Zhou Mengchao 1,Geng Deqin 2,Wang Dunjing 2 ,Mu Yingfeng 2,Dong Jing 1,Lin Xiaoqian1.
1. The Graduate School, Xuzhou Medical University
2.Department of Neurology，Affiliated Hospital of Xuzhou Medical University
Abstract
Objective
To investigate the correlation between serum Corin protein levels and cerebral small vessel disease （CSVD）.To elucidate the pathophysiological roles of Corin in regulating CSVD.
Methods
415 cases of patients，ranging from 45 to 75 years，were randomly selected from Department of Neurology，Affiliated Hospital of Xuzhou Medical University，from April 2015 to November 2015. They were divided into cerebral small vessel disease group（CSVD）and non-cerebral vascular disease group（control），according to the inclusion and exclusion criteria. A total of 415 cases，were enrolled，including 279 cases in the CSVD group，136 cases in the control group. Medical history and general information were collected，as well as laboratory parameters. Brain MRI（including T1，T2，DWI，FLAIR sequences）were done on the patients. The cerebral small vessel diseases were divided into Mild，Moderate and Severe groups，respectively according to the degree of white matter lesions and the lacune numbers of brain. And serum levels of soluble Corin protein（unit ng/ml）were measured using the enzyme-linked immunosorbent assay（ELISA）. Statistical analysis was done by SPSS 19.0. Serum Corin levels and general information for the patients in the CSVD group and the control group were compared to determine the effects of serum Corin levels，as well as other risk factors，on the incidence of CSVD. The subgroup studies of CSVD group were done to determine the relationships between the serum Corin levels and the disease severity evaluations and the clinical type classification. For CSVD diseases, Serum Corin levels were compared between hypertension group and non-hypertension group. There was also a comparision between women and men，for serum Corin levels. Spearman correlation analysis method was used to determine the relationship between the the serum Corin levels and serum LDL levels，as well as age.
Results
1.Serum Corin levels in the CSVD group were lower than those in the control group（0.589±0.439 ng/ml VS 0.870±0.522 ng/ml）, there were statistically significant（P<0.05）.
2.Multivariate logistic regression analysis results showed that lower serum Corin levels，higher age，hypertension，higher cystatin C are independent risk factors for the incidence of CSVD.
3.For patients in the different stages of CSVD : Serum levels of soluble Corin protein were dropping from Mild WML group to Moderate WML group to Severe WML group（0.742±0.393 ng/ml to 0.575±0.467 ng/ml to 0.508±0.420 ng/ml）..Samely, serum Corin levels reduced while the lesion number of ALI and lacune raising（0.696±0.412 ng/ml to 0.543±0.454ng/ml to 0.494±0.409ng/ml）.
4.For patients with the different clinical manifestation of CSVD: Serum Corin levels in the lacunar stroke group were lower than those in the asymptomatic CSVD group（0.539±0.417 ng/ml VS 0.650±0.459 ng/ml）, there were statistically significant（P<0.05）
5.In the CSVD group，women had lower Corin levels（0.528±0.403 ng/ml VS 0.640±0.463 ng/ml）, there were statistically significant（P<0.05）；the age had no effect on the serum Corin levels（P>0.05）.
6.For Serum Corin levels of CSVD deseases，there was no statistical difference between hypertension group and non-hypertension group（0.559±0.480 ng/ml VS 0.622±0.387 ng/ml，P>0.05）.Spearman correlation analysis method showed that serum LDL levels were related to serum Corin levels，in CSVD deseases（r=0.120、P<0.05）.
Conclusion
1.Serum Corin levels were independently correlated to CSVD. Corin might be involved in the pathogenesis of CSVD and could be used as a biomarker..
2.The serum Corin levels may provide the basis and forecast for the disease severity evaluations.
3.The serum Corin levels may also provide the basis and forecast for the lacunar stroke.
4.The mechanisms behind might be the feedback increase of Corin while LDL raising.
