Utility of intestinal intravital microscopy for the study of CNS injury-induced immunodepression syndrome (CIDS)

2021 ◽  
pp. 1-11
Author(s):  
Bashir Bietar ◽  
Juan Zhou ◽  
Christian Lehmann
Keyword(s):  
Intravital Microscopy ◽  
Capillary Density ◽  
Experimental Stroke ◽  
Cns Injury ◽  
Capillary Perfusion ◽  
Local Inflammatory Response ◽  
Lps Challenge ◽  
Intestinal Microcirculation ◽  
Secondary Damage ◽  
Immune Depression

BACKGROUND: Stroke, traumatic brain injury, or other forms of central nervous system (CNS) injury initiate a local inflammatory response. Compensatory anti-inflammatory pathways are activated to limit secondary damage due to inflammation. The associated release of immunosuppressing neuromodulators can result in system-wide immune dysregulation (CNS injury-induced immune-depression syndrome –CIDS). OBJECTIVE: To establish an experimental stroke model where CIDS can be studied by intravital microscopy (IVM). METHODS: We used the photothrombotic stroke (PTS) model in C57BL/6 mice and studied its effects on peripheral immunity following challenge with lipopolysaccharide (LPS). Leukocyte activation, as well as capillary perfusion of the microcirculation, were assessed using intestinal intravital microscopy (IVM). RESULTS: PTS caused a significant reduction in the number of adhering leukocytes in submucosal venules of the terminal ileum of mice challenged with LPS compared to LPS-challenged animals without stroke. Leukocyte rolling was also impacted by PTS in the submucosal venules. Following stroke, we also observed decreased mucosal functional capillary density (FCD). CONCLUSIONS: Our results suggest that PTS with subsequent LPS challenge poses as a viable model to further study CIDS using intravital microscopy of the intestinal microcirculation.

Argatroban administration reduces leukocyte adhesion and improves capillary perfusion within the intestinal microcirculation in experimental sepsis

2010 ◽  
Vol 104 (11) ◽  
pp. 1022-1028 ◽  
Author(s):  
Christian Fuchs ◽  
Elena Ladwig ◽  
Juan Zhou ◽  
Dragan Pavlovic ◽  
Kristina Behrend ◽  
...  
Keyword(s):  
Leukocyte Adhesion ◽  
Capillary Density ◽  
Functional Capillary Density ◽  
Experimental Sepsis ◽  
Capillary Perfusion ◽  
Vascular Contractility ◽  
Intestinal Microcirculation ◽  
Sham Surgery ◽  
The Impact

SummaryCo-activation of pro-coagulatory pathways in sepsis may result in disseminated intravascular coagulation and contributes to microvascular dysfunction. We investigated the effects of the direct thrombin inhibitor, argatroban (ARG), on the sepsis-induced impairment of the intestinal microcirculation (capillary perfusion, leukocyte adhesion) and the vascular contractility in rats. Forty male Lewis rats were randomly assigned to one of four groups: sham surgery (SHAM), experimental sepsis (colon ascendens stent peritonitis – CASP), CASP+ARG, and SHAM+ARG. At 16 hours after colon stent insertion (or sham surgery), 2 mg/kg argatroban or buffer were given intravenously, and 1 hour thereafter, intravital microscopy was performed. In addition, experiments to study the impact of ARG on vascular contractility were conducted in vitro. ARG administration in CASP rats significantly increased functional capillary density in mucosal (+128%) and muscular layers (longitudinal: +42%; circular: +64%) and decreased the number of firmly adhering leukocytes in the intestinal submucosa compared to untreated animals. In vitro findings indicated a vasodilating effect of ARG. ARG administration during experimental sepsis improved intestinal microcirculation by preserving functional capillary density, an indicator of microvascular perfusion, and by reducing leukocyte adherence to the endothelium in submucosal venules.

