Decoding the role of serum cholinesterase among type 2 diabetes mellitus patients at RIMS, Ranchi – A hospital-based study

ABSTRACT

Introduction:

Diabetes mellitus (DM) type 2 is a group of conditions which is defined by persistent hyperglycemia along with various metabolic disturbances in carbohydrate, protein, and lipid metabolism. This condition is characterized by variable levels of insulin resistance, impaired insulin secretion, and elevated glucose production. There are various complications like cardiovascular diseases, neuropathy, nephropathy, and retinopathy which are associated with type 2 DM (T2DM). Serum cholinesterase (ChE) is an alpha glycoprotein enzyme which is formed in the liver and associated with various chronic inflammatory conditions like diabetes, hypertension, and cardiovascular diseases. So, we want to evaluate the correlation between serum cholinesterase concentrations in T2DM and also to evaluate the utility of serum cholinesterase as a diagnostic and prognostic marker in T2DM.

Material and Methods:

A cross-sectional study was conducted involving 176 participants, divided into two groups: 88 patients diagnosed with T2DM and 88 nondiabetic individuals matched for age and sex who attended at OPD and IPD of Medicine department of RIMS, Ranchi. The height and weight are taken, BMIs are calculated, and blood pressures are recorded. Blood samples are collected from all participants to measure serum cholinesterase levels; FBS, PPBS, HbA1c, serum urea, creatinine, lipid profile, thyroid profile, liver function tests (total bilirubin, direct bilirubin, AST, ALT, alkaline phosphatase), and urine protein creatinine ratio are assessed.

Results and Discussion:

The activity of serum cholinesterase raised in T2DM compared to the nondiabetic group. The cholinesterase levels in the diabetic group revealed 9272.18 ± 3062.92, median 9661.65, and in nondiabetics, it was 3504.93 ± 1520.74, median 3267.50, with a P value of < 0.001, which is statistically significant. Our study showed a positive correlation between serum cholinesterase and HbA1c with a correlation coefficient of 0.357 and a P value of 0.001, which is statistically significant. There is also a statistically significant correlation found between serum cholinesterase and serum triglyceride and also between serum cholinesterase and very low-density lipoprotein. But in our study, we have found that there is no statistical significant correlation between serum cholinesterase and overweight (BMI ≥ 25.0). The ROC plot analysis in this study demonstrates the higher diagnostic accuracy, 89.77, of serum cholinesterase with a sensitivity of 96.59 and a specificity of 82.95.

Conclusion:

So, serum cholinesterase concentrations are significantly increased in patients with T2DM and are also associated with poor glycemic control with increased insulin resistance. These study findings demonstrate the potential of serum cholinesterase as a biomarker to monitor the disease progression and its therapeutic effectiveness for interventions in T2DM.

Introduction

Acetylcholine acts as a transmitter at the neuromuscular junction and is present in autonomic ganglia, postganglionic sympathetic and parasympathetic nerves, and target organ junctions. For the repolarization to happen, acetylcholine must be rapidly cleared off from the synapse. This removal occurs due to hydrolysis of acetylcholine and forms choline and acetate, and the reaction is catalyzed by acetylcholinesterase enzyme present in the synaptic cleft.[1] It is of two types:

1. Acetylcholinesterase (AChE, EC 3.1.1.7) – Acetylcholinesterase is a specific esterase that hydrolyses predominantly choline esters. It is present in many types of conducting tissues like sensory fibers, motor fibers, cholinergic fibers, noncholinergic fibers, and central and peripheral tissues. The acetylcholinesterase activity is increased in motor neurons rather than sensory neurons. This enzyme constitutes the Yt blood group antigen, present in red blood cell membranes.

