表没食子儿茶素没食子酸酯对早期压疮大鼠模型缺血再灌注损伤的保护作用及NLRP3炎症小体相关机制研究
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摘要
目的:探讨表没食子儿茶素没食子酸酯(EGCG)对早期压疮的保护作用及初步机制。方法:雌性SD大鼠随机分为空白对照组、模型对照组和EGCG 10、20、40mg/kg剂量组,灌胃给予相应药物,连续7d后,用自制装置对大鼠进行早期压疮造模。造模完成后观察各组大鼠受压局部组织情况,并眼眶采血,组织取材,酶联免疫吸附法测定血清中IL-1β、LDH、CK的含量;HE染色观察各组大鼠组织病变,判断压疮病理变化严重程度;蛋白质印迹法测定大鼠受压组织中NLRP3、caspase-1、IL-1β蛋白的表达水平。结果:造模成功率为100%;实验组大鼠均存在皮肤层次欠清晰,皮下结缔组织和肌肉组织有炎性反应,病理变化严重程度随EGCG剂量的增加而减轻,大鼠血清中IL-1β、LDH、CK含量随EGCG剂量的增加明显降低。EGCG各剂量组大鼠受压组织中NLRP3、IL-1β、caspase-1蛋白表达显著降低,呈一定的剂量依赖性。结论:EGCG预处理可通过抑制炎症而减轻早期压疮缺血再灌注损伤,其作用机制可能主要与抑制NLRP3炎症小体的表达有关。
Objective: To investigate the protective effects of epigallocatechin gallate( EGCG) on ischemia-reperfusion injury( IRI) in early stage pressure ulcers and its possible mechanism. Methods: Female SD rats were randomly divided into the control group,the model group and EGCG groups at the doses of 10 mg/kg、20 mg/kg and 40 mg/kg,10 rats in each group. All animals were intraperitoneally injected with corresponding drugs for 7 days,then the pressure ulcer model was established by using a self-made simple modeling device. The compressed tissues of rats in each group were observed 4 hours after the modeling was completed,and the incidence and index of pressure ulcers were calculated. The blood was collected from the eyelids,then the rats were sacrificed and the tissues were taken. ELISA Kit was used to detect the serum levels of IL-1β,LDH and CK. Western-blot methods were used to determine the expression levels of NLRP3,caspase-1,and IL-1β in tissues. Results: The early stage pressure ulcer model was successfully established. Compared with the model group,the inflammatory responses in EGCG groups were significantly alleviated with the increase of the dose of EGCG. The skin level of the rats was not clear,and inflammatory reaction was observed in the subcutaneous connective tissue and muscle tissue. The pathological changes were improved with the increase of the concentration. The levels of IL-1β,LDH and CK in rats serum were significantly decreased with the increase of doses of EGCG.The expressions of Nlrp3,IL-1β and caspase-1 in the pressed tissues in EGCG groups was significantly decreased in a dose-dependent manner. Conclusion: EGCG pretreatment can reduce the ischemia-reperfusion injury in pressure ulcer by inhibiting inflammation. Its mechanism may be related to the inhibition of expression of Nlrp3 inflammatory bodies.
Objective: To investigate the protective effects of epigallocatechin gallate( EGCG) on ischemia-reperfusion injury( IRI) in early stage pressure ulcers and its possible mechanism. Methods: Female SD rats were randomly divided into the control group,the model group and EGCG groups at the doses of 10 mg/kg、20 mg/kg and 40 mg/kg,10 rats in each group. All animals were intraperitoneally injected with corresponding drugs for 7 days,then the pressure ulcer model was established by using a self-made simple modeling device. The compressed tissues of rats in each group were observed 4 hours after the modeling was completed,and the incidence and index of pressure ulcers were calculated. The blood was collected from the eyelids,then the rats were sacrificed and the tissues were taken. ELISA Kit was used to detect the serum levels of IL-1β,LDH and CK. Western-blot methods were used to determine the expression levels of NLRP3,caspase-1,and IL-1β in tissues. Results: The early stage pressure ulcer model was successfully established. Compared with the model group,the inflammatory responses in EGCG groups were significantly alleviated with the increase of the dose of EGCG. The skin level of the rats was not clear,and inflammatory reaction was observed in the subcutaneous connective tissue and muscle tissue. The pathological changes were improved with the increase of the concentration. The levels of IL-1β,LDH and CK in rats serum were significantly decreased with the increase of doses of EGCG.The expressions of Nlrp3,IL-1β and caspase-1 in the pressed tissues in EGCG groups was significantly decreased in a dose-dependent manner. Conclusion: EGCG pretreatment can reduce the ischemia-reperfusion injury in pressure ulcer by inhibiting inflammation. Its mechanism may be related to the inhibition of expression of Nlrp3 inflammatory bodies.
