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OBJECTIVE To evaluate whether the ultrashort-acting neuromuscular blocking agent gantacurium can be used to blunt evoked laryngospasm in anesthetized cats and to determine the duration of apnea without hemoglobin desaturation. ANIMALS 8 healthy adult domestic shorthair cats. PROCEDURES Each cat was anesthetized with dexmedetomidine and propofol, instrumented with a laryngeal mask, and allowed to breathe spontaneously (fraction of inspired oxygen, 1.0). The larynx was stimulated by spraying sterile water (0.3 mL) at the rima glottidis; a fiberscope placed in the laryngeal mask airway was used to detect evoked laryngospasm. Laryngeal stimulation was performed at baseline; after IV administration of gantacurium at doses of 0.1, 0.3, and 0.5 mg/kg; and after the effects of the last dose of gantacurium had terminated. Duration of apnea and hemoglobin oxygen saturation (measured by means of pulse oximetry) after each laryngeal stimulation were recorded. Neuromuscular block was monitored throughout the experiment by means of acceleromyography on a pelvic limb. RESULTS Laryngospasm was elicited in all cats at baseline, after administration of 0.1 mg of gantacurium/kg, and after the effects of the last dose of gantacurium had terminated. The 0.3 and 0.5 mg/kg doses of gantacurium abolished laryngospasm in 3 and 8 cats, respectively, and induced complete neuromuscular block measured at the pelvic limb; the mean ± SE duration of apnea was 2 ± I minutes and 3 ± 1.5 minutes, respectively. Hemoglobin oxygen saturation did not decrease significantly after administration of any dose of gantacurium. CONCLUSIONS AND CLINICAL RELEVANCE Gantacurium may reduce tracheal intubation-associated morbidity in cats breathing oxygen. [ABSTRACT FROM AUTHOR]

Background: Degenerative myelopathy (DM) in dogs is a progressive neurodegenerative condition that causes white matter spinal cord lesions. These lesions are undetectable on standard magnetic resonance imaging (MRI), limiting diagnosis and monitoring of the disease. Spinal cord lesions cause disruption to the structural integrity of the axons causing water diffusion to become more random and less anisotropic. These changes are detectable by the technique of diffusion tensor imaging (DTI) which is highly sensitive to diffusion alterations secondary to white matter lesion development. Objective: Perform spinal DTI on cohorts of dogs with and without DM to identify if lesions caused by DM will cause a detectable alteration in spinal cord diffusivity that correlates with neurological status. Animals Thirteen dogs with DM and 13 aged‐matched controls. Methods: All animals underwent MRI with DTI of the entire spine. Diffusivity parameters fractional anisotropy (FA) and mean diffusivity (MD) were measured at each vertebral level and statistically compared between groups. Results: Dogs with DM had significant decreases in FA within the regions of the spinal cord that had high expected lesion load. Decreases in FA were most significant in dogs with severe forms of the disease and correlated with neurological grade. Conclusions and Clinical Importance: Findings suggest that FA has the potential to be a biomarker for spinal cord lesion development in DM and could play an important role in improving diagnosis and monitoring of this condition. [ABSTRACT FROM AUTHOR]

