Mesenteric Lymph Drainage Research Articles

Recombinant VEGF-C (Cys156Ser) improves mesenteric lymphatic drainage and gut immune surveillance in experimental cirrhosis

Lymphatic vessels (LVs) are crucial for maintaining abdominal fluid homoeostasis and immunity. In cirrhosis, mesenteric LVs (mLVs) are dilated and dysfunctional. Given the established role of vascular endothelial growth factor-C (VEGF-C) in improving LVs, we hypothesised that VEGF-C treatment could ameliorate the functions of mLVs in cirrhosis. In this study, we developed a nanoformulation comprising LV-specific growth factor, recombinant human VEGF-C (Cys156Ser) protein (E-VEGF-C) and delivered it orally in different models of rat cirrhosis to target mLVs. Cirrhotic rats were given nanoformulation without VEGF-C served as vehicles. Drainage of mLVs was analysed using tracer dye. Portal and systemic physiological assessments and computed tomography were performed to measure portal pressures and ascites. Gene expression and permeability of primary mesenteric lymphatic endothelial cells (LyECs) was studied. Immune cells in mesenteric lymph nodes (MLNs) were quantified by flow cytometry. Endogenous and exogenous gut bacterial translocation to MLNs was examined. In cirrhotic rats, mLVs were dilated and leaky with impaired drainage. Treatment with E-VEGF-C induced proliferation of mLVs, reduced their diameter, and improved functional drainage. Ascites and portal pressures were significantly reduced in E-VEGF-C rats compared with vehicle rats. In MLNs of E-VEGF-C animals, CD8+CD134+ T cells were increased, whereas CD25+ regulatory T cells were decreased. Both endogenous and exogenous bacterial translocation were limited to MLNs in E-VEGF-C rats with reduced levels of endotoxins in ascites and blood in comparison with those in vehicle rats. E-VEGF-C treatment upregulated the expression of vascular endothelial-cadherin in LyECs and functionally improved the permeability of these cells. E-VEGF-C treatment ameliorates mesenteric lymph drainage and portal pressure and strengthens cytotoxic T-cell immunity in MLNs in experimental cirrhosis. It may thus serve as a promising therapy to manage ascites and reduce pathogenic gut bacterial translocation in cirrhosis. A human recombinant pro-lymphangiogenic growth factor, VEGF-C, was encapsulated in nanolipocarriers (E-VEGF-C) and orally delivered in different models of rat liver cirrhosis to facilitate its gut lymphatic vessel uptake. E-VEGF-C administration significantly increased mesenteric lymphatic vesselproliferation and improved lymph drainage, attenuating abdominal ascites and portal pressures in the animal models. E-VEGF-C treatment limited bacterial translocation to MLNs only with reduced gut bacterial load and ascitic endotoxins. E-VEGF-C therapy thus holds the potential to manage ascites and portal pressure and reduce gut bacterial translocation in patients with cirrhosis.

Innate immunity and immunotherapy for hemorrhagic shock.

Hemorrhagic shock (HS) is a shock result of hypovolemic injury, in which the innate immune response plays a central role in the pathophysiology ofthe severe complications and organ injury in surviving patients. During the development of HS, innate immunity acts as the first line of defense, mediating a rapid response to pathogens or danger signals through pattern recognition receptors. The early and exaggerated activation of innate immunity, which is widespread in patients with HS, results in systemic inflammation, cytokine storm, and excessive activation of complement factors and innate immune cells, comprised of type II innate lymphoid cells, CD4+ T cells, natural killer cells, eosinophils, basophils, macrophages, neutrophils, and dendritic cells. Recently, compelling evidence focusing on the innate immune regulation in preclinical and clinical studies promises new treatment avenues to reverse or minimize HS-induced tissue injury, organ dysfunction, and ultimately mortality. In this review, we first discuss the innate immune response involved in HS injury, and then systematically detail the cutting-edge therapeutic strategies in the past decade regarding the innate immune regulation in this field; these strategies include the use of mesenchymal stem cells, exosomes, genetic approaches, antibody therapy, small molecule inhibitors, natural medicine, mesenteric lymph drainage, vagus nerve stimulation, hormones, glycoproteins, and others. We also reviewed the available clinical studies on immune regulation for treating HS and assessed the potential of immune regulation concerning a translation from basic research to clinical practice. Combining therapeutic strategies with an improved understanding of how the innate immune system responds to HS could help to identify and develop targeted therapeutic modalities that mitigate severe organ dysfunction, improve patient outcomes, and reduce mortality due to HS injury.

