Two-minute versus 6-minute walk distances during 6-minute walk test in neuromuscular disease: Is the 2-minute walk test an effective alternative to a 6-minute walk test?

Functional tests such as Motor Function Measure-32 (MFM-32), supine to stand, ascend/descend stairs permit the assessment of task-specific motor function in neuromuscular disease (NMD). The 6-min walk test (6MWT), though functional, is primarily used to assess endurance and disease progression in children with neuromuscular disorders. Barriers to 6MWT administration, in this population, can include reduced attention span due to age and inability to tolerate test length due to weakness. We propose task-specific functional deficits are related to endurance. Additionally, the 2-min walk test (2MWT) could effectively replace the 6MWT in this population. Seventy-seven participants, ages 5-18, with a variety of neuromuscular disorders performed the 6MWT, timed functional tests (TFT), and the MFM-32. Correlation and paired t-test analyses were used to compare the distance walked in the first 2 min (2MWD) to the distance walked in the entire 6 min (6MWD) and to the functional outcome measures above. The 2MWD strongly correlated with 6MWD and the other outcome measures. Paired t-test analysis also showed that the 2MWD did not differ from the distance walked in the last 2 min of the 6MWT. Although equivalence testing could not reject the claim that this difference exceeded the upper practical limit of 9.5 m, it only showed a modest overestimation of the 4-6MWD compared with the 2MWD. Together, our results support the ability of the 2MWD to predict the 6MWD, specifically in the pediatric neuromuscular disease population.

Improved gold chloride staining method for anatomical analysis of sensory nerve endings in the shoulder capsule and labrum as examples of loose and dense fibrous tissues.

Consistency in gold chloride staining is essential for anatomical analysis of sensory nerve endings. The gold chloride stain for this purpose has been modified by many investigators, but often yields inconsistent staining, which makes it difficult to differentiate structures and to determine nerve ending distribution in large tissue samples. We introduce additional steps and major changes to the modified Gairns' protocol. We controlled the temperature and mixing rate during tissue staining to achieve consistent staining and complete solution penetration. We subjected samples to sucrose dehydration to improve cutting efficiency. We then exposed samples to a solution containing lemon juice, formic acid and paraformaldehyde to produce optimal tissue transparency with minimal tissue deformity. We extended the time for gold chloride impregnation 1.5 fold. Gold chloride was reduced in the labrum using 25% formic acid in water for 18 h and in the capsule using 25% formic acid in citrate phosphate buffer for 2 h. Citrate binds gold nanoparticles, which minimizes aggregation in the tissue. We stored samples in fresh ultrapure water at 4° C to slow reduction and to maintain color contrast in the tissue. Tissue samples were embedded in Tissue Tek and sectioned at 80 and 100 µm instead of using glycerin and teasing the tissue apart as in Gairns' modified gold chloride method. We attached sections directly to gelatin subbed slides after sectioning with a cryostat. The slides then were processed and coverslipped with Permount. Staining consistency was demonstrated throughout the tissue sections and neural structures were clearly identifiable.