Key words
cerebral small vessel disease；Corin；acute lacunar infarction；lacune；white matter lesion
Cerebral small vessel including small perforators of the brain artery and small arteries, capillaries and venules, which constitutes the basic unit of the brain blood supply, maintain plays an important role of brain function. Small brain vascular disease (CSVD) refers to the small vascular lesions caused by the clinical, imaging and pathological syndrome, including all kinds of the cause caused by a variety of performance [1]. With the popularity of neural imaging examination and the arrival of an aging population, the detection rate of CSVD is rising.The occurrence and development mechanism and its prevention treatment strategy research is one of the important tasks of the current domestic neural science [2].
Corin is a kind of type II transmembrane serine protease, soluble Corin in the blood is very stable and easy to detect. The study found that there were close relationship betweenCorin and high blood pressure, heart failure, renal insufficiency [3-5].A new study shows that serumCorin levels of patients with acute cerebral apoplexy were significantly reduced[6]. Research on the correlation between serum corin and small brain vascular disease has not yet been reported.Our research, based on the small brain vascular disease patients with normal cranial MRI of cerebrovascular patients as control group, was designed to explore the relationship between serum corin levels with small brain vascular disease.
1.Materials and methods
1.1 Participants
Patients，ranging from 45 to 75 years，were randomly selected from Department of Neurology，Affiliated Hospital of Xuzhou Medical University，from April 2015 to November 2015. They were divided into cerebral small vessel disease group（CSVD）and non-cerebral vascular disease group（control），according to the inclusion and exclusion criteria.
1.1.1；The inclusion and exclusion criteria of the CSVD group：
CSVD diagnosis with reference to the latest version of the "C
erebral small vessel disease diagnosis and expert consensus"
[7].Imaging findings of brain white matter lesions (WMLs) with cerebral small lacuna (including ALI and acute).The clinical manifestations including lacunar syndrome and nonspecific symptoms such as dizziness, headache, memory declining. Eliminate imaging change caused by specific reasons such as multiple sclerosis, carbon monoxide poisoning and severe sleep apnea syndrome; Eliminate artery atherosclerosis, cardiac embolism, and other small vessel disease caused by fresh and pulmonary cavity; Rule out > 15 mm diameter ischemic lesions; Exclusion of thrombolysis therapy; Exclude vascular cognitive dysfunction and/or dementia patients; Eliminate coma patients; Eliminate craniocerebral trauma, nervous system tumor diseases; Exclusion of severe heart failure, acute myocardial infarction, unstable angina, patients with aortic stenosis; Exclusion of patients with severe liver and kidney insufficiency.
1.1.2； The inclusion and exclusion criteria of the control group：Selecting the non-cerebral vascular diseasers over the same period as the control.With clinical manifestations such as dizziness, headache, facial nerve palsy;Without nerve dysfunction;Without positive signs of nervous system;Without history of cerebrovascular disease;Without abnormal head MRI imageing. Exclusion criteria was followed by CSVD group.
1.2 Clinical data
1.2.1General situation and history of collecting, the project including: gender, age, smoking history, drinking history, history of hypertension, diabetes mellitus, heart disease, blood pressure on admission.
1.2.2Laboratory testing, including FBG, HbAlc, TC,TG, LDL - c, HDL - c,APOA1, APOB , LP - A, UA ,Cr, CysC, TSHand Hcy.
1.2.3 Imaging examination: Head MRI of 415 patients were done(including the T1, T2, DWI, and FLAIR).There was no abnormal MRI image in the control group.There were both white matter lesions (WMLs) and cerebral small lacuna in the CSVD group.According to the Fazekas scale evaluation
[8], cerebral white matter lesions can be divided into 1-3 level;Referencing to foreign literature
[9,10], according to the number of small lacunar ,there were 3 levels : level 1 / mild (1 ~ 3) ;level 2 / moderate (4 ~ 10) ;level 3 / moderate (> 10). Two kinds of classification simultaneously.
1.2.4 Determination of serum corin: Using the method of enzyme-linked immunosorbent (ELISA) to detect serum level of soluble corin, unit ng/ml. Kits were bought from wuhan USCN company. Steps were done strictly according to the instructions.
