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1 Oxygen therapy reduces secondary hemorrhage after thrombolysis in thromboembolic cerebral ischemia Li Sun, Wei Zhou, Christian Mueller, Clemens Sommer, Sabine Heiland, Alexander T Bauer, Hugo H Marti and Roland Veltkamp Journal of Cerebral Blood Flow & Metabolism 30, 16511660, 2010 Impact factor 5.478 Slide 2 2 Introduction Slide 3 3 Stroke Stroke is a brain injury. It occurs when the brain's blood supply is interrupted. Without oxygen and nutrients from blood, brain tissue dies quickly (less than 10 minutes). This causes a sudden function loss. Slide 4 4 Hemorrhagic stroke V.S. Ischemic stroke Hemorrhagic StrokeIschemic Stroke Hemorrhage/ blood leaks into brain tissue Clot stops blood supply to an area of the brain Slide 5 5 Chan P.H., 2001 Slide 6 6 The activation and proteolytic activity of matrix metalloproteinases (MMPs), particularly MMP-9, are key factors in the proteolytic disruption of the basal lamina and tight junctions of the BBB. Hawkins and Davis, 2005; Wang et al., 2003. Slide 7 7 Therapy for acute ischemic stroke can be approached in two basic ways: First, by an attempt to restore or improve blood flow in an occluded vascular territory. Nighoghossian and Trouillas. 1997 Therapy for acute ischemic stroke Second, via therapy directed at the cellular and metabolic targets. Tissue plasminogen activator (t-PA) should be given within the first 3 hours after the first symptoms of stroke start. Slide 8 8 Tissue plasminogen activator (tPA) Jaspreet Kaur, et al., 2004 (C)Beneficial effects of tPA are successful thrombolysis and restoration of CBF. (D) The deleterious effects come into play through the NMDA receptors, upregulation of MMPs, enhanced accumulation of PMNLs, and free radicals, which results in exacerbation of ischemic injury via excito-neurotoxicity, edema, and hemorrhage. Slide 9 9 HBO HBO might be more effective in stroke within the first few hours and at a pressure of 23 ATA. HBO is to increase the solubility of oxygen in plasma to a level sufficient to support tissues with minimal extraction of oxygen carried on hemoglobin. Ann K et al. 2005 Slide 10 10 John H. Zhang et al. 2005 Slide 11 11 Hyperbaric oxygen (HBO) and normobaric hyperoxia (NBO) attenuated BBB permeability, edema and do not increase oxidative stress after focal and global ischemia. (Mink and Dutka, 1995; Singhal et al, 2002; Veltkamp et al., 2005a) NBO and HBO attenuates early BBB disruption and inhibition of MMP-9 mediated occludin degradation is an important mechanism for this protection. (Liu et al., 2009) Slide 12 12 AIMS Examine the differences in the effectiveness of oxygen therapy on postischemic BBB damage and secondary hemorrhage after thrombolysis. Slide 13 13 Materials and Methods Slide 14 14 Brain Ischemia thrombin-induced thromboemboli (TT-tMCAO) calcium-rich thromboemboli (CT-tMCAO) Toomey et al., 2002 It was allowed to coagulate spontaneously for 2 hours at 37 . It was exposed to a 20 mmol/L calcium solution for 1 minute. Mixed with 1.0 National Institutes of Health (NIH) units of human thrombin and 5 L of 1 mol/L CaCl 2 Twelve thrombi each 0.35mm in diameter and 1.5mm in length Slide 15 15 Spontaneously hypertensive Rat (300-350g) TT-tMCAO CT-tMCAO 60 minutes air 100% O 2 at ambient pressure (NBO)100% O 2 at 3 bar (HBO) 60 minutes Gelatin ZymographyMRI assay T1W, T2* DWI, PWI MMP-9 and MMP-2 Physiology parameter MABP,heart rate, blood gas Hemoglobin Assay rt-PA (9mg/kg) Histologic staining 30 minutes Slide 16 16 1. MRI assay PWI document the perfusion status after thrombolysis and at the end of the experiment Lesion volume was quantified in Diffusion-weighted imaging (DWI) brain sections at 2.5 and 24h after tMCAO. tMCAO 1hr Air HBO NBO 0hr 24hr 2.5 hr 2hr rt-PA DWI, PWI Slide 17 17 1. MRI assay tMCAO 1hr Air HBO NBO 0hr 24hr 2.5 hr 2hr rt-PA T1w, T2* Postischemic BBB damage was analyzedon postcontrast T1w images. T2* MR imaging was used to detect macroscopic intracerebral hemorrhage. Slide 18 18 2. Spectrophotometric Hemoglobin and Gelatin Zymography Assay Spectrophotometric assay the hemoglobin content of brains. Gelatin zymography protein expression of MMP-2 and MMP-9 at ischemic and nonischemic hemispheres. tMCAO 1hr Air HBO NBO 0hr 24hr 2hr rt-PA Hemoglobin content MMP-2 and MMP-9 Slide 19 19 3. Histologic staining- secondary hemorrage Coronal cryosections the secondary hemorrage of brains was compared with T2* on corresponding sections. Erythrocytic extravasations assess the severity of erythrocytic extravasations 24hr