人胶原和自体脂肪声带注射的实验研究

目的 探讨国产人胶原狗声带注射的组织学转归及其在矫治声门闭合不全中的应用价值和前景。     

人胶原和自体脂肪声带注射的实验研究

目的探讨国产人胶原狗声带注射的组织学转归及其在矫治声门闭合不全中的应用价值和前景。方法实验动物分为４组，Ａ组２只分别注射人胶原和自体脂肪矫正左侧声带麻痹引起的声门闭合不全，并对治疗效果进行主、客观观察，术后５个月取材，透射电镜观察。Ｂ、Ｃ、Ｄ组９只狗（每组３只）于右侧声带注入人胶原、左侧声带注入自体脂肪各０．３ｍｌ，分别于术后１、３、５个月取材并观察其组织学变化和体积变化。结果自体脂肪能在声带组织中存活，但吸收率很高，１、３、５个月体积平均减少速度为３８．５１％，疗效不能持久。人胶原易于注射，组织相容性好，并能被宿主成纤维细胞和血管移生，同时伴有宿主胶原的再生，显示出最终被宿主组织完全取代的趋势，１～５个月体积平均减少速度为０．６５％。矫治效果长期稳定。结论自体脂肪应用有局限性，只能作为短期的声带植入材料；人胶原可用于各种原因所致声门闭合不全的长期矫治材料，具有良好的应用前景。     

Vocal Cord Injection With Autogenous Fat

Since 1987, the senior author has injected autogenous fat into paralyzed or atrophic vocal cords as an alternative to alloplastic substances for vocal cord augmentation and medialization. To determine the fate of the injected autogenous fat, the injected vocal cords of 10 patients were evaluated by laryngeal magnetic resonance imaging (MRI) in the sagittal, coronal, and axial planes. Imaging studies were performed as early as 17 hours after surgery to as long as 31 months after fat injection. In 9 patients, identification of a fat signal within the previously injected vocal cords was observed (including the 31-month postoperative follow-up). In 1 patient, no fat signal was identified 13 months after surgery, but the vocal cord was noted to have a bulging, enlarged contour. The results of this imaging study provide further evidence that autogenous fat, which has not been damaged during harvesting or microinjection, can survive transplantation into the vocal cord. The bulk of the vocal cord is maintained by microlipocytes and fibrous connective tissue, both of which replace the damaged fat cells that are gradually being reabsorbed.     

自体脂肪声带内注射术治疗单侧声带麻痹的远期疗效观察

OBJECTIVE: To study the long-term results of autogenous fat injection for unilateral vocal cord paralysis. METHODS: Twenty cases with unilateral vocal cord paralysis were treated by autogenous fat injection into the thyroarytenoid muscle to achieve medialization. The patients were divided into 3 groups by hoarse degree before operation, all of them were followed more than 12 months with serial video laryngoscope and voice evaluation. The ratio between paralyzed vocal cord upper surface and that of the normal vocal cord were adopted as the measurement for the vocal cord volume changes before and after operation. RESULTS: 1. The volume of paralyzed vocal cord was increased. The degree of hoarse and normalized noise energy (NNE) were evaluated by objective methods after operation. 2. The hoarse symptom was less severe after operation than that before operation. The cure cases 3 to 6 months and over 12 months after operation were nearly the same. 3. NNE of over 12 months and 3 to 6 months after operation were not significantly different, but the postoperative NNE were different with that before operation. CONCLUSIONS: Autologous fat injection was an effective method for treating unilateral vocal cord paralysis, and the long term effects were reliable.     