Mesenteric Lymph Drainage Improves Cardiac Papillary Contractility and Calcium Sensitivity in Rats with Hemorrhagic Shock

Myocardial dysfunction is an important adverse factor of hemorrhagic shock that induces refractory hypotension, and post-hemorrhagic shock mesenteric lymph (PHSML) return is involved in this adverse effect. This study investigated whether mesenteric lymph drainage (MLD) improves PHSML return-induced cardiac contractile dysfunction via the restoration of cardiomyocyte calcium sensitivity. A hemorrhage shock model was established by using a controlled hemorrhage through the femoral artery that maintained a mean arterial pressure of 40 ± 2 mmHg for 3 h. MLD and mesenteric lymph duct ligation (MLDL) were performed from 1 to 3 h during hypotension. The papillarymuscles of the heart were collected for measurement of calmodulin expression and for determining contractile responses to either isoprenaline or calcium. The results showed that either MLD or MLDL reversed the hemorrhagic shock-induced downregulation of calmodulin expression, a marker protein of cardiomyocyte calcium sensitization, in papillary muscles. MLD also improved the decreased contractile response and ±df/dt of the papillary muscle strip to gradientisoprenaline or calcium caused by hemorrhagic shock. These findings indicate that increased cardiac contractibility may be associated with the restoration of calcium sensitivity produced by PHSML drainage.

Mesenteric Lymph Duct Drainage Attenuates Lung Inflammatory Injury and Inhibits Endothelial Cell Apoptosis in Septic Rats.

The present study was to investigate the effect of mesenteric lymph duct drainage on lung inflammatory response, histological alteration, and endothelial cell apoptosis in septic rats. Animals were randomly assigned into four groups: control, sham surgery, sepsis, and sepsis plus mesenteric lymph drainage. We used the colon ascendens stent peritonitis (CASP) procedure to induce the septic model in rats, and mesenteric lymph drainage was performed with a polyethylene (PE) catheter inserted into mesenteric lymphatic. The animals were sacrificed at the end of CASP in 6 h. The mRNA expression levels of inflammatory mediators were measured by qPCR, and the histologic damage were evaluated by the pathological score method. It was found that mesenteric lymph drainage significantly reduced the expression of TNF-α, IL-1β, and IL-6 mRNA in the lung. Pulmonary interstitial edema and infiltration of inflammatory cells were alleviated by mesenteric lymph drainage. Moreover, increased mRNA levels of TNF-α, IL-1β, IL-6 mRNA, and apoptotic rate were observed in PMVECs treated with septic lymph. These results indicate that mesenteric lymph duct drainage significantly attenuated lung inflammatory injury by decreasing the expression of pivotal inflammatory mediators and inhibiting endothelial apoptosis to preserve the pulmonary barrier function in septic rats.

Why is SARS-CoV-2 infection more severe in obese men? The gut lymphatics – Lung axis hypothesis

Consistent observations report increased severity of SARS-CoV-2 infection in overweight men with cardiovascular factors. As the visceral fat possesses an intense immune activity, is involved in metabolic syndrome and is at the crossroad between the intestines, the systemic circulation and the lung, we hypothesized that it plays a major role in severe forms of SARS-CoV-2 infection. SARS-CoV2 presents the ability to infect epithelial cells of the respiratory tract as well as the intestinal tract. Several factors may increase intestinal permeability including direct enterocyte damage by SARS-CoV2, systemic inflammatory response syndrome (SIRS) and epithelial ischemia secondary to SARS-CoV2- associated endothelial dysfunction. This increase permeability further leads to translocation of microbial components such as MAMPs (microbial-associated molecular pattern), triggering an inflammatory immune response by TLR-expressing cells of the mesentery fat (mostly macrophages and adipocytes). The pro-inflammatory cytokines produced by the mesentery fat mediates systemic inflammation and aggravate acute respiratory distress syndrome (ARDS) through the mesenteric lymph drainage.

