Muscle stem cell therapy with satellite cells (SC) is improved by the reduction in muscle fibrosis following treatment with andrographolide. (A) Fibrosis was augmented as explained in the legend of Figure 1. During this period, mice were treated with andrographolide or vehicle (intraperitoneal (ip) injections three times per week, six animals per group). One week after the last drug administration (seven-month-old mice), 500 freshly isolated satellite cells purified from wild type (WT) mice were transplanted into both tibialis anterior (TA) muscles of each mdx mouse. At four weeks after engraftment, the number of fibers expressing dystrophin and collagen I was determined by immunofluorescence analysis of cryosections. The images are representative of two experimental groups with six mice per group. Bar corresponds to 200 μm. (B) Quantification of the data obtained in (A) showing the number of myofibers expressing dystrophin per TA muscle in each case. (C) The number of satellite cells (PAX-7-positive nuclei) was determined for isolated single muscle fibers from extensor digitorum longus muscle (EDL) of each case as an indicator of endogenous SC survival. (D) 500 WT SC freshly isolated as in (A), from the C57-EGFP mice were engrafted in mice treated as in (A) (six animals per group). Both TA muscles were dissected immediately after engraftment or after 2 or 15 days from two animals in each case. Total genomic DNA was purified. EGFP transgene present in the engrafted muscle was detected by qPCR. Values are expressed as mean ± SD of three independent experiments. (*P<0.05 relative to vehicle-treated mice, **P<0.01 relative to vehicle-treated mice at day 15).