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Lymphatic reactivity has been shown to exhibit a biphasic change following hemorrhagic shock, and nitric oxide (NO) is involved in this process. However, the precise mechanism responsible for NO regulation of the lymphatic reactivity along with the progression of hemorrhagic shock is unclear. Therefore, the present study was to investigate how NO participates in regulating the shock-induced biphasic changes in lymphatic reactivity and its underlying mechanisms. First, the expressions or contents of inducible NO synthase, nitrite plus nitrate, and elements of cAMP-PKA-KATP and cGMP-PKG-KATP pathway in thoracic ducts tissue were assessed. The results revealed that levels of nitrite plus nitrate, cAMP, cyclic guanosine monophosphate (cGMP), p-PKA, and p-PKG were increased gradually along with the process of shock. Second, the roles of cAMP-PKA-KATP and cGMP-PKG-KATP in NO regulating lymphatic response to gradient substance P were evaluated with an isolated lymphatic perfusion system. The results showed that the NOS substrate (L-Arg), PKA donor (8-Br-cAMP) decreased the reactivity of shock 0.5 h-lymphatics, and that the PKA inhibitor (H-89) and KATP inhibitor (glibenclamide) restrained the effects of L-Arg while glibenclamide abolished the effects of 8-Br-cAMP. Meanwhile, NOS antagonist (L-NAME), protein kinase G (PKG) inhibitor (KT-5823), and soluble guanylate cyclase inhibitor (ODQ) increased the reactivity of shock 2 h-lymphatics, whereas KATP opener (pinacidil) inhibited these elevated effects induced by either L-NAME, ODQ, or KT-5823. Taken together, these results indicate that NO regulation of lymphatic reactivity during shock involves both cAMP-PKA-KATP and cGMP-PKG-KATP pathways. These findings have potential significance for the treatment of hemorrhagic shock through regulating lymphatic reactivity.

Background: Excessively inflammatory response is one of mechanisms that underlie the acute kidney injury (AKI) induced by severe hemorrhagic shock, which could be ameliorated by post-hemorrhagic shock mesenteric lymph (PHSML) blockage. Recent studies demonstrate that high mobility group box 1 (HMGB1) and the receptor for advanced glycation end products (RAGE) are critical mediators of local inflammations. The present study was sought to investigate whether the PHSML drainage inhibits the HMGB1 and RAGE in mouse kidney to ameliorate the renal inflammatory responses., Methods: A mouse hemorrhagic shock model (40 ± 2 mmHg for 90 min, fluid resuscitation for 30 min) was employed, and the PHMSL drainage was performed at the end of the resuscitation. After 3 h of resuscitation, the expressions of mRNA and protein for the renal HMGB1 and RAGE and the levels of interleukin (IL)-1β and IL-18 were assessed by the real-time reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay, respectively., Results: Hemorrhagic shock elicited significant increases in the mRNA expressions of HMGB1 and RAGE and in the protein expressions of HMGB1, RAGE, IL-1β and IL-18 in kidney. The PHSML drainage abolished these potentiating effects., Conclusion: The present study demonstrates that PHSML blockade reduces the increased HMGB1 and RAGE and pro-inflammatory factors following hemorrhagic shock, suggesting that the PHSML elicits the inflammatory responses via enhancing the HMGB1 and RAGE production in the kidney.

Purpose: To evaluate the changes of contractility and reactivity in isolated lymphatics from hemorrhagic shock rats with resuscitation., Methods: Six rats in the shock group suffered hypotension for 90 min by hemorrhage, and resuscitation with shed blood and equal ringer's solution. Then, the contractility of lymphatics, obtained from thoracic ducts in rats of the shock and sham groups, were evaluated with an isolated lymphatic perfusion system using the indices of contractile frequency (CF), tonic index (TI), contractile amplitude (CA) and fractional pump flow (FPF). The lymphatic reactivity to substance P (SP) was evaluated with the different volume of CF, CA, TI and FPF between pre- and post-treatment of SP at different concentrations., Results: The CF, FPF, and TI of lymphatics obtained from the shocked rats were significantly decreased than that of the sham group. After SP stimulation, the ∆CF (1 × 10(-8), 3 × 10(-8), 1 × 10(-7), 3 × 10(-7) mol/L), ∆FPF (1 × 10(-8), 3 × 10(-8), 1 × 10(-7) mol/L), and ∆TI (1 × 10(-8) mol/L) of lymphatics in the shock group were also obviously lower compared with the sham group. In addition, there were no statistical differences in CA and ∆CA between two groups., Conclusion: Lymphatic contractility and reactivity to substance P appears reduction following hemorrhagic shock with resuscitation.

Purpose: To determine the role of mesenteric lymph reperfusion (MLR) on endotoxin translocation in brain to discuss the mechanism of brain injury subjected to superior mesenteric artery occlusion (SMAO) shock., Methods: Twenty-four rats were randomly assigned to MLR, SMAO, MLR+SMAO and sham groups. MLR was performed by clamping the mesenteric lymph duct (MLD) for 1 h and then allowing reperfusion for 2 h in the MLR group; SMAO involved clamping the superior mesenteric artery (SMA) for 1 h, followed by reperfusion for 2 h in the SMAO group; occlusion of both the SMA and MLD for 1 h was followed by reperfusion for 2 h in the MLR+SMAO group rats., Results: SMAO shock induced severe increased levels of the endotoxin, lipopolysaccharide receptor, lipopolysaccharide-binding protein, intercellular adhesion molecule-1 and tumor necrosis factor-α. Concurrently, MLR after SMAO shock further aggravates these deleterious effects., Conclusion: Mesenteric lymph reperfusion exacerbated the endotoxin translocation in brain; thereby increased inflammatory response occurred, suggesting that the intestinal lymph pathway plays an important role in the brain injury after superior mesenteric artery occlusion shock.

