Current Gene Therapy - Volume 14, Issue 2, 2014

Histidine-proline-rich glycoprotein (HPRG) is a plasma protein of vertebrates, which has potent antiangiogenic and tumor vessel normalization properties. Attenuated Salmonella Typhimurium strain VNP20009 preferentially accumulates and replicates in hypoxic tumor regions. In this study, we engineered VNP20009 to express HPRG under the control of a hypoxia-induced NirB promoter and evaluated the efficacy of the VNP20009-mediated targeted expressionof HPRG (VNP-pNHPRG) on tumor growth in primary and metastatic tumor models. When VNP-pNHPRG was administered to melanoma tumor mice by intraperitoneal injection, the NirB promoter controlled HPRG expression in tumor, which inhibited tumor vessel density and areas as well as regulated vascular normalization. VNP-pNHPRG significantly delayed tumor growth and enhanced survival time in primary B16F10 mice model and markedly suppressed lung metastatic tumor growth and prolonged survival time in B16F10 metastatic tumor models. Furthermore, VNP-pNHPRG down-regulated the HIF-1α-VEGF/Ang-2 signal pathway by altering the hypoxic tumor microenvironment. These results showed that VNP20009-mediated targeted expression of HPRG provides a novel cancer gene therapeutic approach for the treatment of primary and metastatic cancer.

Adeno-associated virus (AAV) based vectors have emerged as important tools for gene therapy in humans. The recent successes seen in Phase I/II clinical trials have also highlighted the issues related to the host and vector-related immune response that preclude the universal application of this promising vector system. A fundamental insight into the biological mechanisms by which AAV infects the host cell and a thorough understanding of the immediate and long-lived cellular responses to AAV infection is likely to offer clues and help design better intervention strategies to improve the therapeutic efficiency of AAV vectors. This article reviews the biology of AAV-host cellular interactions and outlines their application in the development of novel and improved AAV vector systems.

Hearing impairment is primarily attributed to inner ear hair cell (HC) defects that subsequently lead to spiral ganglion neuron (SGN) loss. The HC loss cannot be self-repaired because of the HCs’ limited capacity to regenerate in mammals. Atoh1, also known as Math1, Hath1, and Cath1, is a proneural basic helix–loop–helix (bHLH) transcription factor that played a major role in HC differentiation. Atoh1 activity at various developmental stages can sufficiently drive HC differentiation in the cochlea. Recent issues of a certain publication have identified that Atoh1 is essential for inner ear development, such as cell growth, morphogenesis, differentiation, cellular maintenance, and survival. We summarize the new findings in Atoh1 research and identify the mechanisms underlying the role of Atoh1 in HC regeneration to launch the future of Atoh1 therapy.

For years researchers have exerted every effort to improve the influential roles of microRNA (miRNA) in regulating genes that direct mammalian cell development and function. In spite of numerous advancements, many facets of miRNA generation remain unresolved due to the perplexing regulatory networks. The biogenesis of miRNA, eminently endures as a mystery as no universal pathway defines or explicates the variegation in the rise of miRNAs. Early evidence in biogenesis ignited specific steps of being omitted or replaced that eventuate in the individual miRNAs of different mechanisms. Understanding the basic foundation concerning how miRNAs are generated and function will help with diagnostic tools and therapeutic strategies. This review encompasses the canonical and the non-canonical pathways involved in miRNA biogenesis, while elucidating how miRNAs regulate genes at the nuclear level and also the mechanism that lies behind circulating miRNAs.

Gene therapy is a promising approach for treating a wide range of human pathologies such as genetic disorders as well as diseases acquired over time. Viral and non-viral vectors are used to convey sequences of genes that can be expressed for therapeutic purposes. Plasmid DNA is receiving considerable attention for intramuscular gene transfer due to its safety, simplicity and low cost of production. Nevertheless, strategies to improve DNA uptake into the nucleus of cells for its expression are required. Cytoskeleton plays an important role in the intracellular trafficking. The mechanism regulating this process must be elucidated. Here, we propose a new methodological approach based on the coupling of biology assays and predictive mathematical models, in order to clarify the mechanism of the DNA uptake and its expression into the cells. Once these processes are better clarified, we will be able to propose more efficient therapeutic gene transfer protocols for the treatment of human patients.

Endothelial progenitor cells (EPCs) and angiopoietin-1 (Ang-1) play important roles in vasculogenesis and angiogenesis, respectively. Thus, targeting both aspects of cardiovascular tissue regeneration may offer promising therapeutic options for cardiovascular disorders. To this end, we constructed a lentiviral vector (pNL) with the Ang-1 gene and transfected EPCs with it (Ang-1-EPCs) to investigate vasculogenesis in both cellular and animal models. Compared to controls, intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) increased significantly in both untreated EPCs and in the pNL vector group. After Ang-1 transcription, ICAM-1 and VCAM-1 decreased considerably in those treatment groups. Ang-1-modified EPCs alleviated inflammatory responses induced by tumornecrosis factor-α (TNF-α) in vitro. Moreover, Ang-1-EPC implantation inhibited neointimal hyperplasia after balloon catheter injury in rats, dramatically diminishing the intimal-media (I/M) ratio and decreasing the neointimal area. Proliferating cell nuclear antigen expression in the Ang-1-EPC group was lower than the EPC non-treatment group as well, suggesting that Ang-1-EPC improved cell survival during inflammation and promoted endothelialization in damaged blood vessels.

Purpose: Magnetic resonance imaging (MRI) has been used to track magnetically labeled human bone marrowderived mesenchymal stem cells (hBMSCs) in vivo after COX-2 silencing and transplantation into nude rats via tail vein injection. Methods: In the present study, we knocked down COX-2 expression in hBMSCs through lentivirus transduction. The COX-2 knockdown was confirmed by real-time PCR and Western blotting analyses. Subsequently, we labeled cells with the novel reagent SPIO-Molday ION Rhodamine-B™ (MIRB). The viability, proliferation and differentiation of these cells were assessed in vitro. Labeled lenti-shCOX2 hBMSCs, unlabeled hBMSCs and phosphate-buffered saline (PBS) were individually injected into the tail veins of nude rat models, forming three treatment groups. All nude rats underwent GRE T2*-weighted MRI at 1 h, 7 days and 14 days post-injection. After MRI examination, the animals were sacrificed, and the brain and liver were examined by fluorescence microscopy and Prussian Blue staining. Results: Our results confirmed the successful down-regulation of COX-2 at the mRNA and protein levels in hBMSCs by lentivirus transduction. The viability and differentiation of hBMSCs were not affected by MIRB labeling. After 7 days, hypointense signal void areas in the rat livers were observed on MRI. After 14 days, iron particles were detected in the blood vessels, sinusoids, interlobular septum and capsule tissues of the liver. Conclusion: The MIRB-labeled lenti-shCOX2 hBMSCs transplanted into nude rat models via tail vein injection can be detected and monitored in vivo using 3.0 T clinical MRI for up to 14 days after cell transplantation.