Amitabha Mukhopadhyay
Research interest.
Mechano-transduction in
circulating innate immune cells
Immediate early immune responder, neutrophils, need a final mechanical stimulation during its passage through restrictive endothelial junction – before entering lung (or other) tissue. Without endothelium derived mechano-sensation, such as in acute respiratory distress syndromes or pneumonia, caused due to the relaxation of endothelial adherens junction, neutrophils do not reach to the activation threshold. Thus, immune responses become dampened, leading to prolonged recovery or mortality .
I focus on understanding the mechanism of cellular mechanical signal reception, relay, mechano-chemical coupling processes and resultant cellular responses to pathogen elimination or inflammation.
Experience
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Post-doctoral research fellow.
Biophysical Dynamics; Dept. of Biochemistry & Molecular Biology.
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Mechanism of light signal relay from photo-sensing LOV domain to the effector kinase domain – in signaling photo-receptor Phototropin, applying serial pump-probe femto-second crystallography.
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Molecular cloning.
Protein engineering.
Protein purification.
Protein crystallization.
Single crystal spectroscopy
UV/Vis spectroscopy.
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Post-doctoral research fellow.
Feinberg School of Medicine, Dept. of Pharmacology.
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Mechanical signal relay through TRP channels, located in cilia.
Ciliopathy and polycystic kidney disease.
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Large-scale mutagenesis.
Lentiviral transgenesis and stable cell-line generation.
Immortalization of patient and animal derived cells.
Viral injection in mouse kidney and development of polycystic kidney disease model.
Calcium-calmodulin signaling through TRPP2 channels.
Peptide mass fingerprinting; mass spectrometry.
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Research Instructor.
College of Medicine; Pharmacology and Regenerative Medicine; Center for Lung and Vascular Biology.
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Mechano-transduction in neutrophils.
Translational mechano-immuno-modulatory therapeutics for acute pulmonary infection and lung disease.
Innate immunology.
Targeting of disease associated neutrophils and innate immune cells by nanoparticles.
Neutrophil functional subtype categorization.
Assessment of tightness of vascular junction and it’s implication on (i) vascular leakiness, (ii) barrier protection and (iii) inflammation resolution – by altering CHFR-mediated ubiquitination and degration of VE-cadherin.
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Applying mechanical shear force to neutrophils and other innate immune cells using microfluidic devices, rheometer, transwell pores and genetic mice models.
Generation of acute lung infection models in mice using LPS inhalation, bacterial delivery through trachea and retro-orbital path; generation of bleomycin-induced pulmonary fibrosis in mouse.
Isolation of neutrophil and macrophages from mouse lung and bone-marrow.
In vivo fluorescent labeling of immune cells by retro-orbital delivery of antibodies.
Targeted elimination and enrichment of immune cells in vivo upon drug-loaded albumin nanoparticle delivery through intra-venous and intra-tracheal path in mice.
Assessment of bacterial killing ability of phagocytic cells, in vivo and in vitro.
Multi-color flow cytometry, sorting, mass-cytometry (Cytof), spectral flow cytometry.
Generation of acute lung infection mice models.
Generation of mice model with engineered vascular-junction tension.
Restrictiveness of endothelial junction in vivo and in vitro.
Bulk RNAseq data generation, interpretation and validation by real-time PCR and flow cytometry.
Biochemical protein-protein interaction.
IP coupled mass spectrometry and di-gly mass spectrometry for identification and validation of Ubiquitin binding sites of VE-cadherin.
Imaging in microfluidic device.
Calcium ratiometric imaging and analysis.
Intra-vital imaging in mice cremaster muscle.
Fluorescence confocal life-time imaging ; transmission electron microscopy.
Graphic design, statistical analysis, data plotting.
Education
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ACTREC (Advanced Center for Treatment Research & Education in Cancer).
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Role of 14-3-3 protein in cell cycle check-point (G2/M) regulation.
Centrosome duplication.
Relationship between centrosome hyper-duplication (amplification), override of checkpoint function and neoplastic progression.
Identifying mechanism to kill aggressive metastatic cancer cells and reduce tumor burden by altering centrosome amplification and inducing mitotic catastrophe.
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Cloning, vector designing & engineering, tissue culture, transfection, ectopic protein expression, immuno-precipitation (IP), sucrose density gradient, 2D-electrophoresis, IP - mass spectrometry, peptide-mass fingerprinting, fluorescence confocal imaging, FRET live imaging, generation of mouse tumor xenograft models, immuno-histochemistry, karyotyping, cell cycle analysis by flow cytometry.
Generation of expression cDNA library, bio-panning of filamentous phage antibody library, expression and purification of sc-FV (single-chain antibody) for hypo-glycosylated MUC1 peptide by FPLC , ELISA to test the highest binding affinity (kd) scFVs.