Pathophysiologie der Sepsis

Präsentation zum Thema: "Pathophysiologie der Sepsis"—  Präsentation transkript:

1 Pathophysiologie der Sepsis
Pathophysiologie in Verbindung zu moeglichen therapeutischen Ansaetzen. Anhand ausgewaehlter Beispiele. Sylvia Knapp Klinik f. Innere Medizin 1 - Intensivstation 13i2 - Med. Univ. Wien

2 Infektion, Sepsis & SIRS
This conceptual framework shows the interrelationships between infection, non-infectious disorders, SIRS, sepsis, and severe sepsis. Components of the process not discussed on the following slides include: Infection: a microbial phenomenon characterized by an inflammatory response to the presence of microorganisms or the invasion of normally sterile host tissue by those organisms Bacteremia: the presence of viable bacteria in the blood stream Septic shock: sepsis-induced hypotension despite adequate fluid resuscitation along with the presence of perfusion abnormalities that may include, but are not limited to, lactic acidosis, oliguria, or an acute alteration in mental status Multiple organ dysfunction syndrome (MODS): presence of altered organ function in an acutely ill patient such that homeostasis cannot be maintained without intervention Inflammation and hemostasis are tightly linked. Therefore, although not shown on this slide, sepsis and severe sepsis lie on a background of disturbed hemostasis. Bone RC, Balk RA, Cerra FB, et al. Definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis. Chest. 1992;101: Opal SM, Thijs L, Cavaillon JM, et al. Relationships between coagulation and inflammatory processes. Crit Care Med. 2000;28:S81-2.

3 Pathophysiologie der Sepsis
You have to treat ONE part without impairing the other part!! Riedemann et al. Nature Medicine 2003;9:517

4 Ausgangspunkt- primäre Infektion & Sepsis
Figure 3. Frequency of Sites of Infection Giving Rise to Severe Sepsis in 455 Patients Enrolled in a Trial of Intravenous Ibuprofen for Sepsis. More than 15 percent of the patients had an unknown or uncertain site of primary infection. Data are from Bernard et al.26 Wheeler, N Engl J Med 1999

5 Entzündungsmodulation bei Sepsis
Lokalisierte Infektion Lokale Entzündung unterstützt ‘host defense’ Gestörte bakterielle Clearance Kompensatorische Mechanismen Anti-inflammatorische Zytokine (anti-TNF) Reduktion der systemischen Toxizität Schädlich Fulminante Sepsis Systemische Entzündung erhöht die Mortalität Immunparalyse Mangelhafte Entzündung prädisponiert für nosokomiale Infektionen

6 Potentielle, pathopysiologische Anatzpunkte zur Beeinflussung der Sepsis
Bacteria PRR (e.g. TLRs) Coagulation system Complement Innate immune cells (e.g.C5a) Early inflammatory Mediators (e.g. TNF, IL-1) Late inflammatory Mediators (e.g. HMGB-1) Septic Shock

7 Wie beginnt die Immunantwort / Entzündung?

8 Toll-like Rezeptoren Gram+ Bakterien Gram- Bakterien Viren LTA PGN
Lipopeptid LAM Zymosan Flagellin dsRNA LPS CpG Imiquimod TLR3 CD14 TLR4 ??? TLR5 CD14 TLR9 TLR2 TLR6 TLR7 TLR8 TLR10 TLR1 MyD88 IRAK TRAF6 NFkB Proinflammatorische Zytokine

9 Peptidoglykan – Teichonsäure
Zellwand Gram-positiver Bakterien Toll-like Rezeptor 2 (TLR2) Lipoteichonsäure Peptidoglykan – Teichonsäure Lipoprotein

