阻塞性睡眠呼吸暂停低通气综合征动物模型的建立及腭咽扩张肌病理基础和临床外科治疗研究
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摘要
研究背景和目的建立某一疾病的合理可靠的动物模型是深入研究这种疾病的前提和必要手段。目前国内外对阻塞性睡眠呼吸暂停低通气综合征(obstructive sleep apnea—hypopnea syndrome,OSAHS)动物模型方面的研究仍在初步阶段,因为OSAHS发病机制复杂,不易从病因、病理等方面均得以模拟。颏舌肌是上气道最强有力的扩张肌,其肌力松弛会引起舌根后气道狭窄,是OSAHS发病原因之一。肉毒杆菌毒素这一药物注射于肌肉能引起较持久的肌力松弛作用,且在安全剂量范围内几乎无任何不良反应。我们因此设想通过在动物的颏舌肌内注射肉毒杆菌毒素引起颏舌肌肌力慢性松弛,造成舌根后气道狭窄的OSAHS动物模型,既为研究者对OSAHS的病因、病理、病生理及治疗等各方面研究提供较为可靠的工具,也为本课题第二部分“OSAHS病程中软腭部咽扩张肌不同时相病理改变”的研究奠定基础。
已经明确OSAHS患者均存在上气道结构性狭窄,并主要位于软腭平面或/和舌根平面。在临床中发现轻度患者多只存在或软腭或舌根的一个平面的狭窄,而重度患者却基本两个平面的狭窄同时存在。人的机体在疾病初期器官或系统一般都有代偿功能,如果没有及早进行治疗则组织器官会因过度代偿出现疾病,如地方性甲状腺肿可以发展成为结节性甲状腺肿大甚至甲亢,高血压久之可形成左心衰等等。同理,是否一个平面气道狭窄后另一平面气道咽扩张肌进行代偿性用力收缩,扩大气道截面积以增加通气量,但久之肌肉因过度劳损而发生松弛,反而出现继发性的这一部位气道的狭窄,使OSAHS的病情更加加重呢?到目前为止尚未查阅到是否一个平面气道狭窄会继发性引起另一个平面气道狭窄的研究。本实验以舌根后气道狭窄的OSAHS动物模型为基础连续观察了软腭部咽扩张肌的病理变化来研究这一问题。
根据整形外科原则,OSAHS是组织的过多和移位。我们根据软腭肌的解剖结构和治疗腭裂的经验设计了在软腭正中菱形切除部分软腭组织以短缩和上提软腭来扩大口咽腔的术式。并对这一术式进行了随访以明确手术效果。
研究方法1.以纯种成年雄性新西兰大白兔为实验动物,建立实验组和二组正常对照组,对实验组行颏舌肌内注射肉毒杆菌毒素,对一组对照组行颏舌肌注射生理盐水,另一对照组行股二头肌注射肉毒杆菌毒素,用药后均行动脉血氧饱和度监测,人工计算氧减饱和度指数(ODI_1),采用单因素方差分析,以P值＜0.05作为动物模型成功建立的标准;2.通过上述方法成功建立动物模型后,选取大白兔40只分成5组,每组8只。对照组不做任何处理,余4组均行颏舌肌注射肉毒杆菌毒素,每隔10天处死一组动物行软愕部咽扩张肌取材作病理学检查。通过电镜和光镜观察组织变化,以Nachlas法行肌组织中琥珀酸脱氢酶组织化学染色,研究酶含量变化。3.选择单纯存在软腭平面狭窄的OSAHS患者13例,运用整形外科原则中组织多多少去除多少的原则,根据软腭松弛程度在软腭正中菱形切除部分软腭粘膜及少许肌组织,使软腭短缩前翻,扩大口咽和鼻咽通气道。
结果1.动物行颏舌肌注射肉毒杆菌毒素后出现明显缺氧症状,血氧饱和度监测显示睡眠状态下出现间歇性低氧,其氧减饱和度指数与对照组进行统计学分析具有显著性差异;2.对行颏舌肌注射肉毒杆菌毒素后的动物,其软腭部咽扩张肌连续病理观察发现在第20天左右时显示出了琥珀酸脱氢酶染色加深,线粒体功能加强,肌纤维的增粗;第40余天时肌纤维组织出现渐萎缩的改变,线粒体肿胀嵴减少,琥珀酸脱氢酶染色变浅;3.应用软腭正中菱形切除短缩术共治疗了13例患者,术后随访至少6个月,效果良好,无明显并发症,软腭运动功能无明显障碍。
结论运用颏舌肌注射肉毒杆菌毒素能够成功地建立兔舌根后气道阻塞的OSAHS动物模型,这一模型更符合OSAHS病人发病原因和自然病程;OSAHS疾病过程中咽气道扩张肌具有代偿功能,但随病情的发展会发生失代偿的肌肉萎缩和松驰,从而发生继发平面气道的狭窄,所以要早期干预治疗;软腭正中菱形切除短缩术能够有效治疗软腭平面的气道狭窄,并减少了并发症的发生。
Objective To set up a reliable and dependable animal model is the precondition and instrument to study this disease. Now the research about obstructive sleep apnea—hypopnea syndrome (OSAHS)is still on the beginning,because pathogeny、pathology and pathophysiology are so complexed that construct a perfect model is difficult. Genioglossus muscle is the most powerful muscle of all the palatopharyngeal tensor muscles. When genioglossus is relaxed, the body of the tongue will be drop backwards to obstruct the upper airway and this is the main cause of OSAHS. Botulinum toxin is such a drug who can cause muscle laxation for a long time, and without any complication in the safe dosage. So we have such a presumption