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PureFection Transfection Reagent

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[#LV750A-1] PureFection Transfection Reagent

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(1) The Distinct Role of Small Heat Shock Protein 20 on HCV NS3 Expression in HEK-293T Cell Line.[TOP]

Pubmed ID :30090208
Publication Date : //
Hepatitis C (HCV) is known as a serious blood-borne disease that infects millions of people globally. NS3 is a conserved non-structural sequence of hepatitis C virus which has a major role in activating specific CTL responses. As known, there is no effective vaccine against HCV infection, thus it is required to design a specific regimen of vaccination. Recently, the strong immunological properties of Heat shock proteins (Hsps) led to their use as immunomodulators and an antigen carrier for subunit vaccine candidates. In the current study, the role of Hsp20 was evaluated as a HCV NS3 gene carrier in mammalian cell line.

Authors : Basirnejad Marzieh, Bolhassani Azam, Sadat Seyed Mehdi,



(2) Enhanced Intercellular Delivery of cRGD-siRNA Conjugates by an Additional Oligospermine Modification.[TOP]

Pubmed ID :30087937
Publication Date : //
Small interfering RNA (siRNA), consisting a 21-mer duplex molecule, is often modified by conjugation with specific ligands to enhance its capacity for tissue-specific delivery. However, these attempts are hampered by the low permeability of negatively charged RNA molecules to enter the cell membrane. In this study, we designed and synthesized siRNA conjugates modified with cationic oligospermine and cyclic RGD (cRGD) to overcome the low-membrane permeability of siRNA. The siRNA conjugate, which contains 15 spermines and a cRGD peptide, showed sufficient gene-silencing activity at 250 nM final concentration without a transfection reagent. Under these conditions, the cationic oligospermine and cRGD-siRNA conjugate did not show any cytotoxicity.

Authors : Nakamoto Kosuke, Akao Yukihiro, Furuichi Yasuhiro, Ueno Yoshihito,



(3) Enhanced Th1/Th2 mixed immune responses elicited by polyethyleneimine adjuvanted influenza A (H7N9) antigen HA1-2 in chickens.[TOP]

Pubmed ID :30085299
Publication Date : //
Influenza A (H7N9) viruses have caused severe human infections and deaths every year in China since 2013. To reduce the risk of human infection and prevent a new influenza pandemic, there is a pressing need to develop safe and effective anti-H7N9 vaccines for poultry. Polyethyleneimine (PEI) is an organic polycation used extensively as a transfection reagent for decades. Although the adjuvant potential of PEI is well studied in mammals, its applicability and immune characteristics to avian species are still very rare. Here, to investigate the adjuvant activity of PEI, we analyzed the immune responses in chicken peripheral blood mononuclear cells in vitro. PEI significantly upregulated the expression of immune-related cytokines (IFN-γ, IL-2, IL-4, IL-6, IL-18, and IL-1β) and chemokines (CXCLi1, CXCLi2, MIP-1β, and MCP-3), suggesting that PEI promoted immune responses of avian cells. We also assessed the in vivo immune responses to PEI in a chicken model. After the second and third vaccinations, significantly higher IgG titers were observed in the chickens immunized with HA1-2+PEI than that of HA1-2 alone. The HA1-2+PEI group also increased percentages of CD4+ and CD8+ T cells and improved PBMC proliferation. The significantly upregulated IFN-γ and IL-4 levels of splenocytes from HA1-2+PEI vaccinated chickens further indicated that PEI promoted a Th1/Th2 mixed immune responses. This study not only demonstrates the adjuvant potential of PEI when co-administered with influenza H7N9 antigen HA1-2 in chickens, but also supports the use of PEI as a versatile systemic adjuvant platform in poultry.

