Objective To investigate the effectiveness of facial nerve-sublingual nerve parallel bridge anastomosis for facial nerve injury resulting from closed temporal bone fractures. Methods Between January 2017 and December 2019, 9 patients with facial nerve injury resulting from closed temporal bone fracture caused by head and face trauma were treated. Among them, 5 patients were treated with facial nerve-sublingual nerve parallel bridge anastomosis (operation group), and 4 patients were treated with neurotrophic drugs combined with rehabilitation exercise (conservative group). There was no significant difference in gender, age, side, cause of injury, duration of facial nerve injury before surgery, House-brackmann grading (hereinafter referred to as HB grading) of facial nerve injury, and other general information between 2 groups (P>0.05). HB grading was used to evaluate the improvement of facial nerve function before and after treatment. At the same time, facial nerve neuroelectrophysiological test was performed to evaluate the electrical activity of facial muscles before and after treatment. Tongue function, atrophy, and tongue deviation were evaluated after nerve anastomosis according to the tongue function scale proposed by Martins et al. Results Patients in both groups were followed up 12-30 months, with an average of 25 months. None of the 5 patients in the operation group showed symptoms such as tongue muscle atrophy, tongue extension deviation, hypoglossal nerve dysfunction (mainly including slurred speech, choking with water), postoperative infection, bleeding, lower limb muscle atrophy or lower limb motor dysfunction after sural nerve injury. Postoperative skin sensory disturbance in lateral malleolus area was found, but gradually recovered to normal. During the follow-up, facial nerve and sublingual motor neurons were innervated to paralyzed facial muscle in the operation group. At last follow-up, the HB grading of 5 patients in the operation group improved from preoperative grade Ⅴ in 2 cases, grade Ⅵ in 3 cases to grade Ⅱ in 3 cases, grade Ⅲ in 1 case, and grade Ⅳ in 1 case. And in the conservative group, there were 1 patient with grade Ⅴ and 3 patients with grade Ⅵ before operation, facial asymmetry continued during follow-up, and only 2 patients improved from grade Ⅵ to grade Ⅴ at last follow-up. There was significant difference in prognosis HB grading between the two groups (t=5.693, P=0.001). In the operation group, the amplitude and frequency of F wave were gradually improved, and obvious action potential could be collected when the facial muscle was vigorously contracted. On the contrary, there was no significant difference in neuroelectrophysiological results before and after treatment in the conservative group. ConclusionFacial nerve-sublingual nerve parallel bridge anastomosis can effectively retain the integrity of the facial nerve, while introducing the double innervation of the sublingual nerve opposite nerve, which is suitable for the treatment of severe incomplete facial nerve injury caused by closed fracture.
ObjectiveTo investigate the early effects of acellular xenogeneic nerve combined with adipose-derived stem cells (ADSCs) and platelet rich plasma (PRP) in repairing facial nerve injury in rabbits.MethodsThe bilateral sciatic nerves of 15 3-month-old male Sprague-Dawley rats were harvested and decellularized as xenografts. The allogeneic ADSCs were extracted from the neck and back fat pad of healthy adult New Zealand rabbits with a method of digestion by collagenase type Ⅰ and the autologous PRP was prepared by two step centrifugation. The 3rd generation ADSCs with good growth were labelled with CM-Dil living cell stain, and the labelling and fluorescence attenuation of the cells were observed by fluorescence microscope. Another 32 New Zealand rabbits were randomly divided into 4 groups and established the left facial nerve defect in length of 1 cm (n=8). The nerve defects of groups A, B, C, and D were repaired with CM-Dil-ADSCs composite xenogeneic nerve+autologous PRP, CM-Dil-ADSCs composite xenogeneic nerve, xenogeneic nerve, and autologous nerve, respectively. At 1 and 8 weeks after operation, the angle between the upper lip and the median line of the face (angle θ) was measured. At 4 and 8 weeks after operation, the nerve conduction velocity was recorded by electrophysiological examination. At 8 weeks after operation, the CM-Dil-ADSCs at the distal and proximal ends of regenerative nerve graft segment in groups A and B were observed by fluorescence microscopy; after toluidine blue staining, the