to construct the OSAHS model—injecting Botulinum toxin to genioglossus to cause the tongue fall backwars,then the space post the root of the tongue will change to stricture or closed. The objection to set up this animal model :the first is to provide an instrument for other one to research OSAHS,then make the base for our the second study—the continuous pathological changes of the soft palate in the OSAHS model obstructive post the root of the tongue.
Now it is definite that there are anatomy stricture in all the patients of OSAHS, and the position of the structure are mainly at the plate of soft palate or at the plate of the root of tongue. We have found there is only one plate happened structure in most mild patients while both of two plates have structure in all heavy patients. We all know any organs and systems have the function of compensation when at the beginning of the diseases,but if therapy hasnot been done at this time, excessive compensation will lead to real disease,for example simple goiter can develope to nodular goiter even hyperthyroidism; hypertension can lead to left cardiac ventricle to failure et al. we nature to think in the progress of OSAHS when one plate of the upper airway occurs structure then another plate's tensor m.will be construct strengthly to enlarge the size of the upper airway to increase ventilation capability, but the muscle will developed strain and laxation if having not any therapy. And the muscles relaxtion of the second plate will make this position to structure and decrease the ventilation capability, then the disease developed seriously. We want to research this question to continuously observe the soft palate histological change of the OSAHS model we make in the first part of our experiment.
As a plastic surgeon, we realize OSAHS and the cleft palate belong commonly to palatopharyngeal closed diseases, but OSAHS is the ventilation closed excessively and the cleft palte is the ventilation cannot closed entirely. Recording the principal of pastic surgery, the cleft palate is the tissue decrease and displace; and OSAHS is the tissue increase and displace, our operation is do a rhombus shape excision of the soft palate to therapy obstructive sleep apnea-hypopnea syndrome .Methods is resect of partial tissue in rhombus shape in the middle of soft palate and suture in transverse to lift soft palate to enlarge pharyngeal cavity, while without resecting the uvula. And we do a follow-up of 6 monthes at least.
