【摘要】：一.研究目的本研究通過觀察200mmol/L、300mmol/L與400mmol/L等不同濃度的高滲鈉鹽液(HS)和乳酸鈉林格液(LR)對嚴重燙傷大鼠液體復(fù)蘇的效果,并從炎癥因子、氧化應(yīng)激反應(yīng)和信號轉(zhuǎn)導(dǎo)通道等方面探討不同濃度的高滲鈉鹽復(fù)蘇液對嚴重燙傷大鼠腸損傷影響的初步機制。二.研究方法健康成年SD大鼠104只(購自安徽醫(yī)科大學(xué)實驗動物中心),雌雄不限,體重在200-250g。大鼠適應(yīng)性飼養(yǎng)一周后隨機分成五組:正常對照組(8只)、乳酸鈉林格液組(LR組,24只)、200mmol/L高滲鈉鹽液組(HS200組,24只)、300mmol/L高滲鈉鹽液組(HS300組,24只)、400mmol/L高滲鈉鹽液組(HS400組,24只)。所有大鼠均尾靜脈置管,剃除背部毛發(fā),對照組的大鼠不予燙傷和補液,LR組與各HS組大鼠背部經(jīng)98℃水燙傷12s造成30%體表面積III度燒傷。燙傷后分別采用LR、200 mmol/L HS、300mmol/L HS與400mmol/L HS進行復(fù)蘇。LR組與各HS組的大鼠均在傷后2h、8h和24h處死取血和腸組織。第一部分:不同濃度的高滲鈉鹽復(fù)蘇液對嚴重燙傷大鼠血鈉、腸含水量以及炎性細胞因子表達的影響將抽取的血標本用生化分析儀測量血清鈉濃度;分別稱取對照組、LR組、HS200組、HS300組與HS400組的大鼠的腸濕重(W)與干重(D),并計算腸含水量(以“濕重/干重”表示);使用酶聯(lián)免疫吸附法(ELISA)測量血漿中腫瘤壞死因子α(TNF-α)、白介素1β(IL-1β)和高遷移率蛋白B1(HMGB1)含量。將取得數(shù)據(jù)進行統(tǒng)計分析。第二部分:不同濃度的高滲鈉鹽復(fù)蘇液對嚴重燙傷大鼠腸道氧化應(yīng)激反應(yīng)以及腸血管內(nèi)皮細胞的影響取對照組、LR組、HS200組、HS300組與HS400組大鼠的一部分腸組織進行丙二醛(MDA)含量、超氧化物歧化酶(SOD)和二胺氧化酶(DAO)活性等的測定。另一部分腸組織在使用10%甲醛固定后使用免疫組化法測定血管性血友病因子(v WF)的表達。第三部分:不同濃度的高滲鈉鹽復(fù)蘇液對嚴重燙傷大鼠腸組織p38MAPK以及JNK通道的影響采用蛋白質(zhì)印跡法(Western Blot)測定五組大鼠腸組織中p38MAPK以及JNK通道的活性并進行統(tǒng)計學(xué)分析。三.結(jié)果第一部分LR組血鈉濃度各時間點明顯低于HS200組、HS300組與HS400組和對照組,差異具有統(tǒng)計學(xué)意義(P0.01)。與對照組血鈉濃度相比,HS200組在傷后2h、24h增高(P0.05),在傷后8h無明顯差異(P0.05),HS300組和HS400組在各時間點均明顯增高,差異具有統(tǒng)計學(xué)意義(P0.01)。與HS200組相比,HS300組血鈉濃度各時間點無明顯差異(P0.05),HS400組血鈉濃度各時間點均明顯增高,差異具有統(tǒng)計學(xué)意義(P0.01)。與HS300組相比,HS400組僅在傷后24h顯著增高(P0.05),其他時間點無統(tǒng)計學(xué)差異。LR組和HS200組在各時間點的腸濕干重比(W/D)均高于對照組,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01),HS300組的傷后8h、24h以及HS400組的傷后8h的W/D明顯高于對照組(P0.05或P0.01),其他時間點無明顯統(tǒng)計學(xué)差異。與LR組的腸W/D相比,HS200組和HS300組在傷后2h、8h明顯降低(P0.05或P0.01),傷后24h無統(tǒng)計學(xué)差異,HS400組在各時間點均明顯降低,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。HS200組、HS300組和HS400組之間的腸W/D無明顯差異(P0.05)。與對照組相比,LR組的各時間點的血TNF-α和IL-1β含量均顯著增高,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01);HS200組和HS300組在傷后2h、8h以及HS400組的傷后8h的血TNF-α含量均顯著增高(P0.05或P0.01);三個HS組的各時間點的IL-1β含量均明顯增高,差異具有統(tǒng)計學(xué)意義(P0.01);LR組、HS200組和HS300組在傷后8h、24h以及HS400組的傷后24h的血HMGB1含量均顯著增高,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01),它們在傷后2h的血HMGB1含量均無統(tǒng)計學(xué)差異。與LR組相比,HS200組、HS300組與HS400組在傷后2h、8h的血TNF-α含量均明顯降低,差異具有統(tǒng)計學(xué)意義(P0.01);它們在各時間點的血IL-1β含量均顯著減低(P0.01),它們在傷后8h和24h血中HMGB1含量均明顯降低,差異具有統(tǒng)計學(xué)意義(P0.01);它們在傷后24h的血TNF-α含量以及在傷后2h的血HMGB1含量均無明顯差異(P0.05)。除HS400組在傷后24h的血HMGB1的含量明顯高于HS200組(P0.05)外,三個HS組在各時間點的血TNF-α、IL-1β、血HMGB1的含量均無明顯差異(P0.05)。第二部分LR組、HS200組、HS300組與HS400組在各時間點的腸組織MDA含量均高于對照組,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。與LR組的腸組織MDA含量相比,HS300組與HS400組在各時間點均顯著降低,HS200組在傷后2h、8h明顯減少,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。與HS200組相比,HS300組在各時間點的腸組織MDA含量無明顯差異(P0.05),HS400組在各時間點的腸組織MDA含量均明顯降低(P0.05或P0.01);與HS300組相比,HS400組在各時間點的腸組織MDA含量無統(tǒng)計學(xué)差異(P0.05)。與對照組的腸組織SOD活性相比,LR組各時間點、HS200組和HS300組在傷后8h均明顯增大,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。HS300組和HS400組在傷后8h的腸組織SOD活性顯著低于LR組,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。HS200組、HS300組和HS400組之間在各時間點的腸組織SOD活性無明顯差異(P0.05)。LR組、HS200組、HS300組與HS400組的腸組織DAO活性均低于對照組,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。與LR組的腸組織DAO活性相比,HS200組在各時間點無統(tǒng)計學(xué)差異,HS300組在傷后2h、24h以及HS400組在各時間點均顯著增高,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。除HS400組和HS300組在傷后2h的DAO活性比HS200組增高(P0.05或P0.01)外,三個HS組之間在傷后其他時間點的DAO活性均無明顯差異(P0.05)。腸組織v WF免疫組化結(jié)果顯示:與LR組比較,HS200組、HS300組與HS400組在各時間點的腸組織v WF表達有不同程度的增強,且隨著HS濃度的增加而更明顯。第三部分與對照組相比,LR組和HS200組腸組織p38MAPK的活性在各時間點均明顯增強(P0.05或P0.01),HS300組在傷后8h、24h增強,HS400組在傷后8h增強,差異具有統(tǒng)計學(xué)意義(P0.01)。與LR組相比,HS200組、HS300組與HS400組腸組織p38MAPK的活性在各時間點均明顯降低,差異具有統(tǒng)計學(xué)意義(P0.01)。HS400組在各時間點的腸組織p38MAPK活性均顯著低于HS200組,在傷后2h、8h明顯低于HS300組,差異具有統(tǒng)計學(xué)意義(P0.01)。