Experimental Studies on the Reinnervation of Posterior Cricoarytenoid Muscle by the Upper Branch of Phrenic Nerve

The purpose of this work was to reestablish the respiratory abduction of theparalyzed vocal cord through reinnervation of the posterior cricoarytenoid(PCA)mus-cle by part of phrenic fibres.In fifteen adult cats the adductor branch of the recurrentlaryngeal nerve(RLN)of the right side was cut and its distal end ligated,while the pro -ximal end was implanted into the PCA muscle belly.The whole RLN was then transectedin the tracheoesophageal groove and its distal stump anastomosed to the upper branchof the phrenic nerve.Various techniques for observation were used on day 40,80 and 150after operation.Direct laryngoscopy showed that the inspiratory abduction of the para-lyzed vocal cord recovered within 40 d in all cats.Eighty days later,a larger abducentmotion of the glottis was observed on the reinnervated side.Abduction was caused byreinnervation of the PCA muscle from phrenic motoneurons,as demonstrated by laryn-geal electromyogram,and the function of diaphragm maintained as revealed by monito-ring of the intrathoracic pressure.

Effects of targeted muscle reinnervation on spinal cord motor neurons in rats following tibial nerve transection

Targeted muscle reinnervation(TMR)is a surgical procedure used to transfer residual peripheral nerves from amputated limbs to targeted muscles,which allows the target muscles to become sources of motor control information for function reconstruction.However,the effect of TMR on injured motor neurons is still unclear.In this study,we aimed to explore the effect of hind limb TMR surgery on injured motor neurons in the spinal cord of rats after tibial nerve transection.We found that the reduction in hind limb motor function and atrophy in mice caused by tibial nerve transection improved after TMR.TMR enhanced nerve regeneration by increasing the number of axons and myelin sheath thickness in the tibial nerve,increasing the number of anterior horn motor neurons,and increasing the number of choline acetyltransferase-positive cells and immunofluorescence intensity of synaptophysin in rat spinal cord.Our findings suggest that TMR may enable the reconnection of residual nerve fibers to target muscles,thus restoring hind limb motor function on the injured side.

Key changes in denervated muscles and their impact on regeneration and reinnervation

The neuromuscular junction becomes progressively less receptive to regenerating axons if nerve repair is delayed for a long period of time. It is difficult to ascertain the denervated muscle＇s residual receptivity by time alone. Other sensitive markers that closely correlate with the extent of denervation should be found. After a denervated muscle develops a fibrillation potential, muscle fiber conduction velocity, muscle fiber diameter, muscle wet weight, and maximal isometric force all decrease; remodeling increases neuromuscular junction fragmentation and plantar area, and expression of myogenesis-related genes is initially up-regulated and then down-regulated. All these changes correlate with both the time course and degree of denervation. The nature and time course of these denervation changes in muscle are reviewed from the literature to explore their roles in assessing both the degree of detrimental changes and the potential success of a nerve repair. Fibrillation potential amplitude, muscle fiber conduction velocity, muscle fiber diameter, mRNA expression levels of myogenic regulatory factors and nicotinic acetylcholine receptor could all reflect the severity and length of denervation and the receptiveness of denervated muscle to regenerating axons, which could possibly offer an important clue for surgical choices and predict the outcomes of delayed nerve repair.

Autonomic reinnervation and functional regeneration in autologous transplanted submandibular glands in patients with severe keratoconjunctivitis sicca

