Induced Gut Injury Research Articles

Mesenchymal stem cell extracellular vesicles mitigate vascular permeability and injury in the small intestine and lung in a mouse model of hemorrhagic shock and trauma.

Hemorrhagic shock and trauma (HS/T)-induced gut injury may play a critical role in the development of multi-organ failure. Novel therapies that target gut injury and vascular permeability early after HS/T could have substantial impacts on trauma patients. In this study, we investigate the therapeutic potential of human mesenchymal stem cells (MSCs) and MSC-derived extracellular vesicles (MSC EVs) in vivo in HS/T in mice and in vitro in Caco-2 human intestinal epithelial cells. In vivo, using a mouse model of HS/T, vascular permeability to a 10-kDa dextran dye and histopathologic injury in the small intestine and lungs were measured among mice. Groups were (1) sham, (2) HS/T + lactated Ringer's (LR), (3) HS/T + MSCs, and (4) HS/T + MSC EVs. In vitro, Caco-2 cell monolayer integrity was evaluated by an epithelial cell impedance assay. Caco-2 cells were pretreated with control media, MSC conditioned media (CM), or MSC EVs, then challenged with hydrogen peroxide (H2O2). In vivo, both MSCs and MSC EVs significantly reduced vascular permeability in the small intestine (fluorescence units: sham, 456 ± 88; LR, 1067 ± 295; MSC, 765 ± 258; MSC EV, 715 ± 200) and lung (sham, 297 ± 155; LR, 791 ± 331; MSC, 331 ± 172; MSC EV, 303 ± 88). Histopathologic injury in the small intestine and lung was also attenuated by MSCs and MSC EVs. In vitro, MSC CM but not MSC EVs attenuated the increased permeability among Caco-2 cell monolayers challenged with H2O2. Mesenchymal stem cell EVs recapitulate the effects of MSCs in reducing vascular permeability and injury in the small intestine and lungs in vivo, suggesting MSC EVs may be a potential cell-free therapy targeting multi-organ dysfunction in HS/T. This is the first study to demonstrate that MSC EVs improve both gut and lung injury in an animal model of HS/T.

Electrical stimulation of the vagus nerve improves intestinal blood flow after trauma and hemorrhagic shock

BackgroundGut damage after trauma/hemorrhagic shock contributes to multiple organ dysfunction syndrome. Electrical vagal nerve stimulation is known to prevent gut damage in animal models of trauma/hemorrhagic shock by altering the gut inflammatory response; however, the effect of vagal nerve stimulation on intestinal blood flow, which is an essential function of the vagus nerve, is unknown. This study aimed to determine whether vagal nerve stimulation influences the abdominal vagus nerve activity, intestinal blood flow, gut injury, and the levels of autonomic neuropeptides. MethodsMale Sprague Dawley rats were anesthetized, and the cervical and abdominal vagus nerves were exposed. One pair of bipolar electrodes was attached to the cervical vagus nerve to stimulate it; another pair of bipolar electrodes were attached to the abdominal vagus nerve to measure action potentials. The rats underwent trauma/hemorrhagic shock (with maintenance of mean arterial pressure of 25 mmHg for 30 min) without fluid resuscitation and received cervical vagal nerve stimulation post-injury. A separate cohort of animals were subjected to transection of the abdominal vagus nerve (vagotomy) just before the start of cervical vagal nerve stimulation. Intestinal blood flow was measured by laser Doppler flowmetry. Gut injury and noradrenaline level in the portal venous plasma were also assessed. ResultsVagal nerve stimulation evoked action potentials in the abdominal vagus nerve and caused a 2-fold increase in intestinal blood flow compared to the shock phase (P < .05). Abdominal vagotomy eliminated the effect of vagal nerve stimulation on intestinal blood flow (P < .05). Vagal nerve stimulation protected against trauma/hemorrhagic shock -induced gut injury (P < .05), and circulating noradrenaline levels were decreased after vagal nerve stimulation (P < .05). ConclusionCervical vagal nerve stimulation evoked abdominal vagal nerve activity and relieved the trauma/hemorrhagic shock–induced impairment in intestinal blood flow by modulating the vasoconstriction effect of noradrenaline, which provides new insight into the protective effect of vagal nerve stimulation.