1.3 Statistical treatment
SPSS19.0 software was used to analysis. This study sample size is big (n = 415), the measurement data were described by mean and standard deviation , the comparison between the two groups was done bytwo independent sample t-test , multiple comparison using One - way ANOVA; The count data were described by rate (%), using the chi square test comparison; Multivariate analysis using stepwise Logistic regression analysis; Correlation analysis were done by Pearsonor Spearman correlation test. Using
α=0.05 as inspection standard,
P < 0.05 for the difference was statistically significant.
-
Results
2.1 The general data and laboratory indexes of control group and CSVD group are shown in table 1;
2.2 Single factor analysis of risk factors for CSVD.
Proportion of male in CSVD group and the control group were 53.8% (150/279) and 53.8% (71/136), there was no statistical difference (P > 0.05). The average age of the CSVD group is higher than the control group (62.1±7.7years, versus 54.4±7.0), the difference was statistically significant (P < 0.05). Compared with control group, CSVD group had a higher prevalence of hypertension, diabetes prevalence, heart disease and drinking , the differences were statistically significant (P < 0.05); In addition, CSVD group has
higher low-density lipoprotein，higher creatinine，higher cystatin C and higher homocysteine, the difference was statistically significant (P < 0.05),too.The specific contents are shown in table 1.
Table 1 the baseline data and single factor analys of risk factors for CSVD
|
Risk factors |
CSVD（n=279） |
control（n=136） |
P |
|
age（years） |
62.1±7.7 |
54.4±7.0 |
0.000& |
|
Male n（%） |
150（53.8%） |
71（52.2%） |
0.765# |
|
EH n（%） |
141（52.7%） |
34（25.0%） |
0.000# |
|
DM n（%） |
110（39.4%） |
37（27.2%） |
0.015# |
|
Heart diseasen（%） |
32（11.5%） |
6（4.4%） |
0.019# |
|
Smokingn（%） |
102（36.6%） |
41（30.1%） |
0.197# |
|
Drinkingn（%） |
70（25.1%） |
19（14.0%） |
0.010# |
|
CHOL（mmol/L） |
4.75±1.28 |
4.64±1.00 |
0.681* |
|
TG（mmol/L） |
1.79±1.19 |
1.63±1.13 |
0.197& |
|
LDL（mmol/L） |
2.63±0.91 |
2.40±0.67 |
0.038* |
|
HDL（mmol/L） |
1.40±0.40 |
1.43±0.38 |
0.396& |
|
Apo-a1（mmol/L） |
1.30±0.27 |
1.30±0.25 |
0.905& |
|
Apo-b（mmol/L） |
0.82±0.22 |
0.79±0.19 |
0.153& |
|
Lp（a）（mmol/L） |
184.84±197.82 |
179.39±213.86 |
0.798& |
|
Cr（umol/L） |
64.98±15.78 |
61.48±12.15 |
0.023& |
|
UA（umol/L） |
275.65±79.70 |
279.15±77.31 |
0.671& |
|
CysC（mg/L） |
0.92±0.48 |
0.80±0.11 |
0.006& |
|
TSH（mIU/L） |
2.75±2.91 |
2.34±2.46 |
0.174& |
|
Hcy（umol/L） |
14.99±6.10 |
13.20±5.72 |
0.010& |
&Measurement data, variance, two independent samples t-test;
* measurement data, the variance is not neat, logarithmic transformation was done before independent sample t-test;
# count data, chi-square test
2.3 the correlation between Corin serum level and CSVD
Serum Corin levels in the CSVD group were lower than those in the control group（0.589±0.439 ng/ml VS 0.870±0.522 ng/ml）, there were statistically significant（P<0.05）； the levels of Corin were negatively correlated to CSVD with the correlation coefficient of－0.303 using Spearman correlation analysis method（P<0.05）.
2.4 The logistics regression analysis of risk factors for CSVD
Multivariate logistic regression analysis results showed that lower serum Corin levels，higher age，hypertension，higher cystatin C are independent risk factors for the incidence of CSVD.The specific contents are shown in table 2.
