Comparison of Serum MMP-9 Value in Spondylitis Tuberculous with

Degenerative Spine Disease

Yohanes Augustinus

, Otman Siregar

, and Benny

Resident of Orthopaedic and Traumatology, Faculty of Medicine Universitas Sumatera Utara/ Haji Adam Malik Hospital-

Medan

Consultant of Orthopaedic and Traumatology, Spine Division, Faculty of Medicine Universitas Sumatera Utara/ Haji

Adam Malik Hospital-Medan

Staff of Orthopaedic and Traumatology, Spine Division, Faculty of Medicine Universitas Sumatera Utara/ Haji Adam

Malik Hospital-Medan

Keywords: Matrix metalloproteinase, MMP-9, Spondylitis Tuberculous, Degenerative Spine Disease

Abstract: Fourteen (14) subjects were divided into 2 groups, with seven (7) subjects of spondylitis tuberculous and 7

subjects of degenerative spine disease in the period from December 2017 to November 2018 who were

included in the inclusion criteria included in this study and blood sampling was taken for examination of

serum MMP-9 levels. There were significant differences in serum MMP-9 levels between spondylitis

tuberculous and degenerative spine diseases with a significance value of 0.002 (p<0.05) with low serum

MMP-9 levels in the spondylitis tuberculous study group 1857.14 ± 377.96 and mean in the control group

857.14 ± 243.97. There were significant differences in serum MMP-9 levels between spondylitis

tuberculous and degenerative spine diseases with a significance value of 0.002 (p<0.05) with low serum

MMP-9 levels in the spondylitis tuberculous study group 1857.14 ± 377.96 and mean in the control group

857.14 ± 243.97. Patients suffering from spondylitis tuberculous have higher serum MMP-9 levels than

patients with degenerative spine disease, although MMP-9 is not a specific marker examination for

spondylitis tuberculous, the results of this study can be suggestive into that can help to evaluate enzyme

activity in patients with spondylitis tuberculous disease.

1 INTRODUCTION

Tuberculous (TB) is one of the long-known diseases

and is still the leading cause of death in the world.

The prevalence of TB in Indonesia and other

developing countries is quite high. In 2006, new

cases in Indonesia amounted to more than 600,000

and most of them were suffered by people in

productive age (15–55 years).

About 20% of infections with pulmonary TB will

spread out of the lungs (extrapulmonary TB). Eleven

percent of extrapulmonary TB is osteoarticular TB,

and nearly half of patients suffer from spinal TB

infection. Half have lesions in the spine with

neurological deficits of 10% - 45% of sufferers.

Spondylitis tuberculous is an infection of the

spine caused by Mycobacterium tuberculous

(Lindsay et. Al., 1991; Martini and Welch, 2001;

Savant and Rajamani, 1997; Tachdjian, 1990).

Spondylitis tuberculous results in damage to the

body in the form of a defect that causes spinal

instability and disruption of surrounding structures

(Lindsay, et. Al., 1991; Tachdjian, 1990). The

occurrence of infection in spondylitis tuberculous

can originate from primary infection (Graham and

Kozak, 1993; Martini and Welch, 2001; Savant and

Rajamani, 1997), bacteria directly infect the corpus,

or secondary infections (Graham and Kozak, 1993;

Savant and Rajamani, 1997), namely bacteria spread

hematogenously or lymphogens from the location of

the primary disease to the spinal cord (Martini and

Welch, 2001). Infections that occur in spondylitis

tuberculous are generally secondary infections of the

lungs, but in some cases are primary infections. Pro-

inflammatory and anti-inflammatory cytokines play

an important role in the development and control of

Mycobacterium tuberculous infection (Patil, et. Al.,

2015). It has been demonstrated that cytokine

profiles will differ in each degree of disease

(Graham and Kozak, 1993; Lindsay, et. Al., 1991).

Augustinus, Y., Siregar, O. and Benny, .

Comparison of Serum MMP-9 Value in Spondylitis Tuberculous with Degenerative Spine Disease.

DOI: 10.5220/0009863602170220

In Proceedings of the 2nd International Conference on Tropical Medicine and Infectious Disease (ICTROMI 2019), pages 217-220

ISBN: 978-989-758-469-5

217

Several cytokines have been known to be

biomarkers of disease activity in tuberculous

infections. Matrix metalloproteinase (MMP) is a

zinc-dependent protease, which plays a role in the

process of degradation of the extracellular matrix

and modulates the inflammatory response by

facilitating and inhibiting different cytokines

(Salgame, 2011).

In determining the form of treatment for

spondylitis tuberculous, it is generally divided into

two groups of patients, namely groups of patients

accompanied by complications in the way of

neurological disorders and groups that are not

accompanied by neurological disorders. In patients

without neurological complications, medical

treatment is the primary choice in treatment, and

surgical treatment is only needed in some special

cases. But in patients who are accompanied by

neurological complications, the combination of

medical treatment and surgical management is the

most appropriate treatment.