HS300組在傷后8h、24h的腸組織p38MAPK活性均明顯低于HS200組,差異具有統(tǒng)計學(xué)意義(P0.01)。與對照組相比,LR組在傷后各時間點、HS200組在傷后8h、24h以及HS300組和HS400組的腸組織JNK活性在傷后8h均顯著增強,差異具有統(tǒng)計學(xué)意義(P0.01)。采用三組HS復(fù)蘇,在各時間點的腸組織JNK活性均明顯低于LR組,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。與HS200組的腸組織JNK活性比較,HS300組在HS傷后24h明顯降低,HS400組在傷后8h、24h明顯下降,差異具有統(tǒng)計學(xué)意義(P0.01);與HS300組比較,HS400組在傷后2h、8h的腸組織JNK活性明顯降低,差異具有統(tǒng)計學(xué)意義(P0.05或P0.01)。四.結(jié)論1.三組高滲鈉鹽復(fù)蘇液與LR比較,均能防止低鈉血癥的發(fā)生,減輕嚴重燙傷大鼠傷后的腸道水腫并能抑制相關(guān)炎性因子的表達,三組作用上無明顯差異。但是,使用400mmol/L的HS進行液體復(fù)蘇,其血鈉濃度明顯增高。2.三組高滲鈉鹽復(fù)蘇液與LR比較,均能減輕嚴重燙傷大鼠傷后腸道的氧化應(yīng)激反應(yīng),可以有效保護腸血管內(nèi)皮細胞,其中使用400mmol/L的HS復(fù)蘇的效果最好。3.三組高滲鈉鹽復(fù)蘇液與LR比較,均能降低嚴重燙傷大鼠傷后腸組織中p38MAPK和JNK的活性表達,其中使用400mmol/L的HS復(fù)蘇的效果最好。
[Abstract]:The purpose of this study is to investigate the effect of high osmotic sodium salt solution (HS) and sodium lactate (LR) on the fluid resuscitation of severely scalded rats by observing the different concentrations of 200mmol/L, 300mmol/L and 400mmol/L, and to discuss the severe scald with different concentrations of hypertonic sodium salt resuscitation from the inflammatory factors, oxidative stress reactions and signal transduction pathways. The preliminary mechanism of the effect of intestinal injury in rats. Two. 104 healthy adult SD rats (purchased from the experimental animal center of Medical University Of Anhui) were randomly divided into five groups: normal control group (8 rats), sodium lactate solution group (group LR, 24), and 200mmol/L hypertonic sodium salt solution group (group HS200, 2). 4, 300mmol/L hypertonic sodium salt group (group HS300, 24), 400mmol/L hypertonic sodium salt solution group (group HS400, 24). All rats were treated with tail vein, shaving the back hair, the rats in the control group were not scalded and rehydration, and the back of the LR group and each HS group were scalded at 98 degrees centigrade to 30% body surface area III degree burn. LR, 200 mmol/ were used after the scald, respectively. L HS, 300mmol/L HS and 400mmol/L HS were used to resuscitate the.LR group and the rats of each HS group. The blood and intestinal tissue were killed in 2H, 8h and 24h after injury. Part 1: the effects of different concentrations of hypertonic sodium salt resuscitation on blood sodium, intestinal water content and expression of inflammatory cytokines in severely scalded rats were measured by biochemical analyzer. The concentration of sodium was called the intestinal wet weight (W) and dry weight (D) of the rats in the control group, the LR group, the HS200 group, the HS300 group and the HS400 group, and the calculation of the intestinal water content ("wet weight / dry weight"), and the measurement of the tumor necrosis factor alpha (TNF- a) in plasma with enzyme linked immunosorbent assay (ELISA), the content of interleukin 1 beta (IL-1 beta) and high mobility protein B1 (HMGB1). Statistical analysis of the data. The second part: the effect of different concentrations of hypertonic sodium salt resuscitation on the intestinal oxidative stress reaction and intestinal vascular endothelial cells in severely scalded rats, group LR, HS200, group HS300 and part of the rats of group HS400, the content of malondialdehyde (MDA), superoxide dismutase (SOD) and the oxidation of amines The determination of enzyme (DAO) activity. The other part of the intestinal tissue was immobilized with 10% formaldehyde to determine the expression of vascular hemophilia factor (V WF). The third part: the effect of different concentrations of hypertonic sodium salt resuscitation on the intestinal tissue p38MAPK and JNK channel in severely scalded rats was determined by western blot (Western Blot) five The activity of p38MAPK and JNK channel in the intestinal tissue of