Changes in Serum Growth Factors in Stroke Rehabilitation Patients and Their Relation to Hemiparesis Improvement Hideto Okazaki, MD, PhD,* Hidehiko Beppu, PhD,† Kenmei Mizutani, PhD,† Sayaka Okamoto, MD, PhD,* and Shigeru Sonoda, MD, PhD*
Predicting recovery from hemiparesis after stroke is important for rehabilitation. A few recent studies reported that the levels of some growth factors shortly after stroke were positively correlated with the clinical outcomes during the chronic phase. The aim of this study was to examine the relationships between the serum levels of growth factors (vascular endothelial growth factor [VEGF], insulin-like growth factor-I [IGF-I], and hepatocyte growth factor [HGF]) and improvement in hemiparesis in stroke patients who received rehabilitation in a postacute rehabilitation hospital. Subjects were 32 stroke patients (cerebral infarction: 21 and intracerebral hemorrhage [ICH]: 11). We measured serum levels of VEGF, IGF-I, and HGF and 5 items of the Stroke Impairment Assessment Set (SIAS) for hemiparesis on admission and at discharge. Age-matched healthy subjects (n 5 15) served as controls. Serum levels of VEGF and HGF in cerebral infarct patients on admission were higher than those in control subjects, and the serum levels of IGF-I in stroke patients were lower than those in controls. The level of HGF in ICH patients on admission was negatively correlated with gains in SIAS, and higher outliers in HGF concentration were correlated with lower gains in SIAS. Focusing on the extremely high levels of these factors may be a predictor of the low recovery from hemiparesis after stroke. Key Words: Growth factor—stroke—hemiparesis—rehabilitation. Ó 2014 by National Stroke Association
Introduction Stroke is one of the major diseases causing disabilities, such as hemiparesis or dysphagia. Hemiparesis is the major symptom from which stroke patients seek recovery. There are many recent reports of new treatments, including robot-assisted therapy and constraint-induced
From the *Department of Rehabilitation Medicine II, School of Medicine, Fujita Health University, Tsu Mie; and †Division of Biochemistry, Fujita Memorial Nanakuri Institute, Fujita Health University, Tsu Mie, Japan. Received October 31, 2013; revision received January 9, 2014; accepted January 16, 2014. Grant support: This study was supported by a Grants-in-Aid for Scientific Research No.1765017, No.1870046. Address correspondence to Hideto Okazaki, MD, PhD, Department of Rehabilitation Medicine II, Fujita Health University, School of Medicine, 424-1 Ootorichou Tsu Mie 514-1295, Japan. E-mail: [email protected]. 1052-3057/$ - see front matter Ó 2014 by National Stroke Association http://dx.doi.org/10.1016/j.jstrokecerebrovasdis.2014.01.015
therapy; however, recovery is still limited.1,2 Hence, it is important that the recovery from hemiparesis be accurately estimated, and rehabilitation strategies are designed based on the prognosis for recovery. However, it is difficult to predict to what degree the hemiparesis will improve. In clinical practice, it is not uncommon that stroke patients show more or less recovery from hemiparesis than physiatrists expected. Stroke patients requiring rehabilitation are usually admitted to the subacute rehabilitation hospital about 2-4 weeks after stroke onset in Japan.3 Therefore, if some data collected on admission to the rehabilitation hospital can be used to predict prognosis of hemiparesis, such data would be very useful for evaluation in a clinical context. It is known that growth factors contribute to neuroprotection and neurogenesis and, moreover, that some of these factors contribute not only to neurons but also to muscle cells as well.4-7 This led us to the hypothesis that higher levels of growth factors may lead to higher recovery from hemiparesis. Some researchers reported positive relationships between blood levels of growth
Journal of Stroke and Cerebrovascular Diseases, Vol. -, No. - (---), 2014: pp 1-6
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Table 1. Characteristics of each group Characteristic Age (y) Gender (male) Side of stroke (right) Handedness (right) Stroke subtype
Time from onset to admission (d) Length of stay (d) SOP on admission SOP on discharge
Cerebral infarction (n 5 21)
Intracerebral hemorrhage (n 5 11)
71.0 6 9.0 13 10 21 Large-artery atherosclerosis (n 5 7) Cardioembolism (n 5 6) Small-vessel occlusion (n 5 8) 31.7 6 16.2 70.5 6 17.2 10.5 6 7.1 14.3 6 6.7
58.5 6 14.0 6 7 11 Putamen (n 5 5) Thalamus (n 5 4) Frontal subcortex (n 5 2) 37.2 6 15.4 81.8 6 26.3 10.9 6 8.4 13.8 6 7.8
Abbreviation: SOP, sum of scores on 5 items of SIAS. Data were mean 6 SD.