引文
[1]Thomas DR. Clinical management of pressure ulcers. Clin Geriatr Med, 2013, 29 (2)∶397～413.
[2]Van Gilder C, Amlung S, Harrison P, et al. Results of the 2008-2009 international pressure ulcer prevalence survey and a 3- year, acute care, unit-specific analysis. Ostomy Wound Manage, 2009, 55 (11)∶39～45.
[3]Tannen A, Dassen T, Halfens R. Differences in prevalence of pressure ulcers between the Netherlands and Germany-associations between risk, prevention and occurrence of pressure ulcers in hospitals and nursing homes. J Clin Nurs, 2009, 18(10)∶1511～1512.
[4]Lyder C H, Wang Y, Metersky M, et al. Hospital-acquired pressure ulcers: results from the national medicare patient safety monitoring system study. J Am Geriatr Soc, 2012, 60(9)∶1603～1608.
[5]Black J, Baharestani M M, Cuddigan J, et al. National Pressure Ulcer Advisory Panel's updated pressure ulcer staging system. Adv Skin Wound Care, 2007, 20(5)∶269～274.
[6]Keelaghan E, Margolis D, Zhan M, et al. Prevalence of pressure ulcers on hospital admission among nursing home residents transferred to the hospital. Wound Repair Regen, 2008, 16( 3) ∶331～336.
[7]Sibbald R G, Goodman L, Norton L, et al. Prevention and treatment of pressure ulcers. Skin Therapy Lett, 2012, 17(8)∶4～7.
[8]Sunn G. Spinal cord injury pressure ulcer treatment: an experience-based approach. Phys Med Rehabil Clin N Am, 2014, 25 (3)∶671～680.
[9]Thomas D R. Role of nutrition in the treatment and prevention of pressure ulcers. Nutr Clin Pract, 2014, 29(4)∶466～472.
[10]Pae M, Wu D. Immunomodulating effects of epigallocatechin-3-gallate from green tea: mechanisms and applications. Food Funct, 2013, 4(9)∶1287～1303.
[11]Camfield D A, Stough C, Farrimond J, et al. Acute effects of tea-constituents L-theanine, caffeine, and epigallocatechin gallate on cognitive function and mood: a systematic review and meta-analysis. Nutr Rev, 2014, 72(8)∶507～522.
[12]Akizawa Y, Kitazato T, Kishimoto H, et al. Effects of antioxidants on drug absorption in in vivo intestinal ischemia /reperfusion. Eur J Drug Metab Pharmacokinet, 2011, 35(3-4)∶89～95.
[13]Zhang F, Li N, Jiang L, et al. Neuroprotective effects of (-)-Epigallocatechin-3-Gallate against focal cerebral ischemia/ reperfusion injury in rats through attenuation of inflammation. Neurochem Res, 2015, 40(8)∶1691～1698.
[14]Lee S K, Kim J H, Kim J S, et al. Polyphenol (-)-epigallocatechingallate-induced cardioprotection may attenuate ischemia-reperfusion injury through adenosine receptor activation: a preliminary study. Korean J Anesthesiol, 2012, 63(4)∶340～345.
[15]郭玉怀, 孙忠人, 孙琦, 等. 简易大鼠压疮缺血再灌注模型造模装置的设计及应用. 护理研究, 2015, 29(5)∶592～593.
[16]Zweier J L,Talukder M A. The role of oxidants and free radicals in reperfusion injury. Cardiovas Res, 2006, 70(2)∶181～190.
[17]姜丽萍,蔡福满,杨晔琴,等.局部皮肤持续受压致压疮的实验研究.解放军护理杂志, 2007, 24(12A)∶4～8.
[18]Baroja-Mazo A, Martín-Sánchez F, Gomez AI, et al. The NLRP3 inflammasome is released as a particulate danger signal that amplifies the inflammatory response. Nat Immunol, 2014, 15(8)∶738～748.
[19]Minton K. Inflammasome: fishing for anti-inflammatory mechanisms. Nat Rev Immunol, 2013, 13(8)∶545～545.
[20]Savage C D, Lopez-Castejon G, Denes A, et al. NLRP3 inflammasome activating DAMPs stimulate an inflammatory response in glia in the absence of priming which contributes to brain inflammation after injury. Front Immunol, 2012, 3∶Article 288(1～11).