Dexmedetomidine Attenuates the Microcirculatory Derangements Evoked by Experimental Sepsis

2015 ◽  
Vol 122 (3) ◽  
pp. 619-630 ◽  
Author(s):  
Marcos L. Miranda ◽  
Michelle M. Balarini ◽  
Eliete Bouskela
Keyword(s):  
Blood Gases ◽  
Capillary Density ◽  
Sepsis Syndrome ◽  
In Vivo Studies ◽  
Experimental Sepsis ◽  
Leukocyte Rolling ◽  
Capillary Perfusion ◽  
Arterial Blood ◽  
Baseline Values

Abstract Background: Dexmedetomidine, an α-2 adrenergic receptor agonist, has already been used in septic patients although few studies have examined its effects on microcirculatory dysfunction, which may play an important role in perpetuating sepsis syndrome. Therefore, the authors have designed a controlled experimental study to characterize the microcirculatory effects of dexmedetomidine in an endotoxemia rodent model that allows in vivo studies of microcirculation. Methods: After skinfold chamber implantation, 49 golden Syrian hamsters were randomly allocated in five groups: (1) control animals; (2) nonendotoxemic animals treated with saline; (3) nonendotoxemic animals treated with dexmedetomidine (5.0 μg kg−1 h−1); (4) endotoxemic (lipopolysaccharide 1.0 mg/kg) animals treated with saline; and (5) endotoxemic animals treated with dexmedetomidine. Intravital microscopy of skinfold chamber preparations allowed quantitative analysis of microvascular variables and venular leukocyte rolling and adhesion. Mean arterial blood pressure, heart rate, arterial blood gases, and lactate concentrations were also documented. Results: Lipopolysaccharide administration increased leukocyte rolling and adhesion and decreased capillary perfusion. Dexmedetomidine significantly attenuated these responses: compared with endotoxemic animals treated with saline, those treated with dexmedetomidine had less leukocyte rolling (11.8 ± 7.2% vs. 24.3 ± 15.0%; P < 0.05) and adhesion (237 ± 185 vs. 510 ± 363; P < 0.05) and greater functional capillary density (57.4 ± 11.2% of baseline values vs. 45.9 ± 11.2%; P < 0.05) and erythrocyte velocity (68.7 ± 17.6% of baseline values vs. 54.4 ± 14.8%; P < 0.05) at the end of the experiment. Conclusions: Dexmedetomidine decreased lipopolysaccharide-induced leukocyte–endothelial interactions in the hamster skinfold chamber microcirculation. This was accompanied by a significant attenuation of capillary perfusion deficits, suggesting that dexmedetomidine yields beneficial effects on endotoxemic animals’ microcirculation.

Microvascular ischemia-reperfusion injury in striated muscle: significance of "no reflow"

1992 ◽  
Vol 263 (6) ◽  
pp. H1892-H1900 ◽  
Author(s):  
M. D. Menger ◽  
D. Steiner ◽  
K. Messmer
Keyword(s):  
Shear Rate ◽  
Striated Muscle ◽  
Ischemia Reperfusion ◽  
Reactive Oxygen ◽  
Capillary Density ◽  
Wall Shear Rate ◽  
Reactive Oxygen Metabolites ◽  
Capillary Perfusion ◽  
No Reflow ◽  
Oxygen Metabolites

“No reflow” has been implicated as prominent phenomenon in microvascular injury associated with ischemia-reperfusion (I/R). The objectives of this study were 1) to elucidate the significance of no reflow in microvascular I/R injury of striated muscle and 2) to determine whether reactive oxygen metabolites play a role in the development of postischemic no reflow. By use of the hamster dorsal skinfold preparation and intravital microscopy, microvascular perfusion of capillaries and postcapillary venules of striated muscle was quantitatively assessed before and 30 min, 2 h, and 24 h after 4 h of tourniquet-induced ischemia. I/R was characterized by a significant reduction (P < 0.01) in functional capillary density to 35% of baseline values during initial reperfusion, with incomplete recovery after 24 h (n = 9). In addition, capillary perfusion was found to be extremely heterogeneous, and wall shear rate in postcapillary venules was significantly decreased (P < 0.01). Treatment with either superoxide dismutase (SOD; n = 9) or allopurinol (n = 9) resulted in maintenance of capillary density of 60% of baseline (P < 0.05). Furthermore, I/R-induced capillary perfusion inhomogeneities and decrease of wall shear rate in venules were attenuated significantly (P < 0.01) by SOD and allopurinol. Thus part of capillary perfusion disturbances during I/R in striated muscle may be caused by increased postcapillary vascular resistance, probably mediated by reactive oxygen metabolites. However, the fact that in SOD- and allopurinol-treated animals 40% of the capillaries were still found to be nonperfused indicates that mechanisms other than oxygen radicals play an important role in the development of postischemic no reflow.

