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In JoVE (1)
Other Publications (12)
- Journal of Cellular Physiology
- Langenbeck's Archives of Surgery / Deutsche Gesellschaft Für Chirurgie
- Shock (Augusta, Ga.)
- Critical Care Medicine
- Pflügers Archiv : European Journal of Physiology
- Infection and Immunity
- Respiration; International Review of Thoracic Diseases
- Langenbeck's Archives of Surgery / Deutsche Gesellschaft Für Chirurgie
- Langenbeck's Archives of Surgery / Deutsche Gesellschaft Für Chirurgie
- Critical Care Medicine
- Blood
- Journal of Cellular Physiology
Articles by Wolfram Kessler in JoVE
JoVE Immunology and Infection
Colon Ascendens Stent Peritonitis (CASP) - a Standardized Model for Polymicrobial Abdominal Sepsis
Department of Surgery, University of Greifswald
The Colon Ascendens Stent Peritonitis (CASP) is a highly standardized model for polymicrobial abdominal sepsis in rodents. This article describes the surgical procedure of CASP. The CASP model and its variants allow the systematic investigation of various problems concerning the subject of sepsis.
Other articles by Wolfram Kessler on PubMed
Subcellular Distribution of Calcium-sensitive Potassium Channels (IK1) in Migrating Cells
Cell migration is crucial for wound healing, immune defense, or formation of tumor metastases. In addition to the cytoskeleton, Ca2+ sensitive K+ channels (IK1) are also part of the cellular "migration machinery." We showed that Ca2+ sensitive K+ channels support the retraction of the rear part of migrating MDCK-F cells by inducing a localized shrinkage at this cell pole. So far the molecular nature and in particular the subcellular distribution of these channels in MDCK-F cells is unknown. We compared the effect of IK1 channel blockers and activators on the current of a cloned IK1 channel from MDCK-F cells (cIK1) and the migratory behavior of these cells. Using IK1 channels labeled with a HA-tag or the enhanced green fluorescent protein we studied the subcellular distribution of the canine (cIK1) and the human (hIK1) channel protein in different migrating cells. The functional impact of cIK1 channel activity at the front or rear part of MDCK-F cells was assessed with a local superfusion technique and a detailed morphometric analysis. We show that it is cIK1 whose activity is required for migration of MDCK-F cells. IK1 channels are found in the entire plasma membrane, but they are concentrated at the cell front. This is in part due to membrane ruffling at this cell pole. However, there appears to be only little cIK1 channel activity at the front of MDCK-F cells. In our view this apparent discrepancy can be explained by differential regulation of IK1 channels at the front and rear part of migrating cells.
The Vagal Nerve As a Link Between the Nervous and Immune System in the Instance of Polymicrobial Sepsis
The role of the vagal nerve in the autonomic nervous system is widely well known. Recently, an additional function was revealed serving as a connector between the nervous and immune system. This connection is called the "cholinergic inflammatory pathway." Through stimulation of the acetylcholine receptors located upon the macrophages, the "unspecific" immune system can be directly influenced.
Sepsis Affects Cardiac Expression of Multidrug Resistance Protein 5 (MRP5, ABCC5), an ABC-type CGMP Export Pump
One of the clinical characteristics associated with septic shock is heart failure. Several lines of evidence indicate that functional consequences of heart failure in septic shock are linked to the activated NO-cyclic guanosine monophosphate (NO-cGMP) pathway. We have previously shown that the high-affinity cGMP export transporter, multidrug resistance protein 5 (MRP5), is expressed in the heart, which modulates intracellular concentrations and, hence, the effects of cGMP. Thus, modified expression of cardiac MRP5 in septic shock can alter cGMP concentrations and contribute to the development of heart failure. We therefore investigated MRP5 expression in the heart using two established murine models of septic shock (intraperitoneal LPS injection and surgical implantation of a stent into the ascending colon, resulting in a multibacterial peritonitis [CASP, colon ascendens stent peritonitis] in C57BL/6N mice, respectively; n = 38). Cardiac MRP5 was assessed by quantitative polymerase chain reaction and immunofluorescence. The protein was localized in the endothelial wall, smooth muscle, and cardiac myocytes. MRP5 mRNA expression was significantly reduced compared with controls both in the LPS (31.9 +/- 16.8 x 10(-4) vs. 54.1 +/- 14.8 x 10(-4), P = 0.025) and CASP model (18.3 +/- 9.4 x 10(-4) vs. 42.8 +/- 12.1 x 10(-4), P = 0.009; MRP5/glyceraldehyde 3-phosphate dehydrogenase copy numbers, respectively). In parallel, IL-6 plasma levels were significantly increased in both models. Incubation of cultured murine cardiomyocytes (HL1) with 5 ng/mL IL-6 resulted in decreased expression of MRP5 (54% of control), as did incubation of the cells with serum from septic mice (LPS serum, 22% of control; CASP serum, 11% of control). In conclusion, cardiac expression of the cGMP export transporter MRP5 is decreased in two murine models of septic shock, most likely by a transcriptional mechanism. Reduced cGMP export as a consequence of decreased MRP5 expression can attenuate heart failure in sepsis.
