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Cerebrospinal Fluid Biomarkers in the Evaluation and

Treatment of Normal Pressure Hydrocephalus

by

Tae Sung Lim

Major in Medicine

Department of Medical Sciences

The Graduate School, Ajou University

Cerebrospinal Fluid Biomarkers in the Evaluation and

Treatment of Normal Pressure Hydrocephalus

by

Tae Sung Lim

A Dissertation Submitted to The Graduate School of

Ajou University in Partial Fulfillment of the Requirements for the Degree

of Ph.D. in Medicine

Supervised by

So Young Moon, M.D., Ph.D.

Major in Medicine

Department of Medical Sciences

The Graduate School, Ajou University

August, 2012

ACKNOWLEDGEMENTS

I would like to express my gratitude to all those who gave me the possibility to

complete this thesis. I am greatly grateful to Prof. So Young Moon (Ajou University, Korea)

for encouraging me to finish my experiments and analyses before leaving to St. Louis and

providing me a lot of valuable comments and criticism about this thesis; Prof. In Soo Joo

(Ajou University, Korea) for accepting the chairman of the supervisory committee and

giving me the direction of this thesis and especially, the opportunity to study abroad; Prof.

Sun Ah Park (Soonchunhyang University, Korea) for teaching me the laboratory technique

of ELISA analysis and providing me her valuable samples as normal and disease controls;

Prof. Kyoon Huh (Ajou University, Korea) for giving me inspiration for my thesis and

letting me know the weak points in the connection between part I and II of my thesis; Prof.

Byung Gon Kim (Ajou University, Korea) for letting me use his laboratory facility for my

ELISA analysis and giving me valuable comments to the context of my thesis. I specially

thank Prof. Young Chul Yoon (Chung-Ang University, Korea) for giving me his valuable

sample and Dr. Jun Young Choi for helping me to perform ELISA analyses.

Most of all, I really appreciate the assistance of my wife, Jae Won Hyun and my 3-year

old son, Seungchan Lim to let me study late everyday and go to school for preparation of this

thesis even in weekends.

Jun 29th 2012

Tae Sung Lim from St. Louis

i

- ABSTRACT -

Cerebrospinal Fluid Biomarkers in the Evaluation and Treatment of

Normal Pressure Hydrocephalus

Normal pressure hydrocephalus (NPH) is characterized by slowly progressive gait

disturbance, cognitive impairment and urinary incontinence. It is a well-known potentially

reversible cause of dementia by the shunt operation. However, it is not simple to select

candidates for the shunt operation due to combined pathology and poor general medical

conditions in the aged population and the high complication rate of the procedure. In PART I,

a retrospective study, we reported patients with NPH whose symptomatic response to

repetitive lumbar puncture (LP) was maintained for longer than one year without the need

for the shunt operation and analyzed the predictor for the prolonged symptomatic response to

repetitive LP in patients with NPH. In PART II, a prospective study, we investigated levels

of the β-amyloid 1–42 (Aβ42), total tau protein (T-tau) and tau phosphorylated at position

threonine 181 (P-tau) in cerebrospinal fluid (CSF) of NPH patients and tried to find their

clinical implications in the evaluation and treatment of NPH.

PART I: A retrospective study

Thirty-one NPH patients were retrospectively evaluated. Gait disturbance, urinary

incontinence, and cognitive impairment were semi-quantified. We divided the patients into

ii

three groups (non-responders, temporary responders, and prolonged responders) according to

their responses after LP. We analyzed the characteristics of the groups. Gait disturbance (p=

0.046) and urinary incontinence (p= 0.040) scores and total NPH symptom score (p= 0.007)

immediate after CSF drainage were more significantly improved in prolonged responders

than in temporary responders. On multiple logistic regression analysis, total NPH score

improvement immediate after LP was the only predictor of the prolonged responders (p=

0.038, odds ratio= 9.718). Our study showed that some NPH patients could maintain

favorable courses for at least one year after LP without the shunt operation. Repetitive LP

could be an alternative treatment in selected NPH patients.

PART II: A prospective study

Twenty-five possible NPH patients were prospectively enrolled and their CSF were

collected to analyze levels of Aβ42, T-tau and P-tau using ELISA method. Gait disturbance,

urinary incontinence, and cognitive impairment were semi-quantified and detailed

neuropsychological test was performed. The CSF of 17 Alzheimer’s disease (AD) patients

and 10 normal control subjects were tested to determine a cutoff level of the possible

coexistence of AD pathology in NPH patients. Eight NPH patients were classified into NPH

with lower CSF Aβ42 group and 17 patients were classified into NPH with higher CSF Aβ42

group (cutoff value = 490.13 pg/ml, Aβ42). There was no difference in the NPH grading

score and improvement rate after LP between the two groups. In the evaluation of 18 patients

who underwent the detailed neuropsychological tests (7 with lower CSF Aβ42 and 11 with

high), the NPH with lower CSF Aβ42 group had more deficit in attention (p=0.018),

visuospatial function (p=0.043) and verbal memory (p=0.009). In the evaluation of 10

iii

patients who underwent NP test twice before and after LPs, with lower CSF Aβ42 group

showed less improvement in phonemic categorical naming (p=0.008) and frontal inhibitory

function (p=0.018) after LP compared to NPH with higher CSF Aβ42 group. Our study

suggested that the incidence of Aβ42 pathology in NPH patients was not different from

normal elderly population and the coexistence of Aβ42 pathology in NPH patients might be

a contributing factor for lumbar puncture or shunt unresponsiveness especially in the field of

cognitive dysfunction.

Key words: normal pressure hydrocephalus, lumbar puncture, Alzheimer’s disease,

cerebrospinal fluid, neuropsychological test

iv

TABLE OF CONTENTS

ABSTRACT ··········································································································· ⅰ

TABLE OF CONTENTS ························································································· iv

LIST OF FIGURES ································································································· vi

LIST OF TABLES ·································································································· vii

PART I. Repetitive Lumbar Punctures as Treatment for Normal Pressure

Hydrocephalus: A Retrospective Study

. INTRODUCTIONⅠ ································································································ 2

. MATERIALS AND METHODS Ⅱ ········································································· 4

A. Subjects ··········································································································· 4

B. Evaluation of NPH Symptoms ········································································· 5

C. Classification of Patients ················································································· 6

D. Magnetic Resonance Image ············································································· 7

E. Statistical Analysis ··························································································· 9

F. Representative Cases of Prolonged Responders ·············································· 10

. RESULTS Ⅲ ········································································································· 13

A. Demographic and Clinical Characteristics of the Subject Groups ··················· 13

B. Demographic and Clinical Characteristics of the Subject Subgroups ·············· 15

C. Logistic Regression Analysis ········································································· 17

. DISCUSSION Ⅳ ··································································································· 19

v

PART II. Evaluation of Coexistence of Alzheimer’s Disease Pathology in

Normal Pressure Hydrocephalus using CSF ELISA analyses: A Prospective

Study

Ⅰ. INTRODUCTION ······························································································ 23

Ⅱ. MATERIALS AND METHODS ······································································· 25

A. Subjects ········································································································· 25

B. Evaluation of NPH Symptoms ······································································· 27

C. Sample Collection ························································································· 28

D. ELISA Methods ···························································································· 29

E. Magnetic Resonance Image ··········································································· 31

F. Neuropsychological Tests ·············································································· 33

G. Statistical Analysis ························································································ 34

Ⅲ. RESULTS ········································································································· 35

A. Demographic and Clinical Characteristics of the Subject Groups ··················· 35

B. Classification of NPH Patients According to Response to Lumbar Puncture ··· 38

C. Classification of NPH Patients According to ELISA Results ·························· 40

D. Demographic and Clinical Characteristics of the NPH Patient Subgroups ········ 43

E. Neuropsychological Tests Results ·································································· 45

Ⅳ. DISCUSSION ··································································································· 48

REFERENCES ······································································································· 52

국 요약 ·············································································································· 60

vi

LIST OF FIGURES

PART I. Repetitive Lumbar Punctures as Treatment for Normal Pressure

Hydrocephalus: A Retrospective Study

Fig. 1. Template image for grading white matter hyperintensity ··································· 7