Matrix Metalloproteinase is a zinc-dependent

proteinase that has an important role in the

degradation and rotation of the extracellular matrix.

[4] Since it was first reported, 24 members of MMP

have been identified with specificity and function of

overlapping substrates.4 Following each substrate

and function specification, MMP is divided into six

classes: stromelysins, collagenases, matrilysins,

gelatinases, membrane-type MMPs, and others.

Two gelatinases, MMP-2, and MMP-9 can reduce

original type IV collagen and denaturation of type I

collagen (gelatin). Both circulation and resident

inflammatory cells can synthesize MMP-9.

Experimental studies have provided evidence that

MMP-9 levels were significantly higher in

bronchoalveolar fluid patients with cavitary active

tuberculous, and lung extracts of mice infected with

M. tuberculous, compared to the control group. An

increased significance of MMP-9 was also observed

in tuberculous cerebrospinal fluid (CSF) meningitis

patients and also compared with people suffering

from viral meningitis, where usually these enzymes

are generally not found in cerebrospinal fluid.

Many previous studies have shown that M.

tuberculous can stimulate MMP-9 expression in the

lungs of infected organisms, but no studies have

examined how the expression of MMP-9 in

tuberculous spondylitis when compared to the control

group. Therefore, researchers are interested in trying

to evaluate and compare the expression of MMP-9 in

tuberculous spondylitis using serum levels in the

blood of patients suffering from tuberculous

spondylitis and compared to the control group, in this

case, the control group in this study were patients

with degenerative diseases of the spine.

2 METHODS

This cross-sectional study was conducted at the

Faculty of Medicine, Universitas of North Sumatra /

Haji Adam Malik Hospital, North Sumatra,

Indonesia for 11 months from December 2017 to

November 2018 by taking patient data and

examining serum values of MMP-9, 14 subjects met

the inclusion criteria.

Patients who met the inclusion criteria recorded

age, sex and blood tests to determine serum MMP-9

levels.

The study sample was divided consecutively

with equal numbers into two groups, namely:

spondylitis tuberculous, degenerative spine disease.

Patients were obtained from outpatient or inpatient

care diagnosed with spondylitis tuberculous and

degenerative spine disease. Patients who were

included in the inclusion criteria were taken for

blood sampling to examine levels of MMP-9.

The MMP-9 examination in this study used

MMP-9 (Matrix Metallo Proteinase 9) ELISA Kit

from Fine Test with the catalog number ERB0080,

size 48T / 96T and reactivity to humans. The scale

value used is 3.125-200ng / ml with sensitivity:

<1.875ng / ml. The application of this dosage

application is a quantitative detection of MMP-9 in

serum, plasma, tissue homogenates, and other

biological fluids. Differences in serum MMP-9

values in spondylitis tuberculous with the

degenerative disease in the spine were analyzed

using the Mann-Whitney test because the data

obtained were abnormally distributed. All statistical

calculations are carried out using a computer-based

statistical program. The study was approved by the

Health Research Ethics Committee of the Medical

Faculty of the University of North Sumatra / Haji

Adam Malik Hospital, and informed consent was

obtained from all subjects.

3 RESULTS

Before discussing the results of the study, because

this study had no references, it was conducted with a

small-scale preliminary study using 10 balanced

numbers of research subjects (5 subjects with

spondylitis tuberculous, 5 subjects with degenerative

spine disease) with women as many as 6 people

ICTROMI 2019 - The 2nd International Conference on Tropical Medicine and Infectious Disease

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(60%) and men as many as 4 people (40%). With a

mean and standard deviation of 41.6 ± 18.8 years.

While the mean of subjects with spondylitis

tuberculous (ST) 1800 ± 447.2 and mean

degenerative spine disease (DSD) 800 ± 273.8.

Table 1. Preliminary Study Distribution

Variable

Disease

ST DSD

Gender M/F 1/4 3/2

MMP-9 (nm/ml) 1800 ± 447.2 800 ± 273.8

Mean age 41.6 ± 18.8

ST, Spondylitis Tuberculous; DSD, Degenerative

Spine Disease; n, Subject

A total of 14 subjects were studied until the final

analysis. Data retrieval and examination of serum

MMP-9 values in spondylitis tuberculous (ST) and

degenerative spine disease (DSD) were carried out

in stages with the initial phase of selecting samples

of subjects included in the inclusion criteria. 7

subjects with diagnosed spondylitis tuberculous

(ST), 7 subjects with diagnoses of degenerative

spine disease (DSD), 8 women (57.1%) and 6 men

(42.9%), 13 years youngest research subjects and 73

years old oldest subjects with mean and standard

deviations amounting to 44.79 ± 16.98 years. From

the results of the analysis, it was found that the mean

and standard deviation of serum ST MMP-9 values

were 1857.14 ± 377.96 while the mean and standard

deviation of serum DSD levels of MMP-9 were

857.14 ± 243.97.

