Peripheral Nerve Injury After Deoxycholic Acid (ATX-101) Injection in an Experimental Rat Model.

BACKGROUND: Deoxycholic acid (ATX-101) is a drug administered by subcutaneous injection for local fat reduction. However, ATX-101 treatment has been reported to cause marginal mandibular nerve injury with noticeable functional deficits when targeting submental fat. As a cytolytic agent with some selectivity for adipocytes, ATX-101 may damage the lipid-rich myelin surrounding peripheral nerves. OBJECTIVES: This study seeks to characterize the nerve injection injury from ATX-101 in an experimental rat model. METHODS: Using a rat sciatic nerve injection model, intrafascicular and extrafascicular injections of deoxycholic acid (ATX-101) were compared to lidocaine (positive control) and saline (negative control). Nerves were harvested at a 2-week endpoint for histomorphometric analysis. RESULTS: Cross-sectional area of nerve injury was significantly increased by ATX-101 injection at 75±15% with intrafascicular ATX-101 (p<0.001), 41±21% with extrafascicular ATX-101 (p<0.01), and 38±20% with positive control lidocaine (p<0.01) compared to 7±13% with negative control saline. Demyelinating injury was a significant mechanism of injury in the affected nerve fibers compared to uninjured nerve fibers (p<0.04), but there was no difference in axon-to-myelin area ratio between the lidocaine and ATX-101 cohorts. After two weeks, Wallerian degeneration was evident with only small regenerating nerve fibers present in the ATX-101-injured groups compared to saline (2.54±0.26um vs 5.03±0.44um, p<0.001) in average width. CONCLUSIONS: Deoxycholic acid (ATX-101) is capable of extensive nerve injury in rats. The mechanism of action for ATX-101 does not preferentially target myelin more than other common neurotoxic agents. Appropriate knowledge of surgical anatomy and injection technique is necessary for any practitioners providing ATX-101 injections.

Tumor Regression Following Engineered Polyomavirus-Specific T Cell Therapy in Immune Checkpoint Inhibitor-Refractory Merkel Cell Carcinoma.

Although immune check-point inhibitors (CPIs) revolutionized treatment of Merkel cell carcinoma (MCC), patients with CPI-refractory MCC lack effective therapy. More than 80% of MCC express T-antigens encoded by Merkel cell polyomavirus, which is an ideal target for T-cell receptor (TCR)-based immunotherapy. However, MCC often repress HLA expression, requiring additional strategies to reverse the downregulation for allowing T cells to recognize their targets. We identified TCRMCC1 that recognizes a T-antigen epitope restricted to human leukocyte antigen (HLA)-A*02:01. Seven CPI-refractory metastatic MCC patients received CD4 and CD8 T cells transduced with TCRMCC1 (TTCR-MCC1) preceded either by lymphodepleting chemotherapy or an HLA-upregulating regimen (single-fraction radiation therapy (SFRT) or systemic interferon gamma (IFNγ)) with concurrent avelumab. Two patients who received preceding SFRT and IFNγ respectively experienced tumor regression. One experienced regression of 13/14 subcutaneous lesions with 1 'escape' lesion and the other had delayed tumor regression in all lesions after initial progression. Although TTCR-MCC1 cells with an activated phenotype infiltrated tumors including the 'escape' lesion, all progressing lesions transcriptionally lacked HLA expression. While SFRT/IFNγ did not immediately upregulate tumor HLA expression, a secondary endogenous antigen-specific T cell infiltrate was detected in one of the regressing tumors and associated with HLA upregulation, indicating in situ immune responses have the potential to reverse HLA downregulation. Indeed, supplying a strong co-stimulatory signal via a CD200R-CD28 switch receptor allows TTCR-MCC1 cells to control HLA-downregulated MCC cells in a xenograft mouse model, upregulating HLA expression. Our results demonstrate the potential of TCR gene therapy for metastatic MCC and propose a next strategy for overcoming epigenetic downregulation of HLA in MCC.

Limited Nerve Regeneration across Acellular Nerve Allografts (ANAs) Coincides with Changes in Blood Vessel Morphology and the Development of a Pro-Inflammatory Microenvironment.

