Growth Factor Gene Transfer Research Articles

VEGF-Encoding, Gene-Activated Collagen-Based Matrices Promote Blood Vessel Formation and Improved Wound Repair

Disruption in vascularization during wound repair can severely impair healing. Proangiogenic growth factor therapies have shown great healing potential; however, controlling growth factor activity and cellular behavior over desired healing time scales remains challenging. In this study, we evaluated collagen-mimetic peptide (CMP) tethers for their capacity to control growth factor gene transfer and growth factor activity using our recently developed gene-activated hyaluronic acid-collagen matrix (GAHCM). GAHCM was comprised of DNA/polyethyleneimine (PEI) polyplexes that were retained on hyaluronic acid (HA)-collagen hydrogels using CMPs. We hypothesized that using CMP-collagen tethers to control vascular endothelial growth factor-A (VEGF-A) gene delivery in fibroblasts would provide a powerful strategy to modulate the proangiogenic behaviors of endothelial cells (ECs) for blood vessel formation, resulting in enhanced wound repair. In co-culture experiments, we observed that CMP-modified GAHCM induced tunable gene delivery in fibroblasts as predicted, and correspondingly, VEGF-A produced by the fibroblasts led to increased growth and persistent migration of ECs for at least 7 days, as compared to non-CMP-modified GAHCM. Moreover, when ECs were exposed to fibroblast-containing VEGF-GAHCM with higher levels of CMP modification (50% CMP-PEI, or 50 CP), high CD31 expression was stimulated, resulting in the formation of an interconnected EC network with a significantly higher network volume and a larger diameter network structure than controls. Application of VEGF-GAHCM with 50 CP in murine splinted excisional wounds facilitated prolonged prohealing and proangiogenic responses resulting in increased blood vessel formation, improved granulation tissue formation, faster re-epithelialization, and overall enhanced repair. These findings suggest the benefits of CMP-collagen tethers as useful tools to control gene transfer and growth factor activity for improved treatment of wounds.

Modified hyaluronic acid-collagen matrices trigger efficient gene transfer and prohealing behavior in fibroblasts for improved wound repair

Growth factor therapy has demonstrated great promise for chronic wound repair, but controlling growth factor activity and cell phenotype over desired time frames remains a critical challenge. In this study, we developed a gene-activated hyaluronic acid-collagen matrix (GAHCM) comprising DNA/polyethylenimine (PEI) polyplexes retained on hyaluronic acid (HA)-collagen hydrogels using collagen mimetic peptides (CMPs). We hypothesized that manipulating both the number of CMP-collagen tethers and the ECM composition would provide a powerful strategy to control growth factor gene transfer kinetics while regulating cell behavior, resulting in enhanced growth factor activity for wound repair. We observed that polyplexes with 50% CMP-modified PEI (50 CP) showed enhanced retention of polyplexes in HCM hydrogels by 2.7-fold as compared to non-CMP modified polyplexes. Moreover, the incorporation of HA in the hydrogel promoted a significant increase in gene transfection efficiency based upon analysis of Gaussia luciferase (GLuc) reporter gene expression, and gene expression could be attenuated by blocking HA-CD44 signaling. Furthermore, when fibroblasts were exposed to vascular endothelial growth factor-A (VEGF-A)-GAHCM, the 50 CP matrix facilitated sustained VEGF-A production for up to 7 days, with maximal expression at day 5. Application of these VEGF-A-50 CP samples stimulated prolonged pro-healing responses, including the TGF-β1-induced myofibroblast-like phenotypes and enhanced closure of murine splinted wounds. Overall, these findings demonstrate the use of ECM-based materials to stimulate efficient gene transfer and regulate cellular phenotype, resulting in improved control of growth factor activity for wound repair. GAHCM has significant potential to overcome key challenges in growth factor therapy for regenerative medicine. Statement of significanceDespite great promise for growth factor therapies in wound treatment, controlling growth factor activity and providing a microenvironment for cells that maximizes growth factor signaling have continued to limit the success of existing formulations. Our GAHCM strategy, combining CMP gene delivery and a hyaluronic acid-collagen matrix, enabled enhanced wound healing efficacy via the combination of controlled and localized growth factor expression and matrix-mediated regulation of cell behavior. Incorporation of CMPs and HA in the same matrix synergistically enhanced VEGF activity as compared with simpler matrices. Accordingly, GAHCM will advance our ability to leverage growth factor signaling for wound healing, resulting in new long-term treatments for recalcitrant wounds.

