The Rabinovitch/Bland Lab

 Ongoing Research in the laboratory of Marlene Rabinovitch, M.D.

DNA DAMAGE RESPONSE AND INFLAMMATION, APOPTOSIS AND ABERRANT PROLIFERATION

GPCRs, AMPHETAMINE AND PULMONARY HYPERTENSION

AUTOIMMUNITY, INFLAMMATION AND PULMONARY HYPERTENSION

TRANSLATIONAL RESEARCH: ELAFIN AS A THERAPY FOR PULMONARY HYPERTENSION AND CARDIOVASCULAR DISEASE

CELL EXTRACELLULAR MATRIX INTERACTIONS AND REGULATION OF ELASTIN ASSEMBLY IN VASCULAR DEVELOPMENT AND PATHOLOGY

GENOMICS AND METABOLOMICS IN PULMONARY HYPERTENSION

GENETICS, CONGENTAL HEART DISEASE AND PULMONARY VASCULAR PATHOLOG

INDUCED PLURIPOTENT STEM CELLS AND PULMONARY HYPERTENSION

GENETICALLY MODIFIED MURINE MODELS OF DISEASE

 

DNA DAMAGE RESPONSE AND INFLAMMATION, APOPTOSIS AND ABERRANT PROLIFERATION

Caiyun (Grace) Li, PhD, postdoctoral fellow, in performing immunoprecipitation followed by mass spectrometry (IP-MS), to identify novel PPARg interactors, discovered a role for PPARg in DNA damage sensing and repair.  This role appears critical in preventing the abnormal proliferation and transformation of vascular cells in response to environmental perturbations that could otherwise lead to pulmonary hypertension specifically, and cardiovascular diseases in general.  This is consistent with more evidence of DNA damage in tissues from patients with PAH that have a reduced BMPR2-PPARg axis.                       

 

 BMPR2:PPARgamma axis in PAH  

 

 

 Jan Hennigs, MD, postdoctoral fellow is pursuing the role of p53 in the vascular DNA damage response.

Isabel Diebold, MD, postdoctoral fellow is pursuing the role of mitochondrial DNA damage in response to failure to activate the BMPR2-PPARg axis.

Immunodflourescence image of a section from a lung of a patient with a BMPR2 mutation

Immunofluorescence image analysis of a section from a lung of a patient with a BMPR2 mutation.  Endothelial cells are stained with von Willebrand factor (green) and nuclei are stained with DAPI (blue). Red shows the protein associated with DNA damage foci, phospho gH2AX.  The image on the right is taken with a x40 magnification.

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GPCRs, AMPHETAMINE AND PULMONARY HYPERTENSION

Pin-I Chen, PhD, postdoctoral fellow focuses on a sub-category of pulmonary hypertension associated with abuse of amphetamine derivatives. She has elucidated the signaling mechanism whereby these compounds subvert cell survival, and amplify inflammatory responses and DNA damage. She is also establishing an animal model with chronic infusion of amphetamine.

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A schematic model showing amphetamine-induced GPCR signaling cross-talk with endothelial cell survival pathways and inflammatory responses via recruitment of β-arrestin platforms.

Endothelial cells treated with Dox and Amphetamine Red:  53BP1, the protein recruited to a focus of DNA damage. Green: phospho gammaH2AX an indicator of DNA damage . Yellow: overlap of 53BP1 and  phospho gammaH2AX.
The micrographs demonstrate that  DNA damage in endothelial cells induced by  Doxorubicin (Dox) is amplified by amphetamine (AMPH).

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AUTOIMMUNITY, INFLAMMATION AND PULMONARY HYPERTENSION

Toshie Saito, MD, postdoctoral fellow, is discovering novel autoantigens, that link viral infection and autoimmunity with pulmonary hypertension. This follows from work by a former postdoctoral fellow, Hirofumi Sawada, MD PhD, who showed that enhanced production of  GMCSF related to dysfunction of BMPR2 plays a critical role In the recruitment of macrophages that may be the source of the autoantigens.

 Lung sections showing GM-CSFR+ monocytes and EC progenitors

 

 

GM-CSF-Receptor (GMCSFR-a)+ Monocytes and EC Progenitors: 
In a control lung, immunoreactivity is not apparent

 

 

 

 

In the IPAH patient, GMCSF receptor was detected in thickened intima, colocalizing with macrophage marker CD68. In a plexiform lesion from the same patient, the GMCSF receptor colocalized with CD34, the endothelial progenitor marker. Our data suggest that GMCSF can recruit monocytes and endothelial cell progenitors to vascular lesions.