group rats was statistically analyzed. Three. Results the blood sodium concentration in group LR was significantly lower than that in group HS200. The difference between group HS300 and HS400 group and control group was statistically significant (P0.01). Compared with the concentration of sodium in the control group, HS200 group was 2h, 24h increased (P0.05) after injury, and 8h was no longer after injury. Significant difference (P0.05), group HS300 and HS400 groups were significantly higher at all time points, the difference was statistically significant (P0.01). Compared with the HS200 group, there was no significant difference in the time points of the concentration of sodium in the HS300 group (P0.05), and the concentration of sodium in the HS400 group increased significantly at each time point, and the difference was statistically significant (P0.01). Compared with the HS300 group, the HS400 group was only after the injury. 24h was significantly higher (P0.05). There was no statistical difference at other time points in.LR group and HS200 group at all time points (W/D). The difference was statistically significant (P0.05 or P0.01). The 8h in 8h, 24h and HS400 group after injury of HS300 group was significantly higher than that of the control group. There was no significant statistics at other time points. Difference. Compared with W/D in group LR, 2h, 8h decreased significantly after injury in group HS200 and HS300 (P0.05 or P0.01), and there was no significant difference in 24h after injury. The HS400 group decreased significantly at all time points, and the difference was statistically significant (P0.05 or P0.01), and there was no significant difference between the group and the group. The content of TNF- and IL-1 beta in the blood of the time point increased significantly, and the difference was statistically significant (P0.05 or P0.01), and the content of TNF- alpha in the 8h of 2h, 8h and HS400 groups after injury in HS200 and HS300 groups increased significantly (P0.05 or P0.01); the difference was statistically significant in each time point of the three groups. The serum HMGB1 content of 24h in 8h, 24h and HS400 groups after injury in group HS200 and group HS300 increased significantly (P0.05 or P0.01), and there was no statistical difference in the HMGB1 content of blood in 2h after injury. The content of blood IL-1 beta in all time points decreased significantly (P0.01), and their HMGB1 content in 8h and 24h blood decreased significantly after injury (P0.01), and there was no significant difference between the serum TNF- alpha content of 24h and the HMGB1 content of 2h in 2h after injury (P0.05). The content of MGB1 was significantly higher than that of group HS200 (P0.05). There was no significant difference in the content of TNF- alpha, IL-1 beta, and blood HMGB1 in the three HS groups at all time points (P0.05). The contents of the intestinal tissue in the second part of the LR group, HS200 group, HS300 group and HS400 group were higher than those of the control group. The content of A in group HS300 and group HS400 decreased significantly at all time points, and in group HS200, 2h, 8h decreased significantly after injury. The difference was statistically significant (P0.05 or P0.01). Compared with the HS200 group, there was no significant difference in the MDA content of intestinal tissue in the HS300 group at all time points (P0.05). 