Methods 1. we selected 28 the New Zealand White rabbits as experimental animals and were randomly classified into three groups ,one is trial group and two are control groups. We injct Botulinum toxin to Genioglossus as to trial group animals and inject equal volume normal sodium(NS) to one control group and inject equal volume BTX(Botulinum toxin ) to biceps femoris to other control group. We do continuous monitoring of all animals of SaO_2(arterial oxygen saturation) and compute the times of ODI_4(oxygen desaturation index four) .we do the results a student test and the standar of the successful animol model is P<0.05; 2. We selected 40 the New Zealand White rabbits and classified into 5 groups randomly and it is 8 animals in every group after we had maken OSAHS animal model successfully. The conrol group havenot any disposition and inject BTX to Genioglossus to another groups .then we put one the trial group to death and cut the sofe palate muscle to do a study of pathology every 10 days. We observe the histologcal changes through the light microscope and the electron microscope and do succinate dehydrogenase(SDH) staining with Nachlase and do a gray grade analyses .3. Recording the principal of the plastic surgery "just resect the tissue that is spare" and the degree of the relaxed soft palate ,we choiced 13 OSAHS patients just with the obstruction lied on the area of oropharynx and do a resection of partial tissue in rhombus shape in the middle of soft palate and suture in transverse to lift soft palate to enlarge pharyngeal cavity, while without resecting the uvula.
Results 1 .the animals were found the signs of hypoxia,cyanosis on the nose ,lips and the edge of ears.they have intermit SaO2 decreasing,and the significant differences exsisted between the trial group and two contral groups(P<0.05); 2.the colour of succinate dehydrogenase(SDH) staining has changed deeply on the trial group on 20days afterwe inject BTX to Genioglossus,and the construct of mitochondria is normal, myofilament change thicker. But succinate dehydrogenase(SDH) staining has changed lighter , myofilament change decreased and atrophia and crista of mitochondria reduced;3.we treated 13 OSAHS patients with A rhombus shape excision of the soft palate, we have done a follow-up of 6 monthes at least,and they all had an effective treatments and without serious complications,and the founction of kinesis of the soft palate is well.
Conclusions With the method of injecting BTX to rabbits' Genilglossus we can set up OSAHS animal model successfully and this kind of model is more to approach etiopathogenisis and pathology of the mankind. The palatopharyngeal tensor muscle has the compensation when one plate of the upper airway occurs stricture , but if therapy hasnot been done at this time, excessive compensation will lead to laxation and the muscles relaxtion of the second plate will make disease developed seriously. It is a vacious cycle. So it is necessary for OSAHS patients to see a doctor at the beginning of the initial stage of the disease. A rhombus shape excision of the soft palate had an effective treatments for OSAHS and without serious complications
