European Journal of Cardiovascular Medicine

According to IDF Diabetes Atlas, global diabetes prevalence in 20–79-year-olds in 2021 was estimated as 10.5% (536.6 million people) but will be rising to 12.2% (783.2 million) in 2045.¹ WHO suggests that over 19% of the world’s diabetic population currently resides in India. According to an ICMR-INDIA national study, India has 62.4 million people with type 2 diabetes (T2DM) and 77 million people with pre-diabetes. These figures are expected to rise to 101 million by 2030.²

T2DM is associated with cell-mediated immune responses and abnormal T-lymphocyte function, further linked to insulin deficiency. Immunological disturbances in T2DM are associated with cell-mediated responses and abnormal T-lymphocyte function, both of which are important in diabetes and are linked to insulin deficiency. Adenosine deaminase (ADA), a cytosolic enzyme of purine metabolism is considered a good marker of cell-mediated immune response. ADA has two major isoenzymes ADA1 and ADA2, which are distributed in human tissues, with its highest activity in T-lymphocytes. Adenosine deaminase (ADA) catalyses the hydrolytic cleavage of adenosine and other adenosine nucleoside analogues into inosine and ammonia. Inosine and 2′-deoxyinosine are metabolically converted into 5 hypoxanthine, xanthine, and uric acid.³ According to Warrier et al., adenosine mimics the action of insulin on glucose and lipid metabolism in adipose tissue and the myocardium.⁴ Adenosine modulates insulin action on different tissues in different ways. Through its powerful antilipolytic properties, ADA reduces the free fatty acids and improves insulin sensitivity in adipose tissue. ADA has been previously identified as an indicator of insulin function. Studies reported increased ADA in T2DM compared with healthy individuals. There were no reports cross-sectionally detected the activity of ADA and glycemic control in subjects with T2DM. Hence this study aims to determine the activity of serum total ADA and correlate these parameters with glycemic control, and biochemical variables in type 2 diabetic individuals.

AIMS & OBJECTIVES:

Aim

To estimate serum adenosine deaminase and its correlation with glycemic status in diabetes mellitus.

Objectives

• To estimate the serum adenosine deaminase levels in type 2 diabetes mellitus patients.
• To analyze the correlation between adenosine deaminase levels and glycemic status in patients with type 2 diabetes mellitus.

Study Design:

Hospital-based analytical cross-sectional study conducted at General Medicine OPD of Sri Venkateswara Ramnarain Ruia Government General Hospital, Tirupati. Study protocol approved by Institutional Ethical Committee, S.V. Medical College, Tirupati. Informed consent was obtained from all participants.

Sample Size:

100 cases of diabetes mellitus and 100 non-diabetic individuals.

Study group of T2DM cases were divided into 3 groups, such as

Controlled/good glycemic control and Uncontrolled/poor glycemic group based on HbA1c level (≤ 7% = Controlled and > 7% = Uncontrolled).

Group A: 100 non-diabetic healthy individuals- age and sex matched were enrolled for comparison and Diabetics with good glycemic control (n=50) with HbA1c ≤ 7%.

Group C: Diabetics with poor glycemic control (n=50) with HbA1c >7 %, who were

on oral hypoglycemic drugs.

Inclusion Criteria for Diabetics:

1. Individuals with diagnosed T2DM in the age group 35 to 70 years (both males and females) visiting medicine OPD of SVRR Government Hospital, Tirupati.
2. Patients who are willing to participate in the study and willing to give informed written consent.

Exclusion Criteria for Diabetics:

• Individuals with a history of tuberculosis, rheumatoid arthritis, viral hepatitis, and HIV.
• Patients on insulin therapy.
• Patients who are not willing to participate.
• SYSTEMIC HYPERTENSION: Patients with Systolic BP > 130 & Diastolic BP > 80mmHg.

Investigations included:

            Primary investigations FBS; HBA1C; serum ADA.

Laboratory Ranges:

• Fasting Blood Sugar (FBS) - 60-100 mg/dl
• Post-Prandial Blood Sugar (PPBS) - 80-140 mg/dl
• HBA1C - < 7%

Serum ADA

Serum ADA was analyzed by the Colorimetric method described by Guiseppe Guisti.

Activity was measured by endpoint colorimetric method. (Kit: Quantitative determination of ADA activity by colorimetric endpoint method Mfg. by Tulip diagnostics).

The data was collected, tabulated and analyzed statistically

In this study, 41 cases were males and 59 were females. The mean age was 55.29±6.2349 years. The mean FBS and PPBS were 146.11 ± 12.567g/dl and 188.88±24.16 mg/dl in T2DM subjects. 64% of T2DM patients were hypertensive, and 31% of non-diabetics had hypertension. Obesity was found in 12% of diabetic patients. Other risk factors such as smoking [32%], alcohol [17%] and dyslipidemia [50%] were present in diabetic cases. A family history of diabetes was seen in 28% of the diabetic population. Patients with controlled glycemic control had a history of the disease since 7.06± 1.12089 years, while poor glycemic control had a history of diabetes since 11.78±2.367 years.

