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Improvement of hematoma absorption and neurological function in patients with acute intracerebral hemorrhage treated with Xueshuantong
Journal of the Neurological Sciences 323 (2012) 236–240
Contents lists available at SciVerse ScienceDirect
Journal of the Neurological Sciences journal homepage: www.elsevier.com/locate/jns
Improvement of hematoma absorption and neurological function in patients with acute intracerebral hemorrhage treated with Xueshuantong Li Gao a, b, c, 1, Haiping Zhao b, 1, Qian Liu a, Juexian Song a, Changmin Xu a, Ping Liu a, Wei Gong b, c, Rongliang Wang b, Ke Jian Liu d,⁎, Yumin Luo b, c,⁎⁎ a
Department of Neurology, Capital Medical University, Beijing, China Cerebrovascular Diseases Research Institute, Xuanwu hospital of Capital Medical University, Beijing, China Ministry of Education Key Laboratory of Neurodegenerative Diseases, Capital Medical University, Beijing, China d Department of Pharmaceutical Sciences, University of New Mexico, Albuquerque, NM, USA b c
a r t i c l e
i n f o
Article history: Received 20 June 2012 Received in revised form 22 September 2012 Accepted 25 September 2012 Available online 10 October 2012 Keywords: Intracerebral hemorrhage Hematoma Neurological function C-reactive protein Xueshuantong Injection
a b s t r a c t Spontaneous intracerebral hemorrhage (ICH) leads to high mortality and morbidity. Currently, there is no effective therapy for ICH. Herein we conducted a clinical study in patients with acute ICH to investigate the efficacy of Xueshuantong Injection, a Chinese herbal prescription known for treatment of ischemic diseases in China. Patients (n = 63) were randomly assigned to control (n = 29) and Xueshuantong Injection treatment (175 mg/d, n = 34) groups. Both groups were evaluated using their history and vital signs. The National Institutes of Health Stroke Scale (NIHSS) scores, hematoma volume by CT scanning, and inflammatory factors were assessed before and after two weeks treatment. There were no significant differences in all parameters between two groups before treatment. The treatment group showed significant decreases in both NIHSS score and hematoma volume, compared to control group after treatment (P b 0.01 and P b 0.05, respectively). Furthermore, the inflammatory factors, as measured by leukocytes, neutrophil percentage and C-reactive protein values, were significantly reduced in treatment group compared to control group after treatment (P b 0.05, P b 0.05, P b 0.01 respectively). Our results showed that treatment with Xueshuantong Injection reduced inflammatory response and increased hematoma absorption, which significantly improved recovery of neurological function. This suggests Xueshuantong Injection as a potential treatment of patients with acute ICH. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Cerebrovascular disease (CVD) is a leading cause of death. There are about 700,000 incidences of CVD in the United States each year, of which nearly 160,000 patients die. In 2004, the economic burden was estimated at 53.6 billion U.S. dollars in the US alone [1]. In Europe and Asia, the incidence of intracerebral hemorrhage (ICH) accounts for 10–15% and 20–30% of all cerebrovascular disease, respectively. Of the top disease killers, cerebral hemorrhage has the highest mortality and disability rates. Its mortality rate during the first month following the first onset of the disease is 35–52% [2]. Currently, there is no effective therapy for acute ICH. Present treatments for ICH include stopping or slowing the bleeding in the
⁎ Corresponding author. Tel.: +1 505 272 9546. ⁎⁎ Correspondence to: Y. Luo, Cerebrovascular Diseases Research Institute, Xuanwu Hospital, Capital Medical University, 45 Changchun Street, Beijing, 100053, China. Tel.: + 86 10 83198129; fax: + 86 10 83194754. E-mail addresses: [email protected] (K.J. Liu), [email protected] (Y. Luo). 1 Co-first author. 0022-510X/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jns.2012.09.028
first few hours after onset; removal of blood from the parenchyma or ventricles to eliminate both mechanical and chemical factors that cause secondary insults [3]; management of secondary complications due to blood in the brain, including increased intracranial pressure and decreased cerebral perfusion pressure; and finally good general supportive management [3]. Good clinical practice for supportive care includes management of airways, oxygenation, circulation, glucose level, fever, and nutrition, as well as prophylaxis for deep vein thrombosis [3]. Due to the lack of effective pharmacologic or surgical therapies for ICH, and consequently the lack of a generally accepted standard form of treatment, there is a great variability in the treatment of ICH worldwide. The outcomes for ICH have not changed much in the last 20 to 30 years [4–6]. At the root of the problem is the fact that presently an effective neuroprotective treatment strategy for ICH does not exist. Basic research in recent years indicates that local tissue deformation, excitotoxicity, apoptosis, and inflammation from hemorrhage and ischemia all contribute to neurological damage [7]. Among them, ICHinduced inflammation appears to be an important factor of secondary brain damage, and many studies have suggested that the use of anti-inflammatory drugs may improve recovery in hemorrhagic stroke [7–11].
L. Gao et al. / Journal of the Neurological Sciences 323 (2012) 236–240
The Chinese medicinal herb, Panax notoginseng (Sanqi), has been used for over a thousand of years in traditional Chinese medicine and is known to possess potent anti-inflammatory properties. Xueshuantong Injection was composed of P. notoginseng saponins (PNS), and has been approved by State Food and Drug Administration of China (sFDA) for treatment of patients with some ischemic diseases. PNS has been reported to inhibit the production of TNF-α and IL-6 by dendritic cells following LPS activation of Toll-like receptor [12]. Additionally, PNS affected the expression of the co-stimulatory molecules CD40 and CD86 on dendritic cells following activation by different Toll-like receptor ligands [12]. A recent study demonstrated that PNS significantly decreased gene expression of some inflammatory factors such as integrins, interleukin (IL)-18, IL-1beta, and matrix metalloproteinases 2 and 9 in the rat model of atherosclerosis [13]. These findings suggest that Xueshuantong Injection has the potential to be utilized as an anti-inflammatory drug for neurological diseases. Currently, some traditional Chinese herbal medicine are used as novel treatment for acute ischemic diseases [14,15]; although, few are used for acute ICH. Since P. notoginseng has been widely applied in China for treatment of ischemic stroke and has been found to be beneficial and safe [16], herein we carried out a pilot clinical study to demonstrate the proof-of-principle in utilizing Xueshuantong Injection as a potential strategy for treatment of acute ICH. The NIHSS score, hematoma volume, and inflammatory response, as determined by total leukocyte count, neutrophil percentage, and C-reactive protein level were compared between regular treatment and regular treatment plus Xueshuantong Injection.
