Delivery Of Insulin-like Growth Factor-1 Research Articles

Delivery of AAV-IGF-1 to the CNS Extends Survival in ALS Mice Through Modification of Aberrant Glial Cell Activity

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease of the motor system. Recent work in rodent models of ALS has shown that insulin-like growth factor-1 (IGF-1) slows disease progression when delivered at disease onset. However, IGF-1's mechanism of action along the neuromuscular axis remains unclear. In this study, symptomatic ALS mice received IGF-1 through stereotaxic injection of an IGF-1-expressing viral vector to the deep cerebellar nuclei (DCN), a region of the cerebellum with extensive brain stem and spinal cord connections. We found that delivery of IGF-1 to the central nervous system (CNS) reduced ALS neuropathology, improved muscle strength, and significantly extended life span in ALS mice. To explore the mechanism of action of IGF-1, we used a newly developed in vitro model of ALS. We demonstrate that IGF-1 is potently neuroprotective and attenuates glial cell-mediated release of tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO). Our results show that delivering IGF-1 to the CNS is sufficient to delay disease progression in a mouse model of familial ALS and demonstrate for the first time that IGF-1 attenuates the pathological activity of non-neuronal cells that contribute to disease progression. Our findings highlight an innovative approach for delivering IGF-1 to the CNS.

Engineering insulin‐like growth factor‐1 for local delivery

Insulin-like growth factor-1 (IGF-1) is a small protein that promotes cell survival and growth, often acting over long distances. Although for decades IGF-1 has been considered to have therapeutic potential, systemic side effects of IGF-1 are significant, and local delivery of IGF-1 for tissue repair has been a long-standing challenge. In this study, we designed and purified a novel protein, heparin-binding IGF-1 (Xp-HB-IGF-1), which is a fusion protein of native IGF-1 with the heparin-binding domain of heparin-binding epidermal growth factor-like growth factor. Xp-HB-IGF-1 bound selectively to heparin as well as the cell surfaces of 3T3 fibroblasts, neonatal cardiac myocytes and differentiating ES cells. Xp-HB-IGF-1 activated the IGF-1 receptor and Akt with identical kinetics and dose response, indicating no compromise of biological activity due to the heparin-binding domain. Because cartilage is a proteoglycan-rich environment and IGF-1 is a known stimulus for chondrocyte biosynthesis, we then studied the effectiveness of Xp-HB-IGF-1 in cartilage. Xp-HB-IGF-1 was selectively retained by cartilage explants and led to sustained chondrocyte proteoglycan biosynthesis compared to IGF-1. These data show that the strategy of engineering a "long-distance" growth factor like IGF-1 for local delivery may be useful for tissue repair and minimizing systemic effects.

IGF-1 exacerbates the neurotoxicity of the mitochondrial inhibitor 3NP in rats

Insulin-like Growth Factor 1 (IGF-1) has broad-range neuroprotective effects and is a therapeutic candidate for Huntington's disease (HD). IGF-1 protects striatal neurons from the toxicity of mutated huntingtin in vitro and improves neuronal survival in vivo in a phenotypic model of HD involving excitotoxic cell death. Because HD is a multifactorial disease, it is important to evaluate the neuroprotective role of IGF-1 in other pathological situations involved in HD progression. We have evaluated the neuroprotective effects of IGF-1 in vivo, using the 3-nitropropionic acid (3NP) rat model which replicates the mitochondrial dysfunction observed in HD. Continuous intracerebroventricular infusion of recombinant IGF-1 at a low dose (0.025 microg/h for 5 days) did not alleviate motor impairment and weight loss induced by 3NP treatment. In addition, histological evaluation and quantification of DNA fragmentation evidenced no improvement in neuronal survival. Of interest, we found that a higher concentration of IGF-1 (0.25 microg/h) resulted in an exacerbation of 3NP toxicity on striatal neurons. These results suggest that intracerebral delivery of IGF-1 may not provide a fully effective therapeutic strategy for HD or other disorders involving mitochondrial impairment.