Background: Kidney injury commonly occurs following hemorrhagic shock. This study aims to observe the effects of mesenteric lymph duct ligation (MLDL) on the adenosine triphosphate (ATP) levels and the cell membrane adenosine triphosphatase (ATPase) activity in the kidneys of rats subjected to hemorrhagic shock., Methods: Wistar rats were assigned into sham, shock, and ligation groups. The hemorrhagic shock model was established in the shock and ligation groups, and MLDL was performed in the ligation group after resuscitation. Renal homogenates were prepared to determine the ATP and ATPase levels at 90 min after hemorrhage and at 0, 1, 3, 6, 12, and 24 h after resuscitation., Results: The ATP levels, and the Na(+)-K(+)-ATPase, Mg(2+)-ATPase, Ca(2+)-ATPase, and Ca(2+)-Mg(2+)-ATPase activities in the renal tissue of the shock group were lower than those in the sham group at the multiple time points. Furthermore, the corresponding values in the ligation group were significantly higher than those in the shock group at multiple time points., Conclusion: MLDL improves energy metabolism and enhances the ATPase activity in the kidneys of hemorrhagic shock rats, along with other mechanisms that alleviate renal injury after hemorrhagic shock.

Background: Acute kidney injury (AKI), a common pathological process following hemorrhagic shock, can lead to an internal milieu disorder, which is an important factor of multiple organ failure (MOF). It has been shown that the mesenteric lymph return plays a deleterious effect on MOF induced by hemorrhagic shock. In this study, we investigated the effects of mesenteric lymph duct ligation (MLDL) on gene expression profiles of renal tissue following hemorrhagic shock with fluid resuscitation., Methods: After establishment of hemorrhagic shock model and fluid resuscitation in rats of shock and shock ligation groups, the MLDL was performed in shock ligation group, and only threading under the mesenteric lymph duct in the shock group. Then, the fixed position renal tissue was taken out for homogenate in two groups at 3 h after resuscitation, the total mRNA was extracted, reversely transcribed into cDNAs and marked with Cy3 and Cy5. The cDNAs were subjected for microarray scanning with 12,028 cDNA probes; the differentially expressed genes between two groups were analyzed., Results: In the 5812 valid dates of rat genomes transcription, there were 34 known differentially expressed genes between the two groups, of which 11 genes were up-regulated whereas 23 genes were down-regulated by MLDL. These different expressed genes encoding protein function were mainly involved in signal transduction, transcription regulation, metabolism, transport, cell growth, cell cycle, cell adhesion, cell movement, cellular component, and biological process., Conclusions: The mechanism of MLDL alleviating the AKI aftershock might be associated with up- or down-regulation of the above gene expressions.

This study aimed to investigate the effect of mesenteric lymph drainage on the acute kidney injury induced by hemorrhagic shock without resuscitation. Eighteen male Wistar rats were randomly divided into sham, shock, and drainage groups. The hemorrhagic shock model (40 mmHg, 3 h) was established in shock and drainage groups; mesenteric lymph drainage was performed from 1 h to 3 h of hypotension in the drainage group. The results showed that renal tissue damage occurred; the levels of urea, creatinine, and trypsin in the plasma as well as intercellular adhesion molecule-1 (ICAM-1), receptor of advanced glycation end-products (RAGE), tumor necrosis factor-α (TNF-α), malondialdehyde (MDA), lactic acid (LA), and 2,3-DPG in the renal tissue were increased in the shock group after 3 h of hypotension. Mesenteric lymph drainage lessened the following: renal tissue damage; urea and trypsin concentrations in the plasma; ICAM-1, RAGE, TNF-α, MDA, and LA levels in the renal tissue. By contrast, mesenteric lymph drainage increased the 2,3-DPG level in the renal tissue. These findings indicated that mesenteric lymph drainage could relieve kidney injury caused by sustained hypotension, and its mechanisms involve the decrease in trypsin activity, suppression of inflammation, alleviation of free radical injury, and improvement of energy metabolism.