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glucose-lowering treatment for hyperglycemia, antihypertensive treatment for hypertension, anti-inflammatory treatment for infection, acid inhibitor for peptic ulcer, and (iii) supportive therapy, such as physical cooling, nutritional support, fluid, and electrolyte balance, which was provided as needed. 2.3. NIHSS score and general information collection History and physical findings, including detailed neurological assessments, were recorded before treatment. In particular, patient information (age, gender, height, weight, and body weight index), vital signs (body temperature, heart rate, respiratory rate, and mean arterial pressure), previous disease history hypertension, heart disease, hyperlipidemia, and diabetes), and personal history (smoking and alcohol consumption) were noted. The scores of the National Institutes of Health Stroke Scale (NIHSS) were evaluated in both groups before and after treatment. 2.4. Laboratory parameters
2. Material and methods
Venous blood samples were collected before and after treatment and tested for C-reactive protein (CRP), as well as leukocyte and neutrophil number. The total circulating leukocyte number and neutrophil number were determined from whole blood using an automatic hematology analyzer (Bayer Healthcare, Siemens). Automated neutrophil counts were provided by Quest Diagnostics Clinical Trials (Van Nuys, CA USA). A high sensitivity sandwich enzyme-linked immunoassay (ELISA) (Aushon Biosystems, Inc., USA) was used to measure CRP.
2.1. Participants
2.5. Measurement of hematoma by CT scanning
The study protocol had been approved by the institute's committee on human research, and informed consent was obtained from all participating patients. Patients at the onset of their first intracerebral hemorrhagic stroke, were confirmed by computer tomography (CT), and randomly recruited into the study in the Department of Neurology, Xuanwu Hospital of Capital Medical University, from February 2008 to February 2009. Inclusion criteria were: 35 to 75 years old; the site of hematoma located in the cerebral hemisphere, hematoma volume ≤ 30 ml, and no blood in the ventricles; intracerebral hemorrhage within 24–72 h (including 72 h); no loss of consciousness (drowsiness acceptable); no heart, liver, kidney and other major organ diseases, no anticoagulants; no infectious or autoimmune diseases; not on immunosuppressive agents, anti-inflammatory drugs or hormones. Informed consent was obtained from the patients or their close relatives. Exclusion criteria were: cerebellar or brainstem hemorrhage; cerebral infarct detected by CT; intracerebral hemorrhage caused by bleeding diathesis, aneurysms, vascular malformations, improperly using anticoagulant drugs, or suspicious amyloid angiopathy. Additional exclusion criteria included patients with poor compliance, insufficient information; and patients with other contraindications for participation in the study.
Hematoma was determined by CT scanning (Siemens 64-slice CT machine) and calculated in accordance with the following equation: hematoma volume (ml) = π/6× length (cm)× width (cm)× high (cm). We performed all CT studies using the following parameters: section thickness, 5 mm; gap, 5 mm; pitch, 1; tube current, 304 mA; and voltage, 120 kV.
2.2. Patient groups and treatment The ICH patients were randomly assigned into two groups: control group (regular treatment plus 250 ml saline as placebo) and Xueshuantong Injection group (regular treatment plus Xueshuantong Injection 175 mg/5 ml in 250 ml saline). Xueshuantong Injection (Lot: Z44020284, Limin Pharmaceutical Group, China) was given to the patient by intravenous infusion once a day for two weeks. Regular treatment consists of: (i) dehydration therapy by 20% mannitol (Tianjin Baite Medical Drugs Ltd., Co., China.) with the dosage from 125 to 250 ml every 8 h for 3–7 days depending on their clinically presumed intracranial pressure, (ii) therapy to deal with complications including
2.6. Statistical analysis Data were subsequently analyzed with the software SPSS 11.5 for Windows package. Continuous values are expressed as a mean ± SD (range). Pearson Chi-square test was used to analyze the basic characteristics between control and treatment group. Values of pre- and post-treatment were analyzed by paired-samples t-test; otherwise the independent-samples t-test was used to compare values in different groups. A two sided P-value b 0.05 was considered statistically significant. 3. Results 3.1. Basic health conditions and cerebrovascular disease risk factors A total of 63 patients were enrolled and randomly assigned either to the control (n = 29) or the Xueshuantong Injection (n = 34) group (Fig. 1). The control group had 14 male and 15 female with average age of 54.0 ± 13.0 years; and the Xueshuang tong group, 22 male and 12 female with average age of 54.1 ± 10.6 years. No statistical significance was observed in age and sex between the groups and there were no statistical differences between the two groups in height, weight, body mass index, body temperature, heart rate, respiration rate, or mean arterial pressure (data not shown). Smoking, drinking, high blood pressure, pathoglycemia, heart disease, and dyslipidemia were considered to be the main risk factors for cerebrovascular diseases. These major risk factors were analyzed and no
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was no significant difference of hematoma volume before treatment between the two groups (Fig. 3). After two weeks of treatment, the hematoma volume in the Xueshuantong Injection group was significantly smaller compared to the control group (P b 0.05). The change between pre- and post-treatment for the hematoma volume was significantly greater in the Xueshuantong Injection group than that in the control group (P b 0.01). These results suggest that Xueshuantong Injection could promote the absorption of hematoma in ICH patients. 3.4. Leukocyte count To investigate the possible neuroprotective mechanism of Xueshuantong Injection treatment and to assess the effect on ICHinduced inflammation, the blood leukocytes and the neutrophil percentage of the patients were determined in the two groups before and after treatment. There was no significant difference in blood leukocytes count (Fig. 4) and neutrophil (Fig. 5) percentage before treatment between the two groups. Although no significant difference was observed between the two groups after the treatment, the changes between pre- and post-treatment of blood leukocytes count and neutrophil percentage in the Xueshuantong Injection group was significantly greater compared to the control group (Pb 0.05). Fig. 1. Schematic of clinical study design.
