What Works in Collaboration? Identifying Key Ingredients to Improve Service Delivery in Schools.

PURPOSE: This survey study examined factors that may influence interprofessional collaboration in schools to support children with oral and written language impairments, namely, knowledge, collaborative beliefs and practices, and resources. METHOD: A survey was conducted across 319 school-based professionals, in a partnering public school district, to examine these constructs within the context of each individual participant's professional role. RESULTS: Between-groups comparisons were made between special educators, general educators, paraprofessionals, and indirect educators (i.e., those whose roles do not explicitly include language-based instruction). Special educators had significantly higher levels of learning experiences and knowledge about language disorders compared to other groups. Those who engaged in the most independent learning (i.e., learning outside of pre- or in-service learning) exhibited the highest levels of knowledge. Collaborative beliefs among professionals were best predicted by access to resources and overall professional learning experiences. More positive collaborative beliefs, in turn, predicted higher rates of collaborative behaviors. Resources were predicted by a small negative relationship with years of experience and by working in specific school buildings within the district. Those with less experience in specific buildings reported more resources. CONCLUSION: Individuals with higher levels of individual learning and positive attitudes toward collaboration may enhance the interprofessional collaborative practices of teams supporting children with language disorders. SUPPLEMENTAL MATERIAL: https://doi.org/10.23641/asha.24059178.

Abnormal prostaglandin E2 production blocks myogenic differentiation in myotonic dystrophy.

The congenital form of myotonic dystrophy type 1 (DM1) is the most severe type of the disease associated with CTG expansions over 1500 repeats and delayed muscle maturation. The mechanistic basis of the congenital form of DM1 is mostly unknown. Here, we show that muscle satellite cells bearing large CTG expansions (>3000) secrete a soluble factor that inhibits the fusion of normal myoblasts in culture. We identified this factor as prostaglandin E2 (PGE(2)). In these DM1 cells, PGE(2) production is increased through up-regulation of cyclooxygenase 2 (Cox-2), mPGES-1 and prostaglandin EP2/EP4 receptors. Elevated levels of PGE(2) inhibit myogenic differentiation by decreasing the intracellular levels of calcium. Exogenous addition of acetylsalicylic acid, an inhibitor of Cox enzymes, abolishes PGE(2) abnormal secretion and restores the differentiation of DM1 muscle cells. These data indicate that the delay in muscle maturation observed in congenital DM1 may result, at least in part, from an altered autocrine mechanism. Inhibitors of prostaglandin synthesis may thus offer a powerful method to restore the differentiation of DM1 muscle cells.

Absence of a differentiation defect in muscle satellite cells from DM2 patients.

Myotonic dystrophy type 1 (DM1) and type II (DM2) are dominantly inherited multisystemic disorders. DM1 is triggered by the pathological expansion of a (CTG)(n) triplet repeat in the DMPK gene, whereas a (CCTG)(n) tetranucleotide repeat expansion in the ZNF9 gene causes DM2. Both forms of the disease share several features, even though the causative mutations and the loci involved differ. Important distinctions exist, such as the lack of a congenital form of DM2. The reason for these disparities is unknown. In this study, we characterized skeletal muscle satellite cells from adult DM2 patients to provide an in vitro model for the disease. We used muscle cells from DM1 biopsies as a comparison tool. Our main finding is that DM2 satellite cells differentiate normally in vitro. Myotube formation was similar to unaffected controls. In contrast, fetal DM1 cells were deficient in that ability. Consistent with this observation, the myogenic program in DM2 was intact but is compromised in fetal DM1 cells. Although expression of the ZNF9 gene was enhanced in DM2 during differentiation, the levels of the ZNF9 protein were substantially reduced. This suggests that the presence of a large CCTG tract impairs the translation of the ZNF9 mRNA. Additionally, DM2 muscle biopsies displayed the altered splicing of the insulin receptor mRNA, correlating with insulin resistance in the patients. Finally, CUGBP1 steady-state protein levels were unchanged in DM2 cultured muscle cells and in DM2 muscle biopsies relative to controls, whereas they are increased in DM1 muscle cells. Our findings suggest that the myogenic program throughout muscle development and tissue regeneration is intact in DM2.

