Figure 1.
Expression of SMPX plasmids, in vitro and in vivo.
A) Western blot of SMPX-EGFP expression in HEK-293 cells stained with antibodies against EGFP. Lane 1: Negative control. Lane 2: EGFP only. Lane 3-6: SMPX-EGFP. B) In vivo fluorescence image of EDL muscle expressing pCMS-EGFP-Smpx. Scalebar is 500 microns. Inset: High magnification of single fibers. Scalebar is 50 microns. C) β-gal (left) and myosin heavy chain type 2A (right) stain on neighboring cross-sections from the same muscle as in B). β-gal expressing fibers marked with asterisks. Scale bar is 50 microns.
Figure 2.
SMPX-EGFP was excluded from all myonuclei.
A) C2C12 myoblasts expressing either EGFP or SMPX-EGFP stained with Hoechst 33342 to visualize nuclei. B) In vitro confocal image of dissected single rat EDL muscle fibers expressing EGFP or SMPX-EGFP stained with DAPI to visualize nuclei. C) Confocal images of EDL muscle fibers in situ, after no treatment (CON) or functional overload for 18 hours (LOAD), expressing SMPX-EGFP stained with Hoechst 33342 to visualize nuclei. Myonuclei are labeled with arrowheads. Scale bar is 10 microns.
Figure 3.
SMPX-EGFP was localized in bands flanking the z-disc.
A) In situ confocal image of EGFP or SMPX-EGFP expression from an isolated EDL rat muscle in Ringer-solution. Scale bar is 10 microns. B) In vitro confocal image of EGFP or SMPX-EGFP stained with Alexa Fluor 680 Phalloidin (red) to visualize actin filaments. Scale bar is 10 microns (inset 1 micron).
Figure 4.
Muscle fiber type and cross sectional area (CSA) in mouse EDL.
Single fibers from muscles transfected with either pCMS-EGFP (left leg sham control, only expressing EGFP) or pCMS-EGFP-Smpx (expressing SMPX and EGFP). A) CSA with n = 226/254 cells for sham/SMPX. Values are means +/- SD. B) Fiber type composition of the same fibers as in A.