Degradable hydrogel scaffolds for in vivo delivery of single and dual growth factors in cartilage repair

As our population ages, treatment for joint pain associated with articular cartilage damage is becoming a prevalent challenge. Accordingly, this work investigates local delivery of two regulatory proteins - transforming growth factor-beta1 (TGF-beta1) and insulin-like growth factor-1 (IGF-1) - to cartilage defects from degradable scaffolds as a potential strategy for improving cartilage repair. The effects of TGF-beta1 and/or IGF-1 delivery on osteochondral repair in adult rabbits were examined through histomorphometric analysis of 11 markers of osteochondral repair. Complete scaffold degradation occurred allowing for assessment of the healing response at 12 weeks post-surgery. When compared to untreated defects, higher scores were observed with IGF-1-treated defects for the six markers of neo-surface repair: neo-surface morphology, cartilage thickness, surface regularity, chondrocyte clustering, and the chondrocyte/glycosaminoglycan content of the neo-surface and the cartilage surrounding the defect. Surprisingly, the benefits of IGF-1 delivery were not maintained when this growth factor (GF) was co-delivered with TGF-beta1, despite numerous in vitro reports of the combinatory actions of these GFs. While localized delivery of IGF-1 may be a promising repair strategy, further in vivo assessment is necessary, since fibrous tissue was commonly observed in the neo-surface of all treatment groups. More importantly, this study highlights the need to rigorously examine GF interactions in the wound healing environment and demonstrates that in vitro observations do not directly translate to the in vivo setting.

Local myocardial insulin-like growth factor 1 (IGF-1) delivery with biotinylated peptide nanofibers improves cell therapy for myocardial infarction

Strategies for cardiac repair include injection of cells, but these approaches have been hampered by poor cell engraftment, survival, and differentiation. To address these shortcomings for the purpose of improving cardiac function after injury, we designed self-assembling peptide nanofibers for prolonged delivery of insulin-like growth factor 1 (IGF-1), a cardiomyocyte growth and differentiation factor, to the myocardium, using a "biotin sandwich" approach. Biotinylated IGF-1 was complexed with tetravalent streptavidin and then bound to biotinylated self-assembling peptides. This biotin sandwich strategy allowed binding of IGF-1 but did not prevent self-assembly of the peptides into nanofibers within the myocardium. IGF-1 that was bound to peptide nanofibers activated Akt, decreased activation of caspase-3, and increased expression of cardiac troponin I in cardiomyocytes. After injection into rat myocardium, biotinylated nanofibers provided sustained IGF-1 delivery for 28 days, and targeted delivery of IGF-1 in vivo increased activation of Akt in the myocardium. When combined with transplanted cardiomyocytes, IGF-1 delivery by biotinylated nanofibers decreased caspase-3 cleavage by 28% and increased the myocyte cross-sectional area by 25% compared with cells embedded within nanofibers alone or with untethered IGF-1. Finally, cell therapy with IGF-1 delivery by biotinylated nanofibers improved systolic function after experimental myocardial infarction, demonstrating how engineering the local cellular microenvironment can improve cell therapy.

IGF-I increases bone marrow contribution to adult skeletal muscle and enhances the fusion of myelomonocytic precursors

Muscle damage has been shown to enhance the contribution of bone marrow–derived cells (BMDCs) to regenerating skeletal muscle. One responsible cell type involved in this process is a hematopoietic stem cell derivative, the myelomonocytic precursor (MMC). However, the molecular components responsible for this injury-related response remain largely unknown. In this paper, we show that delivery of insulin-like growth factor I (IGF-I) to adult skeletal muscle by three different methods—plasmid electroporation, injection of genetically engineered myoblasts, and recombinant protein injection—increases the integration of BMDCs up to fourfold. To investigate the underlying mechanism, we developed an in vitro fusion assay in which co-cultures of MMCs and myotubes were exposed to IGF-I. The number of fusion events was substantially augmented by IGF-I, independent of its effect on cell survival. These results provide novel evidence that a single factor, IGF-I, is sufficient to enhance the fusion of bone marrow derivatives with adult skeletal muscle.

