Hemmerla, A. J. | Search bioRxiv

bioRxiv2024

Cytoprotective and Neuroinductive Effects of Thiol-Containing Simple Signaling Molecules

Around 20 million Americans, mostly young adults, have suffered from a peripheral nerve injury due to trauma or a medical condition. However, prevailing treatments, including those that employ exogenous growth factors, often exhibit drawbacks outweighing their benefits, highlighting the need for novel solutions. Non-complex agents that govern bioactivity, termed simple signaling molecules (SSMs), offer an exciting alternative when compared to traditional approaches due to their small size, ready availability, and influence on cellular pathways. This study explores the cytoprotective and neuroinductive effects of three thiol-containing SSMs: hydrogen sulfide (H2S), n-acetyl cysteine (NAC), and glutathione (GSH). Neural stem cells (NE-4Cs) were exposed to toxic levels of hydrogen peroxide (H2O2) while supplemented with different concentrations of H2S, NAC, or GSH to assess their cytoprotective effects. To study the neuroinductivity of H2S, NAC, and GSH with NE-4Cs, immunofluorescence microscopy for an early neuronal marker, {beta}3-tubulin, was employed to determine stem cell neural differentiation. Significant cytoprotection was found for NAC (2 - 8 mM) and GSH (2 - 12 mM), while H2S showed limited protective effects. Interestingly, the opposite result was observed for neuroinductivity as only H2S (7.75 - 125 M) achieved a desirable effect while NAC and GSH had minimal impact on neural differentiation. This research establishes therapeutic concentration ranges of H2S, NAC, and GSH for future drug delivery systems targeting neural regeneration, especially for peripheral nerve injuries. LAY SUMMARYApproximately 20 million Americans, mainly young adults, suffer from peripheral nerve injuries, necessitating new effective treatments. Currently emerging approaches, including growth factor-based therapies, unfortunately face considerable limitations. This study explores the potential of simple signaling molecules (SSMs) - small, readily available agents - specifically hydrogen sulfide (H2S), n-acetyl cysteine (NAC), and glutathione (GSH) for their potential to be used to address issues underlying peripheral nerve injuries. Our findings reveal NAC and GSH have cytoprotective effects, whereas H2S is neuroinductive. This research identifies optimal concentrations for future drug delivery targeting neural regeneration, especially beneficial for peripheral nerve injuries. FUTURE WORKSUpcoming investigation will concentrate on exploiting the potential of novel SSM-releasing monomers within degradable polymeric systems. This involves defining therapeutic windows and optimizing polymer chemistry to induce desired cytoprotective and neuroinductive effects in neural stem cells, crucial for peripheral nerve injury repair. Emphasizing controlled release, this approach holds promise for developing advanced therapeutic strategies utilizing H2S, NAC, and/or GSH in nerve guidance conduits. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/597935v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@187657dorg.highwire.dtl.DTLVardef@17ffa9org.highwire.dtl.DTLVardef@191dc67org.highwire.dtl.DTLVardef@11d0fd2_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2022

Spinal Fusion Properties of Mechanically-Reinforced, Osteomodulatory Chitosan Hydrogels

Lower back pain is a considerable medical problem that will impact 80% of the U.S. population at some point in their life. For the most severe cases, surgical repair is necessary and is associated with costs upwards of $10.2 billion annually in the United States. To alleviate back pain, spine fusions are a common treatment in which two or more vertebrae are biologically fused together often through the use of a graft material. Unfortunately, iliac crest bone autograft, the current gold standard graft material, can yield insufficient fusion and is associated with considerable donor site morbidity and pain as well as limited supply. Therefore, new materials need to be developed in order to better coordinate healing and new bone growth in the affected area to reduce unnecessary patient burden. In order to address this issue, the incorporation of allograft and one of two types of cellulose (i.e., 0CNCs and CNFs) into a dual-crosslinked chitosan hydrogel loaded with bioactive calcium phosphate was investigated. Hydrogels were then tested for both their material and biological properties. Specifically, hydrogel swelling ratio, mass loss, ion release profile, compressive strength, in vitro biocompatibility and osteoinduction as well as in vivo biocompatibility, and effectiveness in a spine fusion model were determined. Cellulose and allograft incorporation significantly improved hydrogel compressive strength and biocompatibility and CNFs were found to be a significantly more biocompatible form of cellulose than 0CNCs. Additionally, through the controlled delivery of osteoinductive simple signaling molecules (i.e., calcium and phosphate ions), DCF-loaded CNF/Chitosan hydrogels were able to induce osteoblast-like activity in murine mesenchymal stem cells. When evaluated in vivo, these hydrogels were found to be non-toxic though the subacute phase (14 days). A 6-week rabbit spine fusion found these materials to achieve near complete fusion when assessed radiographically. This research provides considerable support for the utility of our novel material for spine fusion procedures as well as other future bone applications.

bioengineering↗