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Journal of Molecular Pathology and Biochemistry

April, 2021: 22(6) ISSN:232111373
https://doi.org/10.1155/2021/8863283

THE ROLE OF BETA TRACE PROTEIN (BTP) IN THE DETECTION OF DIABETIC

NEPHROPATHY
Ohiri J. U.1 Orluwene C. G.

1
Department of Chemical Pathology, Federal Medical Center, Owerri, Imo State, Nigeria.
2
College of Medicine, Rivers state University, Port Harcourt, Rivers state.
ABSTRACT
Background: Type 2 Diabetes Mellitus (T2DM) is a common metabolic medical problem
worldwide. It is associated with adverse multisystemic complications such as nephropathy,
neuropathy, retinopathy amongst others.
Objective: The study evaluated the serum concentration Beta Trace Protein (BTP) in non-
diabetics, diabetics without nephropathy and in diabetics with nephropathy at the University of
Port Harcourt Teaching Hospital. The study aims to assess the association between the serum
concentration BTP and glycemic control and to show how BTP define diabetics into nephropathy
by using ROC curve.
Method: Two hundred and forty (240) age and sex matched experimental and control subjects for
the study. BTP was estimated using Elabscience ELISA Kit, creatinine was estimated using Jaffe
Method (kinetic), glycated haemoglobin was done using fluorescence immunoassay, fasting
plasma glucose was estimated by using glucose oxidase method, microalbumin was estimated by
using turbidimetry and glomerular function rate were calculated using the Modified Diet in Renal
Disease Equation (MDRD).
Results: There was a relatively weak positive correlation (r = 0.30) between FPG and BTP,
indicating that there is a corresponding increase in BTP as FPG increases in subjects with diabetic
nephropathy. The BTP assay had an AUC of 0.812. The cut-off for BTP was 4.16ng/mL and the
assay had an 88.1% sensitivity and an 81.0% specificity.
Conclusion: The data strongly suggest that Beta Trace Protein BTP is a good biomarker for the
detection of diabetic nephropathy despite the less than desirable specificity and sensitivity of the
BTP assay as shown in the study.
Keywords; Neuropathy, Diabetes, Beta trace protein, Diabetic Nephropathy

INTRODUCTION condition with growing public health concern

Type 2 diabetes mellitus (T2DM) consists of
an array of metabolic dysfunctions
characterized by chronic hyperglycaemia.1
This is a result of a combination of resistance
to insulin action, inadequate insulin
secretion, and excessive or inappropriate
glucagon secretion.2 It is a common and
costly chronic disease which is associated
with significant premature mortality and
morbidity.3 It is a progressive disease
both nationally and worldwide.4,5
Glycaemic control in T2DM patients is
monitored by HBA1C assay, ideally people
with well-controlled diabetes should have an
HBA1C level of below 6.5% (48
mmol/mol).6 Biochemical markers play an
important role in accurate diagnosis of
kidney injury.5,6 Beta-trace protein (BTP),
also known as prostaglandin D synthase, is a