自体脂肪声带内注射术治疗单侧声带麻痹的远期疗效观察

目的　探讨自体脂肪声带内注射术治疗单侧声带麻痹的远期疗效。方法　 2 0例单侧声带麻痹患者将取自体腹部的脂肪颗粒注射到麻痹侧声带 ,使声带隆起 ,声门闭合 ,恢复嗓音功能 ;所有病例均随访 12个月以上 ,并按术前及术后即刻、近期 (3～ 6个月 )、远期 (>12个月 )通过电子喉镜以计算机软件测得患侧与健侧声带上表面面积比为参数代表患侧声带大小变化指标 ,并用嗓音疾病评估仪进行嗓音声学测试分析。按术前嗓音声学测试结果分为轻度、中度、重度 3组 ,采用自身对照统计分析结果。结果　注射后麻痹侧声带相应体积较术前明显增大 ,但随时间推移 ,麻痹侧声带内的脂肪部分吸收 ,麻痹侧声带体积逐渐减小 ,而术后近期各组及远期中重度声嘶组测得的麻痹侧声带相应体积与术前差异有显著性 ,轻度声嘶组远期测得的麻痹侧声带相应体积与术前差异无显著性。声学测试客观分析 ,注射后较注射前嘶哑程度明显好转 ,随时间延长嘶哑程度有所下降 ,但术后近期及远期嘶哑程度接近。直接反映声门闭合程度的噪声能量在注射脂肪后较注射前明显下降 ,达到正常或接近正常 ;随时间延长噪声能量稍升高 ,但与术前差异有显著性 ,且术后近期及远期的差异无显著性。结论　自体脂肪声带内注射治疗单侧声带麻痹是可选择的有效方法 ,其远期     

Vocal cord augmentation with autogenous fat.

Since 1987, the authors have used a new technique for the augmentation and medialization of the paralyzed vocal cord. Eleven patients with paralyzed vocal cords and one patient with a surgical defect of the vocal cord have been treated with intracordal injections of autogenous fat harvested by suction from the abdominal wall. Treatment efficacy was evaluated by both subjective and objective (videostroboscopic) analyses. Patient follow-up has ranged from 5 to 42 months. All patients showed significant improvement in voice characteristics. Marked reduction in air flow and increased vocal intensity were observed. Correction of glottic insufficiency has been lasting, indicating that the implanted fat cells are viable. At this time, there has been no evidence of loss of vocal cord substance. The natural qualities of autogenous fat appear to make it an excellent alternative to Teflon or collagen. The fat is readily available, and easily harvested and injected. A larger patient population and longer follow-up periods will help determine if these initial findings are consistent.     

Histologic study of an autologous fat graft in the larynx of dogs with unilateral vocal fold paralysis

OBJECTIVE: To determine the histologic finds of autologous fat graft (AFG) in 24 paralyzed canine left vocal folds 12 weeks after introduction. STUDY DESIGN: Animal model. Experimental prospective study. METHODS: Twenty-four mongrel dogs of both genders weighing 15 to 20 kg were divided into three study groups (SGs) of eight dogs each, whose larynges were grafted by vocal fold lipoinjection (VFL-SGA), vocal fold lipoinjection plus insulin (VFLI-SGB), and by fat graft medialization laryngoplasty (FGML-SGC), respectively, and observed for 12 weeks, followed by immediate sacrifice. All 24 dogs were submitted to left vagal and recurrent laryngeal nerve resection. All animals were confirmed to have a left vocal fold paralysis four weeks later, when 2 mL of autologous fat graft were placed inside the left paraglottic space. The larynges were removed at pre-established times and, after they were studied for remaining fat graft volume by magnetic resonance imaging, they were studied histologically to evaluate the influence of the different techniques on the histologic behavior of the graft. RESULTS: Each of the three study groups had its specific finds for fat graft detection and fat absorption, and its characteristics identified and compared statistically. The groups were not statistically different. CONCLUSION: The histologic findings for the AFG applied to a vocal fold were similar in groups VFL-SGA, VFLI-SGB and FGML-SGC 12 weeks after grafting, with no statistically significant differences between groups, and revealed an almost total loss of the grafted fat.     

