Investigating olfactory dysfunction in patients and human olfactory organoids

Summary. Project 2. Investigating olfactory dysfunction in patients and human olfactory organoids. Olfactory dysfunction (OD) describes disorders of smell ranging from complete loss of smell (anosmia) to reduced or altered smell perception (hyposmia and parosmia, respectively). Over 100 million individuals worldwide and 1.4% of the US population are impacted by OD, with etiologies including normal aging, neurodegenerative disorders, congenital conditions, head trauma, cancer treatments, sinonasal inflammatory disease, and upper respiratory viral infection. The SARS-CoV-2 pandemic has emphasized the significant rates and implications of OD. Deficits in olfaction contribute to cognitive problems, decreased overall quality of life, and increased mortality. While many patients with SARS-CoV-2-induced OD recover quickly, some develop long-term smell loss that could become permanent. The underlying causes of long-term smell loss remain poorly understood. This project will identify causes of long-term OD triggered by SARS-CoV-2, using deep molecular and cellular analyses of olfactory tissue samples from patients with OD and establishing causal relationships using a novel in vitro model of human olfactory tissue. Our central hypotheses are the following: (1) persistent inflammation in olfactory tissue accounts for long-term post-viral smell loss, and (2) human olfactory epithelium can be grown in three-dimensional organoid culture as an in vitro model to investigate mechanisms and potential treatments of viral-induced smell loss. Project 2 will test these hypotheses in two Specific Aims. Aim 2.1 will test the hypothesis that inflammation persists in olfactory tissue in patients with long-term post-COVID OD. We will also determine whether SARS-CoV-2 RNA can be detected in the olfactory mucosa of patients with post-COVID smell loss. This part of the project will generate comprehensive datasets from a large cohort of patients with post-COVID OD to fill a critical knowledge gap. Aim 2.2 will utilize a novel human olfactory organoid model we have developed that has been characterized molecularly, cellularly, and functionally. With this model we will investigate viral- and inflammatory cytokine-induced changes for smell loss. We will (1) infect the organoids with SARS-CoV-2 WA1, Delta, and Omicron variants and (2) expose the organoids to inflammatory cytokines. We will then investigate the effects of both exposures on gene expression, cell survival, differentiation, proliferation, and function in human olfactory organoids - disruption of any of these processes can lead to OD. This project will recruit patients with long-term OD, control participants with normal olfaction, and skull base surgery patients to donate a steady supply of olfactory biopsy tissues for the in vitro organoid model. Project 2 will synergize with other projects in the P50 program by sharing resources and expertise and collaborating on patient recruitment, sensory testing, and experimental designs. This study will provide a mechanistic understanding of long-term post-viral smell loss that will direct efforts at prevention and treatment in the future. The organoid model can be used to test treatments and will also benefit studies on human olfactory tissue regeneration.