number of myelinated nerve fibers in regenerated nerve was calculated; the structure of regenerated nerve fibers was observed by transmission electron microscope.ResultsADSCs labelled by CM-Dil showed that the labelling rate of cells was more than 90% under fluorescence microscope, and the labelled cells proliferated well, and the fluorescence attenuated slightly after passage. All the animals survived after operation, the incision healed well and no infection occurred. At 1 week after operation, all the animals in each group had different degrees of dysfunction. The angle θ of the left side in groups A, B, C, and D were (53.4±2.5), (54.0±2.6), (53.7±2.4), and (53.0±2.1)°, respectively; showing significant differences when compared with the healthy sides (P<0.05). At 8 weeks after operation, the angle θ of the left side in groups A, B, C, and D were (61.9±4.7), (56.8±4.2), (54.6±3.8), and (63.8±5.8)°, respectively; showing significant differences when compared with the healthy sides and with the values at 1 week (P<0.05). Gross observation showed that the integrity and continuity of regenerated nerve in 4 groups were good, and no neuroma and obvious enlargement was found. At 4 and 8 weeks after operation, the electrophysiological examination results showed that the nerve conduction velocity was significantly faster in groups A and D than in groups B and C (P<0.05), and in group B than in group C (P<0.05); no significant difference was found between groups A and D (P>0.05). At 8 weeks after operation, the fluorescence microscopy observation showed a large number of CM-Dil-ADSCs passing through the distal and proximal transplants in group A, and relatively few cells passing in group B. Toluidine blue staining showed that the density of myelinated nerve fibers in groups A and D were significantly higher than those in groups B and C (P<0.05), and in group B than in group C (P<0.05); no significant difference was found between groups A and D (P>0.05). Transmission electron microscope observation showed that the myelinated nerve sheath in group D was large in diameter and thickness in wall. The morphology of myelin sheath in group A was irregular and smaller than that in group D, and there was no significant difference between groups B and C.ConclusionADSCs can survive as a seed cell in vivo, and can be differentiated into Schwann-like cells under PRP induction. It can achieve better results when combined with acellular xenogeneic nerve to repair peripheral nerve injury in rabbits.
ObjectiveTo establish an efficient method of isolating and culturing high activity and high purity of Schwann cells, and to identify the cells at the levels of transcription and translation. MethodsThe sciatic nerves harvested from a 4-week-old Sprague Dawley rat were digested in the collagenase I for 15 minutes after dissecting, and then the explants were planted in culture flask directly. The cells were cultured and passaged in vitro, the growth state and morphological changes of the cells were observed under inverted phase contrast microscope. MTT assay was used to test the proliferation of cells and the cells growth curve was drawn. RT-PCR and immunohistochemistry staining were used to detect S100 and glial fibrillary acidic protein (GFAP) at the levels of transcription and translation, respectively. The purity of cells was caculated under microscope. ResultsAfter the digestion of collagenase I, fibroblast-like cells appeared around explants within 24 hours, with slender cell body and weak refraction. After tissues were transferred to another culture flask, a large number of dipolar or tripolar cells were seen after 48 hours, with slender ecphyma, plump cell body, and b refraction, and the cells formed colonies within 72 hours. The cells were covered with the bottom of culture flask within 48-72 hours after passaging at a ratio of 1∶2, and spiral colonies appeared. Cells showed vigorous growth and full cytoplasm after many passages. MTT assay results showed that the cells at passage 3 entered the logarithmic growth phase on the 3rd day, reached the plateau phase on the 7th day with cell proliferation, and the growth curve was “S” shape. RT-PCR results showed that the cells expressed S100 gene and GFAP gene, and immunohistochemistry staining showed that most of the cells were positively stained, indicating that the majority of cells expressing S100 protein and GFAP protein. The purity of Schwann cells was 98.37% ± 0.30%. ConclusionHigh activity and high purity of Schwann cells can be acquired rapidly by single-enzyme digestion and explant-culture method.
