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Electron microscopy of trypsin-digested peripheral nerve myelin
Abstract:
Sciatic nerves from mice were removed and soaked in either PBS (phosphate buffered saline) or PBS plus I% trypsin (Sigma Type III) for various periods of time. Specimens were soaked at either room temperature or 37-degrees C at pH's ranging from 7.5 to 8.0. The epineural and perineural sheaths were split to allow the trypsin to penetrate the nerve. Tissue was prepared for electron microscopy by fixation in cacodylate buffered formaldehyde-glutaraldehyde solutions, post-fixed in OSO4 and embedded in Epon 812 or in glutaraldehyde-urea resin without osmication. After four h incubation at 37-degrees C or eight h at room temperature, the basement membranes of the Schwann cells became fragmented and detached and the myelin intraperiod band lost some density. After 18 h, myelin with swollen intraperiod bands displaying a loss of electron density and split main period bands was noted adjacent to normal myelin. Other areas had been transformed into vesicles indicating that the membranes of these vesicles appeared to have been derived from the detachment of both the intraperiod and main period bands within the myelin. Evidence is presented for the presence of trypsin digestable proteins in both the main period and intraperiod bands of peripheral nervouse system myelin.
Insights
Trypsin digestion fragmented Schwann cell basement membranes and altered myelin structure in mouse sciatic nerves. This indicates trypsin-digestible proteins are present in peripheral nervous system myelin.
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Peripheral nervous system (PNS) myelin provides insulation for nerve axons.
- Understanding the molecular composition of myelin is crucial for studying nerve health and disease.
Purpose of the Study:
- To investigate the effects of trypsin on the structural integrity of mouse sciatic nerve myelin.
- To identify the presence of trypsin-digestible proteins within PNS myelin sheaths.
Main Methods:
- Mouse sciatic nerves were incubated with trypsin (Sigma Type III) in phosphate-buffered saline (PBS) at varying temperatures and durations.
- Nerve tissues were prepared for electron microscopy using standard fixation and embedding techniques.
- Ultrastructural changes in myelin and Schwann cell basement membranes were analyzed.
Main Results:
- Trypsin digestion led to fragmentation and detachment of Schwann cell basement membranes.
- Myelin intraperiod bands lost density, and main period bands showed splitting after prolonged incubation.
- Myelin structures transformed into vesicles, suggesting membrane detachment.
Conclusions:
- Trypsin effectively digests components of the Schwann cell basement membrane.
- The study provides evidence for trypsin-digestible proteins within both the intraperiod and main period bands of PNS myelin.