The use of acellular nerve allografts (ANAs) to reconstruct long nerve gaps (>3 cm) is associated with limited axon regeneration. To understand why ANA length might limit regeneration, we focused on identifying differences in the regenerative and vascular microenvironment that develop within ANAs based on their length. A rat sciatic nerve gap model was repaired with either short (2 cm) or long (4 cm) ANAs, and histomorphometry was used to measure myelinated axon regeneration and blood vessel morphology at various timepoints (2-, 4- and 8-weeks). Both groups demonstrated robust axonal regeneration within the proximal graft region, which continued across the mid-distal graft of short ANAs as time progressed. By 8 weeks, long ANAs had limited regeneration across the ANA and into the distal nerve (98 vs. 7583 axons in short ANAs). Interestingly, blood vessels within the mid-distal graft of long ANAs underwent morphological changes characteristic of an inflammatory pathology by 8 weeks post surgery. Gene expression analysis revealed an increased expression of pro-inflammatory cytokines within the mid-distal graft region of long vs. short ANAs, which coincided with pathological changes in blood vessels. Our data show evidence of limited axonal regeneration and the development of a pro-inflammatory environment within long ANAs.

Acellular Nerve Allografts in Major Peripheral Nerve Repairs: An Analysis of Cases Presenting With Limited Recovery.

BACKGROUND: Acellular nerve allografts have been used successfully and with increasing frequency to reconstruct nerve injuries. As their use has been expanded to treat longer gap, larger diameter nerve injuries, some failed cases have been reported. We present the histomorphometry of 5 such cases illustrating these limitations and review the current literature of acellular nerve allografts. METHODS: Between 2014 and 2019, 5 patients with iatrogenic nerve injuries to the median or ulnar nerve reconstructed with an AxoGen AVANCE nerve allograft at an outside hospital were treated in our center with allograft excision and alternative reconstruction. These patients had no clinical or electrophysiological evidence of recovery, and allograft specimens at the time of surgery were sent for histomorphological examination. RESULTS: Three patients with a median and 2 with ulnar nerve injury were included. Histology demonstrated myelinated axons present in all proximal native nerve specimens. In 2 cases, axons failed to regenerate into the allograft and in 3 cases, axonal regeneration diminished or terminated within the allograft. CONCLUSIONS: The reported cases demonstrate the importance of evaluating the length and the function of nerves undergoing acellular nerve allograft repair. In long length, large-diameter nerves, the use of acellular nerve allografts should be carefully considered.

Electrical stimulation or tacrolimus (FK506) alone enhances nerve regeneration and recovery after nerve surgery, while dual use reduces variance and combines strengths of each in promoting enhanced outcomes.

INTRODUCTION/AIMS: Repaired nerve injuries can fail to achieve functional recovery. Therapeutic options beyond surgery, such as systemic tacrolimus (FK506) and electrical stimulation (E-stim), can improve recovery. We tested whether dual administration of FK506 and E-stim enhances regeneration and recovery more than either therapeutic alone. METHODS: Rats were randomized to four groups: E-stim, FK506, FK506 + E-stim, and repair alone. All groups underwent tibial nerve transection and repair. Two sets of animals were created to measure outcomes of early nerve regeneration using nerve histology (n = 36) and functional recovery (n = 42) (21- and 42-day endpoints, respectively). Functional recovery was measured by behavioral analyses (walking track and grid walk) and, at the endpoint, muscle mass and force. RESULTS: Dual E-stim and FK506 administration produced histomorphometric measurements of nerve regeneration no different than either therapeutic alone. All treatments were superior to repair alone (FK506, P < .0001; E-stim, P < .05; FK506 + E-stim, P < .05). The E-stim and FK506 + E-stim groups had improved behavioral recovery compared with repair alone (at 6 weeks: E-stim, P < .05; FK506 + E-stim, P < .01). The FK506 group had improved recovery based on walking-track analysis (at 6 weeks: P < .001) and muscle force and mass (P < .05). The concurrent use of both therapies ensured earlier functional recovery and decreased variability in functional outcomes compared with either therapy alone, suggesting a moderate benefit. DISCUSSION: Dual administration of FK506 and E-stim showed minimal additive effects to further improve regeneration or recovery compared with either therapy alone. The data suggest the combination of FK506 and E-stim appears to combine the relative strengths of each therapeutic.

Controlling axonal regeneration with acellular nerve allograft limits neuroma formation in peripheral nerve transection: An experimental study in a swine model.