10 TLR2-blocking Antikörper reduziert die Entzündungsantwort gegen Gram+ Bakterien
control LPS Pam3CSK4 Bacillus subtilis LPS: Lipopolysaccharide LTA: Lipoteichoic Acid Figure 3 Inhibitory effect of mAb T2.5 on cell activation in vitro. (A–D) NF- B–dependent luciferase activities in HEK293 cells overexpressing either murine (A) or human TLR2 (B), as well as TNF- concentrations in supernatants of RAW264.7 (C) or primary murine macrophages (D) challenged with inflammatory agonists. Rel. lucif. activity, relative luciferase activity; ND, not detectable. Cells were incubated with T2.5 or conT2 only (white bars), or additionally challenged with IL-1ß (A and B, light gray bars), ultrapure LPS (C and D, medium gray bars), P3CSK4 (black bars), or h.i. B. subtilis (A–D, dark gray bars). (E) NF- B/p65 nuclear translocation dependent on mAb, P3CSK4 challenge, or LPS challenge in human macrophages was analyzed by cytochemical staining. Unstim., unstimulated. Scale bar: 20 m; magnification was equal for all recordings. (F and G) NF- B–dependent EMSA was analyzed by application of nuclear extracts from RAW264.7 macrophages, and phosphorylation of MAPKs Erk1/2 (pErk1/2), p38 (pP38), and Akt (pAkt) was analyzed by application of total extracts from RAW264.7 macrophages. Cells were preincubated with the indicated amounts of mAb T2.5 or conT2 ( g/ml) and challenged with P3CSK4 or LPS subsequently for 90 minutes (F; arrows indicate specific NF- B–DNA complexes) or 30 minutes (G; phosphorylation-independent p38-specific immunoblot analysis as positive control). Untreated cells were analyzed as controls (Control). NF-B/p65 nuclear translocation Knapp et al. J. Immunol 2004; 172:3132 Meng et al. J. Clin. Invest. 2004; 113:1473

11 Anti-TLR2 verbessert Überleben bei Gram+ septischem Schock
0.5 – 1mg T2.5 -0.5h T2.5 +1h: T2.5 0.25mg T2.5 Wild type TLR2-/- Figure 7 Effects of mAb T2.5 administration on viability after TLR2-specific systemic challenge. (A) IFN- – and D-galactosamine–sensitized mice received no mAb, 1 mg of mAb T2.5, or 1 mg of conT2 i.p. 30 minutes prior to microbial challenge with bacterial lipopeptide analogue P3CSK4 (open circles, no mAb, n = 4; open triangles, mAb conT2, n = 3; filled squares, mAb T2.5, n = 4). (B–D) Mice challenged with a high dose of h.i. B. subtilis were left untreated, treated 1 hour later with the indicated dosages of mAb T2.5 (B; filled diamonds, 1 mg, n = 3; open squares, 0.5 mg, n = 3; open triangles, 0.25 mg, n = 4; ’s, 0.13 mg, n = 4; open circles, no mAb T2.5, n = 4), or treated with 1 mg of mAb’s at the different time points indicated below (C and D). (C) TLR2-specific mAb was administered before (–) or after (+) bacterial challenge (filled inverted triangles, no mAb, n = 8; open circles, mAb conT2, –1 hour, n = 3; filled diamonds, mAb T2.5, –1 hour, n = 4; open squares, mAb T2.5, +1 hour, n = 3; ’s, mAb T2.5, +2 hours, n = 3; open diamonds, mAb T2.5, +3 hours, n = 4; open triangles, mAb T2.5, +4 hours, n = 3). (D) TLR2-specific mAb T2.5 was administered before (–) bacterial challenge (open triangles, no mAb; filled squares, mAb T2.5, –3 hours; open diamonds, mAb T2.5, –4 hours; open circles, mAb T2.5, –5 hours; filled inverted triangles, mAb T2.5, –6 hours; n = 3 for all groups). Knapp et al. J. Immunol 2004; 172:3132 Meng et al. J. Clin. Invest. 2004; 113:1473

12 S. pneumoniae Pneumonie in TLR2-/-: Kein Einfluss auf Überleben
Lunge: Bakterien 6h 48h Knapp et al. J. Immunol 2004; 172:3132