2. Pseudocholinesterase (BChE, EC 3.1.1.8) – It is also called as plasma cholinesterase or butyrylcholinesterase (BuChE). This is an alpha-glycoprotein present in various tissues like peripheral and central nervous systems and the liver and some peripheral tissues. Butyrylcholinesterase shows lower affinity for acetylcholine and high concentration of acetylcholine unable to inhibit this enzyme.[2] It is synthesized by the liver and hydrolyzes choline esters as well as other esters.[3] The half-life of butyrylcholinesterase is about 12 days.[4,5] Its normal range is between 5900 and 13,200 IU/L. This enzyme’s increased activity is shown in many conditions like diabetes, hyperlipidemia, obesity, uremia, and hyperthyroidism. Its concentration decreases in acute and chronic inflammations and protein-energy malnutrition and during stress. It is found ten times more than AChE in human blood.[6,7,8]

Diabetes mellitus (DM) is a rapidly growing disease globally with its huge social, health, and economic impacts. The data show that the prevalence of DM in India has increased from 7.1% (2009) to 8.9% (2019). India stands in the second position in global diabetes epidemics after China with 77 million peoples suffering from diabetes (according to International Diabetes Federation, 2019).

DM can be defined by a group of some common metabolic disorders, presented with hyperglycemia. There are several different types of DM due to complex interaction of various environmental and genetic factors. Mainly, two broad classifications of DM are there, type 1 and type 2. Type 1 diabetes is due to complete insulin deficiency or near-total insulin deficiency whether type 2 diabetes is a heterogeneous form due to variable levels of insulin resistance, impairment in insulin secretion, and elevated glucose production.[9]

Increased serum cholinesterase activity due to inadequate insulin secretion may adversely influence lipid and glucose metabolism.[10] Insulin deficiency results in increased flux of nonesterified fatty acids to the liver from peripheral tissues and increased synthesis of hepatic triacylglycerol and secretion of very low density lipoprotein.[11,12,13] This increased nonesterified fatty acids in circulation causes tissue resistance to uptake glucose in response to insulin, leading to hyperglycemia in type 2 DM (T2DM).[13,14]

T2DM is a type of low-grade systemic inflammatory condition. Though acetylcholine is a major neurotransmitter, it also exhibits anti-inflammatory properties. When the concentration of cholinesterase is increased, it causes the reduction of the level of Ach. Thus, an increased level of serum cholinesterase becomes a unique, reliable marker with greater specificity to identify acute, chronic, and also low-grade systemic inflammation.[15]

Due to its relatively lower cost with increased clinical informative power, serum cholinesterase assessment might be included in routine diagnostic procedures for DM. This is for the evaluation of clinical conditions of diabetic patients as diagnostic and prognostic parameters with the assessment of cardiovascular risk.[15]

Increased cholinesterase activity results in hydrolysis of the choline esters which are present in circulation. Thus, a rapid hydrolysis of acetylcholine diminishes the rate of action of Ach in the neighboring endothelium, affects endothelium functions adversely, and causes Ach induced relaxation. This increases blood pressure.[16]

Possibly, there is an association between the increase in serum cholinesterase and vascular complications in diabetic patients (which is strongly stimulated by the level of glycemia and also dyslipidaemia).

So, we intended to do a study expressing the relationship between serum cholinesterase and T2DM patients for the benefit of treating practitioners as well as patients.

Aim and Objectives

Aim

To evaluate the correlation between serum cholinesterase level in T2DM patients and healthy individuals.

Primary objective

To evaluate the utility of serum cholinesterase level in T2DM patients as a diagnostic and prognostic marker.

Secondary objectives

1. To estimate serum cholinesterase level in T2DM patients.

2. To correlate serum cholinesterase level in T2DM patients with dyslipidemia.

Material and Methodology

Study design: Analytical cross-sectional study.

Study duration: February 2023 to April 2024.

Study population: The patients of T2DM who attended at Medicine OPD and IPD, Rajendra Institute of Medical Sciences, Ranchi, between 40 and 70 years of age of either sex.

Study site: The study was carried out in the Department of Biochemistry at RIMS, Ranchi, Jharkhand.

