Conference: 2012 International PHA Conference and Scientific Sessions
Release Date: 06.22.2012
Presentation Type: Abstracts
Hatch A1, Hansmann G2, Patel R1 and Murthy S1
1. Northeastern University USA
2. Children’s Hospital Boston USA
BACKGROUND: Peripheral blood contains a subtype of circulating cells called endothelial progenitor cells (EPCs). Number of EPCs in circulating blood is inversely correlated with endothelial dysfunction, cardiovascular risk and clinical outcome. For these reasons EPC number has been suggested as a biomarker for cardiovascular disease. Typically flow cytometry is used for EPC enumeration however the ability to apply flow cytometry in routine use is limited by the number and complexity of non-automated preprocessing steps. This study proposes a rapid bedside test for selectively isolating and counting EPCs from patient blood samples in a routine manner by selectively capturing and immunostaining them within a disposable microfluidic platform.
METHODS: Peripheral blood was collected in 6 mL EDTA tubes at the 2010 PHA conference research room from healthy controls and PAH patients. 200 microliters of the blood samples were directly injected into the polymeric microfluidic chips at a flow rate of 0.6 mL/h. Following capture cells were identified and enumerated by immunofluorescent staining. Data were tabulated for cells that expressed CD34, KDR and CD31 antigens. Results were correlated with flow cytometry by immunostaining for the same three markers to verify a linear correlation between number of cells captured on the chip and number of cells counted by flow cytometry.
RESULTS: Cell numbers for EPCs (CD34+/CD31+) captured with the microfluidic device did not differ significantly between genders among the control subjects. Cell numbers were also similar among the control group and among the PAH patients even between differing types of PAH (i.e. idopathic, drug induced, connective tissue disease). However cell numbers differed greatly between the control group (n=6) and the PAH patients (n=43). Cells captured among the controls were 28.5±1.0 per chip vs. 17.0±0.4 per chip in the PAH patients. Interestingly EPC numbers were inversely correlated with age in the control group but no such correlation existed among the PAH patients.
CONCLUSIONS: The described EPC capture chip is capable of capturing a large percentage of circulating EPCs from a small blood volume with no preprocessing. Patients with idiopathic PAH, as well as those associated with drug use or connective tissue disease have about half the number of EPCs captured relative to healthy age and gender matched controls. This EPC capture chip requires only minimal blood volume and no preprocessing, and thus has the potential to become a routine diagnostic tool for risk assessment and clinical monitoring of PAH patients.