Ohiri and Orluwene, 2021 JMPB


low-molecular-mass protein which belongs
to the lipocalin protein family, with 168
amino acids and a molecular weight of
23000-29000, depending on the degree of
glycosylation. BTP has been reported to be a
better indicator of reduced glomerular
filtration rate than serum creatinine.7,8 Serum
β-trace protein has been found to be elevated
in patients with renal diseases.9,10 Beta Trace
Protein (BTP) plasma levels rise and virtually
all BTP appears to be freely filtered through
the glomerulus. It is constantly produced and
stable in the body and found to be increased
in the serum of patients with renal
diseases.11,12 It is, thus, useful in the detection
of renal dysfunction and has been proposed
as an endogenous marker for estimation of
glomerular filtration rate (GFR).13,14 This
study assessed the potential of BTP in the
detection and diagnosis of diabetic
nephropathy in T2DM patients.
METHODS
Study Area
The study was conducted at the diabetic
clinic of the University of Port Harcourt
Teaching Hospital, Port Harcourt, Rivers
state. It is a tertiary health care facility with
about 500 beds and serves as a major referral
center in Rivers State with clinics all through
the week.
Study Population and Sample
The study population consists of type 2
diabetes patients and non-diabetic attending
the University of Port Harcourt Teaching
Hospital, Port Harcourt, Rivers state. Two
groups of individuals were recruited into the
study as follows; Group A – Type II diabetes
mellitus subjects, Group B – Non diabetes
patients as controls. The group A and group
B will be age and sex matched.
The sample size for the study was determined
from the formula.15
n =Z2pq
Ohiri and Orluwene, 2021
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d2
where ;
n = sample size for Case and Control
Z = 95% confidence interval= 1.96
P = proportion of the target population
used=6.8%16
q = 1.0 – p = 1.0 - 0.068 = 0.932
d = degree of accuracy desired (usually
set at 0.05)
n = {(1.96)2 (0.068) (1.0 -
0.068)}
0.052
={3.841 × 0.068× 0.932}
0.0025
=97.4 Approximately 97
However taking into consideration that some
patients may opt out in the course of the
study, an attrition rate of 10% was used.
97 × 10 = 9.7, Approximately
10
100
= this gives a minimum sample size of 97 +
10
= 107
Therefore, a projection of 110 study participants
(with type II diabetes) and 110 apparently healthy
participants as control, giving a total of 220
participants were used for the study. Sequence
generation randomization method was employed in
this study such that all eligible study participants
were recruited from the diabetic clinic of the
University of Port Harcourt teaching Hospital on a
daily basis over a period of three months to obtain
the required sampling frame and the control of 110
participants each.
Ethical Consideration
Approval for the study was obtained from
ethical committee of University of Port
Harcourt Teaching Hospital and willing
informed consent obtained from each
participant.
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Data collection Elabscience Enzyme-linked Immunosorbent

A study questionnaire was used and it dealt
with socio- demographic variables, risk
factors for type II DM, including past
medical histories and information on clinical
and laboratory variables.
Specimen collection and analysis
Ten milliliters (10mls) of fasting venous
blood was collected by venipuncture from the
antecubital vein of each study participant.
5mls of venous whole blood was transferred
into a well labelled plain specimen tube
where it was allowed to clot for at least 60
minutes undisturbed at room temperature.
The separated serum was stored frozen
immediately at -200C until it was subjected to
Beta Trace Protein (BTP) assessment using
assay according to Manufacturers
Instruction. Creatinine expression was
estimated using the modified Jaffe method
(kinetic methods).
Data Analysis
Data obtained was analyzed using standard
statistical methods with the statistical
package for social science (SPSS Version
25). Differences of the mean NGAL between
diabetics and non-diabetics was compared
using the students’ t-test. A P-value equal or
lower than 0.05 was taken to be significant.
A Pearson Correlation Coefficient was used
to determine the association between NGAL
with the plasma glucose control.

RESULTS

Table 1: Socio-demographic Data

Variables Control Subject Chi-square

n = 120 (%) n = 120 (%) (p-value)
Gender
Male 60 (50.0) 60 (50.0) 0.00 (1.000)**
Female 60 (50.0) 60 (50.0)
Age
Mean age± SD 47.9 ±11.5 44.7 ± 11.7
**Difference is not statistically significant (p > 0.05)

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Table 2: Serum levels of biochemical parameters

Subject Control t-test

(n=120) (n=120)
FPG (mmol/l) 11.18 ±3.0 5.36 ±0.81 0.001*
HBA1C (%) 9.42 ±2.27 5.14 ±0.81 0.001*
Creatinine (µmol/l) 99.08 ±40.31 76.96 ±11.9 0.001*
eGFR (ml/min) 84.4 ±25.3 108.06 ±23.5 0.001*
BTP (ng/mL) 4.03 ±0.79 3.95 ±0.72 0.508**
All values are presented in mean ±standard deviation
FPG: Fasting plasma glucose, eGFR: Estimated glomerular filtration rate.
A/Cr: Albumin-Creatinine ratio
*Difference is statistically significant (p <0.05),

25 r = 0.30, p = 0.1368
y = 0.7764x + 2.1554
R² = 0.0937

20

15
FPG

10

0
0 2 4 6 8 10 12 14 16 18
BTP

Figure 1: Association of FPG and BTP in DN Subjects

The figure shows a relatively weak positive correlation (r = 0.30) between FPG and BTP,
indicating that there is a corresponding increase in BTP as FPG increases in subjects with diabetic
nephropathy.