Mesenteric lymph drainage alleviates hemorrhagic shock-induced spleen injury and inflammation.

Purpose:To investigate the effect of mesenteric lymph drainage on the spleen injury and the expressions of inflammatory cytokines in splenic tissue in mice following hemorrhagic shock.Methods:Male C57 mice were randomly divided into the sham shock, shock and shock+drainage groups. The mice in both shock and shock+drainage groups suffered femoral artery bleeding, maintained mean arterial pressure (MAP) of 40±2 mmHg for 90 min, and were resuscitated. And mesenteric lymph drainage was performed in the shock+drainage group at the time of resuscitation. After three hours of resuscitation, the splenic tissues were harvested for the histological observation and protein and mRNA expression analysis of cytokines.Results:The spleen in the shock group revealed a significantly structural damage and increased mRNA expressions of MyD88 and TRAF6 and protein expressions of TIPE2, MyD88, TRIF and TRAF3 compared to the sham group. By contrast, the splenic pathological injury in the shock+drainage group was alleviated significantly, and the mRNA and protein expressions of TIPE2, MyD88, TRIF, TRAF3 and TRAF6 were significantly lower than those in the shock group.Conclusion:These results indicate that post-hemorrhagic shock mesenteric lymph drainage alleviates hemorrhagic shock-induced spleen injury and the expressions of inflammatory cytokines.

TNFΔARE Mice Display Abnormal Lymphatics and Develop Tertiary Lymphoid Organs in the Mesentery

Chronic inflammatory diseases are associated with a persistent and enhanced response to environmental antigens. As an adaptive response to this exaggerated immune state, affected tissue typically develops tertiary lymphoid organs. Studies of Crohn disease (CD), a chronic inflammatory disease of the intestinal tract, report tertiary lymphoid organs present within the mucosal wall, along with other lymphatic diseases, such as lymphangiogenesis and obstructed lymphatic vessels. These observations suggest that downstream mesenteric lymphatic vessels and lymph drainage into mesenteric lymph nodes may be compromised. However, information is lacking on the morphologic features and functional status of mesenteric lymphatics in CD. Using confocal imaging, PCR, flow cytometry, and functional strategies, we addressed these questions in the established TNFΔARE mouse model of CD and found that this mouse model had many lymphatic abnormalities reminiscent of human CD. These abnormalities include intestinal lymphangiectasia, mesenteric lymph node lymphadenopathy, and lymphangiogenesis in both the mesentery and mucosa. Critically, TNFΔARE mice also present mesenteric tertiary lymphoid organs and have altered lymphatic transport of dendritic cells to mesenteric lymph nodes, two features likely to actively modulate immunity. Our findings provide key insights into lymphatic remodeling in the TNFΔARE mouse model. They shed light on the involvement of these lymphatic changes in immune dysfunctions observed in CD and suggest the lymphatic system as new target for therapeutic options.

Role of mesenteric lymph drainage in the balance of ACE/ACE2 in murine myocardium following hemorrhagic shock

To observe the change of angiotensin converting enzyme (ACE) and ACE2 in the murine myocardium followed hemorrhagic shock and the role of post-hemorrhagic shock mesenteric lymph (PHSML) drainage. Twenty-four male mice were ran-domly divided into control, sham, shock, and shock + drainage groups. A hemorrhagic shock model was established and then fluid resuscita-tion was performed to the mice in the shock and shock + drainage groups, and the PHMSL was drained in the shock + drainage group after fluid resuscitation. After 6 h of resuscitation in the shock and shock + drainage groups or corresponding time in the sham group, or after anesthesia in the control group, the myocardial tissues were harvested for the determination of the mRNA expressions of ACE, ACE2, angiotensin Ⅱ (Ang Ⅱ) type 1 receptor (AT1R), and Mas related G protein coupled receptor (Mas1R) using the method of qRT-PCR, and the levels of Ang Ⅱ and Ang (1-7) using the method of ELISA. In the myocardial tissue of shock group, the ACE and AT1R mRNA expressions and Ang Ⅱ level were significantly increased than those of the control and sham groups, the ACE2 and Mas1R mRNA expressions were significantly de-creased than that of the control and sham groups, the Ang (1-7) level was decreased compared with the control group, the ratios of ACE/ACE2, Ang Ⅱ/Ang (1-7), and AT1R/Mas1R in the shock group were significantly increased than the control and sham groups. Meanwhile, PHSML drainage obviously suppressed the effects of hemorrhagic shock on these indices. Hemorrhagic shock up-regulated the ACE-Ang Ⅱ-AT1R axis, down-regulated the ACE2-Ang (1-7)-Mas1R axis, and induced the unbalance of ACE and ACE2 in myocardial tis-sue. PHSML drainage decreased the ACE-Ang Ⅱ-AT1R axis and increased the ACE2-Ang (1-7)-Mas1R axis, resulted in the balance of ACE and ACE2.