The aim of this present study was to examine changes in RhoA protein levels and the role in RhoA in lymphatic contractility and reactivity after hemorrhagic shock. Levels of RhoA and phospho-RhoA in lymphatic tissue isolated from hemorrhagic shock rats were measured, and the contractility and reactivity to substance P of lymphatics isolated from control rats and rats subjected to shock 0.5 and 2 h were determined with an isolated lymphatic perfusion system at a transmural pressure of 3 cmH2O. At the same time, lymphatics isolated from rats subjected to shock 0.5 and 2 h were incubated with agonists and antagonists of RhoA/Rho kinase signaling. Contractile frequency, end-diastolic and end-systolic diameter, and passive diameter were recorded and used to calculate lymphatic tonic index, contractile amplitude, and fractional pump flow. After stimulation with a gradient of substance P, the differences between the preadministration and postadministration values of contractile frequency, contractile amplitude, tonic index, and fractional pump flow were calculated to further assess lymphatic reactivity. RhoA protein levels were significantly increased at 0.5 h after shock but decreased at 2 and 3 h after shock; p-Rho levels were initially increased after shock and subsequently decreased. The contractility and reactivity of 0.5-h-shocked lymphatics were significantly reduced by the RhoA antagonist C3 transferase and the Rho kinase antagonist Y-27632. The RhoA agonist U-46619 increased the contractility and reactivity of 2-h-shocked lymphatics, whereas Y-27632 suppressed the effect of U-46619. Okadaic acid, an inhibitor of myosin light-chain phosphatase, had no effect on the contractility of 2-h-shocked lymphatics, but improved lymphatic reactivity. These results suggest that RhoA is involved in the modulation of lymphatic pump function during hemorrhagic shock and that its effects may be mediated by Rho kinase and MLCP.

Vascular hyporeactivity is an important factor in irreversible shock, whereas calcium desensitization is one of the mechanisms of vascular hyporeactivity, and the intestinal lymphatic pathway plays an important role in multiple organ injury after severe hemorrhagic shock (HS). In this study, our aims were to determine the effects of mesenteric lymph on vascular reactivity during HS and the mechanisms involved. First, the in vivo pressor response was observed by intravenous injection of norepinephrine (3 μg/kg) at different time points after HS. We found that mesenteric lymph duct ligation (MLDL) and mesenteric lymph drainage (MLD) enhanced the pressor response at multiple time points after shock. Next, vascular reactivity and calcium sensitivity in superior mesenteric artery (SMA) vascular rings were examined using an isolated organ perfusion system. Vascular reactivity and calcium sensitivity were higher for SMA rings from rats that had undergone HS plus MLDL or MLD that those from rats that had undergone only HS. The effects of MLDL and MLD on vascular reactivity and calcium sensitivity were significantly increased following incubation with the calcium sensitizer angiotensin II and were reduced after incubation with the calcium sensitivity inhibitor insulin. When SMA rings from normal rats were incubated with mesenteric lymph from rats subjected to HS, lymph obtained 0 to 0.5 h after shock enhanced vascular reactivity and calcium sensitivity, whereas lymph obtained 1 to 3 h after shock blunted these effects. We finally examined vascular reactivity and calcium sensitivity in HS rats subjected to MLD at 0 to 3 h or 1 to 3 h after shock. We found that contractile activity of SMAs in response to norepinephrine or Ca was higher in HS rats subjected to MLD at 1 to 3 h after shock compared with rats subjected to MLD at 0 to 3 h after shock. These results indicate that mesenteric lymph return plays an important role in biphasic changes in vascular reactivity during HS. Even more importantly, mesenteric lymph 1 h after shock was an important contributor to vascular hyporeactivity, and its mechanism of action was related to calcium desensitization. Targeting lymph may therefore have therapeutic potential in the treatment of severe shock-induced hypotension.

Objective: To investigate the mechanism of ATP-sensitive potassium channel (K(ATP)) on nitride oxide (NO) regulating contractile activity of isolated lymphatics from hemorrhagic shock (HS) rats., Methods: Eighty - four Wistar rats were randomly divided into control group (n=6), HS 0.5-hour group (n=36), HS 2-hour group (n=42). A segment of thoracic duct of rats was adopted for isolated lymphatics after HS, then the HS 0.5-hour and 2-hour lymphatics were incubated combined or respectively with K(ATP) inhibitor glibenclamide (Gli), opener of K(ATP) pinacidil (Pin), NO donor L-arginine (L-Arg), protein kinase A (PKA) inhibitor H-89, PKA donor 8-bromine-cyclic adenosine monophosphate (8-Br-cAMP), nitricoxide synthase (NOS) inhibitor N-nitro-L-arginine methyl ester (L-NAME), soluble guanylyl cyclase (sGC) inhibitor 1 h-[1,2,4]-oxadiazole-[4,3-a]- quinoxalin-1-one (ODQ), protein kinase G (PKG) inhibitor KT-5823 (named as HS 0.5 h, HS 0.5 h + L-Arg, HS 0.5 h + L-Arg + H-89, HS 0.5 h + L-Arg + Gli, HS 0.5 h + 8-Br-cAMP, HS 0.5 h + 8-Br-cAMP + Gli and HS 2 h, HS 2 h + L-NAME, HS 2 h + L-NAME + Pin, HS 2 h + ODQ, HS 2 h + ODQ + Pin, HS 2 h + KT-5823, HS 2 h + KT-5823 + Pin, n=6). By lymphatic perfusion in vitro, contractile frequency (CF) was recorded, and contractile amplitude (CA), tonic index (TI) and fractional pump flow (FPF) were calculated., Results: The results suggested that the CF, TI, FPF of HS 0.5-hour lymphatics were significantly increased compared with control group, and the CF, TI, FPF decreased significantly when incubated with L-Arg. H-89 could deteriorate the decreased effect of L-Arg on CF and FPF, and Gli could deteriorate the decreased effect of L-Arg on FPF. When the HS 0.5-hour lymphatics incubated with 8-Br-cAMP, the CF and FPF were all decreased significantly, and when the HS 0.5-hour lymphatics incubated with 8-Br-cAMP and Gli, the CF was significantly higher than HS 0.5 h + 8-Br-cAMP group, and the TI and FPF decreased significantly compared with HS 0.5-hour group. The CF, FPF, TI of HS 2-hour lymphatics were significantly decreased compared with control group. L-NAME could increase the CF, TI, FPF; ODQ could increase the CF, TI; KT-5823 could increase the CF and FPF; when incubated with Pin respectively, the CF and FPF when incubated with L-NAME were decreased, the CF, TI and FPF when incubated with ODQ were decreased, in addition, the CA was recovered as level of control group, the CF and FPF when incubated with KT-5823 were decreased., Conclusions: K(ATP) involved in NO modulating pumping function of isolated lymphatics of HS rats, and the effect may be relative to the signal pathway of NO-cAMP-PKA and NO-cGMP-PKG.