OBJECTIVE To create a model of transient unilateral laryngeal paralysis (LP) that will allow the study of cricoarytenoideus dorsalis dysfunction and a method for quantification of varying degrees of LP in dogs. ANIMALS 5 castrated male research Beagles. PROCEDURES Between January and February 2018, dogs were anesthetized and instrumented with a laryngeal mask airway and a flexible endoscope to record the rima glottidis. The left or right recurrent laryngeal nerve (RLn) was localized using ultrasonography and electrical stimulation, then conduction blockade was induced with perineural lidocaine. The normalized glottal gap area (NGGA) was measured before and every 15 minutes after the block. Inspired 10% carbon dioxide (CO2) was administered for 1 minute at each sampling time. The inspiratory increase in NGGA (total and each side) was measured at peak inspiration. The change in hemi-NGGA for the control side versus the anesthetized side was evaluated with a mixed-effect model. RESULTS During CO2 stimulation, the increase in inspiratory hemi-NGGA was consistently less (P < .001) for the treated side (–8% to 13%) versus the control side (49% to 82%). A compensatory increase (larger than at baseline) in the control hemi-NGGA was observed. The total NGGA remained unaffected. CLINICAL RELEVANCE Unilateral local anesthesia of the RLn produced transient unilateral LP with a compensatory increase in the hemiNGGA for the contralateral side. This model could facilitate the evaluation of respiratory dynamics, establishment of a grading system, and collection of other important information that is otherwise difficult to obtain in dogs with LP. [ABSTRACT FROM AUTHOR]

The rat surgical anterior cruciate ligament transection (ACLT) model is commonly used to investigate intra‐articular osteoarthritis (OA) therapies, and histological assessment is often the primary outcome measure. However, histological changes do not always correlate well with clinical outcomes. Therefore, this study evaluated functional outcomes in the rat surgical ACLT model and compared intra‐articular injection volumes ranging from 20 to 50 μl. Unilateral ACLT was surgically induced and static weight‐bearing, mechanical allodynia, motor function, and gait were assessed in four groups of male, Sprague‐Dawley rats (n = 6 per group). Intra‐articular injections of 20 µl Dulbecco's phosphate‐buffered saline (DPBS), 50 µl DPBS, or 50 µl of synthetic biomimetic boundary lubricant were administered once weekly for 3 weeks postoperatively. Structural changes were evaluated histologically at 20 weeks. Rat cadaver knees were injected with 20, 30, 40, or 50 µl of gadolinium solutions and were imaged using magnetic resonance imaging (MRI). Static weight‐bearing, mechanical allodynia, and gait parameters in ACLT groups revealed differences from baseline and naïve controls for 4 weeks post‐ACLT; however, these differences did not persist beyond 6 weeks. Different intra‐articular DPBS injection volumes did not result in functional or histological changes; however, peri‐articular leakage was documented via MRI following 50, 40, and 30 µl but not 20 µl gadolinium injections. Statement of clinical significance: Differences in functional parameters were predominantly restricted to early, postoperative changes in the rat surgical ACLT model despite evidence of moderate histologic OA at 20 weeks. Injection volumes of 20–30 µl are more appropriate for investigating intra‐articular therapies in the rat knee. [ABSTRACT FROM AUTHOR]

Introduction/Aims: While the peripheral nervous system has the inherent ability to recover following injury, results are often unsatisfactory, resulting in permanent functional deficits and disability. Therefore, methods that enhance regeneration are of significant interest. The present study investigates an injectable nerve‐tissue‐specific hydrogel as a biomaterial for nerve regeneration in a rat nerve crush model. Methods: Nerve‐specific hydrogels were injected into the subepineurial space in both uninjured and crushed sciatic nerves of rats to assess safety and efficacy, respectively. The animals were followed longitudinally for 12 wk using sciatic functional index and kinematic measures. At 12 wk, electrophysiologic examination was also performed, followed by nerve and muscle histologic assessment. Results: When the hydrogel was injected into an uninjured nerve, no differences in sciatic functional index, kinematic function, or axon counts were observed. A slight reduction in muscle fiber diameter was observed in the hydrogel‐injected animals, but overall muscle area and kinematic function were not affected. Hydrogel injection following nerve crush injury resulted in multiple modest improvements in sciatic functional index and kinematic function with an earlier return to normal function observed in the hydrogel treated animals as compared to untreated controls. While no improvements in supramaximal compound motor action potential were observed in hydrogel treated animals, increased axon counts were observed on histologic assessment. Discussion: These improvements in functional and histologic outcomes in a rapidly and fully recovering model suggest that injection of a nerve‐specific hydrogel is safe and has the potential to improve outcomes following nerve injury. [ABSTRACT FROM AUTHOR]