肝细胞生长因子对喉返神经损伤后再生的作用

目的探讨肝细胞生长因子(hepatocyte growth factor,HGF)局部应用对受损喉返神经再生的作用。方法 20只健康杂种犬分成HGF组及对照组,每组10只。每只犬切断左侧喉返神经后,用聚乳酸聚羟基乙酸共聚物[Poly(DL lactide co glycolide),PLGA]管桥接左侧喉返神经断端,HGF组于PLGA管内及局部应用HGF,对照组同法应用等量生理盐水。于术后2、3、4个月分别行电子喉镜、喉返神经电生理检查,最后一次检查完后处死动物,取神经干作神经组织学和超微结构观察。结果术后2个月,两组动物术侧声带未恢复运动;术后3个月,HGF组有4只犬的左侧声带观察到内收、外展运动,而对照组均未恢复运动;术后4个月,HGF组存活的8只犬中7只术侧声带恢复运动,对照组7只存活犬中5只术侧声带恢复运动,但运动幅度均明显小于HGF组。术后各时间点HGF组与对照组喉返神经电生理检查结果相比较,HGF组自发肌电图、诱发电位幅度明显增大,潜伏期明显缩短,差异均有统计学意义(P<0.05)。HGF组有髓再生神经纤维数、神经纤维的直径及髓鞘的厚度均明显大于对照组,差异均有统计学意义(P<0.05)。电镜观察示HGF组的再生神经纤维超微结构形态接近正常,而对照组形态与正常差异较大,可见较多的无髓神经纤维。结论喉返神经损伤后应用外源性HGF对其再生有明显的促进作用。     

自体脂肪、筋膜移植用于声带填充术的实验研究

目的 应用自体脂肪、筋膜行声带填充治疗单侧声带麻痹,探讨这两种填充材料的疗效及应用前景。方法 实验动物为狗,分为2组,A组2只分别以自体脂肪、筋膜行声带填充矫治左侧事麻痹而致的声门闭合不全,于术后8个月取材行透射电镜及光镜观察。B组9只分别于左侧声带植入自体脂肪,右侧声带植入自体筋膜各0.4ml,分别手术后1、3、5个月以材行光镜观察。结果 自体脂肪能存活于声带组织中,但吸收较明显,功能观察显示,疗效维持不超过5个月;自体筋膜可成功存活于声带,疗效可维持8个月以上。结论 自体脂肪吸收明显,疗效不能持久,仅能人为短期的声带填充材料,自体膜性质较为稳定,疗效持久,具有良好的应用前景。     

经环甲间隙自体阔筋膜加脂肪声带注射治疗单侧声带麻痹

目的 观察经环甲间隙自体阔筋膜加脂肪声带注射治疗周围性单侧声带麻痹的效果.方法在全麻支撑喉镜下,对12例周围性单侧声带麻痹患者经环甲间隙声带注射自体阔筋膜加脂肪,评估注射前后声带位置、声嘶改善状况及声学参数变化.结果 12例患者术后声嘶明显改善,最大声时延长.动态喉镜检查患侧声带移向正中位,声门闭合良好,3个月后注射侧声带体积保持稳定,未发现任何并发症.结论经环甲间隙自体阔筋膜加脂肪声带注射治疗单侧声带麻痹简单安全、疗效确切.     

Endolaryngeal lipoaugmentation of the vocal cords

Unilateral vocal cord paralysis or the loss of vocal cord tissue results in incomplete glottic closure (internal-paresis) with a consequent hoarseness and poor voice quality. Improving glottic incompetence, instead of the previously used paraffin, teflon is a widely accepted and most commonly used substance at present for vocal cord medialization. Using intracordal teflon injection of for decades had proven that it has limitations and potential complications to the human body. Beside the well-known external "phono-surgical" methods, a wide interest has been shown in endolaryngeal phono-surgery and in finding the ideal and most available biocompatible substances for the procedure. Papers have been issued about the promising results of autologus fat injection for medialization of the paralyzed vocal fold in the early '90s. In this paper the authors report on their slightly modified endolaryngeal intracordal autologus fat injection procedure, and its promising results. The first three patients (a left side glottic paralysis, a bilateral internal paresis and a left side internal paresis) experienced an improvement in their voice right after the medialization procedure, what remained the same during the 11 months follow-up period. Using supraglottic jet ventilation during general anaesthesia provides very good access to the operating field. Monitoring of neuro-muscular block makes possible an intraoperative examination of glottic closure by the protective reflex of the larynx. Incouraged by this initial results the authors suggest the autologus fat as an easy available, ideal substance for increasing (augmenting) the loss of vocal cord tissue.     