Fig. 2. MRI of three representative prolonged responders ··········································· 12

PART II. Evaluation of Coexistence of Alzheimer’s Disease Pathology in

Normal Pressure Hydrocephalus using CSF ELISA analyses: A Prospective

Study

Fig. 1. Plates of CSF ELISA analyses ·········································································· 30

Fig. 2. Schematic drawing showing linear measures of hippocampal atrophy ············· 32

Fig. 3. Levels of CSF β-amyloid 1–42, total tau protein and tau phosphorylated at position

threonine 181 in the subject groups ································································ 37

Fig. 4. Receiver operating characteristic (ROC) curves for CSF β-amyloid 1–42 in the

control and AD patient groups ·········································································· 41

vii

LIST OF TABLES

PART I. Repetitive Lumbar Punctures as Treatment for Normal Pressure

Hydrocephalus: A Retrospective Study

Table 1. Demographic and clinical characteristics of the subject groups ······················· 14

Table 2. Comparison of demographic and clinical characteristics between temporary

responders and prolonged responders ···························································· 16

Table 3. Logistic regression analysis results for predicting prolonged responder group 18

PART II. Evaluation of Coexistence of Alzheimer’s Disease Pathology in

Normal Pressure Hydrocephalus using CSF ELISA analyses: A Prospective

Study

Table 1. Visual rating of hippocampal atrophy ··························································· 32

Table 2. Demographic and clinical characteristics of the subject groups ······················· 36

Table 3. Demographic and clinical data of the NPH patient subgroups according to

response to lumbar puncture ··········································································· 39

Table 4. Stratificatioin of subject by cut-off level using ROC analysis ······················· 42

Table 5. Demographic and clinical data of the NPH patient subgroups according to CSF

β-amyloid 1–42 level ···················································································· 44

viii

Table 6. Neuropsychological test results of the NPH patient subgroups before CSF

drainage ········································································································ 46

Table 7. Neuropsychological test performance change of the NPH patient subgroups

before and after CSF drainage ········································································· 47

1

PART I.

Repetitive Lumbar Punctures as Treatment for

Normal Pressure Hydrocephalus

: A Retrospective Study

2

I. INTRODUCTION

Normal pressure hydrocephalus (NPH) is characterized by slowly progressive gait

disturbance, cognitive decline, and urinary disturbance. It can be diagnosed by characteristic

clinical features and laboratory findings, especially brain imaging showing enlarged

ventricles with relatively less significant periventricular white matter changes (Adams et al.,

1965). However, in the clinical setting, making an NPH diagnosis is not straightforward. It is

necessary to rule out other possible causes and refer to response to lumbar punctures (LP)

and imaging findings to make a diagnosis and plan for the treatment. Dementia from NPH is

treatable by either a ventriculoperitoneal shunt (VPS) or ventriculo-atrial shunt (VAS). The

response to the shunt operation can be predicted by an LP or external cerebrospinal fluid

drainage prior to the operation (Graff-Radford, 2007). However, it is not simple to select

candidates for the shunt operation due to combined pathology and poor general medical

conditions in the aged population. In addition, according the long-term follow-up studies,

about six to eight percents of patients experience serious and permanent neurological deficits

such as hemiparesis or even death. Various complications such as subdural hematoma,

intracranial hemorrhage, or shunt infection were reported in more than 30% of patients

(Hebb and Cusimano, 2001). These problems underscore the need for other treatment

options.

A few studies reported that patients who underwent an LP once or twice without the

shunt operation showed prolonged symptomatic improvement (Tsakanikas and Relkin, 2007).

However, there have been no studies that evaluated if repetitive LP can be an alternative

treatment to the shunt operation in NPH patients. In this study, we reported patients with

3

NPH whose symptomatic response to repetitive LP was maintained for longer than one year

without the need for a shunt operation and analyzed the predictor for the prolonged

symptomatic response to repetitive LP in patients with NPH.

4

II. MATERIALS AND METHODS

A. Subjects

Among patients who visited the department of neurology at the Ajou Medical Center,

Suwon, Korea from January 2001 to December 2007, we recruited 31 patients who satisfied

with the criteria for NPH and were observed for at least one year after admission treatment.

All patients were fully informed that there were two options of repetitive lumbar punctures

or shunt surgery and agreed to be treated by repetitive lumbar punctures at least for one year.

All patients had had brain MRIs and an LP. The clinical criteria for NPH included following:

(1) insidious onset, age of onset older than 40, disease duration longer than three to six

months, progressive course, and no evidence of other possible diseases that could explain

clinical symptoms and imaging findings, (2) MRI showing ventricular enlargement not

entirely attributable to cerebral cortical atrophy, (3) gait disturbance with or without

cognitive deficit or urinary disturbance, (4) no evidence for the increased intracranial

pressure (70~ 245 mmH2O) (Relkin et al., 2005). After making a diagnosis of NPH, one or

two LPs were performed to drain 30 to 50 ml of the cerebrospinal fluid (CSF).

5

B. Evaluation of NPH Symptoms

An NPH scale modified from Larsson et al. (Larsson et al., 1991) and Krauss et al.

(Krauss et al., 1997) that assessed gait [1 = normal, 2 = walk without any assistive device but

insecure, 3 = walk with cane, 4 = walk with bimanual support(walker), 5 = walk aided by an

assistant, 6 = wheelchair-bounded], urinary disturbance (0 = normal, 1 = sporadic

incontinence or urge phenomena, 2 = frequent incontinence or urge phenomena, and 3 = no

or minimal control of bladder function) and cognitive deficit (0 = normal, 1 = minimal

attention or memory deficits, 2 = considerable attention or memory deficits but oriented to

situational context, and 3 = not or only marginally oriented to situational context) was used

to characterize and grade the clinical syndrome. Patients were evaluated both before and 4 to

6 hours after the LP.

6

C. Classification of Patients

After one-year of observation, patients were grouped into non-responders, temporary

responders, and prolonged responders. Non-responders were patients who showed neither

improvement in any scores nor subjective improvement after the LP. Temporary responders

were patients whose score in any category improved by at least one point or who reported

subjective improvement but whose improvement was not sustained up to three months.

Prolonged responders were patients whose improvement was as above but also was sustained

up to three months.

7

D. Magnetic Resonance Image

The Evans index and white matter hyperintensity were assessed using MRI. The Evans

index was defined as the maximal frontal horn ventricular width divided by the transverse

inner diameter of the skull and signifies ventriculomegaly if it is 0.3 or greater (Relkin et al.,

2005). White matter hyperintensity was evaluated by the method designed by Clinical

research for dementia of South Korea study (CREDOS). Both periventricular (1-3) and deep

(1-3) white matter hyperintensities were assessed using template image (Fig. 1) (Moon et al.,

2011). Each longest-diameter white matter change around the lateral ventricles (capping or

banding on the periventricular areas) or deep in white matter (especially the centrum

semiovale) were evaluated separately. The deep white matter changes were rated as 1 ( < 10

mm), 2 ( ≥ 10 mm, < 25 mm), or 3 ( ≥ 25 mm) and periventricular white matter changes

were rated as 1 ( < 5 mm) or 2 ( ≥ 5 mm, < 10 mm), or 3 ( ≥ 10 mm).

8

Fig. 1 Template image for grading white matter hyperintensity. The deep white matter

changes were rated as 1 ( < 10 mm, A), 2 ( ≥ 10 mm, < 25 mm, B), or 3 ( ≥ 25 mm, C) and

periventricular white matter changes were rated as 1 ( < 5 mm, D) or 2 ( ≥ 5 mm, < 10 mm,

E), or 3 ( ≥ 10 mm, F).