Tissue-Expressed B7-H1 Critically Controls Intestinal Inflammation

B7-H1 (PD-L1) on immune cells plays an important role in Tcell coinhibition by binding its receptor PD-1. Here, we show that both human and mouse intestinal epithelium express B7-H1 and that B7-H1-deficient mice are highly susceptible to dextran sodium sulfate (DSS)- or trinitrobenzenesulfonic acid (TNBS)-induced gut injury. B7-H1 deficiency during intestinal inflammation leads to high mortalityand morbidity, which are associated with severe pathological manifestations in the colon, including loss of epithelial integrity and overgrowth of commensal bacteria. Results from bone marrow chimeric and knockout mice show that B7-H1 expressed on intestinal parenchyma, but not on hematopoietic cells, controls intestinal inflammation in an adaptive immunity-independent fashion. Finally, we demonstrate that B7-H1 dampened intestinal inflammation by inhibiting tumor necrosis factor α (TNF-α) production and by stimulating interleukin 22 secretion from CD11c(+)CD11b(+) lamina propria cells. Thus, our data uncover a mechanism through which intestinal tissue-expressed B7-H1 functions as an essential ligand for innate immune cells to prevent gut inflammation.

Oral pretreatment with recombinant human lactoferrin limits trauma-hemorrhagic shock–induced gut injury and the biological activity of mesenteric lymph

BackgroundLactoferrin (LF) is a pleiotropic glycoprotein that is found in bodily secretions and is postulated to enhance the gastrointestinal barrier and promote mucosal immunity. Thus, the ability of talactoferrin, an oral recombinant form of human LF, to limit gut injury and the production of biologically active gut-derived products was tested using a rat model of trauma–hemorrhagic shock (T/HS). MethodsMale rats were orally dosed with vehicle or talactoferrin (1000 mg/kg, every day) for 5 d before being subjected to T/HS or trauma–sham shock (T/SS). Subsequently, rats were subjected to a laparotomy (trauma) and hemorrhagic shock (mean arterial pressure, 30–35 mm Hg × 90 min) or to T/SS, followed by resuscitation with their shed blood. Before inducing shock, the mesenteric lymphatic duct was catheterized for collection of mesenteric lymph. Four hours after the end of the shock or sham-shock period, rats were sacrificed, a segment of the distal ileum was collected for morphologic analysis, and lymph samples were processed and frozen. Subsequently, lymph samples were tested in several pharmacodynamic assays, including endothelial cell permeability, neutrophil respiratory burst activity, and red blood cell (RBC) deformability. Total white blood cell counts in lymph samples were also quantified. ResultsPretreatment with talactoferrin reduced the incidence of T/HS-induced morphologic injury of ileum to T/SS levels. Post-T/HS lymph from vehicle-treated rats increased endothelial monolayer permeability and neutrophil priming for an augmented respiratory burst, and induced loss of RBC deformability, compared with T/SS groups. Talactoferrin pretreatment significantly reduced the biological activity of T/HS lymph on respiratory burst activity and RBC deformability, but had no effect on the lymph cell count or endothelial cell permeability. ConclusionsThese results provide a proof of principle that prophylactic dosing of oral talactoferrin can potentially protect the gut in a T/HS model and limit the production of biologically active factors in rat gastrointestinal tissue subjected to ischemia-reperfusion–type injuries.

Colostrum - Its Composition, Benefits As A Nutraceutical : A Review

Colostrumis ‘the first milk secreted at the time of parturition, differing from the milk secreted later, by containing more lactalbumin and lactoprotein, and also being rich in antibodies that confer passive immunity to the newborn, also called “foremilk”. Human Colostrum (HC) and Bovine Colostrum (BC) are rich in protein, immunoglobulin, lactoferrin and growth factors. Recent studies suggest that colostrum components, immunoglobulin and growth factor benefits physically active personandin treatment of autoimmune disorders.It isused for the treatment of a wide variety of gastrointestinal conditions, including non-steroidalanti-inflammatory drug–induced gut injury, H pylori infection,immunedeficiency related diarrhea as well as infective diarrhea. This review explores the current knowledge on the beneficial effect of colostrum supplementation in the above condition.