Authors : Song Li, Xiong Dan, Hu Maozhi, Jiao Xinan, Pan Zhiming,



(4) Transfection reagent Lipofectamine triggers type I interferon signaling activation in macrophages.[TOP]

Pubmed ID :30084169
Publication Date : //
The commercial transfection reagent Lipofectamine has been widely used for cytoplasmic delivery of nucleic acids and for cytosolic engagement with intracellular innate immune sensors to trigger type I interferon (IFN) production. However, the effect of Lipofectamine alone on type I IFN response has not been studied in detail. Here, we show that Lipofectamine induced type I IFN signaling in both RAW 264.7 macrophage-like cells and primary bone marrow-derived macrophages. Type I IFN induction was dependent on IRF3 and IRF7 and partially required the TIR-domain-containing adapter-inducing interferon-β. In contrast, the transfection reagent Xfect did not activate type I IFN signaling. Our study highlights the potential confounding experimental interpretation when using Lipofectamine-based transfection for delivering intracellular ligands and provides important insights into lipid signaling in innate immune response. This article is protected by copyright. All rights reserved.

Authors : Guo Xiaomin, Wang Huan, Li Yang, Leng Xiaopeng, Huang Weiwei, Ma Yanbing, Xu Tao, Qi Xiaopeng,



(5) Restoration of miR-152 expression suppresses cell proliferation, survival, and migration through inhibition of AKT-ERK pathway in colorectal cancer.[TOP]

Pubmed ID :30076720
Publication Date : //
MiR-152 has been reported as a tumor suppressor microRNA that is downregulated in a number of cancers, including colorectal cancer (CRC). A recent study suggested that miR-152 could be one of the key regulators of CRC. The aim of this study is to investigate the role of miR-152 in CRC and its mechanisms.

Authors : Ghazanchaei Ardavan, Mansoori Behzad, Mohammadi Ali, Biglari Alireza, Baradaran Behzad,



(6) Antibody-mediated oral delivery of therapeutic DNA for type 2 diabetes mellitus.[TOP]

Pubmed ID :30065848
Publication Date : //
Diabetes mellitus (DM) is a chronic progressive metabolic disease that involves uncontrolled elevation of blood glucose levels. Among various therapeutic approaches, GLP-1 prevents type 2 diabetes mellitus (T2DM) patients from experiencing hyperglycemic episodes. However, the short half-life (< 5 min) and rapid clearance of GLP-1 often limits its therapeutic use. Here, we developed an oral GLP-1 gene delivery system to achieve an extended antidiabetic effect.

Authors : Cha Seungbin, Lee Sun Hwa, Kang Sung Hun, Hasan Mohammad Nazmul, Kim Young Jun, Cho Sungpil, Lee Yong-Kyu,



(7) BMP-2 gene transfection of bone marrow stromal cells to induce osteoblastic differentiation in a rat calvarial defect model.[TOP]

Pubmed ID :30033316
Publication Date : //
Gene therapy for bone tissue engineering has been widely developed. Recently, non-viral DNA-based gene therapy has been reported to be a safer and more efficient method of delivering DNA into target cells. We used a non-viral gene transfection reagent to delivery bone morphogenetic protein-2 (BMP-2) gene into bone marrow stromal cells (BMSCs). Primary BMSCs were isolated from rat femurs and transfected with BMP-2 plasmids. The transfection rate was analyzed using flow cytometry. The concentration of BMP-2 protein was quantified using an enzyme-linked immunosorbent assay. Levels of osteopontin and osteocalcin were measured to evaluate osteogenic differentiation. In vivo, we designed a critical-size calvarial defect rat model to study new bone regeneration, using Matrigel as a scaffold to carry BMP-2-transfected bone marrow stromal cells into the defect site. New bone formation was assessed by micro-computed tomography, X-ray, immunohistochemical staining and histomophometry. The transfection rate after 72 h was 31.5%. The BMP-2 protein level as well as osteopontin and osteocalcin expressions were higher in the experimental group (transfected with BMP-2) than the control group (transfected with green fluorescent protein, GFP). The in vivo study suggested that bone healing occurred 12 weeks after scaffold implantation. In addition, BMP-2-transfected bone marrow stromal cells provided better osteogenic differentiation than primary bone marrow stromal cells. Our findings suggest that non-viral gene therapy may be useful in bone tissue engineering.

