中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2014年
14期
1087-1091
,共5页
马丽%柴家科%褚万立%段红杰%李大伟%郁永辉
馬麗%柴傢科%褚萬立%段紅傑%李大偉%鬱永輝
마려%시가과%저만립%단홍걸%리대위%욱영휘
丹曲林%烧伤%肌,骨骼%大鼠%钙蛋白酶
丹麯林%燒傷%肌,骨骼%大鼠%鈣蛋白酶
단곡림%소상%기,골격%대서%개단백매
Dantrolene%Burns%Muscle,skeletal%Rats%Calpain
目的：探讨丹曲林对严重烧伤大鼠骨骼肌损害的治疗作用及可能机制。方法选取清洁级Wistar雄性大鼠56只，按照随机数字表法分为对照组8只、致伤组及丹曲林组各24只（两组3个时相点各8只）。致伤组和丹曲林组大鼠在麻醉后与其背部及腹部行50％总体表面积（ TBSA ）Ⅲ度烫伤，伤后腹腔注射林格平衡液抗休克，致伤组尾静脉注射5％甘露醇，丹曲林组尾静脉注射丹曲林钠2 mg／kg（溶于5％甘露醇）。对照组行假烫伤，其余操作同致伤组。于伤后第1、4、7天收集各组大鼠胫骨前肌组织，透射电镜观察组织超微结构，电子探针X射线显微分析法检测肌细胞微区Ca2＋含量，蛋白印迹方法检测胫骨前肌总蛋白中钙蛋白酶1、2表达，酶法检测胫骨前肌中钙蛋白酶的活性。结果致伤组胫骨前肌肌丝伤后第1天出现排列混杂，第7天出现局部Z线消失。而丹曲林组第1、4天肌组织超微结构与对照相比无显著变化，仅在第7天肌丝存现排列卷曲、轻度断裂。伤后第1、4天致伤组胫骨前肌细胞质Ca2＋水平显著高于对照组[（0.964±0.060）、（0.639±0.067）比（0.266±0.029）μmol／L]，肌浆网Ca2＋水平显著低于对照组[（0.368±0.060）、（0.814±0.089）比（1.337±0.112）μmol／L]（均P＜0.05）；而丹曲林组细胞质和肌浆网Ca2＋水平[（0.310±0.069）、（0.490±0.039）和（1.241±0.073）、（1.161±0.094）μmol／L]与对照组差异均无统计学意义（均P＞0.05）。伤后第1、4天致伤组胫骨前肌钙蛋白酶1和钙蛋白酶2相对表达量均显著高于对照组（1.371±0.034、1.214±0.030比0.838±0.017和1.464±0.015、1.390±0.023比0.806±0.026）（均P＜0.05），而丹曲林组钙蛋白酶1和钙蛋白酶2相对表达量（0.984±0.031、0.935±0.023和0.836±0.014与0.741±0.020）均显著低于致伤组（均P＜0.05）。伤后第1、4、7天致伤组和丹曲林组钙蛋白酶活性分别为（8.33±0.21）、（9.33±0.21）、（10.59±0.18）和（7.76±0.28）、（7.86±0.20）、（7.91±0.22）μmol／L，对照组为（7.62±0.19）μmol／L，致伤组均显著高于丹曲林组及对照组（均P＜0.05），但丹曲林组与对照组差异均无统计学意义（均P＞0.05）。结论丹曲林能够明显减弱严重烧伤引起的骨骼肌超微结构变化，减轻肌组织损伤。其可能是通过抑制严重烧伤后肌细胞肌浆网Ca2＋的过度释放，降低严重烧伤后肌细胞质内钙蛋白酶1、2的表达及其活性而发挥作用。
目的：探討丹麯林對嚴重燒傷大鼠骨骼肌損害的治療作用及可能機製。方法選取清潔級Wistar雄性大鼠56隻，按照隨機數字錶法分為對照組8隻、緻傷組及丹麯林組各24隻（兩組3箇時相點各8隻）。緻傷組和丹麯林組大鼠在痳醉後與其揹部及腹部行50％總體錶麵積（ TBSA ）Ⅲ度燙傷，傷後腹腔註射林格平衡液抗休剋，緻傷組尾靜脈註射5％甘露醇，丹麯林組尾靜脈註射丹麯林鈉2 mg／kg（溶于5％甘露醇）。對照組行假燙傷，其餘操作同緻傷組。于傷後第1、4、7天收集各組大鼠脛骨前肌組織，透射電鏡觀察組織超微結構，電子探針X射線顯微分析法檢測肌細胞微區Ca2＋含量，蛋白印跡方法檢測脛骨前肌總蛋白中鈣蛋白酶1、2錶達，酶法檢測脛骨前肌中鈣蛋白酶的活性。結果緻傷組脛骨前肌肌絲傷後第1天齣現排列混雜，第7天齣現跼部Z線消失。而丹麯林組第1、4天肌組織超微結構與對照相比無顯著變化，僅在第7天肌絲存現排列捲麯、輕度斷裂。傷後第1、4天緻傷組脛骨前肌細胞質Ca2＋水平顯著高于對照組[（0.964±0.060）、（0.639±0.067）比（0.266±0.029）μmol／L]，肌漿網Ca2＋水平顯著低于對照組[（0.368±0.060）、（0.814±0.089）比（1.337±0.112）μmol／L]（均P＜0.05）；而丹麯林組細胞質和肌漿網Ca2＋水平[（0.310±0.069）、（0.490±0.039）和（1.241±0.073）、（1.161±0.094）μmol／L]與對照組差異均無統計學意義（均P＞0.05）。傷後第1、4天緻傷組脛骨前肌鈣蛋白酶1和鈣蛋白酶2相對錶達量均顯著高于對照組（1.371±0.034、1.214±0.030比0.838±0.017和1.464±0.015、1.390±0.023比0.806±0.026）（均P＜0.05），而丹麯林組鈣蛋白酶1和鈣蛋白酶2相對錶達量（0.984±0.031、0.935±0.023和0.836±0.014與0.741±0.020）均顯著低于緻傷組（均P＜0.05）。傷後第1、4、7天緻傷組和丹麯林組鈣蛋白酶活性分彆為（8.33±0.21）、（9.33±0.21）、（10.59±0.18）和（7.76±0.28）、（7.86±0.20）、（7.91±0.22）μmol／L，對照組為（7.62±0.19）μmol／L，緻傷組均顯著高于丹麯林組及對照組（均P＜0.05），但丹麯林組與對照組差異均無統計學意義（均P＞0.05）。結論丹麯林能夠明顯減弱嚴重燒傷引起的骨骼肌超微結構變化，減輕肌組織損傷。其可能是通過抑製嚴重燒傷後肌細胞肌漿網Ca2＋的過度釋放，降低嚴重燒傷後肌細胞質內鈣蛋白酶1、2的錶達及其活性而髮揮作用。