9

E. Statistical Analysis

We compared demographic features, CSF findings, Evans Index, white matter

hyperintensity, medications taken, total and categorized scores for the NPH before and after

the LP, and the amount of the improvement in the scores after the LP between temporary

responders and prolonged responders using the Mann-Whitney test. We also used logistic

regression analysis to search for independent predictors to discriminate between the two

groups. A p value of <0.05 was regarded as statistically significant. The statistical analyses

were performed using commercially available software SPSS 13.0 (SPSS Inc, Chicago, IL,

USA).

10

F. Representative Three Cases of Prolonged Responders

Case 1

A 71-year-old man presented with slowly progressive gait disturbance and frequent falls

that had appeared nine months earlier. In addition, he showed slowly progressive memory

impairment and urinary disturbance such as nocturia and frequency up to 10 times a day. His

gait posture was normal. However, he showed reduction in the overall velocity and the

step/stride length with wide base and ataxia in turning. It took him 13 s to walk 7 meters.

Parkinsonism was not observed. On the Korean Mini-Mental State Examination (K-MMSE),

he scored 18. His total NPH score was 5 with 3 in gait, 1 in cognition, and 1 in urination.

The Evans index was 0.347 and white matter hyperintensity scored 2 in periventricular and 1

in deep white matter (Figure 1A). At six hours after 50 ml of CSF was removed, his 7 m

walking improved to 8 s. His NPH score also improved to 4 due to the improvement of the

gait score from 3 to 2. He was satisfied with his symptomatic improvement, which was

sustained for more than three months up to one year without further LP.

Case 2

A 75-year-old man presented with gait disturbance which had appeared one year prior

to the visit. His gait was characterized by slow velocity, short stride, hesitant initiation, and

ataxia. Parkinsonism was not observed. On the K-MMSE, he scored 25. His NPH score was

6 with 5 in gait, 1 in cognition and 0 in urination. On his brain MRI, his Evans index was

11

0.340 with 2 in the periventricular and 1 in the deep white matter hyperintensity (Fig. 1B).

At six hours after 50 ml of CSF were removed, his NPH score improved to 2 because of

improvement of the gait score from 5 to 2 and cognition score from 1 to 0. His improvement

was sustained up to 3 months. However, his gait started to aggravate with 5 in the gait score

3 months after the LP. He received a second LP removing 50 more ml of CSF, which

resulted in significant improvement in the gait score, dropping it to 2 again. After that time,

he received an LP every three months for one and a half years, allowing him to maintain

independent gait. Finally, he got a VPS with favorable outcome with the NPH score of 2.

Case 3

An 82-year-old man presented with gait disturbance for 6 months. He also noted

frequency and urgency for 3 months. His gait was characterized hesitant initiation, slow

velocity, but normal base. He had trouble walking without assistance. No parkinsonism was

observed. On the K-MMSE, he scored 25. His NPH score was 8 with 5 in gait, 1 in cognition,

and 2 in urination. His Evans index was 0.343 with 1 in the periventricular and 1 in the deep

white matter hyperintensity (Fig. 1C). After 50 ml of CSF removal, his NPH score improved

to 5 because of improvement of the gait score to 3 and urination score to 1. We observed that

for the next three years he sustained his response with a little waxing and waning. His final

NPH score was 5.

12

Fig. 2. MRI of three representative prolonged responders. T2 weighted images show

dilated ventricles with various degrees of periventricular white matter hyperintensities.

13

III. RESULTS

A. Demographic and Clinical Characteristics of the Subject Groups

We recruited 31 patients (21 men and 10 women). The mean age of onset was 72.5 and

the mean disease duration was 453 days. Twenty-two out of 31 patients underwent

cisternography and all of them showed mild to severe communicating hydrocephalus. The

average of Evans index was 0.34 and the mean score of periventricular white matter

hyperintensity was 1.70 and that of deep white matter hyperintensity was 1.67. The mean

value of CSF protein was 38.4 mg/dl and that of CSF glucose was 72.8 mg/dl. The mean

score of NPH score before LP was 5.25 which was improved by 1.45 after LP. According to

the response to LP, the patients were divided into 6 non-responders, 25 responders (12

temporary responders and 13 prolonged responders). We summarized these features in Table

1.

14

Table 1. Demographic and clinical characteristics of the subject groups.

Non- responders

(n=6)

Responders

(n=25)

Total

(n=31)

Sex (Men:Women) 5:1 16:9 21:10

Age (years) 72.8 ± 6.7 73.7 ± 5.8 72.5 ± 5.8

Duration (days) 334 ± 353 481 ± 412 453 ± 400

Evans index 0.34 ± 0.03 0.34 ± 0.02 0.34 ± 0.02

PVWMH 2.00 ± 0.50 1.62 ± 0.88 1.70 ± 0.86

DWMH 1.87 ± 0.87 1.61 ± 0.61 1.67 ± 0.93

CSF protein (mg/dl) 42.3 ± 13.2 37.5 ± 10.9 38.4 ± 11.3

CSF glucose (mg/dl) 68.0 ± 13.7 78.2 ± 21.6 72.8 ± 16.7

Anti-Parkinson drugs (n) 3 13 16

Pre-gait score 2.0 ± 0.00 3.00 ± 1.25 2.80 ± 1.19

Pre-urinary score 1.16 ± 0.40 1.44 ± 0.76 1.38 ± 0.71

Pre-cognition score 0.83 ± 0.40 1.12 ± 0.78 1.06 ± 0.72

Pre-NPH score sum 4.00 ± 0.63 5.56 ± 2.02 5.25 ± 1.93

Post-gait score 2.00 ± 0.00 2.04 ± 0.73 2.03 ± 0.65

Post-urinary score 1.16 ± 0.40 0.88 ± 0.60 0.93 ± 0.57

Post-cognition score 0.83 ± 0.40 0.84 ± 0.47 0.83 ± 0.45

Post-NPH score sum 4.00 ± 0.63 3.76 ± 1.56 3.80 ± 1.42

PVWMH: Periventricular White Matter Hyperintensity; DWMH: Deep White Matter

Hyperintensity; CSF: Cerebrospinal Fluid; NPH: Normal Pressure Hydrocephalus.

15

B. Demographic and Clinical Characteristics of the Subject Subgroups

We statistically compared temporary and prolonged responders using the Mann-

Whitney test. There was no difference in age, sex and duration of illness. Among the

variables tested, the symptom scores before and after LP did not differ between the groups,

while the amount of changes in total NPH scores (p= 0.007), gait score changes (p= 0.046),

and urination score changes (p= 0.040) after LP were significantly different (Table 2).

16

Table 2. Comparison of demographic and clinical characteristics between temporary

responders and prolonged responders.

Temporary

responders

(n=12)

Prolonged

responders

(n=13)

p-value

Sex (Men:Women) M : 8 / F :4 M : 8 / F : 5 0.852

Age (years) 71.6 ± 5.8 73.2 ± 5.9 0.503

Duration (days) 521 ± 458 444 ± 381 0.852

Evans index 0.34 ± 0.02 0.33 ± 0.02 0.650

PVWMH 1.42 ± 0.90 1.81 ± 0.85 0.295

DWMH 1.68 ± 0.96 1.55 ± 0.88 0.728

CSF protein (mg/dl) 36.5 ± 11.4 38.4 ± 10.8 0.689

CSF glucose (mg/dl) 78.2 ± 21.6 70.1 ± 11.8 0.437

Anti-Parkinson drugs (n) 7 6 0.825

Pre-gait score 2.83 ± 1.33 3.15 ± 1.21 0.437

Pre-urinary score 1.33 ± 0.77 1.53 ± 0.77 0.470

Pre-cognition score 1.08 ± 0.79 1.15 ± 0.80 0.810

Pre-NPH score sum 5.25 ± 2.34 5.84 ± 1.72 0.225

Post-gait score 2.25 ± 0.86 1.84 ± 0.55 0.347

Post-urinary score 1.08 ± 0.66 0.69 ± 0.48 0.247

Post-cognition score 0.91 ± 0.51 0.76 ± 0.43 0.611

Post-NPH score sum 4.25 ± 1.86 3.30 ± 1.10 0.270

Gait score improvement 0.58 ± 0.90 1.30 ± 0.94 0.046¶

Urinary score improvement 0.25 ± 0.45 0.84 ± 0.68 0.040¶

Cognition score improvement 0.16 ± 0.38 0.38 ± 0.65 0.538

NPH score improvement 1.00 ± 0.95 2.53 ± 1.50 0.007¶

PVWMH: Periventricular White Matter Hyperintensity; DWMH: Deep White Matter

Hyperintensity; CSF: Cerebrospinal Fluid; NPH: Normal Pressure Hydrocephalus.

