Lin-Ling Zhang

Vibration-induced disruption of retrograde axoplasmic transport in peripheral nerve.

Hand‐arm vibration syndrome (HAVS) results from excessive exposure to hand‐transmitted vibration. Whether the peripheral nerve damage characteristic of HAVS is a direct result of vibration or is secondary to vascular insufficiency remains unclear. The purpose of this study was to explore the effect of vibration exposure on axoplasmic transport in peripheral nerves and soleus motor neurons. Sciatic nerves and motor neurons from rats following two 5‐h periods of vibration exposure demonstrated disruption in retrograde transport compared to normal. After 10 days of vibration (5 h/day), axoplasmic transport failed to recover within 24–48 h in most rats. This study demonstrates that disrupted axoplasmic transport is an early consequence of short‐term vibration exposure. The effects of vibration on axoplasmic transport also appear to be cumulative. This study provides a new biological way to evaluate measures to prevent early vibration injury. Muscle Nerve, 2005

Upregulation of Specific mRNA Levels in Rat Brain After Cell Phone Exposure

Adult Sprague-Dawley rats were exposed to regular cell phones for 6 h per day for 126 days (18 weeks). RT-PCR was used to investigate the changes in levels of mRNA synthesis of several injury-associated proteins. Calcium ATPase, Neural Cell Adhesion Molecule, Neural Growth Factor, and Vascular Endothelial Growth Factor were evaluated. The results showed statistically significant mRNA up-regulation of these proteins in the brains of rats exposed to cell phone radiation. These results indicate that relative chronic exposure to cell phone microwave radiation may result in cumulative injuries that could eventually lead to clinically significant neurological damage.

Neural systemic impairment from whole-body vibration

Insidious brain microinjury from motor vehicle‐induced whole‐body vibration (WBV) has not yet been investigated. For a long time we have believed that WBV would cause cumulative brain microinjury and impair cerebral function, which suggests an important risk factor for motor vehicle accidents and secondary cerebral vascular diseases. Fifty‐six Sprague‐Dawley rats were divided into seven groups (n = 8): 1) 2‐week normal control group, 2) 2‐week sham control group (restrained in the tube without vibration), 3) 2‐week vibration group (exposed to whole‐body vibration at 30 Hz and 0.5g acceleration for 4 hr/day, 5 days/week, for 2 weeks), 4) 4‐week sham control group, 5) 4‐week vibration group, 6) 8‐week sham control group, and 7) 8‐week vibration group. At the end point, all rats were evaluated in behavior, physiological, and brain histopathological studies. The cerebral injury from WBV is a cumulative process starting with vasospasm squeezing of the endothelial cells, followed by constriction of the cerebral arteries. After the 4‐week vibration, brain neuron apoptosis started. After the 8‐week vibration, vacuoles increased further in the brain arteries. Brain capillary walls thickened, mean neuron size was obviously reduced, neuron necrosis became prominent, and wide‐ranging chronic cerebral edema was seen. These pathological findings are strongly correlated with neural functional impairments.

The effect of calcium modulating agents on peripheral nerve recovery after crush

After a nerve injury, calcium concentration in the intra-nerve fiber drastically increases. The purpose of our study was to test an implantable micro-osmotic pump to deliver medications to accelerate calcium absorption, thereby greatly improving nerve regeneration. Twenty-four SD rats were divided into four groups of six each: (1) Sham control: crush injury to sciatic nerve only; (2) Crush injury with a Nifedipine pump; (3) Crush injury with a Calcitonin pump; (4) Crush injury with a Saline pump. Each rats right sciatic nerve was crushed. The micro-osmotic pump was implanted in the neck, and the dripping tube was routed to the injured nerve. After four weeks of survival time, compound muscle action potential (CMAP), tetanic muscle force (TMF), myelinated nerve fiber area (NFA), nerve calcium concentration (NCC), and calcified spots (CS) were evaluated. The calcium absorption rate (CAR) was also determined. The order from highest to lowest recovery rate was Nifedipine>Calcitonin>Sham control>Saline. Differences among the groups were statistically significant (P<0.001, ANOVA test), and the difference between Nifedipine/Calcitonin and Saline/Sham control were all statistically significant (P<0.001, t-test). The correlation rate of NCC with CMAP/TMF and with NFA/CS and CAR were calculated to be 0.99 (all P<0.001, Pearsons Correlation). We conclude from this study that nerve regeneration strongly correlated with calcium absorption; our new data has shown greatly improved nerve functional recovery, and this can potentially be translated into clinical applications.

Negative Effect of High Calcium Levels on Schwann Cell Survival

Our previous studies showed that the functional recovery of an injured nerve correlates with calcium absorption, and that acceleration of this absorption can greatly improve regeneration. In our study, we examined effects of the calcium concentration on cultured Schwann cells obtained from the rat sciatic nerve. Using published methods and simple calculations, we measured the calcium concentration inside normal and injured (crushed) nerve fibers and found that the injured-nerve internal calcium concentration is 16.8 mM, on average. The cells, including Schwann cells, were isolated from an intact sciatic nerve and cultured with various Ca2+ concentrations of calcium-spiked media, from the normal (1.8 mM) to 5.0–16.8 mM. The cells were fixed, blocked, and incubated with Anti-S-100, Goat-Anti-Mouse, and Propidium Iodide. They were viewed under fluorescent conditions to identify the Schwann cells. All the cells in the control dishes were viable, and there were large numbers of Schwann cells present. All the experimental dishes showed the profound concentration-dependent harmful effect of excessive calcium on the number and type of cells present. The high level of calcium corresponding to nerve injury severely affected Schwann and all other cell survival and growth in this cell culture study. Thus, Schwann cell survival and growth are very sensitive to and negatively affected by higher calcium levels than that in the normal media.