scholarly journals Imaging of the Intestinal Microcirculation during Acute and Chronic Inflammation

Biology ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 418
Author(s):  
Kayle Dickson ◽  
Hajer Malitan ◽  
Christian Lehmann
Keyword(s):  
Blood Flow ◽  
Multiorgan Failure ◽  
Capillary Density ◽  
Intestinal Microcirculation ◽  
Inflammatory Conditions ◽  
Microcirculatory Disturbances ◽  
Vessel Integrity ◽  
Irritable Bowel ◽  
Acute And Chronic Inflammation

Because of its unique microvascular anatomy, the intestine is particularly vulnerable to microcirculatory disturbances. During inflammation, pathological changes in blood flow, vessel integrity and capillary density result in impaired tissue oxygenation. In severe cases, these changes can progress to multiorgan failure and possibly death. Microcirculation may be evaluated in superficial tissues in patients using video microscopy devices, but these techniques do not allow the assessment of intestinal microcirculation. The gold standard for the experimental evaluation of intestinal microcirculation is intravital microscopy, a technique that allows for the in vivo examination of many pathophysiological processes including leukocyte-endothelial interactions and capillary blood flow. This review provides an overview of changes in the intestinal microcirculation in various acute and chronic inflammatory conditions. Acute conditions discussed include local infections, severe acute pancreatitis, necrotizing enterocolitis and sepsis. Inflammatory bowel disease and irritable bowel syndrome are included as examples of chronic conditions of the intestine.

Plasma viscosity regulates capillary perfusion during extreme hemodilution in hamster skinfold model

1998 ◽  
Vol 275 (6) ◽  
pp. H2170-H2180 ◽  
Author(s):  
Amy G. Tsai ◽  
Barbara Friesenecker ◽  
Michael McCarthy ◽  
Hiromi Sakai ◽  
Marcos Intaglietta
Keyword(s):  
Molecular Weight ◽  
Shear Stress ◽  
Capillary Density ◽  
High Viscosity ◽  
Plasma Viscosity ◽  
Capillary Perfusion ◽  
Low Viscosity ◽  
Arterial Blood ◽  
Wall Shear ◽  
Stress Dependent

Effect of increasing blood viscosity during extreme hemodilution on capillary perfusion and tissue oxygenation was investigated in the awake hamster skinfold model. Two isovolemic hemodilution steps were performed with 6% Dextran 70 [molecular weight (MW) = 70,000] until systemic hematocrit (Hct) was reduced by 65%. A third step reduced Hct by 75% and was performed with the same solution [low viscosity (LV)] or a high-molecular-weight 6% Dextran 500 solution [MW = 500,000, high viscosity (HV)]. Final plasma viscosities were 1.4 and 2.2 cP (baseline of 1.2 cP). Hct was reduced to 11.2 ± 1.1% from 46.2 ± 1.5% for LV and to 11.9 ± 0.7% from 47.3 ± 2.1% for HV. HV produced a greater mean arterial blood pressure than LV. Functional capillary density (FCD) was substantially higher after HV (85 ± 12%) vs. LV (38 ± 30%) vs. baseline (100%).[Formula: see text] levels measured with Pd-porphyrin phosphorescence microscopy were not statistically changed from baseline until after the third hemodilution step. Wall shear rate (WSR) decreased in arterioles and venules after LV and only in arterioles after HV. Wall shear stress (WSR × plasma viscosity) was substantially higher after HV vs. LV. Increased mean arterial pressure and shear stress-dependent release of endothelium-derived relaxing factor are possible mechanisms that improved arteriolar and venular blood flow and FCD after HV vs. LV exchange protocols.