Proper approval has been taken from Institutional Ethical Committee, Rajendra Institute of Medical Sciences, Ranchi, vide Memo no. 26/IEC, RIMS, dated 08.02.2023.

Inclusion criteria:

1. Patients who attended at medicine OPD and IPD of RIMS, Ranchi, between 40 and 70 years of age of either sex with T2DM.

2. Patients who wanted to participate after explaining the purpose of study and had given the consent for the study.

Exclusion criteria:

1. The patients of type 1 diabetes mellitus.

2. High alcohol consumption (>30 g/d)

3. Patients with known chronic liver or chronic inflammatory G.I. diseases

4. Patients with liver enzyme (AST, ALT, ALP) concentrations higher than 3 times the upper limit.

5. Patients on corticosteroids, methotrexate, amiodarone, tamoxifen, or other hepatotoxic drugs.

6. Organophosphorus poisoning.

7. Gestational diabetes.

8. Any chronic infections like TB and sarcoidosis were excluded from this study.

Sample size in number: 176

  (diabetic cases = 88)

  (nondiabetic cases = 88)

Method of sample collection: The patients of 40–70 years of age, males and females, diagnosed as T2DM from Medicine OPD and IPD of RIMS, Ranchi, were enrolled in the study. At the onset of study, the subject was briefed about the purpose of study and informal informed consent was taken from all the participants.

The nonrandomized sampling method was used.

The study subjects were divided into two groups, which include:

Group 1: T2DM patients

Group 2: Nondiabetic patients

Interview method:

The interview was conducted based on questionnaires. The interview was designed in two parts:

The first part related to subject identity, age, sex, ethnicity, occupation, socioeconomic status, and family history of diseases like hypertension, diabetes, and thyroid disorder.

The second part of the scheduled content items related to addictions like chewing tobacco, smoking, and alcoholism. The interview was followed by measurement of anthropometric parameters and collection of blood samples. The variables of the study that where inquired, measured, or calculated are discussed below in a stepwise manner.

1. Age – age was recorded in number of years already completed. All the subjects were able to say their correct age.

2. Sex – it was recorded as male or female.

3. Ethnicity – all the subjects were able to tell their ethnicity as tribal or nontribal.

4. Occupation – all the subjects were able to state their occupation.

5. Medical history – history of any comorbidities or drug history like antihypertensives, hypoglycemic drugs, drugs for thyroid disorder, and history of taking oral contraceptive pills in the case of women were stated by all participants.

6. History of addiction – any history of addiction like smoking, chewing tobacco, and alcoholism were recorded.

7. Family history – family history of diabetes, hypertension, and thyroid disorders were recorded.

Anthropometric measurements

Weight – weight was measured in kilograms (kg) in nearest number.

Height – height was measured with the subject standing by the side of the wall upright with heels close to each other and arms hanging by the side without any footwear. Height was marked against the wall on the horizontal plane touching the head of the subject. Length was then measured by a measuring tape, and the reading was recorded in meters.

Blood pressure – blood pressure was measured using a standard mercury sphygmomanometer.

Calculated parameters

Body Mass Index (BMI) – It is defined as the individual’s weight in kg divided by the square of their height in m with the unit of kg/m².

Study tools

1. Consent from the subject and an interview based on questionnaires.

2. Collection of blood samples: an overnight fasting of at least 12 hours prior to blood collection and 2 hours after the meal with caution to avoid hemolysis and contamination.

3. Processing and biochemical analysis of blood samples:

   1. FBS, PPBS was estimated using a biochemical automated analyzer on an ERBA EM360 clinical chemistry analyzer using absorptive spectrophotometric assay.

   2. Serum cholinesterase was estimated using a biochemical semiautomated analyzer on Metrolab 1600 DR using spectrophotometric assay.

   3. HbA1c was estimated using a Bio-Rad D10 hemoglobin testing system using ion-exchange high-performance liquid chromatography (HPLC).

   4. Serum urea and creatinine was estimated using a biochemical automated analyzer on an ERBA EM360 clinical chemistry analyzer using absorptive spectrophotometric assay.