Background and Aims: Peripheral nerve injury is common with poor functional recovery and consequent high personal and societal costs. Sciatic nerve transection and assessment of recovery using sciatic functional index (SFI) are widely used. SFI is biologically limited as axonal misdirection of axons supplying flexors and extensors in the hindlimb, after nerve injury can lead to synkinetic innervation and function which does not correspond to the degree of axonal regeneration. Methods: We reevaluated the use of traditional metrics such as print length (PL), toe spread (TS), and intermediate toe spread (ITS) as well as hock angle at mid‐swing as approaches for determining recovery. We used two alternative approaches in discrete cohorts of rats following common peroneal crush injury, transection with repair and critical gap, using transection with ligation as a negative control. We compared walking track analysis (print) with digital capture and kinematics. Results: PL, TS, and ITS varied as expected after injury. The traditional functional index for common peroneal injury using inked prints failed to describe recovery and we derived new indices to describe recovery (all R2 > 0.88, p <.0001) although pre‐injury PFI was never attained by any of the models. Kinematic analysis identified hock angle at mid‐swing as a useful predictor of recovery (p <.0001). Interpretation: Using complementary approaches. [ABSTRACT FROM AUTHOR]

Currently available software tools for automated segmentation and analysis of muscle cross-section images often perform poorly in cases of weak or non-uniform staining conditions. To address these issues, our group has developed the MyoSAT (Myofiber Segmentation and Analysis Tool) image-processing pipeline. MyoSAT combines several unconventional approaches including advanced background leveling, Perona-Malik anisotropic diffusion filtering, and Steger's line detection algorithm to aid in pre-processing and enhancement of the muscle image. Final segmentation is based upon marker-based watershed segmentation. Validation tests using collagen V labeled murine and canine muscle tissue demonstrate that MyoSAT can determine mean muscle fiber diameter with an average accuracy of ~92.4%. The software has been tested to work on full muscle cross-sections and works well even under non-optimal staining conditions. The MyoSAT software tool has been implemented as a macro for the freely available ImageJ software platform. This new segmentation tool allows scientists to efficiently analyze large muscle cross-sections for use in research studies and diagnostics. [ABSTRACT FROM AUTHOR]

Exercise induced intermittent dorsal displacement of the soft palate (DDSP) is a common cause of airway obstruction and poor performance in racehorses. The definite etiology is still unclear, but through an experimental model, a role in the development of this condition was identified in the dysfunction of the thyro-hyoid muscles. The present study aimed to elucidate the nature of this dysfunction by investigating the spontaneous response to exercise of the thyro-hyoid muscles in racehorses with naturally occurring DDSP. Intramuscular electrodes were implanted in the thyro-hyoid muscles of nine racehorses, and connected to a telemetric unit for electromyographic monitoring implanted subcutaneously. The horses were recruited based on upper airway function evaluated through wireless endoscopy during exercise. Five horses, with normal function, were used as control; four horses were diagnosed as DDSP-affected horses based on repeated episodes of intermittent dorsal displacement of the soft palate. The electromyographic activity of the thyro-hyoid muscles recorded during incremental exercise tests on a high-speed treadmill was analyzed to measure the mean electrical activity and the median frequency of the power spectrum, thereafter subjected to wavelet decomposition. The affected horses had palatal instability with displacement on repeated exams prior to surgical implantation. Although palatal instability persisted after surgery, only two of these horses displaced the palate after instrumentation. The electromyographic traces from this group of four horses showed, at highest exercise intensity, a decrease in mean electrical activity and median power frequency, with progressive decrease in the contribution of the high frequency wavelets, consistent with development of thyro-hyoid muscle fatigue. The results of this study identified fatigue as the main factor leading to exercise induced palatal instability and DDSP in a group of racehorses. Further studies are required to evaluate the fiber type composition and metabolic characteristics of the thyro-hyoid muscles that could predispose to fatigue. [ABSTRACT FROM AUTHOR]