목적：탐토단곡림대엄중소상대서골격기손해적치료작용급가능궤제。방법선취청길급Wistar웅성대서56지，안조수궤수자표법분위대조조8지、치상조급단곡림조각24지（량조3개시상점각8지）。치상조화단곡림조대서재마취후여기배부급복부행50％총체표면적（ TBSA ）Ⅲ도탕상，상후복강주사림격평형액항휴극，치상조미정맥주사5％감로순，단곡림조미정맥주사단곡림납2 mg／kg（용우5％감로순）。대조조행가탕상，기여조작동치상조。우상후제1、4、7천수집각조대서경골전기조직，투사전경관찰조직초미결구，전자탐침X사선현미분석법검측기세포미구Ca2＋함량，단백인적방법검측경골전기총단백중개단백매1、2표체，매법검측경골전기중개단백매적활성。결과치상조경골전기기사상후제1천출현배렬혼잡，제7천출현국부Z선소실。이단곡림조제1、4천기조직초미결구여대조상비무현저변화，부재제7천기사존현배렬권곡、경도단렬。상후제1、4천치상조경골전기세포질Ca2＋수평현저고우대조조[（0.964±0.060）、（0.639±0.067）비（0.266±0.029）μmol／L]，기장망Ca2＋수평현저저우대조조[（0.368±0.060）、（0.814±0.089）비（1.337±0.112）μmol／L]（균P＜0.05）；이단곡림조세포질화기장망Ca2＋수평[（0.310±0.069）、（0.490±0.039）화（1.241±0.073）、（1.161±0.094）μmol／L]여대조조차이균무통계학의의（균P＞0.05）。상후제1、4천치상조경골전기개단백매1화개단백매2상대표체량균현저고우대조조（1.371±0.034、1.214±0.030비0.838±0.017화1.464±0.015、1.390±0.023비0.806±0.026）（균P＜0.05），이단곡림조개단백매1화개단백매2상대표체량（0.984±0.031、0.935±0.023화0.836±0.014여0.741±0.020）균현저저우치상조（균P＜0.05）。상후제1、4、7천치상조화단곡림조개단백매활성분별위（8.33±0.21）、（9.33±0.21）、（10.59±0.18）화（7.76±0.28）、（7.86±0.20）、（7.91±0.22）μmol／L，대조조위（7.62±0.19）μmol／L，치상조균현저고우단곡림조급대조조（균P＜0.05），단단곡림조여대조조차이균무통계학의의（균P＞0.05）。결론단곡림능구명현감약엄중소상인기적골격기초미결구변화，감경기조직손상。기가능시통과억제엄중소상후기세포기장망Ca2＋적과도석방，강저엄중소상후기세포질내개단백매1、2적표체급기활성이발휘작용。
Objective To explore the effect and mechanism of ryanodine receptor antagonist dantrolene on skeletal muscle of rats with severe scald injury.Methods A total of 56 Wistar rats were divided into control , scald and dantrolene treatment groups according to a random digital table.Rats in scald and dantrolene treatment groups were subject to 50%total body surface area ( TBSA) full-thickness scald by a 12-second immersion of back and a 6-second immersion of abdomen in 94 ℃water and then received an intraperitoneal injection of Ringer′s solution.At the same time , the rats in scald group received 5%mannitol through caudal vein while those in dantrolene treatment group received dantrolene 2 mg/kg ( dissolved in 5%mannitol ).Rats in control group were sham-injured through an immersion of back and abdomen into 37 ℃warm water.Tibialis anterior muscle samples were harvested at Days 1, 4 and 7 post-scalding.Changes of skeletal muscle ultrastructure were observed by transmission electron microscope , subcellular calcium ion ( Ca2+) contents of skeletal muscle ( including cytoplasm , mitochondria & sarcoplasm reticulum ) were detected by electron probe X-ray microanalysis ( EPMA) and the levels of calpain-1 and calpain-2 protein