Effects of nanoparticle-mediated growth factor gene transfer to the injured microenvironment on the tendon-to-bone healing strength.

The tendon-to-bone healing after trauma is usually slow and weak, and the repair site is easily disrupted during early mobilization exercise. bFGF and VEGFA gene therapy may hold promise in augmenting the tendon-to-bone healing process through enhancing cell proliferation and angiogenesis. This study is conducted to determine the effects of nanoparticle-mediated co-delivery of bFGF and VEGFA genes to the tendon-to-bone repair interface on the healing strength and biological responses in a chicken model. The PLGA nanoparticle/pEGFP-bFGF + pEGFP-VEGFA plasmid complexes were prepared and were characterized in vitro and in vivo. The nanoparticle/plasmid complexes can effectively transfer bFGF and VEGFA genes to the tendon-to-bone interface. Nanoparticle-mediated co-delivery of bFGF and VEGFA genes significantly improved the tendon-to-bone healing in terms of healing strengths and histology in a chicken flexor tendon repair model. Our results suggest a new biological approach to accelerate the tendon-to-bone healing.

Vascular endothelial growth factor gene transfer therapy for coronary artery disease: A systematic review and meta-analysis.

It is not clear whether treatment by vascular endothelial growth factor (VEGF) gene transfer can improve myocardial ischemia through a proangiogenesis mechanism and is effective against coronary artery disease (CAD). We aimed to perform a systematic review and meta-analysis of randomized controlled trials (RCTs) that compared VEGF gene therapy and standard treatments in CAD. We systematically searched the PubMed, Embase, and Cochrane databases and relevant references for RCTs (published up to May 2018; no language restrictions) and performed meta-analysis using both fixed and random effects models. Our primary outcome measures were mortality and serious cardiac events. The secondary outcome measures were follow-up left ventricular ejection fraction (LVEF), change in LVEF (ΔLVEF), and angina outcomes. The registration number is CRD42017058430. Of 524 identified studies, 14 were eligible and were included in our analysis. At a mean follow-up of 6months, VEGF gene therapy demonstrated a decreased risk of serious cardiac events (11.7% vs 21.2%, relative risk: 0.56; 95% confidence interval (CI): 0.37, 0.84; P=0.005) and a slight improvement in follow-up LVEF (weighted mean difference: 1.95; 95%CI: 1.28, 2.62). Furthermore, VEGF gene therapy using adenoviral vectors showed more potential benefit in terms of the risk of serious cardiac events, ΔLVEF, and Canadian Cardiovascular Society angina class. Nevertheless, mortality and angina frequency scores were not different. Vascular endothelial growth factor gene therapy appears to be safe and effective regarding serious cardiac events, with greater benefit when using adenoviral vectors. This meta-analysis highlights the need for further exploration in these areas.

Abstract WMP74: Therapeutic Effect of Human Dental Pulp Stem Cells Overexpressing Hepatocyte Growth Factor in Experimental Ischemic Stroke