Introduction: Controversy exists over the effects of functional electrical stimulation (FES) on reinnervation. We hypothesized that intramuscular FES would not delay reinnervation after recurrent laryngeal nerve (RLn) axonotmesis.Methods: RLn cryo-injury and electrode implantation in ipsilateral posterior cricoarytenoid muscle (PCA) were performed in horses. PCA was stimulated for 20 weeks in eight animals; seven served as controls. Reinnervation was monitored through muscle response to hypercapnia, electrical stimulation and exercise. Ultimately, muscle fiber type proportions and minimum fiber diameters, and RLn axon number and degree of myelination were determined.Results: Laryngeal function returned to normal in both groups within 22 weeks. FES improved muscle strength and geometry, and induced increased type I:II fiber proportion (p = 0.038) in the stimulated PCA. FES showed no deleterious effects on reinnervation.Discussion: Intramuscular electrical stimulation did not delay PCA reinnervation after axonotmesis. FES can represent a supportive treatment to promote laryngeal functional recovery after RLn injury. Muscle Nerve 59:717-725, 2019. [ABSTRACT FROM AUTHOR]

Study Design: Ovine in vivo study.Objective: To perform lateral approach lumbar surgery in an ovine model to administer an injectable riboflavin cross-linked high-density collagen (HDC) gel and to assess its ability to mitigate intervertebral disc (IVD) degeneration after induced annulus fibrosus (AF) injury.Summary Of Background Data: Biological-based injectable gels have shown efficacy in restoring biomechanical, radiographic, and histological parameters in IVD-injured animal models. Riboflavin cross-linked HDC gel has previously demonstrated retention of nucleus pulposus (NP) tissue, reduced loss of disc height, and prevention of terminal cellular degenerative changes in rat-tail spines. However, this biological therapy has never been tested in large animal models.Methods: Forty lumbar IVDs were accessed from eight sheep via lateral approach surgery. IVDs were randomly assigned to healthy control, injury and HDC treatment, or negative control with injury and no treatment. IVD injury was carried out using a drill-bit through the AF followed by needle puncture of the NP. Sheep were followed for 16 weeks and underwent qualitative/quantitative magnetic resonance imaging, x-ray, and histological analyses of collagen and proteoglycan content.Results: The lateral approach to the ovine lumbar spine to deliver HDC gel proved to be safe and reproducible. IVDs treated with the HDC gel revealed less degenerative changes at the microscopic level based on AF and NP histology. However, mean Pfirrmann grade, T2 relaxation time, NP voxel size, and disc height index were not significantly different between the two injury groups.Conclusion: Injectable HDC gel can be administered safely via lateral approach surgery in an ovine AF injury model. IVDs treated with HDC gel demonstrated less degeneration at the microscopic level though radiographic changes were slight when comparing treated to untreated IVDs. Future studies will need to elucidate the role of injury technique and time frame for follow-up in correlating histological and radiographical outcomes.Level Of Evidence: N /A. [ABSTRACT FROM AUTHOR]

