Additional file 1 of Expressing a Z-disk nebulin fragment in nebulin-deficient mouse muscle: effects on muscle structure and function
Creators: name: Li, Frank
name: Kolb, Justin
name: Crudele, Julie
name: Hourani, Zaynab
name: Smith, John
name: Chamberlain, Jeffrey
name: Granzier, Henk
Publisher: name: figshare
Year: 2020
Subjects: subject: Biochemistry
subject: Medicine
subject: Microbiology
subject: FOS: Biological sciences
subject: FOS: Biological sciences
subject: Cell Biology
subject: Molecular Biology
subject: Physiology
subject: Pharmacology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: FOS: Chemical sciences
subject: Immunology
subject: FOS: Clinical medicine
subject: FOS: Clinical medicine
subject: Cancer
subject: Computational Biology

description: Additional file 1: Figure S1. Effect of AAV treatment on muscle weights (TC and EDL) and physiological cross-sectional area (PCSA) in EDL muscle. A) Vehicle-treated and AAV-treated, CTRL and Neb cKO TC weights in the 1x (left) and 3x (right) dosage groups. Paired t-test revealed a small but significant decrease in the Neb cKO TC weights of the 1x dosage group. This difference was not replicated in the 3x dosage group. B) Vehicle-treated and AAV-treated, CTRL and Neb cKO EDL weights in the 1x (left) and 3x (right) dosage groups. No significant differences were found using paired t-tests between AAV-treated EDLs and their contralateral, vehicle-treated muscles. C) PCSAs of the EDL muscles in both 1x (left) and 3x (right) dosage groups (see Methods for details). AAV treatment had no significant effect on the cross-sectional areas. (1x: n = 9,12 mice; 3x: n = 4,9 mice). Figure S2. Localization of the Z-disk AAV construct in the 3x AAV dosage group. Figure S3. High magnification TEM images of Z-disk structure in Neb cKO TCs. Even at high 43,000x magnification, the differences in Z-disk widths are difficult to discern. Z-disks were precisely measured using the full-width, half-max values of a gaussian fit of the gray values. Figure S4. Quantification of MHC composition in 3x dosage treatment group. A) Quantification of MHC composition in vehicle-treated and AAV-treated CTRL TCs and EDLs (3x dosage). Paired t-tests reveal a significant increase in Type IIA/X MHC and a significant decrease in Type IIB MHC in both muscle types. (n = 4 mice). B) Quantification of MHC composition in vehicle-treated and AAV-treated Neb cKO TCs and EDLs (3x dosage). Paired t-tests reveal a significant increase in Type IIB MHC and a significant decrease in Type I MHC. (n = 8 mice). Table S1. Specific force results at each stimulation frequency were compared at the 1x dose (A) and 3x dose (B). (1x: n = 9,11 mice; 3x: n = 4,9 mice). Figure S5. Normalized force-frequency curves. A) Normalized force-frequency (FF) curves at 1x (left) and 3x (right) dosages. The Neb cKO curves are left-shifted from CTRL curves. No consistent AAV-effect is seen. (Forces at each stimulation frequency were divided by the maximal tetanic force). B) Quantification of frequency that results in half-maximal force at 1x (left) and 3x (right) dosages. A significant reduction in the ‘half-max frequency’ in Neb cKO mice treated with the 1x dosage (paired t-test) occurs. This decrease is lost in the 3x dosage group and no effect is present in the CTRL group. (Paired t-tests were used in this analysis). (1x: n = 9,11 mice; 3x: n = 4,9 mice). Figure S6. Analysis of contraction kinetics in both 1x and 3x dosage groups. A) Time to max force during a twitch (left) and a 200 Hz tetanus (right). Neb cKO mice have a significantly longer time to maximal twitch force in the 3x dosage group and a significantly longer time to max tetanic force in the 1x dosage group. B) Half-relaxation time after the twitch (left) and the 200 Hz tetanus (right). While Neb cKO mice have a significantly faster relaxation time in the 3x dosage group, none of the other comparisons had a change in twitch relaxation time. Comparatively, all groups except for the Neb cKO mice in the 3x dosage group had significantly longer relaxation times following a maximal tetanus. (Paired t-tests were used in these analyses. 1x: n = 9,11 mice; 3x: n = 4,9 mice). Figure S7. Expression of nebulin regulatory proteins in 3x dosage group. (n = 4,9 mice). Figure S8. Representative blots pertaining to different components of the Z-disks. (n = 3–5 mice). Figure S9. Analyses of Z-disk protein expression. (n = 3–5 mice).