<0.0001) higher than the plasma level of HER-2 of healthy females (mean: 0.06 ± 0.09 ng/mL), with no overlap. Therefore, we suggest that more studies are required to re-check the cutoff values for HER-2 plasma levels based on gender since the clinical implications of a unique HER-2 cutoff for both genders may be seriously concerning.