Abstract
Statement of Purpose: Pathogens have evolved over time to evade the host immune system in various ways. For instance, the fungal species Cryptococcus neoformans, following engulfment by phagocytes, has been observed to stay alive within the acidic phagolysosome and escape through a process called vomocytosis1. Using this phenomenon, C. neoformans utilizes host immune cells to disseminate intection throughout the body. One condition due to infection of this microbe is cryptococcal meningitis (CM). Primarily affecting immunocompromised individuals, including an estimated 220,000 HIV/AIDS patients, CM causes -181,000 deaths/ year worldwide2.Greater understanding of vomocytosis could lead to the development of new CM treatments for patients, as well as novel biomaterial particulate vaccines. However, the underlying mechanisms of this phenomenon are unknown.To study vomocytosis, a method for quantifying phagocytosis and expulsion rates is required. Current studies use manual counting of vomocytic events3 (Fig. 1) or multi-step flow cytometry staining4 with limited success. This work characterizes a novel dual fluorescent reporter system for precise monitoring of phagocytic entry and vomocytic expulsion. The molecular signaling system is composed of tethered Enzyme-Labeled Fluorescence (ELF-97) and Fluorogen Activating Peptide (FAP). ELF-97 is a molecule that fluorescently activates upon cleavage by lysosomal phosphatases, indicating successful phagosomal uptake.The second indicator, FAP, is a protein that activates a cell- impermeable fluorogen (either aRED-np or MG Beta Tau) while outside of the cytoplasm and will be used to confirm escape from phagocytes. Here, poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) are used to demonstrate proof-of-concept for the transfer of the reporter system to C. neoformans and particulates.
| Original language | English (US) |
|---|---|
| Title of host publication | Society for Biomaterials Annual Meeting and Exposition 2019 |
| Subtitle of host publication | The Pinnacle of Biomaterials Innovation and Excellence - Transactions of the 42nd Annual Meeting |
| Publisher | Society for Biomaterials |
| Number of pages | 1 |
| ISBN (Electronic) | 9781510883901 |
| State | Published - Jan 1 2019 |
| Event | 42nd Society for Biomaterials Annual Meeting and Exposition 2019: The Pinnacle of Biomaterials Innovation and Excellence - Seattle, United States Duration: Apr 3 2019 → Apr 6 2019 |
Publication series
| Name | Transactions of the Annual Meeting of the Society for Biomaterials and the Annual International Biomaterials Symposium |
|---|---|
| Volume | 40 |
| ISSN (Print) | 1526-7547 |
Conference
| Conference | 42nd Society for Biomaterials Annual Meeting and Exposition 2019: The Pinnacle of Biomaterials Innovation and Excellence |
|---|---|
| Country | United States |
| City | Seattle |
| Period | 4/3/19 → 4/6/19 |
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ASJC Scopus subject areas
- Biochemistry
- Biophysics
- Biotechnology
- Biomaterials
- Materials Chemistry
Cite this
Environment-responsive two-fluorophore reporter system : A potential tool to monitor particulate vomocytosis. / Pacifici, Noah; Bolandparvaz, Amir; Shams, Shahin; Stilhanou, Roberta; Silva, Eduardo; Lewis, Jamal.
Society for Biomaterials Annual Meeting and Exposition 2019: The Pinnacle of Biomaterials Innovation and Excellence - Transactions of the 42nd Annual Meeting. Society for Biomaterials, 2019. (Transactions of the Annual Meeting of the Society for Biomaterials and the Annual International Biomaterials Symposium; Vol. 40).Research output: Chapter in Book/Report/Conference proceeding › Conference contribution
}
TY - GEN
T1 - Environment-responsive two-fluorophore reporter system
T2 - A potential tool to monitor particulate vomocytosis
AU - Pacifici, Noah
AU - Bolandparvaz, Amir
AU - Shams, Shahin
AU - Stilhanou, Roberta
AU - Silva, Eduardo
AU - Lewis, Jamal
PY - 2019/1/1
Y1 - 2019/1/1
N2 - Statement of Purpose: Pathogens have evolved over time to evade the host immune system in various ways. For instance, the fungal species Cryptococcus neoformans, following engulfment by phagocytes, has been observed to stay alive within the acidic phagolysosome and escape through a process called vomocytosis1. Using this phenomenon, C. neoformans utilizes host immune cells to disseminate intection throughout the body. One condition due to infection of this microbe is cryptococcal meningitis (CM). Primarily affecting immunocompromised individuals, including an estimated 220,000 HIV/AIDS patients, CM causes -181,000 deaths/ year worldwide2.Greater understanding of vomocytosis could lead to the development of new CM treatments for patients, as well as novel biomaterial particulate vaccines. However, the underlying mechanisms of this phenomenon are unknown.To study vomocytosis, a method for quantifying phagocytosis and expulsion rates is required. Current studies use manual counting of vomocytic events3 (Fig. 1) or multi-step flow cytometry staining4 with limited success. This work characterizes a novel dual fluorescent reporter system for precise monitoring of phagocytic entry and vomocytic expulsion. The molecular signaling system is composed of tethered Enzyme-Labeled Fluorescence (ELF-97) and Fluorogen Activating Peptide (FAP). ELF-97 is a molecule that fluorescently activates upon cleavage by lysosomal phosphatases, indicating successful phagosomal uptake.The second indicator, FAP, is a protein that activates a cell- impermeable fluorogen (either aRED-np or MG Beta Tau) while outside of the cytoplasm and will be used to confirm escape from phagocytes. Here, poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) are used to demonstrate proof-of-concept for the transfer of the reporter system to C. neoformans and particulates.
AB - Statement of Purpose: Pathogens have evolved over time to evade the host immune system in various ways. For instance, the fungal species Cryptococcus neoformans, following engulfment by phagocytes, has been observed to stay alive within the acidic phagolysosome and escape through a process called vomocytosis1. Using this phenomenon, C. neoformans utilizes host immune cells to disseminate intection throughout the body. One condition due to infection of this microbe is cryptococcal meningitis (CM). Primarily affecting immunocompromised individuals, including an estimated 220,000 HIV/AIDS patients, CM causes -181,000 deaths/ year worldwide2.Greater understanding of vomocytosis could lead to the development of new CM treatments for patients, as well as novel biomaterial particulate vaccines. However, the underlying mechanisms of this phenomenon are unknown.To study vomocytosis, a method for quantifying phagocytosis and expulsion rates is required. Current studies use manual counting of vomocytic events3 (Fig. 1) or multi-step flow cytometry staining4 with limited success. This work characterizes a novel dual fluorescent reporter system for precise monitoring of phagocytic entry and vomocytic expulsion. The molecular signaling system is composed of tethered Enzyme-Labeled Fluorescence (ELF-97) and Fluorogen Activating Peptide (FAP). ELF-97 is a molecule that fluorescently activates upon cleavage by lysosomal phosphatases, indicating successful phagosomal uptake.The second indicator, FAP, is a protein that activates a cell- impermeable fluorogen (either aRED-np or MG Beta Tau) while outside of the cytoplasm and will be used to confirm escape from phagocytes. Here, poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) are used to demonstrate proof-of-concept for the transfer of the reporter system to C. neoformans and particulates.
UR - http://www.scopus.com/inward/record.url?scp=85065400778&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85065400778&partnerID=8YFLogxK
M3 - Conference contribution
AN - SCOPUS:85065400778
T3 - Transactions of the Annual Meeting of the Society for Biomaterials and the Annual International Biomaterials Symposium
BT - Society for Biomaterials Annual Meeting and Exposition 2019
PB - Society for Biomaterials
ER -