已经明确OSAHS患者均存在上气道结构性狭窄,并主要位于软腭平面或/和舌根平面。在临床中发现轻度患者多只存在或软腭或舌根的一个平面的狭窄,而重度患者却基本两个平面的狭窄同时存在。人的机体在疾病初期器官或系统一般都有代偿功能,如果没有及早进行治疗则组织器官会因过度代偿出现疾病,如地方性甲状腺肿可以发展成为结节性甲状腺肿大甚至甲亢,高血压久之可形成左心衰等等。同理,是否一个平面气道狭窄后另一平面气道咽扩张肌进行代偿性用力收缩,扩大气道截面积以增加通气量,但久之肌肉因过度劳损而发生松弛,反而出现继发性的这一部位气道的狭窄,使OSAHS的病情更加加重呢?到目前为止尚未查阅到是否一个平面气道狭窄会继发性引起另一个平面气道狭窄的研究。本实验以舌根后气道狭窄的OSAHS动物模型为基础连续观察了软腭部咽扩张肌的病理变化来研究这一问题。
根据整形外科原则,OSAHS是组织的过多和移位。我们根据软腭肌的解剖结构和治疗腭裂的经验设计了在软腭正中菱形切除部分软腭组织以短缩和上提软腭来扩大口咽腔的术式。并对这一术式进行了随访以明确手术效果。
研究方法1.以纯种成年雄性新西兰大白兔为实验动物,建立实验组和二组正常对照组,对实验组行颏舌肌内注射肉毒杆菌毒素,对一组对照组行颏舌肌注射生理盐水,另一对照组行股二头肌注射肉毒杆菌毒素,用药后均行动脉血氧饱和度监测,人工计算氧减饱和度指数(ODI_1),采用单因素方差分析,以P值＜0.05作为动物模型成功建立的标准;2.通过上述方法成功建立动物模型后,选取大白兔40只分成5组,每组8只。对照组不做任何处理,余4组均行颏舌肌注射肉毒杆菌毒素,每隔10天处死一组动物行软愕部咽扩张肌取材作病理学检查。通过电镜和光镜观察组织变化,以Nachlas法行肌组织中琥珀酸脱氢酶组织化学染色,研究酶含量变化。3.选择单纯存在软腭平面狭窄的OSAHS患者13例,运用整形外科原则中组织多多少去除多少的原则,根据软腭松弛程度在软腭正中菱形切除部分软腭粘膜及少许肌组织,使软腭短缩前翻,扩大口咽和鼻咽通气道。
结果1.动物行颏舌肌注射肉毒杆菌毒素后出现明显缺氧症状,血氧饱和度监测显示睡眠状态下出现间歇性低氧,其氧减饱和度指数与对照组进行统计学分析具有显著性差异;2.对行颏舌肌注射肉毒杆菌毒素后的动物,其软腭部咽扩张肌连续病理观察发现在第20天左右时显示出了琥珀酸脱氢酶染色加深,线粒体功能加强,肌纤维的增粗;第40余天时肌纤维组织出现渐萎缩的改变,线粒体肿胀嵴减少,琥珀酸脱氢酶染色变浅;3.应用软腭正中菱形切除短缩术共治疗了13例患者,术后随访至少6个月,效果良好,无明显并发症,软腭运动功能无明显障碍。
结论运用颏舌肌注射肉毒杆菌毒素能够成功地建立兔舌根后气道阻塞的OSAHS动物模型,这一模型更符合OSAHS病人发病原因和自然病程;OSAHS疾病过程中咽气道扩张肌具有代偿功能,但随病情的发展会发生失代偿的肌肉萎缩和松驰,从而发生继发平面气道的狭窄,所以要早期干预治疗;软腭正中菱形切除短缩术能够有效治疗软腭平面的气道狭窄,并减少了并发症的发生。
Objective To set up a reliable and dependable animal model is the precondition and instrument to study this disease. Now the research about obstructive sleep apnea—hypopnea syndrome (OSAHS)is still on the beginning,because pathogeny、pathology and pathophysiology are so complexed that construct a perfect model is difficult. Genioglossus muscle is the most powerful muscle of all the palatopharyngeal tensor muscles. When genioglossus is relaxed, the body of the tongue will be drop backwards to obstruct the upper airway and this is the main cause of OSAHS. Botulinum toxin is such a drug who can cause muscle laxation for a long time, and without any complication in the safe dosage. So we have such a presumption to construct the OSAHS model—injecting Botulinum toxin to genioglossus to cause the tongue fall backwars,then the space post the root of the tongue will change to stricture or closed. The objection to set up this animal model :the first is to provide an instrument for other one to research OSAHS,then make the base for our the second study—the continuous pathological changes of the soft palate in the OSAHS model obstructive post the root of the tongue.