Microvascular perfusion deficits are not a prerequisite for mucosal injury in septic rats

1999 ◽  
Vol 276 (4) ◽  
pp. G933-G940 ◽  
Author(s):  
R. R. Nevière ◽  
M. L. Pitt-Hyde ◽  
R. D. Piper ◽  
W. J. Sibbald ◽  
R. F. Potter
Keyword(s):  
Cell Injury ◽  
Mucosal Injury ◽  
Endotoxin Tolerance ◽  
Capillary Density ◽  
Microvascular Perfusion ◽  
Myeloperoxidase Activity ◽  
Capillary Perfusion ◽  
Ileal Mucosa ◽  
Mucosal Permeability ◽  
Perfusion Deficits

Our major objective was to investigate whether injury to the mucosa of the small intestine occurred in a normotensive model of sepsis and whether such injury was associated with microvascular perfusion deficits. Using fluorescence intravital microscopy, we show direct evidence of cell injury within the mucosa (pneumonia 12.4 ± 2.6 cells/field, sham 2.2 ± 0.7 cells/field), whereas use of51Cr-labeled EDTA showed evidence of increased mucosal permeability (pneumonia 1.90 ± 0.67 ml · min−1 · 100 g−1; sham 0.24 ± 0.04 ml · min−1 · 100 g−1), 48 h following induction of pneumonia. Despite such injury the capillary density in the ileal mucosa and submucosa of pneumonic rats (1,027 ± 77 and 1,717 ± 86 mm2) was not significantly different compared with sham (998 ± 63 and 1,812 ± 101 mm2). However, a modest albeit significant decrease in capillary perfusion was measured in the muscularis layer of pneumonia (11.0 ± 1.3 mm) compared with sham (13.9 ± 0.63 mm) and appeared to be associated with leukocyte entrapment. Pretreatment using low doses of endotoxin to induce endotoxin tolerance not only increased muscularis capillary density but reduced the number of leukocytes trapped within the microvasculature, decreased myeloperoxidase activity within the ileum in pneumonic rats, and prevented mucosal injury. In conclusion, we have shown that pneumonia results in remote injury to the mucosa of the ileum and that such injury was not associated with concurrent mucosal perfusion deficits.

Hemorrhagic hypotension induces arteriolar vasomotion and intermittent capillary perfusion in rat pancreas

1994 ◽  
Vol 267 (5) ◽  
pp. H1936-H1940 ◽  
Author(s):  
B. Vollmar ◽  
G. Preissler ◽  
M. D. Menger
Keyword(s):  
Blood Cell ◽  
Microscopic Analysis ◽  
Capillary Density ◽  
Functional Capillary Density ◽  
Intravital Fluorescence Microscopy ◽  
Capillary Perfusion ◽  
Induced Hypotension ◽  
Rat Pancreas ◽  
Hemorrhagic Hypotension ◽  
Acinar Tissue

Hemorrhage-induced intermittent capillary perfusion and its relation to arteriolar vasomotion was studied in rat pancreatic acinar tissue using intravital fluorescence microscopy. During prehemorrhage conditions, microscopic analysis of the pancreatic microcirculation displayed neither arteriolar vasomotion nor intermittency of capillary perfusion (n = 22 animals). Hemorrhage-induced hypotension of 40 mmHg provoked arteriolar vasomotion in 18 of 22 animals and 59 of 115 arterioles studied. The maximum relative amplitude of arteriolar vasomotion was 44 +/- 8% (range 12–81%), and vasomotion frequency averaged 4.73 +/- 0.11 cycles/min. Hemorrhagic hypotension was further accompanied by 1) a decrease of functional capillary density [length of red blood cell-perfused capillaries per area of tissue under investigation (cm/cm2)] from 515 +/- 3 cm-1 at baseline to 386 +/- 3 cm-1 (P < 0.05) and 2) the instantaneous occurrence of intermittency of capillary perfusion in all observation areas (N = 220) of the 22 animals studied. The frequency of intermittency of capillary perfusion (4.72 +/- 0.14 cycles/min) did not differ from the frequency of arteriolar vasomotion, which implies a causal relationship between these two hemorrhage-induced microvascular mechanisms with the probable aim to counteract the decrease of functional capillary density.