¶ p < 0.05.

17

C. Logistic Regression Analysis

To look for predictors to be able to discriminate between temporary and prolonged

responders, we used multiple logistic regression analysis. We categorized several variables

as follows: age of the onset (61-70, 71-80, older than 81), sex (Male, Female), disease

duration (< 1 year, ≥ 1 year), scores for periventricular white matter hyperintensity (< 2, ≥ 2),

and changes in the total NPH scores (< 2, ≥ 2). The analysis showed that the amount of

changes in the total NPH scores was the only independent predictor to discriminate two

groups [Odds ratio, 9.718 (95% confidence interval, 1.128 –83.709), p = 0.038] (Table 3).

18

Table 3. Logistic regression analysis results for predicting prolonged responder group.

B S.E Wald df Sig. OR (95% CI)

Age 1.928 2 0.381

71-80 1.413 1.614 0.766 1 0.382 4.106 (0.173-97.20)

>80 3.096 2.232 1.924 1 0.165 22.101 (0.278-1753)

Female -0.107 1.697 0.004 1 0.950 0.898 (0.032-24.99)

Duration -0.272 1.041 0.068 1 0.794 0.762 (0.099-5.860)

PVWMH 1.818 1.272 2.045 1 0.153 6.162 (0.510-74.47)

NPH improvement 2.274 1.099 4.284 1 0.038* 9.718 (1.128-83.70)

Constant -0.815 1.929 2.130 1 0.144 0.060

PVWMH: Periventricular White Matter Hyperintensity; NPH; Normal Pressure

Hydrocephalus

* p < 0.05

19

IV. DISCUSSION

This study showed that prolonged responders had significantly more improvement in

the total NPH score, gait score, and urination score. In addition, we found that the amount of

the total NPH score change was an independent predictor to discriminate between temporary

and prolonged responders. Although it is impossible to distinguish responses to the LP from

the natural course of the NPH, the patients who showed sustained response with repeated LP

could be evidences for the therapeutic responses. In addition, it is difficult to exclude some

disease responsive to the anti-parkinsonian drugs. However, there was no significant

difference in the drug history among the three groups.

Since Adams et al. first reported on NPH in the 1960s, there have been lots of

controversies surrounding this disease. Even up to now, prospective studies have not come to

a consensus about the disease criteria and pathologic findings (Adams et al., 1965; Graff-

Radford, 2007). The symptomatic triad, gait disturbance, cognitive decline, and urinary

disturbance, are very common complaints in the aged population and can be caused by

various conditions and diseases. Therefore, it is very difficult to avoid false positive or false

negative to make diagnostic criteria (Graff-Radford, 2007). In addition, the ventricle size

increases with age and degenerative diseases such as Alzheimer’s disease are also seen

frequently (Barron et al., 1976; Jack et al., 2004). Therefore, with the current diagnostic

criteria including symptomatic triad and enlarged ventricles, the prevalence of the NPH can

be overestimated. Currently, the main treatment for NPH is a shunt operation. The decision

for the shunt operation is made after one or two LPs. However, long term follow-up studies

20

reported high complication rates and symptomatic aggravation within a few years after the

shunt operation (Pujari et al., 2008). So, in many cases, it is difficult to make diagnoses for

the NPH and make decisions for the treatment. Although a few studies have reported that

patients showed prolonged response to LP, these studies have not been given full attention.

In addition, good response to the LP, which is revealed to be an independent predictor for the

prolonged responders in this study, is also the predictor for favorable outcome of the shunt

operation (Vanneste, 2000). Therefore, repeated LP has not been systematically studied since

the shunt operation was introduced in 1960s.

At an early stage of NPH, patients can compensate for abnormalities in CSF flow

through both reduced production and increased absorption of the CSF due to increased

intracranial pressure (Bateman, 2004). However, as the ventricles get larger, they lose the

elasticity with the even intracranial pressure according to the Laplace’s law. If the ventricles

are slightly shrunken by CSF removal, the elasticity of the ventricles can recover along with

symptomatic improvement for a certain period. It seems that prolonged responders to LP did

not acquire irreversible damage to the brain although patients showed symptoms. However,

if appropriate CSF removal is not done at the threshold, irreversible damage to the cerebrum

occurs and can produce non-responders or temporary responders to the LP.

We should accept that this study has several limitations. This study was retrospective

and could not quantify the severity of each symptom and determine the order of change

accurately. Each symptom was evaluated every 3 months at the outpatient clinic. Therefore,

we could not assess exactly when patients started to worsen. Furthermore, lots of patients

failed to be followed with just one year of observation. Future studies are needed to compare

clinical courses according to the treatment modality (the shunt operation/ repeated LP)

21

among non-responders, temporary responders, and prolonged responders.

In conclusion, this study showed that good responders to the LP should be observed for

at least 3 months to see if their responses are prolonged. If they are prolonged and patients

agree, they can be observed without an immediate shunt operation, especially in cases where

they are not in appropriate condition for the operation or have the possibility of having

several degenerative diseases.

22

PART II.

Evaluation of Coexistence of Alzheimer’s Disease

Pathology in Normal Pressure Hydrocephalus

using CSF ELISA analyses

: A Prospective Study

23

I. INTRODUCTION

Normal pressure hydrocephalus (NPH) is characterized by clinical triad of symptoms

including cognitive impairment, gait difficulty, and urinary incontinence along with

ventricular enlargement in brain imaging (Relkin et al., 2005). It is a potentially reversible

cause of cognitive and motor impairment in older adults using ventriculo-peritoneal (VP)

shunt or ventriculo-atrial (VA) shunt operation. While the treatment with VP shunts are

widely used and encouraging, there exists a high rate of complications and the factors that

predict shunt unresponsiveness remain poorly understood. One potential contributor to shunt

unresponsiveness is the presence of comorbid neurologic conditions that are common in the

aged, such as Alzheimer’s disease (AD) (Hamilton et al., 2010).

NPH and AD have been considered to have definitely different pathomechanism but

these days there have been a number of studies about sharing pathomechanism between the

two diseases (Silverberg et al., 2003). However there is still controversy about the clinical

implication of existence of AD pathology in NPH patients. Some insisted that it was only a

bystander and the rate of coexistence of AD was similar to that of normal population (Bech

et al., 1999; Golomb et al., 2000; Bech-Azeddine et al., 2007) while the others reported the

poor shunt response was possibly due to AD pathology (Hamilton et al., 2010; Patel et al.,

2012). Since the introduction of ELISA method to detect cerebrospinal fluid (CSF)

biomarkers for AD, there have been a lot of studies about the differential role of levels of the

β-amyloid 1–42 (Aβ42), T-tau protein (T-tau) and tau phosphorylated at position threonine

181 (P-tau) in CSF of NPH patients (Agren-Wilsson et al., 2007; Kapaki et al., 2007;

Tarnaris et al., 2009; Leinonen et al., 2011; Tarnaris et al., 2011a). However, there is still

24

conflicting results of the level of each biomarker between the two diseases and the

predictability of outcome in patients who underwent VP shunt.