电子喉镜下脂肪注射术治疗单侧声带麻痹

目的 探讨电子喉镜监控下声带黏膜下自体脂肪注射术治疗单侧声带麻痹的疗效.方法 32例单侧声带麻痹患者,取自体脂肪颗粒在电子喉镜监控下自环甲膜穿刺,患侧声带内多角度分层次多点注射,术前、术后即刻、术后6个月、术后12个月分别测量患侧声带的表面积和声门裂隙大小,嗓音质量评估仪对嗓音进行评估.结果注射术后麻痹侧声带表面积增加,声门裂隙缩小,随时间推移,麻痹侧声带内的脂肪部分吸收,声带表面积减小,声门裂隙稍增大,术后各阶段和术前比较差异有显著性统计学意义;但术后6个月和术后12个月比较差异无显著性意义.嗓音声学测试发现,术后声门噪声能量下降,声时延长,与术前比较差异有显著性统计学意义;术后6个月和术后12个月比较,差异无显著性意义.结论电子喉镜下声带黏膜下自体脂肪注射术方法简单、方便,疗效确切,是治疗单侧声带麻痹的有效方法.     

脂肪干细胞复合脱细胞真皮基质用于声带注射的实验研究

目的 探索脂肪干细胞(adipose-derived stem cell,ASC)与脱细胞真皮基质微粒(micronized acellular dermal matrix,MADM)复合培养后用于声带内注射的可行性.方法 体外分离培养兔ASC,取第3代细胞,用细胞膜红色荧光探针(DiI)标记,并与已制备好的MADM复合培养,形成细胞和材料复合体,应用荧光显微镜、扫描电镜了解ASC黏附材料情况,四唑类化合物3-(4,5-dimethylthizazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfonyl)-2H-tetrazolium,inner salt(MTS)比色法检测细胞增殖情况.ASC-MADM复合体培养3 d后与适量胶原混合后注入异体兔单侧声带内,术后2、4、8周内镜观察,并分别处死动物,检测ASC在声带中存活、分布情况.结果 MADM所含胶原纤维松散、多孔隙,具有良好三维结构;ASC可黏附其生长并能不断增殖,与没有复合MADM的对照组比较细胞增殖水平差异有统计学意义(P值均＜0.05或＜0.01).ASC-MADM复合体注入兔声带肌后均存活良好,连续内镜观察、冰冻切片、HE染色显示声带局部无不良反应,ASC-MADM复合体在兔声带内基本无炎症细胞浸润,ASC可在声带内存活8周.结论 MADM是较理想的细胞支架材料,ASC能黏附MADM生长并增殖.ASC-MADM复合体异体动物声带内注射可存活且无不良反应.

Abstract：

Objective To explore the feasibility of adipose-derived stem cells (ASC) combined with micronized acellular dermal matrix (MADM) for vocal cord injection. Methods The adipose-deprived stem cells were harvested from rabbit adipose tissue in vitro. The 3rd generation of ASC was labeled with DiI ( 1, 1-dioctadecyl-3,3,3,3-tetramethylindocarbocyanine perchlorate) and cultured with MADM to form a complex. The adhesion of ASC to MADM was observed by fluorescence microscope and electron microscope.The proliferation of ASC on MADM was evaluated by 3-(4, 5-dimethylthizazol-2-yl)-5-( 3-carboxy methoxyphenyl)-2-(4-sulfonyl )-2H- tetrazolium, inner salt (MTS). Three days after the culture, the complex was mixed with appropriate amount of collagen, and then injected into the unilateral vocal cord of the rabbit. The animals were sacrificed 2, 4, 8 weeks after injection, the survival time and distribution of ASC in vocal fold were tested, and the responses of vocal cord to ASC-MADM and the degradation of MADM were observed. Results The ASC adhered to MADM and grew well (P ＜0. 05 or ＜0. 01 ), showing good compatibility with MADM in vocal cord tissue. The complex of ASC-MADM could be injected into the rabbit vocal cords, while no adverse reactions was observed in the vocal cord by endoscope, frozen section and HE staining. ASC could survive for 8 weeks in vocal cords, and no inflammatory cell infiltration was observed.Conclusions MADM is an ideal scaffold material and shows perfect compatibility with ASC which can adhere and proliferate well on it. The complex of ASC-MADM can be injected into the vocal cord and can survive. There is no adverse reaction in vocal cords.     