Table 1. Mean values of glycemic variables in diabetics and non-diabetics.

Mean of FBS was significantly higher in poor glycemic control (Group C) (164.26±25.93mg/dl) compared to good glycemic control (Group B) (140.56±15.0714mg/dl) and (Group A) (107.7±9.8005mg/dl) with p < 0.001. Mean HbA1C was significantly higher in poor glycemic control (Group C) (8.756±1.1607%) compared to good glycemic control (Group B) (6.866±0.1458%) and non-diabetics (Group A) (5.365±0.336%) with p < 0.001.

Table 2: Comparison of Mean HbA1c in three groups

Mean HbA1C was significantly raised in poor glycemic control (Group C) (8.756±1.1607%) compared to good glycemic control (Group B) (6.866±0.1458%) and non-diabetics (Group A) (5.365±0.336%) with p < 0.001. The levels of HbA1c in diabetics are higher than in non-diabetics and the difference is highly significant (P<0.001).

Mean FBS was significantly raised in poor glycemic control (Group C) (164.26±25.93mg/dl) compared to good glycemic control (Group B) (140.56±15.0714mg/dl) and non-diabetics (Group A) (107.7±9.8005mg/dl) with p < 0.001. The study also reported that the mean PPBS in good glycemic control (173.16±15.7808) was lower than with poor glycemic control (203.36±28.5619 mg/dl) with statistically significant (P<0.001).

Table 3: Mean ADA of parameters in Diabetics and Non-Diabetics

ADA activity was significantly higher in uncontrolled (Group C) diabetic patients (55.428±3.736U/L) compared to the controlled (Group B) population (42.63±4.59 U/L) and non-diabetics (22.0581±5.1968 U/L) with p value 0.001. Patients who have a longer duration of diabetes history had higher serum ADA levels than newly diagnosed patients with significant differences, p < 0.001.

Table 4: Association between mean ADA level and HbA1c

A significant association was observed between HbA1C levels and ADA. Patients with HbA1>7 had high serum ADA levels compared to patients with HbA1c <7, and healthy individuals with significant differences, p<0.05.

Pearson’s correlation coefficient analysis for the relationships between serum ADA, FBS, and HbA1c levels in poor glycemic control (group C) depicted a strong positive correlation between HbA1c and ADA (r=0.3256; p=0.0012), and between FBS and serum HbA1c levels (r=0.610). Good glycemic control cases (group B) depicted a weak positive correlation between the HbA1c and ADA (r=0.2546; p=0.0079) and between FBS and HbA1c (r=0.66), and ADA and FBS (r=0.062).

Adenosine deaminase is (ADA) an important enzyme for modulating the bioactivity of insulin, on glucose/lipid metabolism via decreased amounts of tissue adenosine content. ADA is widely distributed in human tissues, the highest being in lymphoid tissue, and is involved in immune system development, particularly in the proliferation and differentiation of lymphoid cells and plays a major role in various stages of lymphocyte maturation. Two major isoforms of ADA such as ADA1, exist in all human tissues and account for the main ADA activity of most of the tissues. ADA2 is an isoenzyme in serum originating mainly from the monocyte-macrophage system.

This current study aims to correlate ADA as a marker of altered glycemic status in T2DM to glycemic control as analyzed by HbA1C. Relevant clinical history was noted, clinical examination and laboratory investigations were done on 100 patients of T2DM, correlation of HbA1c and serum ADA was done in these subjects taking into consideration the glycemic control.

Among 100 T2DM patients in our study, 41 were males and 59 were females. The mean age ± SD for the whole population was 55.29±6.234953 years. Whereas a study by Suhail Ahmed Almani et al.,⁵ reported that the mean ±SD of type 2 DM cases was 52.87±8.85 with 35 male and 15 female populations. In patients with T2DM cases, the mean ±SD for fasting and PPBS was 146.11 ± 12.56730969 and 188.38±37.43 mg/dl. Whereas the study by Suhail Ahmed Almani et al.⁵ reported the mean ±SD for fasting blood glucose (FBG) and random blood glucose (RBG) was reported as131.21±4.82 and 230.63±9.94 mg/dl.

Comparison of groups-FBS:

In our study, the mean FBS in Group A was 107.7±9.8005 mg/dl, Group B was 140.56±15.0714 mg/dl and in Group C subjects was 164.26±25.936 mg/dl. The mean FBS in the controlled diabetic (HbA1c<7) group and uncontrolled diabetic (HbA1c>7) group was more significant than Group A (p <0.001). Kundu et.al.,⁶ reported that the mean FBS levels were in the control (88.00±10.00), the controlled diabetic group (129.18±21.67) and in uncontrolled T2DM group (137.04±20.97). The study by Amandeep Kaur et.al.,⁷ reported FBS in control was 82.00 ± 13.00 mg/dl, HbA1c<7% group was 126.12 ± 22.71 mg/dl and in 136.97 ± 24.88 mg/dl in group HbA1c>7%. Ray D. et. al.,⁸ study shows that the mean FBS of control was 88.00±10.00 mg/dl, controlled DMT2 group was 129.18±21.67 mg/dl and in uncontrolled DMT2 group subjects was 137.04±20.97 mg/dl.

