There are an estimated 300 million people worldwide who are affected by asthma, a respiratory condition in which a person has inflammation and swelling in the airways. Asthma patients also experience increased vasodilation in their lungs, i.e. the widening of blood vessels, which causes increased blood flow and results in increased inflammation. The goal of this project is to create a device that makes free standing hydrogel rings, modelling the structure of blood vessels, offering a simple approach to better understand asthma and potential treatments. The device used to form the rings is 3D printed and can be printed in a range of sizes. The rings are composed of collagen I that has been seeded with smooth muscle cells. Once the hydrogel rings are cast, they can be transferred to a 96 well plate and be free standing of any rigid structures. The ability to be free standing allows us to measure the ring diameter and wall thickness, as well as measure any change in size when a vasodilator is added. Future steps to be taken with this project include optimizing the size for biological relevance, introduce endothelial cells to create multiple layers of cells that are involved in signaling for vasodilation, and increase the responsiveness that the rings have to constriction factors as well as dilators.

Traumatic brain injury (TBI) refers to brain damage resulting from an external force resulting in temporary or permanent impairment of cognitive, physical, and psychosocial functions. Following TBI, widespread neuronal loss occurs, and ischemic and inflammatory processes can greatly increase the extent of neural injury beyond the initial mechanical injury. Microglia are specialized immune cells in the brain that constantly surveil the extracellular environment and respond rapidly to damage by proliferating, phagocytosing debris, and releasing cytokines and chemokines that orchestrate recruitment and regulation of peripheral immune cells to the injured brain post-TBI. With the emergence of chemogenetics, a method by which engineered proteins interact with previously unrecognized chemical activators, inhibitory control can be exerted over microglia activation in a highly specific fashion allowing for precise targeting of brain regions and fewer off-target effects relative to traditional pharmacological approaches. The Weinstein lab aims to examine the effects of targeted inhibition of microglia activation using G-protein coupled receptors called Designer Receptors Exclusively Activated by Designer Drugs (DREADDs). Normally, following traumatic brain injury, the CD68 promoter region is upregulated, resulting in increased microglia expression. However, the inserted HM4Di DREADD gene utilizes this promoter to express the DREADD receptor, and the downstream effects result in neural inflammatory response inhibition. We hypothesize that microglial inhibition will reduce proliferation and local cytokine levels after TBI, thus modulating the inflammatory microenvironment, especially when inhibition is initiated early after TBI. To determine efficacy of DREADDs, we quantify microglia number and proliferation using immunohistochemistry and stereology. We use computer software to capture multi-channel fluorescent images and montages for use in cell counting following stereological methods for random, unbiased sampling of cortical tissue across the TBI epicenter and penumbra. We anticipate that regions expressing activated DREADDs, which should inhibit microglial activation, will have reduced microglia post-TBI relative to controls.