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 A24
y = 0.6872x + 0.9693
18 r = 0.40, p = R² = 0.2189
16
14
12

HBA1c
10
8
6
4
2
0
0 5 10 15 20
BTP
Figure 2: Association of HBA1c and BTP in DN Subjects

The figure shows a weak positive correlation (r = 0.40) between HBA1cs and BTP, indicating that
there is a corresponding increase in BTP as HBA1c increases in subjects with diabetic
nephropathy.

1.2

0.8
Sensitivity

0.6

0.4

0.2

0
0 0.2 0.4 0.6 0.8 1
Specificity
Figure 3: Reciever Operator Characteristics Curve for BTP
The Receiver Operating Characteristic (ROC) curve for BTP is generated by plotting the
sensitivity and specificity of the BTP assay for each subject using the ROC function in the SPSS
v25 software. The ROC curve is an indication of the specificity and sensitivity of the assay in
predicting or indicating the presence of diabetic nephropathy. The BTP assay had an AUC of
0.812. The cut-off for BTP was 4.16ng/mL and the assay had a 88.1% sensitivity and a 81.0%
specificity.

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DISCUSSION
Beta-trace protein (BTP), is a novel low-
molecular-weight glycoprotein that is being
investigated for its use as a marker of GFR. It
exhibits similar advantages to cystatin-C over
serum creatinine, because it is independent of
height, sex, age, and muscle mass, and
exhibits increased sensitivity.2,3 It is
increased in cases of increased glomerular
permeability owing to glomerular capillary
lesion. BTP can be used as a marker of kidney
damage alternative to albuminuria as it may
detect renal injury earlier than albuminuria
because of its lower molecular mass, its
constant production rate, its ionic property,
and its stability.6–8 The serum concentration
of BTP was observed to be elevated among
subjects with Diabetes, while subjects with
diabetic nephropathy had an average BTP of
4.32 ±0.44mg/L that is significantly elevated
compared to diabetic subjects with no
nephropathy. Several studies have verified an
increase in the concentration of BTP in
diabetic patients who had already presented
other signs of reduced renal function.9–11
The fractional clearance of BTP
progressively increases with the reduction of
GFR, suggesting a decrease in its tubular
reabsorption. In fact, urinary excretion of
BTP, as demonstrated in multiple regression
analysis, is determined by its serum
concentration, by albumin excretion, and,
inversely, by GFR. A positive correlation
between serum BTP and U-Alb had been
found in diabetic patients and also in renal
patients.6,12 The current study shows a strong
correlation of HbA1c and BTP levels in
patients with microalbuminuria. This is
consistent with the findings of similar
studies, which reported elevated levels of
BTP as HbA1c levels increased in
individuals with diabetes.8,9 The
concentration of HbA1c predicts diabetes
complications because it reflects more
harmful glycation sequelae of diabetes, such
as retinopathy and nephropathy, which are
Ohiri and Orluwene, 2021
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understood to be due to harmful advanced
glycation end products.6–8 The results
showed that the BTP assay was 88.1%
sensitive and 81.0% specific. The half-life of
BTP is approximately 1.2 hours and it is
virtually exclusive excreted via the
kidneys.10–12 There have been varying
sensitivities and specificities of BTP assays
in diabetic nephropathy reported in several
studies.67 The sensitivity of BTP have been
reported to be between 73 – 90% and assay
specificity ranging from 74 – 92% in
previous studies.5,7,12
CONCLUSION
The data strongly suggest that Beta Trace
Protein BTP is a good biomarker for the
detection of diabetic nephropathy despite the
less than desirable specificity and sensitivity
of the BTP assay as shown in the study.
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