With the development of molecular and cellar cardiology, gene therapy to cardiovascular disease has become the hot spot and the direction of study. Now, preclinical studies on ultrasound-mediated gene delivery (UMGD) in cardiovascular disease have achieved some success, but it is still hindered by a series of practical challenges for clinical translation. Even so, UMGD still holds the promise to cardiovascular disease in gene therapy for its non-invasiveness, accuracy, safety and ability to deliver multiple genes with repeated deliveries. In this review, we will focus on the basic principle, the current development, the future prospect and drawbacks of UMGD in the therapeutic applications of cardiovascular disease.
Objective To study the effect of immunogene therapy on the improvement of the outcome of malignant tumor. Methods Literatures about immunogene therapy of malignant tumor were collected and reviewed. Results The major methods of immunogene therapy of malignant tumor were as follows： ①Transference of cytokin genes into lymphocytes. ②Genetically engineered autologous tumor cells used as vaccine. ③Combined gene therapy.Conclusion The therapeutic effect of malignant tumor can be improved by immunogene therapy.
Selective recognition of double strands DNA (dsDNA) has been a research hot spot in molecular biology and biomedicine for a couple decades. Based on the selective interaction between natural nucleic acid/synthetic molecular ligands and dsDNA, gene diagnosis, gene therapy and gene editing would be realized. Hairpin oligopolyamide is a molecular ligand with excellent cellular permeability and nucleases-resistance which can target dsDNA sequence with high affinity and specificity at minor groove. This paper reviews the binding properties and biomedical applications of hairpin oligopolyamide targeting dsDNA, which provide references for further design and application of hairpin oligopolyamide.
ObjectiveTo construct tumor specific tubercle bacillus antigen Ag85A gene lentiviral vector driven by murine telomerase catalytic subunit promoter (PmTERT), paving the way for further research in tumor targeting immuno-gene therapy. MethodsPmTERT was amplified by PCR method, with murine genomic DNA as template. Then, transcriptional activities of PmTERT in various murine and human cell strains were studied by luciferase assay. Ag85A expression lentiviral vectors driven by cytomegalo virus (CMV) promoter and PmTERT respectively (pLVX-Ag85ACMV and pLVX-Ag85A-PmTERT) were constructed with nucleic acid cloning approach. And above recombinants were verified with DNA sequencing and Western blot. ResultsLucifease assay revealed that 331 bp PmTERT cloned in present research had transcriptional activity in murine Lewis lung cancer cells, human lung adenocarcinoma cells A549, and human esophageal cancer cells EC-109, while no transcriptional activity in murine fibroblasts NIH3T3 and human embryo fibroblasts MRC-5. Western blot revealed expression of Ag85A in pLVX-Ag85A-CMV transfected Lewis and NIH3T3 cells, pLVX-Ag85A-PmTERT transfected Lewis cells, no expression in pLVX-Ag85A-PmTERT transfected NIH3T3 cells. ConclusionPmTERT has tumor specific transcriptional activity. Ag85A gene can express selectively in tumor cells, driven by PmTERT.