Inhibitory effect of post-hemorrhagic shock mesenteric lymph drainage on the HMGB1 and RAGE in mouse kidney

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.

Hydrogen sulfide in posthemorrhagic shock mesenteric lymph drainage alleviates kidney injury in rats.

Posthemorrhagic shock mesenteric lymph (PHSML) is a key factor in multiple organ injury following hemorrhagic shock. We investigated the role of hydrogen sulfide (H2S) in PHSML drainage in alleviating acute kidney injury (AKI) by administering D,L-propargylglycine (PPG) and sodium hydrosulfide hydrate (NaHS) to 12 specific pathogen-free male Wistar rats with PHSML drainage. A hemorrhagic shock model was established in 4 experimental groups: shock, shock+drainage, shock+drainage+PPG (45 mg/kg, 0.5 h prehemorrhage), and shock+drainage+NaHS (28 µmol/kg, 0.5 h prehemorrhage). Fluid resuscitation was performed after 1 h of hypotension, and PHMSL was drained in the last three groups for 3 h after resuscitation. Renal function and histomorphology were assessed along with levels of H2S, cystathionine-γ-lyase (CSE), Toll-like receptor 4 (TLR4), interleukin (IL)-10, IL-12, and tumor necrosis factor (TNF)-α in renal tissue. Hemorrhagic shock induced AKI with increased urea and creatinine levels in plasma and higher H2S, CSE, TLR4, IL-10, IL-12, and TNF-α levels in renal tissue. PHSML drainage significantly reduced urea, creatinine, H2S, CSE, and TNF-α but not TLR4, IL-10, or IL-12. PPG decreased creatinine, H2S, IL-10, and TNF-α levels, but this effect was reversed by NaHS administration. In conclusion, PHSML drainage alleviated AKI following hemorrhagic shock by preventing increases in H2S and H2S-mediated inflammation.

Mesenteric lymph duct drainage attenuates acute lung injury in rats with severe intraperitoneal infection.

The purpose of this study is to investigate the hypothesis that the mesenteric lymphatic system plays an important role in acute lung injury in a rat model induced by severe intraperitoneal infection. Male Wistar rats weighing 250∼300 g were randomly divided into 3 groups and subjected to sham operation, intraperitoneal infection, or mesenteric lymphatic drainage. The activity of diamine oxidase (DAO) and myeloperoxidase (MPO) were measured by enzymatic assay. The endotoxin levels in plasma, lymph, and bronchoalveolar lavage fluid (BALF) were evaluated using the limulus amoebocyte lysate reagent. The cytokines, adhesion factors, chemokines, and inflammatory factors were detected by ELISA. TLR-4, NF-kB, and IRAK-4 were analyzed by Western blotting. Compared with sham-operated rats, rats with intraperitoneal infection had increased MPO and decreased DAO activity in intestinal tissues. Mesenteric lymph drainage reduced the alterations in MPO and DAO activity induced by intraperitoneal infection. The MPO activity in pulmonary tissue and the permeability of pulmonary blood vessels were also increased, which were partially reversed by mesenteric lymph drainage. The endotoxin levels in lymphatic fluid and alveolar perfusion fluid were elevated after intraperitoneal infection but decreased to control levels after lymph drainage. No alterations in the levels of plasma endotoxin were observed. The number of neutrophils was increased in BALF and lymph in the infected rats, and was also reduced after drainage. Lymph drainage also decreased the levels of inflammatory cytokines, chemokines, and adhesion factors in the plasma, lymph, and BALF, as well as the levels of TLR-4, NF-kB, and IRAK-4 in pulmonary and intestinal tissues. The mesenteric lymphatic system is the main pathway involved in early lung injury caused by severe intraperitoneal infection, in which activation of the TLR-4 signal pathway may play a role.