   5. The lipid profile was estimated using a biochemical automated analyzer on an ERBA EM360 clinical chemistry analyzer using absorptive spectrophotometric assay.

   6. Liver function test was done using a biochemical automated analyzer on an ERBA EM360 clinical chemistry analyzer using absorptive spectrophotometric assay.

   7. Urine routine examination for presence of sugar and albumin was qualitatively done by the dipstick method.

   8. The urine–protein creatinine ratio was determined by measuring urine protein and creatinine using a DIRUI CS-1300B Auto Chemistry Analyzer.

4. Collection of blood samples – Subjects were instructed to be in fasting for at least 12 hours for all the tests except PPBS. For PPBS, subjects were instructed to give blood samples after 2 hours of meal. For urine tests, random samples were taken. Proper aseptic precautions were taken while collecting blood samples to ensure safety of self and the patients. Standard procedures were implemented in order to obtain good quality serum and plasma. Urine samples were collected in noncontaminated containers.

5. Procedure – Blood samples were allowed to clot by placing in the rack at room temperature for at least 30 minutes, and then, it was centrifuged at 3000 rpm for 10 minutes and serum was separated.

The serum, plasma, and urine samples were analyzed on the same day using the abovementioned machines with specific methodology.

Statistical methods

Categorical variables are expressed as number of patients and percentage of patients and compared across the groups using Pearson’s Chi square test for independence of attributes/Fisher’s exact test as appropriate.

Continuous variables are expressed as mean, median, and standard deviation and compared across the groups using Mann–Whitney U test/Kruskal–Wallis test as appropriate since the data do not follow normal distribution.

Association between continuous variables is captured by Spearman’s rank correlation coefficient since the data do not follow normal distribution.

ROC curve was used to understand the predictive power as measured by area under the curve. The optimum cutoff is determined at the point where sensitivity + specificity is the highest. Sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy were calculated at the optimum cutoff.

The statistical software SPSS version 25 has been used for the analysis.

An alpha level of 5% has been taken; that is, if any P value is less than 0.05, it has been considered as significant.

Results

Statistically significant results are found in height, weight, serum cholinesterase, fasting blood sugar, post prandial blood sugar, glycated hemoglobin, serum urea, serum cholesterol, triglyceride, HDL, LDL, VLDL, SGOT, ALP, urine protein creatinine ratio, systolic blood pressure, and diastolic blood pressure among diabetics and nondiabetics groups.

As shown in Figure 1 and Table 1, mean serum cholinesterase among diabetics is 9272.18, the median is 9661.65, and SD is 3062.92, and mean serum cholinesterase is 3504.93, the median is 3267.50, and SD is 1520.74. The P value is < 0.001, which is strongly statistically significant.

Figure 1.

Mean serum cholinesterase level in diabetic and nondiabetic groups

Table 1.

Comparison between mean, median, and standard deviation among diabetic cases and nondiabetic control groups with statistical significance