ObjectiveTo investigate the effect of zinc finger protein A20 on lumbar intervertebral disc degeneration in rabbits.MethodsTwenty-six 3-month-old New Zealand rabbits, 2.0-2.5 kg in weight, were used to establish the model of intervertebral disc degeneration at L3, 4, L4, 5, and L5, 6 by transabdominal needle puncture. At 4 weeks after operation, the 24 rabbits were randomly divided into 4 groups after successful modeling, which checked by MRI. The target intervertebral discs of each group were injected with zinc finger protein A20 overexpressed adenovirus (Ov-A20 group), empty carrier adenovirus (NC group), phosphate buffer saline (control group), and shRNA-A20 adenovirus (Sh-A20 group). The biological responses of animals in each group were comprehensive scored before 1 day of injection and after 1, 2, 3, and 6 days of injection. At 2, 4, and 8 weeks after injection, the animals in each group were observed by MRI to obtain the exact T2 relaxation time (T2 signal value). After MRI examination, the animals were killed to take the degenerative intervertebral disc tissue; and the tissue was detected by Alcian blue staining to observed the intervertebral disc degeneration. The expressions of zinc finger protein A20, collagen Ⅱ, and aggrecan were detected by immunohistochemistry staining. The expressions of zinc finger protein A20, nuclear factor κB binding protein [P65, phosphate P65 (P-P65), collagen Ⅱ, aggrecan], inflammatory factors [tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β)], autophagy-related protein [LC3 (LC3Ⅱ/LC3Ⅰ) and P62] were detected by Western blot.ResultsThe comprehensive score of biological response in each group after injection was significantly lower than that before injection (P<0.05). At 6 days after injection, the comprehensive score of biological response in the Sh-A20 group was significantly lower than that in other groups (P<0.05), and there was no significant difference among other groups (P>0.05). The detection of MRI showed that the T2 signal value in the Ov-A20 group was the highest at 2, 4, and 8 weeks after injection (P<0.05), and the T2 signal value in the Sh-A20 group was the lowest at 2 and 4 weeks after injection (P<0.05). There was no significant difference between other groups (P>0.05). Alcian blue staining showed that the expression of aggrecan was the highest in Ov-A20 group and the lowest in Sh-A20 group at 4 weeks (P<0.05); the expression of aggrecan in Ov-A20 group was the highest at 8 weeks (P<0.05), and there was no significant difference between other groups (P>0.05). Immunohistochemical staining showed that the expressions of zinc finger protein A20, collagen Ⅱ, and aggrecan were the highest in Ov-A20 group and lowest in Sh-A20 group (P<0.05). Western blot showed that the expressions of zinc finger protein A20, collagen Ⅱ, aggrecan, and LC3 (LC3Ⅱ/LC3Ⅰ) proteins were the highest in the Ov-A20 group and the lowest in Sh-A20 group (P<0.05), while the expressions of P-P65, TNF-α, IL-1β, and P62 proteins were the lowest in Ov-A20 group and the highest in Sh-A20 group (P<0.05). There was no significant difference in the expression of p65 protein between groups (P>0.05).ConclusionZinc finger protein A20 can effectively regulate the process of lumbar intervertebral disc degeneration in rabbits by inhibiting inflammation.
Objective To summarize the research progress of microRNA (miRNA) and its non-viral vector in intervertebral disc degeneration (IDD) and to investigate the potential of non-viral vector delivery of miRNA in clinical application. Methods The related literature about the role of miRNA in IDD and its non-viral delivery system was reviewed and analyzed. Results MiRNA can regulate the related gene expression level and further participate in the pathophysiologic process in degenerated intervertebral disc, miRNA delivered by various non-viral vectors has obtained an ideal effect in some diseases. Conclusion MiRNA plays a great role in the cellular and molecular mechanisms of IDD, as a safe and effective strategy for gene therapy, non-viral vector provides new possibilities for IDD treated with miRNA.
Objective To investigate the effect and mechanism of calcitonin gene-related peptide (CGRP) on the prevention and treatment of transplant vein graft disease. Methods The 25 New Zealand white rabbits were divided into three groups: an experimental group [n=8, the rabbit jugular veins transfected with adeno-associated virus vector tipe 2/1 containing CGRP gene (AAV2/1-CGRP)], a carrier group [n=9, transfected with mosaic adeno-associated virus vector tipe 2/1 containing LacZ gene (AAV2/1-LacZ)] and a control group (n=8, saline) and then the cervical veins were implantated into the ipsilateral carotid artery by reverse end-side anastomosis. At 4 weeks after surgery, the pathology of the specimens, CD68 immunohistochemistry, in situ β-galactosidase staining were obtained. The expression of CGRP gene was detected by reverse transcription-polymerase chain reaction (RT-PCR). Monocyte chemoattractant protein-1(MCP-1), tumour necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS) and matrix metalloproteinase-9 (MMP-9) were detected by real-time polymerase chain reaction (real-time PCR). Results The CGRP and LacZ gene expression was positive at postoperative 4 weeks. The intima/media ratio was significantly inhibited in the experimental group. Macrophage infiltration and expression of inflammatory mediators including MCP-1, TNF-α, iNOS and MMP-9 were also significantly inhibited in the experimental group. Conclusion Transfection of AAV2/1-CGRP inhibits inflammatory mediator expression, macrophage infiltration and neointimal hyperplasia in experimental vein graft disease.
