Abstract
Background
Administration of Botulinum toxin type A (BTX-A) in patients with spastic cerebral palsy aims to improve mobility by increasing joint range of motion and decreasing passive resistance. However, our recent animal experiments indicated that BTX-A can decrease muscle”s length range of force exertion (Lrange), and increase its passive forces and extracellular matrix (ECM) collagen content. Moreover, BTX-A injected into the tibialis anterior (TA) was shown to spread into non-injected synergistic muscles in the whole anterior crural compartment. These effects that contradict the treatment aims deserve further investigation.
Aim
To test in a rat model if: (1) BTX-A injected into the medial and lateral gastrocnemius (GM&GL) muscles spreads into the synergistic soleus (SOL) as well as antagonistic TA and extensor digitorum longus (EDL). (2) The muscles exposed show a wider Lrange, decreased muscle passive force and reduced ECM collagen.
Methods
2×0.1U/20µl of BTX-A (BTX-A group, n=6) or only 2×20µl of saline (Control group, n=6) were prepared and each was injected into the mid-belly of the GM and GL separately. 5 days post injection, forces of all muscles were measured in passive state and also on activation. The GM&GL length was changed whereas; all other muscles were kept at constant length. After biomechanical testing, the muscles were histologically analyzed using Gomori trichrome stain to detect ECM collagen. Two-way ANOVA (factors: GM&GL length and animal group) was used to assess BTX-A effects on forces, and the Kruskal-Wallis test was used to test the change in proportion of collagenous tissue for each muscle. Differences were considered significant at p<0.05.
Results
Injected muscles: ANOVA showed significant main effects of both factors on GM&GL total forces and a significant interaction. Force reductions are more pronounced at shorter lengths (increase from 80.8% to 88.4% with decreasing length). Lrange decreased (by 24.1%). ANOVA showed significant main effects of only muscle length on GM&GL passive forces and no significant interaction. Non-injected muscles: ANOVA showed significant main effects of both factors (for SOL), or only of BTX-A (for TA and EDL) only on muscle total forces, but no significant interaction. Force drops for the SOL (89.8%) and anterior crural muscles (57.0% and 51.0% for TA and EDL) do indicate spread of BTX-A intra- and extra-compartmentally. Histological analyses showed increased ECM collagen contents of BTX-A group for the GM&GL, TA, and EDL.
Conclusion
Narrowed Lrange and increased ECM collagen content are not in accord with the clinical purpose of the treatment. BTX-A did not reduce passive forces, but did not cause an increase either. Remarkably, the results show that BTX-A leakage is a major issue that can affect muscles of even antagonistic muscle compartments. Hence, our animal experiments indicate much more complex BTX-A effects than considered, which requires further testing in patients.