factors at particular time points after stroke and clinical outcomes in ischemic stroke patients.8-12 Serum levels of vascular endothelial growth factor (VEGF) at acute stroke were proportional to improvement in National Institutes of Health Stroke Scale (NIHSS) scores after 3 months.8 Insulin-like growth factor-I (IGF-I) levels at the acute phase of stroke and after 3 months were positively correlated with improvements in the modified Rankin scale (mRS) between 3 and 24 months.9 IGF-I levels were positively correlated with gains in the Functional Independence Measure between admission and discharge10 and negatively correlated with mRS scores at 1-3 months and 12-15 months after stroke.11 Hepatocyte growth factor (HGF) levels within 3 hours of symptom onset were negatively correlated to NIHSS scores at 2 hours and 12 hours after onset.12 However, except for 1 study,10 there are no reports of serial changes in growth factors between admission and discharge from the rehabilitation hospital. The aim of this study was to evaluate serum levels of VEGF, IGF-I, and HGF in stroke patients undergoing rehabilitation in a postacute rehabilitation hospital and then to evaluate the relationships between the changes in serum growth factor levels on admission and at discharge and the clinical outcomes.
cause hemiparesis or motor weakness were excluded. The mean age was 66.7 6 12.4 years. The mean time from onset to admission was 33.6 6 15.9 days, and the mean hospitalization stay was 74.4 6 21.1 days. The strokes were because of cerebral infarction (CI) and intracerebral hemorrhage (ICH). See Table 1 for descriptions of the patients in these 2 groups. The CI strokes were because of large-artery atherosclerosis, cardioembolism, and small-vessel occlusion. The ICHs were located in the putamen, thalamus, and frontal subcortex. All the subjects underwent Full-time Integrated Treatment designed for rehabilitation patients during hospitalization.13 All patients had 40 minutes of physical therapy (gait training) and 40 minutes of occupational therapy (activity of daily living training) per day daily during hospitalization. The aim of the rehabilitation was improvement in daily living and walking, so the patients did not have any specific exercises for hemiparesis. Fifteen normal healthy volunteers (3 men and 12 women; mean age: 63.0 6 7.6 years) participated as control subjects for the serum growth factors. All subjects provided written, informed consent before participating in the study, and the ethics committee of our hospital approved all protocols.
Blood Samples and Clinical Outcomes
Methods Subjects Stroke patients whose admission is requested by other hospitals and for whom improvement is expected by the physician are admitted to rehabilitation hospitals from about 2-4 weeks after onset in Japan.3 Our subjects comprised 32 patients (19 men and 13 women) with stroke and hemiparesis hospitalized in the postacute rehabilitation ward of our hospital from July 2004 to October 2006. Patients who had a medical history of stroke, stroke involving the brain stem, cognitive dysfunction such as dementia, or other neurologic or muscular diseases that could
We measured serum levels of VEGF, IGF-I, and HGF on admission and at discharge (VEGF-Ad, VEGF-Dis, IGFI-Ad, IGF-I-Dis, HGF-Ad, and HGF-Dis). As these particular growth factors display diurnal fluctuations,14 medical laboratory technicians drew blood at the time of awakening in the morning (7:00 AM). VEGF, IGF-I, and HGF were measured by enzyme-linked immunosorbent assays (VEGF: BioSource Immunoassay Kit; IGF-I and HGF: R&D Quantikine Set). The severity of hemiparesis was evaluated by the Stroke Impairment Assessment Set (SIAS)15 on the same day as the blood withdrawals. SIAS is a 22-item evaluation of stroke impairment designed to comprehensively assess motor
GROWTH FACTORS IN STROKE REHABILITATION
functioning, sensory function, visuospatial perception, trunk control, grip strength, etc. Each item has either 4 or 6 grades, and higher points indicate higher function.15 The SIAS has high inter-rater reliability,16,17 high correlations with the NIHSS,18,19 significant correlation between its motor items and the Motricity Index or the Brunnstrom stage, and is more responsive to change than the Motricity Index, Brunnstrom stage, or NIHSS.19 For hemiparesis, the SIAS has 5 items (knee mouth, finger function, hip flexion, knee extension, and foot pat), and we used the sum of these 5 items (SOP) and the sound side grip strength (GS) as a measure of hemiparesis and muscle strength.
Statistical Analysis The number of patients in each of the CI and ICH subtypes was too small for statistical analyses, and so we compared values between the 2 major groups. The Kruskal–Wallis test was used to compare the serum levels of growth factors between the control subjects, and the patients at the time of admission and discharge, and when necessary, were followed by Mann–Whitney U tests for pairwise comparisons. The Wilcoxon signed rank test was used to compare the clinical outcomes between admission and discharge. Changes in SAIS scores were calculated by subtracting the SOP on admission (SOPAd) from SOP at discharge (SOP-Dis). The Spearman rank correlation coefficient was then used to investigate the relationship between growth factor levels on admission and changes in the SOP scores, growth factor levels on admission and SOP-Ad, and growth factor levels at discharge and SOP-Dis. These scatter plots were drawn and their characteristics investigated (IBMÒ; SPSS, ver21, Armonk, NY). The level of significance was set at P less than .05 for all analyses.
Results The comparisons of growth factor levels in control subjects and in stroke patients with CI and ICH on admission
Figure 1. Serum levels of VEGF, IGF-I, and HGF in control subjects and at admission and discharge in patients with strokes because of CB and CI. Data represent means (vertical bars) and 95% confidence intervals (error bars). Circles represent outliers (defined as outside the 95% confidence interval). *P , .05 (significantly different from control). Abbreviations: CB, cerebral bleeding; CI, cerebral infarction; HGF, hepatocyte growth factor; IGF-I, insulin-like growth factor-I; VEGF, vascular endothelial growth factor.
