Using bimodal MRI/fluorescence imaging to identify host angiogenic response to implants.

Therapies that promote angiogenesis have been successfully applied using various combinations of proangiogenic factors together with a biodegradable delivery vehicle. In this study we used bimodal noninvasive monitoring to show that the host response to a proangiogenic biomaterial can be drastically affected by the mode of implantation and the surface area-to-volume ratio of the implant material. Fluorescence/MRI probes were covalently conjugated to VEGF-bearing biodegradable PEG-fibrinogen hydrogel implants and used to document the in vivo degradation and liberation of bioactive constituents in an s.c. rat implantation model. The hydrogel biodegradation and angiogenic host response with three types of VEGF-bearing implant configurations were compared: preformed cylindrical plugs, preformed injectable microbeads, and hydrogel precursor, injected and polymerized in situ. Although all three were made with identical amounts of precursor constituents, the MRI data revealed that in situ polymerized hydrogels were fully degraded within 2 wk; microbead degradation was more moderate, and plugs degraded significantly more slowly than the other configurations. The presence of hydrogel degradation products containing the fluorescent label in the surrounding tissues revealed a distinct biphasic release profile for each type of implant configuration. The purported in vivo VEGF release profile from the microbeads resulted in highly vascularized s.c. tissue containing up to 16-fold more capillaries in comparison with controls. These findings demonstrate that the configuration of an implant can play an important role not only in the degradation and resorption properties of the materials, but also in consequent host angiogenic response.

TVP1022 protects neonatal rat ventricular myocytes against doxorubicin-induced functional derangements.

Our recent studies demonstrated that propargylamine derivatives such as rasagiline (Azilect, Food and Drug Administration-approved anti-Parkinson drug) and its S-isomer TVP1022 protect cardiac and neuronal cell cultures against apoptotic-inducing stimuli. Studies on structure-activity relationship revealed that their neuroprotective effect is associated with the propargylamine moiety, which protects mitochondrial viability and prevents apoptosis by activating Bcl-2 and protein kinase C-epsilon and by down-regulating the proapoptotic protein Bax. Based on the established cytoprotective and neuroprotective efficacies of propargylamine derivatives, as well as on our recent study showing that TVP1022 attenuates serum starvation-induced and doxorubicin-induced apoptosis in neonatal rat ventricular myocytes (NRVMs), we tested the hypothesis that TVP1022 will also provide protection against doxorubicin-induced NRVM functional derangements. The present study demonstrates that pretreatment of NRVMs with TVP1022 (1 microM, 24 h) prevented doxorubicin (0.5 microM, 24 h)-induced elevation of diastolic [Ca(2+)](i), the slowing of [Ca(2+)](i) relaxation kinetics, and the decrease in the rates of myocyte contraction and relaxation. Furthermore, pretreatment with TVP1022 attenuated the doxorubicin-induced reduction in the protein expression of sarco/endoplasmic reticulum calcium (Ca(2+)) ATPase, Na(+)/Ca(2+) exchanger 1, and total connexin 43. Finally, TVP1022 diminished the inhibitory effect of doxorubicin on gap junctional intercellular coupling (measured by means of Lucifer yellow transfer) and on conduction velocity, the amplitude of the activation phase, and the maximal rate of activation (dv/dt(max)) measured by the Micro-Electrode-Array system. In summary, our results indicate that TVP1022 acts as a novel cardioprotective agent against anthracycline cardiotoxicity, and therefore potentially can be coadmhence, theinistered with doxorubicin in the treatment of malignancies in humans.

TVP1022 and propargylamine protect neonatal rat ventricular myocytes against doxorubicin-induced and serum starvation-induced cardiotoxicity.

We recently reported that propargylamine derivatives such as rasagiline (Azilect) and its S-isomer TVP1022 are neuroprotective. The aim of this study was to test the hypothesis that the neuroprotective agents TVP1022 and propargylamine (the active moiety of propargylamine derivatives) are also cardioprotective. We specifically investigated the protective efficacy of TVP1022 and propargylamine in neonatal rat ventricular myocytes (NRVM) against apoptosis induced by the anthracycline chemotherapeutic agent doxorubicin and by serum starvation. We demonstrated that pretreatment of NRVM cultures with TVP1022 or propargylamine attenuated doxorubicin-induced and serum starvation-induced apoptosis, inhibited the increase in cleaved caspase 3 levels, and reversed the decline in Bcl-2/Bax ratio. These cytoprotective effects were shown to reside in the propargylamine moiety. Finally, we showed that TVP1022 neither caused proliferation of the human cancer cell lines HeLa and MDA-231 nor interfered with the anti-cancer efficacy of doxorubicin. These results suggest that TVP1022 should be considered as a novel cardioprotective agent against ischemic insults and against anthracycline cardiotoxicity and can be coadministered with doxorubicin in the treatment of human malignancies.

Protein composition alters in vivo resorption of PEG-based hydrogels as monitored by contrast-enhanced MRI.

We report on the use of magnetic resonance imaging (MRI)-based non-invasive monitoring to document the role of protein adjuvants in hydrogel implant integration in vivo. Polyethylene glycol (PEG) hydrogels were formed with different protein constituents, including albumin, fibrinogen and gelatin. The hydrogels were designed to exhibit similar material properties, including modulus, swelling and hydrolytic degradation kinetics. The in vivo resorption properties of these PEG-based hydrogels, which contained a tethered gadolinium contrast agent, were characterized by MRI and histology, and compared to their in vitro characteristics. MRI data revealed that PEG-Albumin implants remained completely intact throughout the experiments, PEG-Fibrinogen implants lost about 10% of their volume and PEG-Gelatin implants underwent prominent swelling and returned to their initial volume by day 25. Fully synthetic PEG-diacrylate (PEG-DA) control hydrogels lost about half of their volume after 25 days in vivo. Transverse MRI cross-sections of the implants revealed distinct mechanisms of the hydrogel's biodegradation: PEG-Fibrinogen and PEG-Albumin underwent surface erosion, whereas PEG-Gelatin and PEG-DA hydrogels mainly underwent bulk degradation. Histological findings substantiated the MRI data and demonstrated significant cellular response towards PEG-DA and PEG-Gelatin scaffolds with relatively low reaction towards PEG-Fibrinogen and PEG-Albumin hydrogels. These findings demonstrate that PEG-protein hydrogels can degrade via a different mechanism than PEG hydrogels, and that this difference can be linked to a reduced foreign body response.

Fabrication of PEGylated fibrinogen: a versatile injectable hydrogel biomaterial.

Hydrogels are one of the most versatile biomaterials in use for tissue engineering and regenerative medicine. They are assembled from either natural or synthetic polymers, and their high water content gives these materials practical advantages in numerous biomedical applications. Semisynthetic hydrogels, such as those that combine synthetic and biological building blocks, have the added advantage of controlled bioactivity and material properties. In myocardial regeneration, injectable hydrogels premised on a semisynthetic design are advantageous both as bioactive bulking agents and as a delivery vehicle for controlled release of bioactive factors and/or cardiomyocytes. A new semisynthetic hydrogel based on PEGylated fibrinogen has been developed to address the many requirements of an injectable biomaterial in cardiac restoration. This chapter highlights the fundamental aspects of making this biomimetic hydrogel matrix for cardiac applications.