Objective To determine if neuromuscular monitoring at the pelvic limb accurately reflects neuromuscular function in the larynx after administration of rocuronium in anesthetized dogs. Study design Prospective experimental study. Animals Six healthy Beagle dogs. Methods Anesthesia was maintained in dogs with isoflurane and a continuous infusion of dexmedetomidine. Rocuronium (0.6 mg kg-1) was administered intravenously to induce neuromuscular block. Train-of-four (TOF) impulses were applied to the left recurrent laryngeal nerve (RLn) and the peroneal nerve (Pn). The evoked TOF ratio (TOFR; T4:T1) was measured with electromyography (EMG) simultaneously at the larynx and at the pelvic limb. Spontaneous recoveries of T1 to 25% (T125%) and 75% (T175%) of twitch height, and to TOFR of 0.70 and 0.90 (TOFR0.90) at each EMG site were compared. Results Data from five dogs were analyzed. Times to T125% were similar at the pelvic limb and larynx when measured by EMG; time to T175% was slower at the larynx by 6 ± 4 minutes (p = 0.012). The larynx had a slower recovery to TOFR0.70 (41 ± 13 minutes) and TOFR0.90 (45 ± 13 minutes) than did the pelvic limb [29 ± 8 minutes (p = 0.011) and 33 ± 9 minutes (p = 0.003), respectively]. When the pelvic limb EMG returned to TOFR0.70 and TOFR0.90, the larynx EMG TOFR0.70 and TOFR0.90 values were 0.32 ± 0.12 (p = 0.001) and 0.38 ± 0.13 (p = 0.001), respectively. Conclusions and clinical relevance After administration of rocuronium, neuromuscular function assessed by EMG recovered approximately 36% slower at the larynx than at the pelvic limb. The results in these dogs suggest that quantitative neuromuscular monitoring instrumented at a pelvic limb may be unable to exclude residual block at the larynx in anesthetized dogs. [ABSTRACT FROM AUTHOR]

Background Quantification of the number of axons reinnervating a target organ is often used to assess regeneration after peripheral nerve repair, but because of axonal branching, this method can overestimate the number of motor neurons regenerating across an injury. Current methods to count the number of regenerated motor neurons include retrograde labeling followed by cryosectioning and counting labeled motor neuron cell bodies, however, the process of sectioning introduces error from potential double counting of cells in adjacent sections. New method We describe a method, retroDISCO, that optically clears whole mouse spinal cord without loss of fluorescent signal to allow imaging of retrograde labeled motor neurons using confocal microscopy. Results Complete optical clearing of spinal cords takes four hours and confocal microscopy can obtain z-stacks of labeled motor neuron pools within 3–5 min. The technique is able to detect anticipated differences in motor neuron number after cross-suture and conduit repair compared to intact mice and is highly repeatable. Comparison with existing method RetroDISCO is inexpensive, simple, robust and uses commonly available microscopy techniques to determine the number of motor neurons extending axons across an injury site, avoiding the need for labor-intensive cryosectioning and potential double counting of motor neuron cell bodies in adjacent sections. Conclusions RetroDISCO allows rapid quantification of the degree of reinnervation without the confounding produced by axonal sprouting. [ABSTRACT FROM AUTHOR]

Objective To evaluate the hypothesis that epidural morphine (0.1 mg kg−1) decreases pain in horses after laparoscopic surgery without adversely affecting gastrointestinal ( GI) motility. Study design Randomized clinical trial. Animals Eighteen horses undergoing laparoscopic cryptorchidectomy under general anesthesia. Methods Horses were randomly assigned to receive either epidural morphine (0.1 mg kg−1) or no epidural before the start of surgery. Pain behaviors were assessed during the first two post-operative days using a numerical rating scale. Barium-filled spheres were administered through a nasogastric tube before anesthesia. GI motility was assessed by recording manure production, by quantitating the spheres in the manure, and by abdominal auscultation of intestinal sounds. Heart rates and cortisol concentrations were also measured during the post-operative period. Results Pain scores increased for 12 hours after surgery in the control group and were significantly higher than in the morphine group for the first 6 hours. Pain scores remained unaltered in the morphine group throughout the observation period. Heart rate and plasma cortisol concentrations did not differ between groups or with time. No signs of colic were observed in any horse. Conclusion and clinical relevance Epidural morphine (0.1 mg kg−1) did not adversely affect GI motility in horses after laparoscopic surgery under general anesthesia. [ABSTRACT FROM AUTHOR]