Microfluidics-based high-throughput single cell transcriptional analysis of single cells isolated and sorted by FACS. Cells were obtained from lung tissues explanted from an IPAH patient that underwent lung transplantation.CD31+ GMCSFRalpha+ cells from IPAH lung tissue - gene expression

Increased expression is designated by yellow color, decreased expression by blue. Grey indicates unchanged expression. Two subpopulations of cells were identified that expressed the receptor for GMCSF, one expressed genes of endothelial lineage (left panel), the other expressed genes of monocyte macrophage lingeage (right panel).

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TRANSLATIONAL RESEARCH: ELAFIN AS A THERAPY FOR PULMONARY HYPERTENSION AND CARDIOVASCULAR DISEASE

Nils Nickel, MD, postdoctoral fellow investigates the role of elafin, the elastase inhibitor and immunomodulatory agent in regenerating lost microvessels as well as reversing pathological features of pulmonary hypertension.

Schema of the role of elastase in inflammation, SMC proliferation and EC dysfunction

 

 

 

 

 

 

 

  

Elafin treatment reverses occluded lesions in the lung that show increased alpha actin positive staining cells compromising the vessel lumen.

Elafin treatment reverses occluded lesions in the lung that show increased alpha actin positive staining cells compromising the vessel lumen.

 

 

 

 

 

 

 

 

 


 

 

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CELL EXTRACELLULAR MATRIX INTERACTIONS AND REGULATION OF ELASTIN ASSEMBLY IN VASCULAR DEVELOPMENT AND PATHOLOGY

Nancy Ferreira Tojais, PhD, postdoctoral fellow investigates the factors that cause susceptibility of elastin to degradation and has discovered the role of BMPR2 signaling in regulating appropriate scaffolding of the elastin fiber via fibrillin1 and the role of the proteoglycan decorin in restricting elastin assembly in development and impairing elastin repair in disease. Figure shows that BMP stimulates fluorescent elastin fiber formation in lung fibroblasts.

Figure showing that BMP stimulates fluorescent elastin fiber formation in lung fibroblasts.

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GENOMICS AND METABOLOMICS IN PULMONARY HYPERTENSION

Rachel Hopper, MD, postdoctoral fellow uses induced induced pluripotent stem cells to study the genetic association of  congenital heart disease and pulmonary hypertension.  Her work also follows from RNA Seq analysis carried out by former postdoctoral fellow, Chris Rhodes, PhD that demonstrated gene expression changes in endothelial cells from patients with pulmonary hypertension. One of those genes being studied by Dr. Hopper is a transforming factor HMGA1. Schema of genomic findings

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GENETICS, CONGENTAL HEART DISEASE AND PULMONARY VASCULAR PATHOLOGY

Nathaly Sweeney, MD, Academic Research Staff is addressing the genetic basis for the changes in vascular morphology  observed  in  patients with familial Tetralogy of Fallot.  Tetralogy of Fallot (TOF).  This is the number one cyanotic congenital heart disease and is seen in 3.3 of 10,000 live births.  TOF is associated with abnormal development of the pulmonary valve and in most severe cases the patient’s develop extensive aorto-pulmonary collaterals (MAPCAs), frequently with obstruction to pulmonary flow.  

schema of genetics of a family with DiGeorge syndrome

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INDUCED PLURIPOTENT STEM CELLS AND PULMONARY HYPERTENSION

Silin Sa, PhD, LSRA, is developing and improving strategies to differentiate endothelial cells from iPSCs.  Lung, blood and skin-derived iPSCs from pulmonary hypertension patients who undergo lung transplantation are compared to those from control (unused donor) lungs.

iPSC trasformed to ECs

 Aiqin Cao, PhD, LSRA, is addressing the functional significance of aberrant collagen IV and ephrin A1 signaling in endothelial cells, following up on the RNA-Seq observations made by former post-doctoral fellow Chris Rhodes, PhD.

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GENETICALLY MODIFIED MURINE MODELS OF DISEASE

Lingli Wang, MD, Lab Manager is creating genetically modified mice to conditionally delete and fate map cells to be used in investigating pulmonary hypertension pathophysiology.  She has investigated the role of BMPR2 in the smooth muscle cells of the vessel wall and found that these cells are hypocontractile and hyperproliferative owing to impaired RhoA and heightened beta catenin.  

Impaired contractility of BMPR2 knock-out smooth muscle cells

 

 

 

 

 

 

 

 

 

 

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