Targeted expression of IGF-1 in the central nervous system fails to protect mice from experimental autoimmune encephalomyelitis

Insulin-like growth factor 1 (IGF-1) has been identified as a critical molecule in the induction of myelination in the central nervous system (CNS). Systemic injection of IGF-1 has been shown to have a varied and transiently protective effect on the clinical course of experimental autoimmune encephalomyelitis (EAE). Since systemic IGF-1 can also modulate peripheral immune lymphocytes, we examined whether a sustained and local delivery of IGF-1 into the spinal cord would have any influence on the chronic course of EAE in C57/BL6 mice. The capability of adeno-associated virus (AAV) to be retrogradely transported efficiently from muscle to motor neurons of the spinal cord was used to overcome the difficulty routinely encountered when attempting chronic delivery of molecules into the CNS. We demonstrate that AAV-mediated delivery of IGF-1 in CNS did not have any beneficial effect on the clinical course of EAE. Injection of AAV-IGF1 after induction of the disease worsened the clinical symptoms. Furthermore, CNS expression of IGF-1 did not affect the pathogenic anti-MOG T cell response, as examined by proliferation and cytokine secretion. Thus, enhanced expression of IGF-1 in the CNS during inflammation does not have a significant effect on myelination. These data have important implications for the potential use of IGF-1 in the treatment of multiple sclerosis.

Dual growth factor delivery from degradable oligo(poly(ethylene glycol) fumarate) hydrogel scaffolds for cartilage tissue engineering

This work describes the development of a non-invasive means of simultaneously delivering insulin-like growth factor-1 (IGF-1) and transforming growth factor-β1 (TGF-β1) to injured cartilage tissue in a controlled manner. This novel delivery technology employs the water-soluble polymer, oligo(poly(ethylene glycol) fumarate) (OPF), in the fabrication of biodegradable hydrogels which encapsulate gelatin microparticles. Release studies first examined the effect of gelatin isoelectric point (IEP) and crosslinking extent on IGF-1 release from these microparticles. In the presence of collagenase, highly crosslinked, acidic gelatin (IEP=5.0) provided sustained release of IGF-1, 95.2±2.9% cumulative release at day 28, while less crosslinked microparticles and microparticles of alternate IEP exhibited similar release values after only 6 days. Encapsulation of these highly crosslinked microparticles in a network of OPF provided a means to further control release, reducing final cumulative release to 70.2±4.7% in collagenase-containing PBS. Final release values from OPF–gelatin microparticle composites could be altered by incorporating less crosslinked, non-loaded microparticles within these constructs. Finally, this technology was extended to the dual delivery of IGF-1 and TGF-β1 by loading these growth factors into either the OPF hydrogel phase or gelatin microparticle phase of composites. Release profiles were successfully manipulated by altering the phase of growth factor loading and microparticle crosslinking extent. For instance, by loading TGF-β1 into the gelatin microparticle phase, a burst release of 10.8±0.7% was achieved, while loading this growth factor into the OPF hydrogel phase resulted in a burst release of 25.2±1.5%. With either system, simultaneous, slow release of IGF-1 over a 4-week period was accomplished by selectively loading this protein into highly crosslinked, encapsulated microparticles. These results demonstrate the utility of these systems in future studies to assess the interplay and time course of multiple growth factors in cartilage repair.

Use of a polymeric device to deliver growth factors to a healing fracture.

Fracture healing is a cascade of events regulated by systemic and local factors. Local growth factors are believed to play an integral role. The present study evaluates the effects of basic fibroblast growth factor (bFGF) and insulin-like growth factor-1 (IGF-1) using a controlled delivery system in a closed rodent femoral fracture model. Female Wistar rats (n = 144) were used in the present study. Animals were randomly allocated to six groups, with each group divided into three time-points of 2, 4 and 8 weeks. Two groups had growth factor administered. The others had no operation or sham operations. Growth factors were delivered to the fracture site using a specialised delivery system. This consisted of a Kirschner-wire coated with ethylene vinyl acetate co-polymer embedded with growth factors, inserted as an intramedullary nail. Fractures were effected with a three-point bending device. Femurs were tested in four-point bending and structural properties of peak load and stiffness were obtained from the load-displacement graphs. Specimens were prepared for qualitative analysis under a light microscope and using immunohistochemistry, specimens were analysed for expression of bFGF, IGF-1 and transforming growth factor beta (TGF-beta). The growth factor-treated groups exhibited larger calluses at 2 and 4 weeks. Four-point bending showed weaker structural properties (stiffness and peak load) at 4 weeks in both growth factor-treated groups. Administration of bFGF or IGF-1 increased the ratio of cartilaginous to mesenchymal tissues in the fracture callus compared with non-treated animals at 2 and 4 weeks. Immunostaining intensity and distribution of both growth factors in the treated groups was greater than the non-treated groups. Exogenous delivery of bFGF or IGF-1 alters the course of fracture healing.