3
and at discharge are shown in Figure 1. VEGF and HGF levels in controls were lower than in patients, and VEGF-Ad and HGF-Ad levels were significantly higher in patients with CI than in control subjects (VEGF: P , .01, HGF: P 5 .04). IGF-I levels in all stroke patients were significantly lower than those in control subjects (P , .01 for all comparisons). The SOP and GS scores of patients with both stroke types were significantly higher at discharge than they were on admission (SOP for all patients: P , .01, GS with CI: P , .01, GS with ICH: P 5 .02; Fig 2). Scatter plots of growth factor levels and SOP scores showed that patients with higher outlier levels of VEGF tended to have higher SOP scores, whereas those with higher outlier levels of IGF-I and HGF tended to have lower SOP scores (Fig 3). Spearman rank correlation coefficients showed that HGF-Ad and increased SOP at discharge were significantly correlated in the cerebral bleeding stroke patients (R 5 2.71, P 5 .01; Fig 4). Furthermore, patients with higher outlier levels of the growth factors tended to show less improvement in SOP scores (Fig 4).
Discussion There are 2 major findings from this study: (1) that serum HGF levels on admission to a subacute rehabilitation hospital negatively correlate with improved SOP scores at discharge in hemiparetic patients with strokes because of ICH and (2) that patients with higher outlier levels of growth factors showed less improvement in the SOP scores at discharge. These findings suggest that extremely high levels of some serum growth factors on admission to rehabilitation are not necessarily positively associated with the recovery from hemiparesis after admission. We focused on the hospitalization of subacute rehabilitation patients, and so patients with mild and severe strokes (eg, only mild motor weakness without daily living limitation or loss of consciousness) were excluded from this study. Because of comorbidities and different severities, the patients had variable latencies to admission and different lengths of rehabilitation. This is realistic as
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Figure 2. Clinical outcomes of patients with strokes because of CB and CI at admission and discharge. Data represent means (vertical bars) and 95% confidence intervals (error bars). Circles represent outliers (defined as outside the 95% confidence interval). *P , .05 (significantly different from control). Abbreviations: CB, cerebral bleeding; CI, cerebral infarction.
it is the general situation in subacute rehabilitation hospitals. In this study, we used 5 items (knee mouth, finger function, hip flexion, knee extension, and foot pat) of the SIAS as a measure of the severity of hemiparesis. The SIAS has high enough reliability, validity, and sensitivity to detect changes in hemiparesis.16,17 Moreover, the SIAS is highly correlated with the NIHSS18 and is more responsive to changes than the NIHSS.19 Therefore, using SIAS was approximate for this study. We included not only patients with ischemic stroke, on, which previous reports focused, but also those with ICH. Our study also examined patients during the subacute phase of their
strokes in contrast to those during the acute phase examined in previous studies. Previous studies showed that the levels of VEGF and HGF were elevated both after ischemic and hemorrhagic stroke,20,21 whereas our results showed significantly increased levels of VEGF and HGF in ischemic stroke patients and only a tendency to be increased in hemorrhagic patients. There is also a discrepancy between our findings of lower levels of IGF-I in stroke patients and the results of previous studies showing that serum levels of IGF-I at 3 months were not lower than controls9 and that serum levels of IGF-I did not change
Figure 3. Correlations between serum growth factor levels and the sum of the 5 paresis items in the SIAS measured at admission (SOP-Ad) and at discharge (SOP-Dis). Circles represent data from patients with strokes because of CB and crosses those because of CI. Abbreviations: Ad, admission; CB, cerebral bleeding; CI, cerebral infarction; Dis, discharge; HGF, hepatocyte growth factor; IGF-I, insulin-like growth factor-I; VEGF, vascular endothelial growth factor.
GROWTH FACTORS IN STROKE REHABILITATION
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Figure 4. Correlations between serum growth factor levels and improvement in SOP scores at discharge. Circles represent data from patients with strokes because of CB and crosses those because of CI. Abbreviations: CB, cerebral bleeding; CI, cerebral infarction; HGF, hepatocyte growth factor; IGF-I, insulinlike growth factor-I; SOP, sum of scores on 5 items of SIAS; VEGF, vascular endothelial growth factor.