Autologous submandibular gland（SMG） transplantation has been proved to ameliorate the discomforts in patients with severe keratoconjunctivitis sicca. The transplanted glands underwent a hypofunctional period and then restored secretion spontaneously.This study aims to investigate whether autonomic nerves reinnervate the grafts and contribute to the functional recovery, and further determine the origin of these nerves. Parts of the transplanted SMGs were collected from the epiphora patients, and a rabbit SMG transplantation model was established to fulfill the serial observation on the transplanted glands with time. The results showed that autonomic nerves distributed in the transplanted SMGs and parasympathetic ganglionic cells were observed in the stroma of the glands. Low-dense and unevenly distributed cholinergic axons, severe acinar atrophy and fibrosis were visible in the patients＇ glands 4–6 months post-transplantation, whereas the cholinergic axon density and acinar area were increased with time. The acinar area or the secretory flow rate of the transplanted glands was statistically correlated with the cholinergic axon density in the rabbit model, respectively. Meanwhile, large cholinergic nerve trunks were found to locate in the temporal fascia lower to the gland, and sympathetic plexus concomitant with the arteries was observed both in the adjacent fascia and in the stroma of the glands. In summary, the transplanted SMGs are reinnervated by autonomic nerves and the cholinergic nerves play a role in the morphological and functional restoration of the glands. Moreover, these autonomic nerves might originate from the auriculotemporal nerve and the sympathetic plexus around the supplying arteries.

Experience of Cervical Plexus Reinnervation for Patients with Unilateral Recurrent Laryngeal Nerve Invasion or Injury

OBJECTIVE To explore the clinical and therapeutic effects of cervical plexus reinnervation for infiltrated or injured unilateral recurrent laryngeal nerve （URLN）. METHODS Functional neck dissection for removal of differentiated thyroid carcinoma （DTC） in patients was performed, in which cervical plexus reinnervation was adopted for patients with stage I disease and URLN with injury or with tumor invasion. Outcomes of surgery were evaluated by examination under fibrolaryngoscope, and the patients＇ voices were evaluated before and after surgery. RESULTS All cases were followed up for 3 mon-2 years （average 8 mon）. Abductory motion of the vocal cords of 15 patients was completely or partly restored, but 3 patients＇ vocal cords were immovable. The recovery rate of abductory motion of the paralyzed vocal cords was 83.33% （15/18）. The function of phonation in the 16 patients was restored to normal or near normal limits, and their hoarseness was improved significantly. CONCLUSION Cervical plexus-URLN reinnervation should be considered when treating patients with unilateral vocal cord paralysis. Removing the tumor simultaneously with cervical plexus reinnervation during surgery for repair of unilateral recurrent laryngeal nerve injury was convenient and easy to perform with less functional damage compared with other methods of reinnervation. The abductory motion of vocal cord could be satisfactorily restored by this reinnervation. Surgical performance skills and application of neurotrophic drugs were important for the success of the surgery.

Nerve function restoration following targeted muscle reinnervation after varying delayed periods

Targeted muscle reinnervation has been proposed for reconstruction of neuromuscular function in amputees.However,it is unknown whether performing delayed targeted muscle reinnervation after nerve injury will affect restoration of function.In this rat nerve injury study,the median and musculocutaneous nerves of the forelimb were transected.The proximal median nerve stump was sutured to the distal musculocutaneous nerve stump immediately and 2 and 4 weeks after surgery to reinnervate the biceps brachii.After targeted muscle reinnervation,intramuscular myoelectric signals from the biceps brachii were recorded.Signal amplitude gradually increased with time.Biceps brachii myoelectric signals and muscle fiber morphology and grooming behavior did not significantly differ among rats subjected to delayed target muscle innervation for different periods.Targeted muscle reinnervation delayed for 4 weeks can acquire the same nerve function restoration effect as that of immediate reinnervation.

Preemptive targeted muscle reinnervation:the single incision approach should be avoided in trans‑tibial traumatic amputation

Dear Editor,Chronic pain is a significant concern after major lower limb amputations that often preclude prosthetic fitting,decrease ambulation,and impact the quality of life[1,2].In the last decade,targeted muscle reinnervation(TMR)has been proposed as a surgical strategy for treating or preventing symptomatic neuromas and phantomlimb phenomena in major amputees[1].This technique involves the transfer of an amputated mixed-motor and sensory nerve to a nearby recipient motor nerve[1,2].Unlike most surgical strategies that aim to hide or protect the neuroma,TMR gives the amputated nerves“somewhere to go and something to do”[2].In a randomized clinical trial on neuroma and phantom pain,Dumanian et al.[1]demonstrated that TMR reduces amputationrelated chronic pain at 1-year post-intervention when compared with the excision and muscle-burying technique,which remains the current gold standard.Valerio et al.[2]also proposed applying TMR at the time of major limb amputation for preventing chronic pain and found that TMR patients experienced less residual limb pain(RLP)and phantom limb pain(PLP)when compared with untreated amputee controls.