The aim of the present study was to investigate whether protein kinase C (PKC) was involved in the effect of mesenteric lymph duct ligation or mesenteric lymph drainage on vascular calcium sensitivity in hemorrhagic shock rats. Male Wistar rats were randomly divided into Sham, Shock (hemorrhagic shock), Shock+Ligation (mesenteric lymph duct ligation plus shock) and Shock+Drainage (mesenteric lymph drainage plus shock) groups. After being in shock (hypotension 40 mmHg) for 3 h, the tissue of superior mesenteric artery (SMA) was taken out for detecting the PKC expression and phospho-PKC (p-PKC) activity, and the vascular rings of SMA were prepared and used to measure the response to gradient calcium concentration for assaying the calcium sensitivity, the parameters of which including tension, maximum tension (E(max)) and negative logarithm of EC(50), called the pD(2). Other vascular rings from Shock+Ligation and Shock+Drainage groups were incubated with PKC regulator PMA or Staurosporine before the measurement of calcium sensitivity. The results showed that, PKC expression, p-PKC activity and calcium sensitivity of SMA in Shock group was significantly lower than that of Sham group, whereas the above-mentioned indexes were significantly elevated in Shock+Ligation and Shock+Drainage groups compared with those in Shock group. PKC agonist PMA enhanced the contractile activity of vascular rings to gradient calcium ions, and increased E(max) of SMA in Shock+Ligation and Shock+Drainage groups. On the contrary, PKC inhibitor Staurosporine significantly decreased the response to gradient calcium ions and E(max) of SMA in Shock+Ligation and Shock+Drainage groups. These results suggest that PKC plays a role in the improvement of vascular calcium sensitivity by blockade of mesenteric lymph return in hemorrhagic shock rats.

Objective: To investigate the role of mesenteric lymph reperfusion (MLR) aggravates multiple organs injury in superior mesenteric artery occlusion (SMAO) shock and its mechanism., Methods: Twenty four Wistar rats were randomly divided into 4 groups (n = 6): Sham group (only anesthetized and operated), MLR group rats performed 1 h occlusion of mesenteric lymph duct (MLD), then followed by 2 h of reperfusion, SMAO group (rats performed 1 h occlusion of superior mesenteric artery (SMA) and then followed by 2 h of reperfusion), SMAO + MLR group (rats performed 1 h occlusion of SMA and MLD and then followed by 2 h of reperfusion). The blood sample was taken out from abdominal aortic for plasma and the liver, kidney, myocardium, lung tissues in fixed position were prepared for making homogenate after reperfusion of 2 h respectively. And the levels of endotoxin (ET) in plasma and homogenates were determined with kinetic turbidimetric technique of tachypleus amebocyte lysate, the contents of lipopolysaccharide-binding protein (LBP), lipopolysaccharide receptor (CD14) and tumor necrosis factor-alpha (TNF-alpha) in homogenates were determined with enzyme-linked immunosorbent assay method., Results: The indices have no statistics difference between sham group and MLR group. The ET levels of the plasma and hepatic, renal, myocardial, pulmonary homogenates in SMAO and SMAO + MLR groups were significant higher than that of sham and MLR groups, and these indices in SMAO + MLR were increased significantly than those in SMAO group. The CD14, LBP and TNF-alpha contents of the hepatic, renal, myocardial and pulmonary homogenates in SMAO and SMAO + MLR groups were significant higher than those in sham and MLR groups, and these indices in SMAO+ MLR were higher than SMAO group significantly., Conclusion: The mechanism of MLR aggravates multiple organs injury in SMAO shock may be associated with enterogenous ET through intestinal lymphatic pathway to translocate, activate the LBP/CD14 as endotoxin sensitizing system and promote inflammatory response.

