Decellularized allografts as an alternative for reconstruction of large inferior alveolar nerve defects: a systematic review

Background Inferior alveolar nerve (IAN) injuries are a clinical problem with devastating consequences, causing temporary or permanent paresthesia, significantly affecting the patient's quality of life. Despite morbidity, side effects and controversy regarding its results, autologous nerve grafting is still the main treatment for these type of lesions. However, due to advances in knowledge about nerve damage and with the aim of preventing the described problems of autografts, new treatment alternatives based on decellularized allografts have emerged. The aim of this systematic review was to evaluate the reported efficacy of decellularized allografts for the treatment of IAN damage. Material and Methods We performed a systematic search in Pubmed, Scopus and Web of Science databases following the PRISMA guidelines. Cohort studies, randomized or non-randomized clinical studies, prospective or retrospective studies, without age limits and language restriction that included human subjects who received decellularized allograft as treatment for IAN damage were included. Results Six articles met the inclusion criteria and were included for data analysis. In all 6 articles, resolution of IAN damage was observed in more than 85% of patients after a 12-month follow-up period, and in 2 of them, complete resolution was observed in 100% of their patients at longer follow-ups. Conclusions Decellularized allograft appears to be a promising alternative to resolve IAN lesions, without requiring a nerve autograft procedure. However, more randomized clinical trials are needed to validate adequate treatment modalities with decellularized allografts. Key words:Decellularized allograft, alveolar inferior nerve, allograft, injury.


Introduction
The inferior alveolar nerve (IAN) is one of the most important and voluminous terminal sensory nerves of the mandibular nerve (V3) of the trigeminal nerve. It runs inferiorly and anteriorly and penetrates the mandibular canal. The latter can present two dispositions. In the most frequent, the IAN runs to the mentonian foramen, dividing into two terminal branches, the mentonian nerve and the inferior dental plexus. The second arrangement is more infrequent, where the nerve divides into two terminal branches at the entrance of the mandibular canal; the mentonian nerve and the inferior dental nerve. Both arrangements innervate the buccal and vestibular mucosa, the vermilion of the lower lip and the cutaneous skin overlying the chin region (1,2). Injuries to the IAN are a clinical problem with devastating consequences, as they affect the function of the nerve involved and may cause temporary or permanent paresthesia. This significantly affects the patient's quality of life, altering speech, taste, chewing, tooth brushing and the ability to maintain lip competence for food retention and swallowing (1,(3)(4)(5)(6)(7), resulting in severe disability with important social, personal and psychological effects (4)(5)(6)(8)(9)(10)(11)(12)(13). According to Seddon's classification there are three types of nerve injury: (i) neuropraxia, where nerve conduction is interrupted at the site of injury preserving the anatomy of the nerve and axon, (ii) axonotmesis, which corresponds to complete interruption of the axon and myelin sheath preserving the endoneurium, (iii) neurotmesis, where complete interruption of the axon and its myelin sheath is evident, without preservation of the endoneurium, preventing spontaneous regeneration (14)(15)(16)(17) IAN lesions can be caused by a great variety of injuries, being the extraction of impacted teeth the most frequent cause, corresponding to 22% of the cases (18). Of these, 25% do not achieve complete recovery during the first year, and 0.9% remain with permanent alterations (19). Other causes that can injure this nerve include the placement of dental implants, orthognathic surgical procedures (6), removal of benign or malignant tumors (6,20), endodontic therapy, injections of local anesthesia and as a direct consequence of maxillofacial trauma and/or surgical interventions for its repair (3). There are several treatment options for peripheral nerve injuries when there is a loss of nerve continuity at both ends. When a tension-free repair is desired, direct neurorrhaphy should be the procedure of choice (1). However, if it is not possible to perform this technique, the first option is the use of an autologous nerve graft (1,5,6,8), considered the gold standard for nerve grafts. This graft acts as a scaffold that does not produce immunologic reactions and provides neurotrophic factors and Schwann cells, both important for axonal regeneration. For this procedure, donor tissue is usually ob-tained from the sural nerve and/or the greater auricular nerve (1,5,6,8,14,15,17,18,21), reporting a range of nerve recovery between 87.3% and 100% (22,23). However, this technique is associated with high donor site morbidity, as it requires a secondary surgical procedure to remove the donated nerve tissue (1,5,6,8,15,24,25). Because of the need to minimize autograft complications, such as the risk of neuromas formation, cutaneous scarring, and loss of sensation, the use of conduits as scaffolds to bridge nerve gaps without the interference of a nerve graft has been explored (8,15,17,26). However, this technique has limited applications due to variability in the reported results and its difficulty to be used in small nerve gaps (1,5,8,14,15,19). Other alternative for the treatment of peripheral nerve defects are nerve allografts (4,15,17). These can cover a nerve gap of up to 70 mm in length, and due to the neurotrophic effect they provide, they seem to be more effective than conduits. In addition, they do not need a donor site, thus have a reduced morbidity compared autologous nerve grafts (4,15,20). In recent years, non-immunogenic nerve allografts have been used with promising results (8,15,20). The term non-immunogenic refers to decellularized nerve allografts, which retain the nerve tissue framework but are inert to the body, since they were previously processed (4,8,17,20). The process of forming these allografts consists of repeated freeze-thaw cycles, exposure to radiation, prolonged storage in University of Wisconsin cold solution, and decellularization with detergents (8,17). The resulting processed allograft retains the native architecture within the original nerve fascicle and epineural scaffold, which comprises extracellular matrix proteins (laminin, fibronectin, and glycosaminoglycans) (8,20). These proteins, in addition to the native microscopic structure, provide natural axonal growth signals for guided growth, which are not currently found in hollow tube conduits (20). Because the studies reported to date are few and heterogeneous, this study aims to analyze and synthesize the information reported in the scientific literature through a qualitative systematization, in relation to the use of decellularized allografts in IAN defects, and their application as a promising alternative for optimal sensory recovery in the maxillofacial area.