Group	Group						P	Significance
	Diabetic			Nondiabetic
	Mean	Median	Std. Deviation	Mean	Median	Std. Deviation
Age - years	54.31	55.00	8.77	50.76	48.00	8.63	0.008	Significant
Ht - m	1.61	1.61	0.07	1.64	1.65	0.05	0.011	Significant
Wt - kg	64.78	63.45	7.49	65.42	65.75	6.03	0.528	Not Significant
BMI - kg/m2	24.93	24.81	2.27	24.43	24.44	1.64	0.219	Not Significant
ChE - U/I	9272.18	9661.65	3062.92	3504.93	3267.50	1520.74	0.000	Significant
FBS - mg/dl	160.83	153.00	46.96	83.44	83.55	8.41	0.000	Significant
PPBS - mg/dl	241.92	231.75	82.13	125.58	127.75	10.98	0.000	Significant
HbA1C - %	8.55	7.90	2.23	4.82	4.80	0.39	0.000	Significant
s.Ur - mg/dl	26.97	23.95	10.25	18.87	19.20	3.69	0.000	Significant
s.Cr - mg/dl	1.03	1.06	0.21	0.97	0.98	0.20	0.075	Not Significant
Chl - mg/dl	185.09	182.00	33.59	158.88	158.00	11.74	0.000	Significant
Tg - mg/dl	165.51	147.50	72.37	138.40	141.00	13.94	0.005	Significant
HDL - mg/dl	42.50	41.40	7.59	47.85	48.30	5.56	0.000	Significant
LDL - mg/dl	111.34	110.78	27.94	82.20	83.55	13.06	0.000	Significant
VLDL - mg/dl	33.10	29.50	14.48	27.68	28.20	2.79	0.005	Significant
Bil (T) - mg/dl	0.65	0.64	0.28	0.67	0.68	0.16	0.249	Not Significant
Bil (D) - mg/dl	0.25	0.24	0.10	0.27	0.27	0.10	0.398	Not Significant
SGOT - U/L	26.03	23.65	10.36	29.57	28.00	11.55	0.011	Significant
SGPT - U/L	25.59	24.40	11.17	26.54	25.90	8.82	0.178	Not Significant
ALP - U/L	101.86	103.00	20.96	112.87	112.75	19.50	0.000	Significant
UPCR	0.36	0.14	0.82	0.10	0.08	0.06	0.003	Significant
SBP - mm of Hg	125.16	124.00	13.40	115.86	116.00	7.32	0.000	Significant
DBP - mm of Hg	80.50	80.00	6.04	75.98	76.00	5.29	0.000	Significant

As shown in Figure 2 and Table 1, mean serum cholesterol in the diabetic group is 185.09, median is 182.00, and SD is 33.59, and mean serum cholesterol in the nondiabetic group is 158.88, the median is 158.00, and SD is 11.74, and the P value is <0.001, which is strongly statistically significant.

Figure 2.

Mean serum cholesterol in diabetic and nondiabetic groups

As shown in Figure 3 and Table 1, mean serum triglyceride in the diabetic group is 165.51, the median is 147.50, and SD is 72.37, and mean serum triglyceride in the nondiabetic group is 138.40, the median is 141.00, and SD is 13.94, and the P value is 0.005, which is statistically significant.

Figure 3.

Mean serum triglyceride in diabetic and nondiabetic groups

As shown in Figure 4 and Table 1, mean serum HDL in the diabetic group is 42.50, median is 41.40, SD is 7.59, and mean serum HDL in the nondiabetic group is 47.85, median is 48.30, SD is 5.56, and the P value is < 0.001, which is strongly statistically significant.

Figure 4.

Mean serum HDL in diabetic and nondiabetic groups

As shown in Figure 5 and Table 1, mean serum LDL in the diabetic group is 111.34, median is 110.78, SD is 27.94, and mean serum LDL in the nondiabetic group is 82.20, median is 83.55, SD is 13.06, and the P value is <0.001, which is strongly statistically significant.

Figure 5.

Mean serum LDL in diabetic and nondiabetic groups

As shown in Figure 6 and Table 1, mean serum VLDL in the diabetic group is 33.10, median is 29.50, SD is 14.48, and mean serum VLDL in the nondiabetic group is 27.68, median is 28.20, SD is 2.79, and the P value is 0.005, which is statistically significant.

Figure 6.

Mean serum VLDL in diabetic and nondiabetic groups

As shown in Figure 7 and Table 1, mean UPCR in the diabetic group is 0.36, median is 0.14, SD is 0.82, and mean UPCR in the nondiabetic group is 0.10, median is 0.08, SD is 0.06, and the P value is 0.003, which is statistically significant.

Figure 7.

Mean urine protein creatinine ratio in diabetic and nondiabetic groups

As shown in Figure 8, the mean serum cholinesterase level among diabetic group in females is 9382.80; in males, it is 9151.03, and among the nondiabetic group, the mean serum cholinesterase level in females is 3442.32; in males, it is 3564.75.