声带血管扩张症21例临床分析

目的探讨声带血管扩张症的诊断和治疗。方法总结分析21例患者的临床资料。所有患者均有不同程度的声音嘶哑，其中女性15例，男性6例；年龄23～68岁，中位年龄44岁；血管扩张位于右侧声带12例，左侧声带9例。单纯声带血管扩张10例；声带血管扩张伴发声带息肉8例，伴发声带小结2例，伴发一侧声带麻痹1例。所有患者随访6个月以上。结果在电视喉镜下显示扩张的小血管从声带前端开始向后延伸至声带中部的6例，表现为扩张的血管在声带中部进一步扩张成类似小的血管瘤者3例，或表现点状、片状或短线状血管扩张12例。8例伴声带息肉的患者摘除声带息肉手术后复查见2例声带表面血管扩张消失，6例血管扩张无变化，声嘶明显改善；其余13例患者进行声带休息治疗，12例复查病变无变化，但声休后主观感觉声音嘶哑症状好转，1例患者未注意声带休息，3个月时因声音嘶哑加重，就诊发现对侧声带黏膜下出血，6个月复查时血管扩张未见变化，但对侧声带黏膜下出血吸收消失。结论声带血管扩张症好发于女性患者，做喉镜检查可明确诊断，治疗以声休为主，反复出血或持续声嘶者可考虑手术。     

Newer technique of laryngeal reinnervation: superior laryngeal nerve (motor branch) as a driver of the posterior cricoarytenoid muscle

This report analyzes the experience gained using two different techniques to reinnervate the paralyzed vocal cord. In the neurotization group, the superior laryngeal nerve (SLN) motor branch-cricothyroid muscle pedicle was used to reinnervate the posterior cricoarytenoid muscle. In the direct nerve anastomosis group, the SLN was anastomosed to the abductor branch of the recurrent laryngeal nerve (RLN), and the ansa hypoglossi (AH) to the adductor branch of the RLN. A third group of animals (control) had the right RLN sectioned without any anastomosis. About 5 to 6 months postoperatively the animals were killed painlessly and evaluated. The neurotization group revealed vocal fold mobilization on the right side to have an average of about half of the mobility of the left, normal side. After the RLN and SLN on the left were severed as well as the AH bilaterally, the vocal cord mobility was reduced to about one fourth. The direct nerve anastomosis group showed about fourfold less vocal cord mobility than the neurotization group. After the SLN, RLN, and AH were severed bilaterally, the control group showed no vocal cord mobility. The neurotization technique has been selected for further experimentation in human adults.     

Adipose tissue formation in response to basic fibroblast growth factor

CONCLUSION: Formation of new fat cells may occur in injected adipose tissue in response to basic fibroblast growth factor (bFGF). OBJECTIVE: In an effort to prevent loss of fat volume and generation of additional adipose tissue after intracordal injection of autologous fat, the effects of injecting fat together with basic fibroblast growth factor bFGF were evaluated. MATERIALS AND METHODS: Models of bilateral recurrent laryngeal nerve paralysis were prepared using 12 beagle dogs. Autologous fat was injected into one vocal cord, and a mixture of autologous fat with bFGF, gelatin microspheres, and collagen sponge was injected into the other. Histologic reactions were examined over time. RESULTS: In vocal cords where autologous fat was injected together with bFGF, fusiform immature adipocytes were found in the injected fat at 8 weeks after injection. The volume of the injected fat was maintained almost completely even at 24 weeks after injection. Vocal cords where only autologous fat was injected showed a marked decrease in volume of injected fat over time.     

Phonosurgical studies: fat-graft reconstruction of injured canine vocal cords

Damage to the vocal cords can result in scarring and impaired vibration and can manifest clinically as hoarseness and loss of vocal power. If the vibratory characteristics could be restored in these scarred vocal cords, the vocal intensity and efficiency of phonation also should improve. In an effort to enhance the vibration of damaged vocal cords, we implanted a submucosal fat autograft within the injured vocal cord cover layer of dogs 6 weeks after unilateral mucosal excision had been performed. Three months postoperatively these animals were compared to normal dogs and those with mucosal excision but no fat-grafting. Acoustic and biomechanical measures of phonation were collected from an excised larynx preparation. We found that the fat-augmented vocal cords had lower threshold pressures for phonation, greater vocal intensity, and more efficient acoustic output than injured vocal cords without the fat-grafting. These results provide a foundation for further research on reconstructive surgery of damaged vocal cords.     