BACKGROUND: Symptomatic neuromata are a common indication for revision surgery following amputation. Previously described treatments, including traction neurectomy, nerve transposition, targeted muscle re-innervation, and nerve capping, have provided inconsistent results or are technically challenging. Prior research using acellular nerve allografts (ANA) has shown controlled termination of axonal regrowth in long grafts. The purpose of this study was to determine the ability of a long ANA to prevent neuroma formation following transection of a peripheral nerve in a swine model. MATERIALS AND METHODS: Twenty-two adult female Yucatan miniature swine (Sus scrofa; 4-6 months, 15-25 kg) were assigned to control (ulnar nerve transection only, n = 10), treatment (ulnar transection and coaptation of 50 mm ANA, n = 10), or donor (n = 2) groups. Nerves harvested from donor group animals were treated to create the ANA. After 20 weeks, the transected nerves including any neuroma or graft were harvested. Both qualitative (nerve architecture, axonal sprouting) and quantitative histologic analyses (myelinated axon number, cross sectional area of nerve tissue) were performed. RESULTS: Qualitative histologic analysis of control specimens revealed robust axon growth into dense scar tissue. In contrast, the treatment group revealed dwindling axons in the terminal tissue, consistent with attenuated neuroma formation. Quantitative analysis revealed a significantly decreased number of myelinated axons in the treatment group (1232 ± 540) compared to the control group (44,380 ± 7204) (p < .0001). Cross sectional area of nerve tissue was significantly smaller in treatment group (2.83 ± 1.53 mm2 ) compared to the control group (9.14 ± 1.19 mm2 ) (p = .0012). CONCLUSIONS: Aberrant axonal growth is controlled to termination with coaptation of a 50 mm ANA in a swine model of nerve injury. These early results suggest further investigation of this technique to prevent and/or treat neuroma formation.

An In Vivo Model of Human Macrophages in Metastatic Melanoma.

Despite recent therapeutic progress, advanced melanoma remains lethal for many patients. The composition of the immune tumor microenvironment (TME) has decisive impacts on therapy response and disease outcome, and high-dimensional analyses of patient samples reveal the heterogeneity of the immune TME. Macrophages infiltrate TMEs and generally associate with tumor progression, but the underlying mechanisms are incompletely understood. Because experimental systems are needed to elucidate the functional properties of these cells, we developed a humanized mouse model reconstituted with human immune cells and human melanoma. We used two strains of recipient mice, supporting or not supporting the development of human myeloid cells. We found that human myeloid cells favored metastatic spread of the primary tumor, thereby recapitulating the cancer-supportive role of macrophages. We next analyzed the transcriptome of human immune cells infiltrating tumors versus other tissues. This analysis identified a cluster of myeloid cells present in the TME, but not in other tissues, which do not correspond to canonical M2 cells. The transcriptome of these cells is characterized by high expression of glycolytic enzymes and multiple chemokines and by low expression of gene sets associated with inflammation and adaptive immunity. Compared with humanized mouse results, we found transcriptionally similar myeloid cells in patient-derived samples of melanoma and other cancer types. The humanized mouse model described here thus complements patient sample analyses, enabling further elucidation of fundamental principles in melanoma biology beyond M1/M2 macrophage polarization. The model can also support the development and evaluation of candidate antitumor therapies.

Targeting an alternate Wilms' tumor antigen 1 peptide bypasses immunoproteasome dependency.

Designing effective antileukemic immunotherapy will require understanding mechanisms underlying tumor control or resistance. Here, we report a mechanism of escape from immunologic targeting in an acute myeloid leukemia (AML) patient, who relapsed 1 year after immunotherapy with engineered T cells expressing a human leukocyte antigen A*02 (HLA-A2)-restricted T cell receptor (TCR) specific for a Wilms' tumor antigen 1 epitope, WT1126-134 (TTCR-C4). Resistance occurred despite persistence of functional therapeutic T cells and continuous expression of WT1 and HLA-A2 by the patient's AML cells. Analysis of the recurrent AML revealed expression of the standard proteasome, but limited expression of the immunoproteasome, specifically the beta subunit 1i (ß1i), which is required for presentation of WT1126-134. An analysis of a second patient treated with TTCR-C4 demonstrated specific loss of AML cells coexpressing ß1i and WT1. To determine whether the WT1 protein continued to be processed and presented in the absence of immunoproteasome processing, we identified and tested a TCR targeting an alternative, HLA-A2-restricted WT137-45 epitope that was generated by immunoproteasome-deficient cells, including WT1-expressing solid tumor lines. T cells expressing this TCR (TTCR37-45) killed the first patients' relapsed AML resistant to WT1126-134 targeting, as well as other primary AML, in vitro. TTCR37-45 controlled solid tumor lines lacking immunoproteasome subunits both in vitro and in an NSG mouse model. As proteasome composition can vary in AML, defining and preferentially targeting these proteasome-independent epitopes may maximize therapeutic efficacy and potentially circumvent AML immune evasion by proteasome-related immunoediting.

