3.12 Preclinical research on EPCs in PAH
Because EPCs have numerous capacities such as proliferation, migration,
and adhesion to protect the normal structure of ECs and PASMCs,
scientists use EPCs to attenuate and reverse PAH. Zhao et al. injected
fluorescently labeled EPCs into monocrotaline (MCT)-induced PAH mice,
the EPCs integrated into the distal pulmonary artery endothelium and
limited the progression of PAH through differentiation to mature ECs and
alleviation of neointima formation(Yang et al., 2013). Zhao et al. also
detected EPC-derived proangiogenic growth factors (such as IL-8 and
adrenomedullin) in the adjacent area of EPCs through autocrine and
paracrine methodology. Zheng et al. found that EVs of EPCs transport
mRNA to ECs, and express a series of proteins such as chemokines,
pathway proteins, and growth factors to enhance EC proliferation and
migration and decrease angiogenesis(Deregibus et al., 2007; Ingram et
al., 2004; X. Li et al., 2016). Zhao et al. used EPCs to treat
monocrotaline (MCT)-induced PAH mice at two different time points to
explore the effect of EPCs on different disease stages. The results
seemed inspiring, as compared with the MCT group, the hemodynamic
improvement in both groups was significant, but both treatment groups
still had a higher mean pulmonary arterial pressure (mPAP) than the
control group, indicating that EPCs can prevent but cannot reverse the
occurrence of PAH. Zhao et al. also investigated the microstructure of
the pulmonary artery under EPC therapy. They stained the pulmonary
artery with fluorescent microspheres and alpha-smooth muscle actin
(α-sma) to visualize the structure of PASMCs and microvasculature
perfusion. The results showed that the 21-day and 35-day groups
dramatically increased their microvascular perfusion and
microvasculature after EPC treatment(Y. D. Zhao et al., 2005).
As a nonoral therapy, use frequency is also an important indicator of
feasibility. Therefore, Harper et al. and Zhou et al. investigated the
stay time and stay location after EPC transplantation. Harper et al.
selected cells expressing CD34, CD306, CD146, and CD45 by
fluorescence-activated cell sorting (FACS) and transfected adenoviral
vectors carrying the luciferase and GFP reporter gene. The
immunofluorescence results showed that at 1 h and 7 h after EPC
transplantation, most of the luminescence was located in the chest
region, and there was little or no luminescence in other areas, the
luciferase results were same. Compared with the control group, the
concentration of EPCs in the lung was two times higher than that in the
spleen. This shows that EPCs tend to aggregate in damaged endothelial
tissues, but rarely aggregate in integrated tissues. According to the
above evidence, EPC therapy seems to be a targeting therapy that has
little impact on other organs, at least in concentration
relationships(Harper et al., 2019; Zhou et al., 2013). However, the
experiment by Zhou et al. produced some opposite results. He found that
the retention rate of EPCs was still nearly 50% 25 days after
transplantation(Zhou et al., 2013). These contrary results were caused
by different screening conditions during FACS.
In order to improve the effect of EPC therapy, scientists use gene
editing technology. Wei et al. used Ad.CMV-human endothelialNO synthase
enzymes (Ad.CMV-heNOS), Ad.CMV, or Ad.CMV-enhanced green fluorescent
protein (Ad.CMV-EGFP) to infect EPCs for 72 h. Two weeks after the
injection, the systolic arterial pressure (ASP) of mice did not
significantly change, but the systolic pulmonary artery pressure (sPAP)
significantly decreased in both the simple EPC group and the heNOS-EPC
group. Furthermore, the heNOS-EPC group was significantly lower than the
vector-EPC-treated group and the normal EPC-treated group. The histology
results also showed the same trend, with a significant reduction in the
number of muscular pulmonary arteries and thickness of the muscular
coat(Wei et al., 2013).
Cao et al. selected the human hypoxia inducible factor-1 alpha (hHIF-1α)
gene and transfected it into EPCs. Compared with the EPC group and blank
group, hHIF-1-EPCs significantly reversed the vascular remodeling, and
decreased the sPAP, mPAP, and right ventricular/left ventricular +
septum (RV/LV+S). Xu et al. chose to disturb the metabolic process of
the pulmonary artery to control the progression of PAH. They used
lentiviral vectors to inhibit the expression of E2F transcription factor
1 (E2F1) and subsequently increase oxidative metabolism, endothelial
differentiation, vascular repair, and decrease the pyruvate
dehydrogenase kinase 4/2 (PDK4/2) expression. The metabolic level was
measured by the oxygen consumption rate (OCR), extracellular
acidification rate (ECAR), and lactate level. The results showed that
E2F1-/- mice had a greater OCR, a lower ECAR, and a lower level of
lactate (which represents a lower anaerobic respiration level),
indicating that E2F1-/-EPC enhanced vascular growth, reduced infarct
size, and improved vascular function through increasing the oxygen
utilization rate of pulmonary ECs(Cao et al., 2015; S. Xu et al., 2018).
Dysfunctions of the prostacyclin (PGI) pathway are also an important
factor in the occurrence of PAH. Therefore, Zhou et al. tried to
engineer the cyclooxygenase-prostacyclin (COX-PGI) pathway to improve
the effect of EPCs. The engineered EPCs significantly attenuated the
RVSP increase, RV hypertrophy, and intimal and medial smooth muscle
layer cell proliferation, and enhanced adventitial pulmonary vessel wall
apoptosis(Zhou et al., 2013).