Objective: To observe the change of lymphatic reactivity to substance P (SP) during the process of hemorrhagic shock (HS) with a technique of lymphatic perfusion in vitro in this study., Methods: Male Wistar rats were randomly divided into control group (surgical procedure only) and HS group (the rats in this group were further divided into five subgroups: shock 0 h, 0.5 h, 1 h, 2 h and 3 h groups after duplicating the HS model with method of bloodletting to mean arterial blood pressure was 40 mmHg through the femoral venous). Thoracic ducts were separated from HS rats at the corresponding time points in each group. A segment of thoracic duct was pressed and perfused in vitro at transmural pressure of 3 cm H2O, and then stimulated with gradient SP respectively. The end systolic diameter, end diastolic diameter, contraction frequency (CF) and passive diameter of isolated lymphatics were measured, while the contraction amplitude (CA), tonic index (TI) and fractional pump flow (FPF) were calculated, and the different values between pre- and post- administration of SP of CF, CA, TI and FPF were calculated and expressed as Delta CF, Delta TI, Delta CA and Delta FPF to further assess the reactivity of lymphatics., Results: After SP incubation, the Delta CF, Delta TI, Delta CA and Delta FPF of 0 h- and 0.5 h shocked lymphatics were significantly increased when compared with that of control group on one or several concentrations. The Delta CF (at 3 x 10(-7) mol/L of SP) and Delta TI (1 x 10(-7) mol/L) of 2 h- shocked lymphatics and the Delta CF (1 x 10(-7) mol/L, 3 x 10(-7) mol/L), Delta TI (1 x 10(-7) mol/L) and Delta CA (1 x 10(-7) mol/L) of 3 h- shocked lymphatics were all significantly reduced when compared with control group., Conclusion: The reactivity of lymphatics to SP presented a biphasic change during the process of HS: increase in early phase and decline in later stage.

Objective: To observe the effects of mesenteric lymph drainage and transfusion on pump activity and oxygen free radical of erythrocyte membrane of rats following hemorrhagic shock or normal, respectively., Methods: Twenty-four male Wistar rats were randomly divided into sham-shock group, shock group, shock mesenteric lymph drainage group (drainage group) and shock mesenteric lymph transfusion group (transfusion group) by randomization, with 6 rats in each group. Model of hemorrhagic shock was reproduced both in shock group and drainage group after anesthesia and operation, mesenteric lymph was drained 1 hour after shock in drainage group. Sham-shock group only received anesthesia and laparotomy, and the obtained mesenteric lymph was transfused in transfusion group. Blood was obtained from abdominal aorta, and red cell membrane suspension was prepared after 3 hours of shock or corresponding time, then the indices of erythrocyte membrane pump activity and oxygen free radical were determined., Results: Compared with sham-shock group, the activity of Na(+)-K(+)-ATPase (μmol×mg(-1)×h(-1)), Ca(2+)-ATPase (μmol×mg(-1) ×h(-1) ) and superoxide dismutase (SOD) (NU/mg)of red cell membrane were decreased markedly (Na(+)-K(+)-ATPase: 0.039±0.011 vs. 0.068±0.010; Ca(2+)-ATPase: 0.035±0.016 vs. 0.087±0.015; SOD: 0.785±0.289 vs. 1.202±0.328, P<0.05 or P<0.01), and the content of malondialdehyde (MDA, nmol/mg) was increased markedly in shock group (1.914±0.225 vs. 0.913±0.138, P<0.01). Compared with shock group, the activity of Na(+)-K(+)-ATPase (0.056±0.009), Ca(2+)-ATPase (0.079±0.025) and SOD (1.220±0.380) of red cell membrane were increased in drainage group, and the level of MDA (1.214±0.127) was decreased markedly (P<0.05 or P<0.01). Compared with sham-shock group, the activity of Na(+)-K(+)-ATPase (0.050±0.013), Ca(2+)-ATPase (0.056±0.023) of red cell membrane in transfusion group were significantly lower (both P<0.05), the content of MDA (1.456±0.270) was significantly higher (P<0.01), respectively, the activity of SOD (0.862±0.288) showed a lowering trend, but there was no statistical significance (P>0.05)., Conclusion: Mesenteric lymph return is a key factor in decreasing pump activity while aggravating free radical damage injury of erythrocyte membrane.

Objective: To observe the effects of mesenteric lymph duct (MLD) ligation on erythrocyte rheology in acute hemorrhagic rats., Methods: Twenty male Wistar rats were randomly divided into hemorrhage group and ligation group (n = 10). Blood (one fourth of body whole blood volume) was withdrawn through right common carotid arteries after rats were anesthetized. In ligation group, the MLD was ligated after hemorrhage, and only threading under the MLD in hemorrhage group. The survival situation at 24 h was recorded. After 24 h, survival rats were anesthetized again, blood sample was withdrawn through left common carotid artery rapidly. And the erythrocyte sedimentation rate (ESR), electrophoresis of erythrocytes, hematocrit (Hct) were determined in blood samples of before and after hemorrhage, the erythrocytes aggregation and deformability indices were calculated., Results: It showed that the ligation group survival (9 rats alive) was slightly better than that in hemorrhage group (6 rats alive). The results of erythrocyte rheology indices showed that the ESR, K value of equation, K value of emendation and electrophoresis time in hemorrhage group and ligation group were higher or longer than those before hemorrhage, the erythrocyte deformability was reduced significantly, respectively. And the erythrocytes aggregation index in hemorrhage group was increased, the electrophoresis length and migration of erythrocyte in hemorrhage group were lower than those before hemorrhage, respectively. But compared with hemorrhage group, the ESR, K value of equation, K value of emendation, erythrocytes aggregation index and electrophoresis time in ligation group were lower, the electrophoresis lenght, migration and deformability of erythrocyte were increased significantly., Conclusion: The results indicate that the higher erythrocyte aggregation ability, lower electrophoresis function and deformability are caused by acute hemorrhage in rats, and the MLD ligation can improve the abnormal erythrocyte rheology.