Short-Duration, Pulsatile, Electrical Stimulation Therapy Accelerates Axon Regeneration and Recovery following Tibial Nerve Injury and Repair in Rats.

BACKGROUND: Repair of nerve injuries can fail to achieve adequate functional recovery. Electrical stimulation applied at the time of nerve repair can accelerate axon regeneration, which may improve the likelihood of recovery. However, widespread use of electrical stimulation may be limited by treatment protocols that increase operative time and complexity. This study evaluated whether a short-duration electrical stimulation protocol (10 minutes) was efficacious to enhance regeneration following nerve repair using rat models. METHODS: Lewis and Thy1-green fluorescent protein rats were randomized to three groups: 0 minutes of electrical stimulation (no electrical stimulation; control), 10 minutes of electrical stimulation, and 60 minutes of electrical stimulation. All groups underwent tibial nerve transection and repair. In the intervention groups, electrical stimulation was delivered after nerve repair. Outcomes were assessed using immunohistochemistry, histology, and serial walking track analysis. RESULTS: Two weeks after nerve repair, Thy1-green fluorescent protein rats demonstrated increased green fluorescent protein-positive axon outgrowth from the repair site with electrical stimulation compared to no electrical stimulation. Serial measurement of walking tracks after nerve repair revealed recovery was achieved more rapidly in both electrical stimulation groups as compared to no electrical stimulation. Histologic analysis of nerve distal to the repair at 8 weeks revealed robust axon regeneration in all groups. CONCLUSIONS: As little as 10 minutes of intraoperative electrical stimulation therapy increased early axon regeneration and facilitated functional recovery following nerve transection with repair. Also, as early axon outgrowth increased following electrical stimulation with nerve repair, these findings suggest electrical stimulation facilitated recovery because of earlier axon growth across the suture-repaired site into the distal nerve to reach end-organ targets. CLINICAL RELEVANCE STATEMENT: Brief (10-minute) electrical stimulation therapy can provide similar benefits to the 60-minute protocol in an acute sciatic nerve transection/repair rat model and merit further studies, as they represent a translational advantage.

IL-4 expressing cells are recruited to nerve after injury and promote regeneration.

Interleukin-4 (IL-4) has garnered interest as a cytokine that mediates regeneration across multiple tissues including peripheral nerve. Within nerve, we previously showed endogenous IL-4 was critical to regeneration across nerve gaps. Here, we determined a generalizable role of IL-4 in nerve injury and regeneration. In wild-type (WT) mice receiving a sciatic nerve crush, IL-4 expressing cells preferentially accumulated within the injured nerve compared to affected sites proximal, such as dorsal root ganglia (DRGs), or distal muscle. Immunohistochemistry and flow cytometry confirmed that eosinophils (CD45+, CD11b+, CD64-, Siglec-F+) were sources of IL-4 expression. Examination of targets for IL-4 within nerve revealed macrophages, as well as subsets of neurons expressed IL-4R, while Schwann cells expressed limited IL-4R. Dorsal root ganglia cultures were exposed to IL-4 and demonstrated an increased proportion of neurons that extended axons compared to cultures without IL-4 (control), as well as longer myelinated axons compared to cultures without IL-4. The role of endogenous IL-4 during nerve injury and regeneration in vivo was assessed following a sciatic nerve crush using IL-4 knockout (KO) mice. Loss of IL-4 affected macrophage accumulation within injured nerve compared to WT mice, as well as shifted macrophage phenotype towards a CD206- phenotype with altered gene expression. Furthermore, this loss of IL-4 delayed initial axon regeneration from the injury crush site and subsequently delayed functional recovery and re-innervation of neuromuscular junctions compared to wild-type mice. Given the role of endogenous IL-4 in nerve regeneration, exogenous IL-4 was administered daily to WT mice following a nerve crush to examine regeneration. Daily IL-4 administration increased early axonal extension and CD206+ macrophage accumulation but did not alter functional recovery compared to untreated mice. Our data demonstrate IL-4 promotes nerve regeneration and recovery after injury.

Brief Electrical Stimulation Accelerates Axon Regeneration and Promotes Recovery Following Nerve Transection and Repair in Mice.