ObjectiveTo construct recombinant adenovirus expressing nerve growth factor (NGF) and myelin associated glycoprotein (MAG) (Ad-NGF-MAG) and to investigate its effect on repair and regeneration of sciatic nerve injury in rats. MethodsNGF and MAG gene sequences were cloned into shuttle plasmid pCA13 of adenovirus type 5. After packed in HEK293 cells, the recombinant Ad-NGF-MAG underwent sequence and identification. Thirty-two male Sprague Dawley rats were randomly divided into 4 groups (n=8): control group (normal control), adenovirus vector group (Ad group), Ad-NGF group, and Ad-NGF-MAG group. The sciatic nerve injury model was established by transection of the right sciatic nerve; then, the empty adenovirus vector, Ad-NGF, and Ad-NGF-MAG were injected into the gastrocnemius muscle of the affected limb at a dose of 1×108 PFU every other day for 3 times in Ad group, AdNGF group, and Ad-NGF-MAG group, respectively. The right sciatic nerve was exposed only, and then the incision was closed in the control group. The sciatic nerve function index (SFI) was measured, and neuro-electrophysiology was observed; mRNA and protein expressions of NGF and MAG were detected by RT-PCR and Western blot; and histological examination was performed at 31 days after operation. ResultsRecombinant adenovirus vectors of Ad-NGF and Ad-NGF-MAG were constructed successfully. All rats survived and incision healed by first intension. The SFI, nerve conduction velocity, evoked potential amplitude, and latent period of Ad-NGF-MAG group were significantly better than those of Ad group and Ad-NGF group (P < 0.05). MAG mRNA and protein expressions of Ad-NGF-MAG group were the highest in all the groups (P < 0.05). The expressions of NGF mRNA and protein increased in Ad-NGF group and AdNGF-MAG group when compared with control group and Ad group (P < 0.05). Histological examination showed that the nerve had good continuity in control group; nerve fibers disarranged in Ad group; neurons connections formed in some nerve fibers of Ad-NGF group, but nerve fibers arrange disorderly; and the growth of the nerve were ordered and wellstructured in Ad-NGF-MAG group. ConclusionAd-NGF-MAG can effectively promote the growth of the nerve and inhibit the form of abnormal branches, facilitating the repair of sciatic nerve injury in rats.
ObjectiveTo investigate the effect of folic acid coated-crosslinked urethane-doped polyester elastomer (fCUPE) nerve conduit in repairing long distance peripheral nerve injury. MethodsThirty-six 3-month-old male Sprague Dawley rats weighing 180-220 g were randomly assigned to 3 groups, each consisting of 12 rats: CUPE nerve conduit transplantation group (group A), fCUPE nerve conduit transplantation group (group B), and autologous nerve transplantation group (group C), the contralateral healthy limb of group C served as the control group (group D). A 20-mm-long sciatic nerve defect model was established in rats, and corresponding materials were used to repair the nerve defect according to the group. The sciatic function index (SFI) of groups A-C was calculated using the Bain formula at 1, 2, and 3 months after operation. The nerve conduction velocity (NCV) of the affected side in groups A-D was assessed using neuroelectrophysiological techniques. At 3 months after operation, the regenerated nerve tissue was collected from groups A-C for S-100 immunohistochemical staining and Schwann cell count in groups A and B to compare the level of nerve repair and regeneration in each group. ResultsAt 3 months after operation, the nerve conduits in all groups partially degraded. There was no significant adhesion between the nerve and the conduit and the surrounding tissues, the conduit was well connected with the distal and proximal nerves, and the nerve-like tissues in the conduit could be observed when the nerve conduit stents were cut off. SFI in group A was significantly higher than that in group C at each time point after operation and was significantly higher than that in group B at 2 and 3 months after operation (P<0.05). There was no significant difference in SFI between groups B and C at each time point after operation (P>0.05). NCV in group A was significantly slower than that in the other 3 groups at each time point after operation (P<0.05). The NCV of groups B and C were slower than that of group D, but the difference was significant only at 1 month after operation (P<0.05). There was no significant difference between groups B and C at each time point after operation (P>0.05). Immunohistochemical staining showed that the nerve tissue of group A had an abnormal cavo-like structure, light tissue staining, and many non-Schwann cells. In group B, a large quantity of normal neural structures was observed, the staining was deeper than that in group A, and the distribution of dedifferentiated Schwann cells was obvious. In group C, the nerve bundles were arranged neatly, and the tissue staining was the deepest. The number of Schwann cells in group B was (727.50±57.60) cells/mm2, which was significantly more than that in group A [(298.33±153.12) cells/mm2] (t=6.139, P<0.001). ConclusionThe fCUPE nerve conduit is effective in repairing long-distance sciatic nerve defects and is comparable to autologous nerve grafts. It has the potential to be used as a substitute material for peripheral nerve defect transplantation.