Objective To explore the tumor suppressive effect of superantigen staphylococcal enterotoxin B (SEB) for mice beared by Lewis lung cancer cell (LLCC). Methods SEB gene expressive plasmid PCDH-SEB-GFP was constructed and transfected into LLCC and it was detected by Western blot. Tumor-bearing mice model was established by subcutaneous injection of LLCC and SEB expressive plasmid PCDH-SEB-GFP was successfully injected into the tumor to observe its suppressive effect on the growth of Lewis lung cancer in mice. Results SEB expression was detected after transfection of LLCC with SEB gene plasmid PCDH-SEB-GFP. Intratumoral injection of plasmid PCDH-SEB-GFP could significantly inhibit the growth of Lewis lung cancer in tumor-bearing mice. Conclusions Intratumoral injection of superantigen SEB expressive plasmid can inhibit the growth of Lewis lung cancer in mice. It deserves a further study that SEB gene can work as an exogenous antigen gene in immune gene therapy for lung cancer.
Objective To summarize the research progress of gene-based therapeutic angiogenesis in lower limb ischemia, so as to provide a new method for non-invasive treatment of lower limb ischemia. Method The literatures on studies of gene-based therapeutic angiogenesis in lower limb ischemia in recent years were read and reviewed. Results The incidence of peripheral arterial disease had been increasing annually. How to effectively reduce the amputation rate and mortality rate of patients with critical limb ischemia was still a clinical problem that needs to be solved urgently. A large number of basic and clinical studies had shown that gene-based therapeutic angiogenesis could effectively induce angiogenesis and collateral circulation in ischemic tissue of lower limb, leading to the significant improvements of blood perfusion in ischemic areas. Additionally, the construction of many kinds of new non-viral gene delivery vectors could also improve the safety and effectiveness of gene therapy to a certain extent. Conclusion Although promising therapeutic effect of gene-based therapeutic angiogenesis brings new ideas and strategies for the treatment of lower limb ischemia, issues still exist that have not been solved.
Survivin-D53A (SVV-D53A) is a dominant-negative mutant survivin, which represents a potential promising target for cancer gene therapy. The present study was designed to determine whether SVV-D53A plasmid encapsuled by DOTAP: Chol liposome would have the anti-tumor activity against SPC-A1 lung adenocarcinoma, and to detect the possible mechanisms. In our experiment, SPC-A1 cells were transfected in vitro with SVV-D53A plasmid and examined for protein expression by Western blot, then flow cytometric analysis was used to detect apoptosis. SPC-A1 lung adenocarcinoma xenografts were established in vivo in the nude mice, which received the i.v. administrations of SVV-D53A plasmid/liposome complexes. After mice were sacrificed, the paraffin-embedded tumor tissue sections were used for proliferating cell nuclear antigen (PCNA) expression and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)assay. Compared with the control group, the mice treated with SVV-D53A plasmid had an obviously reduced tumor volume, with high level of apoptosis and decreased cell proliferation in tumor tissue. The research results proved that the administration of SVV-D53A plasmid resulted in significant inhibition of SPC-A1 cells both in vitro and in vivo. The functional mechanism is that the anti-tumor response causes and induces tumor cell apoptosis.