after hemorrhagic stroke.22 Why were IGF-I levels low in our study? The expression of IGF-I is affected by at least 2 factors. One is physical activity. Ardawi et al23 reported that higher physical activity led to higher serum levels of IGF-I. Lower knee extension strength was associated with lower serum IGF-I 1 month after stroke onset.24,25 Remaining sedentary in an acute hospital for 1 month is long enough to make muscles weak, as demonstrated by the fact that almost of our subjects could not walk at admission, in contrast with the fact that more than 60% of ischemic stroke patients in Bondanelli study had mRS scores of 2 or less 1-3 months after stroke.11 The other factor affecting IGF-I levels is malnutrition. Some stroke patients have dysphagia that may induce malnutrition. Caregalo et al26 reported that malnutrition decreased serum IGF-I. Hence, immobilization and malnutrition may have contributed to our results. How may we explain that high HGF levels were related to severe paralysis? Because growth factors contribute to neuroprotection and neurogenesis, perhaps something such as brain lesion size was responsible. The serum levels of VEGF during acute stroke were reported to be correlated with infarct volume,8 and chronic high serum IGF-I correlates with increased brain infarct size in middle cerebral artery occlusion mice.27 Furthermore, massive brain damage tends to cause large damage in motor pathways, resulting in severe paralysis with little hope of recovery. Hence, our results may possibly be explained by the hypothesis that large brain damage causes both elevation of growth factors in serum and restriction of the degree of paralysis to the severe level. Both these hypotheses can be tested in future studies. There were different results between the various subtypes of stroke patients. The patients with CI had three subtypes, large-artery atherosclerosis, cardioembolism,
and small-vessel occlusion. Large-artery artherosclerosis and cardioembolism usually have larger stroke lesions than those with small-vessel occlusion.28 On the other hand, four patients in the ICH group had hematoma removal, and so it is possible that the size of the lesions were quite variable and much larger in the infarction group than in the hemorrhage group. This difference might have led to the different results found in this study. The influence of lesion size on growth factor levels and their relation to rehabilitation can be tested in future studies. Because we focused on hemiparesis, these results do not comprehensively represent all after-stroke conditions, and the sample size was also a bit small. To test some of the possible hypotheses that may explain our results, in the future we should evaluate the sizes and locations of stroke lesions, including the data at onset. In conclusion, the low IGF-I levels that we observed in these patients may be explained by low physical activity and malnutrition during the acute phase of stroke, and the serum levels of VEGF, IGF-I, and HGF may be influenced by the stroke lesion size. Hence, focusing on the extremely high levels of these factors may predict a low recovery of hemiparesis from stroke.
References 1. Viana R, Teasell R. Barriers to the implementation of constraint-induced movement therapy into practice. Top Stroke Rehabil 2012;19:104-114. 2. Krebs HI, Hogan N. Robotic therapy: the tipping point. Am J Phys Med Rehabil 2012;91(11 Suppl 3):S290-S297. 3. Miyai I, Sonoda S, Nagai S, Takayama Y, Inoue Y, Kakehi A, et al. Results of new policies for inpatient rehabilitation coverage in Japan. Neurorehabil Neural Repair 2011;25:540-547. 4. Puche JE, Castilla-Cort azar I. Human conditions of insulin-like growth factor-I (IGF-I) deficiency. J Transl Med 2012;10:224.
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6 5. Shang J, Deguchi K, Ohta Y, Liu N, Zhang X, Tian F, et al. Strong neurogenesis, angiogenesis, synaptogenesis, and antifibrosis of hepatocyte growth factor in rats brain after transient middle cerebral artery occlusion. J Neurosci Res 2011;89:86-95. 6. Nakamura T, Sakai K, Matsumoto K. Hepatocyte growth factor twenty years on: Much more than a growth factor. J Gastroenterol Hepatol 2011;26(Suppl 1):188-202. 7. Rosenstein JM, Krum JM, Ruhrberg C. VEGF in the nervous system. Organogenesis 2010;6:107-114. 8. Lee SC, Lee KY, Kim YJ, Kim SH, Koh SH, Lee YJ. Serum VEGF levels in acute ischaemic strokes are correlated with long-term prognosis. Eur J Neurol 2010;17:45-51. 