Background: The kidney is a common target in multiple organ dysfunction syndrome (MODS). The aim of this study is to determine the role of intestinal lymphatic pathway on renal injury in hemorrhagic shock rats., Methods: Wistar rats were divided into sham, shock, and ligation groups. The hemorrhagic shock model was induced in the shock and ligation groups. After resuscitation, the mesenteric lymph ducts were ligated in the ligation group. Blood from the carotid artery was taken to determine renal functional indices. The kidneys were used to observe histomorphological changes at 6 h after resuscitation. In addition, kidney homogenate was used to determine malondialdehyde (MDA), superoxide dismutase (SOD), tumor necrosis factor alpha (TNFalpha), interleukin-6 (IL-6), and myeloperoxidase (MPO) levels at 90 min after shock and 0, 1, 3, 6, 12, and 24 h after resuscitation. And the survival rate of 24 h was recorded., Results: The survival rate in shock group was obviously lower than sham and ligation groups. The urea and creatinine contents in the serum of shock and ligation groups were significantly higher than the sham group; the indices in the ligation group were significantly lower than the shock group. Histological studies showed various degrees of renal injury in the shock and ligation groups with a lesser severity in the ligation group. MDA, TNFalpha, IL-6, and MPO in renal homogenate of the shock group were raised, and the activity of SOD was lower in comparison to the sham group. Further, MDA, TNFalpha, IL-6, and MPO in renal homogenate of the ligation group at 6, 12, and 24 h were lower, and the SOD activity was higher than that of the shock group at the same time points., Conclusion: The mesenteric lymph duct ligation could be used to attenuate renal injury in shock rats. Its mechanism might be related to reducing the polymorph nuclear (PMN) and decreasing inflammatory mediator and free radical.

Objective: To explore the effect of mesenteric lymph duct ligation in protecting myocardial injury in rats after hemorrhage and lipopolysaccharide (LPS) challenge., Methods: Wistar rats were randomly divided into the sham group, non-ligation group and ligation group, and the "two-hit" injury model was reproduced by hemorrhage and LPS administration. Mesenteric lymph was blocked by ligating mesenteric lymph duct in ligation group after hemorrhage. After 24 hours of the assault, myocardial tissue was harvested and homogenized. The activity of myeloperoxidase (MPO) and ATPase and the contents of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) were determined in myocardial homogenate. At the same time, myocardium was pathologically studied. Apoptosis cell rate of myocardium was determined by method of terminal-deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL), the expression of bcl-2 and bax protein was determined by immunohistochemical method., Results: After hemorrhage and LPS challenge, the level of MPO [(0.23+/-0.08) U/g], TNF-alpha [(9.99+/-2.74) microg/g] and IL-6 [(31.57+/-12.71) microg/g] in myocardial homogenate of non-ligation group were significantly higher than those of sham group [MPO: (0.12+/-0.03) U/g, TNF-alpha: (4.17+/-1.35) microg/g, IL-6: (17.86+/-5.17) microg/g, respectively, all P<0.01], and the activity of ATPase was significantly lower. MPO [(0.13+/-0.03) U/g], TNF-alpha [(5.57+/-1.65) microg/g] and IL-6 [(23.24+/-5.95) microg/g] in myocardial homogenate of ligation group were significantly lower (P<0.05 or P<0.01), and the ATPase activity was significantly higher compared with non-ligation group. The apoptosis rate [(22.7+/-6.9)%] and expression of bax protein (104.5+/-11.4) of myocardial cells in non-ligation group were significantly higher compared with sham group [apoptosis rate: (3.8+/-1.2)%, bax protein: 142.1+/-10.9] and ligation group [apoptosis rate: (8.4+/-2.8)%, bax protein: 128.4+/-9.6], and expression of bcl-2 protein of non-ligation group (196.4+/-19.3) was significantly lower than that of sham group (132.2+/-12.3) and ligation group (165.1+/-11.6, all P<0.01)., Conclusion: The results demonstrate that the mesenteric lymph duct ligation ameliorate the myocardial injury in rats subjected to hemorrhage and LPS by reducing the inflammatory mediators of TNF-alpha and IL-6 levels, upregulating the bcl-2 protein expression and improving membrane ATPase activity of myocardium.