Now it is definite that there are anatomy stricture in all the patients of OSAHS, and the position of the structure are mainly at the plate of soft palate or at the plate of the root of tongue. We have found there is only one plate happened structure in most mild patients while both of two plates have structure in all heavy patients. We all know any organs and systems have the function of compensation when at the beginning of the diseases,but if therapy hasnot been done at this time, excessive compensation will lead to real disease,for example simple goiter can develope to nodular goiter even hyperthyroidism; hypertension can lead to left cardiac ventricle to failure et al. we nature to think in the progress of OSAHS when one plate of the upper airway occurs structure then another plate's tensor m.will be construct strengthly to enlarge the size of the upper airway to increase ventilation capability, but the muscle will developed strain and laxation if having not any therapy. And the muscles relaxtion of the second plate will make this position to structure and decrease the ventilation capability, then the disease developed seriously. We want to research this question to continuously observe the soft palate histological change of the OSAHS model we make in the first part of our experiment.
As a plastic surgeon, we realize OSAHS and the cleft palate belong commonly to palatopharyngeal closed diseases, but OSAHS is the ventilation closed excessively and the cleft palte is the ventilation cannot closed entirely. Recording the principal of pastic surgery, the cleft palate is the tissue decrease and displace; and OSAHS is the tissue increase and displace, our operation is do a rhombus shape excision of the soft palate to therapy obstructive sleep apnea-hypopnea syndrome .Methods is resect of partial tissue in rhombus shape in the middle of soft palate and suture in transverse to lift soft palate to enlarge pharyngeal cavity, while without resecting the uvula. And we do a follow-up of 6 monthes at least.
Methods 1. we selected 28 the New Zealand White rabbits as experimental animals and were randomly classified into three groups ,one is trial group and two are control groups. We injct Botulinum toxin to Genioglossus as to trial group animals and inject equal volume normal sodium(NS) to one control group and inject equal volume BTX(Botulinum toxin ) to biceps femoris to other control group. We do continuous monitoring of all animals of SaO_2(arterial oxygen saturation) and compute the times of ODI_4(oxygen desaturation index four) .we do the results a student test and the standar of the successful animol model is P<0.05; 2. We selected 40 the New Zealand White rabbits and classified into 5 groups randomly and it is 8 animals in every group after we had maken OSAHS animal model successfully. The conrol group havenot any disposition and inject BTX to Genioglossus to another groups .then we put one the trial group to death and cut the sofe palate muscle to do a study of pathology every 10 days. We observe the histologcal changes through the light microscope and the electron microscope and do succinate dehydrogenase(SDH) staining with Nachlase and do a gray grade analyses .3. Recording the principal of the plastic surgery "just resect the tissue that is spare" and the degree of the relaxed soft palate ,we choiced 13 OSAHS patients just with the obstruction lied on the area of oropharynx and do a resection of partial tissue in rhombus shape in the middle of soft palate and suture in transverse to lift soft palate to enlarge pharyngeal cavity, while without resecting the uvula.
Results 1 .the animals were found the signs of hypoxia,cyanosis on the nose ,lips and the edge of ears.they have intermit SaO2 decreasing,and the significant differences exsisted between the trial group and two contral groups(P<0.05); 2.the colour of succinate dehydrogenase(SDH) staining has changed deeply on the trial group on 20days afterwe inject BTX to Genioglossus,and the construct of mitochondria is normal, myofilament change thicker. But succinate dehydrogenase(SDH) staining has changed lighter , myofilament change decreased and atrophia and crista of mitochondria reduced;3.we treated 13 OSAHS patients with A rhombus shape excision of the soft palate, we have done a follow-up of 6 monthes at least,and they all had an effective treatments and without serious complications,and the founction of kinesis of the soft palate is well.
Conclusions With the method of injecting BTX to rabbits' Genilglossus we can set up OSAHS animal model successfully and this kind of model is more to approach etiopathogenisis and pathology of the mankind. The palatopharyngeal tensor muscle has the compensation when one plate of the upper airway occurs stricture , but if therapy hasnot been done at this time, excessive compensation will lead to laxation and the muscles relaxtion of the second plate will make disease developed seriously. It is a vacious cycle. So it is necessary for OSAHS patients to see a doctor at the beginning of the initial stage of the disease. A rhombus shape excision of the soft palate had an effective treatments for OSAHS and without serious complications
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