statistical significance was observed between the two groups (data not shown). 3.2. NIHSS scores In order to determine the effect of treatment with Xueshuantong Injection on neurological function of the patients NIHSS score was assessed. Before treatment there was no significant difference in NIHSS scores between the two groups of patients. Following the two-week treatment, the NIHSS score in both groups were found to be reduced and the Xueshuantong Injection treated group was significantly lower compared with the control group (Pb 0.05). Furthermore, the change of the NIHSS scores between pre- and post-treatment in the Xueshuantong Injection group was significantly greater than that in the control group (Pb 0.01) (Fig. 2). These results indicate that the Xueshuantong Injection group had a measurably better recovery of neurological functions than the control group. 3.3. Hematoma volume Hematoma volume is an important predictor of patient prognosis after acute ICH, and was assessed for all patients in this study. There
Fig. 2. Xueshuantong Injection improved the NIHSS scores of patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups after 2 week treatment. **P b 0.01 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N = 34).
3.5. Serum CRP level In order to further investigate the effects of Xueshuantong Injection on the inflammatory factors in the patients with acute ICH, the amount of serum C-reactive protein in the two groups were analyzed. There was no significant difference in CRP before treatment between the two groups (Fig. 6). After two-week treatment, the values of CRP were decreased in both control group and Xueshuantong Injection group when compared with pretreatment, but the serum CRP level was significantly lower in the Xueshuantong Injection group compared to the control group (P b 0.05). Furthermore, the change between preand post-treatment of serum CRP in Xueshuantong Injection group was significantly greater than that in the control group (P b 0.01). These results demonstrate that the serum concentration of CRP in the Xueshuantong Injection group was significantly decreased compared to the control group, suggesting that the anti-inflammation property of Xueshuantong Injection could be a possible mechanism of its neuroprotective effects. 4. Discussion Intracerebral hemorrhage (ICH) is a common neurological disorder, and is a major threat to the health of people in middle-age and
Fig. 3. Xueshuantong Injection decreased the hematoma volumes of patients with intracerebral hemorrhage after 2-week treatment determined by CT scanning. *P b 0.05 treatment vs. control group after 2 week treatment. **P b 0.01 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N= 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N= 34).
L. Gao et al. / Journal of the Neurological Sciences 323 (2012) 236–240
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Fig. 4. Xueshuantong Injection reduced the blood leukocyte count in patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N= 34).
Fig. 6. Xueshuantong Injection reduced the serum CRP level of patients with intracerebral hemorrhage after 2 week treatment detected by ELISA. *Pb 0.05 treatment vs. control groups. **Pb 0.01 treatment vs. control groups for the change between pre- and posttreatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between preand post-treatment. Control (regular treatment group, N=29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N=34).
the elderly [17]. The current treatment of ICH is far from desirable, and is mainly limited to the management of increased arterial blood pressure and intracranial pressure, the medical complications of intracerebral hemorrhage, and the prevention of recurrent intracerebral hemorrhage [3]. A clinically effective treatment is urgently needed. Recently, basic research has demonstrated that thrombin inhibitors (hirudin, Argatroban) and a complement activation system inhibitor (N-acetyl heparin) can significantly reduce the brain edema and improve the prognosis following ICH [18,19]. The beneficial effects of the traditional Chinese medicine have been reported in a rat model of cerebral hemorrhage [20]. Xueshuantong Injection, a crude extraction from the widely used traditional Chinese medicine P. notoginseng, has been approved by Chinese FDA for treatment of certain ischemic diseases in China. However, currently there is a lack of sufficient clinical evidence to support its routine use for stroke, especially hemorrhagic stroke. In the present study, we investigated the effects of Xueshuantong Injection in patients with intracerebral hemorrhage. We found that patients given Xueshuantong Injection treatment showed significantly better improvement in hematoma absorption and behavioral function recovery. After two weeks of treatment, NIHSS score of the Xueshuantong Injection group showed a significant decrease when compared with the control group. The magnitude of change of the NIHSS scores between pre- and post-treatment in the Xueshuantong Injection group was significantly greater than that in the control group indicating that
the Xueshuantong Injection group had a measurably better recovery of neurological functions than the control group. The beneficial effect of Xueshuantong Injection in ICH patients could be due to several mechanisms [16,21]. It has been reported that the main active ingredients of Xueshuantong Injection, PNS, have been found to have functions in anti-inflammatory effects. PNS could reduce leukocyte adhesion in rat mesentery venules under the inhibitory effect on the expression of adhesion molecules (CD11b and CD18) on neutrophils [22]. PNS also exerted their therapeutic effects on atherosclerosis through an anti-inflammatory action and regulation of the blood lipid profile via inhibiting NF-kappaB signaling pathway [13]. These reports highlight the anti-inflammatory property of PNS. Mounting evidence has shown that there are significant inflammatory responses following cerebral hemorrhage, including leukocyte infiltration, microglial cell activation, the activation of the complement system, cytokines and adhesion molecule expression. The interactions among these mechanisms will undoubtedly contribute toward the subsequent pathological processes of brain edema and brain damage that are seen after cerebral hemorrhage [23,24]. A prominent inflammatory response, which includes the activation of resident brain microglia and inflammatory infiltration into the brain, is initiated by neutrophils followed by macrophages [25]. This process is accompanied by the massive rapid activation of the peripheral immune system and the dynamic and widespread activation of inflammatory cytokines and chemokines in the spleen [26]. TNF-α is one of the major inflammatory mediators in stroke [27], and it is largely produced by infiltrating macrophages [28]. In our study, we investigated the changes of inflammatory factors in ICH patients after Xueshuantong Injection treatment. We found that the total leukocyte count, neutrophil percentage, and CRP level were significantly decreased in patients with Xueshuantong Injection treatment compared to the control group. These findings indicate that the anti-inflammatory function of Xueshuantong Injection may be one of the underlying mechanisms responsible for the neuroprotective effects in patients with intracerebral hemorrhage. Hematoma is an important predictor of poor outcome for ICH. Therapies designed to stabilize hematoma growth and reduce hematoma burden have been shown to improve outcomes [29]. Iron accumulates in the brain and contributes to brain injury after intracerebral hemorrhage. Iron overload occurs in the brain after intracerebral hemorrhage in rats. Experiments have indicated that iron and clot lysis play an important role in intracerebral hemorrhage-induced brain injury. Iron chelation has been suggested as an effective therapy for intracerebral hemorrhage [30]. Our results presented here show that the hematoma in the Xueshuantong Injection group was significantly smaller compared to the control group after treatment. The magnitude of change between pre- and post-treatment for the
Fig. 5. Xueshuantong Injection decreased the neutrophil percentage in patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N = 34).
