Increasing myosin light chain 3f (MLC3f) protects against a decline in contractile velocity

Figures

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  Fig 1. Experimental timeline to highlight day of rAd-MLC3f injection. The experimental timeline in days is divided into two phases [Acclimatization of at least 7 days and Hindlimb Unloading (HU) for 7 days]. The study evaluated three titers of the rAd-MLC3f 1×1012 ifu/ml, 2.5 x 1011, 3.75 x 1011 or 5 x 1011 ifu/ml and three injection timepoints. 250 μl total of the rAd-MLC3f was injected into five areas of the SM muscle of both hindlimbs and at one of the three-time points: one-day before HU (-1), two-days after HU (2), or four-days after HU (4).

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  Fig 2. Representative MLC isoforms of single fibers in CON, HU, and HUM groups. The MLC isoforms within each individual skeletal muscle fiber were detected by 12% SDS-PAGE and silver staining technique. Each lane contains a single skeletal muscle fiber (lanes 2, 3 for CON, 4, 5 for HU, and 6, 7 for HUM). The ratio of MLC3f/MLC1f, MLC3f/MLC2f, and MLC3f/(MLC1f+MLC3f) was calculated by determining the relative content of MLC1f, MLC2f, and MLC3f with densitometry. Tibialis anterior (TA) muscle was used as a protein standard (STD), lane 1. The representative image was selected from control (CON), hindlimb unloading (HU), and HU for 7 days plus rAd-MLC3f injections (HUM; four-days after HU with 5 x 1011 ifu/ml). The dashed lines between the CON, HU, and HUM groups indicate that the gel lanes were taken from three separate gels.

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  Fig 3. Percent of MLC3f protein content and Vo in single MHC type IIB fibers. The mean percent of MLC3f protein content (Panel A) and the mean unloaded shortening velocity (Vo, Panel B) of single MHC type IIB fibers from control (CON), HU for 7 days (HU), and HU for 7 days plus rAd-MLC3f injections (HUM). Vo was determined by the slack test and the percent of MLC3f protein content of each fiber was determined by 12% SDS-PAGE. The number of MHC type IIB fibers is shown in the boxes. * indicates a significant difference from CON group. † indicates a significant difference from HU group. Significance was set at P<0.05 and values are expressed as mean ± SEM.

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  Fig 4. Relationship between Percent MLC3f content and Vo. The percent of MLC3f protein content and the corresponding Vo in each single MHC type IIB fiber investigated from CON, HU, and HUM (3.75 x 1011–5 x 1011 ifu/ml rAd-MLC3f injection) (Panel A). The relationship between percent of MLC3f and the corresponding Vo of single fibers where the fibers were grouped based on 1% intervals of MLC3f content and their respective mean Vo (Panel B). The simple linear regression analyses are presented.