Objective: Human dental pulp stem cell (DPSC) is an attractive cell source because they can be easily obtained as medical waste without ethical and logistical problems. Transplantation of DPSC decreases tissue injury in the rat brain and enhances motor function recovery after the middle cerebral artery occlusion (MCAO). We aimed increasing therapeutic effects combining DPSC and ex vivo human hepatocyte growth factor (HGF) gene transfer with adeno-associated virus (AAV) vector. Methods: Male Sprague-Dawley rats were subjected to transient 90 minute MCAO, followed by intravenous administration of 1 x 10 6 cells of DPSC, AAV-1/HGF transferred DPSC (DPSC/HGF) or vehicle immediately after reperfusion (each, n=8). Infarct volumes, neurological deficits and immunochemistry were assessed at 24 and 72 h after reperfusion. We also detected concentrations of TNF-α and IL-1β in the brain tissue extracts from the ischemic hemisphere by ELISA. Results: Infarct volumes at 24 h after reperfusion were diminished in both DPSC (155.7 ± 35.6 mm 3 , p < 0.01) and DPSC/HGF (110.2 ± 34.7 mm 3 , p < 0.01) transplantation compared with vehicle (213.1 ± 29.1 mm 3 ). DPSC/HGF transplantation showed more reduction of infarct volume compared with DPSC (110.2 ± 34.7 mm 3 vs 155.7 ± 35.6 mm 3 , p < 0.05). Motor function recovery was also observed at 24 and 72 h by both DPSC and DPSC/HGF transplantation. Furthermore, we determined that DPSC/HGF transplantation significantly suppressed expression of Iba-1 and TNF-α compared with DPSC in the cortical ischemic boundary zone (p < 0.05, p < 0.01). Also, DPSC/HGF markedly decreased brain tissue TNF-α (11.5 ± 4.2 pg/ml) and IL-1β (22.2 ± 4.2 pg/ml) concentrations compared with vehicle (TNF-α, 54.6 ± 9.8 pg/ml, p < 0.01; IL-1β, 95.2 ± 9.8 pg/ml, p < 0.01) and DPSC (TNF-α, 26.8 ± 11.2 pg/ml, p < 0.05; IL-1β, 40.6 ± 11.2 pg/ml, p < 0.05) in the ischemic hemisphere. Both DPSC and DPSC/HGF group improved neuronal degeneration compared with vehicle (p < 0.05, p < 0.01). Conclusions: These findings show that overexpression of HGF by AAV vector would enhance the neuroprotective effects of DPSC transplantation through the modulation of inflammation after acute ischemic stroke.

Cardiac Nerve Growth Factor Overexpression Induces Bone Marrow–derived Progenitor Cells Mobilization and Homing to the Infarcted Heart

Reparative response by bone marrow (BM)-derived progenitor cells (PCs) to ischemia is a multistep process that comprises the detachment from the BM endosteal niche through activation of osteoclasts and proteolytic enzymes (such as matrix metalloproteinases (MMPs)), mobilization to the circulation, and homing to the injured tissue. We previously showed that intramyocardial nerve growth factor gene transfer (NGF-GT) promotes cardiac repair following myocardial infarction (MI) in mice. Here, we investigate the impact of cardiac NGF-GT on postinfarction BM-derived PCs mobilization and homing at different time points after adenovirus-mediated NGF-GT in mice. Immunohistochemistry and flow cytometry newly illustrate the temporal profile of osteoclast and activation of MMP9, PCs expansion in the BM, and liberation/homing to the injured myocardium. NGF-GT amplified these responses and increased the BM levels of active osteoclasts and MMP9, which were not observed in MMP9-deficient mice. Taken together, our results suggest a novel role for NGF in BM-derived PCs mobilization/homing following MI.

Mutant MMP-9 and HGF gene transfer enhance resolution of CCl4-induced liver fibrosis in rats: role of ASH1 and EZH2 methyltransferases repression.

Hepatocyte growth factor (HGF) gene transfer inhibits liver fibrosis by regulating aberrant cellular functions, while mutant matrix metalloproteinase-9 (mMMP-9) enhances matrix degradation by neutralizing the elevated tissue inhibitor of metalloproteinase-1 (TIMP-1). It was shown that ASH1 and EZH2 methyltransferases are involved in development of liver fibrosis; however, their role in the resolution phase of liver fibrosis has not been investigated. This study evaluated the role of ASH1 and EZH2 in two mechanistically different therapeutic modalities, HGF and mMMP-9 gene transfer in CCl4 induced rat liver fibrosis. Liver fibrosis was induced in rats with twice a week intraperitoneal injection of CCl4 for 8 weeks. Adenovirus vectors encoding mMMP-9 or HGF genes were injected through tail vein at weeks six and seven and were sacrificed one week after the second injection. A healthy animal group was likewise injected with saline to serve as a negative control. Rats treated with mMMP-9 showed significantly lower fibrosis score, less Sirius red stained collagen area, reduced hydroxyproline and ALT concentration, decreased transforming growth factor beta 1 (TGF-β1) mRNA and lower labeling indices of α smooth muscle actin (α-SMA) and proliferating cell nuclear antigen (PCNA) stained cells compared with HGF- or saline-treated rats. Furthermore, TIMP-1 protein expression in mMMP-9 group was markedly reduced compared with all fibrotic groups. ASH1 and EZH2 protein expression was significantly elevated in fibrotic liver and significantly decreased in mMMP-9- and HGF-treated compared to saline-treated fibrotic livers with further reduction in the mMMP-9 group. Conclusion: Gene transfer of mMMP-9 and HGF reduced liver fibrosis in rats. ASH1 and EZH2 methyltransferases are significantly reduced in mMMP-9 and HGF treated rats which underlines the central role of these enzymes during fibrogenesis. Future studies should evaluate the role of selective pharmacologic inhibitors of ASH1 and EZH2 in resolution of liver fibrosis.