13 Acinetobacter Pneumonie: TLR2 hemmt bakterielle Clearance via reduziertem PMN Einstrom
Increased DC and lung TLR2 expression in postseptic mice (Kunkel). TLR2 high expressing DCs favor a Th2 driven immune response! Wt TLR2-/- Knapp et al. AJRCCM 2006

14 Entzündungsmodulation via TLR2
Kein Einfluss auf bakterielle Clearance Lokalisierte Infektion Lokale Entzündung unterstützt ‘host defense’ Kompensatorische Mechanismen Anti-TLR2 Reduktion der systemischen Toxizität Kein Einfluss Fulminante Sepsis Systemische Entzündung erhöht die Mortalität Immunparalyse Mangelhafte Entzündung prädisponiert für nosokomiale Infektionen

15 Disseminated Intravascular Coagulation (DIC) und Sepsis
Figure 2. Pathogenetic Pathways Involved in Disseminated Intravascular Coagulation. In patients with disseminated intravascular coagulation, fibrin is formed as a result of the generation of thrombin mediated by tissue factor. Tissue factor, expressed on the surface of activated mononuclear cells and endothelial cells, binds and activates factor VII. The complex of tissue factor and factor VIIa can activate factor X directly (black arrows) or indirectly (white arrows) by means of activated factor IX and factor VIII. Activated factor X, in combination with factor V, can convert prothrombin (factor II) to thrombin (factor IIa). Simultaneously, all three physiologic means of anticoagulation -- antithrombin III, protein C, and tissue factor-pathway inhibitor (TFPI) -- are impaired. The resulting intravascular formation of fibrin is not balanced by adequate removal of fibrin because endogenous fibrinolysis is suppressed by high plasma levels of plasminogen-activator inhibitor type 1 (PAI-1). The high levels of PAI-1 inhibit plasminogen-activator activity and consequently reduce the rate of formation of plasmin. The combination of increased formation of fibrin and inadequate removal of fibrin results in disseminated intravascular thrombosis. FDPs denotes fibrin-degradation products. Levi. NEJM 1999; 341:586

16 Konsequenz der DIC Levi M. NEJM 1999; 341:586
Figure 1. The Mechanism of Disseminated Intravascular Coagulation. Systemic activation of coagulation leads to widespread intravascular deposition of fibrin and depletion of platelets and coagulation factors. As a result, thrombosis of small and midsize vessels may occur, contributing to organ failure, and there may be severe bleeding. Levi M. NEJM 1999; 341:586

17 Angriffspunkte von aktiviertem Protein C
Hemmt die Gerinnung APC VIIIa IXa Tissue factor/VIIa TAFI PAI-1 Profibrinolytisch Va Xa Inhibiert ‘rolling’ Inhibition der TNF Produktion APC Thrombin Fibrin Tissue factor/VIIa

18 Long-term survival – APACHE II score
Treatment effect depends on severity of illness n.s. 52.1% 45.6% 41.3% n.s. 33.8% P=.002 Figure 2. Survival for overall cohort. Kaplan-Meier survival curves in PROWESS primary analysis population (Peto-Wilcoxon, p = .10; Mantel-Haenszel, p = .15 for the comparison of drotrecogin alfa [activated] and placebo survival curves). The dashed line is placebo and the solid line is drotrecogin alfa (activated). The curves are truncated at 2½ yrs where the proportion of the overall cohort still alive and in follow-up was 20%, as per recent recommendations (25). The numbers of subjects at risk are indicated below the x-axis. Figure 3. Survival for prespecified subgroups. Kaplan-Meier survival curves in PROWESS subgroups of age (<60 yrs, n = 741 vs. >=60 yrs, n = 949), functional status (independent, n = 1220 vs. dependent, n = 470, as measured by preenrollment ADL score = 0 or not) (32), Acute Physiology and Chronic Health Evaluation (APACHE) II score (<25, n = 873 vs. >=25, n = 817), and degree of organ dysfunction (single, n = 419 vs. multiple, n = 1271). The dashed line is placebo and the solid line is drotrecogin alfa (activated). All four subgroup analyses were prespecified. There was a statistically significant qualitative difference in treatment effect by baseline APACHE II score evaluated dichotomously (<25 or >=25, p = .0008). There were no significant treatment assignment-by-subgroup interactions for age, functional status, or degree of organ dysfunction subgroups (p values ranging from .55 to .91 for all interaction terms introduced into the Cox models). OD, organ dysfunction. P=.001 Significantly improved hospital survival No difference in long-term survival Significantly improved long-term survival when APACHEII >25 at baseline Angus DL et al. Crit Care Med 2004, 32:2199