[21]Gefen A, Gefen N, LinderGanz E, et al. In vivo muscle tiffening under bone compression promotes deep pressure sores. J Biomech Eng, 2005, 127(3)∶512～524.
[2]Van Gilder C, Amlung S, Harrison P, et al. Results of the 2008-2009 international pressure ulcer prevalence survey and a 3- year, acute care, unit-specific analysis. Ostomy Wound Manage, 2009, 55 (11)∶39～45.
[3]Tannen A, Dassen T, Halfens R. Differences in prevalence of pressure ulcers between the Netherlands and Germany-associations between risk, prevention and occurrence of pressure ulcers in hospitals and nursing homes. J Clin Nurs, 2009, 18(10)∶1511～1512.
[4]Lyder C H, Wang Y, Metersky M, et al. Hospital-acquired pressure ulcers: results from the national medicare patient safety monitoring system study. J Am Geriatr Soc, 2012, 60(9)∶1603～1608.
[5]Black J, Baharestani M M, Cuddigan J, et al. National Pressure Ulcer Advisory Panel's updated pressure ulcer staging system. Adv Skin Wound Care, 2007, 20(5)∶269～274.
[6]Keelaghan E, Margolis D, Zhan M, et al. Prevalence of pressure ulcers on hospital admission among nursing home residents transferred to the hospital. Wound Repair Regen, 2008, 16( 3) ∶331～336.
[7]Sibbald R G, Goodman L, Norton L, et al. Prevention and treatment of pressure ulcers. Skin Therapy Lett, 2012, 17(8)∶4～7.
[8]Sunn G. Spinal cord injury pressure ulcer treatment: an experience-based approach. Phys Med Rehabil Clin N Am, 2014, 25 (3)∶671～680.
[9]Thomas D R. Role of nutrition in the treatment and prevention of pressure ulcers. Nutr Clin Pract, 2014, 29(4)∶466～472.
[10]Pae M, Wu D. Immunomodulating effects of epigallocatechin-3-gallate from green tea: mechanisms and applications. Food Funct, 2013, 4(9)∶1287～1303.
[11]Camfield D A, Stough C, Farrimond J, et al. Acute effects of tea-constituents L-theanine, caffeine, and epigallocatechin gallate on cognitive function and mood: a systematic review and meta-analysis. Nutr Rev, 2014, 72(8)∶507～522.
[12]Akizawa Y, Kitazato T, Kishimoto H, et al. Effects of antioxidants on drug absorption in in vivo intestinal ischemia /reperfusion. Eur J Drug Metab Pharmacokinet, 2011, 35(3-4)∶89～95.
[13]Zhang F, Li N, Jiang L, et al. Neuroprotective effects of (-)-Epigallocatechin-3-Gallate against focal cerebral ischemia/ reperfusion injury in rats through attenuation of inflammation. Neurochem Res, 2015, 40(8)∶1691～1698.
[14]Lee S K, Kim J H, Kim J S, et al. Polyphenol (-)-epigallocatechingallate-induced cardioprotection may attenuate ischemia-reperfusion injury through adenosine receptor activation: a preliminary study. Korean J Anesthesiol, 2012, 63(4)∶340～345.
[15]郭玉怀, 孙忠人, 孙琦, 等. 简易大鼠压疮缺血再灌注模型造模装置的设计及应用. 护理研究, 2015, 29(5)∶592～593.
[16]Zweier J L,Talukder M A. The role of oxidants and free radicals in reperfusion injury. Cardiovas Res, 2006, 70(2)∶181～190.
[17]姜丽萍,蔡福满,杨晔琴,等.局部皮肤持续受压致压疮的实验研究.解放军护理杂志, 2007, 24(12A)∶4～8.
[18]Baroja-Mazo A, Martín-Sánchez F, Gomez AI, et al. The NLRP3 inflammasome is released as a particulate danger signal that amplifies the inflammatory response. Nat Immunol, 2014, 15(8)∶738～748.
[19]Minton K. Inflammasome: fishing for anti-inflammatory mechanisms. Nat Rev Immunol, 2013, 13(8)∶545～545.
[20]Savage C D, Lopez-Castejon G, Denes A, et al. NLRP3 inflammasome activating DAMPs stimulate an inflammatory response in glia in the absence of priming which contributes to brain inflammation after injury. Front Immunol, 2012, 3∶Article 288(1～11).
[21]Gefen A, Gefen N, LinderGanz E, et al. In vivo muscle tiffening under bone compression promotes deep pressure sores. J Biomech Eng, 2005, 127(3)∶512～524.