Effects of Theranekron and alpha-lipoic acid combined treatment on GAP-43 and Krox-20 gene expressions and inflammation markers in peripheral nerve injury

AUTHORS

Leman Sencar, Gülfidan Coşkun, Dilek Şaker, Tuğçe Sapmaz, Samet Kara, Alper Çelenk, Sema Polat, Derviş Mansuri Yılmaz, Y. Kenan Dağlıoğlu & Sait Polat

ABSTRACT

Peripheral nerve injury (PNI) is a major health problem that results in loss of motor and sensory functions. In treatment of PNI, various methods such as anastomosis, nerve grafts, nonneural tissue grafts, and nerve conduits are applied. In the present study, it was aimed to investigate the effects of Theranekron and Alpha-lipoic acid (ALA) combined treatment on nerve healing in experimental PNI by using histomorphometric, electron microscopic, immunohistochemical and molecular biological methods. Sixty-two Wistar rats were divided into six groups; the normal control group, sham operation group, experimental control group having a crush type injury with no treatment, Theranekron treatment group, ALA treatment group and Theranekron+ALA combined treatment group. Sciatic nerve tissue samples were obtained on days 1, 7 and 14 following injury in all groups. GAP-43 expression was upregulated in all PNI received groups compared to the control group. Krox-20 expression was downregulated in all groups that received PNI compared to the control group. While intensely positive TNF-α and IL-6 expressions were observed up to the 1st to the 14th day for the experimental control group, these expressions were seen as “weakly positive” in the treatment groups from the 1st day to the 14th day. The number of myelinated fibers was higher in the control and sham operation groups. Additionally, the number of myelinated nerve fibers increased in the combined treatment group. In conclusion, these findings suggest that combined therapy of Theranekron and ALA promotes structural recovery and it should be considered as an effective treatment protocol following PNI.

Nerve transfer for restoration of lower motor neuron-lesioned bladder function. Part 2: correlation between histological changes and nerve evoked contractions

AUTHORS

Mary F. Barbe, Courtney L. Testa, Geneva E. Cruz, Nagat A. Frara, Ekta Tiwari, Lucas J. Hobson, Brian S. McIntyre, Danielle S. Porreca, Dania Giaddui, Alan S. Braverman, Emily P. Day, Mamta Amin, Justin M. Brown, Michael Mazzei, Michel A. Pontari, Ida J. Wagner, and Michael R. Ruggieri Sr.

ABSTRACT

We determined the effect of pelvic organ decentralization and reinnervation 1 yr later on urinary bladder histology and function. Nineteen canines underwent decentralization by bilateral transection of all coccygeal and sacral (S) spinal roots, dorsal roots of lumbar (L)7, and hypogastric nerves. After exclusions, eight were reinnervated 12 mo postdecentralization with obturator-to-pelvic and sciatic-to-pudendal nerve transfers, then euthanized 8-12 mo later. Four served as long-term decentralized only animals. Before euthanasia, pelvic or transferred nerves and L1–S3 spinal roots were stimulated and maximum detrusor pressure (MDP) recorded. Bladder specimens were collected for histological and ex vivo smooth muscle contractility studies. Both reinnervated and decentralized animals showed less or denuded urothelium, fewer intramural ganglia, and more inflammation and collagen, than controls, although percent muscle was maintained. In reinnervated animals, pgp9.5+ axon density was higher compared with decentralized animals. Ex vivo smooth muscle contractions in response to KCl correlated positively with submucosal inflammation, detrusor muscle thickness, and pgp9.5+ axon density. In vivo, reinnervated animals showed higher MDP after stimulation of L1–L6 roots compared with their transected L7–S3 roots, and reinnervated and decentralized animals showed lower MDP than controls after stimulation of nerves (due likely to fibrotic nerve encapsulation). MDP correlated negatively with detrusor collagen and inflammation, and positively with pgp9.5+ axon density and intramural ganglia numbers. These results demonstrate that bladder function can be improved by transfer of obturator nerves to pelvic nerves at 1 yr after decentralization, although the fibrosis and inflammation that developed were associated with decreased contractile function.

AAV1.NT-3 gene therapy for X-linked Charcot–Marie–Tooth neuropathy type 1

AUTHORS

Burcak Ozes, Morgan Myers, Kyle Moss, Jennifer Mckinney, Alicia Ridgley, Lei Chen, Shasha Bai, Charles K. Abrams, Mona M. Freidin, Jerry R. Mendell & Zarife Sahenk

ABSTRACT

X-linked Charcot-Marie-Tooth neuropathy (CMTX) is caused by mutations in the gene encoding Gap Junction Protein Beta-1 (GJB1)/Connexin32 (Cx32) in Schwann cells. Neurotrophin-3 (NT-3) is an important autocrine factor supporting Schwann cell survival and differentiation and stimulating axon regeneration and myelination. Improvements in these parameters have been shown previously in a CMT1 model, TremblerJ mouse, with NT-3 gene transfer therapy. For this study, scAAV1.tMCK.NT-3 was delivered to the gastrocnemius muscle of 3-month-old Cx32 knockout (KO) mice. Measurable levels of NT-3 were found in the serum at 6-month post gene delivery. The outcome measures included functional, electrophysiological and histological assessments. At 9-months of age, NT-3 treated mice showed no functional decline with normalized compound muscle action potential amplitudes. Myelin thickness and nerve conduction velocity significantly improved compared with untreated cohort. A normalization toward age-matched wildtype histopathological parameters included increased number of Schmidt-Lanterman incisures, and muscle fiber diameter. Collectively, these findings suggest a translational application to CMTX1.