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hematoma was significantly greater in the Xueshuantong Injection group than that of the control group. These findings suggest that Xueshuantong Injection could promote the absorption of hematoma and improve the prognosis of patients with cerebral hemorrhage. In summary, the results presented here demonstrate that Xueshuantong Injection treatment could decrease the hematoma and inflammatory response, leading to improved outcomes of patients with cerebral hemorrhage. Xueshuantong Injection could be an effective clinically viable strategy as a neuroprotective therapy for intracerebral hemorrhage. However, the positive findings observed in this pilot study require further confirmation with clinical trials at multiple centers and with larger patient size. Conflict of interest The authors declare no conflict of interest. Acknowledgment This work was supported by the Projects of Capital Medical Technology Development Foundation (No. 2005-SF-II-038) and the Natural Science Foundation in China (No: 81071058). References [1] Goldstein LB, Adams R, Alberts MJ, Appel LJ, Brass LM, Bushnell CD, et al. Primary prevention of ischemic stroke: a guideline from the American Heart Association/ American Stroke Association Stroke Council: cosponsored by the Atherosclerotic Peripheral Vascular Disease Interdisciplinary Working Group; Cardiovascular Nursing Council; Clinical Cardiology Council; Nutrition, Physical Activity, and Metabolism Council; and the Quality of Care and Outcomes Research Interdisciplinary Working Group: the American Academy of Neurology affirms the value of this guideline. Stroke 2006;37:1583-633. [2] Broderick JP, Adams Jr HP, Barsan W, Feinberg W, Feldmann E, Grotta J, et al. Guidelines for the management of spontaneous intracerebral hemorrhage: a statement for healthcare professionals from a special writing group of the Stroke Council, American Heart Association. Stroke 1999;30:905-15. [3] Broderick J, Connolly S, Feldmann E, Hanley D, Kase C, Krieger D, et al. Guidelines for management of spontaneous intracerebral hemorrhage in adults: 2007 update: a guideline from the American Heart Association/American Stroke Association Stroke Council, High Blood Pressure Research Council, and the Quality of Care and Outcomes in Research Interdisciplinary Working Group. Stroke 2007;38:2001-23. [4] Teernstra OP, Evers SM, Kessels AH. Meta analyses in treatment of spontaneous supratentorial intracerebral haematoma. Acta Neurochir (Wien) 2006;148:521-8. [5] Dennis MS. Outcome after brain haemorrhage. Cerebrovasc Dis 2003;16:9–13 (Suppl.). [6] Lovelock CE, Molyneux AJ, Rothwell PM. Change in incidence and aetiology of intracerebral haemorrhage in Oxfordshire, UK, between 1981 and 2006: a population-based study. Lancet Neurol 2007;6:487-93. [7] Aronowski J, Hall CE. New horizons for primary intracerebral hemorrhage treatment: experience from preclinical studies. Neurol Res 2005;27:268-79.
[8] Jung KH, Chu K, Jeong SW, Han SY, Lee ST, Kim JY, et al. HMG-CoA reductase inhibitor, atorvastatin, promotes sensorimotor recovery, suppressing acute inflammatory reaction after experimental intracerebral hemorrhage. Stroke 2004;35:1744-9. [9] Chu K, Jeong SW, Jung KH, Han SY, Lee ST, Kim M, et al. Celecoxib induces functional recovery after intracerebral hemorrhage with reduction of brain edema and perihematomal cell death. J Cereb Blood Flow Metab 2004;24:926-33. [10] Wang J, Doré S. Inflammation after intracerebral hemorrhage. J Cereb Blood Flow Metab 2007;27:894-908. [11] Sinn DI, Kim SJ, Chu K, Jung KH, Lee ST, Song EC, et al. Valproic acid mediated neuroprotection in intracerebral hemorrhage via histone deacetylase inhibition and transcriptional activation. Neurobiol Dis 2007;26:464-72. [12] Rhule A, Rase B, Smith JR, Shepherd DM. Toll-like receptor ligand-induced activation of murine DC2.4 cells is attenuated by Panax notoginseng. J Ethnopharmacol 2008;116:179-86. [13] Zhang YG, Zhang HG, Zhang GY, Fan JS, Li XH, Liu YH, et al. Panax notoginseng saponins attenuate atherosclerosis in rats by regulating the blood lipid profile and an anti-inflammatory action. Clin Exp Pharmacol Physiol 2008;35:1238-44. [14] Fu XX, Xiao WJ, Lu J, Wu LW, Yang F. Retrospective analysis of thrombolysis therapy for 64 cases of acute myocardial infarction with elevated ST segment. Chin J Integr Med 2009;15:462-5. [15] Wu B, Liu M, Liu H, Li W, Tan S, Zang SH, et al. Meta-analysis of traditional Chinese patent medicine for ischemic stroke. Stroke 2007;38:1973-9. [16] Chen X, Zhou M, Li Q, Yang J, Zhang Y, Zhang D, et al. Sanchi for acute ischemic stroke. Stroke 2009;40:e394-5. [17] Thompson KM, Gerlach SY, Jom HK, Larson JM, Brott TG, Files JA. Advances in the care of patients with intracerebral hemorrhage. Mayo Clin Proc 2007;82:987-90. [18] Hua Y, Schaller T, Keep RF, Wu J, Hoff JT, Xi G. Behavioral tests after intracerebral hemorrhage in the rat. Stroke 2002;33:2478-84. [19] Mayer SA. Ultra-early hemostatic therapy for intracerebral hemorrhage. Stroke 2003;34:224-9. [20] Zhang ZQ, Tang T, Luo JK, Huang JF, Yang QD, Li XQ, et al. Effect of qi-tonifying and stasis-eliminating therapy on expression of vascular endothelial growth factor and its receptors Flt-1, Flk-1 in the brain of intracerebral hemorrhagic rats. Chin J Integr Med 2007;13:285-90. [21] Chen X, Zhou M, Li Q, Yang J, Zhang Y, Zhang D, et al. Sanchi for acute ischaemic stroke. Cochrane Database Syst Rev 2008;8:CD006305. [22] Sun K, Wang CS, Guo J, Liu YY, Wang F, Liu LY, et al. Effect of Panax notoginseng saponins on lipopolysaccharide-induced adhesion of leukocytes in rat mesenteric venules. Clin Hemorheol Microcirc 2006;34:103-8. [23] Hallenbeck JM, Hansson GK, Becker KJ. Immunology of ischemic vascular disease: plaque to attack. Trends Immunol 2005;26:550-6. [24] Power C, Henry S, Del Bigio MR, Larsen PH, Corbett D, Imai Y, et al. Intracerebral hemorrhage induces macrophage activation and matrix metalloproteinases. Ann Neurol 2003;53:731-42. [25] Del Bigio MR, Yan HJ, Buist R, Peeling J. Experimental intracerebral hemorrhage in rats. Magnetic resonance imaging and histopathological correlates. Stroke 1996;27:2312-20. [26] Offner H, Subramanian S, Parker SM, Afentoulis ME, Vandenbark AA, Hurn PD. Experimental stroke induces massive, rapid activation of the peripheral immune system. J Cereb Blood Flow Metab 2006;26:654-65. [27] Zheng Z, Yenari MA. Post-ischemic inflammation: molecular mechanisms and therapeutic implications. Neurol Res 2004;26:884-92. [28] Gregersen R, Lambertsen K, Finsen B. Microglia and macrophages are the major source of tumor necrosis factor in permanent middle cerebral artery occlusion in mice. J Cereb Blood Flow Metab 2000;20:53-65. [29] Nyquist P. Management of acute intracranial and intraventricular hemorrhage. Crit Care Med 2010;38:946-53. [30] Hua Y, Keep RF, Hoff JT, Xi G. Deferoxamine therapy for intracerebral hemorrhage. Acta Neurochir Suppl 2008;105:3-6.