1): compared with the HS300 group, there was no significant difference in intestinal tissue MDA content in group HS400 at all time points (P0.05). Compared with the SOD activity of the intestinal tissue in the control group, the 8h of HS200 group and HS300 group increased significantly at each time point in group LR and HS300 group, and the difference was statistically significant (P0.05 or P0.01) and the activity of intestinal tissue was significantly lower in the.HS300 group and in the group after injury. In group LR, the difference was statistically significant (P0.05 or P0.01) group.HS200, and there was no significant difference in intestinal tissue SOD activity between HS300 and HS400 groups at all time points (P0.05).LR group, HS200 group, HS300 group and HS400 group were lower than the control group, and the difference has statistical significance. There was no statistical difference at all time points in group S200, and in group HS300, 2h, 24h and HS400 were significantly higher in all time points after injury. The difference was statistically significant (P0.05 or P0.01). Except for HS400 and HS300 group, DAO activity of 2H was higher than that of HS200 group. There was no obvious activity between the three groups at other time points after injury. Difference (P0.05). The immunohistochemical results of intestinal tissue V WF showed that: compared with the LR group, the expression of V WF in the intestinal tissue of the HS200 group, the HS300 group and the HS400 group increased in varying degrees, and was more obvious with the increase of HS concentration. The third part, compared with the control group, increased significantly at all time points in the LR group and the HS200 group. P0.05 or P0.01), group HS300 was 8h, 24h increased after injury, and 8h increased in group HS400 after injury. The difference was statistically significant (P0.01). Compared with group LR, the activity of HS200 group, HS300 group and HS400 group decreased significantly at all time points. Lower than HS200 group, 2h, 8h significantly lower than group HS300 after injury, the difference was statistically significant (P0.01).HS300 group 8h, 24h in the intestinal p38MAPK activity was significantly lower than the HS200 group, the difference was statistically significant (P0.01). The activity of NK increased significantly after injury (8h). The difference was statistically significant (P0.01). The activity of JNK in the intestinal tissue at all time points was significantly lower than that in the LR group with three groups of HS resuscitation. The difference was statistically significant (P0.05 or P0.01). The HS300 group decreased obviously after the HS injury. The difference was statistically significant (P0.01). Compared with the HS300 group, the activity of JNK in the intestinal tissue of group HS400 after injury was significantly reduced, and the difference was statistically significant (P0.05 or P0.01). Four. Conclusion 1. the three groups of hypertonic sodium salt resuscitation could prevent the occurrence of hyponatremia and reduce the intestinal edema after severe scald injury and inhibit the intestinal edema in severely scalded rats. There was no significant difference in the expression of related inflammatory factors in the three groups. However, 400mmol/L HS was used to resuscitate the liquid, and the serum sodium concentration was significantly higher than that of the.2. three group of hypertonic sodium salt resuscitation, which could reduce the oxidative stress reaction in the intestinal tract after severe scald injury in rats, and could effectively protect the intestinal vascular endothelial cells, which used 400mmol/L. The best effect of HS resuscitation is.3. three hypertonic sodium salt resuscitation and LR, which can reduce the expression of p38MAPK and JNK in the intestinal tissue of severely scalded rats, and HS resuscitation with 400mmol/L is the best.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2014
【分類號】：R644

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