The aim of this study was to investigate levels of the Aβ42, T-tau and P-tau in CSF of

NPH patients and tried to find their clinical implications in the evaluation and treatment of

NPH.

25

II. MATERIALS AND METHODS

A. Subjects

Among patients who visited the department of neurology at the Ajou Medical Center,

Suwon, Korea from March 2010 to February 2012, we recruited 25 patients who satisfied

with the criteria for possible NPH. All patients had had brain MRIs and an LP. The clinical

criteria for possible NPH included following: (1) Subacute or indeterminate mode of onset,

any age after childhood, non-progressive or not clearly progressive, may follow events, such

as mild head trauma, may coexist with other neurologic, psychiatric, or medical condition

apart from the disease symptoms, (2) MRI showing ventricular enlargement but may be

associated with either cerebral cortical atrophy or structural lesion that may influence

ventricular size (3) Any one symptom from clinical triad (gait disturbance, cognitive deficit

or urinary disturbance) (Relkin et al., 2005). After making a diagnosis of NPH, one or two

LPs were performed to drain 30 to 50 ml of the CSF and 10ml of CSF was collected to

evaluate biomarkers for AD.

The CSF of 17 AD patients and 10 normal control subjects which were collected and

stored previously in two other hospitals (S.C.H. U. H. and C.A. U. H.) were tested to get a

cutoff value of the coexistence of AD pathology. All AD patients met the criteria for

probable AD as proposed by the National Institute of Neurological and Communicative

Diseases and Stroke and Alzheimer’s disease and Related Disorders Association (NINCDS-

ADRDA) (McKhann et al., 1984). All normal control subjects scored in each cognitive

domain test higher than the cutoff value. The cutoff values for each test score were

26

represented as a –1.0 SD below the published norms for their age and education group (Ahn

et al., 2010).

We excluded patients with a history of significant hearing or visual impairment that

could render interview participation difficult, as well as those with a history of other

neurological disorders (e.g., idiopathic Parkinson’s disease, dementia with Lewy bodies, or

active epilepsy), psychiatric illnesses (e.g., schizophrenia, mental retardation, major

depression, or mania), those taking psychotropic medications, and those with a history of

significant alcohol and/or other substance abuse. We obtained informed consent from all

participants after they received a complete written and verbal description of the study. This

study was approved by the hospital's Institutional Review Board.

27

B. Evaluation of NPH symptoms

An NPH scale modified from Larsson et al. (Larsson et al., 1991) and Krauss et al.

(Krauss et al., 1997) that assessed gait [1 = normal, 2 = walk without any assistive device but

insecure, 3 = walk with cane, 4 = walk with bimanual support(walker), 5 = walk aided by an

assistant, 6 = wheelchair-bounded], urinary disturbance (0 = normal, 1 = sporadic

incontinence or urge phenomena, 2 = frequent incontinence or urge phenomena, and 3 = no

or minimal control of bladder function) and cognitive deficit (0 = normal, 1 = minimal

attention or memory deficits, 2 = considerable attention or memory deficits but oriented to

situational context, and 3 = not or only marginally oriented to situational context) was used

to characterize and grade the clinical syndrome. Patients were evaluated both before and 4 to

6 hours after the LP.

28

C. Sample collection

All participants underwent LP in the L3-4 or L4-5 interspace to drain 30~50 ml of CSF

to evaluate response to LP from 10:00 to 12:00. During the procedure, 10 ml of CSF was

collected in polypropylene tubes after discarding the first 3~4ml. Bloody or cloudy samples

were rejected. No serious adverse events were reported. The samples were immediately

centrifuged for 15 minutes at 2000 rpm to remove cells and aliquots were stored in

polypropylene tubes and immediately frozen at −80°C until analysis. They were thawed just

before analysis.

29

D. ELISA Methods

CSF T-tau concentration was determined using a sandwich enzyme-linked

immunosorbent assay ([ELISA] Innotest hTAU-Ag, Innogenetics, Ghent, Belgium)

specifically constructed to measure all tau isoforms irrespective of phosphorylation status, as

previously described (Blennow et al., 1995). P-tau was measured using a sandwich ELISA

method (Innotest Phospho-Tau[181P]), as previously described (Vanmechelen et al., 2000).

Aβ42 levels were determined using a sandwich ELISA (Innotest β-amyloid[1-42]),

specifically constructed to measure amyloid-β containing both the 1st and 42nd amino acids,

as previously described (Andreasen et al., 1999). All biomarker levels were measured in

duplicate according to the manufacturer’s instructions. The CSF samples of NPH patients

were analyzed twice using different aliquot. (Fig. 1).

30

Fig 1. Plates of CSF ELISA analyses used in this study. A: Innotest -amyloid[1-42]; B:

Innotest hTAU-Ag; C: Innotest Phospho-Tau[181P].

31

D. Magnetic Resonance Image

The Evans index and white matter hyperintensity were assessed using MRI. The Evans

index was defined as the maximal frontal horn ventricular width divided by the transverse

inner diameter of the skull and signifies ventriculomegaly if it is 0.3 or greater (Relkin et al.,

2005). White matter hyperintensity was evaluated by the method designed by Clinical

research for dementia of South Korea study (CREDOS). Both periventricular (1-3) and deep

(1-3) white matter hyperintensities were assessed (Moon et al., 2011). Each longest-diameter

white matter change around the lateral ventricles (capping or banding on the periventricular

areas) or deep in white matter (especially the centrum semiovale) were evaluated separately.

The deep white matter changes were rated as 1 ( < 10 mm), 2 ( ≥ 10 mm, < 25 mm), or 3 ( ≥

25 mm) and periventricular white matter changes were rated as 1 ( < 5 mm) or 2 ( ≥ 5 mm, <

10 mm), or 3 ( ≥ 10 mm). Hippocampal arophy was graded by Scheltens’ method (Table 1

and Fig. 2) (Scheltens et al., 1992).

32

Table 1. Visual rating of hippocampal atrophy.

Score Width of choroid

fissure

Width of temporal

horn

Height of

hippocampal

formation

0 Normal Normal Normal

1 ↑ Normal Normal

2 ↑↑ ↑ ↓

3 ↑↑↑ ↑↑ ↓↓

4 ↑↑↑ ↑↑↑ ↓↓↓

↑: increase, ↓: decrease

Fig 2. Schematic drawing showing linear measures of hippocampal atrophy. A: largest

vertical height of hippocampal formation, defined as dentate gyrus, hippocampus proper, and

subiculum together with parahippocampal gyrus; B: largest horizontal width between

hippocampal formation and brainstem; C: largest vertical width of choroid fissure; D: width

of temporal horn.

33

F. Neuropsychological Tests

Neuropsychologists assessed participants' cognitive functioning via the extensive Seoul

Neuropsychological Screening Battery (SNSB) (Ahn et al., 2010) covering five specific

cognitive domains, as follows.

(1) The attention and working memory assessment used the digit span forward and

backward tests.

(2) The language function assessment employed the Korean version of the Boston

Naming Test (K-BNT) (Kim and Na, 1999).

(3) The visuospatial function assessment was the patient's copy score of the Rey

Complex Figure Test (RCFT), neuropsychological assessment in which examinees are asked

to reproduce a complicated line drawing, first by copying and then from memory.

(4) Memory function was divided into verbal and visual memory. We assessed verbal

memory by means of the Seoul Verbal Learning Test (SVLT), the Korean version of the

revised Hopkins Verbal Learning Test (HVLT-R), testing participants on the immediate

recall, delayed recall, and recognition tasks. To assess visual memory, we tested participants

on the RCFT's immediate recall, delayed recall, and recognition tasks.

(5) To assess frontal lobe functioning, we used the contrasting program, go-no go, the

semantic and phonemic aspects of the Controlled Oral Word Association Test (COWAT)

and Stroop test.