Contents lists available at SciVerse ScienceDirect
Journal of the Neurological Sciences journal homepage: www.elsevier.com/locate/jns
Improvement of hematoma absorption and neurological function in patients with acute intracerebral hemorrhage treated with Xueshuantong Li Gao a, b, c, 1, Haiping Zhao b, 1, Qian Liu a, Juexian Song a, Changmin Xu a, Ping Liu a, Wei Gong b, c, Rongliang Wang b, Ke Jian Liu d,⁎, Yumin Luo b, c,⁎⁎ a
Department of Neurology, Capital Medical University, Beijing, China Cerebrovascular Diseases Research Institute, Xuanwu hospital of Capital Medical University, Beijing, China Ministry of Education Key Laboratory of Neurodegenerative Diseases, Capital Medical University, Beijing, China d Department of Pharmaceutical Sciences, University of New Mexico, Albuquerque, NM, USA b c
a r t i c l e
i n f o
Article history: Received 20 June 2012 Received in revised form 22 September 2012 Accepted 25 September 2012 Available online 10 October 2012 Keywords: Intracerebral hemorrhage Hematoma Neurological function C-reactive protein Xueshuantong Injection
a b s t r a c t Spontaneous intracerebral hemorrhage (ICH) leads to high mortality and morbidity. Currently, there is no effective therapy for ICH. Herein we conducted a clinical study in patients with acute ICH to investigate the efficacy of Xueshuantong Injection, a Chinese herbal prescription known for treatment of ischemic diseases in China. Patients (n = 63) were randomly assigned to control (n = 29) and Xueshuantong Injection treatment (175 mg/d, n = 34) groups. Both groups were evaluated using their history and vital signs. The National Institutes of Health Stroke Scale (NIHSS) scores, hematoma volume by CT scanning, and inflammatory factors were assessed before and after two weeks treatment. There were no significant differences in all parameters between two groups before treatment. The treatment group showed significant decreases in both NIHSS score and hematoma volume, compared to control group after treatment (P b 0.01 and P b 0.05, respectively). Furthermore, the inflammatory factors, as measured by leukocytes, neutrophil percentage and C-reactive protein values, were significantly reduced in treatment group compared to control group after treatment (P b 0.05, P b 0.05, P b 0.01 respectively). Our results showed that treatment with Xueshuantong Injection reduced inflammatory response and increased hematoma absorption, which significantly improved recovery of neurological function. This suggests Xueshuantong Injection as a potential treatment of patients with acute ICH. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Cerebrovascular disease (CVD) is a leading cause of death. There are about 700,000 incidences of CVD in the United States each year, of which nearly 160,000 patients die. In 2004, the economic burden was estimated at 53.6 billion U.S. dollars in the US alone [1]. In Europe and Asia, the incidence of intracerebral hemorrhage (ICH) accounts for 10–15% and 20–30% of all cerebrovascular disease, respectively. Of the top disease killers, cerebral hemorrhage has the highest mortality and disability rates. Its mortality rate during the first month following the first onset of the disease is 35–52% [2]. Currently, there is no effective therapy for acute ICH. Present treatments for ICH include stopping or slowing the bleeding in the
⁎ Corresponding author. Tel.: +1 505 272 9546. ⁎⁎ Correspondence to: Y. Luo, Cerebrovascular Diseases Research Institute, Xuanwu Hospital, Capital Medical University, 45 Changchun Street, Beijing, 100053, China. Tel.: + 86 10 83198129; fax: + 86 10 83194754. E-mail addresses: [email protected] (K.J. Liu), [email protected] (Y. Luo). 1 Co-first author. 0022-510X/$ – see front matter © 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.jns.2012.09.028
first few hours after onset; removal of blood from the parenchyma or ventricles to eliminate both mechanical and chemical factors that cause secondary insults [3]; management of secondary complications due to blood in the brain, including increased intracranial pressure and decreased cerebral perfusion pressure; and finally good general supportive management [3]. Good clinical practice for supportive care includes management of airways, oxygenation, circulation, glucose level, fever, and nutrition, as well as prophylaxis for deep vein thrombosis [3]. Due to the lack of effective pharmacologic or surgical therapies for ICH, and consequently the lack of a generally accepted standard form of treatment, there is a great variability in the treatment of ICH worldwide. The outcomes for ICH have not changed much in the last 20 to 30 years [4–6]. At the root of the problem is the fact that presently an effective neuroprotective treatment strategy for ICH does not exist. Basic research in recent years indicates that local tissue deformation, excitotoxicity, apoptosis, and inflammation from hemorrhage and ischemia all contribute to neurological damage [7]. Among them, ICHinduced inflammation appears to be an important factor of secondary brain damage, and many studies have suggested that the use of anti-inflammatory drugs may improve recovery in hemorrhagic stroke [7–11].