Effect of insulin-like growth factor-1 (IGF-1) plus alendronate on bone density during puberty in IGF-1-deficient MIDI mice

Insulin-like growth factor-1 (IGF-1) increases both bone formation and bone resorption processes. To test the hypothesis that treatment with an antiresorber along with IGF-1, during the pubertal growth phase, would be more effective than IGF-1 alone to increase peak bone mass, we used an IGF-1 MIDI mouse model, which exhibits a >60% reduction in circulating IGF-1 levels. We first determined an optimal IGF-1 delivery by evaluating IGF-1 administration (2 mg/kg body weight/day) by either a single daily injection, three daily injections, or by continuous delivery via a minipump during puberty. Of the three regimens, the three daily IGF-1 injections and IGF-1 through a minipump produced a significant increase in total body bone mineral density (BMD) (6.0% and 4.4%, respectively) and in femoral BMD (4.3% and 6.2%, respectively) compared with the control group. Single subcutaneous (s.c.) administration did not increase BMD. We chose IGF-1 administration three times daily for testing the combined effects of IGF-1 and alendronate (100 μg/kg per day). The treatment of IGF-1 + alendronate for a period of 2 weeks increased total body BMD at 1 week and 3 weeks after treatment (21.1% and 20.5%, respectively) and femoral BMD by 29% at 3 weeks after treatment. These increases were significantly greater than those produced by IGF-1 alone. IGF-1, but not alendronate, increased bone length. IGF-1 and/or alendronate increased both periosteal and endosteal circumference. Combined treatment caused a greater increase in the total body bone mineral content (BMC) and periosteal circumference compared with individual treatment with IGF-1 or alendronate. Our data demonstrate that: (1) inhibition of bone turnover during puberty increases net bone density; and (2) combined treatment with IGF-1 and alendronate is more effective than IGF-1 or alendronate alone in increasing peak bone mass in an IGF-1-deficient MIDI mouse model.

Augmentation of Adipofascial Flaps Using the Long-Term Local Delivery of Insulin and Insulin-Like Growth Factor-1

The adipofascial flaps currently described in the literature frequently lack the volume requirements for reconstructive goals. In this study, the authors examined the use of long-term local delivery of insulin and insulin-like growth factor-1 (IGF-1) using polylactic-coglycolic acid/polyethylene glycol (PLGA/PEG) microspheres to augment inguinal adipofascial flaps based on the inferior epigastric vessels in the rat. Two flap models, the island flap and the limited dissection flap, were used to demonstrate simultaneous treatment and pretreatment modalities, respectively. Experimental groups received 12.5 mg of insulin microspheres (carrying 1 IU of insulin) plus 12.5 mg of IGF-1 microspheres (carrying 2.5 microg of IGF-1). A group undergoing the operation only (no treatment with microspheres) and a group treated with blank microspheres (no growth factor) served as external controls for the surgical procedure and the drug delivery device, respectively. In all groups (n = 5 animals in each), the contralateral flap served as an internal control. Upon harvest on postoperative day 28, the insulin and IGF-1-treated flaps in both models weighed statistically more than the internal control flaps and the two external control flaps. Likewise, on gross inspection, the adipogenic growth factor-treated flaps had greater volumes than the internal control flap groups and both of the external control flap groups (operation only and blank microspheres). Other intergroup comparisons suggested the absence of a systemic insulin and IGF-1 effect on adiposity. A histomorphometric analysis suggested (1) that insulin and IGF-1 treatment does not alter flap cell composition and (2) that flap augmentation is secondary to the stimulation of cell proliferation and adipocytic differentiation rather than the hypertrophy of mature adipocytes. Further evidence in favor of cell proliferation and differentiation was the discovery of nonanatomic, ectopic fat islands on the pedicle sheath of the treated flaps and the lack of variation in cell size distribution among groups. The authors concluded that the long-term local delivery of insulin and IGF-1 with PLGA/PEG microspheres is an effective method of adipofascial flap augmentation; this method increases the number of mature adipocytes rather than increasing the size of preexisting cells.