Table 2. Study Subject Distribution

Variable

Disease

ST MMP-9

(nm/ml)

DSD MMP-9

(nm/ml)

Patient

1 2000 1000

2 1000 1000

3 2000 500

4 2000 500

5 2000 1000

6 2000 1000

7 2000 1000

Mean MMP-9

(nm/ml)

1857.14 857.14

SD ± 377.96 ± 243.97

p-Value 0.002

Youngest age 13 year old 43 year old

Oldest age 62 year old 73 year old

Tuberculous drug

consumption

9.14 ± 5.29

weeks

ST, Spondylitis Tuberculous; DSD, Degenerative Spine

Disease; n, Subject

From the results of the statistical analysis of the

comparison of serum MMP-9 values in tuberculous

spondylitis (ST) with degenerative spine disease

(DSD), the results were significant that the serum

MMP-9 values were higher and this was indicated

by a p-value of 0.002 (p <0.05).

4 DISCUSSION

Pott's disease is the most common spinal

granulomatous bacterial infection and is the most

frequent bone TB. [6] The occurrence of infection in

spondylitis tuberculous can originate from primary

infection (Graham and Kozak, 1993; Martini and

Welch, 2001; Savant and Rajamani, 1997), i.e.

bacteria directly infect the corpus, or secondary

infections (Graham and Kozak, 1993; Savant and

Rajamani, 1997), namely bacteria spread

hematogenously or lymphogens from the location of

the primary infection to the spinal cord (Martini and

Welch, 2001). Infections that occur in spondylitis

tuberculous are generally secondary infections of the

lungs, but in some cases are primary infections. Pro-

inflammatory and anti-inflammatory cytokines play

an important role in the development and control of

Mycobacterium tuberculous infection (Patil, et. Al.,

2015). It has been demonstrated that cytokine

profiles will differ in each degree of disease

(Graham and Kozak, 1993; Lindsay, et. Al., 1991).

Several cytokines have been known to be

biomarkers of disease activity in tuberculous

infections. Matrix metalloproteinase (MMP) is a

zinc-dependent protease, which plays a role in the

process

of degradation of the extracellular matrix and

modulates the inflammatory response by facilitating

and inhibiting different cytokines (Salgame, 2011).

Two gelatinases, MMP-2, and MMP-9 have the

ability to reduce original type IV collagen and

denaturation of type I collagen (gelatin). Both

circulation and resident inflammatory cells have the

capacity to synthesize MMP-9. Experimental studies

have provided evidence that MMP-9 levels were

significantly higher in bronchoalveolar fluid patients

with cavitary active tuberculous, and lung extracts of

mice infected with M. tuberculous, compared to the

control group.

In a study conducted by Hrabec et al., 2002 it

was found that serum MMP-9 levels in patients with

active pulmonary tuberculous had significantly

higher levels of the control group with mean MMP-9

levels in the group with pulmonary tuberculous 1.23

± 0.43 µU / µL, while the mean MMP-9 level in the

Comparison of Serum MMP-9 Value in Spondylitis Tuberculous with Degenerative Spine Disease

219

control group was 0.37 ± 0.10 µU / µL and this

result was in accordance with the function of MMP-

9 which could induce enzymes produced by

mononuclear phagocytes and stimulated neutrophils.

This finding is in accordance with the study

conducted that there was a significant difference in

MMP-9 levels in patients with spondylitis

tuberculous with a control group used in this group

with subjects with degenerative spine disease with

mean spondylitis tuberculous patients 1857.14 ±

377.96 while the mean in patients with degenerative

spine disease 857.14 ± 243.97 with p-value 0.002 (p

<0.05), but in this study there was no comparison of

the ratio of white blood cell levels in the two study

groups as did the research conducted by Hrabec et al

2002 with this study obtained MMP-9 rate with an

increase in white blood cells was significantly higher

in the study group with pulmonary tuberculous

0.150 ± 0.054 mU/ml/L while in the control group

0.059 ± 0.023 mU/ml/L. In this study no correlation

was found between the duration of consumption of

anti-tuberculous drug and an increase in serum

MMP-9 levels (p> 0.05) with an average use for

tuberculous medicines 9.14 ± 5.29 weeks, although

this was not a primary goal in this study, but need to

be reviewed, because this assessment only uses 7

research subjects.

5 CONCLUSION

From the results of the comparison of serum

MMP-9 values in spondylitis tuberculous with

degenerative spine diseases, it was found that the

serum MMP-9 values in spondylitis tuberculous

gave higher results so that MMP-9 examination

could be used as a barometer for the diagnosis of

spondylitis tuberculous.

Further studies are needed to find out whether the

increase in serum MMP-9 levels in tuberculous

spondylitis is also accompanied by an increase in the

ratio of white blood cells and further studies are

needed to determine whether an increase in serum

MMP-9 levels can also be influenced by the duration

of use of anti-tuberculous drugs.

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