EXPERIMENTAL STUDY ON INFLUENCE OF MUSCLE TENSION TOWARDS THE FUNCTION OF THE SKELETAL MUSCLE FOLLOWING ITS REINNERVATION

Objective To investigate the influence of tension on the function of the denervated skeletalmuscle after its reinnervation. Methods Fifty-four Sprague-Dawley (SD) rats were randomly divided into 3 groups. The left gastrocnemius muscles of the rats were dissected with only the neurovascular pedicles intact; the tib-ial nerves were cut and immediately repaired by epineurial suture. Then the Achilles tendons were isolated and treated accordingly; the Achilles tendon was lengthened by 0. 5cm in lengthened group, shortened by 0. 5cm in shortened group and left alone in normal (control) group. In the 2nd, 4th and 8th week after operation, the isometric twitch contractile force of both the right and the left gastrocnemius muscles were measured; specimens were taken from gastrocnemius muscle for histological study by light microscope. Results In comparison between the groups, the gastrocnemius muscles in the shortened group showed less severe muscle atrophy and connective tissue proliferation, bigger diameter and cross section dimension of the muscle fiber and greater isometric twitch contractile strength of the bilateral gastrocnemius muscles than those in the normal and lengthened groups in all the postoperative periods. Conclusion A proper high tension of the muscle may improve the function of the denervated skeletal muscle after its reinnervation.

REINNERVATION OF CENTRAL RECEPTIVE NEURONS ON THE ROTATED SKIN IN LEECHES

Neuronal connections with their targets manifest selectivity during the development and regeneration. In 1963, Sperry proposed the chemoaffinity hypothesis. Through changing the relative positions of the neuron and its target, we can test this hypothesis by investigating the regeneration. Results from the experiments of changing the neuronal posi-

Reinnervation of hair cells by neural stem cell-derived neurons

Background Replacement of spiral ganglion neurons would be one prioritized step in an attempt to restore sensory neuronal hearing loss.However,the possibility that transplanted neurons could regenerate new synaptic connections to hair cells has not been explored.The objective of this study was to test whether neural stem cell （NSC）-derived neurons can form synaptic connections with hair cells in vitro.Methods NSCs were mechanically separated from the hippocampus in SD rat embryos （E12-E14） and cultured in a serum-free medium containing basic fibroblast growth factor and epidermal growth factor.Rat NSCs were co-cultured with explants of cochlea sensory epithelia obtained from postnatal Day 3 rats under transway filter membrane.Results At Day 3,the NSCs began to show chemotactic differentiation and grew toward cochlea sensory epithelia.After 9-day co-culture,neurites of NSC-derived neurons predominantly elongated toward hair cells.Immunohistochemical analyses revealed the fibers overlapped with synapsin and hair cells,indicating the formation of new synaptic connections.After 14-day culture,triple staining revealed the fibers overlapped with PSD95 （postsynaptic density） which is juxtaposed with CtBP2 （presynaptic vesicle）,indicating the formation of new ribbon synapse.Conclusions NSC-derived neurons can make synaptic connections with hair cells and provide a model for studying synaptic plasticity and regeneration.Whether the newly forming synapse is functional merits further electrophysiological study.

Using posterior part of the deltoid muscle as receptor and quality control with intra-operative electrophysiological examination in targeted muscle reinnervation for high-level upper extremity amputees

To the Editor:Targeted muscle reinnervation(TMR)is a surgical technique of multiple nerve transfers,providing a potential of improved intuitive prosthetic control via surface electromyography(sEMG)in the high-level upper extremity amputees.[1]However,there is a risk that some of the reinnervations might be unsuccessful,especially for the ulnar nerve.[2]Both the quality control of nerve stumps and the receptor are important factors for the surgery.Assessing the nerve stumps during the surgery and finding more muscles as receptor might address the problem.Biceps,triceps,and brachialis muscles were mostly chosen as receptors for reinnervation in the trans-humeral amputees.Pectoralis major and pectoralis minor were mostly chosen as receptors for reinnervation in the shoulder disarticulation patients.