scholarly journals Imaging the injured beating heart intravitally and the vasculoprotection afforded by haematopoietic stem cells

2019 ◽  
Vol 115 (13) ◽  
pp. 1918-1932 ◽  
Author(s):  
Dean P J Kavanagh ◽  
Adam B Lokman ◽  
Georgiana Neag ◽  
Abigail Colley ◽  
Neena Kalia
Keyword(s):  
Blood Flow ◽  
Progenitor Cells ◽  
Intravital Microscopy ◽  
Causative Factor ◽  
Capillary Density ◽  
Beating Heart ◽  
Laser Speckle ◽  
Microvascular Perfusion ◽  
Contrast Imaging

Abstract Aims Adequate microcirculatory perfusion, and not just opening of occluded arteries, is critical to salvage heart tissue following myocardial infarction. However, the degree of microvascular perfusion taking place is not known, limited primarily by an inability to directly image coronary microcirculation in a beating heart in vivo. Haematopoietic stem/progenitor cells (HSPCs) offer a potential therapy but little is known about their homing dynamics at a cellular level and whether they protect coronary microvessels. This study used intravital microscopy to image the anaesthetized mouse beating heart microcirculation following stabilization. Methods and results A 3D-printed stabilizer was attached to the ischaemia–reperfusion injured (IRI) beating heart. The kinetics of neutrophil, platelet and HSPC recruitment, as well as functional capillary density (FCD), was imaged post-reperfusion. Laser speckle contrast imaging (LSCI) was used for the first time to monitor ventricular blood flow in beating hearts. Sustained hyperaemic responses were measured throughout reperfusion, initially indicating adequate flow resumption. Intravital microscopy confirmed large vessel perfusion but demonstrated poor transmission of flow to downstream coronary microvessels. Significant neutrophil adhesion and microthrombus formation occurred within capillaries with the latter occluding them, resulting in patchy perfusion and reduced FCD. Interestingly, ‘patrolling’ neutrophils were also observed in capillaries. Haematopoietic stem/progenitor cells readily trafficked through the heart but local retention was poor. Despite this, remarkable anti-thromboinflammatory effects were observed, consequently improving microvascular perfusion. Conclusion We present a novel approach for imaging multiple microcirculatory perturbations in the beating heart with LSCI assessment of blood flow. Despite deceptive hyperaemic responses, increased microcirculatory flow heterogeneity was seen, with non-perfused areas interspersed with perfused areas. Microthrombi, rather than neutrophils, appeared to be the major causative factor. We further applied this technique to demonstrate local stem cell presence is not a pre-requisite to confer vasculoprotection. This is the first detailed in vivo characterization of coronary microcirculatory responses post-reperfusion injury.

Anti-inflammatory iron chelator, DIBI, reduces leukocyte-endothelial adhesion and clinical symptoms of LPS-induced interstitial cystitis in mice

2021 ◽  
Vol 79 (3) ◽  
pp. 395-406
Author(s):  
Georg Hagn ◽  
Bruce Holbein ◽  
Juan Zhou ◽  
Christian Lehmann
Keyword(s):  
Interstitial Cystitis ◽  
Intravital Microscopy ◽  
Iron Chelator ◽  
Capillary Density ◽  
Control Group ◽  
Pain Response ◽  
Leukocyte Activation ◽  
Gram Negative ◽  
Pain And Suffering ◽  
Adherent Leukocytes