BACKGROUND: Clinical outcomes following nerve injury repair can be inadequate. Pulsed-current electrical stimulation (ES) is a therapeutic method that facilitates functional recovery by accelerating axon regeneration. However, current clinical ES protocols involve the application of ES for 60 minutes during surgery, which can increase operative complexity and time. Shorter ES protocols could be a strategy to facilitate broader clinical adoption. The purpose of the present study was to determine if a 10-minute ES protocol could improve outcomes. METHODS: C57BL/6J mice were randomized to 3 groups: no ES, 10 minutes of ES, and 60 minutes of ES. In all groups, the sciatic nerve was transected and repaired, and, in the latter 2 groups, ES was applied after repair. Postoperatively, changes to gene expression from dorsal root ganglia were measured after 24 hours. The number of motoneurons regenerating axons was determined by retrograde labeling at 7 days. Histomorphological analyses of the nerve were performed at 14 days. Function was evaluated serially with use of behavioral tests up to 56 days postoperatively, and relative muscle weight was evaluated. RESULTS: Compared with the no-ES group, both ES groups demonstrated increased regeneration-associated gene expression within dorsal root ganglia. The 10-minute and 60-minute ES groups demonstrated accelerated axon regeneration compared with the no-ES group based on increased numbers of labeled motoneurons regenerating axons (mean difference, 202.0 [95% confidence interval (CI), 17.5 to 386.5] and 219.4 [95% CI, 34.9 to 403.9], respectively) and myelinated axon counts (mean difference, 559.3 [95% CI, 241.1 to 877.5] and 339.4 [95% CI, 21.2 to 657.6], respectively). The 10-minute and 60-minute ES groups had improved behavioral recovery, including on grid-walking analysis, compared with the no-ES group (mean difference, 11.9% [95% CI, 3.8% to 20.0%] and 10.9% [95% CI, 2.9% to 19.0%], respectively). There was no difference between the ES groups in measured outcomes. CONCLUSIONS: A 10-minute ES protocol accelerated axon regeneration and facilitated functional recovery. CLINICAL RELEVANCE: The brief (10-minute) ES protocol provided similar benefits to the 60-minute protocol in an acute sciatic nerve transection/repair mice model and merits further studies.

Long Acellular Nerve Allografts Cap Transected Nerve to Arrest Axon Regeneration and Alter Upstream Gene Expression in a Rat Neuroma Model.

BACKGROUND: Treatments to manage painful neuroma are needed. An operative strategy that isolates and controls chaotic axonal growth could prevent neuroma. Using long acellular nerve allograft to "cap" damaged nerve could control axonal regeneration and, in turn, regulate upstream gene expression patterns. METHODS: Rat sciatic nerve was transected, and the distal nerve end was reversed and ligated to generate a model end-neuroma. Three groups were used to assess their effects immediately following this nerve injury: no treatment (control), traction neurectomy, or 5-cm acellular nerve allograft cap attached to the proximal nerve. Regeneration of axons from the injured nerve was assessed over 5 months and paired with concurrent measurements of gene expression from upstream affected dorsal root ganglia. RESULTS: Both control and traction neurectomy groups demonstrated uncontrolled axon regeneration revealed using Thy1-GFP rat axon imaging and histomorphometric measures of regenerated axons within the most terminal region of regenerated tissue. The acellular nerve allograft group arrested axons within the acellular nerve allograft, where no axons reached the most terminal region even after 5 months. At 5 months, gene expression associated with regeneration and pain sensitization, including Bdnf, cfos, and Gal, was decreased within dorsal root ganglia obtained from the acellular nerve allograft group compared to control or traction neurectomy group dorsal root ganglia. CONCLUSIONS: Long acellular nerve allografts to cap a severed nerve arrested axon regeneration within the acellular nerve allograft. This growth arrest corresponded with changes in regenerative and pain-related genes upstream. Acellular nerve allografts may be useful for surgical intervention of neuroma.

Neuroma Management: Capping Nerve Injuries With an Acellular Nerve Allograft Can Limit Axon Regeneration.