The role of peripheral nerve surgery in a tissue reinnervation

In modern neuroscience,the most relevant is the study of the problem of reinnervation of tissues after severe injuries.Complete restoration of lost physiological functions is still impossible with lesions of peripheral nerves with the formation of extensive diastasis between their proximal and distal sites.In this case,the standard neurorrhaphy cannot be carried out because of the eruption of the filaments during tension and convergence of the ends.To solve this problem,a technique was developed for autotransplantation of the nerve sections,which is still the gold standard for the reconstruction of extensive nerve defects.However,the presence of significant shortcomings led to the development of the doctrine of the direction of regeneration with the help of conduits.Currently,the use of nerve channels is the most promising technology for peripheral nerve repair after trauma.The most actively developing now is the direction of reinnervation,such as neurotization.Neurotization,in some way,combined all the methods of restoring nerves.The overall goal of all these methods—the restoration of extensive nerve defects—allows them to be combined into a new industry:reinnervating neurosurgery.

An analysis of the role of targeted muscle reinnervation(TMR)in quality of life and pain outcomes:a case series

Aim:Targeted muscle reinnervation(TMR)surgery has fundamentally changed the management of patients who have suffered or are about to undergo amputation.Providing nerve stumps with a muscle target has been shown to have profound effects on levels of post-amputation pain in relation to phantom limb pain(PLP)and neuroma pain(NP).The primary objective of this report was to quantify pain parameters for this population and to measure the impact on health-related quality of life(HRQol)before and after TMR surgery.In this case series,we evaluate the role of TMR in addressing both pain and the impact of the surgery on the patient’s quality of life.Methods:A retrospective analysis of 15 upper limb amputee patients who underwent TMR by the Relimb Unit in London,UK.Participants’perceptions of pain were determined using the 11-point numerical(Pain)rating scale(NRS)and HRQoL was calculated using the Euroqol EQ-5D-5L questionnaire at two time points,comparing both pain and perceived quality of life pre and post surgery.The Wilcoxon Signed Rank Test was used for the NRS data and a paired sample t-test was used for the EQ-VAS data.Results:A total of 15 patients completed the evaluation.We observed statistically significant reductions in both PLP(pre-operative mean:7.6,post-operative mean:2.7,P<0.05)and NP(pre-operative mean:6.4,post-operative mean:2.5,P<0.05)in these patients.Similarly,HRQoL observed on the EQ-VAS scale demonstrated a significant improvement in quality of life,from 68 pre-operatively to 78 post-procedure(P<0.05).Conclusion:This is the first quantified evaluation of changes in HRQoL after TMR surgery for upper limb amputation.There appears to be a significant improvement in both HRQoL and overall perception of pain.This finding may have important implications for funding and national resource allocation for TMR surgery.

The neurobiology of targeted muscle reinnervation for post-amputation pain

Targeted muscle reinnervation(TMR)is a peripheral nerve procedure that can prevent and treat postamputation pain.The nerve transfer allows for organized nerve regeneration and repair after amputation surgery.The procedure can successfully prevent neuromas despite large size mismatches between the donor and recipient nerves.Here,we discuss the fundamentals of peripheral nerve injury and regeneration as it pertains to TMR.We propose axonal pruning to explain axon behavior when there are large size mismatches between transferred nerves.Given the increasing use of TMR for amputees,future studies should investigate the basic science of peripheral nerves in TMR.Advances in this field have the potential to significantly improve clinical outcomes for these patients.