34

G. Statistical Analysis

We used chi-square, analysis of variance (ANOVA), and Kruskal-Wallis tests to

compare demographic data of each group and analysis of covariance (ANCOVA) to compare

clinical data adjusted for age, sex and duration of education. Post-hoc analyses with Least

Significant Difference (LSD) method were performed for between-group comparisons.

Intraclass correlation coefficient was calculated to show the test-retest reliability of ELISA.

Receiver operating characteristics (ROC) curve analysis was used to identify the cut-off

levels with the optimal combination of specificity and sensitivity. Mann-Whitney test was

used to compare the scores of neuropsychological tests between the subgroups of NPH

patients, adjusted for age, sex and duration of education. All statistical analyses were

performed using SPSS 13.0 (SPSS Inc, Chicago, IL, USA). Statistical significance was

established using a p-value limit of less than 0.05.

35

III. RESULTS

A. Demographic and Clinical Characteristics of the Subject Groups

Demographic and clinical characteristics and results of statistical analyses are

summarized in Table 2 and plotted in Fig. 3. Age of AD group was significantly older than

control group (p=0.029). The intraclass correlation coefficient of Aβ42, T-tau and P-tau were

0.922, 0.908 and 0.960, respectively. The mean value of the two test results of each analysis

was used for statistical analyses. CSF Aβ42 level of AD group was significantly lower than

NPH group (p=0.013) and control group (p=0.001) after adjustment for age, sex and duration

of education. CSF T-tau and P-tau levels were significantly higher in AD group than NPH

group (p=0.003 and p=0.002, respectively).

36

Table 2. Demographic and clinical characteristics of the subject groups (N=52).

NPH AD Control p1 p2 p3

n (M/F) 25 (12/13) 17 (10/7) 10 (3/7)

Age, yr 73.3±7.0 72.2±10.0 63.0±6.7 0.924 0.029* 0.090

Education, yr 8.5±5.2 6.1±5.6 5.5±5.2 0.469 0.498 0.970

K-MMSE 19.5±6.9 18.3±2.1 24.4±5.5 0.843 0.232 0.157

Aβ42, pg/ml 579.8±182.3 409.2±166.1 691.8±212.7 0.013* 0.241 0.001*

T-tau, pg/ml 131.9±77.6 259.6±161.5 196.9±114.4 0.003* 0.312 0.382

P-tau, pg/ml 27.0±9.6 51.3±28.3 43.0±28.5 0.002* 0.123 0.597

NPH: Normal Pressure Hydrocephalus; AD: Alzheimer’s Disease; M: Male; F: Female;

K-MMSE: Korean version of Mini Mental Status Exam; p1 : p-value between NPH and

AD; p2: p-value between NPH and control; p3: p-value between AD and control

* p < 0.05

37

Fig. 3. Levels of CSF Aβ42 (A), T-tau (b) and P-tau (C) in the subject groups.

38

B. Classification of NPH Patients According to Response to Lumbar Puncture

According to the results of Part I, NPH patients were divided into responder group and

non-responder group. Responder group was determined as patients who showed 2 or more

improvement in NPH scale because it was the only independent predictor for prolonged

responder group. There was no difference in age, sex, duration of education, K-MMSE score,

white matter hyperintensity, hippocampal atrophy and CSF biomarkers for AD (Aβ42, T-tau

and P-tau) between the two groups (Table 3).

39

Table 3. Demographic and clinical data of the NPH patient subgroups according to

response to lumbar puncture (N=25).

Non-responders

(n=16)

Responders

(n=9) p-value

Sex (Men:Women) M : 7 / F : 9 M : 5 / F : 4 0.688

Age (years) 73.8 ± 8.4 72.4 ± 3.9 0.638

Education, yr 8.7 ± 5.1 8.2 ± 5.8 0.833

K-MMSE 20.1 ± 6.9 18.5 ± 7.2 0.600

Aβ42, pg/ml 543.8 ± 157.3 643.7 ± 214.7 0.194

T-tau, pg/ml 133.0 ± 92.0 128.5 ± 46.7 0.874

P-tau, pg/ml 25.5 ± 10.7 29.5 ± 7.2 0.329

Evans ratio 0.35 ± 0.03 0.36 ± 0.02 0.507

DWMH 1.38 ± 0.71 1.89 ± 1.05 0.161

PVWMH 1.81 ± 0.65 2.22 ± 0.83 0.186

Hippocampal atrophy 1.13 ± 0.95 0.89 ± 0.78 0.535

Pre-gait score 1.31 ± 1.13 2.22 ± 1.92 0.148

Pre-urinary score 0.63 ± 0.80 1.56 ± 1.01 0.019

Pre-cognition score 1.31 ± 0.79 1.67 ± 0.70 0.278

Pre-NPH score sum 3.19 ± 2.25 5.44 ± 3.28 0.053

Post-gait score 1.06 ± 1.18 0.89 ± 1.53 0.754

Post-urinary score 0.56 ± 0.81 0.44 ± 1.01 0.753

Post-cognition score 1.31 ± 0.79 1.00 ± 0.86 0.369

Post-NPH score sum 2.94 ± 2.35 2.44 ± 2.96 0.651

Gait score improvement 0.25 ± 0.44 1.33 ± 0.86 < 0.001*

Urinary score improvement 0.06 ± 0.25 1.11 ± 0.78 < 0.001*

Cognition score improvement 0 0.67 ± 0.50 < 0.001*

NPH score improvement 0.25 ± 0.44 3.11 ± 1.16 < 0.001*

NPH: Normal Pressure Hydrocephalus: AD: Alzheimer’s Disease: M: Male: F: Female:

K-MMSE: Korean version of Mini Mental Status Exam: PVWMH: Periventricular

White Matter Hyperintensity: DWMH: Deep White Matter Hyperintensity

40

C. Classification of NPH Patients According to ELISA Results

To determine a cut-off level of the coexistence of AD pathology in NPH patients, ROC

curve analysis was performed. CSF Aβ42 was used because it was the only biomarker which

was significantly different between AD and control group. The area under the curve was

0.876 with p-value of 0.001 (Fig. 4). According to the instruction of previous studies,

sensitivity more than 85% was selected in the determination of cutoff level of 490 pg/ml

which was quite similar to the level of the previous study (482 pg/ml) (Mattsson et al., 2009)

(Table 4).

41

Fig. 4. Receiver operating characteristic (ROC) curve of CSF Aβ42 for discrimination

between the control and AD patient groups.

42

Table 4. Stratification of subject by cut-off level using ROC analysis.

CSF Aβ42,

pg/ml Sensitivity Specificity

134.59 1 0

168.58 1 0.059

202.12 1 0.118

204.35 1 0.176

215.92 0.9 0.176

237.89 0.9 0.235

288.09 0.9 0.294

338.27 0.9 0.353

376.85 0.9 0.412

405.49 0.9 0.471

422.42 0.9 0.529

456.4 0.9 0.588

490.13 0.9 0.647

511.76 0.8 0.647

545.09 0.8 0.706

579.8 0.8 0.765

593.08 0.8 0.824

602.37 0.8 0.882

609.14 0.8 0.941

634.36 0.7 0.941

670.2 0.7 1

690.78 0.6 1

734.44 0.5 1

791.41 0.4 1

813.93 0.3 1

859.97 0.2 1

908.33 0.1 1

913.2 0 1

43

D. Demographic and Clinical Characteristics of the NPH Patient Subgroups

We statistically compared NPH with lower CSF Aβ42 group and NPH with higher CSF

Aβ42 group using the Mann-Whitney test. There was no difference in age, sex and duration

of education. Among the variables tested, hippocampal atrophy grade was significantly

higher in NPH with lower CSF Aβ42 group than NPH with higher CSF Aβ42 group (p=0.02).

There was no difference in the NPH grading score and improvement rate after LP between

the two groups (Table 5).

44

Table 5. Demographic and clinical data of the NPH patient subgroups according to

CSF Aβ42 level (N=25).