Objective: To observe the effects of mesenteric lymph duct ligation on platelet function, thrombus formation in vitro and coagulation function in rats with acute loss of blood, and investigate the role of intestinal lymphatic pathway on coagulability change during acute loss of blood., Methods: Twenty male Wistar rats were randomly divided into loss of blood group (n=10) and ligation group (n=10). The acute loss of blood model was reproduced by withdrawing blood (one fourth of body whole blood volume) with an automatic withdrawal-infusion machine through right common carotid artery. In ligation group, the mesenteric lymph duct was ligated after loss of blood, and in loss of blood group only a thread was passed under the mesenteric lymph duct. The rats' survival rate at 24 hours was recorded. After 24 hours, surviving rats were anesthetized again, and 6 ml of blood was with drew from left common carotid artery rapidly. The platelet adhesive rate, platelet aggregation rate, thrombus formation in vitro, coagulation function were determined before and after experiment, and the cerebral blood flow was measured., Results: There were 6 rats alive in loss of blood group (60%), and 9 rats alive in ligation group (90%). The platelet adhesive rate, platelet aggregation rate, fibrinogen (Fib) content were increased in both groups, and cerebral blood flow was lower compared with before experiment significantly. The activated partial thromboplastin time (APTT) in loss of blood group, and thrombin time (TT) in both groups were prolonged. And the length of wet thrombus, wet weight of thrombus, length of dry thrombus, dry weight of thrombus and thrombus formation rate in loss of blood group were significantly increased compared with before experiment (P<0.05 or P<0.01). But in ligation group, the platelet adhesive rate [(15.02+/-1.24)%], platelet aggregation rate (at 1 minute, 3 minutes and max aggregation rate), length of wet thrombus, wet weight of thrombus, length of dry thrombus, dry weight of thrombus, thrombus formation rate [(46.2+/-6.9)%], APTT [(21.04+/-5.53)s] and Fib content [(433.67+/-13.97) g/L] were lower, cerebral blood flow [(485.1+/-41.4) ml x kg(-1) x min(-1)] was higher compared with loss of blood group, respectively [(18.54+/-1.18)%, (69.8+/-6.9)%, (26.35+/-6.26)s, (510.96+/-35.59) g/L, (417.8+/-42.2) ml x kg(-1) x min(-1), P<0.05 or P<0.01]., Conclusion: The results indicate that acute loss of blood leads to coagulation and increased thrombosis, and mesenteric lymph duct ligation can improve hypercoagulability of blood.

Objective: To observe the effect of ligation of mesenteric lymph duct on changes in free radicals and pro-inflammatory mediators in the liver of rats with serious hemorrhagic shock at different periods, and explore the effect of blockage of intestinal lymphatic pathway on inflammation response of liver., Methods: Seventy-eight male Wistar rats were randomly divided into the sham group (n=6), shock group (n=42), and ligation group (n=30). The model of serious hemorrhagic shock was reproduced in shock group and ligation group. Mesenteric lymph was blocked by ligating mesenteric lymph duct in ligation group after resuscitation. Six rats were sacrificed at 0, 1, 3, 6, 12, and 24 hours, and the livers were harvested and homogenized for the determination of malondialdehyde (MDA), nitric oxide (NO), nitric oxide synthase (NOS), superoxide dismutase (SOD), tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and myeloperoxidase (MPO) activity. The expression of inducible nitric oxide synthase (iNOS) mRNA in liver was detected by reverse transcription-polymerase chain reaction (RT-PCR)., Results: The contents of TNF-alpha, IL-6, NO, NOS, MDA, MPO and iNOS mRNA in liver homogenate of shock group were increased after transfusion and resuscitation, and their levels were higher at 6 and 12 hours. The values were significantly higher than those of the sham group, while the activity of SOD was significantly lower than that of sham group (P<0.05 or P<0.01). The contents of TNF-alpha, IL-6, NO, NOS, MDA, MPO and iNOS mRNA in liver homogenate were lower significantly after transfusion and 3, 6, 12 and 24 hours after resuscitation than those of shock group at each time points, and the SOD activity was higher (P<0.05 or P<0.01)., Conclusion: The results demonstrate that the ligation of mesenteric lymph duct could reduce the polymorphonuclear leucocyte (PMN) detain, and its mechanism might relate to reduction of neutrophil aggregation, thus decreases the release of TNF-alpha and IL-6, reduces the NO and expression of iNOS mRNA, reduces the release of free radicals and consumption of SOD, as a result, it reduces the inflammation response of liver in serious hemorrhagic shock rats.

Objective: To explore the effect of mesenteric lymph duct ligation in preventing apoptosis of renal tubule epithelial cells in rats by two-hits including hemorrhage and lipopolysaccharide (LPS) challenge., Methods: Forty-five Wistar rats were randomly divided into three groups: the ligation group, the non-ligation group and sham operation group. The two-hits model was established by withdrawing the blood via the right side carotid artery of rats and intragastric administration 4 mg/kg LPS 6 hours after hemorrhage, and mesenteric lymph flow was blocked by ligating mesenteric lymph duct in ligation group. Twenty-four hours after operation, kidney was harvested for pathological examination, and the apoptosis cells rate of renal tubule epithelial cells were determined by method of terminal-deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL), the expression of apoptosis-correlating gene bcl-2 and bax protein were determined by streptavidin-biotin complex (SABC) immunohistochemical method., Results: After two-hits, the apoptosis rate and expression of bax protein of renal tubule epithelial cells in non-ligation group were significantly increased, and expression of bcl-2 protein was significantly lower as compared with those of sham operation group (all P<0.01). But the apoptosis rate and expression of bax protein of renal tubule epithelial cells in ligation group were significantly lower, and expression of bcl-2 protein was significantly increased as compared with non-ligation group (all P<0.01)., Conclusion: The results demonstrate that the ligation of mesenteric lymph duct could ameliorate the apoptosis of renal tubule epithelial cells in rats as produced by hemorrhage and LPS, and its mechanism might relate to the reduction of the down regulation of gene expression of bax protein and enhancement of the expression of bcl-2 protein after ligation of mesenteric lymph duct.