Background: Management of painful neuromas continues to challenge clinicians. Controlling axon growth to prevent neuroma has gained considerable traction. A logical extension of this idea is to therefore develop an approach to control and arrest axon growth. Given the limits in axonal regeneration across acellular nerve allografts (ANAs), these constructs could provide a means to reliably terminate axon regeneration from an injured nerve. The purpose of this study was to determine if attaching an ANA to an injured nerve could provide a means to control and limit axon regeneration in a predictable manner. Methods: Twenty (20) adult rats received a sciatic nerve transection, where only the proximal nerve was repaired using an ANA of variable length (0.5, 2.5, and 5.0 cm) or left unrepaired (control). The nerves were harvested 5 weeks post-operatively for gross and histomorphometric analysis. The extent of myelinated axons in regenerated tissue was quantified. Results: At 5 weeks, limited axon regeneration within the ANAs was observed. All lengths of ANAs lead to reduced myelinated axon numbers in the most terminal tissue region compared to untreated injured nerve (P = .002). Additionally, ANA length 2.5 cm or greater did not contain any axons at the most terminal tissue region. Conclusions: This study demonstrates a proof of concept that ANAs attached to the proximal end of an injured nerve can limit axon growth in a controlled manner. Furthermore, the extent of axon growth from the injured nerve into the ANA is dependent on the ANA length.

Autoimmunity and NMDA receptor in brain disorders: Where do we stand?

Over the past decades, the identification of autoimmune encephalitis in which patients express autoantibodies directed against neurotransmitter receptors has generated great hope to shed new light on the molecular mechanisms underpinning neurological and psychiatric conditions. Among these autoimmune encephalitides, the discovery of autoantibodies directed against the glutamatergic NMDA receptor (NMDAR-Ab), in the anti-NMDAR encephalitis, has provided some key information on how complex neuropsychiatric symptoms can be caused by a deficit in NMDAR signalling. Yet, NMDAR-Abs have also been detected in several neurological and psychiatric conditions, as well as in healthy individuals. In addition, these various NMDAR-Abs appear to have different molecular properties and pathogenicities onto receptors and synaptic functions. Here, we discuss the current view on the variety of NMDAR-Abs and, in particular, how these autoantibodies can lead to receptor dysfunction in neuronal networks. Since our mechanistic understanding on patients' NMDAR-Abs is still in its infancy, several complementary processes can be proposed and further in-depth molecular and cellular investigations will surely reveal key insights. Autoantibodies represent a great opportunity to gain knowledge on the etiology of neuropsychiatric disorders and pave the way for innovative therapeutic strategies. ONE SENTENCE SUMMARY: Current view on patients' autoantibody against NMDAR.

New in vitro highly cytotoxic platinum and palladium cyanoximates with minimal side effects in vivo.

Several biologically active bivalent Pd and Pt complexes with two structurally similar cyanoxime ligands abbreviated as H(DECO): 2-oximino-2-cyano-N,N'-diethylacetamide, and H(PyrCO): 2-oximino-2-cyan-N-pyrrolidine acetamide were synthesized and characterized using spectroscopic methods, thermal analysis and X-ray crystallography. Structures revealed planar cis-geometry of studied complexes. Freshly obtained Pt(DECO)2, Pd(DECO)2, Pt(PyrCO)2 and Pd(PyrCO)2 complexes were used in for in vitro cytotoxicity assays using two different etiology human cancer cell lines HeLa and WiDr cells. Investigated compounds showed cytotoxicity levels at or above cisplatin. Pt(DECO)2 was also tested in vivo in healthy C57BL/6 mice. The complex was administered at three different dosage (0, 7.5, 15 mg/kg, i.p. once/week), over a total period of 8 weeks. No changes were observed in the animal weight in the treated mice compared to the control dextrose-treated group. The levels of erythrocytes, leukocytes, and hemoglobin were within the normal level suggesting low myelotoxicity. Negligible cardiotoxicity was observed from the histological evaluation of the hearts from the treated animals. Results from the tail nerve conduction velocity (NCV) and nerve histomorphometry suggested no impact of Pt(DECO)2 on peripheral nerves. The complex, however, induced certain hepatotoxicity and lead to the elevation of IL-6, a pro-inflammatory cytokine. Overall, Pt(DECO)2 showed minimal in vivo toxicity, thus presenting a promising candidate for future testing in animal models of cancer.

Evaluation for Late Nerve Transfer Surgery in Spinal Cord Injury: Predicting the Degree of Lower Motor Neuron Injury.