NPH with

lower CSF Aβ42

NPH with

higher CSF Aβ42 p-value

n (M/F) 8 (3/5) 17 (9/8) 0.673

Age, yr 76.1±7.3 72.0±6.7 0.187

Education, yr 7.7±5.6 8.7±5.2 0.687

K-MMSE 16.8±6.6 20.8±6.9 0.193

Disease duration, day 724.1±513.6 769.4±520.4 0.857

Aβ42, pg/ml 368.7±72.3 679.1±121.7 <0.001*

T-tau, pg/ml 145.7±103.9 125.3±64.6 0.551

P-tau, pg/ml 28.2±13.8 26.4±7.3 0.682

Evans ratio 0.34±0.02 0.36±0.03 0.277

DWMH 1.6±0.9 1.5±0.8 0.804

PVWMH 2.0±0.7 1.9±0.7 0.856

Hippocampal atrophy 1.6±0.9 0.7±0.6 0.020*

Pre-gait score 2.75±1.48 2.59±1.54 0.807

Pre-urinary score 1.00±1.06 0.94±0.96 0.892

Pre-cognition score 1.50±0.75 1.41±0.79 0.795

Pre-NPH score sum 4.13±2.94 3.94±2.86 0.883

Post-gait score 2.00±1.69 2.00±1.11 1.000

Post-urinary score 0.63±1.06 0.47±0.80 0.689

Post-cognition score 1.38±0.91 1.12±0.78 0.474

Post-NPH score sum 3.00±3.38 2.65±2.14 0.753

Gait score improvement 0.75±0.70 0.59±0.87 0.651

Urinary score improvement 0.38±0.74 0.47±0.71 0.761

Cognition score improvement 0.13±0.35 0.29±0.47 0.377

NPH score improvement 1.25±1.48 1.29±0.68 0.950

NPH: Normal Pressure Hydrocephalus: AD: Alzheimer’s Disease: M: Male: F: Female:

K-MMSE: Korean version of Mini Mental Status Exam: PVWMH: Periventricular

White Matter Hyperintensity: DWMH: Deep White Matter Hyperintensity

45

E. Neuropsychological Tests Results

A total of 18 NPH patients underwent detailed neuropsychological tests as an initial

evaluation before CSF drainage. Seven patients belonged to NPH with lower CSF Aβ42

group and 11 belonged to NPH with higher CSF Aβ42 group. Among the

neuropsychological test scores, the NPH with lower CSF Aβ42 group had lower score in

Digit span forward test (p=0.018), RCFT copy test (p=0.043) and SVLT immediate recall

test (p=0.009) after adjustment for age, sex and duration of education (Table 6)

In a subset analysis of 10 patients who underwent follow-up neuropsychological tests

after CSF drainage, NPH with lower CSF Aβ42 group showed significantly less

improvement in phonemic COWAT (p=0.008) and color reading test in stroop test (p=0.018)

after LP compared to NPH with higher CSF Aβ42 group after adjustment for age, sex and

duration of education. The mean interval between the first and the second

neuropsychological test was 13.2 ± 9.4 days (Table 7).

46

Table 6. Neuropsychological test results of the NPH patient subgroups according to

CSF Aβ42 level before CSF drainage (N=18).

NPH with

lower CSF Aβ42

(n=7)

NPH with

higher CSF Aβ42

(n=11)

p-value

Attention

Digit span forward test 4.7±1.1 6.2±1.4 0.018*

Digit span backward test 2.0±1.5 2.7±1.1 0.271

Language function

K-BNT 25.3±6.6 36.1±14.3 0.157

Visuospatial function

RCFT Copy 12.8±10.7 23.7±12.5 0.043*

Memory function

SVLT immediate recall 6.8±1.9 13.3±3.6 0.009*

SVLT delayed recall 0.1±0.3 1.8±2.0 0.204

SVLT recognition 15.5±2.9 15.9±4.0 0.820

RCFT immediate recall 2.9±4.3 7.9±6.7 0.129

RCFT delayed recall 1.8±2.8 6.6±6.2 0.232

RCFT recognition 16.0±3.3 16.9±3.1 0.480

Frontal function

Contrasting program 15.0±8.3 14.2±9.1 0.691

Go-no go 9.6±10.2 11.0±9.1 0.499

Semantic COWAT 6.3±3.9 8.8±6.1 0.269

Phonemic COWAT 10.0±6.0 14.4±11.4 0.926

Stroop test: color reading 26.0±13.2 44.5±35.9 0.905

NPH: Normal Pressure Hydrocephalus; AD: Alzheimer’s Disease; K-BNT: Korean

version of Boston Naming Test; RCFT: Rey Complex Figure Test; SVLT: Seoul Verbal

Learning Test; COWAT: Controlled Oral Word Association Test.

47

Table 7. Neuropsychological test performance change of the NPH patient subgroups

according to CSF Aβ42 level before and after CSF drainage (N=10).

NPH with

lower CSF Aβ42

(n=5)

NPH with

higher CSF Aβ42

(n=5)

p-value

Attention

Digit span forward test 0.2±1.3 -0.6±0.8 0.131

Digit span backward test 0.0±0.7 -0.2±0.8 0.911

Language function

K-BNT 2.4±6.5 0.6±3.6 0.602

Visuospatial function

RCFT Copy 0.1±4.5 -1.0±2.1 0.577

Memory function

SVLT immediate recall 2.0±3.0 4.2±3.8 0.117

SVLT delayed recall 0.0±0.0 2.0±1.8 0.054

SVLT recognition -1.0±4.1 1.4±2.8 0.602

RCFT immediate recall -0.8±2.5 2.2±3.5 0.245

RCFT delayed recall 0.3±2.3 3.0±3.2 0.245

RCFT recognition -2.6±3.2 2.0±3.0 0.465

Frontal function

Contrasting program -0.2±0.4 0.2±0.4 0.180

Go-no go 3.2±8.9 2.8±6.2 0.451

Semantic COWAT -1.4±3.3 5.4±6.1 0.117

Phonemic COWAT -1.0±1.7 5.4±6.1 0.018*

Stroop test: color reading -4.4±9.5 8.2±8.1 0.008*

NPH: Normal Pressure Hydrocephalus; AD: Alzheimer’s Disease; K-BNT: Korean

version of Boston Naming Test; RCFT: Rey Complex Figure Test; SVLT: Seoul Verbal

Learning Test; COWAT: Controlled Oral Word Association Test.

48

Ⅳ. DISCUSSION

Although there have been a few studies about the coexistence of AD pathology in NPH

patients and its clinical implication, to our best knowledge, this is the first study which

reported the difference in detailed neuropsychological tests before and after CSF drainage

according to CSF biomarkers for AD. The major findings of this study were as follows: (1)

All the three biomarkers for AD diagnosis were significantly different between AD and NPH

groups indicating there was not enough AD pathology compatible to AD in NPH patients

group. (2) There was no difference in CSF biomarkers for AD between responder and non-

responder group. (3) There was no difference in demographic and clinical characteristics

including the response to CSF drainage using NPH grading system between NPH with lower

CSF Aβ42 group and NPH with higher CSF Aβ42 group. (4) Comparison of initial

neuropsychological tests showed deficit in attention, visuospatial function and verbal

memory are more prominent in NPH with lower CSF Aβ42 group than NPH with higher

CSF Aβ42 group. (5) Neuropsychological performance improvement before and after CSF

drainage were significantly less in phonemic categorical naming and frontal inhibitory

function in NPH with lower CSF Aβ42 group than NPH with higher CSF Aβ42 group.

In comparison of Aβ42, T-tau and P-tau between NPH and AD groups, Aβ42 was lower

in AD group than in NPH group and T-tau and P-tau were higher in AD group than in NPH

group. These results correspond well with those of earlier studies which reported various

CSF biomarkers indicating less AD pathology in NPH patients than AD patients group (Lins

et al., 2004; Kapaki et al., 2007; Wilson and Williams, 2010). After determination of cutoff

49

level of AD pathology using ROC analysis of CSF biomarkers in AD and control group, 8

out of 25 NPH patients (32%) were classified as having AD co-pathology. This fits well with

the overall incidence of AD pathology in normal elderly population (Aizenstein et al., 2008).

