Sustained delivery of insulin-like growth factor-1/hepatocyte growth factor stimulates endogenous cardiac repair in the chronic infarcted pig heart

Abstract

Activation of endogenous cardiac stem/progenitor cells (eCSCs) can improve cardiac repair after acute myocardial infarction. We studied whether the in situ activation of eCSCs by insulin-like growth factor 1 (IGF-1) and hepatocyte growth factor (HGF) could be increased using a newly developed hydrogel in chronic myocardial infarction (MI). One-month post-MI pigs underwent NOGA-guided intramyocardial injections of IGF-1/HGF (GF: both 0.5 μg/mL, n = 5) or IGF-1/HGF incorporated in UPy hydrogel (UPy-GF; both 0.5 μg/mL, n = 5). UPy hydrogel without added growth factors was administered to four control (CTRL) pigs. Left ventricular ejection fraction was increased in the UPy-GF and GF animals compared to CTRLs. UPy-GF delivery reduced pathological hypertrophy, led to the formation of new, small cardiomyocytes, and increased capillarization. The eCSC population was increased almost fourfold in the border zone of the UPy-GF-treated hearts compared to CTRL hearts. These results show that IGF-1/HGF therapy led to an improved cardiac function in chronic MI and that effect size could be further increased by using UPy hydrogel.

Fig. 1

Study design. a Schematic study design showing the targeted intramyocardial delivery in the MI border zone of empty UPy-hydrogel as control (1, CTRL); IGF-1/HGF dissolved in saline, denoted as GF (2); or UPy hydrogel with IGF-1/HGF, denoted as UPy-GF (3)

Fig. 2

UPy-IGF-1/HGF therapy improves cardiac function in chronic MI. a, b LV end-diastolic and end-systolic volumes at baseline, 1 month after injection, and the relative change between both time points. c LV ejection fraction. d FAS measured by 2DE at the level of the papillary muscles. e Preload recruitable stroke work (PRSW) measured by intracardiac pressure–volume loop recordings. *p < 0.05 (vs. CTRL). All data are the mean ± SD; n = 3, 5, and 5 for CTRL, GF, and UPy-GF, respectively

Fig. 3

IGF-1/HGF treatment reduced pathological hypertrophy in the MI border zone. a, b Representative MI border zone sections (hematoxylin and eosin staining) showing adverse cardiac hypertrophy in the control-treated animals (a), which was not observed in the UPy-GF-treated animals (b). c–h Picric Sirius red staining in bright-field images (c–e) and under polarized light (f–h) showing extensive scar tissue in all groups depicted as red staining in bright-field microscopy. Under polarized light, color depended on the collagen fiber density (yellow for higher intensity, green for lower intensity). In both growth factor-treated groups, small myocardial islands were visible in the infarct area (see arrowheads). Quantification of cardiomyocyte diameter in the MI border zone (i) and fibrosis (j). *p < 0.05 (vs. CTRL). All data are the mean ± SD; n = 3, 4, and 5 for CTRL, GF, and UPy-GF, respectively. MI myocardial infarction

Fig. 4

IGF-1/HGF administration leads to the formation of new cardiac myocytes. a, b Expression of cellular proliferation marker Ki67 (green) showed increased proliferation index of cells (arrowheads) in the UPy-GF-treated animals compared to CTRL. c, d Increased newly formed Ki67^pos (green) cardiomyocytes (arrowheads, asterisk; see inset) after GF treatment compared to CTRL in the peri-infarct/border zone. e Ki67^pos cardiac myocytes were smaller than the quiescent Ki67^neg cardiomyocyte fraction, indicative of their immature, newly formed nature. *p < 0.05 (vs. CTRL); ^† p < 0.05 (vs. Ki67^neg cardiac myocytes). All data are the mean ± SD; n = 3, 4, and 5 for CTRL, GF, and UPy-GF, respectively

Fig. 5

IGF-1/HGF leads to increased capillerization and reduces microvascular resistance. a Staining for von Willebrand factor (vWF) shows small capillary structures (red arrowheads, asterisk; see inset) in the border zone of the UPy-GF-treated heart. b Number of capillaries in the peri-infarct/border zone area. c Relative change, compared to baseline, in simultaneously measured intracoronary pressure and flow-derived hyperemic microvascular resistance (HMR). *p < 0.05 (vs. CTRL). All data are the mean ± SD; n = 3, 4, and 5 for CTRL, GF, and UPy-GF, respectively

Fig. 6

IGF-1/HGF treatment increases the epCSC compartment and drives their cardiac commitment in chronic MI. a The infarct area harbors various cell types, such as (i) c-kit^pos CD45^neg epCSCs, (ii) c-kit^neg CD45^pos cells, or (iii) c-kit^pos CD45^pos cells (including mast cells). b Endogenous epCSCs were a morphologically distinct subset of small cells showing perinuclear expression of c-kit (green, arrowheads) and negative for CD45. c Quantification of epCSCs in the peri-infarct/border and infarct zone. d A c-kit^pos (green) myogenic progenitor (arrowhead, asterisk; see inset) expressing the early cardiac transcription factor, Nkx2.5 (white). e Quantification of Nkx2.5^pos epCSCs in the peri-infarct/border and infarct zone. *p < 0.05 (vs. CTRL). All data are the mean ± SD; n = 3, 4, and 5 for CTRL, GF, and UPy-GF, respectively. f Some c-kit^pos epCSCs also expressed the transcription factor ETS-1 (arrowhead, asterisk; see inset)