9. Aberg D, Jood K, Blomstrand C, Jern C, Nilsson M, Isgaard J, et al. Serum IGF-I levels correlate to improvement of functional outcome after ischemic stroke. J Clin Endocrinol Metab 2011;96:E1055-E1064. 10. Bondanelli M, Ambrosio MR, Onofri A, Bergonzoni A, Lavezzi S, Zatelli MC, et al. Predictive value of circulating insulin-like growth factor I levels in ischemic stroke outcome. J Clin Endocrinol Metab 2006;91:3928-3934. 11. Bondanelli M, Ambrosio MR, Carli A, Bergonzoni A, Bertocchi A, Zatelli MC, et al. Predictors of pituitary dysfunction in patients surviving ischemic stroke. J Clin Endocrinol Metab 2010;95:4660-4668. 12. Navarro-Sobrino M, Rosell A, Hern andez-Guillamon M, Penalba A, Boada C, Domingues-Montanari S, et al. A large screening of angiogenesis biomarkers and their association with neurological outcome after ischemic stroke. Atherosclerosis 2011;216:205-211. 13. Sonoda S, Saitoh E, Nagai S, Kawakita M, Kanada Y. Fulltime integrated treatment program, a new system for stroke rehabilitation in Japan: comparison with conventional rehabilitation. Am J Phys Med Rehabil 2004; 83:88-93. 14. Itoh E, Iida K, del Rincon JP, Kim DS, Thorner MO. Diurnal variation in growth hormone receptor messenger ribonucleic acid in liver and skeletal muscle of lit/1 and lit/lit mice. Endocr J 2004;51:529-535. 15. Chino N, Sonoda S, Domen K, Saitoh E, Kimura A. Stroke Impairment Assessment Set (SIAS)—a new evaluation instrument for stroke patients. Jpn J Rehabil Med 1994; 31:119-125. 16. Sonoda S. Reliability and validity of Stroke Impairment Assessment Set (SIAS) (2): the items comprise the trunk, higher cortical function, and sensory function and effectiveness as outcome predictor. Jpn J Rehabil Med 1995; 32:123-132. 17. Domen K. Reliability and validity of Stroke Impairment Assessment Set (SIAS) (1): items of affected-side motor function, muscle tone, deep tendon reflex, and
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unaffected-side function. Jpn J Rehabil Med 1995; 32:113-122. Seki M, Hase K, Takahashi H, Liu M. Comparison of three instruments to assess changes of motor impairment in acute hemispheric stroke: the Stroke Impairment Assessment Set (SIAS), the National Institutes of Health Stroke Scale (NIHSS) and the Canadian Neurological Scale (CNS). Disabil Rehabil 2013 [epub ahead of print]. Liu M, Chino N, Tuji T, Masakado Y, Hase K, Kimura A. Psychometric properties of the Stroke Impairment Assessment Set (SIAS). Neurorehabil Neural Repair 2002;16:339-351. Slevin M, Krupinski J, Slowik A, Kumar P, Szczudlik A, Gaffney J. Serial measurement of vascular endothelial growth factor and transforming growth factor-beta1 in serum of patients with acute ischemic stroke. Stroke 2000;31:1863-1870. Paczkowska E, Go1a˛b-Janowska M, Bajer-Czajkowska A, Machali nska A, Ustianowski P, Rybicka M, et al. Increased circulating endothelial progenitor cells in patients with haemorrhagic and ischaemic stroke: the role of endothelin-1. J Neurol Sci 2013;325:90-99. Huttner HB, Kiphuth IC, Teuber L, L€ ucking H, Kloska SP, Staykov D, et al. Neuroendocrine changes in patients with spontaneous supratentorial intracerebral hemorrhage. Neurocrit Care 2013;18:39-44. Ardawi MS, Rouzi AA, Qari MH. Physical activity in relation to serum sclerostin, insulin-like growth factor-1, and bone turnover markers in healthy premenopausal women: a cross-sectional and a longitudinal study. J Clin Endocrinol Metab 2012;97:3691-3699. Cappola AR, Bandeen-Roche K, Wand GS, Volpato S, Fried LP. Association of IGF-I levels with muscle strength and mobility in older women. J Clin Endocrinol Metab 2001;86:4139-4146. Kostka T, Arsac LM, Patricot MC, Berthouze SE, Lacour JR, Bonnefoy M. Leg extensor power and dehydroepiandrosterone sulfate, insulin-like growth factor-I and testosterone in healthy active elderly people. Eur J Appl Physiol 2000;82:83-90. Caregaro L, Favaro A, Santonastaso P, Alberino F, Di Pascoli L, Nardi M, et al. Insulin-like growth factor 1 (IGF-1), a nutritional marker in patients with eating disorders. Clin Nutr 2001;20:251-257. Endres M, Piriz J, Gertz K, Harms C, Meisel A, Kronenberg G, et al. Serum insulin-like growth factor I and ischemic brain injury. Brain Res 2007;1185:328-335. Adams HP, Bendixen BH, Kappelle LJ, Biller J, Love BB, Gordon DL, et al. Classification of subtype of acute ischemic stroke. Definitions for use in a multicenter clinical trial. TOAST. Trial of Org 10172 in Acute Stroke Treatment. Stroke 1993;24:35-41.