The effects of insulin-like growth factor-1 gene therapy and cell transplantation on rat acute wound model.

Background:Wound healing is a complex process. Different types of skin cells, extracellular matrix and variety of growth factors are involved in wound healing. The use of recombinant growth factors in researches and production of skin substitutes are still a challenge.Objectives:Much research has been done on the effects of gene therapy and cell therapy on wound healing. In this experimental study, the effect of insulin-like growth factor (IGF-1) gene transfer in fibroblast cells was assessed on acute dermal wound healing.Materials and Methods:Fibroblasts were cultured and transfected with IGF-1. Lipofectamine 2000 was used as a reagent of transfection. Transgene expression levels were measured by the enzyme linked immunosorbent assay (ELISA). To study in vivo, rats (weighing 170-200 g) were randomly divided into three groups (five/group) and full-thickness wounds were created on the dorsum region. Suspensions of transfected fibroblast cells were injected into the wound and were compared with wounds treated with native fibroblast cells and normal saline. For the microscopic examination, biopsy was performed on day seven.Results:In vitro, the maximum expression of IGF1 (96.95 pg/mL) in transfected fibroblast cells was 24 hours after gene transfer. In vivo, it was clear that IGF-1 gene therapy caused an increase in the number of keratinocyte cells during the wound healing process (mean of group A vs. group B with P value = 0.01, mean of group A vs. group C with P value = 0.000). Granulation of tissue formation in the transfected fibroblast group was more organized when compared with the normal saline group and native fibroblast cells.Conclusions:This study indicated that the optimization of gene transfer increases the expression of IGF-1. High concentrations of IGF-1, in combination with cell therapy, have a significant effect on wound healing.

GW25-e0833 Acidic fibroblast growth factor promotes the function of endothelial progenitor cells through Akt/FOXO3a signaling pathway

Endothelial progenitor cells (EPCs) contribute to angiogenesis and acidic fibroblast growth factor (FGF1) gene transfer enhances EPC function. The Forkhead box O transcription factors (FOXOs) play an important role in the regulation of various cellular processes and EPCs mainly express FOXO3a. Here

The effects of adenoviral transfection of the keratinocyte growth factor gene on epidermal stem cells: an in vitro study.

Epidermal stem cells (ESCs) are characterized as slowcycling, multi-potent, and self-renewing cells that not only maintain somatic homeostasis but also participate in tissue regeneration and repair. To examine the feasibility of adenoviral vector-mediated keratinocyte growth factor (KGF) gene transfer into in vitro-expanded ESCs, ESCs were isolated from samples of human skin, cultured in vitro, and then transfected with recombinant adenovirus (Ad) carrying the human KGF gene (AdKGF) or green fluorescent protein gene (AdGFP). The effects of KGF gene transfer on cell proliferation, cell cycle arrest, cell surface antigen phenotype, and β-catenin expression were investigated. Compared to ESCs transfected with AdGFP, AdKGFtransfected ESCs grew well, maintained a high proliferative capacity in keratinocyte serum-free medium, and expressed high levels of β-catenin. AdKGF infection increased the number of ESCs in the G0/G1 phase and promoted ESCs entry into the G2/M phase, but had no effect on cell surface antigen phenotype (CD49f(+)/CD71(-)). The results suggest that KGF gene transfer can stimulate ESCs to grow and undergo cell division, which can be applied to enhance cutaneous wound healing.

