UNLOCKING THE GENETIC CODE TO THE HEART AND LUNG DISEASES.

Every 4 minutes, a baby is born with a birth defect. Rapid advances in next-generation sequencing have identified candidate gene variants. However, many variants remain of unknown significance (VUS) and a huge burden on birth defect diagnosis. Clinicians cannot use VUS in decision-making and, therefore, cannot 1) assess patients’ risk of complications, 2) provide the best clinical therapy, and 3) provide recurrence risk counseling for parents. Our long-term goal is to improve clinical outcomes for patients with birth defects by increasing the rate of genetic diagnosis.

RESEARCH

VISION

To be at the forefront of the cutting-edge field of functional genomics, enabling scientific curiosity, excellence in research, and groundbreaking discoveries to improve the clinical outcomes for babies born with heart and lung defects.

RESEARCH APPROACH

We use in vivo frog and mouse models and stem cell-based 2D and 3D (organoid) models to functionally characterize VUS in birth defects. We employ an interdisciplinary approach combining CRISPR/Cas9 genome editing, super-resolution microscopy, mechanobiology, single-cell/nuclei transcriptomics, and computational modeling.

PROJECTS

Discovering new genes and signaling pathways essential in cilia assembly and function.

Cilia are microtubule-based organelles present on almost all the cells in the human body and serve multiple roles during development and homeostasis. Defects in the assembly or function of cilia can lead to an array of diseases from heart, respiratory to skeletal and metabolic diseases. We have projects focused on identifying novel genes and signaling pathways essential in cilia assembly and function in the context of tissue and organ development.

Uncovering the mechanochemical gene regulatory networks (GRNs) in lung development and function.

Mechanical forces play a critical role in respiratory physiology. Our lungs continuously expand and compress as we breathe, exerting mechanical tension on the lung epithelium. This mechanical tension is translated into chemical signals essential for the differentiation of airway and lung epithelial cells (multiciliated cells, mucus-secreting cells, ionocytes, etc.). We have projects focused on uncovering mechanochemical GRNs essential for proper cell differentiation and maintaining epithelium integrity in the airway and lungs.

High-throughput translational genomics of pulmonary disorders.

Respiratory diseases (RDs) are caused by defects in the structure or function of motile cilia and multiciliated cells in the airway and lungs. These patients are at risk of recurrent respiratory infections, bronchiectasis, or even respiratory failure. About half of all these patients also develop cardiac defects. RDs are often challenging to diagnose due to the lack of a “gold standard” diagnostic tool and clinically variable presentation. Genetic testing has the potential to address these shortcomings and yet, the rate of genetic diagnosis remains very low. We are addressing this shortcoming by developing and implementing high-throughput functional genomics models of frog embryos and human airway cell culture to classify variants robustly and at an unprecedented rate. Our research has an immediate positive global impact by increasing the rate of genetic diagnosis in RD patients in a clinically consequential time frame.

Bench to Bedside PRECISION MEDICINE program for congenital heart disease (CHD).

CHD is a leading cause of infant death. Nearly 1 in 100 babies are born with a heart defect, and about 25% have critical CHD. CHD remains a leading cause of infant death because phenotypically similar patients can have dramatically different outcomes. Therefore, we need clinical treatment driven by the genotype instead of a phenotype to treat these patients effectively. The NGS has identified candidate disease variants rapidly and cost-effectively. However, the yield of current clinical genetic testing is less than 30%, and determining the pathogenicity of the remaining candidate genes remains a major obstacle. Dr. Kulkarni has established a Bench-to-Bedside (B2B) CHD program at UVA Children’s to address this challenge. Our B2B project brings together basic science and clinical expertise in functional genomics human genome analysis , genetic counseling, pediatric cardiology, and pediatric pulmonology to facilitate patient-driven research. Our research directly helps patients and families with aggressive management of clinical complications, including the risk of developing life-threatening conditions like respiratory problems and hydrocephalus. Further, our research enables recurrence risk counseling and reproductive decision-making for parents.