Predicting recovery from hemiparesis after stroke is important for rehabilitation. A few recent studies reported that the levels of some growth factors shortly after stroke were positively correlated with the clinical outcomes during the chronic phase. The aim of this study was to examine the relationships between the serum levels of growth factors (vascular endothelial growth factor [VEGF], insulin-like growth factor-I [IGF-I], and hepatocyte growth factor [HGF]) and improvement in hemiparesis in stroke patients who received rehabilitation in a postacute rehabilitation hospital. Subjects were 32 stroke patients (cerebral infarction: 21 and intracerebral hemorrhage [ICH]: 11). We measured serum levels of VEGF, IGF-I, and HGF and 5 items of the Stroke Impairment Assessment Set (SIAS) for hemiparesis on admission and at discharge. Age-matched healthy subjects (n 5 15) served as controls. Serum levels of VEGF and HGF in cerebral infarct patients on admission were higher than those in control subjects, and the serum levels of IGF-I in stroke patients were lower than those in controls. The level of HGF in ICH patients on admission was negatively correlated with gains in SIAS, and higher outliers in HGF concentration were correlated with lower gains in SIAS. Focusing on the extremely high levels of these factors may be a predictor of the low recovery from hemiparesis after stroke. Key Words: Growth factor—stroke—hemiparesis—rehabilitation. Ó 2014 by National Stroke Association
Introduction Stroke is one of the major diseases causing disabilities, such as hemiparesis or dysphagia. Hemiparesis is the major symptom from which stroke patients seek recovery. There are many recent reports of new treatments, including robot-assisted therapy and constraint-induced
From the *Department of Rehabilitation Medicine II, School of Medicine, Fujita Health University, Tsu Mie; and †Division of Biochemistry, Fujita Memorial Nanakuri Institute, Fujita Health University, Tsu Mie, Japan. Received October 31, 2013; revision received January 9, 2014; accepted January 16, 2014. Grant support: This study was supported by a Grants-in-Aid for Scientific Research No.1765017, No.1870046. Address correspondence to Hideto Okazaki, MD, PhD, Department of Rehabilitation Medicine II, Fujita Health University, School of Medicine, 424-1 Ootorichou Tsu Mie 514-1295, Japan. E-mail: [email protected]. 1052-3057/$ - see front matter Ó 2014 by National Stroke Association http://dx.doi.org/10.1016/j.jstrokecerebrovasdis.2014.01.015
therapy; however, recovery is still limited.1,2 Hence, it is important that the recovery from hemiparesis be accurately estimated, and rehabilitation strategies are designed based on the prognosis for recovery. However, it is difficult to predict to what degree the hemiparesis will improve. In clinical practice, it is not uncommon that stroke patients show more or less recovery from hemiparesis than physiatrists expected. Stroke patients requiring rehabilitation are usually admitted to the subacute rehabilitation hospital about 2-4 weeks after stroke onset in Japan.3 Therefore, if some data collected on admission to the rehabilitation hospital can be used to predict prognosis of hemiparesis, such data would be very useful for evaluation in a clinical context. It is known that growth factors contribute to neuroprotection and neurogenesis and, moreover, that some of these factors contribute not only to neurons but also to muscle cells as well.4-7 This led us to the hypothesis that higher levels of growth factors may lead to higher recovery from hemiparesis. Some researchers reported positive relationships between blood levels of growth
Journal of Stroke and Cerebrovascular Diseases, Vol. -, No. - (---), 2014: pp 1-6
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Table 1. Characteristics of each group Characteristic Age (y) Gender (male) Side of stroke (right) Handedness (right) Stroke subtype
Time from onset to admission (d) Length of stay (d) SOP on admission SOP on discharge
Cerebral infarction (n 5 21)
Intracerebral hemorrhage (n 5 11)
71.0 6 9.0 13 10 21 Large-artery atherosclerosis (n 5 7) Cardioembolism (n 5 6) Small-vessel occlusion (n 5 8) 31.7 6 16.2 70.5 6 17.2 10.5 6 7.1 14.3 6 6.7
58.5 6 14.0 6 7 11 Putamen (n 5 5) Thalamus (n 5 4) Frontal subcortex (n 5 2) 37.2 6 15.4 81.8 6 26.3 10.9 6 8.4 13.8 6 7.8
Abbreviation: SOP, sum of scores on 5 items of SIAS. Data were mean 6 SD.
factors at particular time points after stroke and clinical outcomes in ischemic stroke patients.8-12 Serum levels of vascular endothelial growth factor (VEGF) at acute stroke were proportional to improvement in National Institutes of Health Stroke Scale (NIHSS) scores after 3 months.8 Insulin-like growth factor-I (IGF-I) levels at the acute phase of stroke and after 3 months were positively correlated with improvements in the modified Rankin scale (mRS) between 3 and 24 months.9 IGF-I levels were positively correlated with gains in the Functional Independence Measure between admission and discharge10 and negatively correlated with mRS scores at 1-3 months and 12-15 months after stroke.11 Hepatocyte growth factor (HGF) levels within 3 hours of symptom onset were negatively correlated to NIHSS scores at 2 hours and 12 hours after onset.12 However, except for 1 study,10 there are no reports of serial changes in growth factors between admission and discharge from the rehabilitation hospital. The aim of this study was to evaluate serum levels of VEGF, IGF-I, and HGF in stroke patients undergoing rehabilitation in a postacute rehabilitation hospital and then to evaluate the relationships between the changes in serum growth factor levels on admission and at discharge and the clinical outcomes.
cause hemiparesis or motor weakness were excluded. The mean age was 66.7 6 12.4 years. The mean time from onset to admission was 33.6 6 15.9 days, and the mean hospitalization stay was 74.4 6 21.1 days. The strokes were because of cerebral infarction (CI) and intracerebral hemorrhage (ICH). See Table 1 for descriptions of the patients in these 2 groups. The CI strokes were because of large-artery atherosclerosis, cardioembolism, and small-vessel occlusion. The ICHs were located in the putamen, thalamus, and frontal subcortex. All the subjects underwent Full-time Integrated Treatment designed for rehabilitation patients during hospitalization.13 All patients had 40 minutes of physical therapy (gait training) and 40 minutes of occupational therapy (activity of daily living training) per day daily during hospitalization. The aim of the rehabilitation was improvement in daily living and walking, so the patients did not have any specific exercises for hemiparesis. Fifteen normal healthy volunteers (3 men and 12 women; mean age: 63.0 6 7.6 years) participated as control subjects for the serum growth factors. All subjects provided written, informed consent before participating in the study, and the ethics committee of our hospital approved all protocols.
