Application Study Of Stable Isotope Technique On Burn Metabolism

和稳定同位素碳13标记的谷氨酸(13C-Glutamic acid)作为示踪剂,以严重烧伤患者为研究对象,以首剂冲击与维持给药为示踪氨基酸输注方式,以口服和静脉7小时交叉给药途径,在血液中示踪氨基酸达到稳态是采集标本,加入内标13C-Glutamic acidM+5、13C-GlutamineM+5。衍生,GC-MC分析,量化谷氨酰胺代谢效能。结果：1.(1)第一批5组大鼠烫伤创面深度差异有统计学意义,随着烫伤时间增加,烫伤深度成线性加深趋势(p<0.05)。(2)第一批5组大鼠烫伤创面愈合时间差异有统计学意义(F=167.3411,p=0.0000<0.05),随着烫伤时间增加,烫伤创面愈合时间成线性延长趋势。(3)3D扫描组烫伤面积与解剖测量面积无统计学差异(t=0.689,p=0.5008>0.05)。2.(1)烫伤组血浆游离氨基酸2H丰度明显高于假伤组,烫伤组较假伤组增加了48.24%,差异有统计学意义(p<0.05)。(2)烫伤组亮氨酸更新速率为(236.2±13.1)μmol/(kg·h),假伤组为(132.2±6.5)μmol/(kg·h),烧伤组较对照组增加了78.6%,差异有统计学意义(p<0.05)。(3)烫伤组大鼠整体蛋白分解速率为(9.1±1.6)g/kg·d,较假伤组(5.1±0.5)g/kg·d有明显增加,前者较后者增高78.4%(p<0.05)。(4)烫伤组大鼠整体蛋白合成速率为(5.1±1.8)g/kg·d,与假伤组(3.8±1.1)g/kg·d差异无统计学意义(p=0.067>0.05)。(5)烫伤大鼠为负氮平衡状态(-4.0±1.0)g/kg·d。3.(1)在以3.26mg/kg.h的标准维持胃肠给予患者谷氨酸的情况下,胃肠吸收的谷氨酸有将有(1.35±0.28)%(n=12,曲线为正态分布)转化为体内的谷氨酰胺。(2)在以1.32mg/kg-h的标准维持静脉给予患者谷氨酰胺的情况下,外源性的谷氨酰胺的(81.4±8.1)%(n=12,曲线为正态分布)参与代谢被消耗。结论：应用94℃热水接触大鼠背部12s可以建立稳定的深Ⅱ度～Ⅲ度烫伤大鼠动物模型,3D扫描技术能够精确计算大鼠背部烫伤面积；在严重烫伤大鼠模型的基础上,烫伤4d后,大鼠总体蛋白分解率明显增高,超过假伤对照组50%以上,而总体蛋白质合成率差异不显著；严重烧伤患者,口服使用谷氨酸能够部分转化为血浆中的谷氨酰胺,但比例较小,效能低,静脉使用谷氨酰胺时,仍会有部分谷氨酰胺直接转化为谷氨酸,以谷氨酸的形式起到营养作用。 14s respectively. The burnt skin samples were collected24h after the injury to investigatethe depth of the scalded wound, skin appendages remaining. Furthermore, wound healingtime after constant rearing was also recorded. The rest9rats were left for3D scanning.The backs of these rats are immersed in94℃water for12s and the burned skin area wasidentified by the3D scanner with binocular vision24h later. The result was comparedwith the data measured through anatomy.2. Measurement of overall protein metabolic rate of burned SD rats: The aboveburned animal model was further used. Twenty male SD rats were randomly divided into2groups:(1) sham scald group (n=10),(2) scald group (n=10). All the rats were injectedwith the tracer amino acid solution,2H–leucine,4d post injury., Blood samples weretaken from celiac axis0.5h after the injection. Thereafter, serum were treated with leucinderivation, KIC derivation, amino acid combination after protein precipitation. Then, thetreated serum were subjected to GC-MC detection. The plasma free-2H amino acidabundance, leucine updating rate, protein degradation rate and protein incorporation rateof the two groups of rats were calculated and analyzed.3. Study on efficacy of glutamine metabolism:15N-Glutamine and13C-Glutamicacid were used as tracers in studying on severely burned patients. The amino acid tracerswere given pulse infusion on the first use, followed by sustaining dose as instillation. Thetracer solution were given per os and through vein alternately every7h. Samples werecollected when the amino acid tracer was stable in the blood, followed bysupplementation with13C-Glutamic acidM+5,13C-GlutamineM+5. The samples werederived and analyzed by GC-MC to quantify the efficacy of glutamine metabolism.Results:1.(1) The differences in depth of scalded wound was significant among thefirst five groups. With the prolonged burning time, the depth of wound increased inlinear dependence.(p<0.05).(2) There was notable differences in wound healing timeamong the first five groups (F=167.3411，p=0.0000＜0.05). The burning time wascorrelated with the healing time in linear manner.(3) There was no statistical differencebetween the3D scannning and the anatomical measurement(t=0.689，p=0.5008＞0.05).2.(1) The plasma free-2H amino acid abundance in the scald group was48.24% higher than that in the sham scald group. The difference was remarkable (p<0.05).(2)The leucine updating rate in scald group was (236.2±13.1)μmol/kg·h, while that of thesham scald group was (132.2±6.5)μmol/kg·h. The former was78.6%higher than thelatter and the difference had statistical significance (p<0.05).(3) The overall proteindegradation rate in the scald group was (9.1±1.6)g/kg·d much higher than that in thesham scald group,(5.1±0.5)g/kg·d. The former was78.4%more than the latter (p<0.05).(4) As for the protein incorporation rate, there was no statistical difference (p=0.067﹥0.05) between (5.1±1.8)g/kg·d of the scald group and (3.8±1.1)g/kg·d of the sham scaldgroup.(5) Negative nitrogen balance was found in the scald group (-4.0±1.0).g/kg·d3.(1) When the patient was given glutamic acid to the gastrointestinal tract with asustained flow of3.26mg/kg.h,(1.35.±0.28)%of the glutamic acid absorbed throughgastrointestinal tract was transformed to the blood glutamine(n=12, normal distribution).(2) When the patient was given glutamine through vein with a sustained infusion of1.32mg/kg.h,(81.4±8.1)%of the exogenous glutamine was consumed(n=12, normaldistribution).Conclusion: A stable deep Ⅱ-III degree burned SD rat model can be established byimmersing the back of rats to the94℃water for12s.3D scanning technology can beused to calculate the size of the back burning area precisely. On4d post burn, the proteindegradation rate of rats increases evidently, over50%higher than the sham scald group,while the protein incorporation rate shows no difference,. The glutamic acid taken per osis transformed to the glutamine in the plasma, though the percentage is small and theefficacy is low. The intravenous infused glutamine is partly transformed into glutamicacid directly and serve as direct nutrition in the form of glutamic acid.