MEET THE TEAM

  • Saurabh Kulkarni

    Assistant Professor

  • Chris Courtney

    Lab manager

  • Venkatraman Rao

    Senior Postdoctoral Associate

  • Vani Narayan

    Postdoctoral Associate

  • Vignesh Aravind

    Postdoctoral Associate

  • Angelo Arrigo

    Graduate Student

  • Victoria Hua

    Undergraduate Researcher

  • Kranti Kaur

    Undergraduate Researcher

  • Aarathi Manchikalapudi

    Undergraduate Researcher

  • Krishna Girish Kumar

    Undergraduate Researcher

RESOURCES

MICROSCOPY

NIKON AX-R confocal: This new state-of-the-art microscope is equipped with 8K Galvo and 2K resonant scanners, 4 solid-state laser lines (405, 488, 561, and 640), and 4 PMT detectors with 2 GaAsP detectors. The software capabilities include advanced 2D tracking of the cells or organelles, 3D measurement analysis, deconvolution, and tile scanning.

The LEICA SP8 confocal microscope: The microscope is built off a DMi8 inverted research microscope and comes equipped with a white light laser scan head that is tunable within the range of 470-670nm with up to eight laser lines; a 405nm laser; a filter-free spectral detector for up to five individually regulatable channels.

NIKON SMZ1270 Stereomicroscope: The lab has two sets of stereomicroscopes equipped with a camera and computer for high-speed imaging.

OTHER

Mechanical stretcher:  Lab is equipped with a state-of-the-art custom-made radial mechanical stretcher controlled by the software and can stretch and compress the tissue. It also enables live imaging of cellular events like the apical expansion of MCCs and centriole amplification using a confocal microscope while applying mechanical forces.

Microinjection room: Four sets of microinjection stations that include picospritzers, micromanipulators, needle holders, and stereomicroscopes.

TECHNIQUES

Genome manipulation: CRISPR-Cas9 and morpholino oligo in frog embryos, skin organoids, and cell culture

Imaging: Confocal and Super-resolution microscopy, live imaging, Scanning Electron Microscopy (SEM), Transmission electron microscopy (TEM), and Electron Tomography (ET)

Cell culture: Generation of stem cell explants and organoids

Transcriptomics: Single cell and single nuclei sequencing

Computational biology: Mathematical modeling and machine learning

Mechanobiology: Biomechanical manipulations of cells and skin organoids

UVA FACILITIES

Advanced Microscopy Facility

The Keck Center for Cellular Imaging

The Molecular Electron Microscopy Facility

Genome Analysis and Technology Core

AFFILIATIONS

DEVELOPMENTAL GENOMICS CENTER

The Developmental Genomics Center at UVA will bridge developmental biologists with genomic and clinical translational scientists across grounds and with nearby Inova Health System and the NIH NICHD. The Center aims to integrate genomic technologies and next-generation sequencing datasets from human and animal model systems to address cutting-edge research questions in cell and developmental biology.

CENTER FOR MEMBRANE AND CELL PHYSIOLOGY

UVA’s Center for Membrane and Cell Physiology strives to understand fundamental biological processes at the highest possible spatial and temporal resolution. Our ultimate goal is to use high-end imaging, structural, biophysical, and biological and chemical probe technologies to make impactful discoveries on understanding the causes, development, and cures of diseases ranging from cardiovascular to cancer to neurological and infectious diseases.

CHILD HEALTH RESEARCH CENTER

The center's mission is to support scientists engaged in basic and clinical research to discover innovative therapies for childhood diseases. We bridge the gap between the laboratory and the bedside with cutting-edge research that improves the lives of children.

GRADUATE PROGRAMS AND TRAINING GRANTS

The Kulkarni lab is affiliated with postdoctoral and graduate T-32 training grants supported by the National Institutes of Health at the University of Virginia. Post-doctoral and Ph.D. applicants who are interested in the Kulkarni lab may be good candidates for applying for financial support from these training grants.

CONTACT

Kulkarni lab

Department of Cell Biology

Department of Biology,

University of Virginia

Charlottesville, VA 22908

Email: sk4xq@virginia.edu

Phone No: +14342976833