Blood Samples and Clinical Outcomes
Methods Subjects Stroke patients whose admission is requested by other hospitals and for whom improvement is expected by the physician are admitted to rehabilitation hospitals from about 2-4 weeks after onset in Japan.3 Our subjects comprised 32 patients (19 men and 13 women) with stroke and hemiparesis hospitalized in the postacute rehabilitation ward of our hospital from July 2004 to October 2006. Patients who had a medical history of stroke, stroke involving the brain stem, cognitive dysfunction such as dementia, or other neurologic or muscular diseases that could
We measured serum levels of VEGF, IGF-I, and HGF on admission and at discharge (VEGF-Ad, VEGF-Dis, IGFI-Ad, IGF-I-Dis, HGF-Ad, and HGF-Dis). As these particular growth factors display diurnal fluctuations,14 medical laboratory technicians drew blood at the time of awakening in the morning (7:00 AM). VEGF, IGF-I, and HGF were measured by enzyme-linked immunosorbent assays (VEGF: BioSource Immunoassay Kit; IGF-I and HGF: R&D Quantikine Set). The severity of hemiparesis was evaluated by the Stroke Impairment Assessment Set (SIAS)15 on the same day as the blood withdrawals. SIAS is a 22-item evaluation of stroke impairment designed to comprehensively assess motor
GROWTH FACTORS IN STROKE REHABILITATION
functioning, sensory function, visuospatial perception, trunk control, grip strength, etc. Each item has either 4 or 6 grades, and higher points indicate higher function.15 The SIAS has high inter-rater reliability,16,17 high correlations with the NIHSS,18,19 significant correlation between its motor items and the Motricity Index or the Brunnstrom stage, and is more responsive to change than the Motricity Index, Brunnstrom stage, or NIHSS.19 For hemiparesis, the SIAS has 5 items (knee mouth, finger function, hip flexion, knee extension, and foot pat), and we used the sum of these 5 items (SOP) and the sound side grip strength (GS) as a measure of hemiparesis and muscle strength.
Statistical Analysis The number of patients in each of the CI and ICH subtypes was too small for statistical analyses, and so we compared values between the 2 major groups. The Kruskal–Wallis test was used to compare the serum levels of growth factors between the control subjects, and the patients at the time of admission and discharge, and when necessary, were followed by Mann–Whitney U tests for pairwise comparisons. The Wilcoxon signed rank test was used to compare the clinical outcomes between admission and discharge. Changes in SAIS scores were calculated by subtracting the SOP on admission (SOPAd) from SOP at discharge (SOP-Dis). The Spearman rank correlation coefficient was then used to investigate the relationship between growth factor levels on admission and changes in the SOP scores, growth factor levels on admission and SOP-Ad, and growth factor levels at discharge and SOP-Dis. These scatter plots were drawn and their characteristics investigated (IBMÒ; SPSS, ver21, Armonk, NY). The level of significance was set at P less than .05 for all analyses.
Results The comparisons of growth factor levels in control subjects and in stroke patients with CI and ICH on admission
Figure 1. Serum levels of VEGF, IGF-I, and HGF in control subjects and at admission and discharge in patients with strokes because of CB and CI. Data represent means (vertical bars) and 95% confidence intervals (error bars). Circles represent outliers (defined as outside the 95% confidence interval). *P , .05 (significantly different from control). Abbreviations: CB, cerebral bleeding; CI, cerebral infarction; HGF, hepatocyte growth factor; IGF-I, insulin-like growth factor-I; VEGF, vascular endothelial growth factor.
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and at discharge are shown in Figure 1. VEGF and HGF levels in controls were lower than in patients, and VEGF-Ad and HGF-Ad levels were significantly higher in patients with CI than in control subjects (VEGF: P , .01, HGF: P 5 .04). IGF-I levels in all stroke patients were significantly lower than those in control subjects (P , .01 for all comparisons). The SOP and GS scores of patients with both stroke types were significantly higher at discharge than they were on admission (SOP for all patients: P , .01, GS with CI: P , .01, GS with ICH: P 5 .02; Fig 2). Scatter plots of growth factor levels and SOP scores showed that patients with higher outlier levels of VEGF tended to have higher SOP scores, whereas those with higher outlier levels of IGF-I and HGF tended to have lower SOP scores (Fig 3). Spearman rank correlation coefficients showed that HGF-Ad and increased SOP at discharge were significantly correlated in the cerebral bleeding stroke patients (R 5 2.71, P 5 .01; Fig 4). Furthermore, patients with higher outlier levels of the growth factors tended to show less improvement in SOP scores (Fig 4).