PURPOSE: Nerve transfer surgery is used to restore upper extremity function following cervical spinal cord injury (SCI) with substantial variation in outcomes. The injury pattern in SCI is complex and can include isolated upper motor neuron (UMN) and combined UMN/lower motor neuron (LMN) dysfunction. The purpose of the study was to determine the most effective diagnostic technique for determining suitable candidates for nerve transfer surgery in SCI. METHODS: Medical records were reviewed of patients who had nerve transfers to restore upper extremity function in SCI. Data collected included (1) preoperative clinical examination and electrodiagnostic testing; (2) intraoperative neuromuscular stimulation (NMS); and (3) nerve histopathology. Preoperative, intraoperative, and postoperative data were compared to identify predictors of isolated UMN versus combined UMN/LMN injury patterns. RESULTS: The study sample included 22 patients with 50 nerve transfer surgeries and included patients ranging from less than 1 year to over a decade post-SCI. Normal recipient nerve conduction studies (NCS) before surgery corresponded to the intraoperative presence of recipient NMS and postoperative histopathology that showed normal nerve architecture. Conversely, abnormal recipient NCS before surgery corresponded with the absence of recipient NMS during surgery and patterns of denervation on postoperative histopathology. Normal donor preoperative manual muscle testing corresponded with the presence of donor NMS during surgery and normal nerve architecture on postoperative histopathology. An EMG of corresponding musculature did not correspond with intraoperative donor or recipient NMS or histopathological findings. CONCLUSIONS: NCS better predict patterns of injury in SCI than EMG. This is important information for clinicians evaluating people for late nerve transfer surgery even years post-SCI. TYPE OF STUDY/LEVEL OF EVIDENCE: Diagnostic II.

T cell receptor gene therapy targeting WT1 prevents acute myeloid leukemia relapse post-transplant.

Relapse after allogeneic hematopoietic cell transplantation (HCT) is the leading cause of death in patients with acute myeloid leukemia (AML) entering HCT with poor-risk features1-3. When HCT does produce prolonged relapse-free survival, it commonly reflects graft-versus-leukemia effects mediated by donor T cells reactive with antigens on leukemic cells4. As graft T cells have not been selected for leukemia specificity and frequently recognize proteins expressed by many normal host tissues, graft-versus-leukemia effects are often accompanied by morbidity and mortality from graft-versus-host disease5. Thus, AML relapse risk might be more effectively reduced with T cells expressing receptors (TCRs) that target selected AML antigens6. We therefore isolated a high-affinity Wilms' Tumor Antigen 1-specific TCR (TCRC4) from HLA-A2+ normal donor repertoires, inserted TCRC4 into Epstein-Bar virus-specific donor CD8+ T cells (TTCR-C4) to minimize graft-versus-host disease risk and enhance transferred T cell survival7,8, and infused these cells prophylactically post-HCT into 12 patients ( NCT01640301 ). Relapse-free survival was 100% at a median of 44 months following infusion, while a concurrent comparative group of 88 patients with similar risk AML had 54% relapse-free survival (P = 0.002). TTCR-C4 maintained TCRC4 expression, persisted long-term and were polyfunctional. This strategy appears promising for preventing AML recurrence in individuals at increased risk of post-HCT relapse.

Comparison of Myelin-Associated Glycoprotein With Vincristine for Facial Nerve Inhibition After Bilateral Axotomy in a Transgenic Thy1-Gfp Rat Model.

IMPORTANCE: Aberrant synkinetic movement after facial nerve injury can lead to prominent facial asymmetry and resultant psychological distress. The current practices of neuroinhibition to promote greater facial symmetry are often temporary in nature and require repeated procedures. OBJECTIVE: To determine whether myelin-associated glycoprotein (MAG), a specific neuroinhibitor, can prevent neuroregeneration with efficacy comparable with that of vincristine, a well-established neurotoxin. DESIGN, SETTING, AND PARTICIPANTS: Rats transgenic for Thy-1 cell surface antigen-green fluorescent protein (Thy1-Gfp) were randomized into 3 groups. Each rat received bilateral crush axotomy injuries to the buccal and marginal mandibular branches of the facial nerves. The animals received intraneural injection of saline, MAG, or vincristine. MAIN OUTCOMES AND MEASURES: The animals were imaged via fluorescent microscopy at weeks 1, 3, 4, and 5 after surgery. Quantitative fluorescent data were generated as mean intensities of nerve segments proximal and distal to the axotomy site. Electrophysiological analysis, via measurement of compound muscle action potentials, was performed at weeks 0, 3, 4, and 5 after surgery. RESULTS: A total of 12 rats were included in the study. Administration of MAG significantly reduced fluorescent intensity of the distal nerve in comparison with the control group at week 3 (mean [SD], MAG group: 94 [11] intensity units vs control group: 130 [11] intensity units; P < .001), week 4 (MAG group: 81 [19] intensity units vs control group: 103 [9] intensity units; P = .004), and week 5 (MAG group: 76 [10] intensity units vs control group: 94 [10] intensity units; P < .001). In addition, rats treated with MAG had greater fluorescent intensity than those treated with vincristine at week 3 (mean [SD], MAG group: 94 [11] intensity units vs vincristine group: 76 [6] intensity units; P = .03), although there was no significant difference for weeks 4 and 5. At week 5, both MAG and vincristine demonstrated lower distal nerve to proximal nerve intensity ratios than the control group (control group, 0.94; vs MAG group, 0.82; P = .01; vs vincristine group; 0.77; P < .001). There was no significant difference in amplitude between the experimental groups at week 5 of electrophysiological testing. CONCLUSIONS AND RELEVANCE: Lower facial asymmetry and synkinesis are common persistent concerns to patients after facial nerve injury. Using the Thy1-Gfp rat, this study demonstrates effective inhibition of neuroregeneration via intraneural application of MAG in a crush axotomy model, comparable with results with vincristine. By potentially avoiding systemic toxic effects of vincristine, MAG demonstrates potential as an inhibitor of neural regeneration for patients with synkinesis. LEVEL OF EVIDENCE: NA.

