Identification and characterization of dystrophin-locus-derived testis-specific protein: A testis-specific gene within the intronic region of the rat dystrophin gene.

The mammalian X chromosome exhibits enrichment in genes associated with germ cell development. Previously, we generated a rat model of Becker muscular dystrophy (BMD) characterized by an in-frame mutation in the dystrophin gene, situated on the X chromosome and responsible for encoding a protein crucial for muscle integrity. Male BMD rats are infertile owing to the absence of normal spermatids in the epididymis. Within the seminiferous tubules of BMD rats, elongated spermatids displayed abnormal morphology. To elucidate the cause of infertility, we identified a putative gene containing an open reading frame situated in the intronic region between exons 6 and 7 of the dystrophin gene, specifically deleted in male BMD rats. This identified gene, along with its encoded protein, exhibited specific detection within the testes, exclusively localized in round to elongated spermatids during spermiogenesis. Consequently, we designated the encoded protein as dystrophin-locus-derived testis-specific protein (DTSP). Given the absence of DTSP in the testes of BMD rats, we hypothesized that the loss of DTSP contributes to the infertility observed in male BMD rats.

Macroglossia and less advanced dystrophic change in the tongue muscle of the Duchenne muscular dystrophy rat.

BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked muscle disease caused by a complete lack of dystrophin, which stabilizes the plasma membrane of myofibers. The orofacial function is affected in an advanced stage of DMD and this often leads to an eating disorder such as dysphagia. Dysphagia is caused by multiple etiologies including decreased mastication and swallowing. Therefore, preventing the functional declines of mastication and swallowing in DMD is important to improve the patient's quality of life. In the present study, using a rat model of DMD we generated previously, we performed analyses on the masseter and tongue muscles, both are required for proper eating function. METHODS: Age-related changes of the masseter and tongue muscle of DMD rats were analyzed morphometrically, histologically, and immunohistochemically. Also, transcription of cellular senescent markers, and utrophin (Utrn), a functional analog of dystrophin, was examined. RESULTS: The masseter muscle of DMD rats showed progressive dystrophic changes as observed in their hindlimb muscle, accompanied by increased transcription of p16 and p19. On the other hand, the tongue of DMD rats showed macroglossia due to hypertrophy of myofibers with less dystrophic changes. Proliferative activity was preserved in the satellite cells from the tongue muscle but was perturbed severely in those from the masseter muscle. While Utrn transcription was increased in the masseter muscle of DMD rats compared to WT rats, probably due to a compensatory mechanism, its level in the tongue muscle was comparable between WT and DMD rats and was similar to that in the masseter muscle of DMD rats. CONCLUSIONS: Muscular dystrophy is less advanced in the tongue muscle compared to the masseter muscle in the DMD rat.

In utero transplantation of myoblasts and adipose-derived mesenchymal stem cells to murine models of Duchenne muscular dystrophy does not lead to engraftment and frequently results in fetal death.

Introduction: Duchenne muscular dystrophy (DMD) is a progressive disease that leads to damage of muscle and myocardium due to genetic abnormalities in the dystrophin gene. In utero cell transplantation that might facilitate allogenic transplantation is worth considering to treat this disease. Methods: We performed allogeneic in utero transplantation of GFP-positive myoblasts and adipose-derived mesenchymal stem cells into murine DMD model animals. The transplantation route in this study was fetal intraperitoneal transplantation and transplacental transplantation. Transplanted animals were examined at 4-weeks old by immunofluorescence staining and RT-qPCR. Results: No GFP-positive cells were found by immunofluorescence staining of skeletal muscle and no GFP mRNA was detected by RT-qPCR in any animal, transplantation method and cell type. Compared with previous reports, myoblast transplantation exhibited an equivalent mortality rate, but adipose-derived stem cell (ASC) transplantation produced a higher mortality rate. Conclusions: In utero transplantation of myoblasts or ASCs to murine models of DMD does not lead to engraftment and, in ASC transplantation primarily, frequently results in fetal death.

Loss of p16/Ink4a drives high frequency of rhabdomyosarcoma in a rat model of Duchenne muscular dystrophy.

Rhabdomyosarcoma (RMS) is an aggressive type of soft tissue sarcoma, and pleomorphic RMS is a rare subtype of RMS found in adult. p16 is a tumor suppressor which inhibits cell cycle. In human RMS, p16 gene is frequently deleted, but p16-null mice do not develop RMS. We reported that genetic ablation of p16 by the crossbreeding of p16 knock-out rats (p16-KO rats) improved the dystrophic phenotype of a rat model of Duchenne muscular dystrophy (Dmd-KO rats). However, p16/Dmd double knock-out rats (dKO rats) unexpectedly developed sarcoma. In the present study, we raised p16-KO, Dmd-KO, and dKO rats until 11 months of age. Twelve out of 22 dKO rats developed pleomorphic RMS after 9 months of age, while none of p16-KO rats and Dmd-KO rats developed tumor. The neoplasms were connected to skeletal muscle tissue with indistinct borders and characterized by diffuse proliferation of pleomorphic cells which had eosinophilic cytoplasm and atypical nuclei with anisokaryosis. For almost all cases, the tumor cells immunohistochemically expressed myogenic markers including desmin, MyoD, and myogenin. The single cell cloning from tumor primary cells gained 20 individual Pax7-negative MyoD-positive RMS cell clones. Our results demonstrated that double knock-out of p16 and dystrophin in rats leads to the development of pleomorphic RMS, providing an animal model that may be useful to study the developmental mechanism of pleomorphic RMS.