Role of mesenteric lymph pathway in the effects of exogenous somatostatin against early intestinal injury after ischemia–reperfusion in rats

Intestinal ischemia–reperfusion (I/R)-induced gut injury remains a challenge for critically ill patients. This study aims to test whether mesenteric lymph pathway is involved in intestinal I/R injury and whether somatostatin (SST) affects mesenteric lymph pathway after mesenteric reperfusion. Intestinal I/R rats were treated with SST-14 by intravenous injection combined with intraperitoneal injection before occlusion of the SMA until the end of the experiment. When intestinal I/R injury treated with SST, the volumes of mesenteric lymph flow at the 6th h after reperfusion following intestinal ischemia were increased ([0.55±0.24] ml/h vs [0.25±0.09] ml/h, p<0.05) and the number of intestinal lymphocytes per milliliter ([2.30±0.72]×107/ml vs [1.16±0.63]×107/ml, p<0.05) was also increased, which caused the number of intestinal lymphocytes output at the same period of time was significantly increased compared with intestinal I/R group ([1.33±0.88]×107/h vs [0.28±0.15]×107/h, p<0.05). Meanwhile, the number of 51Cr-lymphocytes migration from systemic circulation to the effector sites in GALT was significantly increased ([1.93±0.23]×105/h vs [0.90±0.25]×105/h, p<0.05), although the percentage of 51Cr-lymphocytes in the effector sites ([1.45±0.26]% vs [3.23±1.69]%, p<0.05) was sharply decreased compared with intestinal I/R group. The accompanying decreases of the endotoxin concentration ([0.038±0.017] EU/mL vs [0.110±0.028] EU/mL, compared with intestinal I/R group p<0.05) and the TNF-α levels ([37.50±10.45] ρg/ml) vs ([74.93±14.77] ρg/ml), compared with intestinal I/R group p<0.05) in mesenteric lymph and the improvement of vital organ dysfunction happened during the early intestinal I/R injury. Suppression of gut-derived toxic mediators reaching systemic circulation and increases of the number of lymphocytes homing to the effector sites in GALT to strengthen the effective immune responses in intestinal mucosa account for the protective effects of exogenous SST against early intestinal I/R injury.

Uncovering the neuroenteric–pulmonary axis: Vagal nerve stimulation prevents acute lung injury following hemorrhagic shock

AimsTrauma/hemorrhagic shock (T/HS) induced gut injury is known to initiate a systemic inflammatory response which can lead to secondary lung injury. We have shown that vagal nerve stimulation (VNS) protects intestinal epithelial integrity after a severe burn insult. We hypothesize that VNS will protect the lung from injury following T/HS by preventing intestinal barrier failure. Main methodsMale Balb/c mice were subjected to a T/HS model with and without cervical VNS. Intestinal injury was evaluated by measuring changes in gut barrier function and tight junction protein localization. Lung injury was evaluated using histology and markers of lung inflammation. Using NF-kB-luciferase (NF-kB-luc) transgenic mice, NF-kb–DNA binding was measured by photon emission analysis at 4 after injury. Key findingsT/HS is associated gut injury characterized by histologic injury, increased epithelial permeability, and altered localization of gut tight junction proteins. Cervical VNS prevented the T/HS-induced changes in gut barrier integrity. Gut injury after T/HS was associated with acute lung injury at 24h characterized by histologic injury, increased number of MPO positive stained cells and MPO enzymatic activity, and increased ICAM-1 expression in lung endothelium. VNS decreased T/HS-induced lung injury with a marked decrease in lung inflammation compared to T/HS alone. Lungs harvested from NF-kB-luc mice at 4h post VNS+T/HS demonstrated decreased DNA binding of NF-kB compared to T/HS alone as measured by changes in bioluminescence. SignificanceVNS is effective in protecting against acute lung injury caused by hemorrhagic shock through its ability to prevent gut barrier dysfunction.

