Figure 1c:

General study design and workflow. (a) Schematic for the TF reporter gene construct fluc2-egfp-sr39ttk driven by the ubiquitin C promoter. The TF reporter gene consists of fluc2, a 14–amino acid spacer, egfp, an eight–amino acid spacer, and sr39ttk. (b) Schematic for lentiviral transduction of MSC. The ubiquitin (U) TF reporter gene construct was excised and cloned into a second-generation self-inactivating lentiviral vector system. High-titer lentivirus containing the ubiquitin TF reporter gene was produced, and MSC were transduced with lentivirus containing the ubiquitin TF reporter gene in cell culture, generating MSC-TF. (c) MSC-TF were expanded for 2 weeks and were then sorted for cells showing high enhanced green fluorescent protein (EGFP) (GFP+) and those showing low EGFP (GFP−). This was referred to as sort 1. After expansion in culture for 2 weeks, the EGFP^high cells from sort 1 were sorted again; this was referred to as sort 2. Again, after expansion in culture, the EGFP^high cells from sort 2 were sorted for high-expressing, EGFP^high cells; this was referred to as sort 3. The EGFP^high cells from sort 3 were expanded in culture, and the resulting MSC-TF were frozen in aliquots and used in all mouse, rat, and swine studies. (d) Schematic for study design for CCT after MI in mice. MSC-TF were divided into four groups, including MSC-TF with no MI (n = 5), MSC-TF with MI (n = 8), mock injection (n = 3), or MSC with no reporter gene (n = 3). Each group underwent MI induction, injection of 5 × 10⁵ MSC-TF, and bioluminescence imaging on days 2, 4, 8, and 14.