Research

CRISPR/Cas9 for Musculoskeletal Disorders

At the moment, patients suffering from degenerative disc disease (DDD) and numerous other musculoskeletal disorders are treated conservatively, i.e. with physiotherapy and analgesic medication, but may have to undergo surgery if the symptoms prevail. Novel treatment options may arise from using the CRISPR/Cas9 gene editing technique that can precisely target specific DNA sequences, allowing to correct an unfavorable genetic background in affected individuals. As such, CRISPR/Cas9 has great potential to change the traditional symptom-driven treatment of age-related musculoskeletal disorders towards therapies that target the underlying mechanisms in a personalized manner. For DDD, CRISPR/Cas9 could be used to target genetic factors associated with DDD, inflammation receptors, catabolic molecules or senescence genes. Furthermore, stem and progenitor cells can be modified by CRISPR to better withstand the harsh microenvironment of the intervertebral disc. In this project, the therapeutic potential of CRISPR/Cas9 knock-out, knock-down and/or activation is explored.

Electrosprayed Biomaterials for Drug Delivery

While oral delivery of pharmaceutical compounds generally has the highest compliance rates, local delivery of drugs can often be more effective. To prolong the effects of the selected therapeutics, such as substances with anti-inflammatory, anti-catabolic, and/or anti-oxidant properties, slow-release systems that provide sustained delivery and enhanced protection against degradation are needed. In this context, electrospraying is particularly useful as it is a gentle electrohydrodynamic encapsulation method that can be conducted under less harsh conditions than most other encapsulation techniques and is hence specifically suitable for sensitive substances such as natural polyphenols. In this project, we aim to create electrosprayed nano-/microparticles for improved drug delivery. We currently focus on encapsulating the polyphenol resveratrol for improved wound healing, but this approach is also relevant for other drugs and application areas.

Electrospun 3D Skin Models

3D skin models have become increasingly relevant over the past years, especially due to society’s goal to reduce animal testing. Their fabrication has been significantly advanced as a result of enhanced biofabrication techniques, such as 3D bioprinting and electrospinning, hence representing an alternative approach to classical, yet flawed, collagen hydrogel models. Electrospinning allows for the cost-effective creation of nanofibrous 3D skin models that bear high similarity to the native ECM, promoting cell adhesion, proliferation, and differentiation. However, classical electrospinning results in membranes with a relatively low pore size that hinders cell infiltration. One of the methods to enhance pore size and hence cell infiltration is through cryogenic electrospinning. The goal of this project is to combine normal and cryogenic electrospinning to create 3D organotypic skin models seeded with dermal cells. While the basic model mimics healthy skin, incorporation of diseased cells and/or addition of biological cues allow advancing it into specific disease models, e.g. for inflammatory skin diseases or systemic sclerosis.

EVs from CRISPR-Modified Stem Cells

Degenerative disc disease (DDD) is a major cause of low back pain, which is the leading cause of activity limitation and work absence and results in a high economic burden. In the US, the yearly costs related to back pain are around US$ 100 to 200 billion. Despite the relevance of DDD, current treatments are neither satisfactory in outcome, nor do they target the underlying pathophysiological mechanisms, i.e. degeneration and inflammation. While mesenchymal stem cells (MSCs) have in theory great potential for the treatment of DDD due to their regenerative and anti-inflammatory capacities, their success has been limited due to the harsh microenvironment of the degenerated disc that hampers their survival and functionality. On the other hand, extracellular vesicles (EVs) produced by MSCs have recently been highlighted as alternative treatments. With the latest developments of the CRISPR technique, MSCs can be genetically modified in ways that enhance their therapeutic potential. The aim of this project is to investigate the potential of EVs produced by CRISPR-modified MSCs to treat DDD. In collaboration with Prof. Gaborski (BME, RIT), sophisticated nanomembranes will be used for size sorting of EVs.

TLR-Associated MicroRNAs

Dysregulation of inflammation in degenerative disc disease, which is believed to be associated with pain development, is based on complex molecular mechanisms. One of the main regulators of inflammation are Toll-like receptors (TLRs), amongst which TLR2 is specifically relevant in disc pathophysiology. Importantly, inflammation is closely linked with mechanical loading in the IVD as high loads not only induce inflammation themselves, but also modulate TLR expression and lead to the accumulation of damage-associated molecular patterns with inflammatory/TLR activating capacity. As TLRs are an essential component to tissue inflammation, their tight control is crucial. In numerous cell types, microRNAs (= small noncoding RNAs) have been shown to modulate TLR signaling and the downstream inflammation and catabolism, both in health and disease. This project will be the first to identify and comprehensively investigate miRNAs in the context of TLR activation in the IVD and will thus elucidate posttranscriptional regulation of genes associated with IVD pathophysiology under the linked concepts of inflammation and mechanoregulation.

Substrate Stiffness, Topography, and TRPV4 in Disc Mechanotransduction

Over the past decade(s), research has highlighted that substrate stiffness and architecture/topography can be recognized by cells and serve as mechanical and topographical cues that ultimately drive cell behavior through mechanoreceptors. Substrate changes can also affect the mechanical stimulation of cells and thus their response to loading. These cell responses are largely governed through mechanosensitive ion channels, such as the transient receptor potential (TRP) channels. This project aims to determine the relevance of substrate stiffness and topography on TRPV4 activation in intervertebral disc cells (AF cells) in response to a pharmacological TRPV4 agonist as well as cyclic stretching and its relevance in regulating extracellular matrix synthesis and remodeling. Overall, we hope to reveal the relevance of cell-substrate processes in intervertebral disc health and disease and to use this knowledge in the development of regenerative approaches.

TRPC6 Modulation to Treat Scleroderma

Scleroderma is a rare disease commonly characterized by chronic hardening (fibrosis) of body tissues including the skin due to excessive accumulation of collagen. Despite intensive research, no therapies are effective in treating scleroderma-related skin disease. The ion channel TRPC6 is involved in numerous fibrotic diseases, but its role in scleroderma has not yet been investigated. Therefore, the goal of this project is to investigate the potential of TRPC6 inhibition in treating scleroderma. Experiments will first be conducted in classical 2D culture, thereafter in a new 3D human fibrosis model, and subsequently in a well-established mouse model. By investigating a new therapeutic approach with high potential, this project will hopefully result in the first effective treatment for scleroderma patients suffering from skin hardening and tightening. 

TRCP6 in Intervertebral Disc Mechanosensing

Mechanical loading is involved in many degenerative and regenerative processes in mechanosensitive tissues, including the intervertebral disc. While physiological mechanostimulation is favorable and even necessary to maintain intervertebral disc homeostasis, high levels of mechanical loading are a well-known contributor to intervertebral disc degeneration. However, the underlying mechanotransduction processes are not well understood to date. The overall objective of this project is to elucidate the role of the ion channel TRPC6 in sensing and transducing mechano-osmotic stress in the intervertebral disc and its contribution to degenerative disc disease. To achieve this goal, bioreactors for cell stretching and compression as well as adjustment of medium osmolarity are used. This research has the potential to not only better understand intervertebral disc mechanobiology, but also to further highlight TRPC6 as a therapeutic target in degenerative disc disease.