Objective--To determine the effects of hypoglossal nerve block and electrical stimulation of the thyrohyoideus muscles on position of the larynx and hyoid apparatus in resting horses. Animals--16 healthy horses that underwent hypoglossal nerve block and 5 healthy horses that underwent electrical stimulation of the thyrohyoideus muscles. Procedures--Horses underwent bilateral hypoglossal nerve block or electrical stimulation of the thyrohyoideus muscles. Positions of the basihyoid bone, ossified part of the thyroid cartilage, and articulations of the thyrohyoid bones and thyroid cartilage were determined in radiographic images obtained before and after performance of hypoglossal nerve blocks or during thyrohyoideus muscle stimulation. Radiographic images were obtained with the heads of horses in neutral (thyrohyoideus muscle stimulation) or neutral and extended (hypoglossal nerve block) positions. Radiographic images of horses obtained after performance of hypoglossal nerve blocks were also evaluated to detect dorsal displacement of the soft palate. Results--Hypoglossal nerve blocks did not induce significant changes in the positions of evaluated anatomic sites in radiographic images obtained in neutral or extended head positions. Hypoglossal nerve block did not induce dorsal displacement of the soft palate in horses at rest. Bilateral thyrohyoideus muscle stimulation induced significant dorsal movement (mean ± SD change in position, 18.7 ± 6.8 mm) of the ossified part of the thyroid cartilage; rostral movement of evaluated anatomic structures was small and not significant after thyrohyoideus muscle stimulation. Conclusions and Clinical Relevance--Bilateral electrical stimulation of the thyrohyoideus muscles in horses in this study induced dorsal laryngeal movement. [ABSTRACT FROM AUTHOR]

Objective--To compare effects of 3 methods of topically applied cold treatment (cryotherapy) on digital laminar and venous temperatures in horses. Animals--9 healthy adult Thoroughbreds. Procedures--Thermocouples were placed in palmar digital veins and digital laminae of both forelimbs of horses. Three methods of cryotherapy were applied to the distal aspects of the limbs: wader boot (63-cm-tall vinyl boot filled with ice and water [ice slurry]), ice bag (5-L fluid bag filled with ice slurry), and a gel pack boot (boot containing frozen gel packs). Gel packs and ice slurries were replenished every hour during cryotherapy. The forelimb that received the first treatment was randomly assigned; thereafter, control and treated forelimbs were alternated for each treatment. For each treatment, temperatures were recorded every minute during 15-minute pretreatment, 2-hour treatment, and ≥ 30 minute rewarming periods. Once temperatures had returned to within 3°C below pretreatment values, the experiment was repeated in a similar manner for other cryotherapy methods. Results--Digital venous temperatures were similar to laminar temperatures during each treatment. Ice bag and wader boot treatments caused similar cooling of digits. Gel boot treatment did not cause substantial cooling of digits. Conclusions and Clinical Relevance--Ice bag treatment caused laminar and digital venous cooling equivalent to that of wader boot treatment. Cryotherapy by use of 5-L fluid bags with an ice slurry may be a readily available, practical, and efficient method for prevention of laminitis in horses. Digital laminar and venous temperatures were similar in forelimbs of horses before and during cryotherapy. [ABSTRACT FROM AUTHOR]

Objective-To investigate the efficacy and safety of a low-volume, single-catheter, continuous peripheral neural blockade (CPNB) technique to locally deliver bupivacaine to alleviate signs of severe forelimb pain resulting from experimentally induced tendonitis in horses. Design-Randomized controlled experimental trial. Sample-14 horses and 5 forelimbs from equine cadavers. Procedures-Horses underwent collagenase-induced superficial digital flexor tendonitis in the midmetacarpal region of 1 forelimb. To deliver analgesia, a closed-tip catheter was placed from lateral to medial, approximately 12 cm distal to the accessory carpal bone, between the suspensory ligament and accessory ligament of the deep digital flexor tendon. Success of catheter placement and anesthetic delivery was documented ex vivo in 5 forelimbs from equine cadavers. Effective analgesia in affected forelimbs of horses from continuous (n = 7) versus intermittent (7) local anesthetic delivery (intermittent peripheral neural blockade; IPNB) was compared over a 3-day period. Results-Horses that received CPNB in the affected forelimb were less lame than horses that received IPNB. A lower proportion of CPNB-treated horses had behavioral and physiologic signs of pain, compared with IPNB-treated horses. Neither technique completely blocked the sensation of pain or resulted in swelling in the distal portion of the forelimb, vasodilation, or an increase in lameness. After removal, Staphylococcus aureus was cultured from 1 catheter tip. Conclusions and Clinical Relevance-For short-term treatment, CPNB was more effective than IPNB for reduction in signs of severe pain in the distal aspect of the forelimb of horses. [ABSTRACT FROM AUTHOR]