after CT-tMCAO. tMCAO 1hr1hr Air HBO NBO 0hr0hr 24hr 2hr2hr rt-PA coronal cryosections Erythrocytic extravasations Slide 20 20 Results Slide 21 21 Slide 22 22 Figure 1. Hyperintense lesion volumes on magnetic resonance diffusion-weighted images at 2.5 and 24 hours after thromboembolic middle cerebral artery occlusion (tMCAO) (mm3). TT-tMCAO CT-tMCAO MRI-DWI (Lesion volume) Slide 23 23 In TT-tMCAO, NBO and HBO significantly reduced lesion volume on DWI compared with air. In CT-tMCAO, only a transient trend toward reduced lesion volume was detected in the HBO group at 2.5 hours but no differences were seen at 24 hours after tMCAO. Oxygen therapy can reduce lesion volume after thrombolysis. SUMMARY 1 Slide 24 24 Figure 2. Volume of enhancement on postcontrast T1w magnetic resonance images 2.5 and 24 hours after thromboembolic middle cerebral artery occlusion (tMCAO) (mm3). MRI-T1w (BBB damage) TT-tMCAO CT-tMCAO Slide 25 25 In TT-tMCAO, HBO significantly reduced postischemic BBB damage on T1w images. In CT-tMCAO, HBO also significantly reduced postischemic, at 24 hours after tMCAO, whereas NBO failed to attenuate the enhancing volume on T1w Images. Hyperbaric oxygen therapy significantly reduces BBB damage after thrombolysis. SUMMARY 2 Slide 26 26 TT-tMCAO Figure 3. Correspondence of bloodbrain barrier damage, infarct lesion, and secondary hemorrhage. T1w T2* cryosections BBB damage intracerebral hemorrhage 2.5hr24hr Slide 27 27 Figure 4. Multimodal magnetic resonance imaging images showing the topography of the parenchymal infarct (diffusion weighted imaging (DWI) at 24 hours), bloodbrain barrier permeability (postcontrast T1w at 2.5 hours), and hemorrhage (T2* at 24 hours). 24 hr 2.5 hr 24 hr Hemorrhage BBB damage Lesion volume TT-tMCAO MRI assay Slide 28 28 Figure 5. Erythrocytic extravasation on trichrome-stained coronal brain sections at the level of the bregma +0.26mm (anterior commissure) 24 hours after calcium-induced thromboemboli-middle cerebral artery occlusion without recanalization. CT-tMCAO 24hr Ischemic striatum (score 4) Contralateral cortex without erythrocytic extravasation Ischemic cortex (score 2) Histologic staining- erythrocytic extravasation Slide 29 29 Figure 6. Hemoglobin spectrophotometry of perfused ischemic brain hemisphere after thromboembolic middle cerebral artery occlusion (tMCAO). Hemoglobin content Slide 30 30 Figure 7. Expression of matrix metalloproteinase (MMP)-2 and MMP-9 24 hours after thromboembolic middle cerebral artery occlusion (tMCAO). TT-tMCAO 24hr Expression of MMP-2 and MMP-9 MMP-9 MMP-2 Slide 31 31 Thus, both NBO and HBO induced a significant reduction in macroscopic hemorrhage on T2* MR image. The area of hypointense T2* signal at 24 hours after MCAO was located within the area of intense postcontrast enhancement on T1w images at 2.5 hours after MACO. In TT-t MCAo and CT-tMCAO, HBO significantly decreased the mean hemoglobin volume. 1.Early increase in BBB permeability appeared to indicate a risk for later secondary hemorrhage. 2.HBO significantly reduces infarct volume, BBB permeability, hemorrhage and MMP-9 activity after thrombolysis. SUMMARY 3 Slide 32 32 Conclusion Slide 33 33 Oxygen therapy can decrease infarct size and BBB damage after thromboembolic ischemia and reduce postthrombolytic intracerebral hemorrhage. Slide 34 34 Thank you for attention Slide 35 35 Oxygen therapy in combination with thrombolytic therapy only affects infarct size if recanalization is successful. Oxygen therapy reduce size and frequency of gross parenchymal hemorrhage after thrombolysis- induced reperfusion. HBO and NBO reduce early BBB permeability after tMCAO, which is a marker for subsequent hemorrhagic complications of thrombolysis. Oxygen therapy improves microvascular integrity even in regions that undergo parenchymal infarction Slide 36 36 (cerebral thrombosis) (coagulation) (cerebral embolism) cerebral thrombosis cerebral embolism Slide 37 37 1. 2. Slide 38 38 1. (anticoagulant drugs) 2. (thrombolytic drugs) 3. (antiplatelet drugs) Slide 39 39 Slide 40 40 (plasmin) (fibrinolysin) (plasminogen) Slide 41 41 (thrombolysis agents) (plasminogen) (fibrinolytics) 3~6 72 Slide 42 42 Alteplase Recombinant 1. (t-PA) 2. Slide 43 43 V O 2 /V B = 1.39 Hb S O 2 Oxygen Content in Blood on HBO PO 2, mmHg O 2 content, ml/dl Dissolved Hgb-bound PO 2 + 1.39 Hb SO 2 Dissolved O 2 = P O 2 = 0.003 x 2000 [3 ATA] = 6.0 ml/dl Hb bound O 2 = 1.39 Hb SO 2 = 1.39 x 14 x 100% 19.7 ml/dl 3 ATA Total Slide 44 44 Hyperbaric Oxygen Therapy HBO 1. HBO 1 3 100% 2. Leach et al., 1998