Figure 8.

Graphical representation of relation of serum cholinesterase with gender in diabetic and nondiabetic groups

Figure 9 shows that the mean serum cholinesterase level among the diabetic group in overweight is 9619.01; in normal weight, it is 9020.56, and among the nondiabetic group, the mean serum cholinesterase level in overweight is 3444.46; in normal weight, it is 3534.65.

Figure 9.

Graphical representation of relation of serum cholinesterase with weight in diabetic and nondiabetic groups

Table 2 shows the relation of serum cholinesterase with gender in diabetic and nondiabetic groups and analyses that the P value among diabetics is 0.719, and among nondiabetics, it is 0.265. Both are statistically not significant.

Table 2.

Relation of serum cholinesterase with gender in diabetic and nondiabetic groups

ChE - U/I
Group	Gender	Mean	Median	Std. Deviation
Diabetic	Female	9382.80	9684.00	3244.79
	Male	9151.03	9661.65	2884.86
	P	0.719
	Significance		Not Significant
Nondiabetic	Female	3442.32	3201.60	1685.24
	Male	3564.75	3481.20	1361.74
	P	0.265
	Significance		Not Significant

Table 3 shows the relation of serum cholinesterase with BMI (≥25.0) in diabetic and nondiabetic groups which analyses that the P value among diabetics with is 0.410, which is not significant, and the P value among nondiabetics is 0.912, which is also not significant.

Table 3.

Relation of serum cholinesterase with BMI (≥25.0) in diabetic and nondiabetic groups

ChE - U/I
Group	Overweight	Mean	Median	Std. Deviation
Diabetic	NO	9020.56	9358.00	2956.00
	YES	9619.01	9854.10	3212.88
	P	0.410
	Significance		Not Significant
Nondiabetic	NO	3534.65	3274.90	1510.83
	YES	3444.46	3203.40	1565.83
	P	0.912
	Significance		Not Significant

Table 4 shows the association between continuous variables like HbA1c, Tg, VLDL and diabetic and nondiabetic group by Spearman’s Rank correlation coefficient with a significant P value.

Table 4.

Association between continuous variables by Spearman’s rank correlation coefficient

GROUP				ChE - U/I
Spearman’s rho	Diabetic	HbA1C - %	Correlation Coefficient	0.357
			P	0.001
		Tg - mg/dl	Correlation Coefficient	-0.217
			P	0.042
		VLDL - mg/dl	Correlation Coefficient	-0.217
			P	0.042
	Nondiabetic	HbA1C - %	Correlation Coefficient	0.199
			P	0.063
		Tg - mg/dl	Correlation Coefficient	-0.298
			P	0.005
		VLDL - mg/dl	Correlation Coefficient	-0.300
			P	0.005

The ROC plot analysis in our study demonstrates the higher serum cholinesterase activity in diabetic groups compared with the nondiabetic group.

Table 5 shows that the area under the curve shows the area is 0.952, the standard error is 0.016, the P value is < 0.001, and the lower bound and upper bound for asymptotic 95% confidence interval are 0.921 and 0.983, respectively.

Table 5.

Area Under the Curve

Test Result Variable (s): ChE - U/I
Area	Std. Errora	P	Asymptotic 95% Confidence Interval
			Lower Bound	Upper Bound
0.952	0.016	<0.001	0.921	0.983

Table 6 shows that the cutoff of serum cholinesterase from ROC is 4557.35.

Table 6.

Cutoff of serum cholinesterase from ROC

	Cut off from ROC
ChE - U/I	4557.350

Table 7 shows that among 88 diabetic cases, 85 have high serum cholinesterase concentration, whereas among 88 nondiabetic cases, 73 have normal serum cholinesterase concentration.

Table 7.