Authors : Hsieh Ming-Kai, Wu Chia-Jung, Chen Chun-Chieh, Tsai Tsung-Ting, Niu Chi-Chien, Wu Shinn-Chih, Lai Po-Liang,



(8) Simple in Vivo Gene Editing via Direct Self-Assembly of Cas9 Ribonucleoprotein Complexes for Cancer Treatment.[TOP]

Pubmed ID :30028587
Publication Date : //
Cas9 ribonucleoprotein (RNP)-mediated delivery has emerged as an ideal approach for in vivo applications. However, the delivery of Cas9 RNPs requires electroporation or lipid- or cationic-reagent-mediated transfection. Here, we developed a carrier-free Cas9 RNP delivery system for robust gene editing in vivo. For simultaneous delivery of Cas9 and a guide RNA into target cells without the aid of any transfection reagents, we established a multifunctional Cas9 fusion protein (Cas9-LMWP) that forms a ternary complex with synthetic crRNA:tracrRNA hybrids in a simple procedure. Cas9-LMWP carrying both a nuclear localization sequence and a low-molecular-weight protamine (LMWP) enables the direct self-assembly of a Cas9:crRNA:tracrRNA ternary complex (a ternary Cas9 RNP) and allows for the delivery of the ternary Cas9 RNPs into the recipient cells, owing to its intrinsic cellular and nuclear translocation ability with low immunogenicity. To demonstrate the potential of this system, we showed extensive synergistic anti-KRAS therapy (CI value: 0.34) via in vitro and in vivo editing of the KRAS gene by the direct delivery of multifunctional Cas9 RNPs in lung cancer. Thus, our carrier-free Cas9 RNP delivery system could be an innovative platform that might serve as an alternative to conventional transfection reagents for simple gene editing and high-throughput genetic screening.

Authors : Kim Seung Min, Shin Sang Chul, Kim Eunice EunKyeong, Kim Sang-Heon, Park Kwideok, Oh Seung Ja, Jang Mihue,



(9) Polysaccharide-modified nanoparticles with intelligent CD44 receptor targeting ability for gene delivery.[TOP]

Pubmed ID :30022822
Publication Date : //
Hyaluronic acid (HA) and chondroitin sulfate (CD) are endogenous polysaccharides. In recent years, they have aroused the interest of scientists because of specific binding to CD44 receptors, which are overexpressed in several types of tumors.

Authors : Lin Wen Jen, Lee Wei Chi,



(10) Influence of YKL-40 gene RNA interference on the biological behaviors of endometrial cancer HEC-1A cells.[TOP]

Pubmed ID :30008865
Publication Date : //
The present study aimed to investigate the effects of chitinase-3-like protein 1 (YKL-40) gene RNA interference on the biological behaviors and enhanced chemosensitivity of endometrial cancer (EC) HEC-1A cells. YKL-40 small interfering (si)RNA was transduced into EC HEC-1A cells using a lentivirus. The experiment was divided into three groups: The experimental group was transfected with YKL-40 siRNA (si-YKL-40); the mock-treatment group was transfected with transfection reagent only; and the blank control group was left untreated. A reverse transcription-quantitative polymerase chain reaction was performed to investigate the mRNA expression levels of YKL-40. The biological behaviors, including cell proliferation, migration, invasion and apoptosis, were detected by MTT and Transwell assays, and flow cytometry (FCM) analysis, respectively. The results of the present study demonstrated that the mRNA expression levels of YKL-40 were downregulated within HEC-1A cells upon transfection with si-YKL-40 (P<0.05). The proliferative, migratory and invasive abilities of HEC-1A cells were inhibited by si-YKL-40 (P<0.05). The mRNA expression levels of YKL-40 were upregulated within HEC-1A cells following treatment with cisplatin (P<0.05). FCM analysis revealed that the average cellular apoptosis rate increased following the inhibition of YKL-40 gene expression via siRNA (P<0.05). Therefore, the YKL-40 gene may be associated with the proliferative, migratory, invasive and anti-apoptotic ability of HEC-1A cells. YKL-40 downregulation may enhance the sensitivity of human EC HEC-1A cells to chemotherapy.

Authors : Li Lili, Fan Jiangtao, Li Dahai, Liu Yan, Shrestha Poonam, Zhong Chunyan, Xia Xiuhong, Huang Xiaobing,