Table 2 the baseline data and single factor analys of risk factors for CSVD
|
Risk factors |
B |
OR |
95%CI |
P |
|
Corin |
-1.077 |
0.340 |
0.186-0.624 |
0.000 |
|
elder |
0.108 |
1.115 |
1.074-1.157 |
0.000 |
|
EH |
1.025 |
0.359 |
0.202-0.636 |
0.000 |
|
CysC |
3.314 |
27.490 |
3.363-224.734 |
0.002 |
2.5 For patients in the different stages of CSVD :
Serum levels of soluble Corin protein were dropping from Mild WML group to Moderate WML group to Severe WML group（0.742±0.393 ng/ml to 0.575±0.467 ng/ml to 0.508±0.420 ng/ml）.There was a negative correlation between serum Corin levels and the severity of white matter lesion（r=－0.247，P<0.05）.Samely, serum Corin levels reduced while the lesion number of ALI and lacune raising（0.696±0.412 ng/ml to 0.543±0.454ng/ml to 0.494±0.409ng/ml）.There was a negative correlation between serum Corin levels and the lesion numbers of brain（r=－0.225，P<0.05），too.
2.6 For patients with the different clinical manifestation of CSVD:
The diagnosis of lacunar stroke was based on "diagnosis and treatment of acute ischemic stroke in China 2010"
[11].Serum Corin levels in the lacunar stroke group were lower than those in the asymptomatic CSVD group（0.539±0.417 ng/ml VS 0.650±0.459 ng/ml）, there were statistically significant（P<0.05）
2.7 Relationship between levels of serum Corin and gender, age,
In the CSVD group，women had lower Corin levels（0.528±0.403 ng/ml VS 0.640±0.463 ng/ml）, there were statistically significant（P<0.05）；the age had no effect on the serum Corin levels（P>0.05. The same outcome was also shown in the control group：women had lower Corin levels（0.787±0.557 ng/ml VS 0.972±0.451 ng/ml）, there were statistically significant（P<0.05）；the age had no effect on the serum Corin levels（P>0.05）.
2.8 The effect of hypertension to serum Corin levels in CSVD group:
For Serum Corin levels of CSVD deseases，there was no statistical difference between hypertension group and non-hypertension group（0.559±0.480 ng/ml VS 0.622±0.387 ng/ml，P>0.05）.
2.9 Factors affecting CSVD group serum Corin levels
Spearman correlation analysis method showed that serum LDL levels were related to serum Corin levels，in CSVD deseases（r=0.120、P<0.05）.
-
Discussion
Corin respectively turns pro - ANPandpro - BNP into bioactive ANP and BNP
[12], and then downstream of a series of biological effects, including diuretic, platoon sodium, vasodilator, and promote local angiogenesis, inhibiting renin angiotensin aldosterone system, resisting myocardial fibrosis, etc,through the ANP and BNP and cGMP dependent protein kinase signaling pathways.Moreover, there are 8 low density lipoprotein receptor (LDLR) structure in Corin, , may be mediated myocardial and coronary intake of low density lipoprotein
[13.14]; And low density lipoprotein can adjuste the transhipment of cholesterol, play an important role in a variety of disease of heart and head blood-vessel disease. Corin are cracked by metal protease ADAM10 and Corin itself , into segments of the soluble in the blood
[15].
CSVD plays an important rolein cerebrovascular disease, is a main reasonfor stroke, cognitive decline and age related disability
[16] . Pathology, CSVD refers to the pathological process caused by a set of different etiology ;which affectingon the brain capillaries and venules, micro artery and small artery
[17]. The form of CSVD is diversed, pathogenesis is complexed, pathophysiologic mechanism has not yet fully understood.Main hypothesis including micro embolus embolism, low perfusion or ischemic damage, changes of blood vessels , and participation of some metabolic encephalopathy
[18].
Studies have confirmed that the concentration of serumCorin are closely related with high blood pressure, heart failure, chronic kidney disease and acute cerebral apoplexy (including hemorrhagic stroke and ischemic stroke). We hypothesized that Corin likely also participate in the occurrence and development of cerebral small vessel disease.
