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| Nile Red-stained vesicles in A549 cells (📷 B. Hariri) |
A large focus of the lab is studying how airway cells sense and respond to infections. One major focus is on the nose and sinuses, also known as the upper airway.
Chronic upper respiratory infections can result in chronic rhinosinusitis (CRS), a disease affecting 8-10% of the US population with direct healthcare costs of over 6 billion dollars annually. CRS has a major impact on quality of life, with CRS patients often suffering debilitating headaches from congestion and sinus pressure, a lack of ability to smell, and other detrimental symptoms. CRS also has a major public health impact, as it accounts for 1 out of every 5 antibiotic prescriptions in adults in the US, making CRS treatment a major contributor to the emergence of antibiotic-resistant organisms. A continuing goal of our research is to better understand airway physiology to identify new and better therapies to treat CRS and other airway diseases without the use of antibiotics. We are particularly interested in the roles of bitter taste receptors (taste family 2 receptors or T2Rs), which detect bacteria in the airways and stimulate beneficial innate immune responses. We are studying how these receptors might be stimulated to enhance airway innate immunity as well as how genetic variations that alter their function might lead to disease. This work is carried out in collaboration with rhinologist surgeons Nithin Adappa, Jim Palmer, and Noam Cohen.
Another focus of the lab is cystic fibrosis (CF), the most common lethal recessive genetic disease in the US. CF is characterized by altered fluid homeostasis in the lung, resulting in impaired mucociliary clearance and innate defense. Our goal is to better understand the molecular and cellular bases of airway epithelial changes in CF to help elucidate how to better treat the disease. This work intersects with taste receptors but also includes studies of the molecular mechanisms of airway submucosal gland fluid secretion, which is impaired in CF lungs.
A final focus on the lab is head and neck cancer. We recently found that T2R bitter taste receptors are expressed in oral and oropharyngeal cancer cells. Our data suggest that stimulation of these receptors might lead to activation of cell death (apoptosis) and thus stimulating these receptors might be a useful cancer therapeutic strategy. This work is being carried out in partnership with Dr. Ryan Carey, an ENT surgeon who is starting a lab as Penn faculty in August 2023.
A final focus on the lab is head and neck cancer. We recently found that T2R bitter taste receptors are expressed in oral and oropharyngeal cancer cells. Our data suggest that stimulation of these receptors might lead to activation of cell death (apoptosis) and thus stimulating these receptors might be a useful cancer therapeutic strategy. This work is being carried out in partnership with Dr. Ryan Carey, an ENT surgeon who is starting a lab as Penn faculty in August 2023.
The tools we use include molecular biology, biochemistry, electrophysiology, and microbiology. We specialize in using live-cell fluorescence and brightfield imaging to monitor real time cell function (physiology) in minimally invasive ways.
Diversity Statement:
We hope to create a welcoming and inclusive community that values and celebrates diversity, including but not limited to gender identity, sexual orientation, race, ethnicity, nationality, age, disability, religion, marital or parental status, socioeconomic status, etc. We recognize that scientific research is strengthened by a diverse workforce that brings different backgrounds, perspectives, ideas, and insights to the table to enhance productivity and innovation. We will continue to strive to maintain an inclusive, respectful, professional, safe, and open environment in our lab where everyone feels valued and respected. While we value intellectual curiosity, rigorous scientific discussions, and asking hard questions about our data, we do not tolerate discrimination, harassment, or any other types of physical or emotional harm. We work everyday to maintain a "non-toxic" lab environment.
Research Support:
This lab has been generously supported by both private foundation and federal grant funds:
Current Research Support:
- National Institutes of Health (NIH) National Institute for Allergy and Infections Diseases (NIAID) R01AI167971 (2022-2026; MPIs with N. Adappa and J. Palmer)
- National Institutes of Health (NIH) National Institute for Deafness and Other Communication Disorders (NIDCD) R01DC016309 (2018-2024), R21DC020041 (2022-2024)
- Cystic Fibrosis Foundation Research Grant LEE21G0 (2021-2023)
- Start-up funding from the Department of Otorhinolaryngology-Head and Neck Surgery of the University of Pennsylvania Perelman School of Medicine (2015)
- Pilot grant from the Penn Diabetes Research Center (through DK19525; 2016-2018)
- NIH NIDCD R03DC013862 (results); 2015-2018
- NIH NIAID R21AI137484 (results); 2018-2020
- Cystic Fibrosis Foundation LEER16G0 (2016-2018), LEE19G0 (2019-2021), GOPALL19F0 (2019-2021)
Resources and Equipment:
General Molecular Biology, Biochemistry, and Cell Biology Equipment
- Centrifuges: Fisher Marathon Micro A, Eppendorf 5415C, Eppendrof 5402 refrigerated, Sorvall ST-8, Savant Speed Vac
- General: Heatblocks, stir plates, balances, shakers, water bath, vortexers, etc.