Seasonal Variations in Inflammatory Responses to Sepsis and Stress in Mice
In this study, we analyzed seasonal variations of immunoreactivity using a model of septic shock and a model of immunosuppression induced by chronic stress in mice.
Activation of Cell Migration with Fibroblast Growth Factor-2 Requires Calcium-sensitive Potassium Channels
Tumor cell migration is crucial for the formation of tumor metastases and the progression of tumor disease. Fibroblast growth factor-2 (FGF-2) is one of the cytokines involved in the autocrine stimulation of tumor development. FGF-2 also stimulates transcription of Ca(2+)-sensitive K(+) channels (IK1 or K(Ca)3.1), which are part of the migration machinery in many cell types. Here, we tested whether FGF-2 acutely stimulates migration of transformed MDCK cells in a K(Ca)3.1 channel-dependent way. FGF-2 accelerates migration dose dependently. The speed of migration increases almost instantaneously. After 2 min, ERK1/2 phosphorylation has almost doubled. FGF-2 does not stimulate migration when ERK1/2 phosphorylation is inhibited. K(Ca)3.1 channel blockade also prevents the stimulatory effect of FGF-2 on cell migration. In addition, FGF-2 treatment leads to an activation of K(Ca)3.1 channels and a rapid rise of the cell area, which is because of an elevated rate of exocytosis. However, the amount of K(Ca)3.1 channels within the plasma membrane does not change. Our results show that there is a reciprocal interrelation between FGF-2 and K(Ca)3.1 channels. K(Ca)3.1 channels that are under the transcriptional control of FGF-2 are part of the FGF-2-mediated signaling cascade leading to an acceleration of migration.
Detrimental Role of CC Chemokine Receptor 4 in Murine Polymicrobial Sepsis
CC chemokine receptor 4 (CCR4) and its two ligands, CCL17 and CCL22, are critically involved in different immune processes. In models of lipopolysaccharide-induced shock, CCR4-deficient (CCR4(-/-)) mice showed improved survival rates associated with attenuated proinflammatory cytokine release. Using CCR4(-/-) mice with a C57BL/6 background, this study describes for the first time the role of CCR4 in a murine model of polymicrobial abdominal sepsis, the colon ascendens stent peritonitis (CASP). CASP-induced sepsis led to a massive downregulation of CCR4 in lymphoid and nonlymphoid tissues, whereas the expression of CCL17 and CCL22 was independent of the presence of CCR4. After CASP, CCR4(-/-) animals showed a strongly enhanced bacterial clearance in several organs but not in the peritoneal lavage fluid and the blood. In addition, significantly reduced levels of proinflammatory cytokines/chemokines were measured in organ supernatants as well as in the sera of CCR4(-/-) mice. CCR4 deficiency consequently resulted in an attenuated severity of systemic sepsis and a strongly improved survival rate after CASP or CASP with intervention. Thus, our data provide clear evidence that CCR4 plays a strictly detrimental role in the course of polymicrobial sepsis.
Selective Depletion of Alveolar Macrophages in Polymicrobial Sepsis Increases Lung Injury, Bacterial Load and Mortality but Does Not Affect Cytokine Release
Resident tissue macrophages exert important functions during severe systemic infection and contribute to changes in local as well as systemic immune responses. Alveolar macrophages (AM) play a crucial role in airway diseases and in the defense against microorganisms invading the body via the bronchopulmonary tract. It has been postulated that AM are involved in the development of acute local disorders as a consequence of extrapulmonary stimuli like pancreatitis, peritonitis, or trauma.