Testosterone Depletion or Blockade in Male Rats Protects Against Trauma Hemorrhagic Shock-Induced Distant Organ Injury by Limiting Gut Injury and Subsequent Production of Biologically Active Mesenteric Lymph

We tested the hypothesis that testosterone depletion or blockade in male rats protects against trauma hemorrhagic shock-induced distant organ injury by limiting gut injury and subsequent production of biologically active mesenteric lymph. Male, castrated male, or flutamide-treated rats (25 mg/kg subcutaneously after resuscitation) were subjected to a laparotomy (trauma), mesenteric lymph duct cannulation, and 90 minutes of shock (35 mm Hg) or trauma sham-shock. Mesenteric lymph was collected preshock, during shock, and postshock. Gut injury was determined at 6 hours postshock using ex vivo ileal permeability with fluorescein dextran. Postshock mesenteric lymph was assayed for biological activity in vivo by injection into mice and measuring lung permeability, neutrophil activation, and red blood cell deformability. In vitro neutrophil priming capacity of the lymph was also tested. Castrated and flutamide-treated male rats were significantly protected against trauma hemorrhagic shock (T/HS)-induced gut injury when compared with hormonally intact males. Postshock mesenteric lymph from male rats had a higher capacity to induce lung injury, Neutrophil (PMN) activation, and loss of red blood cell deformability when injected into naïve mice when compared with castrated and flutamide-treated males. The increase in gut injury after T/HS in males directly correlated with the in vitro biological activity of mesenteric lymph to prime neutrophils for an increased respiratory burst. After T/HS, gut protective effects can be observed in males after testosterone blockade or depletion. This reduced gut injury contributes to decreased biological activity of mesenteric lymph leading to attenuated systemic inflammation and distant organ injury.

Intestinal Mucus Layer Preservation in Female Rats Attenuates Gut Injury After Trauma-Hemorrhagic Shock

We tested the hypothesis that females are more resistant to trauma-hemorrhagic shock (T/HS)-induced gut injury than males, and this is related to better preservation of their intestinal mucus layer, which is influenced in turn by the estrus cycle stage at the time of injury. Male, proestrus and diestrus female rats underwent a laparotomy (trauma) and 90 minutes of shock ( approximately 35 mm Hg). At 3 hours after reperfusion, terminal ileum was harvested and stained with Carnoy's Alcian Blue for mucus assessment, hematoxylin and eosin, and periodic acid schiff for villous and goblet cell morphology and injury. Ileal permeability was measured in separate intestinal segments using the ex vivo everted gut sac technique. When compared with males, proestrus female rats were significantly more resistant to T/HS-induced morphologic gut injury, as reflected in both a lower incidence of villous injury (14% vs. 22%; p < 0.05) and a lesser grade of injury (1.0 vs. 2.8; p < 0.05) as well as preservation of gut barrier function (17.9 vs. 32.2; p < 0.05). This resistance to gut injury was associated with significant preservation of the mucus layer (87% vs. 62%; p < 0.05) and was influenced by the estrus cycle stage of the female rats. There was a significant inverse correlation between mucus layer coverage and the incidence (r = 0.9; p < 0.0001) and magnitude (r = 0.89; p < 0.0001) of villous injury and gut permeability (r = 0.74; p < 0.001). The resistance of female rats to T/HS-induced intestinal injury and dysfunction was associated with better preservation of the intestinal mucus barrier and was to some extent estrus cycle-dependent. Preservation of the mucus barrier may protect against shock-induced gut injury and subsequent distant organ injury by limiting the ability of luminal contents such as bacteria and digestive enzymes from coming into direct contact with the epithelium.

Somatostatin Limits Intestinal Ischemia-Reperfusion Injury in Macaques via Suppression of TLR4-NF-κB Cytokine Pathway