Changes in renal tissue proteome induced by mesenteric lymph drainage in rats after hemorrhagic shock with resuscitation.

Kidney injury commonly occurs after hemorrhagic shock. Previous studies have shown that post-hemorrhagic shock mesenteric lymph (PHSML) return negatively affects the kidneys and may induce injury. This study investigates the effect of PHSML drainage on the proteome in renal tissue. A controlled hemorrhagic shock model was established in the shock and shock+drainage groups. After 1 h of hypotension, fluid resuscitation was implemented within 30 min. Meanwhile, PHSML was drained in the shock+drainage group. After 3 h of resuscitation, renal tissue was extracted for proteome analysis using two-dimensional fluorescence difference gel electrophoresis. Differential proteins with intensities that either increased or decreased by 1.5-fold or greater were selected for trypsin digestion and analyzed by matrix-assisted laser desorption/ionization time-of-flight (TOF) mass spectrometry and tandem TOF/TOF mass spectrometry. Enzyme-linked immunosorbent assay was used to validate the identified partial proteins. Compared with the sham group, hnRNPC and Starp decreased in the shock group, whereas Hadha, Slc25a13, Atp5b, hnRNPC, Starp, Rps3, and actin were downregulated in the shock+drainage group. Meanwhile, Atp5b and actin decreased in the shock+drainage group relative to the shock group. The identified proteins can be classified into different categories, such as cell proliferation (hnRNPC, Strap, and Rps3), energy metabolism (Hadha, Atp5b, and Slc25a13), cell motility, and cytoskeleton (actin). Moreover, enzyme-linked immunosorbent assay measurement validated the changed levels of Atp5b and Actg2. Our findings provide a starting point for investigating the functions of differentially expressed proteins in acute kidney injury induced by hemorrhagic shock. These findings hold great potential for the development of therapeutic interventions.

Post-shock mesenteric lymph drainage ameliorates cellular immune function in rats following hemorrhagic shock.

Disturbance of immunity is an important factor to modulate inflammatory responses after severe shock. Post-shock mesenteric lymph (PSML) return plays an adverse role in multiple organ injuries induced by the hemorrhagic shock, and the inflammatory factors are involved in this process. However, whether the PSML can exacerbate immune dysfunctions that modulate inflammatory response to the hemorrhagic shock remains unknown. In the present study, the effects of PSML drainage on the distribution of T lymphocyte subgroup, the release of inflammatory factors, and apoptosis of thymocytes were investigated; the effect of PSML on the specific parameters of cellular immune function was also determined. Results showed that PSML drainage reduced the increased levels of CD3+, CD3+CD4+, CD4+CD25+ lymphocytes, IFN-γ, and the ratios of CD3 + CD4+/CD3 + CD4- in blood of the shocked rats at 3 h after resuscitation; PSML drainage also abolished the decreased IL-4 level and restored the higher ratio of IFN-γ/IL-4 to normal levels. Tissue injury, including enlarged intermembrance space and edema with congestion in the medulla, increased apoptotic cells and bax expression, decreased number of cells in the S phase, and bcl-2 expression were observed in the thymus after hemorrhagic shock. PSML drainage reversed these effects. In particular, PSML drainage increased the proliferation index and decreased p53 expression of thymocytes. These results suggest that hyperimmunity occurred at early stages of hemorrhagic shock with resuscitation and that PSML drainage could markedly improve cellular immune function that is responsible for the reduced inflammatory responses.

Mesenteric lymph drainage alleviates acute kidney injury induced by hemorrhagic shock without resuscitation.

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.