Discussion There are 2 major findings from this study: (1) that serum HGF levels on admission to a subacute rehabilitation hospital negatively correlate with improved SOP scores at discharge in hemiparetic patients with strokes because of ICH and (2) that patients with higher outlier levels of growth factors showed less improvement in the SOP scores at discharge. These findings suggest that extremely high levels of some serum growth factors on admission to rehabilitation are not necessarily positively associated with the recovery from hemiparesis after admission. We focused on the hospitalization of subacute rehabilitation patients, and so patients with mild and severe strokes (eg, only mild motor weakness without daily living limitation or loss of consciousness) were excluded from this study. Because of comorbidities and different severities, the patients had variable latencies to admission and different lengths of rehabilitation. This is realistic as
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Figure 2. Clinical outcomes of patients with strokes because of CB and CI at admission and discharge. Data represent means (vertical bars) and 95% confidence intervals (error bars). Circles represent outliers (defined as outside the 95% confidence interval). *P , .05 (significantly different from control). Abbreviations: CB, cerebral bleeding; CI, cerebral infarction.
it is the general situation in subacute rehabilitation hospitals. In this study, we used 5 items (knee mouth, finger function, hip flexion, knee extension, and foot pat) of the SIAS as a measure of the severity of hemiparesis. The SIAS has high enough reliability, validity, and sensitivity to detect changes in hemiparesis.16,17 Moreover, the SIAS is highly correlated with the NIHSS18 and is more responsive to changes than the NIHSS.19 Therefore, using SIAS was approximate for this study. We included not only patients with ischemic stroke, on, which previous reports focused, but also those with ICH. Our study also examined patients during the subacute phase of their
strokes in contrast to those during the acute phase examined in previous studies. Previous studies showed that the levels of VEGF and HGF were elevated both after ischemic and hemorrhagic stroke,20,21 whereas our results showed significantly increased levels of VEGF and HGF in ischemic stroke patients and only a tendency to be increased in hemorrhagic patients. There is also a discrepancy between our findings of lower levels of IGF-I in stroke patients and the results of previous studies showing that serum levels of IGF-I at 3 months were not lower than controls9 and that serum levels of IGF-I did not change
Figure 3. Correlations between serum growth factor levels and the sum of the 5 paresis items in the SIAS measured at admission (SOP-Ad) and at discharge (SOP-Dis). Circles represent data from patients with strokes because of CB and crosses those because of CI. Abbreviations: Ad, admission; CB, cerebral bleeding; CI, cerebral infarction; Dis, discharge; HGF, hepatocyte growth factor; IGF-I, insulin-like growth factor-I; VEGF, vascular endothelial growth factor.
GROWTH FACTORS IN STROKE REHABILITATION
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Figure 4. Correlations between serum growth factor levels and improvement in SOP scores at discharge. Circles represent data from patients with strokes because of CB and crosses those because of CI. Abbreviations: CB, cerebral bleeding; CI, cerebral infarction; HGF, hepatocyte growth factor; IGF-I, insulinlike growth factor-I; SOP, sum of scores on 5 items of SIAS; VEGF, vascular endothelial growth factor.
after hemorrhagic stroke.22 Why were IGF-I levels low in our study? The expression of IGF-I is affected by at least 2 factors. One is physical activity. Ardawi et al23 reported that higher physical activity led to higher serum levels of IGF-I. Lower knee extension strength was associated with lower serum IGF-I 1 month after stroke onset.24,25 Remaining sedentary in an acute hospital for 1 month is long enough to make muscles weak, as demonstrated by the fact that almost of our subjects could not walk at admission, in contrast with the fact that more than 60% of ischemic stroke patients in Bondanelli study had mRS scores of 2 or less 1-3 months after stroke.11 The other factor affecting IGF-I levels is malnutrition. Some stroke patients have dysphagia that may induce malnutrition. Caregalo et al26 reported that malnutrition decreased serum IGF-I. Hence, immobilization and malnutrition may have contributed to our results. How may we explain that high HGF levels were related to severe paralysis? Because growth factors contribute to neuroprotection and neurogenesis, perhaps something such as brain lesion size was responsible. The serum levels of VEGF during acute stroke were reported to be correlated with infarct volume,8 and chronic high serum IGF-I correlates with increased brain infarct size in middle cerebral artery occlusion mice.27 Furthermore, massive brain damage tends to cause large damage in motor pathways, resulting in severe paralysis with little hope of recovery. Hence, our results may possibly be explained by the hypothesis that large brain damage causes both elevation of growth factors in serum and restriction of the degree of paralysis to the severe level. Both these hypotheses can be tested in future studies. There were different results between the various subtypes of stroke patients. The patients with CI had three subtypes, large-artery atherosclerosis, cardioembolism,
and small-vessel occlusion. Large-artery artherosclerosis and cardioembolism usually have larger stroke lesions than those with small-vessel occlusion.28 On the other hand, four patients in the ICH group had hematoma removal, and so it is possible that the size of the lesions were quite variable and much larger in the infarction group than in the hemorrhage group. This difference might have led to the different results found in this study. The influence of lesion size on growth factor levels and their relation to rehabilitation can be tested in future studies. Because we focused on hemiparesis, these results do not comprehensively represent all after-stroke conditions, and the sample size was also a bit small. To test some of the possible hypotheses that may explain our results, in the future we should evaluate the sizes and locations of stroke lesions, including the data at onset. In conclusion, the low IGF-I levels that we observed in these patients may be explained by low physical activity and malnutrition during the acute phase of stroke, and the serum levels of VEGF, IGF-I, and HGF may be influenced by the stroke lesion size. Hence, focusing on the extremely high levels of these factors may predict a low recovery of hemiparesis from stroke.
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