Qualitative Effect on mRNAs of Injury-Associated Proteins by Cell Phone Like Radiation in Rat Facial Nerves

Rats were exposed to cellphone radiation for 6 hours per day for 18 weeks. The buccal and mandibular branches of the facial nerve were evaluated for this study. The mRNA levels of four proteins that are usually up regulated when an injury has occurred were investigated; included were Calcium ATP-ase, Endothelin, Neural Cell Adhesion Molecule, and Neural Growth Factor. These isolated mRNAs were subjected to RT-PCR and all four were up regulated. The mandibular nerve showed a higher and broader level of up regulation than the buccal nerve. All four mRNA up regulations for the mandibular nerve and two for the buccal nerve were also statistically significant. These specific injury-related findings were mild. As the use of these cell phones continues, there most likely will be permanent damage to these tissues over the years and the likelihood of tumors, cancers, and system failures will potentially increase.

Calcitonin pump improves nerve regeneration after transection injury and repair

Introduction: After nerve injury, excessive calcium impedes nerve regeneration. We previously showed that calcitonin improved nerve regeneration in crush injury. We aimed to validate the direct effect of calcitonin on transected and repaired nerve. Methods: Two rat groups (n = 8) underwent sciatic nerve transection followed by direct repair. In the calcitonin group, a calcitonin‐filled mini‐osmotic pump was implanted subcutaneously, with a catheter parallel to the repaired nerve. The control group underwent repair only, without a pump. Evaluation and comparison between the groups included: (1) compound muscle action potential recording of the extensor digitorum longus (EDL) muscle; (2) tetanic muscle force test of EDL; (3) nerve calcium concentration; and (4) nerve fiber count and calcified spot count. Results: The calcitonin pump group showed superior recovery. Conclusions: Calcitonin affects injured and repaired peripheral nerve directly. The calcitonin‐filled mini‐osmotic pump improved nerve functional recovery by accelerating calcium absorption from the repaired nerve. This finding has potential clinical applications. Muscle Nerve 51: 229–234, 2015

A New Computerized Morphometric Analysis for Peripheral Nerve Study

The commonly used methods to quantify axon numbers and mean area include manual and semiautomated procedures. The authors introduce a new fully automated method of morphometric analysis using ImageJ and Paint.net software to improve efficiency and accuracy. A total of six rat sciatic nerves were examined for their axon numbers and mean axon area by comparing the manual method or semiautomated MetaVue method with the new ImageJ method. It was observed that the number of axons for manual counting and ImageJ were 4,630 ± 403 and 4,779 ± 352, respectively, and the difference was not statistically significant (p > 0.5, t-test). The mean axon area measured was 13.44 ± 2.62 µm2 for MetaVue and 8.87 ± 0.78 µm2 for ImageJ, respectively, and the difference was statistically significant (p < 0.01, t-test). The standard error and coefficient of variation of MetaVue were 1.07 and 0.195; and for ImageJ were 0.32 and 0.087. The authors conclude that their new approach demonstrates improved convenience, time efficiency, accuracy, and less operator error or bias.

Effect of calcitonin on cultured schwann cells: Calcitonin and Schwann Cells

Introduction: After nerve injury, calcium concentrations in intranerve fibers quickly increase. We have shown that functional recovery of injured nerves correlates with calcium absorption. A slight increase in calcium reduces the number of Schwann cells present. Calcitonin therapy greatly improves regeneration by accelerating calcium absorption. We examined the effect of adding calcitonin to higher concentration calcium media on cultured Schwann cells. Methods: The cells, isolated from intact sciatic nerves, were cultured with normal or higher concentration calcium media with or without calcitonin. Schwann cells were incubated with anti–S‐100, goat–anti‐mouse, and propidium iodide and then viewed through fluorescent light and phase‐contrast microscopy for observation and analysis. Results: The cells in each calcitonin‐containing medium showed many Schwann cells, however, the cells in the higher concentration calcium media showed fewer and more defective Schwann cells. Conclusion: These results show that calcitonin protects against the harmful effects of excessive calcium encountered in peripheral nerve injury. Muscle Nerve 56: 768–772, 2017

Best time window for the use of calcium‐modulating agents to improve functional recovery in injured peripheral nerves—An experiment in rats

Peripheral nerve injury can have a devastating effect on daily life. Calcium concentrations in nerve fibers drastically increase after nerve injury, and this activates downstream processes leading to neuron death. Our previous studies showed that calcium‐modulating agents decrease calcium accumulation, which aids in regeneration of injured peripheral nerves; however, the optimal therapeutic window for this application has not yet been identified. In this study, we show that calcium clearance after nerve injury is positively correlated with functional recovery in rats suffering from a crushed sciatic nerve injury. After the nerve injury, calcium accumulation increased. Peak volume is from 2 to 8 weeks post injury; calcium accumulation then gradually decreased over the following 24‐week period. The compound muscle action potential (CMAP) measurement from the extensor digitorum longus muscle recovered to nearly normal levels in 24 weeks. Simultaneously, real‐time polymerase chain reaction results showed that upregulation of calcium‐ATPase (a membrane protein that transports calcium out of nerve fibers) mRNA peaked at 12 weeks. These results suggest that without intervention, the peak in calcium‐ATPase mRNA expression in the injured nerve occurs after the peak in calcium accumulation, and CMAP recovery continues beyond 24 weeks. Immediately using calcium‐modulating agents after crushed nerve injury improved functional recovery. These studies suggest that a crucial time frame in which to initiate effective clinical approaches to accelerate calcium clearance and nerve regeneration would be prior to 2 weeks post injury.