Protection against radiation-induced hematopoietic damage in bone marrow by hepatocyte growth factor gene transfer

Purpose: To investigate whether adenovirus-mediated delivery of the human hepatocyte growth factor (HGF) gene could prevent radiation-induced hematopoietic damage.Materials and methods: Thirty C57BL/6 mice were randomized into three groups, in which phosphate buffer saline (PBS), mock adenovirus vector (Ad-null) or adenovirus vector containing HGF (Ad-HGF) were injected into the tail vein of each group, respectively. After 48 hours, the mice received a single irradiation dose of 6.5 Gy 60Co gamma rays. Blood samples were extracted via the tail vein at day 0, 4, 7, 10, 14, 21, 24 and 30 after irradiation, for red blood cell (RBC) and white blood cell (WBC) and cluster of differentiation4 (CD4)/cluster of differentiation8 (CD8) ratio assessment. At weekly intervals following irradiation, serum erythropoietin (EPO), Interleukin-6 (IL-6) and Interferon-gamma (IFN-γ) levels were measured using enzyme-linked immunosorbent assay (ELISA). On post-irradiation day 30, the mice were autopsied and erythroid burst-forming units (BFU-E) were evaluated.Results: Adenovirus-mediated HGF gene transfer could increase human HGF level in serum and have a significant elevation in RBC and WBC count. Ad-HGF increased EPO and IL-6 levels and prompted BFU-E formation. Ad-HGF decreased radiation- induced micronucleus frequency in the mouse bone marrow (BM). Most evidence of radiation-induced hematopoietic damage was observed morphologically in bone marrow specimen four weeks after irradiation. Ad-HGF protected against radiation-induced BM failure and increased survival. Finally, Ad-HGF increased the thymic index and enhanced immune function in the irradiated C57BL/6 mice.Conclusions: This is the first report to date that demonstrates the potential of HGF gene transfer to prevent radiation-induced hematopoietic damage.

Hepatocyte growth factor gene-modified bone marrow-derived mesenchymal stem cells transplantation promotes angiogenesis in a rat model of hindlimb ischemia

Angiogenic gene therapy and cell-based therapy for peripheral arterial disease(PAD) have been studied intensively currently. This study aimed to investigate whether combining mesenchymal stem cells(MSCs) transplantation with ex vivo human hepatocyte growth factor(HGF) gene transfer was more therapeutically efficient than the MSCs therapy alone in a rat model of hindlimb ischemia. One week after establishing hindlimb ischemia models, Sprague-Dawley(SD) rats were randomized to receive HGF gene-modified MSCs transplantation(HGF-MSC group), untreated MSCs transplantation (MSC group), or PBS injection(PBS group), respectively. Three weeks after injection, angiogenesis was significantly induced by both MSCs and HGF-MSCs transplantation, and capillary density was the highest in the HGF-MSC group. The number of transplanted cell-derived endothelial cells was greater in HGF-MSC group than in MSC group after one week treatment. The expression of angiogenic cytokines such as HGF and VEGF in local ischemic muscles was more abundant in HGF-MSC group than in the other two groups. In vitro, the conditioned media obtained from HGF-MSCs cultures exerted proproliferative and promigratory effects on endothelial cells. It is concluded that HGF gene-modified MSCs transplantation therapy may induce more potent angiogenesis than the MSCs therapy alone. Engraftment of MSCs combined with angiogenic gene delivery may be a promising therapeutic strategy for the treatment of severe PAD.

GW24-e3919 Short-term safety and curative effect of recombinant adenovirus carrying hepatocyte growth factor gene on ischaemic cardiac disease

ObjectivesAdenovirus-mediated angiogenic growth factor gene transfer provides a new therapeutic strategy for treatment of coronary heart disease. Preclinical studies showed that direct intramyocardial injection of recombinant adenovirus with hepatocyte growth...

Unrestricted somatic stem cells as vehicle for nerve growth factor gene transfer

Objective: Nerve growth factor (NGF), a member of the neurotrophic factor family, plays a critical role in the maintenance and regeneration of different types of neurons. To overcome drastic challenges in the peripheral delivery of NGF, transplantation of NGF secreting stem cells to the target site of an injury may be an effective procedure. Unrestricted somatic stem cells (USSCs), a subtype of umbilical cord blood (UCB) stem cells, have shown promise for gene therapy purposes, and proper results have been observed from transplantation experiments in neurodegenerative disorders. Based on the considerable potential of USSCs for gene delivery applications, the goal of the current study was to establish a betaNGF gene containing USSCs, which is able to secrete functional recombinant betaNGF protein.Methods: Unrestricted somatic stem cells were isolated from UCB and were cultured in a DMEM medium. The betaNGF gene was cloned in the EFalpha-promoting lentiviral vector, and virus production was performed as a third generation lentivirus packaging system. Enzyme-linked immunosorbent assay, real-time PCR, co-culture, MTT, and immunocytochemistry assays were performed to evaluate the genetically engineered USSCs.Results: Overexpression of betaNGF gene in human USSCs created a USSC line that is able to secrete high amounts of functional betaNGF protein. betaNGF-secreting USSCs showed a high rate of viability along with acceptable immunological and morphological properties for transplantation into the nervous system.Discussion: Long-term expression of functional betaNGF, high viability of betaNGF producing USSCs, and expression of primary neuronal markers suggest that USSCs may be useful to deliver betaNGF into targeted sites of the nervous system in neurodegenerative disorders.