BACKGROUND: Interstitial cystitis (IC) is a prevalent and debilitating chronic inflammatory disease of the urinary bladder. Currently there are no fully effective therapeutic agents available, in part due to the still obscure pathogenesis of IC. Lipopolysaccharide (LPS) also known as endotoxin from Gram negative bacteria elicits IC in mice and has formed the basis of model systems for investigation. Excess free iron plays an important role in inflammation through generation of reactive oxygen species (ROS). The novel iron chelator DIBI has been shown to sequester excess free iron and dampen excess inflammatory responses to systemic LPS administration and also to Gram negative bacterial infections. OBJECTIVE: The overall objective of this study was to evaluate the effects of DIBI on LPS induced IC in mice. Leukocyte activation, endothelial adhesion and functional capillary density were assessed by intravital microscopy of the bladder microcirculation following a single intravesical LPS administration with or without intravesical DIBI treatment. Clinical IC symptoms were also assessed through behavioral and pain threshold force measurements. METHODS: Four groups of female BALB/c mice (n = 5–6/group) were randomized in this study: control group, IC group without therapy, IC group with DIBI therapy and control group with DIBI therapy. The groups were examined using intravital microscopy (IVM) of the bladder for leukocyte-endothelial interactions (adherent leukocytes, temporarily interacting leukocytes) and functional capillary density (FCD). A modified behavioral score by Boucher et al. and Von-Frey-Aesthesiometry were used to evaluate key behavioral indices related to pain and visceral pain perception. RESULTS: LPS introduced intravesically induced an early (≤2h) inflammation of the bladder evidenced by leukocyte activation and adhesion to bladder capillary walls. Intravesical DIBI therapy of mice 30min following LPS administration and assessed after 1.5h treatment showed a significant decrease in the number of adherent leukocytes compared to IC animals without DIBI treatment. DIBI treated mice showed a significantly lowered increase in behavioral distress scores compared to IC mice without therapy. Untreated IC mice exhibited a significantly decreased threshold force value for evoked pain response and DIBI treatment improved the threshold pain response. A significant inverse correlation was found for the two pain and suffering evaluation methods results. CONCLUSION: DIBI reduced inflammatory endothelial leukocyte adhesion and key indices related to pain and suffering over those observed in untreated IC mice. Our findings suggest a potential therapeutic role for DIBI for IC treatment.

Regional transcapillary albumin exchange in rodent endotoxaemia: effects of fluid resuscitation and inhibition of nitric oxide synthase

2000 ◽  
Vol 100 (1) ◽  
pp. 81-89 ◽  
Author(s):  
Suveer SINGH ◽  
Peter B. ANNING ◽  
C. Peter WINLOVE ◽  
Timothy W. EVANS
Keyword(s):  
Nitric Oxide ◽  
Skeletal Muscle ◽  
Surface Area ◽  
Blood Volume ◽  
Fluid Resuscitation ◽  
Volume Ratio ◽  
Capillary Density ◽  
Microvascular Permeability ◽  
Capillary Perfusion ◽  
Dual Isotope

Sepsis is characterized by increased microvascular permeability and regional variations in capillary perfusion, which may be modulated by nitric oxide (NO) and reversed by fluid resuscitation (FR). The effects of saline FR and NO synthase blockade [by NG-nitro-L-arginine methyl ester (L-NAME)] on microvascular albumin transport and perfused capillary density were assessed in anaesthetized Wistar rats with acute normodynamic endotoxaemia. Separate dual-isotope techniques were employed to measure the permeability index (PIA) and the permeability×surface area product index (PIB), which provide different and complementary information regarding blood–tissue albumin exchange. PIA represents the tissue/blood distribution volume ratio of albumin. PIB is a composite measure of endothelial permeability and the vascular surface area available for albumin exchange, and therefore takes into account the effect of altered blood volume. Capillary density was quantified by fluorescence microscopy following circulation of Evans Blue-labelled albumin. Compared with controls, PIA was reduced significantly in lipopolysaccharide (LPS)-treated animals in skeletal muscle and skin, probably due to blood volume redistribution rather than to changes in permeability. PIB was increased significantly in LPS-treated animals in the kidney, mesentery, skeletal muscle, skin and lung, and in the small bowel following FR. FR also improved the LPS-induced metabolic base deficit, but did not alter capillary density. L-NAME significantly attenuated the LPS-induced rise in PIB in the lung. In conclusion, acute endotoxaemia induces tissue-dependent variations in microvascular albumin exchange. FR improves acid–base disturbance in endotoxaemia, through mechanisms other than microvascular recruitment. NO appears to increase microvascular permeability in endotoxaemia, an effect that may be attenuated by L-NAME, particularly in the lung.

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