Material and Methods
-Study design The following review was performed following the recommendations from PRISMA. The research question to be answered was: "Are decellularized allografts an effective alternative for the reconstruction of nerve defects associated with the inferior alveolar nerve?". -Eligibility Criteria The eligibility criteria used in the selection of studies and identified articles eligible for full review. Disagreements were resolved under consensus and discussion by the two reviewers with a third reviewer who acted as a judge to settle any disagreements (D.M.R).
-Data extraction Several variables were considered, which were tabulated in an Excel spreadsheet and presented as tables and/or figures.
-Risk of bias To assess the risk of bias of the included studies, we used the Newcastle-Ottawa scale, which allowed us to analyze and calculate a quality score for each selected manuscript based on three main components: (a) selection of the study groups, (b) comparability of the groups, and (c) assessment of the outcome or exposure. The minimum score corresponds to 0 and the maximum to 9, the latter representing the highest methodological quality.

Results
Initially, we identified 86 potential articles, 15 of which corresponded to duplicates and were eliminated. The remaining 71 studies were subjected to title and abstract review, which left a total of 12 potential manuscripts for full-text evaluation. Finally, 6 studies did not meet the inclusion criteria, so 6 articles were included for analysis (Fig. 1).
were: full text available, no language restriction, studies reporting the use of decellularized allografts for inferior alveolar nerve reconstruction, cohort studies, clinical studies (randomized or nonrandomized), prospective, comparative and retrospective studies without age limits. Animal studies and articles reporting the use of decellularized allografts in other nerves unrelated to the research question, narrative reviews, systematic reviews, and in vitro or animal studies were excluded.  The studies included in our study were published between the years 2011 -2020. Six came from the United States. In terms of study design, 1 study corresponded to a case report, 3 to retrospective cohort studies, 1 to a case-control study, and 1 to a case series study. In 4 of the 6 studies was a predominance of females over males (4,5,27,28) and the age range was between 9 and 67 years (6) ( Table 1). The most common cause of IAN injury was ameloblastoma resection (in 3 out of the 6 articles) (4,20,27), followed by complications related to molar exodontias (5,28), post implant installation iatrogenesis and resection of other tumors (6) ( Table 2).
All included studies reported the use of decellularized allograft from AxoGen Inc (Alachua FL). Regarding the time interval between the lesion and the surgical procedure, in 3/6 articles the IAN reconstruction was immediate (4,20,27), in 2/6 after 8 and 9 months (5,28), and in one study they reported two time periods, immediate reconstruction and after a period of 8 and 9 months (6). Regarding the size of the nerve defect, the gaps ranged from 2 mm (6) to 70 mm (4,6,20,27). To determine functional sensory recovery (FSR), in 5/6 articles the Medical Research Council System (MRCS) scale was used as the preferred classification system (4,5,20,27,28), while in 1/6 the Neurosensory Test    (5,20). In the remaining study, only 11% of patients achieved RSF (n=18) (27). Follow-up time ranged from 3 months (4,6,27) to 17.7 months (20). No postoperative complications associated with the nerve graft were reported ( Table 2). -Risk of bias All studies, except of one (4), determined how the selection of the individuals was performed, so the selection item presented low risk of bias. The results were adequately reported in all studies. One article lacked comparability based on design and analysis (28). The overall risk of bias scheme is presented in Fig. 2.