Growth factor transgenes interactively regulate articular chondrocytes

Adult articular chondrocytes lack an effective repair response to correct damage from injury or osteoarthritis. Polypeptide growth factors that stimulate articular chondrocyte proliferation and cartilage matrix synthesis may augment this response. Gene transfer is a promising approach to delivering such factors. Multiple growth factor genes regulate these cell functions, but multiple growth factor gene transfer remains unexplored. We tested the hypothesis that multiple growth factor gene transfer selectively modulates articular chondrocyte proliferation and matrix synthesis. We tested the hypothesis by delivering combinations of the transgenes encoding insulin-like growth factor I (IGF-I), fibroblast growth factor-2 (FGF-2), transforming growth factor beta1 (TGF-β1), bone morphogenetic protein-2 (BMP-2), and bone morphogenetic protien-7 (BMP-7) to articular chondrocytes and measured changes in the production of DNA, glycosaminoglycan, and collagen. The transgenes differentially regulated all these chondrocyte activities. In concert, the transgenes interacted to generate widely divergent responses from the cells. These interactions ranged from inhibitory to synergistic. The transgene pair encoding IGF-I and FGF-2 maximized cell proliferation. The three-transgene group encoding IGF-I, BMP-2, and BMP-7 maximized matrix production and also optimized the balance between cell proliferation and matrix production. These data demonstrate an approach to articular chondrocyte regulation that may be tailored to stimulate specific cell functions, and suggest that certain growth factor gene combinations have potential value for cell-based articular cartilage repair.

HEPATOCYTE GROWTH FACTOR GENETICALLY MODIFIED BONE MARROW-DERIVED MESENCHYMAL STEM CELLS TRANSPLANTATION PROMOTES ANGIOGENESIS IN A RAT MODEL OF HINDLIMB ISCHAEMIA

ObjectivesAngiogenic gene therapy and cell-based therapy for peripheral arterial disease (PAD) have been studied intensively currently. This study aimed to investigate a new strategy whether combining mesenchymal stem cells (MSCs)...

Pretreatment of Hepatocyte Growth Factor Gene Transfer Mediated by Octaarginine Peptide-Modified Nanoparticles Ameliorates LPS/D-Galactosamine-Induced Hepatitis

We previously reported that an octaarginine- and pH-sensitive fusogenic peptide-modified multifunctional envelope-type nano device (R8-GALA-MEND) produces a high level of gene expression in the liver. In this study, we report on an examination of whether this gene delivery system exerts potent hepatoprotective effects against lipopolysaccharide/D-galactosamine (LPS/D-GalN)-induced acute liver injury. In vivo-jetPEI(™)-Gal, a commercially available in vivo transfection reagent, was used as a reference. The systemic administration of the R8-GALA-MEND or in vivo-jetPEI(™)-Gal showed that the latter was more toxic than the R8-GALA-MEND, indicating that R8-GALA-MEND is a safer system than in vivo-jetPEI(™)-Gal. Pretreatment with R8-GALA-MEND or in vivo-jetPEI(™)-Gal loaded with hepatocyte growth factor (HGF) pDNA inhibited serum GPT and GOT levels from becoming elevated. However, the survival rate of the mice was significantly enhanced in the case of R8-GALA-MEND, but not for the in vivo-jetPEI(™)-Gal treatment. These results demonstrate that R8-GALA-MEND has the potential for use in the pretreatment of an acute liver injury.

Effect of adenovirus-mediated basic fibroblast growth factor gene transfer in vivo on oligodendrocyte cell numbers throughout ventrolateral white matter following spinal cord injury in rats.