This study with cerebral small vessel disease patients as the research object, with normal cranial MRI performance of non-cerebral vascular disease as control group.The correlation between serum Corin and CSVD was discussed for the first time. Two independent sample t-test shows serum soluble Corin levels of CSVD patients was significantly lower than the control group (P = 0.000), the levels of Corin were negatively correlated to CSVD .Further lines of multivariate logistic regression analysis, the results showed that the serum Corin levels are independent risk factors for CSVD, higher age，hypertension，higher cystatin C are the other independent risk factors for the incidence of CSVD. Diabetes, heart disease, alcohol consumption, lowerHDL-c, higher Cr, and higher HCY levels did not enter the regression model. Knocked out risk factors may have originated from existence and synergy between each other;or with sample selection, the use of drugs, the interference of other vascular risk factors and so on.But to some extent, they are not major roles in the CSVD, significantly weaker than the independent risk factors.
As a deep understanding of the inner link of Corin with CSVD, this study further conducted subgroup analysis in patients with CSVD. At present, the cerebral small lacuna, cerebral white matter lesions, perivascular gap enlargement and cerebral microbleed is recognized as imaging signs of CSVD
[1, 4]. These imaging signs can exist at the same time, there is no very strict boundaries .The diagnostic specificity of any one is low; multiple, simultaneous existence can improve the diagnostic specificity. This study , all CSVD patients exist clear focal cerebral small lacuna and cerebral white matter lesions, may also exist EPVS and CMB, but due to the limited imaging methods, this article does not discusses the latter two. According to the degree of cerebral white matter lesions , CSVD patients were divided into mild, moderate and severe ;At the same time ,in accordance with the number of cerebral small lacuna , CSVD patients were also divided into mild, moderate and severe . Results showed that serum corin level and the degree of the cerebral white matter lesions and cerebral small lacunar number are both negatively correlated, The serum Corin levels may provide the basis and forecast for the disease severity evaluations.
Clinical manifestations of CSVD were complexed, including asymptomatic cerebrovascular disease (asymptomatic lacunar cerebral infarction, the CMB, and part of the WML), lacunar stroke and vascular cognitive dysfunction. Cognitive dysfunction and dementia mainly caused by lacunar stroke occurring repeatedly, and asymptomatic cerebrovascular disease progressing, can be understood as CSVD terminal phase.This study did not include this kind of patients. According to the clinical signs and symptoms, medical history and imaging studies, CSVD patients were divided into lacunar stroke patients and asymptomatic CSVDpatients. Results showed that the serumCorin levels in the lacunar stroke group were lower than those in the asymptomatic CSVD group.The serum Corin levels may also provide the basis and forecast for the lacunar stroke.
To explore possible mechanisms behind Corin and CSVD, Factorsmay influence serum Corin levels of patients with CSVDare analyzed in this study.Hypertension is the basis happened causeof most CSVD , We had suspected Corin participated in the occurrence and development of CSVDby influencing the blood pressure .The CSVD group was divided into CSVD patients with hypertension group and without hypertension group, the results showed there is no statistically significant differences of the levels of serum Corin in two groups. At the same time, we found that in patients with CSVD, with the increase of blood LDL levels, serum Corin protein levels also has a certain degree of rise, this could be a feedback protection mechanism, this hypothesis still needs more research to confirm.
To sum up, this study found that Serum Corin levels were independently correlated to CSVD. Corin might be involved in the pathogenesis of CSVD and could be used as a biomarker; The serum Corin levels may provide the basis and forecast for the disease severity evaluations; The serum Corin levels may also provide the basis and forecast for the lacunar stroke; There is correlation between Corin and CSVD ，the mechanisms behind might be the feedback increase of Corin while LDL raising. Corin can become clinical therapeutic targets or only can be used as biological marker of CSVD is unknown. What is the specific pathophysiological mechanism; and how to control the level of serum Corin still needs a lot of problems such as prospective, multicenter clinical trialand related animal experiment to answer .
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