自体脂肪及筋膜植入犬声门旁间隙及固有层的实验研究

目的探讨犬喉自体脂肪及筋膜植入后植入组织生长特性、填充机制及影响填充效果的因素。方法 16只犬32侧声带,除2侧声带作为正常对照组不作任何手术处理外,其余30侧声带通过切断喉返神经建立声带麻痹动物模型后,分为以下4组:脂肪注射组(10侧)、脂肪填充组(5侧)、筋膜注射组(8侧)、筋膜填充组(7侧),除筋膜填充组填充于犬声带的固有层外,其余三组脂肪和筋膜注射分别注射于犬喉的声门旁间隙,于术后第1周、1、3、6、12个月对犬喉植入的脂肪、筋膜分别进行大体及组织病理学观察及比较。结果脂肪植入组在脂肪植入部位可见炎性细胞浸润,于术后第3个月后消失,术后3月时开始有新生血管形成,12月时脂肪植入部位仍可见团状的成活脂肪细胞。筋膜植入组无明显炎性细胞反应,在筋膜植入部位,筋膜组织以密度较高的纤维组织形式存在,均于第1个月时观察到纤维母细胞,筋膜填充组术后第3个月后筋膜组织呈均匀排列,第6个月后椭圆形纤维母细胞密度增大,填充部位新生血管形成,术后12个月其密度与周围组织趋于一致。脂肪植入组纤维母细胞出现比例(4/15)小于筋膜植入组(9/15),而新生血管比例(7/15)大于筋膜植入组(3/15)。结论自体脂肪及筋膜作为声带填充材料具有良好的组织相容性,术后12月时仍在植入部位存活。填充物周围纤维包膜的形成及植入后纤维组织、纤维母细胞、自体血管等的生长,对保持填充效果长期稳定有影响。     

Autologous fat graft absorption in unilateral paralyzed canine vocal folds

OBJECTIVE: The objective of this experimental prospective study on an animal model was to determine the rate of autologous fat graft absorption in 24 paralyzed canine left vocal folds 12 weeks after introduction. METHODS: Forty mongrel dogs of both sexes weighing 15 to 20 kg were divided into three study groups (SGs) of eight dogs each whose larynges were grafted by vocal fold lipoinjection (VFL-SGA), vocal fold lipoinjection plus insulin (VFLi-SGB), and by fat graft medialization laryngoplasty (FGML-SGC) and observed for 12 weeks. Two control groups (CGs) of eight dogs each were submitted to fat graft medialization laryngoplasty (FGML-CG1) and by vocal fold lipoinjection (VFL-CG2), respectively, followed by immediate killing. All 40 dogs were submitted to left vagal and recurrent laryngeal nerve resection. All animals were confirmed to have a left vocal fold paralysis 4 weeks later when 2 mL of autologous fat graft was placed inside the left paraglottic space. The larynges were removed at preestablished times and studied for remaining fat graft volume by magnetic resonance imaging. RESULTS: The mean volume obtained by imaging were 1.9388 mL for CG1, 1.0476 mL for CG2, 0.1924 mL for VFL, 0.336 mL for VFLi, and 0.565 mL for FGML. CG1 and CG2 did not differ significantly from one another but differed from the SGs, with statistical analysis suggesting a lower absorption of the fat graft in FGML. CONCLUSION: The absorption rate was 82% in SGA, 68% in the SGB, and 71% in SGC.     

嗓音训练在单侧声带息肉术后嗓音恢复中的应用

目的 探讨单侧大、小声带息肉术后发声功能恢复规律和嗓音训练最佳时机.方法 选取68例武汉市第三医院耳鼻咽喉科行单侧声带息肉手术患者,根据与声带平行的息肉最大直径占同侧声带膜部1/3以下称为小声带息肉,1/3及以上为大声带息肉,分成大声带息肉组30例,小声带息肉组38例,每组随机分成嗓音训练组和禁声组;分别于术前1d,术...