Discussion
The processed nerve allograft (PNA) is a promising option for IAN reconstruction, since unlike the autograft (considered the gold standard), has greater advantages in terms of morbidity, biocompatibility, convenience of use and supply abundance (29). Currently, Avance Nerve Graft (Axogen Inc, Alachua, FL) is the only PNA available in the market (30). This PNA consists of an extracellular matrix scaffold created from donated human peripheral nerve tissue that has been predegenerated, decellularized and sterilized (1). Decellularization and sterilization of the allograft significantly reduces the risk of immune rejection and thus eliminates the need for immunosuppressive therapy (1,20). The allograft preserves the nerve architecture and extracellular matrix microenvironment, which in turn favors natural guided nerve regrowth, whereas conduits do not have these characteristics (1,20). This review suggests a positive result in surgical therapy with PNA as treatment alternative for IAN damage, even in patients who underwent chemo and radiotherapy prior to surgery after the excision of a malignant lesion (4). In all studies, a significant improvement was observed in more than 85% of the patients (4)(5)(6)20,27,28) and in 2/6 studies a resolution was achieved in 100% of the patients (4,28). Regarding the improvement of the lesions, only 1/6 study reported 1 patient without improvement, nevertheless, this patient evidenced a therapeutic failure in the bone graft that was performed (20). This suggests that the treatment failure was due to infection of the bone graft and not due to the nerve allograft used.  (20). This suggests that time is important in the resolution of IAN lesions treated with PNA. Ziccardi et al. considered the time elapsed since the initial injury as an important factor for success. If the time elapsed between injury and surgery is excessive, exceeding a time limit of up to 10 weeks, it could be a contraindication for surgical intervention with ANP, since repairs that are completed before this period of time have a better prognosis (31). Nevertheless, this is not clear. Shanti et al., intervened patients 8 months after the initial surgery causative of the nerve defect, obtaining S3 results on the MRCS scale (please see Table  3 for more details on the MRCS scale) (28). Yampolsky et al, reported a time range between initial injury to surgery of 2 to 923 days, obtaining successful results with 93.75% of the patients, without detailing which time interval was less successful (5). On the other hand, Zuniga et al. reported that one of their patients had a lesion-treatment interval of 228 days which was considered the cause for, obtaining a "severe" result in the neurosensorial test (please see Table 4 for more details on the neurosensorial test) (6), Thus, it is not possible to determine if time elapsed is a significant determinant for treatment success.    (Table 3) (28). According to these results, in gaps smaller than 70 mm, the success of PNA seems not to be significantly affected by the defect length.

Classification of sensory recovery
None of the studies assess PNA treatment for defects larger than 70 mm ( Table 2). Given the positive results of decellularized allograft, we can compare its advantages and disadvantages versus autologous nerve graft, which is considered the gold standard (5,27,28). The latter is reported to have a success rate ranging from 87.3% to 100% (6), similar to the one reported when using decellularized allograft (85.7% to 100%). However, the latter has the advantage of not generating donor site morbidity, as does the autograft, which is mainly extracted from the greater auricular nerve or sural nerve (4)(5)(6)27). This can generate sensory loss, neuroma formation or neuropathic pain (28). The small number of primary randomized studies describing the use of decellularized allografts, the limited availability of clinical information, and the fact that most of the available studies are from the same authors, limit the conclusions that can be drawn from these results. In addition, the publications included are mostly case report or case series.

Conclusions
Despite the scarce report of primary studies, processed nerve autografting appears to be an effective and promising alternative to achieve positive results of inferior alveolar nerve injuries involving small or wide gaps, without requiring an additional procedure to remove a healthy nerve located elsewhere in the body.