Serum cholinesterase concentration among diabetic and nondiabetic groups

		ChE - U/I		Total
		Normal	High
Diabetic	NO	73	15	88
	YES	3	85	88
Total		76	100	176

Table 8 shows that For serum cholinesterase, true positive is 85, true negative is 73, false positive is 15, false negative is 3, sensitivity is 96.59, specificity is 82.95, positive predictive value is 85.00, and negative predictive value is 96.05, with a diagnostic accuracy of 89.77.

Table 8.

True positive, true negative, false positive, false negative, sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic accuracy of serum cholinesterase

Parameter	TP	TN	FP	FN	Sensitivity	Specificity	PPV	NPV	Diagnostic accuracy
ChE - U/I>=4557.35	85	73	15	3	96.59	82.95	85.00	96.05	89.77

Graph 1 scatter diagram showing correlation of serum cholinesterase with FBS among diabetic and nondiabetic groups.

Graph 1.

Scatter diagram showing correlation of serum cholinesterase with FBS among diabetic and nondiabetic groups

Graph 2 scatter diagram showing correlation of serum cholinesterase with PPBS among diabetic and nondiabetic groups.

Graph 2.

Scatter diagram showing correlation of serum cholinesterase with PPBS among diabetic and nondiabetic groups

Graph 3 scatter diagram showing correlation of serum cholinesterase with serum triglyceride among diabetic and nondiabetic groups.

Graph 3.

Scatter diagram showing correlation of serum cholinesterase with serum triglyceride among diabetic and nondiabetic groups

Graph 4 scatter diagram showing correlation of serum cholinesterase with serum VLDL among diabetic and nondiabetic groups.

Graph 4.

Scatter diagram showing correlation of serum cholinesterase with serum VLDL among diabetic and nondiabetic groups

Graph 5 scatter diagram showing correlation of serum cholinesterase with glycated hemoglobin (HbA1c) among diabetic and nondiabetic groups.

Graph 5.

Scatter diagram showing correlation of serum cholinesterase with glycated hemoglobin (HbA1c) among diabetic and nondiabetic groups

Graph 6 shows ROC plot of distribution of serum cholinesterase activities of diabetic groups and nondiabetic groups.

Graph 6.

ROC plot of distribution of serum cholinesterase activities of diabetic groups and nondiabetic groups

Discussion

We have taken 88 diabetic patients with 88 in the nondiabetic group, and we have compared the serum cholinesterase concentration in both the groups along with serum urea, serum creatinine, lipid profile tests (total cholesterol, triglyceride, HDL-C, LDL-C, VLDL-C), serum total bilirubin, serum direct bilirubin, AST, ALT, alkaline phosphatase, glycated hemoglobin, and urine protein creatinine ratio.

In our study, we observed that serum cholinesterase activity is significantly higher among T2DM compared to the nondiabetic group with mean and standard deviation 9272.18 ± 3062.92 and median 9661.65 with a P value <0.001 that is statistically significant [Table 1], which is in agreement to the many studies conducted by Abbott C.A. et al., Inácio Lunkes G et al., Turecky L et al., Jewad Abdulkareem M. et al., Deepa K et al., and Rashid Hussein M. et al.[10,16,17,18,19,20] This finding suggests that serum cholinesterase may have a role in the pathogenesis of T2DM through changes in amyloid fibril or by modifying other risk variables of insulin resistance. This amyloid fibrils causes β-cells apoptosis by the excessive production of superoxide radicals and lipid peroxidation with nitric oxide inactivation in pancreatic islets as stated by Hemantha Kumara DS et al. and Deepa K et al.[19,21]

Turecky L, et al.,[17] 2005, reported that serum cholinesterase activity was significantly higher in the diabetes mellitus group compared to a controlled group (65.05 vs 73.33 μkat/l).

Dave KR and Katyare SS, 2002, showed that serum butyrylcholinesterase activity was sex-dependent and was increased only in male diabetic rats (2.2- to 2.8-fold), whereas for females, the result was reversed.[22] But in our study, the relation of serum cholinesterase with sex in diabetic and nondiabetic groups is statistically not significant [Table 2].