ObjectiveIntestinal ischemia-reperfusion (IIR)-induced gut injury remains a challenge for critically ill patients despite the oxidative stress theory that has been elaborated. This study aimed to test whether Toll-like receptor 4 (TLR4) is involved in gut injury during IIR and whether somatostatin (SST) affects TLR4-nuclear factor-κB (NF-κB) cytokine pathway in the intestinal mucosa of macaques. DesignFifteen macaques were randomized into control, IIR, and SST + IIR groups. Pieces of isolated ileal epithelium from each animal were incubated with lipopolysaccharide (LPS), interferon-γ, or SST. Expression of TLR4 and NF-κBp65 was evaluated by immunohistochemical staining, Western blot analysis and reverse transcription polymerase chain reaction. Cytokine levels were measured by ELISA. Radioimmunoassay was used to determine of SST levels. Measurements and Main ResultsSignificant overexpression (IIR vs control) of ileal TLR4 (0.17±0.03 vs 0.05 ± 0.02), NF-κBp65 (0.55±0.11 vs 0.15 ± 0.05), and TNF-α (213.2±29.2 vs 56.0 ± 10.04) after IIR was greatly decreased (p < 0.05) by prophylactic use of SST (TLR4: 0.06 ± 0.02; NF-κBp65: 0.26 ± 0.09; TNF-α: 97.1 ± 32.3) in vivo. TLR4 expression in the ileal epithelium treated with LPS and SST (1,330 ± 93) was significantly lower than that in the ileal epithelium treated with LPS alone (2,088 ± 126) in vitro. SST levels in plasma (3.67±0.41 ng/ml) and ileal mucosa (1,402.3 ± 160 ng/mg protein) of the IIR group were significantly lower than those (6.09±1.29 ng/ml, 2,234. 8 ± 301.8 ng/mg protein) in the control group (p < 0.05). ConclusionsEndogenous SST is a crucial inhibitor of massive inflammatory injury in the intestinal mucosa via direct suppression of the TLR4-NF-κB cytokine pathway induced by LPS in ileal epithelium. IIR attacks caused shortages of endogenous SST in the plasma and intestinal mucosa of macaques in this study. Therefore, preventive supplements of SST may limit intestinal injury of macaques by IIR.

PANCREATIC PROTEASES ARE NECESSARY FOR INCREASED GUT EPITHELIAL PERMEABILITY DURING TRAUMA/HEMORRHAGIC SHOCK

Shock-induced distant organ injury is associated with increased gut permeability which contributes to gut-origin multiple organ dysfunction syndrome (MODS). Thus, understanding the pathogenesis of trauma- hemorrhagic shock (T/HS)-induced gut injury is important. In this context, we propose the hypothesis that pancreatic proteases play a key role in loss of gut barrier function after T/HS. To test this hypothesis, male Sprague- Dawley rats were subjected to T/HS (MAP 30-35 mmHg × 90 min) with and without pancreatic duct ligation (PDL). After 3 hours of reperfusion, ileal permeability was assessed using FITC-labeled dextran (in-vivo closed loop technique). The magnitude of gut permeability was approximately 3-fold greater in the T/HS rats as compared to T/HS rats with PDL (18.3 ± 4 vs 6.5 ± 5.1 μg/ml P<0.05). Next, we directly tested whether pancreatic proteases increase enterocyte permeability using an in-vitro enterocyte monolayer model. Caco-2 monolayers were incubated for 3 hr or 18 hr with either fetal bovine serum (FBS) or supernatants from homogenized rat pancreas (1:6 w/v PBS). Permeability was assessed using a 10,000 MW rhodamine probe applied to a 3 μm-pore sized transwell cell culture system. We demonstrated a dose-dependent response at both 3hr (r2 = 0.997) and 18 hr (r2 = 0.998) incubation using 5%, 10%, 25% dilutions of initial homogenized pancreas supernatants. Furthermore, we demonstrated that at 18 hr monolayer permeability was approximately 5-fold greater in the cells treated with 25% pancreatic supernatants as opposed to 25 % FBS (16 ± 5 vs 3.3 ± 0.3 % permeability p<0.05). Conclusion: These results support out hypothesis that pancreatic proteases are involved in the pathogenesis of T/HS-induced gut permeability and are able to directly increase enterocyte monolayer permeability.

The Role of Intestinal Flora in Endotoxin (LPS)-Induced Gut Injury and Group II Phospholipase A2 (PLA2-II) Activation in the Small Intestine

The Role of Intestinal Flora in Endotoxin (LPS)-Induced Gut Injury and Group II Phospholipase A 2 (PLA 2 -II) Activation in the Small Intestine