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The Chinese medicinal herb, Panax notoginseng (Sanqi), has been used for over a thousand of years in traditional Chinese medicine and is known to possess potent anti-inflammatory properties. Xueshuantong Injection was composed of P. notoginseng saponins (PNS), and has been approved by State Food and Drug Administration of China (sFDA) for treatment of patients with some ischemic diseases. PNS has been reported to inhibit the production of TNF-α and IL-6 by dendritic cells following LPS activation of Toll-like receptor [12]. Additionally, PNS affected the expression of the co-stimulatory molecules CD40 and CD86 on dendritic cells following activation by different Toll-like receptor ligands [12]. A recent study demonstrated that PNS significantly decreased gene expression of some inflammatory factors such as integrins, interleukin (IL)-18, IL-1beta, and matrix metalloproteinases 2 and 9 in the rat model of atherosclerosis [13]. These findings suggest that Xueshuantong Injection has the potential to be utilized as an anti-inflammatory drug for neurological diseases. Currently, some traditional Chinese herbal medicine are used as novel treatment for acute ischemic diseases [14,15]; although, few are used for acute ICH. Since P. notoginseng has been widely applied in China for treatment of ischemic stroke and has been found to be beneficial and safe [16], herein we carried out a pilot clinical study to demonstrate the proof-of-principle in utilizing Xueshuantong Injection as a potential strategy for treatment of acute ICH. The NIHSS score, hematoma volume, and inflammatory response, as determined by total leukocyte count, neutrophil percentage, and C-reactive protein level were compared between regular treatment and regular treatment plus Xueshuantong Injection.
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glucose-lowering treatment for hyperglycemia, antihypertensive treatment for hypertension, anti-inflammatory treatment for infection, acid inhibitor for peptic ulcer, and (iii) supportive therapy, such as physical cooling, nutritional support, fluid, and electrolyte balance, which was provided as needed. 2.3. NIHSS score and general information collection History and physical findings, including detailed neurological assessments, were recorded before treatment. In particular, patient information (age, gender, height, weight, and body weight index), vital signs (body temperature, heart rate, respiratory rate, and mean arterial pressure), previous disease history hypertension, heart disease, hyperlipidemia, and diabetes), and personal history (smoking and alcohol consumption) were noted. The scores of the National Institutes of Health Stroke Scale (NIHSS) were evaluated in both groups before and after treatment. 2.4. Laboratory parameters
2. Material and methods
Venous blood samples were collected before and after treatment and tested for C-reactive protein (CRP), as well as leukocyte and neutrophil number. The total circulating leukocyte number and neutrophil number were determined from whole blood using an automatic hematology analyzer (Bayer Healthcare, Siemens). Automated neutrophil counts were provided by Quest Diagnostics Clinical Trials (Van Nuys, CA USA). A high sensitivity sandwich enzyme-linked immunoassay (ELISA) (Aushon Biosystems, Inc., USA) was used to measure CRP.
2.1. Participants
2.5. Measurement of hematoma by CT scanning
The study protocol had been approved by the institute's committee on human research, and informed consent was obtained from all participating patients. Patients at the onset of their first intracerebral hemorrhagic stroke, were confirmed by computer tomography (CT), and randomly recruited into the study in the Department of Neurology, Xuanwu Hospital of Capital Medical University, from February 2008 to February 2009. Inclusion criteria were: 35 to 75 years old; the site of hematoma located in the cerebral hemisphere, hematoma volume ≤ 30 ml, and no blood in the ventricles; intracerebral hemorrhage within 24–72 h (including 72 h); no loss of consciousness (drowsiness acceptable); no heart, liver, kidney and other major organ diseases, no anticoagulants; no infectious or autoimmune diseases; not on immunosuppressive agents, anti-inflammatory drugs or hormones. Informed consent was obtained from the patients or their close relatives. Exclusion criteria were: cerebellar or brainstem hemorrhage; cerebral infarct detected by CT; intracerebral hemorrhage caused by bleeding diathesis, aneurysms, vascular malformations, improperly using anticoagulant drugs, or suspicious amyloid angiopathy. Additional exclusion criteria included patients with poor compliance, insufficient information; and patients with other contraindications for participation in the study.
Hematoma was determined by CT scanning (Siemens 64-slice CT machine) and calculated in accordance with the following equation: hematoma volume (ml) = π/6× length (cm)× width (cm)× high (cm). We performed all CT studies using the following parameters: section thickness, 5 mm; gap, 5 mm; pitch, 1; tube current, 304 mA; and voltage, 120 kV.