Aim: To observe the effect of mesenteric lymph duct ligation on free radical and inflammatory mediator of myocardium with severe hemorrhagic shock in rats at different period, and explore the effect of intestinal lymphatic pathway on myocardium injury pathogenesis in shock rats., Methods: 78 male Wistar rats were divided into the sham group, shock group and ligation group. The model of serious hemorrhagic shock was established in shock group, ligation group, and mesenteric lymph was blocked by ligating mesenteric lymph duct in ligation group after resuscitate. All rats were executed and taken out heart making for homogenate of 10 percent to determine the MDA, SOD, tumor necrosis factor-alpha (TNFalpha), interleukin-6 (IL-6), myeloperoxidase (MPO), NO and NOS at after shock 90 min, after transfusion and resuscitate 0 h, 1 h, 3 h, 6 h, 12 h and 24 h etc. different times, and the expression of inducible nitric oxide synthase (iNOS) mRNA in myocardium was detected by RT-PCR., Results: The contents of MDA, TNFalpha, IL-6, MPO, NO, NOS and iNOS expression in myocardium of shock group were rising after transfusion and resuscitate, and that was higher level at 3 h to 12 h, and that was significantly higher than sham group, the activity of SOD was significantly lower than sham group. The contents of MDA, TNFalpha, IL-6, MPO, NO, NOS and iNOS expression in myocardium of ligation group were significantly lower than that of shock group at sameness points, and the SOD activity was higher., Conclusion: The mesenteric lymph duct ligation and blocking mesenteric lymph could reduce the PMN detaining, decrease the discharging of TNFa and IL-6, reduce the NO and expression of iNOS mRNA, and reduce the releasing of free radical and consuming of SOD.

Objective: To observe the changes of toxic substances in mesenteric lymph and portal vein blood of rats in hemorrhagic shock, and the influence of mesenteric lymph duct ligation on level of endotoxin (ET) in organs and bacterial contents in mesenteric lymph nodes (MLN) and spleen in rats with hemorrhagic shock, and to evaluate the role of lymphatic pathway in pathogenesis of intestine-derived bacteria/endotoxin translocation (BET) in rats with shock., Methods: Twenty-four male Wistar rats were randomly divided into the shock group and control group. A model of serious hemorrhagic shock was reproduced by blood shedding to maintain the blood pressure at 40 mm Hg (1 mm Hg=0.133 kPa) for 90 minutes under aseptic condition, and MLN and portal vein blood were harvested. The specimens were also obtained in control group. The contents of ET, tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) were determined in them. Thirty male Wistar rats were randomly divided into the sham operation group, shock group and lymphatic duct ligation group. Mesenteric lymph ducts were ligated after resuscitation. All rats were sacrificed, and lung, liver, heart and kidney were removed and homogenized for determination of the content of ET. MLN and spleen homogenates were subjected to bacterial culture., Results: The contents of ET, TNF-alpha and IL-6 in lymph were significantly higher than those of plasma in shock group, and also higher than that in normal plasma and normal lymph (all P<0.01). In shock group the contents of ET in lung, liver, heart and renal homogenate 3 and 6 hours after transfusion and resuscitation were significantly higher than those of sham operation group and ligation group (P<0.05 or P<0.01). Bacterial culture of MLN and spleen in shock group rats 3 and 6 hours after transfusion and resuscitation was positive, but it was not in ligation group., Conclusion: The results demonstrate that the intestinal lymphatic pathway plays an important role after compromise of gut barrier function in carrying out BET after hemorrhagic shock.

Objective: To observe the effect of ligation of mesenteric lymph duct on release of oxygen free radical and inflammatory mediator in lung of rat with serious hemorrhagic shock at different periods, and explore the role of intestinal lymphatic pathway on acute lung injury (ALI)., Methods: Seventy-eight male Wistar rats were randomly divided into the sham operation group, shock group, and ligation group. The model of serious hemorrhagic shock was reproduced in shock group, ligation group, and mesenteric lymph flow was occluded by ligating mesenteric lymph duct in ligation group after resuscitation. Eight rats were sacrificed and the lung was harvested and homogenized for the determination of malondialdehyde (MDA), superoxide dismutase (SOD), tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6) and myeloperoxidase (MPO) activity at 90 minutes after shock, 0, 1, 3, 6, 12 and 24 hours after transfusion and resuscitation., Results: The contents of MDA, TNF-alpha, IL-6 and MPO activity in lung homogenate of shock group were elevated after transfusion and resuscitation, and the levels were higher significantly at 3-12 hours, than those of the sham operation group, while the activity of SOD was significantly lower than that of sham operation group (P<0.05 or P<0.01). The contents of MDA, TNF-alpha, IL-6 and MPO in lung homogenate of ligation group at 3, 6, 12 and 24 hours after resuscitation were significantly lower than those of shock group at the same time points, and the SOD activity were higher (P<0.05 or P<0.01)., Conclusion: The results demonstrate that the ligation of mesenteric lymph duct could ameliorate ALI in rats with serious hemorrhagic shock, and its mechanism might relate with reduction of sequestration of neutrophils, thus reduce the release of TNF-alpha and IL-6, and also reduce the release of oxygen free radical and consumption of SOD. Therefore mesenteric lymphatics might play an important role in pathogenesis of ALI.