Mutations in laminin α2-subunit (Lmα2, encoded by LAMA2) are linked to approximately 30% of congenital muscular dystrophy cases. Mice with a homozygous mutation in Lama2 (dy2J mice) express a nonpolymerizing form of laminin-211 (Lm211) and are a model for ambulatory-type Lmα2-deficient muscular dystrophy. Here, we developed transgenic dy2J mice with muscle-specific expression of αLNNd, a laminin/nidogen chimeric protein that provides a missing polymerization domain. Muscle-specific expression of αLNNd in dy2J mice resulted in strong amelioration of the dystrophic phenotype, manifested by the prevention of fibrosis and restoration of forelimb grip strength. αLNNd also restored myofiber shape, size, and numbers to control levels in dy2J mice. Laminin immunostaining and quantitation of tissue extractions revealed increased Lm211 expression in αLNNd-transgenic dy2J mice. In cultured myotubes, we determined that αLNNd expression increased myotube surface accumulation of polymerization-deficient recombinant laminins, with retention of collagen IV, reiterating the basement membrane (BM) changes observed in vivo. Laminin LN domain mutations linked to several of the Lmα2-deficient muscular dystrophies are predicted to compromise polymerization. The data herein support the hypothesis that engineered expression of αLNNd can overcome polymerization deficits to increase laminin, stabilize BM structure, and substantially ameliorate muscular dystrophy.
Authors
Karen K. McKee, Stephanie C. Crosson, Sarina Meinen, Judith R. Reinhard, Markus A. Rüegg, Peter D. Yurchenco
Forelimb (A), hind limb (B), and combined limb (C) measurements were conducted on control, dy2J, and αLNNd plus Tg+dy2J mice between 3 and 11 weeks of age. Values shown represent the mean ± SEM of individual averaged mouse sets of grip strength determinations divided by mouse mass. The number of mice used for each determination is indicated in red. Lowercase red letter pairs indicate the statistical significance ranges, which were determined by 1-way ANOVA with pairwise Holm-Sidak comparisons (P value ranges are indicated in the lower right corner of the top and bottom panels). NS, not significant. Dy2J mice showed reduced forelimb grip strength by 4 weeks of age. Grip strength was improved to near-control levels with the αLNNd transgene and became indistinguishable from that of control mice by 6 weeks of age. For hind limbs, the transgene partially increased grip strength between 3 and 7 weeks, but not thereafter, corresponding to the emergence of hind limb contractures. Note that the peripheral nerve defect, which selectively affected the hind limbs, was not corrected, because a muscle-specific promoter was used to drive αLNNd expression. Combined (all-limb) grip strength was reduced in dy2J mice, with substantial improvement (~50%) detected in the presence of the transgene.