Early muscle reinnervation by means of end-to-side neurorrhaphy in an experimental model

Aim:The aim of the study was to provide early muscular reinnervation to avoid muscle atrophy and functional loss in an experimental model.Methods:Fifty rats were divided into five groups.In group 0(control group)only nerve dissection was performed.Total peroneal nerve section was performed in the remaining groups.Immediate end-to-end neurorrhaphy(EEN)was made in group 1.In group 2,an end-to-side neurorrhaphy(ESN)was performed from the tibial nerve to the peroneal nerve.In group 3,a direct EEN,plus an ESN,were used as a nerve graft as a bridge from the donor nerve(tibial);all nerve coaptations were performed through an epineural window.In group 4,only a neurotomy was made without any type of reconstruction.Results:Neural diameters were similar in groups 0,1,and 3(38±1μm,31±6μm,32±3μm).Neural fibers in group 3 had an 18%increase in the number of axons(P<0.001)when compared to group 0.Group 2(28±1μm)and group 4(19±3μm)had diminished diameters with a lower index of muscle regeneration.Animals in group 4 presented with'clawed'lower extremities and had difficulty with ambulation.Neural graft diameters was similar in groups 2 and 3(33±4μm,31±3μm),but axon density was significantly higher in group 3(53±6μm,39±8μm)(P<0.001).Axon density was 36%higher when the combination of EEN and ESN with a neural graft through an epineural window was performed.Conclusion:This study revealed that the combination of EEN and ESN repairs with the addition of a neural graft provides a lower index of muscle fiber destruction,and can be a reliable method for reconstruction in high neural injuries.

Expanding the top rungs of the extremity reconstructive ladder:targeted muscle reinnervation,osseointegration,and vascularized composite allotransplantation

Osseointegration(OI),targeted muscle reinnervation(TMR),and vascularized composite allotransplantation(VCA)are just a few ways by which our reconstructive ladder is evolving.It is important to recognize that amputation does not necessarily denote failure,but surgeons should strive to find ways to provide these patients with means for obtaining better satisfaction and quality of life postoperatively.TMR and OI have added options for mutilating lower extremity injuries that necessitate amputation.More recently,the senior author(Levin LS)described the"penthouse"floor of the reconstructive ladder being VCA.Despite the advances in VCA over the last 20 years,there are many challenges that face this discipline including indications for patient selection,minimizing immunosuppressive regimens,standardizing outcome measures,establishing reliable protocols for monitoring,and diagnosing and managing rejection.Herein,the authors review TMR,OI,and VCA as additional higher rungs of the reconstructive ladder.

A review of pain outcomes following targeted muscle reinnervation in lower extremity limb pain

Approximately 75%experience phantom(PLP),residual(RLP),or general(GLP)limb pain following lower extremity amputation.Targeted muscle reinnervation(TMR)is a peripheral nerve transfer that reroutes amputated nerves to motor endplates that can prevent or treat limb pain.This systematic review summarizes pain outcomes following primary and secondary treatment of lower extremity PLP,RLP,and GLP.Primary literature review of three databases-PubMed,EMBASE,MEDLINE-were used for all articles related to TMR and lower extremity limb pain,querying the same keywords:“targeted muscle reinnervation”AND“pain”.Citations were then reviewed and eliminated if only upper extremities were studied or the study lacked pain outcomes.Citations were categorized as primary or secondary TMR.Pain outcomes,including Numerical Rating Scales(NRS)and Patient-Reported Outcome Measurement Information System(PROMIS)Pain scores,were aggregated when appropriate.Ten studies met all inclusion and exclusion criteria after formal review for a total of 431 extremities,of which 79.1%(n=341 limbs)were lower extremities.Average primary TMR PROMIS scores for PLP and RLP were lower than amputees without primary TMR.Average NRS scores and PROMIS Pain scores in secondary TMR demonstrated improvements in PLP,RLP,and GLP.Primary and Secondary TMR does prevent and improve PLP,RLP,and GLP;however,a minority of studies report quantifiable pain outcomes.All future TMR studies should include validated pain outcomes to better quantify the expected pain and quality of life improvements after lower extremity TMR.