However some recent studies using ventricular CSF during shunt procedure or external

lumbar drainage (ELD) reported higher T-tau and P-tau level in NPH patients (Tarnaris et al.,

2009; Leinonen et al., 2011; Patel et al., 2012). Elevated T-tau level has been detected in

various diseases causing neuronal injury such as stroke, trauma, hemorrhage and encephalitis

(Hulstaert et al., 1999; Hesse et al., 2001; Paraskevas et al., 2005). Shunt operation and ELD

procedure might result in neuronal injury and elevated T-tau and P-tau level in these studies.

Furthermore, the advanced stage of disease which needed to be treated by shunt operation

might affect this discrepancy.

There was no difference in CSF Aβ42, T-tau and P-tau between non-responder and

responder group. Furthermore, except for the hippocampal atrophy, all the radiological and

clinical symptomatic indicators of NPH patients including response to CSF drainage did not

differ between the two subgroups divided by CSF Aβ42 level. These results are quite similar

to those of previous studies reported that NPH patients benefit equally from shunting

regardless of the presence of AD pathology (Bech et al., 1999; Golomb et al., 2000; Bech-

Azeddine et al., 2007). Recently a study performed to compare CSF biomarkers and

response to ELD showed that none of the biomarker predicted the ELD results (Leinonen et

al., 2011). However there have a few reports that cortical AD pathology and ventricular CSF

biomarker for AD resulted in a less robust response to shunting (Hamilton et al., 2010; Patel

et al., 2012). Further study using simultaneous investigation of AD biomarkers of cortical

pathology and ventricular and lumbar CSF might provide some of these answers.

50

Although there was no difference in general cognitive function between the two

subgroups, the detailed neuropsychological tests revealed more deficits in attention,

visuospatial function and verbal memory in NPH with lower CSF Aβ42 group. These results

suggested that AD pathology impacts an additive AD-related cognitive dysfunction in NPH

patients regardless of underlying cognitive dysfunction caused by NPH (Patel et al., 2012).

Furthermore categorical naming and inhibitory executive dysfunctions in NPH with lower

CSF Aβ42 group, which were considered as NPH-related cognitive dysfunction, were not

improved after lumbar drainage compared to NPH with higher CSF Aβ42 group. The pattern

of improvement in neuropsychological tests in patients with higher CSF Aβ42 was similar to

the result of previous studies which reported improvement of subcortical dysfunction after

shunting but there has been no report about the pattern of improvement after lumbar

puncture (Caltagirone et al., 1982; Tarnaris et al., 2011b; Patel et al., 2012). The difference

between the first and second tests was not the result of learning effect because there was a

report that no learning effect was found in patients with NPH on any of neuropsychological

tests (Solana et al., 2010).

We should accept that this study has several limitations. One limitation of this study is

its relatively small sample size. Another limitation is lack of analysis of long term follow up

data such as shunting history. In addition, normal control subjects were recruited from the

clinic and possibly had cognitive complaint more than those recruited from the cohort.

Subjects with cognitive complaint are known to have more AD pathology than those without

it (Visser et al., 2009). This may affect the lack of difference in T-tau and P-tau between AD

and control groups.

51

In conclusion, our study reported that the incidence of AD pathology in NPH patients

which was demonstrated in the analysis of lumbar CSF ELISA tests was not different from

normal elderly population. However the NPH with lower CSF Aβ42 group showed more

deficits in AD-related cognitive dysfunction such as attention deficit, visuospatial

dysfunction and verbal memory dysfunction. Furthermore NPH with lower CSF Aβ42 group

showed less improvement even in NPH-related cognitive dysfunction such as phonemic

categorical naming and frontal executive function compared to NPH with higher CSF Aβ42

group. These results of the present study suggest that AD co-morbidity in NPH patients

might be a contributing factor for lumbar puncture or shunt unresponsiveness especially in

the field of cognitive dysfunction.

52

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60

- 국 요약 -

상압 증 평가 료에 뇌척 액 생체지 역할

아주 학 학원 학과

임 태

(지도 : 소 )

상압 증 히 진행하는 보행 장애, 인지 장애, 뇨 장애를

특징 하는 질 단락 에 해 있는 매 원인 잘

알 있다. 그러나 노인 자에 는 다른 퇴행 질 인 내과 인

질 이 동 는 경우가 많 며, 단락 자체 높 부작용 인해 에

한 자를 하는 것 쉬운 일이 아니다. 후향 연구인 부분 연구

1에 는 단락 하지 않고 복 인 뇌척 액 액 통해 어도 1 동안

증상 지하는 지속 보인 자들 증 들 보고하고,

지속 군 독립 요인 인자를 찾 한 분 하 다. 향 연구인

부분 연구 2에 는 상압 증 자 뇌척 액에 β-amyloid 1–42 (Aβ42), T-

tau protein (T-tau) and tau phosphorylated at position threonine 181 (P-tau)를 하여

상압 증 자 평가 료에 있어 그 임상 를 견하고자

하 다.

61

부분 연구 1: 후향 연구

31명 상압 증 자들 후향 분 하 다. 보행 장애, 뇨

장애, 인지 장애를 분 하 며 뇌척 액 액 후 에 라 자군

군, 일시 군, 지속 군 나 어 각각 특징에 해 통계

분 시행하 다. 지속 군에 일시 군에 해 뇌척 액 액

후에 보행 장애 (p=0.046) 뇨 장애 (p=0.040), 체 증상

변 (p=0.007)가 하게 높 소견 보 다. 다변량 지스틱 회귀분 에 는

체 증상 항목이 지속 군 일한 요인 인자 (p=0.038, odds

ratio=9.718) 분 었다. 본 논 에 르면 몇몇 상압 증 자들

단락 없이 복 인 뇌척 액 액 통해 어도 1 이상 증상

상태를 지하며 증상 이 클 지속 인 보일 것

상할 있다. 복 인 뇌척 액 액 상압 증 자 단락

체요법 사용 있다.

부분 연구 2: 향 연구

25명 상압 증 자들 향 모집하여 뇌척 액 집하고

상압 증 증상 자 한 신경심리검사를 시행하여 그 결과를

분 하 다. 집 뇌척 액 ELISA 분 통해 Aβ42, T-tau, P-tau를

하 고 상압 증에 알 하이 병 동 여부에 한 값

62

찾 해 17명 알 하이 병 자들과 10명 상 조군 뇌척 액

분 하 다. 8명 상압 증 자들이 알 하이 병 병 군 분 었고

나 지 17명이 알 하이 병 미병 군 분 었다 (cutoff value=490.13 pg/ml,

Aβ42). 군 사이에 상압 증 증상 뇌척 액 액 후

도 차이는 없었다. 신경심리검사를 시행한 18명 자들 분 통하여

알 하이 병 병 군이 알 하이 미병 군에 해 주 집 (p=0.018), 시공간

능 (p=0.043) 언어 억 (p=0.009)에 하를 보이고 있 알

있었다. 뇌척 액 액 후에 시행한 신경심리검사 결과 차이에 한

분 에 는 알 하이 병 병 군에 해 알 하이 병 미병 군에 이름

(p=0.008), 엽 행 능 (p=0.018)에 많 보 다. 본 연구를

통해 상압 증 자들 뇌척 액 상 알 하이 병 병 도는 상

노인 인구집단과 다르지 않았 나 상압 증 자에 알 하이 병

병 뇌척 액 액 또는 단락 에 지 않는 인지 하 연 이 있

알 있었다.

핵심어: 상압 증, 요추 천자, 알 하이 병, 뇌척 액, 신경심리검사