This study also showed positive correlation between serum cholinesterase and HbA1c with a correlation coefficient of 0.357 and P value 0.001, which is strongly significant statistically [Table 4]. This observation is similar with the finding of the study done by Katoh Shuichi et al., 2014.[23]

There is also statistically significant correlation found between serum cholinesterase and serum triglyceride and also between serum cholinesterase and very low density lipoprotein. This finding is also in agreement with the study finding of Ragoobirsingh D et al. in 1992,[24] Abbott C.A. et al. in 1993,[10] Cucuianu M et al. in 2002,[7] and Deepa K et al. in 2019.[19]

Deepa K et al., 2019 showed a positive correlation between serum cholinesterase and serum lipid profile except HDL and a negative correlation in respect of HDL, which was statistically significant among obese T2DM patients by comparing to normal T2DM patients. This suggested that serum cholinesterase is intimately related to obesity along with T2DM.[19] But in our study, we have found no such statistical significant correlation between serum cholinesterase and overweight diabetic patients (BMI ≥ 25.0) [Table 3].

The increased cholinesterase activity among diabetic patients was attributed by Inácio Lunkes G et al.,[16] 2005 to a possible interference of this disease in serum cholinesterase’s catalytic mechanism.

Rao AA et al.,[25] 2007 in a reliable explanation proposed that type-2 diabetes is a low-grade systemic inflammatory condition and acetylcholine has anti-inflammatory properties. Hence, the increased serum cholinesterase concentrations in type-2 DM were enhanced by the high acetylcholine levels that were found in type-2 diabetic patients.

The ROC plot analysis in this study demonstrates the higher diagnostic accuracy and 89.77 of serum cholinesterase with sensitivity 96.59 and specificity 82.95 [Table 8]. The area under the curve shows the area is 0.952, standard error 0.016, and a P value of < 0.001, which is statistically significant. The cutoff of serum cholinesterase concentration from ROC is 4557.35. In this study, among 88 diabetic cases, 85 have high serum cholinesterase concentration, whereas among 88 nondiabetic cases, 73 have normal serum cholinesterase concentration. Thus, with a high clinical informative power and relatively low cost, serum cholinesterase may evaluate the clinical conditions of diabetic patients as diagnostic and prognostic parameters with the assessment of complications. This finding was also stated by Das U.N. in 2012.[15]

The limitations of the present study are by the fact that due to its cross-sectional nature, the data were not sufficient for providing cause–effect analysis. Another limitation of our study includes to generalize the serum cholinesterase as a more sensitive and specific tool for diagnosing T2DM; there is a need of community-based study consisting of a large group of subjects.

For future implication, serum cholinesterase may be considered as a probable new marker to assess the complication and pathophysiology of T2DM, and if sufficient research will be done, it may act as one of the diagnostic test in future. As serum cholinesterase can also be used to estimate risk of cardiovascular diseases among T2DM, this enzyme activity can be considered as a routine parameter in T2DM.

Conclusion

Early diagnosis and prevention of the complications of diabetes mellitus is very much necessary due to its rapidly growing nature with a huge health, social, and economic impact globally.

Our study concluded that serum cholinesterase levels are increased in T2DM patients and also showed that serum cholinesterase concentrations are correlated with serum triglyceride levels and serum very low-density lipoprotein levels. It has been also shown in our result that the serum cholinesterase concentration is very much sensitive and specific for T2DM with a fair diagnostic accuracy. As the relationship of triglyceride and atherogenicity in diabetes is well established, we can generalize our study finding to a possible association between elevated serum cholinesterase and vascular complications in diabetes.

To consider in future, serum cholinesterase as a diagnostic and prognostic marker, there is a need for more researches and data on a basis of a large multicentric study all over the World. We are trying through our study to suggest the role of serum cholinesterase as an independent marker of T2DM in our study population.

Conflicts of interest

There are no conflicts of interest.

Funding Statement

Nil.