Biological Systems Engineering

 

First Advisor

Rebecca Wachs

Date of this Version

12-3-2021

Citation

Piening, L. (2021). Development and Characterization of a Decellularized Neuroinhibitory Scaffold Containing Matrix Bound Nanovesicles [Master's thesis, University of Nebraska-Lincoln].

Comments

A THESIS Presented to the Faculty of The Graduate College at the University of Nebraska In Partial Fulfillment of Requirements For the Degree of Master of Science, Major: Agricultural & Biological Systems Engineering, Under the Supervision of Professor Rebecca Wachs. Lincoln, Nebraska: December, 2021

Copyright © 2021 Logan M. Piening

Abstract

Chronic low back pain (LBP) is a leading cause of disability but treatments for LBP are limited. Degeneration of the intervertebral disc leads to loss of neuroinhibitory sulfated glycosaminoglycans (sGAGs) which allows nerves from dorsal root ganglia (DRG) to grow into the core of the disc, leading to pain. Current treatments for LBP involve drugs that do not target the source of the pain and lack long term efficacy or use invasive surgeries with high complication rates. Treatment with a decellularized tissue scaffold that contains neuroinhibitory components may inhibit nerve growth and prevent disc-associated LBP. Here, a decellularized nucleus pulposus (NP) tissue scaffold was developed to be used as a treatment for low back pain. Results indicated that the decellularized tissue scaffold developed here removed over 99% of DNA and maintained 70% of the native sGAGs. After gelation, it was determined that the gel was not cytotoxic, and it exhibited neuroinhibitory properties. Following the research on a neuroinhibitory gel, the project shifted to the investigation of a recently discovered nanovesicle with active cargo that may be present in NP tissue. Matrix bound nanovesicles (MBVs) were successfully isolated from NP tissue. Analysis of the MBVs found within the decellularized tissue revealed that they polarized macrophages towards an inflammatory phenotype, which is different to previous literature investigating MBVs. Culture of nucleus pulposus cells with MBVs demonstrated that the NP cells started growing on top of each other, although this was the only morphological change. This work also characterized the protein contents of the NP-derived suggesting that MBVs may be one reason that the NP cannot resolve inflammation during degeneration. The work presented here demonstrates that a decellularized NP tissue scaffold can be formulated into an injectable gel that exhibits neuroinhibitory properties for the treatment of low back pain. Along with this, the presence of MBVs were discovered within the decellularized NP and the NP derived MBVs had unique profile of contents and cellular responses.

Advisor: Rebecca Wachs

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