- DNA and protein electrophoresis and western blotting equipment
- Genesys UV/Vis spectrophotometer
- Applied BioSystems QuantStudio 5 Real-Time multi-color 384 well block real time PCR system
- BioRad MyiQ single color real-time PCR detection system
- BioRad GelDoc Imaging system
- 4°C refrigerators, -20°C and -80°C freezers
- Thermo Scientific MaxQ4450 incubated shaker
- VWR Incu-Line and Fisher Isotemp bacteria incubators
- Bead tissue lyser
- 2 Thermo Scientific Forma 370 Stericycle CO2 mammalian cell incubators
- 2 Forma water-jacketed incubators
- Eppendorf refrigerated centrifuge
- Forma Tissue Culture Hood
- Custom configured in collaboration with Olympus. Housed in dedicated microscopy suite within the lab
- Olympus IX-83 two-deck inverted microscope
- 4x, 10x, 20x, 30x (silicone), 40x (LWD), 60x (oil) objectives
- Differential interference contrast
- Z-drift compensation (ZDC) laser focusing system (Olympus)
- Lambda 10-2 dual filter wheels for ratiometric imaging with Smart Shutter (Sutter Instruments)
- Filters for DAPI/SPQ, Fura-2, BCECF, FITC/GFP/Fluo-4/DAF-FM, TRITC/Cy3, CFP/YFP FRET/cameleon, SNARF (Chroma Technologies)
- Lambda LS 300 W Xe arc lamp (Sutter Instruments)
- XCite LED 120 Boost LED light source
- Olympus DSU spinning disk confocal with filters for DAPI, FITC, TRITC, Cy5 (Semrock), SNARF dual emission, and GFP/mCherry FRET
- Automated programmable stage (Prior)
- Hamamatsu ORCA-Flash4.0 sCMOS camera (2Kx2K; 30 fps at full frame)
- Tokai Hit (WSKM-F1) temperature-controlled and CO2-controlled incubation system for long-term time-lapse imaging of multi-well plates, including Neco control software and objective heater
- Vibration isolation table (TMC)
- Heated (TC-344C), 6-channel perfusion system (Warner Instruments) for 35 mm glass bottom dishes (RC-37F) and coverslips (RC-26) with pinch valves
- Peristaltic pump for perfusion
- Bioptechs Delta-T heated chamber with perfusion lid and objective heater
- Imaging acquisition and analysis workstation (Dell Precision 7910 with Intel Xeon E5-2650 v3 processor, 32 GB RAM, 2x 512 GB SATA solid state drives, NVIDIA Quadro K2200 4GB GPU) with Metamorph Advanced for Olympus, MetaMorph NX 2.0, and MetaFluor for Olympus ratiometric imaging software, ImageJ, FIJI, Icy
- Uses: Live-cell imaging of furs-2, fluo-4, DAF-FM, SNARF, BCECF, genetic GFP-based indicators, longer term imaging (5% CO2, 37 °C), 4-color immunofluorescence
- Nikon TS100 inverted microscope
- Phase contrast 10x and 100x objectives, 10x, 40x (LWD), 60x (LWD) PlanFluor objectives, 20x PlanApo objective
- 50W HBO Hg Arc Lamp
- XCite 110 LED light source with instant on/off for shuttering
- GFP/DAF-FM/Fluo-4 filter set; Texas Red filter set
- Photometric's Cool Snap HQ 12-bit cooled CCD camera
- QImaging Retiga R1 USB3 cooled CCD camera
- Color camera for histology
- Micromanager (µManager) software for acquisition
- Uses: Live cell imaging of GFP, calcium (fluo-4), nitric oxide (DAF-FM), green or red GECO, etc.
Nikon Imaging System ("Big Nikon")
- Nikon Eclipse TE2000U inverted microscope
- Prior Lumen Pro 2000S light source
- Retina R6 CCD camera
- Sutter excitation and emission filter wheels, smart shutter
- Micromanager (µManager) software for acquisition
- DAPI, CFP, GFP, YFP, TRITC, TexasRed, filters
- Uses: Live cell imaging of GFP, calcium (fluo-4), nitric oxide (DAF-FM), green or red GECO, etc.; CFY/YFP FRET biosensors
- Absorbance
- Top and bottom fluorescence with monochrometer excitation and monochrometer/filter fusion optics
- High resolution area scan
- Enhanced PMT
- Incubation heating with carbon dioxide and oxygen control (including hypoxia) for real-time, live cell measurements
- Two channel injection system
- NanoQuant Plate for DNA/RNA quantification
- SparkControl Magellan software for kinetic analyses
- Uses: Endpoint assays (ELISA); DNA-RNA quantification; Real-time, live cell imaging of proliferation (XTT), mitochondrial membrane potential (JC-1), H2O2 production (amplex red), calcium (fluo-4), etc.
- Cooled CCD, 6 megapixels
- Excitation options: Trans-UV (302 nm excitation), Epi-white, Trans-white, Trans-blue (450−490 nm ex), Epi-blue (460−490 nm ex), Epi-green (520−545 nm ex), Epi-red (625−650 nm ex), Epi-far red (650−675 nm excitation), Epi-IR (755−777 nm ex)
- Emission options: 590/110 nm filter for protein and DNA gel and chemiluminescence, 518–546 nm for blue-excitable fluorophores and stains, 577–613 nm filter for green-excitable fluorophores and stains, 675–725 nm for red-excitable fluorophores and stains, 700–730 nm filter for far red-excitable fluorophores and stains, 813–860 nm for near IR-excitable fluorophores and stains
- World Precision Instruments DVC-1000 and Warner EC-825 dual channel epithelial voltage clamp amplifiers
- Various Ussing chambers and accessories