19 Rh APC improves outcome in Severe Community Acquired Pneumonia (CAP)
80% 68.2% % survival 67.1% 61.2% Figure 6. Survival for community-acquired pneumonia (CAP) by disease severity. Kaplan-Meier survival curves for the comparison of drotrecogin alfa (activated) and placebo. All curves are truncated at 90 days. The black dashed line is placebo and the solid line is drotrecogin alfa (activated) (DrotAA) in the indicated severe CAP subgroup. In A, the gray dashed line is placebo and the solid line is drotrecogin alfa (activated) in PROWESS. PROWESS intention-to-treat data has been previously published (31). n, number of patients in each subgroup; APACHE, Acute Physiology and Chronic Health Evaluation score; PSI, Pneumonia Severity Index. From:   Laterre: Crit Care Med, Volume 33(5).May Table 1. Most frequently observed pathogens in the severe community-acquired pneumonia subgroup From:   Laterre: Crit Care Med, Volume 33(5).May 35,6% mit severe CAP Laterre et al. Crit Care Med, 2005; 33:952

20 Thrombomodulin (und damit aPC) hat keinen Einfluss auf Pneumokokken- oder Klebsiellen-Pneumonie
S. pneumoniae Klebsiella TM is strongly expressed in lung tissue (capillaries) Figure 2. TM does not influence bacterial outgrowth during pneumococcal or Klebsiella pneumonia. (A) S pneumoniae CFUs in lungs of Wt ( ) and TMpro/pro ( ) mice 48 hours after intranasal inoculation. (B) K pneumoniae CFUs in lungs of Wt and TMpro/pro mice 24 hours after intranasal inoculation. Data are mean ± SEM; n = 8 per group per time point. Thrombomodulin (TM): membrane protein that forms complexes with thrombin Thrombin-TM complex converts Protein C to activated Protein C (facilitated by EPCR) TM and EPCR Rijneveld, A. W. et al. Blood 2004;103:1702

21 Verschiedene Erreger …. Sepsis
Figure 1. Methods of identifying bacterial pathogens, from pre–nineteenth century to the present time. (A) A drawing by Louis Joblot in 1718 of "small animals (animalcules)" seen under the microscope. The appearance is reminiscent of protozoa (including one with a human face). Adapted from Reference 14. (B) Culture plate with bacterial growth and Gram's stain of bacteria. (C) Schema of 16S ribosomal DNA bacterial sequencing from bacterial culture or from patient sample. Baron RM et al American Journal of Respiratory Cell and Molecular Biology. Vol. 34, pp , 2006 Pathobiology of Sepsis

22 Genexpression bei Gram+ bzw. Gram- Infektion
Robert J. Feezor,1 Angela Cheng,1,a Heather N. Paddock,1,a Henry V. Baker,1,2 and Lyle L. Moldawer1 Clinical Infectious Diseases    2005;41:S427-S435 Figure 1CITED IN TEXT  |  HIGH-RESOLUTION IMAGE (245 kB)  K-means cluster analysis (heat map) and gene ontologies of leukocytes when ex vivo whole blood is stimulated with either lipopolysaccharide (LPS) or heat-killed Staphylococcus aureus (SAC). A, K-means cluster analysis on 780 genes whose expression significantly changed in response to ex vivo stimulation. Patterns of gene expression could be classified into bins on the basis of the similar and disparate responses to gram-negative and gram-positive pathogens. CON, control. B, Differences in gene expression between the 2 stimuli based on the ontologies for the 780 genes. Gram-positive (Gram+) stimulus (heat-killed S. aureus) preferentially altered the expression of ribosomal and mitochondrial proteins and cell cycle proteins, whereas the gram-negative (Gram-) stimulus altered the expression of genes involved in signal transduction and the immune response. Figure is modified from Feezor et al. [29]. [29] Feezor RJ, Oberholzer C, Baker HV, et al. Molecular characterization of the acute inflammatory response to infections with gram-negative versus gram-positive bacteria. Infect Immun 2003; 71: Feezor et al. Clin Inf Dis 2005, 41: S427