A Cadaveric Study on the Utility of the Levator Scapulae Motor Nerve as a Donor for Brachial Plexus Reconstruction

AUTHORS

Eliana B.Saltzman, Karthik Krishnan, Mark J.Winston, Soumen Das DeM, Steve K.Lee, Scott W. Wolfe

ABSTRACT

Purpose

The purpose of the study was to evaluate the utility of the levator scapulae motor nerve (LSN) as a donor nerve for brachial plexus nerve transfer. We hypothesized that the LSN could be transferred to the suprascapular nerve (SSN) or long thoracic nerve (LTN) with a reliable tension-free coaptation and appropriate donor-to-recipient axon count ratio.

Methods

Twelve brachial plexus dissections were performed on 6 adult cadavers, bilaterally. We identified the LSN, spinal accessory nerve (SAN), SSN, and LTN. Each nerve was prepared for transfer and nerve redundancies were calculated. Cross-sections of each nerve were examined histologically, and axons counted. We transferred the LSN to target first the SSN and then the LTN, in a tension-free coaptation. For reference, we transferred the distal SAN to target the SSN and LTN and compared transfer parameters.

Results

Three cadavers demonstrated 2 LSN branches supplying the levator scapulae. The axon count ratio of donor-to-recipient nerve was 1:4.0 (LSN:SSN) and 1:2.1 (LSN:LTN) for a single LSN branch and 1:3.0 (LSN:SSN) and 1:1.6 (LSN:LTN) when 2 LSN branches were available. Comparatively, the axon count ratio of donor-to-recipient nerve was 1:2.5 and 1:1.3 for the SAN to the SSN and the LTN, respectively. The mean redundancy from the LSN to the SSN and the LTN was 1.7 cm (SD, 3.1 cm) and 2.9 cm (SD, 2.8 cm), and the redundancy from the SAN to the SSN and the LTN was 4.5 (SD, 0.7 cm) and 0.75 cm (SD, 1.0 cm).

Conclusions

These data support the use of the LSN as a potential donor for direct nerve transfer to the SSN and LTN, given its adequate redundancy and size match.

Ultrastructural effects of nerve growth factor and betamethasone on nerve regeneration after experimental nerve injury

AUTHORS

Leman Sencar, Mustafa Güven, Dilek Şaker, Tuğçe Sapmaz, Abdullah Tuli, and Sait Polat

ABSTRACT

Peripheral nerve injuries (PNI) are an important health problem in the world. In this study, the effects of nerve growth factor (NGF) and betamethasone on nerve regeneration after sciatic nerve crush injury were examined by footprint analysis, electron microscopic, histomorphometric, and biochemical methods. Fifty Wistar rats were divided into five groups as intact control, experimental control, NGF, betamethasone, and NGF+betamethasone combined treatment groups. After the injury, betamethasone was subcutaneously injected into the lesion area of the treatment groups three times during the first day. NGF was subcutaneously injected into the lesion area of treatment groups for 14 days. Footprint analysis was made on 7, 14, 21, 28, and 35 days and after 6 weeks, tissue samples were obtained from all groups. In the experimental control group, there were severe degenerative changes in most of the axons and myelin sheaths of the nerve fibers. Moreover, an increase of MDA levels and a decrease in SOD activities were found in this group. On the other hand, malondialdehyde (MDA) levels decreased, superoxide dismutase (SOD) activities increased and significant motor functional recovery were found in the combined treatment group. The number of axons, axon diameters, and myelin thickness were significantly greater in the combined treatment group when compared with experimental control and other treatment groups. It was thought that nerve regenerative effects of NGF and anti-inflammatory and/or anti-edematous effects of betamethasone could induce functional recovery in the combined treatment group. In conclusion, combined therapy of NGF and betamethasone may be an effective approach for the treatment of PNI.

Ferroptosis Mediates Cuprizone-Induced Loss of Oligodendrocytes and Demyelination

AUTHORS

Priya Jhelum, Eva Santos-Nogueira, Wulin Teo, Alice Haumont, Isadora Lenoël, Peter K. Stys and Samuel David

ABSTRACT

Multiple sclerosis (MS) is a chronic demyelinating disease of the CNS. Cuprizone (CZ), a copper chelator, is widely used to study demyelination and remyelination in the CNS, in the context of MS. However, the mechanisms underlying oligodendrocyte (OL) cell loss and demyelination are not known. As copper-containing enzymes play important roles in iron homeostasis and controlling oxidative stress, we examined whether chelating copper leads to disruption of molecules involved in iron homeostasis that can trigger iron-mediated OL loss. We show that giving mice (male) CZ in the diet induces rapid loss of OL in the corpus callosum by 2 d, accompanied by expression of several markers for ferroptosis, a relatively newly described form of iron-mediated cell death. In ferroptosis, iron-mediated free radicals trigger lipid peroxidation under conditions of glutathione insufficiency, and a reduced capacity to repair lipid damage. This was further confirmed using a small-molecule inhibitor of ferroptosis that prevents CZ-induced loss of OL and demyelination, providing clear evidence of a copper-iron connection in CZ-induced neurotoxicity. This work has wider implications for disorders, such as multiple sclerosis and CNS injury.