Postshock mesenteric lymph drainage ameliorates vascular reactivity and calcium sensitivity through RhoA

BackgroundVascular hyporeactivity plays an important role in the pathogenesis of severe shock. Previous studies have shown that postshock mesenteric lymph (PSML) blockage ameliorates the vascular reactivity and calcium sensitivity, and RhoA is involved in the regulation of vascular reactivity after hemorrhagic shock. Therefore, the present study tested whether small GTPase RhoA mediates the improvement of the vascular reactivity and calcium sensitivity in superior mesenteric artery (SMA) of rats with PSML drainage. Materials and methodsThe hemorrhagic shock model (blood pressure to 40 ± 2 mm Hg) was established, and PSML was drained from immediate hypotension for 3 h, after which SMA was isolated, and the vascular reactivity and calcium sensitivity were tested in the presence of RhoA agonist (U-46619) or inhibitor (C3 transferase). The protein expressions of small GTPase RhoA and phospho-RhoA were also examined in SMA. ResultsThe hemorrhagic shock resulted in a significant decrease in the SMA reactivity and calcium sensitivity, which was enhanced by the application of U-46619 to the SMA. In contrast, the PSML drainage ameliorated the deleterious effect of the hemorrhagic shock on the SMA. This beneficial effect of the PSML drainage was abolished by C3 transferase. Western blotting revealed that the expressions of the RhoA and phospho-RhoA in SMA tissue obtained from the shock group were significantly decreased, and the PSML drainage markedly enhanced these protein expressions. ConclusionsRhoA is an important contributor to the PSML drainage-induced amelioration of the vascular reactivity and calcium sensitivity in rats with hemorrhagic shock.

Myosin light chain kinase is necessary for post-shock mesenteric lymph drainage enhancement of vascular reactivity and calcium sensitivity in hemorrhagic-shocked rats

Vascular hyporeactivity is an important factor in irreversible shock, and post-shock mesenteric lymph (PSML) blockade improves vascular reactivity after hemorrhagic shock. This study explored the possible involvement of myosin light chain kinase (MLCK) in PSML-mediated vascular hyporeactivity and calcium desensitization. Rats were divided into sham (n=12), shock (n=18), and shock+drainage (n=18) groups. A hemorrhagic shock model (40±2 mmHg, 3 h) was established in the shock and shock+drainage groups. PSML drainage was performed from 1 to 3 h from start of hypotension in shock+drainage rats. Levels of phospho-MLCK (p-MLCK) were determined in superior mesenteric artery (SMA) tissue, and the vascular reactivity to norepinephrine (NE) and sensitivity to Ca2+ were observed in SMA rings in an isolated organ perfusion system. p-MLCK was significantly decreased in the shock group compared with the sham group, but increased in the shock+drainage group compared with the shock group. Substance P (1 nM), an agonist of MLCK, significantly elevated the decreased contractile response of SMA rings to both NE and Ca2+ at various concentrations. Maximum contractility (Emax) in the shock group increased with NE (from 0.179±0.038 to 0.440±0.177 g/mg, P<0.05) and Ca2+ (from 0.515±0.043 to 0.646±0.096 g/mg, P<0.05). ML-7 (0.1 nM), an inhibitor of MLCK, reduced the increased vascular response to NE and Ca2+ at various concentrations in the shock+drainage group (from 0.744±0.187 to 0.570±0.143 g/mg in Emax for NE and from 0.729±0.037 to 0.645±0.056 g/mg in Emax for Ca2+, P<0.05). We conclude that MLCK is an important contributor to PSML drainage, enhancing vascular reactivity and calcium sensitivity in rats with hemorrhagic shock.

Mesenteric Lymph Return Is an Important Contributor to Vascular Hyporeactivity and Calcium Desensitization After Hemorrhagic Shock

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.

Role of mesenteric lymph drainage improving the metabolism of red blood cell in hemorrhagic shock rats following fluid resuscitation