23 Zytokin-Dynamik bei Endotoxämie
IL-1 verlauf ca. wie IL-6, i.e. etwas spaeter peak als TNF 4ng/kg LPS i.v.

24 Genexpressionsprofil in Leukozyten bei Endotoxämie
FIGURE 1. Gene expression profiles of circulating leukocytes in response to bacterial endotoxin infusion. Samples from eight healthy volunteers were tested at baseline (0 h) and 2, 4, 6, 9 and 24 h after intravenous administration of endotoxin (four subjects) or vehicle (four subjects). a, Significant (false discovery rate of <0.1%) probe sets (5,093) were subjected to K-means clustering into ten bins (0–9). Probe sets for which the abundance was above the mean are shown in red, below the mean are shown in blue, and equivalent to the mean are in white. b, Principal component plot of the significant probe sets at the indicated times after endotoxin administration. A network-based analysis of systemic inflammation in humans Steve E. Calvano, Wenzhong Xiao, Daniel R. Richards, Ramon M. Felciano, Henry V. Baker, Raymond J. Cho, Richard O. Chen, Bernard H. Brownstein, J. Perren Cobb, S. Kevin Tschoeke, Carol Miller-Graziano, Lyle L. Moldawer, Michael N. Mindrinos, Ronald W. Davis, Ronald G. Tompkins, Stephen F. Lowry and Inflamm and Host Response to Injury Large Scale Collab. Res. Program Nature 437, (13 October 2005) doi: /nature03985 Calvano SE et al. Nature 2005; 437:1032

25 Genexpressionsprofil in Leukozyten bei Endotoxämie
Figure 2: Pathway analysis of representative genes involved in innate immunity A prototypical inflammatory cell was constructed from 292 representative genes involved in inflammation and innate immunity. Genes for which the expression statistically increased from baseline are coloured red, those for which expression decreased are shown in blue. a, Composite changes in apparent expression over 24 h, identifying nodes and interactions. b, Temporal changes in apparent expression. The response to endotoxin administration in blood leukocytes can be viewed as an integrated cell-wide response, propagating and resolving over time. A network-based analysis of systemic inflammation in humans Steve E. Calvano, Wenzhong Xiao, Daniel R. Richards, Ramon M. Felciano, Henry V. Baker, Raymond J. Cho, Richard O. Chen, Bernard H. Brownstein, J. Perren Cobb, S. Kevin Tschoeke, Carol Miller-Graziano, Lyle L. Moldawer, Michael N. Mindrinos, Ronald W. Davis, Ronald G. Tompkins, Stephen F. Lowry and Inflamm and Host Response to Injury Large Scale Collab. Res. Program Nature 437, (13 October 2005) doi: /nature03985 Calvano SE et al. Nature 2005; 437:1032

26 www.gluegrant.org Pseudomonas (nosokomial) S. aureus Pseudomonas (CF)
Anti-inflammatorische Mediatoren Pseudomonas (CF) E. coli Pro-inflammatorische Mediatoren

27 Zukunft: Individualisierte, Patienten-bezogene Diagnose & Therapie
1. “Bedside tests” Infektion oder SIRS? Phase: Hyper- or Hypoinflammation? Erreger? Grunderkrankung? 2. Anti- or pro-inflammatorische Therapie?