Implantation of Alloplastic Material Increases Survival of Mice Subsequently Exposed to Polymicrobial Sepsis
Major surgery can modulate the immune system and by this the clinical course of following complications. Effects of minor surgical treatments on the immune system and septic complications are poorly understood.
Laparoscopic Appendectomy Using a Single Polymeric Clip to Close the Appendicular Stump
The aim of this study was to evaluate the feasibility, morbidity, and cost-effectiveness of the closure of the appendicular stump with a single non-absorbable polymeric clip compared with the closure of the appendicular stump with a stapler.
Tumor Necrosis Factor-related Apoptosis-inducing Ligand (TRAIL) Improves the Innate Immune Response and Enhances Survival in Murine Polymicrobial Sepsis
To investigate the role of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) in postoperative polymicrobial abdominal sepsis.Sepsis is the leading cause of death among critically ill surgical patients. TRAIL is commonly known as an apoptosis-inducing agent in cancer cells. It also plays an important role in the regulation of inflammatory reactions. The role of TRAIL in polymicrobial sepsis is still unclear.
Platelet Factor 4 Binds to Bacteria, [corrected] Inducing Antibodies Cross-reacting with the Major Antigen in Heparin-induced Thrombocytopenia
A clinically important adverse drug reaction, heparin-induced thrombocytopenia (HIT), is induced by antibodies specific for complexes of the chemokine platelet factor 4 (PF4) and the polyanion heparin. Even heparin-naive patients can generate anti-PF4/heparin IgG as early as day 4 of heparin treatment, suggesting preimmunization by antigens mimicking PF4/heparin complexes. These antibodies probably result from bacterial infections, as (1) PF4 bound charge-dependently to various bacteria, (2) human heparin-induced anti-PF4/heparin antibodies cross-reacted with PF4-coated Staphylococcus aureus and Escherichia coli, and (3) mice developed anti-PF4/heparin antibodies during polymicrobial sepsis without heparin application. Thus, after binding to bacteria, the endogenous protein PF4 induces antibodies with specificity for PF4/polyanion complexes. These can target a large variety of PF4-coated bacteria and enhance bacterial phagocytosis in vitro. The same antigenic epitopes are expressed when pharmacologic heparin binds to platelets augmenting formation of PF4 complexes. Boosting of preformed B cells by PF4/heparin complexes could explain the early occurrence of IgG antibodies in HIT. We also found a continuous, rather than dichotomous, distribution of anti-PF4/heparin IgM and IgG serum concentrations in a cross-sectional population study (n = 4029), indicating frequent preimmunization to modified PF4. PF4 may have a role in bacterial defense, and HIT is probably a misdirected antibacterial host defense mechanism.
Dynamic Redistribution of Calcium Sensitive Potassium Channels (hK(Ca)3.1) in Migrating Cells
Calcium-sensitive potassium channels (K(Ca)3.1) are expressed in virtually all migrating cells. Their activity is required for optimal cell migration so that their blockade leads to slowing down. K(Ca)3.1 channels must be inserted into the plasma membrane in order to elicit their physiological function. However, the plasma membrane of migrating cells is subject to rapid recycling by means of endo- and exocytosis. Here, we focussed on the endocytic internalization and the intracellular transport of the human isoform hK(Ca)3.1. A hK(Ca)3.1 channel construct with an HA-tag in the extracellularly located S3-S4 linker was transfected into migrating transformed renal epithelial MDCK-F cells. Channel internalization was visualized and quantified with immunofluorescence and a cell-based ELISA. Movement of hK(Ca)3.1 channel containing vesicles as well as migration of MDCK-F cells were monitored by means of time lapse video microscopy. hK(Ca)3.1 channels are endocytosed during migration. Most of the hK(Ca)3.1 channel containing vesicles are moving at a speed of up to 2 µm/sec in a microtubule-dependent manner towards the front of MDCK-F cells. Our experiments indicate that endocytosis of hK(Ca)3.1 channels is clathrin-dependent since they colocalize with clathrin adaptor proteins and since it is impaired when a C-terminal dileucine motif is mutated. The C-terminal dileucine motif is also important for the subcellular localization of hK(Ca)3.1 channels in migrating cells. Mutated channels are no longer concentrated at the leading edge. We therefore propose that recycling of hK(Ca)3.1 channels contributes to their characteristic subcellular distribution in migrating cells.