Objective To analyze the therapy and effectiveness of ulnar styloid fracture complicated with wrist dorsal branch of ulnar nerve injury. Methods Between October 2005 and October 2012, 16 cases of ulnar styloid fracture complicated with wrist dorsal branch of ulnar nerve injury were treated. There were 14 males and 2 females with an average age of 42 years (range, 22-58 years). Fracture was caused by traffic accident in 8 cases, by mechanical crush in 5 cases, and by falling in 3 cases. According to the anatomical features of the ulnar styloid and imaging findings, ulnar styloid fractures were classified as type I (ulnar styloid tip fracture) in 1 case and type II (ulnar styloid base fracture) in 15 cases. The skin sensation of ulnar wrist was S0 in 5 cases, S1 in 1 case, S2 in 7 cases, and S3 in 3 cases according to the criteria of the British Medical Research Council in 1954 for the sensory functions of the ulnar wrist. The time from injury to operation was 6-72 hours (mean, 18 hours). Fracture was treated by operative fixation, and nerve was repaired by epineurium neurolysis in 13 cases of nerve contusion and by sural nerve graft in 3 cases of complete nerve rupture. Results All incisions healed by first intention. Sixteen patients were followed up for an average time of 14 months (range, 6-24 months). The X-ray films showed that all of them achieved bone union at 4-10 weeks after operation (mean, 6 weeks). No patient had complications such as ulnar wrist chronic pain and an inability to rotate. According to Green-O’Brien wrist scoring system, the results were excellent in 13 cases and good in 3 cases; according to the criteria of the British Medical Research Council in 1954 for the sensory functions of the ulnar wrist, the results were excellent in all cases, including 11 cases of S4 and 5 cases of S3+. Two-point discrimination of the ulnar wrist was 5-9 mm (mean, 6.6 mm). Conclusion For patients with ulnar styloid fracture complicated with wrist dorsal branch of ulnar nerve injury, internal fixation and nerve repair should be performed. It can prevent ulnar wrist pain and promote sensory recovery.
ObjectiveTo investigate the effect of electrospun chitosan/polylactic acid (ch/PLA) nerve conduit for repairing peripheral nerve defect in rats. MethodsNerve conducts loaded with ch/PLA was made by the way of electrospun. The mechanical property, hydrophility, biocompatibility were tested, and the scanning electron microscope was used to observe the ultrastructure. The same experiments were also performed on pure PLA nerve conducts as a comparison. Then, 54 Sprague Dawley rats were divided into 3 groups randomly, 18 rats in each group. Firstly, the 10 mm defects in the right sciatic nerves were made in the rats and were respectively repaired with ch/PLA (group A), autografts (group B), and no implant (group C). At 4, 8, and 12 weeks after operation, general observations, sciatic functional index (SFI), electrophysiological evaluation, wet weight of gastrocnemius and soleus muscles, histological examination, immunohistological analysis, and transmission electron microscopy were performed to evaluate the effects. ResultsCompared with pure PLA nerve conducts, the addition of chitosan could improve the mechanical property, hydrophility, biocompatibility, and ultrastructure of the nerve conducts. At 4 weeks postoperatively, the regenerated nerve bridged the nerve defect in group A. The SFI improved gradually in both group A and group B, showing no significant difference (P>0.05). Compound muscle action potentials and nerve conduction velocity could be detected in both group A and group B at 8 and 12 weeks after operation, and significant improvements were shown in both groups (P<0.05). The wet weight and myocyte cross section of gastrocnemius and soleus muscles showed no significant difference between group A and group B (P>0.05), but there was significant difference when compared with group C (P<0.05) at 12 weeks postoperatively. Immunohistological analysis revealed that S-100 positive Schwann cells migrated in both group A and group B, and axon also regenerated by immunohistological staining for growth associated protein 43 and neurofilaments 160. Transmission electron microscopy showed no significant difference in the diameter of nerve fiber between group A and group B (P>0.05), but the thickness of myelin sheath in group A was significantly larger than that in group B (P<0.05). ConclusionThe electrospun ch/PLA nerve conduits can effectively promote the peripheral nerve regeneration, and may promise an alternative to nerve autograft for repairing peripheral nerve defect.