Efficacy of targeted muscle reinnervation for treating and preventing postamputation pain-a systematic review

Aim:Targeted muscle reinnervation(TMR)is a procedure pioneered to improve control of myoelectric prostheses and was fortuitously found to improve postamputation pain by transferring residual nerve ends from an amputated limb to reinnervate motor nerve units in denervated muscles.This study sought to perform a systematic review of the literature regarding the postamputation pain-related outcomes following TMR.Methods:PubMed database was queried using the key term“targeted muscle reinnervation”.Articles were chosen based on the following criteria:(1)clinical studies on TMR;(2)greater than one subject;(3)studies were case-controls,comparative cohort analyses,controlled trials,or randomized controlled trials;and(4)studies included one or more outcomes of interest:prosthetic use and functionality,improvement or persistence of pain,indications,complications,donor nerves,and technical aspects of TMR.Results:Overall,9 studies including 101 upper extremity and 252 lower extremity nerve transfers were analyzed,with nerve transfer type,amputation location,and specific neurotizations reported.Four studies assessed the efficacy of TMR in addressing phantom limb pain(PLP)and residual limb pain(RLP),with 3 out of 4 studies reporting significant improvements in PROMIS(Patient Reported Outcome Measurement Information System)scores in TMR subjects compared to controls.Five additional studies did not analyze PROMIS scores but reported subjective improvements in pain outcomes.Conclusion:Included studies demonstrated TMR had lower maximal pain and pain intensity,behavior and interference compared to the standard of care.Secondary TMR used to treat patients with established painful neuromas also reported improvement in pain compared to baseline.

Advances in upper limb loss rehabilitation:the role of targeted muscle reinnervation and regenerative peripheral nerve interfaces

Upper limb loss results in significant physical and psychological impairment and is a major financial burden for both patients and healthcare services.Current myoelectric prostheses rely on electromyographic(EMG)signals captured using surface electrodes placed directly over antagonistic muscles in the residual stump to drive a single degree of freedom in the prosthetic limb(e.g.,hand open and close).In the absence of the appropriate muscle groups,patients rely on activation of biceps/triceps muscles alone(together with a mode switch)to control all degrees of freedom of the prosthesis.This is a non-physiological method of control since it is non-intuitive and contributes poorly to daily function.This leads to the high rate of prosthetic abandonment.Targeted muscle reinnervation(TMR)reroutes the ends of nerves in the amputation stump to nerves innervating“spare”muscles in the amputation stump or chest wall.These then become proxies for the missing muscles in the amputated limb.TMR has revolutionised prosthetic control,especially for high-level amputees(e.g.,after shoulder disarticulation),resulting in more intuitive,fluid control of the prosthesis.TMR can also reduce the intensity of symptoms such as neuroma and phantom limb pain.Regenerative peripheral nerve interface(RPNI)is another technique for increasing the number of control signals without the limitations of finding suitable target muscles imposed by TMR.This involves wrapping a block of muscle around the free nerve ending,providing the regenerating axons with a target organ for reinnervation.These RPNIs act as signal amplifiers of the previously severed nerves and their EMG signals can be used to control prosthetic limbs.RPNI can also reduce neuroma and phantom limb pain.In this review article,we discuss the surgical technique of TMR and RPNI and present outcomes from our experience with TMR.

Lower extremity amputation:the emerging role of targeted muscle reinnervation(TMR)and regenerative peripheral nerve interface(RPNI)

Lower extremity amputation is increasingly prevalent in the United States,with growing numbers of patients suffering from diabetes and peripheral vascular disease.Amputation has significant functional sequelae as more than half of patients are unable to ambulate at one year postoperatively.Improving mobility and decreasing chronic post-amputation pain can significantly improve the quality of life for these patients and reduce the cost burden on the healthcare system.Plastic and reconstructive surgery has been at the forefront of“reconstructive amputation”,in which nerve pedicles can be surgically guided to decrease painful neuroma formation as well as provide targets for myoelectric prosthesis use.We herein review post-amputation outcomes,epidemiology of chronic,post-amputation pain,and current treatments,including total muscle reinnervation and regenerative peripheral nerve interface,which are at the forefront of multidisciplinary treatment of lower extremity amputees.