2.2. Patient groups and treatment The ICH patients were randomly assigned into two groups: control group (regular treatment plus 250 ml saline as placebo) and Xueshuantong Injection group (regular treatment plus Xueshuantong Injection 175 mg/5 ml in 250 ml saline). Xueshuantong Injection (Lot: Z44020284, Limin Pharmaceutical Group, China) was given to the patient by intravenous infusion once a day for two weeks. Regular treatment consists of: (i) dehydration therapy by 20% mannitol (Tianjin Baite Medical Drugs Ltd., Co., China.) with the dosage from 125 to 250 ml every 8 h for 3–7 days depending on their clinically presumed intracranial pressure, (ii) therapy to deal with complications including
2.6. Statistical analysis Data were subsequently analyzed with the software SPSS 11.5 for Windows package. Continuous values are expressed as a mean ± SD (range). Pearson Chi-square test was used to analyze the basic characteristics between control and treatment group. Values of pre- and post-treatment were analyzed by paired-samples t-test; otherwise the independent-samples t-test was used to compare values in different groups. A two sided P-value b 0.05 was considered statistically significant. 3. Results 3.1. Basic health conditions and cerebrovascular disease risk factors A total of 63 patients were enrolled and randomly assigned either to the control (n = 29) or the Xueshuantong Injection (n = 34) group (Fig. 1). The control group had 14 male and 15 female with average age of 54.0 ± 13.0 years; and the Xueshuang tong group, 22 male and 12 female with average age of 54.1 ± 10.6 years. No statistical significance was observed in age and sex between the groups and there were no statistical differences between the two groups in height, weight, body mass index, body temperature, heart rate, respiration rate, or mean arterial pressure (data not shown). Smoking, drinking, high blood pressure, pathoglycemia, heart disease, and dyslipidemia were considered to be the main risk factors for cerebrovascular diseases. These major risk factors were analyzed and no
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was no significant difference of hematoma volume before treatment between the two groups (Fig. 3). After two weeks of treatment, the hematoma volume in the Xueshuantong Injection group was significantly smaller compared to the control group (P b 0.05). The change between pre- and post-treatment for the hematoma volume was significantly greater in the Xueshuantong Injection group than that in the control group (P b 0.01). These results suggest that Xueshuantong Injection could promote the absorption of hematoma in ICH patients. 3.4. Leukocyte count To investigate the possible neuroprotective mechanism of Xueshuantong Injection treatment and to assess the effect on ICHinduced inflammation, the blood leukocytes and the neutrophil percentage of the patients were determined in the two groups before and after treatment. There was no significant difference in blood leukocytes count (Fig. 4) and neutrophil (Fig. 5) percentage before treatment between the two groups. Although no significant difference was observed between the two groups after the treatment, the changes between pre- and post-treatment of blood leukocytes count and neutrophil percentage in the Xueshuantong Injection group was significantly greater compared to the control group (Pb 0.05). Fig. 1. Schematic of clinical study design.
statistical significance was observed between the two groups (data not shown). 3.2. NIHSS scores In order to determine the effect of treatment with Xueshuantong Injection on neurological function of the patients NIHSS score was assessed. Before treatment there was no significant difference in NIHSS scores between the two groups of patients. Following the two-week treatment, the NIHSS score in both groups were found to be reduced and the Xueshuantong Injection treated group was significantly lower compared with the control group (Pb 0.05). Furthermore, the change of the NIHSS scores between pre- and post-treatment in the Xueshuantong Injection group was significantly greater than that in the control group (Pb 0.01) (Fig. 2). These results indicate that the Xueshuantong Injection group had a measurably better recovery of neurological functions than the control group. 3.3. Hematoma volume Hematoma volume is an important predictor of patient prognosis after acute ICH, and was assessed for all patients in this study. There
Fig. 2. Xueshuantong Injection improved the NIHSS scores of patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups after 2 week treatment. **P b 0.01 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N = 34).
3.5. Serum CRP level In order to further investigate the effects of Xueshuantong Injection on the inflammatory factors in the patients with acute ICH, the amount of serum C-reactive protein in the two groups were analyzed. There was no significant difference in CRP before treatment between the two groups (Fig. 6). After two-week treatment, the values of CRP were decreased in both control group and Xueshuantong Injection group when compared with pretreatment, but the serum CRP level was significantly lower in the Xueshuantong Injection group compared to the control group (P b 0.05). Furthermore, the change between preand post-treatment of serum CRP in Xueshuantong Injection group was significantly greater than that in the control group (P b 0.01). These results demonstrate that the serum concentration of CRP in the Xueshuantong Injection group was significantly decreased compared to the control group, suggesting that the anti-inflammation property of Xueshuantong Injection could be a possible mechanism of its neuroprotective effects. 4. Discussion Intracerebral hemorrhage (ICH) is a common neurological disorder, and is a major threat to the health of people in middle-age and
Fig. 3. Xueshuantong Injection decreased the hematoma volumes of patients with intracerebral hemorrhage after 2-week treatment determined by CT scanning. *P b 0.05 treatment vs. control group after 2 week treatment. **P b 0.01 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N= 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N= 34).
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Fig. 4. Xueshuantong Injection reduced the blood leukocyte count in patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N= 34).
Fig. 6. Xueshuantong Injection reduced the serum CRP level of patients with intracerebral hemorrhage after 2 week treatment detected by ELISA. *Pb 0.05 treatment vs. control groups. **Pb 0.01 treatment vs. control groups for the change between pre- and posttreatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between preand post-treatment. Control (regular treatment group, N=29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N=34).