• SSchwann cells and macrophages play key roles in the aftermath of peripheral nerve injury. • Previous methods of Schwann cell isolation relied on pre-enrichment through cell culture, which alters phenotype. • This paper describes two methods for accurate separation of live Schwann cells and macrophages from injured murine nerve. • Live cells are sorted immediately after harvest, allowing gene expression analysis. • B6;D2-Tg(S100b-eGFP)1Wtj/J mice express GFP in Schwann cells and macrophages. Schwann cells (SC) and macrophages play key roles in the response to peripheral nerve injury (PNI). Accurate isolation of such cells is essential for further analyses that can lead to better understanding of the repair process after PNI. Separation of live SC from the injury site without culture enrichment is necessary for targeted gene expression analysis. Two flow cytometric techniques are presented for rapid enrichment of live SC and macrophages from injured murine peripheral nerve without the need for culture. SC were isolated by fluorescent activated cell sorting (FACS) using transgenic expression of eGFP in SC, or by exclusion of other cell types collected from the injury site. Gene expression analyses of peripheral nerve repair have commonly used whole nerve lysates. Isolating SC allows more accurate understanding of their specific role in repair. SC are commonly enriched from nerve by culture, however this changes gene expression patterns and limits the utility for transcriptomic analysis. The surface marker p75-NTR has variable expression in different SC phenotypes and during the course of injury and repair. Using p75-NTR for SC isolation might enrich only a subset of SC. More stably expressed lineage markers for SC are intracellular and not suitable for sorting for gene expression. The methods used here avoid the requirement for surface marker labeling of SC. Gene expression analysis of sorted cells from both methods showed successful enrichment of SC. Lineage markers such as Map1b, p75-NTR and S100b were enriched in the sorted SC population. SC sorting by eGFP expression showed improved enrichment, particularly of mature myelinating genes, although this could represent sampling of a subset of SC. [ABSTRACT FROM AUTHOR]

Background: Macrophages play a key role in peripheral nerve repair and demonstrate complex phenotypes that are highly dependent on microenvironmental cues.Methods: We determined temporal changes in macrophage gene expression over time using RNA sequencing after fluorescence-activated cell sorting (FACS) macrophage populations from injured peripheral nerve. We identified key upstream regulators and dominant pathways using ingenuity pathway analysis and confirmed these changes with NanoString technology. We then investigate the effects of extreme polarizers of macrophage phenotype (IL4 and IFNγ) on nerve regeneration. We determined macrophage gene expression in vivo at the site of peripheral nerve injury with NanoString technology, and assessed recovery from sciatic nerve injury by cranial tibial muscle weights and retrograde labeling motor neurons in mice with deletion of IL4 or IFNγ receptors.Results: We demonstrate that IL4R and IFNγR deletions provide complementary responses to polarization, and alter expression of genes associated with angiogenesis and axonal extension, but do not influence recovery from peripheral nerve transection at 8 weeks after repair.Conclusions: Overall, this study provides a framework to evaluate the phenotype of macrophages over time, and provides a broader and more precise assessment of gene expression changes than has previously been commonly used. This data suggests ways in which polarization may be modulated to improve repair. [ABSTRACT FROM AUTHOR]