Replication and expansion of trinucleotide repeats in yeast.

The mechanisms of trinucleotide repeat expansions, underlying more than a dozen hereditary neurological disorders, are yet to be understood. Here we looked at the replication of (CGG)(n) x (CCG)(n) and (CAG)(n) x (CTG)(n) repeats and their propensity to expand in Saccharomyces cerevisiae. Using electrophoretic analysis of replication intermediates, we found that (CGG)(n) x (CCG)(n) repeats significantly attenuate replication fork progression. Replication inhibition for this sequence becomes evident at as few as approximately 10 repeats and reaches a maximal level at 30 to 40 repeats. This is the first direct demonstration of replication attenuation by a triplet repeat in a eukaryotic system in vivo. For (CAG)(n) x (CTG)(n) repeats, on the contrary, there is only a marginal replication inhibition even at 80 repeats. The propensity of trinucleotide repeats to expand was evaluated in a parallel genetic study. In wild-type cells, expansions of (CGG)(25) x (CCG)(25) and (CAG)(25) x (CTG)(25) repeat tracts occurred with similar low rates. A mutation in the large subunit of the replicative replication factor C complex (rfc1-1) increased the expansion rate for the (CGG)(25) repeat approximately 50-fold but had a much smaller effect on the expansion of the (CTG)(25) repeat. These data show dramatic sequence-specific expansion effects due to a mutation in the lagging strand DNA synthesis machinery. Together, the results of this study suggest that expansions are likely to result when the replication fork attempts to escape from the stall site.

Mechanistic features of CAG*CTG repeat contractions in cultured cells revealed by a novel genetic assay.

Trinucleotide repeats (TNRs) undergo high frequency mutagenesis to cause at least 15 neurodegenerative diseases. To understand better the molecular mechanisms of TNR instability in cultured cells, a new genetic assay was created using a shuttle vector. The shuttle vector contains a promoter-TNR-reporter gene construct whose expression is dependent on TNR length. The vector harbors the SV40 ori and large T antigen gene, allowing portability between primate cell lines. The shuttle vector is propagated in cultured cells, then recovered and analyzed in yeast using selection for reporter gene expression. We show that (CAG*CTG)25-33 contracts at frequencies as high as 1% in 293T and 293 human cells and in COS-1 monkey cells, provided that the plasmid undergoes replication. Hairpin-forming capacity of the repeat sequence stimulated contractions. Evidence for a threshold was observed between 25 and 33 repeats in COS-1 cells, where contraction frequencies increased sharply (up 720%) over a narrow range of repeat lengths. Expression of the mismatch repair protein Mlh1 does not correlate with repeat instability, suggesting contractions are independent of mismatch repair in our system. Together, these findings recapitulate certain features of human genetics and therefore establish a novel cell culture system to help provide new mechanistic insights into CAG*CTG repeat instability.

RNA based gene therapy for dominantly inherited diseases.

There are numerous examples in the literature of gene therapy applications for recessive disorders. There are precious few instances, however, of studies conducted to treat dominantly inherited pathologies. The reasons are simple: there are fewer cases of dominantly inherited diseases on one hand, but mostly it is far easier to correct recessive mutations than dominant ones. Typically recessive mutations cause a loss of (or reduced) gene function which can be compensated for by introduction of a replacement allele into the cell. In contrast, dominant negative mutations not only display impaired function, but also exhibit a novel one that is pathologic to the cell. Treating these conditions by gene therapy implies silencing the dominant allele without altering the expression of the wild-type gene. We describe here different strategies aimed at silencing dominant mutations through mRNA destruction and provide examples of their application to known autosomal dominant diseases. An overview of the most common molecular tools (antisense DNA and RNA, ribozymes and RNA interference) suitable to utilize these strategies is also presented and we discuss the relevant aspects involved in the choice of a particular approach in a gene therapy experiment.