Objective To analyze the therapeutic effect of olfactory ensheathing cells (OECs) transplantation for central nervous system diseases. Methods Between November 2001 and January 2008, 1 255 participants with central nervous system diseases were enrolled in this cl inical study for fetal OECs transplantation. There were 928 males and 327 femalesaged 1.2-87 (mean 40) years. The course of disease was (4.52 ± 4.67) years. Among them, 656 participants suffered from chronic spinal cord injury (SCI), 457 amyotrophic lateral sclerosis (ALS), 68 cerebral palsy (CP), 20 multiple sclerosis (MS), 11 the sequelae of stoke, 10 ataxia, and 33 residual diseases. The participants came from 71 countries or regions. Accidentally abortional fetal olfactory bulbs were donated voluntarily and were cultured for 2 weeks, then were transplanted. Results One thousand one hundred and twenty-eight cases were followed up for 2-8 weeks (mean 4 weeks) to obtain integrated data. Among them, the neurological functional amel ioration was noticed in 994 participants with the overall short-term improvement rate of 88.12%. Seventy-six patients experienced the various perioperative compl ications with the incidence rate of 6.74%. One hundred and twenty patients with SCI received over 1 year follow-up. And according to ASIA assessment, motor scores increased from (39.82 ± 20.25) to (44.55 ± 18.99) points, l ight touch scores from (51.56 ± 25.89) to (59.81 ± 27.72) points, pain scores from (50.36 ± 27.44) to (57.09 ± 28.51) points for foreign patients (P lt; 0.05); motor scores increased from (40.52 ± 20.80) to (46.45 ± 20.35) points, l ight touch scores from (55.64 ± 26.32) to (68.64 ± 25.89) points, pain scores from (57.05 ± 26.00) to (66.13 ± 24.29) points for good rehabil itation Chinese patients (overall P lt; 0.05); motor scores from (37.03 ± 18.52) to (38.03 ± 18.50 points (P lt;0.05), l ight touch scores from (45.88 ± 22.56) to (46.63 ± 23.09) points (P gt; 0.05), pain scores from (45.25 ± 23.68) to (45.28 ± 23.63) points (P gt; 0.05) for poor rehabil itation Chinese patients. Compared foreign patients and good rehabil itation Chinese patients with poor rehabil itation Chinese patients, difference in score change was remarkable (P lt; 0.05). One hundred and six cases of ALS, 32 CP, 8 MS, 7 ataxia, and 2 stroke sequelae were followed up for 3-48, 3-36, 2-20, 7-17, 6 and 24 months, One hundred and six cases of respectively. Majority of them (113/155, 72.9%）were benefited from OECs transplantation. Conclusion OECs transplantation into brain and spinal cord is feasible and safe . The therapeutic strategy is valuable treatment for such central nervous system diseases such as chronic SCI, ALS, CP and stroke sequelae and can improve the patients’ neurological functions and/or decrease the progressive deterioration.
Objective To review the clinical applications of functional perforator flaps in restoring human body functions. Methods An extensive literature review was conducted on both domestic and international publications to summarize the clinical use of functional perforator flaps for functional restoration. Results Perforator flaps are among the most commonly used flaps in reconstructive surgery. Beyond providing soft tissue repair, they are increasingly employed to reconstruct diverse bodily functions, leading us to propose the concept of the “functional perforator flap”. Although various forms of functional perforator flaps are currently utilized, reports are predominantly scattered case studies, lacking systematic organization. Commonly used functional perforator flaps can be categorized into five types: chimeric perforator flaps, perforator flaps for nerve function restoration, perforator flaps for lymphatic drainage enhancement, flow-through perforator flaps, and perforator flaps for restoring bone and joint motion. These flaps significantly broaden the application scope of perforator flaps, elevating the goal of reconstruction from mere wound repair to achieving repair concurrent with functional reconstruction. ConclusionThe application of various functional perforator flap designs significantly improves wound reconstruction outcomes and represents an effective approach for managing complex defects. Future developments will undoubtedly see more forms of functional perforator flaps reported to meet increasingly sophisticated reconstructive demands.
Peripheral nerve injury (PNI) is a common neurological dysfunction. In clinical practice, autologous nerve transplantation is used to solve problems related to PNI, such as limited donor resources, neuroma formation and high donor incidence rate. Therefore, searching for new nerve regeneration materials has become a hot research topic. The decellularized extracellular matrix (dECM) hydrogel provides a scaffold for nerve regeneration by removing the cellular components in biological tissues, preserving the extracellular matrix, and is a potential therapeutic material for nerve regeneration. This article reviews the research progress of dECM hydrogel for PNI and looks forward to the clinical prospects of this research direction.