Delivery of biotinylated IGF-1 with biotinylated self-assembling peptides combined with bone marrow stem cell transplantation promotes cell therapy for myocardial infarction

Abstract

Cell therapy is a promising approach for cardiac repair. The aim of the present study was to determine the feasibility of using biotinylated insulin-like growth factor 1 (IGF-1) with biotinylated self-assembling peptides (tethered IGF-1) combined with bone marrow stem cells (BMSCs) transplantation for the treatment of heart failure. Tethered IGF-1 was synthesized and its effect on H9c2 cells was analyzed. Reverse transcription-quantitative polymerase chain reaction and western blot assays demonstrated that tethered IGF-1 did not significantly affect the expression and phosphorylation of AKT, whereas it significantly increased the expression of cardiac troponin T (P<0.01). A rabbit myocardial infarction model was constructed and rabbits were divided into four groups: Control group (no treatment), group 1 (G1; BMSC transplantation), group 2 (G2; BMSCs + non-biotinylated IGF-1) and group 3 (G3; BMSCs + tethered IGF-1). At 4 weeks after modeling, cardiac tissues were obtained for analysis. In the control group, myocardial fibers were disordered, a large number of inflammatory cells infiltrated the cardiac tissues, and apoptosis occurred in ~50% of cells. However, in G1, G2 and G3, muscle cells were well ordered, and a lesser degree of myocardial degeneration and inflammatory cell infiltration was observed. Compared with the control group, the apoptosis rates of myocardial cells in G1-G3 were significantly decreased (P<0.01). Furthermore, compared with G1 and G2, tissue morphology was improved in G3and the number of apoptotic myocardial cells was significantly decreased (P<0.01). These results suggest that treatment with tethered IGF-1 + BMSCs significantly suppresses cell apoptosis and induces the expression of cardiac maturation proteins. These findings provide a novel insight into how the delivery of tethered IGF-1 with BMSCs could potentially enhance the prognosis of patients with heart failure treatment.

Keywords: bone marrow stem cell; cardiac failure; insulin-like growth factor 1; self-assembling peptides.

Figure 1.

Tethered IGF-1 prolonged the biological effect of IGF-1.H9c2 cells were incubated with tethered IGF-1 or non-biotinylated IGF-1, respectively. (A) RT-qPCR results demonstrated that tethering IGF-1 did not affect Akt expression. (B) Western blotting revealed that AKT phosphorylation was not affected by the tethering of IGF-1. (C) Quantified results of western blotting. (D) RT-qPCR results demonstrated that the cTnT expression was increased following incubation with tethered IGF-1 compared with non-biotinylated IGF-1 for 14 days. (E) Western blotting revealed that cTnT protein was upregulated by tethered IGF-1 compared with non-biotinylated IGF-1. GAPDH was used as the internal control. (F) Quantified results of western blotting. **P<0.01 vs. tethered IGF-1. IGF-1, insulin-like growth factor 1; tethered IGF-1, biotinylated IGF-1 with biotinylated self-assembling peptides; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; cTnT, cardiac troponin T; p, phosphorylated.

Figure 2.

Tethered IGF-1 with BMSCs ameliorated the pathological morphology of rabbit MI. Rabbit MI models were constructed and injected with various BMSC treatments: (A) Control; (B) G1; (C) G2; and (D) G3. Heart tissues were extracted after 4 weeks and subjected to hematoxylin and eosin staining. Among the MI groups, tissues from G3 rabbits were observed to have the most improved morphology. Scale bar, 100 µm. IGF-1, insulin-like growth factor 1; tethered IGF-1, biotinylated IGF-1 with biotinylated self-assembling peptides; BMSCs, bone marrow stem cells; MI, myocardial infarction; G1, group 1 (BMSC transplantation); G2, group 2 (BMSCs + non-biotinylated IGF-1); G3, group 3 (BMSCs + tethered IGF-1).

Figure 3.

Tethered IGF-1 with BMSCs suppressed cardiomyocyte apoptosis in vivo. Rabbit MI models were injected with different BMSC treatments and after 4 weeks the heart tissues were isolated. (A) TUNEL staining in the (a) control, (b) G1, (c) G2 and (d) G3 groups indicated that tethered IGF-1 + BMSCs inhibited cell apoptosis. Scale bar, 100 µm. (B) Quantified AI. **P<0.01 vs. control group and ^##P<0.01 vs. G1 and G2. IGF-1, insulin-like growth factor 1; tethered IGF-1, biotinylated IGF-1 with biotinylated self-assembling peptides; BMSCs, bone marrow stem cells; MI, myocardial infarction; AI, apoptosis index; G1, group 1 (BMSC transplantation); G2, group 2 (BMSCs + non-biotinylated IGF-1); G3, group 3 (BMSCs + tethered IGF-1).

Figure 4.

Tethered IGF-1 with BMSCs improved the expression of specific myocardial maturation markers. Total RNA and proteins were extracted from heart tissues. (A) Reverse transcription-quantitative polymerase chain reaction results revealed that mRNA levels of N-cad, β-MHC and CX-43 were increased by tethered IGF-1. GAPDH was used as an internal control. (B) Western blotting demonstrated that N-cad, β-MHC and CX-43 protein expression was increased by tethered IGF-1 treatment. (C) Quantified western blotting results. **P<0.01 vs. control group and ^##P<0.01 vs. G1 and G2. IGF-1, insulin-like growth factor 1; tethered IGF-1, biotinylated IGF-1 with biotinylated self-assembling peptides; BMSCs, bone marrow stem cells; N-cad, N-cadherin; β-MHC, β-myosin heavy chain; CX-43, connexin-43.