were determined by Western blot.And the activities of calpain were detected by enzyme-linked immunosorbent assay.Results In scald group , assorted arrangement appeared immediately at Day 1 post-injury and partial disappearance of Z lines at Day 7 post-injury.There were no significant ultrastructure changes in dantrolene treatment group at Day 1 and 4 post-injury.Curled filament and mild fracture occurred merely in dantrolene treatment group at Day 7 post-injury.The cytoplasmic contents of Ca 2+ were significantly higher in scald group than those in control group at Day 1 and 4 ((0.964 ±0.060), (0.639 ± 0.067) vs (0.266 ±0.029) μmol/L respectively, all P<0.05) while the contents of Ca2+within sarcoplasm reticulum were obviously lower in scald group than those in control group at Day 1 and 4 ((0.368 ±0.060), (0.814 ±0.089) vs (1.337 ±0.112) μmol/L respectively, all P<0.05).However, those subcellular regions in dantrolene treatment group ( ( 0.310 ±0.069 ) , ( 0.490 ±0.039 ) and ( 1.241 ±0.073 ) , (1.161 ±0.094) μmol/L) had no significant difference with control group ( all P>0.05).Caplain-1 and calpain-2 protein levels in scald group increased significantly at Day 1 and 4 post-injury versus control group (1.371 ±0.034, 1.214 ±0.030 vs 0.838 ±0.017 & 1.464 ±0.015, 1.390 ±0.023 vs 0.806 ±0.026 respectively , all P <0.05 ) , whereas caplain-1 and calpain-2 protein levels in dantrolene treatment (0.984 ±0.031, 0.935 ±0.023 and 0.836 ±0.014, 0.741 ±0.020) obviously were lower than those in scald group respectively ( all P<0.05).The activities of calpain in scald and dantrolene treatment groups at Day 1, 4 and 7 post-injury were (8.33 ±0.21), (9.33 ±0.21), (10.59 ±0.18) and (7.76 ±0.28), (7.86 ±0.20), (7.91 ±0.22) μmol/L respectively while the activity of calpain in control group was (7.62 ±0.19) μmol/L.The activities of calpain in scald group were significantly higher than those in dantrolene treatment and control groups ( all P <0.05 ) whereas the activities of calpain in dantrolene treatment group had no obvious change versus control group ( all P>0.05 ) .Conclusions Dantrolene offers significant protection from skeletal muscle tissue damage and minimizes the ultrastructural change of tibialis anterior muscle induced by severe scald injury.The mechanism is probably through inhibiting an excessive release of Ca 2+within sarcoplasm reticulum and down-regulated cytoplasmic expression and activity of calpain-1 and calpain-2.
