Defense Date

5-27-2025

Graduation Date

Summer 8-8-2025

Availability

Immediate Access

Submission Type

thesis

Degree Name

MS

Department

Pharmaceutics

School

School of Pharmacy

Committee Chair

Jelena M. Janjic Ph.D.

Committee Member

James Drennen, III, Ph.D.

Committee Member

Wilson Meng Ph.D.

Keywords

Macrophages, Nanoemulgel, Neuroregeneration

Abstract

Peripheral nerve injury (PNI) refers to damage to the nerve located outside the brain and spinal cord. PNI is commonly caused by a physical accident (trauma) or medical operation1, 2. The impact on the quality of life of PNI patients is directly correlated to neuropathic pain, resultant depression, and increased stress2. Even after recovering from the PNI, patients still experience phantom pain and lose the ability to work due to the loss of tactile sensory or motor function2-4. However, unlike damage to the central nervous system (CNS), peripheral nerves can regenerate because macrophages clean up debris with assistance from Schwann cells5. In the early phases of nerve injury, macrophage recruitment and rapid clearance of myelin debris improve blood-nerve barrier permeabilization and activate nearby Schwann cells to produce new myelin1, 6. These macrophages then help facilitate nerve regeneration. In the process of neuroregeneration, the axotomy results in fragmentation of the distal axon and myelin sheaths7. Nerve injury induces upregulation of collagen VI derived from Schwann cells and macrophages, recruiting macrophages into injured sites in a paracrine and autocrine manner via phosphoinositide 3-kinase and protein kinase A signaling pathways, further enhancing peripheral nerve regeneration8, 9. Schwann cells proliferate, and macrophages invade the distal nerve segment to phagocytose degraded materials6, 10. Based on the injury type and cellular and molecular environment, macrophages can assume two well-studied phenotypes, pro-inflammatory (M1) and anti-inflammatory/pro-regenerative (M2). Therefore, the polarization and migration of the macrophages contribute to peripheral nerve regeneration.

The key enzyme cyclooxygenase-2 (COX-2) plays an important role in the macrophage-driven neuroinflammatory processes11. Infiltrating macrophages overexpress COX-2, leading to an increased production of PGE2. COX-2 is a popular therapeutic target of NSAIDs (such as Celecoxib). Celecoxib (CXB), a non-steroidal anti-inflammatory drug (NSAID), is a selective COX-2 inhibitor and can be used as a non-opioid alternative pain treatment12-14. It belongs to the biopharmaceutical classification system (BCS) class II, which indicates that the in vivo efficacy of CXB is severely limited due to its poor aqueous solubility, leading to low bioavailability15. Due to its poor water solubility, it is challenging to deliver CXB for inflammation treatment through conventional dosage forms (e.g., oral dosage form)16-19. Therefore, we propose a nanoemulgel-based system that can encapsulate celecoxib and locally deliver pain relief at PNI sites to treat pain and neuropathy.

This nanoemulgel (NEG) is composed of nanoemulsion (NE) and thermo-responsive gelling agent (e.g., Pluronic F127). As a lipophilic therapeutic drug carrier, NE is a major component in the NEG. We have previously shown that intravenously delivered CXB-NE modulates macrophage/monocyte infiltrating patterns, and the anti-inflammatory action leads to a six-day pain relief in a rat chronic constriction injury model20-22. We hypothesize that celecoxib-loaded nanoemulgel (CXB-NEG) can provide neuroimmunomodulation at the site of injury, leading to accelerated nerve recovery. CXB-NEG can also sustain anti-inflammatory and analgesic effects locally at the site of nerve injury and/or inflammation. Accumulated at the site of injury, polarized M2 macrophages contribute to neuroregeneration.

The celecoxib-loaded nanoemulgel (CXB-NEG) exhibits excellent colloidal stability and thermos-responsive behavior. We took the nanoemulgel dilution in the biological medium for the sterility test and found no significant change in OD600 absorbance. In in vitro cell work, we did not observe any toxic effects on macrophages and Schwann cells when exposed to nanoemulgels. Moreover, we found that the drug-loaded nanoemulgel can inhibit COX-2 and reduce the production of PGE2 in vitro (Figure 1). Lastly, we evaluated the sciatic nerve transection and repair (SN-TSR) model in rats using in vivo NIRF live imaging to monitor drug delivery and therapeutic efficacy. The NIRF (label infiltrating macrophages) signal emitted by the CXB-NEG treatment group was reduced compared to the rats treated with drug-free nanoemulgel. Our histology data (H&E) showed evidence of axonal preservation and less fascicular disorganization in rat nerve samples that were treated with CXB-NEG. The immunofluorescence data showed that CXB-NEG promotes the expression of several neuroregenerative biomarkers (NFM, S100, Kronx20, Oct6, GFAP, and GAP43) in nerve tissues, which indicates Schwann cell proliferation and re-myelination, then leads to neuroregeneration.

To the best of our knowledge, this nanoemulgel platform demonstrates potential as an effective theranostic tool for the treatment and study of acute post-trauma nerve injury and pain. In this work, we evaluate release kinetics and safety of the nano-formulation with optimized drug payloads in vivo in an inflammatory model of nerve injury and peripheral neuropathy. The presented pain nanomedicines provide a single, long-lasting, low-dose, non-opioid means of pain relief at the point of injury and/or disease through targeted immunomodulation and neurodegeneration.

Language

English

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