To observe the effects of mesenteric lymph drainage on the metabolism of red blood cell (RBC) in hemorrhagic shock (HS) rats following fluid resuscitation. Eighteen male Wistar rats were randomly divided into sham group (n=6), HS group (n=6), HS + drainage group (n=6). After 1.5 hours of HS model prepared, the animals were given fluid resuscitation by lost blood plus equal volume of Ringer solution within 30 minutes in HS and HS + drainage groups, and mesenteric lymph drainage was performed after 1 hour of hypotension in HS + drainage group. At 3 hours after resuscitation or corresponding time, blood samples were obtained from abdominal aorta. Membrane suspensions of RBC prepared from part of whole blood samples were used to measure the activities of adenosine triphosphate ase (ATPase) and contents of ATP and lactic acid (LA), the intracellular fluid of RBC prepared from part of whole blood samples was used to determine the concentration of 2,3-diphosphoglyceric acid (2,3-DPG), Na(+) and K(+), plasma samples isolated from blood by centrifugation were used to determine the concentration of Na(+), K(+), Cl(-) and total Ca. Compared with sham group, the content of ATP (μmol/g), activity of Na(+)-K(+)-ATPase (μmol×mg(-1)×h(-1)) and Ca(2+)-ATPase (μmol×mg(-1)×h(-1)) in RBC membrane and total Ca (mmol/L) in plasma were decreased markedly (ATP: 6.698±0.938 vs. 10.670±1.466, Na(+)-K(+)-ATPase: 0.042±0.010 vs. 0.066±0.019, Ca(2+)-ATPase: 0.054±0.015 vs. 0.081±0.017, total Ca: 2.27±0.18 vs. 2.66±0.21, P<0.05 or P<0.01) in HS group, and the content of LA (mmol/g) in RBC and K(+) (mmol/L), Cl(-) (mmol/L) in plasma were increased observably in HS group (LA: 3.472±0.853 vs. 1.743±0.395, K(+): 5.83±0.34 vs. 5.23±0.37, Cl(-): 113.37±3.63 vs. 106.35±4.99, P<0.05 or P<0.01), there was no significant difference in term of the contents of 2,3-DPG (mmol/L: 2.196±0.609 vs. 2.590±0.574, P>0.05). Compared with HS group, the contents of 2,3-DPG (4.459±0.900) and ATP (8.859±1.189), the activities of Na(+)-K(+)-ATPase (0.089±0.022), Ca(2+)-ATPase (0.082±0.020) of RBC were increased in HS + drainage group, and the level of LA (2.060±0.810) was decreased observably (P<0.05 or P<0.01), there were no significant differences in the other indices. The results indicate that shock mesenteric lymph drainage plays an important role in improving the metabolism of RBC in HS rats following fluid resuscitation, subsequently, may preserve the structure and function of RBC.

Effect of mesenteric lymph obtained during shock on pump activity and oxygen free radical on erythrocyte membrane of rat

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. 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. 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). Mesenteric lymph return is a key factor in decreasing pump activity while aggravating free radical damage injury of erythrocyte membrane.

Morphological Effects of Intraperitoneal Oily Fatty Acids on the Intestine and Peritoneum of Rats

Changes of the mucosa of the small intestine in animal experiments can be produced by a variety of methods. The direct application of numerous chemical agents on the epithelium of the mucosa and parenteral administration of cytostatics as well as an indirect damage by experimental intestinal lymphatic obstruction lead to mucosal lesions with atrophy of the villi. Our paper presents a simple indirect method for induction of structural mucosa changes in rats using experimentally produced peritoneal alterations as a precursor. The first experimental group with 108 male wistar rats was divided into 5 subgroups and treated as follows: 43 rats got continuous daily intraperitoneal injections of 3-3 ml/kg body weight sesame oil DAB 6 which contains 50% oleic acid, 36% linolic acid, 1% free fatty acids, 0.1% lecithin and portions of stearic, palmitic and arachic acid; 6 rats got 6.6 ml sesame oil per kg i.p., 43 animals received daily doses of 3-3 ml oil/kg per orally by a pharyngeal tube, 6 rats of each group were sacrificed after 68 and 92 days, all the rest were killed after 100 days of continuous daily treatment. Aortic blood was used for determination of the total cholesterol serum concentration. Blocks of tissue of every organ were fixed in 4% formalin, sections were stained with hematoxylin and eosin, hematoxylin-sudan and by the periodic acid Schiff (PAS-) method. For the purpose of autoradiographic determination of the 3H-thymidine labelling index the rats were administered i.p. 2 mCi 3H-thymidine/kg 40 minutes prior to death. Autoradiographs were prepared from sections of the small intestine by using liquid photoemulsion Ilford K5. In the second experimental group 7 rats got a single i.p. injection of 5 ml, 5 rats of the 10 ml and 9 rats of 20 ml sesame oil per animal; 4 rats were administered 8 ml daily for 5 days...