the elderly [17]. The current treatment of ICH is far from desirable, and is mainly limited to the management of increased arterial blood pressure and intracranial pressure, the medical complications of intracerebral hemorrhage, and the prevention of recurrent intracerebral hemorrhage [3]. A clinically effective treatment is urgently needed. Recently, basic research has demonstrated that thrombin inhibitors (hirudin, Argatroban) and a complement activation system inhibitor (N-acetyl heparin) can significantly reduce the brain edema and improve the prognosis following ICH [18,19]. The beneficial effects of the traditional Chinese medicine have been reported in a rat model of cerebral hemorrhage [20]. Xueshuantong Injection, a crude extraction from the widely used traditional Chinese medicine P. notoginseng, has been approved by Chinese FDA for treatment of certain ischemic diseases in China. However, currently there is a lack of sufficient clinical evidence to support its routine use for stroke, especially hemorrhagic stroke. In the present study, we investigated the effects of Xueshuantong Injection in patients with intracerebral hemorrhage. We found that patients given Xueshuantong Injection treatment showed significantly better improvement in hematoma absorption and behavioral function recovery. After two weeks of treatment, NIHSS score of the Xueshuantong Injection group showed a significant decrease when compared with the control group. The magnitude of change of the NIHSS scores between pre- and post-treatment in the Xueshuantong Injection group was significantly greater than that in the control group indicating that
the Xueshuantong Injection group had a measurably better recovery of neurological functions than the control group. The beneficial effect of Xueshuantong Injection in ICH patients could be due to several mechanisms [16,21]. It has been reported that the main active ingredients of Xueshuantong Injection, PNS, have been found to have functions in anti-inflammatory effects. PNS could reduce leukocyte adhesion in rat mesentery venules under the inhibitory effect on the expression of adhesion molecules (CD11b and CD18) on neutrophils [22]. PNS also exerted their therapeutic effects on atherosclerosis through an anti-inflammatory action and regulation of the blood lipid profile via inhibiting NF-kappaB signaling pathway [13]. These reports highlight the anti-inflammatory property of PNS. Mounting evidence has shown that there are significant inflammatory responses following cerebral hemorrhage, including leukocyte infiltration, microglial cell activation, the activation of the complement system, cytokines and adhesion molecule expression. The interactions among these mechanisms will undoubtedly contribute toward the subsequent pathological processes of brain edema and brain damage that are seen after cerebral hemorrhage [23,24]. A prominent inflammatory response, which includes the activation of resident brain microglia and inflammatory infiltration into the brain, is initiated by neutrophils followed by macrophages [25]. This process is accompanied by the massive rapid activation of the peripheral immune system and the dynamic and widespread activation of inflammatory cytokines and chemokines in the spleen [26]. TNF-α is one of the major inflammatory mediators in stroke [27], and it is largely produced by infiltrating macrophages [28]. In our study, we investigated the changes of inflammatory factors in ICH patients after Xueshuantong Injection treatment. We found that the total leukocyte count, neutrophil percentage, and CRP level were significantly decreased in patients with Xueshuantong Injection treatment compared to the control group. These findings indicate that the anti-inflammatory function of Xueshuantong Injection may be one of the underlying mechanisms responsible for the neuroprotective effects in patients with intracerebral hemorrhage. Hematoma is an important predictor of poor outcome for ICH. Therapies designed to stabilize hematoma growth and reduce hematoma burden have been shown to improve outcomes [29]. Iron accumulates in the brain and contributes to brain injury after intracerebral hemorrhage. Iron overload occurs in the brain after intracerebral hemorrhage in rats. Experiments have indicated that iron and clot lysis play an important role in intracerebral hemorrhage-induced brain injury. Iron chelation has been suggested as an effective therapy for intracerebral hemorrhage [30]. Our results presented here show that the hematoma in the Xueshuantong Injection group was significantly smaller compared to the control group after treatment. The magnitude of change between pre- and post-treatment for the
Fig. 5. Xueshuantong Injection decreased the neutrophil percentage in patients with intracerebral hemorrhage after 2-week treatment. *P b 0.05 treatment vs. control groups for the change between pre- and post-treatment. Pre-T, pre-treatment; post-T, post-treatment; CB-PPT, change between pre- and post-treatment. Control (regular treatment group, N = 29), treatment (regular treatment plus Xueshuantong Injection group, 175 mg/d, N = 34).
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hematoma was significantly greater in the Xueshuantong Injection group than that of the control group. These findings suggest that Xueshuantong Injection could promote the absorption of hematoma and improve the prognosis of patients with cerebral hemorrhage. In summary, the results presented here demonstrate that Xueshuantong Injection treatment could decrease the hematoma and inflammatory response, leading to improved outcomes of patients with cerebral hemorrhage. Xueshuantong Injection could be an effective clinically viable strategy as a neuroprotective therapy for intracerebral hemorrhage. However, the positive findings observed in this pilot study require further confirmation with clinical trials at multiple centers and with larger patient size. Conflict of interest The authors declare no conflict of interest. Acknowledgment This work was supported by the Projects of Capital Medical Technology Development Foundation (No. 2005-SF-II-038) and the Natural Science Foundation in China (No: 81071058). References [1] Goldstein LB, Adams R, Alberts MJ, Appel LJ, Brass LM, Bushnell CD, et al. Primary prevention of ischemic stroke: a guideline from the American Heart Association/ American Stroke Association Stroke Council: cosponsored by the Atherosclerotic Peripheral Vascular Disease Interdisciplinary Working Group; Cardiovascular Nursing Council; Clinical Cardiology Council; Nutrition, Physical Activity, and Metabolism Council; and the Quality of Care and Outcomes Research Interdisciplinary Working Group: the American Academy of Neurology affirms the value of this guideline. Stroke 2006;37:1583-633. [2] Broderick JP, Adams Jr HP, Barsan W, Feinberg W, Feldmann E, Grotta J, et al. Guidelines for the management of spontaneous intracerebral hemorrhage: a statement for healthcare professionals from a special writing group of the Stroke Council, American Heart Association. Stroke 1999;30:905-15. [3] Broderick J, Connolly S, Feldmann E, Hanley D, Kase C, Krieger D, et al. Guidelines for management of spontaneous intracerebral hemorrhage in adults: 2007 update: a guideline from the American Heart Association/American Stroke Association Stroke Council, High Blood Pressure Research Council, and the Quality of Care and Outcomes in Research Interdisciplinary Working Group. Stroke 2007;38:2001-23. [4] Teernstra OP, Evers SM, Kessels AH. Meta analyses in treatment of spontaneous supratentorial intracerebral haematoma. Acta Neurochir (Wien) 2006;148:521-8. [5] Dennis MS. Outcome after brain haemorrhage. Cerebrovasc Dis 2003;16:9–13 (Suppl.). [6] Lovelock CE, Molyneux AJ, Rothwell PM. Change in incidence and aetiology of intracerebral haemorrhage in Oxfordshire, UK, between 1981 and 2006: a population-based study. Lancet Neurol 2007;6:487-93. [7] Aronowski J, Hall CE. New horizons for primary intracerebral hemorrhage treatment: experience from preclinical studies. Neurol Res 2005;27:268-79.
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