Increasing Nerve Autograft Length Increases Senescence and Reduces Regeneration.

BACKGROUND: Nerve grafting with an autograft is considered the gold standard. However, the functional outcomes of long (>3 cm) nerve autografting are often poor. The authors hypothesized that a factor contributing to these outcomes is the graft microenvironment, where long compared to short autografts support axon regeneration to different extents. METHODS: A rat sciatic nerve defect model was used to compare regeneration in short (2 cm) and long (6 cm) isografts. Axon regeneration and cell populations within grafts were assessed using histology, retrograde labeling of neurons regenerating axons, immunohistochemistry, quantitative reverse transcriptase polymerase chain reaction, and electron microscopy at 4 and/or 8 weeks. RESULTS: At 8 weeks, for distances of both 1 and 2 cm from the proximal coaptation (equivalent regenerative distance), long isografts had reduced numbers of regenerated fibers compared with short isografts. Similarly, the number of motoneurons regenerating axons was reduced in the presence of long isografts compared with short isografts. Considering the regenerative microenvironments between short and long isografts, cell densities and general populations within both short and long isografts were similar. However, long isografts had significantly greater expression of senescence markers, which included senescence-associated ß-galactosidase, p21, and p16, and distinct chromatin changes within Schwann cells. CONCLUSIONS: This study shows that axon regeneration is reduced in long compared with short isografts, where long isografts contained an environment with an increased accumulation of senescent markers. Although autografts are considered the gold standard for grafting, these results demonstrate that we must continue to strive for improvements in the autograft regenerative environment.

Short-Term Effects of Titanium Dioxide Nanofiber on the Renal Function of Male Sprague Dawley Rats.

Titanium dioxide nanofiber (TDNF) is widely used in the manufacture of various household products, including cosmetics. As a result, the possibility exists for TDNFs to affect human health. Because the kidneys are responsible for filtering out waste from the blood, the goal of the present study was to investigate the short-term effects of TDNF on kidney function of male Sprague Dawley rats. To achieve study objectives, 6- to 7-wk-old male rats were exposed via oral gavage to a total of 0, 40, and 60 parts per million of TDNF for 2 wk. The TDNF was fabricated by electrospinning and then dissolved in water. We measured serum concentration of lactate dehydrogenase, renal histopathology, identification of TDNF in kidney tissue via scanning electron microscopy, and quantitative amounts of titanium-47 in kidney tissue. We also measured specific gene-expression analysis of transcripts involved in apoptosis, inflammation, cell-division regulation, cell structure, and motility. Results showed a slight dose-dependent reduction in renal weight. In contrast, a concentration-dependent elevation in titanium-47 amounts was noted in kidney tissue. We found no significant differences in histopathological patterns. Gnat3 and Hepacam3 were up-regulated in TDNF-treated groups. Up-regulation of NF-κB likely indicated the involvement of renal-tissue inflammation via an independent mechanism. Similarly, Gadd45-α was significantly overexpressed in kidney tissues. This transcript was previously increased following stressful growth-arrest conditions and treatment with DNA-damaging agents. Our overall results suggest marginal renal toxicity in Sprague Dawley rats after ingesting TDNF.