To study the effect of adenovirus-mediated basic fibroblast growth factor(FGF-2) gene transfer in vivo on oligodendrocyte cell numbers throughout ventrolateral white matter following spinal cord injury in rats. Thirty-two adult female Sprague Dawley rats were injured with the Infinite Horizon Impactor, and then were randomly assigned to four groups: FGF-2-Adts high-titer group (1.27x10(7) pfu/rat), FGF-2-Adts intermediate-titre group (6.37x10(6) pfu/rat), FGF-2-Adts low-titer group (3.18 x 10(6) pfu/rat), and green fluorescent protein (GFP)-Adts group (5.9x10(7) pfu/rat). The transgenic expression in vivo was detected with fluorescence microscopy. The locomotor function of the hindlimbs of rats was evaluated using Rivlin plate. Slides mounted with tissue sections were processed for immunohistochemical detection and quantification of oligodendrocytes (CC1(+)) in the ventral lateral funiculi (VLF) of injured spinal cords. One week after spinal cord injury, GFP showed that many cells had expressed objective gene in vivo and the angles of the occlusal plane of rats in FGF-2 groups were significantly higher than in GFP-Adts group. Also, there was a significant difference among the FGF-2-Adts treatment groups for the volume of gray matter sparing. However, there were no significant differences for total white matter sparing. Stereological quantification of total CC1(+) cell numbers in the spared VLF showed a significant reduction in numbers with GFP controls compared to all other groups 4 weeks after injury. In contrast, the FGF-2 Adts intermediate-titer group had significantly more CC1(+) cells when compared to both the FGF-2-Adts high- and low-titer groups. Adenovirus-mediated FGF-2 gene transfer can promote the functional recovery of the injured spinal cord by enhancing the proliferation and/or differentiation of oligodendrocytes.

Emerging tools for erectile dysfunction: a role for regenerative medicine

Erectile dysfunction (ED) is the most common sexual disorder reported by men to their health-care providers and the most investigated male sexual dysfunction. Currently, the treatment of ED focuses on 'symptomatic relief' of ED and, therefore, tends to provide temporary relief rather than providing a cure or reversing the cause. The identification of a large population of "difficult-to-treat" patients has triggered researchers to identify novel treatment approaches, which focus on cure and restoration of the underlying cause of ED. Regenerative medicine has developed extensively in the past few decades and preclinical trials have emphasized the benefit of growth factor therapy, gene transfer, stem cells and tissue engineering for the restoration of erectile function. Development of clinical trials involving immunomodulation in postprostatectomy ED patients and the use of maxi-K channels for gene therapy are illustrative of the advances in the field. However, the search for novel treatment targets and a wealth of preclinical studies represent a dynamic and continuing field of enquiry.

Attenuation of hepatic fibrosis through ultrasound-microbubble-mediated HGF gene transfer in rats

ObjectiveThe objective was to explore the feasibility of ultrasound-microbubble-mediated hepatocyte growth factor (HGF) gene transfer for treating rat hepatic fibrosis induced by CCl4. MethodsForty-eight male SD rats were divided into ultrasound-microbubble-HGF group (U-M-HGF group), ultrasound-HGF group (U-HGF group), microbubble-HGF group (M-HGF group), HGF group (HGF group), CCl4 group (control group), and normal group. The serum levels of alanine transaminase (ALT), aspartate transaminase (AST), total protein, albumin (ALB), and globulin (GLB) and the ratio of ALB/GLB were determined after treatment. The degree of hepatic fibrosis was evaluated by histopathological numerical scores. The protein expressions of HGF, collagen I, collagen III, and α-smooth muscle antibody (α-SMA) were detected by immunohistochemistry. ResultsUltrasound-microbubble-mediated HGF therapy significantly reduced the serum level of ALT and AST to 59.88% and 49.18% of the control group, respectively. Ultrasound-microbubble-mediated HGF therapy prevented liver fibrosis, with an obvious decrease in fibrosis areas and extracellular matrix production of collagen I, collagen III, and α-SMA. The gene therapy could induce HGF delivery into the fibrotic liver effectively. ConclusionsUltrasound-microbubble-mediated HGF gene therapy can reduce liver fibrosis, which provides a novel strategy for gene therapy of chronic liver disease.