Comparison of groups-PPBS:

In our study, PPBS levels in Group A, Group B, and Group C were 130.52±14.2647, 173.4±15.713, and 203.36±28.56 mg/dl respectively. Student’s t-test (two-tailed) analysis showed that PPBS levels in T2DM were higher than in non-diabetic individuals with a significant difference (P<0.001). The study by Niraula A, et al.,⁹ showed that PPBS levels were significantly increased in uncontrolled Diabetic patients (242.72±136.63mg/dL) than in healthy individuals (130.54±35.55mg/dL).

Comparison of groups-HbA1C levels

In our study, the mean HbA1c in non-diabetic group A was 5.365±0.336 %, in Group B was 6.866±0.1458% and 8.756±1.1607% in Group C. Mean HbA1c between Group B and Group C subjects were significantly higher than non-diabetic subjects in Group A (p <0.001). Kundu D. et. al.,⁶ studies reported that HbA1c levels in healthy individuals were (5.70±0.40%), controlled diabetic group was (6.10±0.49%) and in the uncontrolled diabetic group was (8.90±1.02 %). In the study by Ray D. et. al.,⁸ reported HbA1c levels in healthy individuals were 5.70±0.40%, in controlled Diabetic patients were 6.10±0.49% and 8.90±1.02 % in uncontrolled Diabetic patients. Amandeep Kaur et.al.,⁷ reported similar findings, which showed HbA1c in Group C subjects was 5.75± 0.46%, in Group B subjects was 6.09± 0.56% and 8.72± 1.35% in Group C subjects. Singh S et al.¹⁰ found that HbA1c levels in the control group were 5.420.38% and 10.08 2.46% in the diabetic group, which was statistically significant (P<0.001).

Comparison of groups- ADA

In our current study, the mean serum ADA in Group A was 22.0581±5.19688 U/L, Group B was 42.63±4.59779 U/L, and Group C was 55.428±3.736 U/L. Statistical analysis revealed that the mean serum ADA in Group C was significantly higher than in Group A and Group B (p<0.001). A study by Amandeep Kaur et.al.,⁷ reported the mean serum ADA in the control Group A was 17.30± 7.28 U/L, 30.04± 10.41 U/L in study Group B, and 44.23± 16.14 U/L in

Group C with significant difference between groups (p<0.001). Kundu D. et.al.,⁶ reported the mean serum ADA in the control (17.30±7.08 U/L), controlled diabetic group (31.05±9.49 U/L), and 44.03±15.10 U/L in the uncontrolled diabetic group.

Spearman's rho correlation was used to compare serum ADA levels to FBS, PPBS, and HbA1c in T2DM patients. Our study found that serum ADA levels (U/L) were significantly higher in uncontrolled (HbA1c >7%) diabetic patients (55.4283.736) than in controlled (HbA1c< 7%) diabetics (42.63±4.5979) and non-diabetics (22.0581±5.196), p-value < 0.001. Serum ADA and HbA1c [r=0.29, p=0.004], Fasting blood sugar [r=0.2, p=0.03], and PPBS [r=0.32, p=0.01] levels all had a significant positive correlation. In a recent study, Spearman's correlation coefficient showed 0.728, indicating a strong correlation between ADA and HbA1C with a statistically significant P = 0.001.¹¹

This Hospital-based cross-sectional study was conducted on 100 patients with T2DM, and 100 non-diabetic individuals (group A). T2DM patients were categorized into HbA1c <7% = good glycemic Controlled Diabetes (group B), and HbA1c > 7% = poor or uncontrolled diabetics (group C). Among 100 T2DM cases, 41 cases were males and 59 cases were females with a mean age of 55.29±6.2349 years. The mean FBS and PPBS were 146.11 ± 12.567g/dl and 188.88±24.16 mg/dl in T2DM cases.

This study's findings clearly show that Adenosine deaminase (ADA) levels are elevated in type 2 diabetics, and the positive correlation of ADA with higher glycemic control suggests that ADA may serve as a prognostic factor in T2DM. ADA had a significant positive correlation with HbA1c, which is regarded as a good marker for long-term glycemic control.

1. Sun H, Saeedi P, Karuranga S, Pinkepank M, Ogurtsova K, Duncan BB, Stein C, Basit A, Chan JC, Mbanya JC, Pavkov ME. IDF Diabetes Atlas: Global, regional and country-level diabetes prevalence estimates for 2021 and projections for 2045. Diabetes research and clinical practice. 2022 Jan 1;183:109119.
2. Whiting DR, Guariguata L, Weil C, Shaw J. IDF Diabetes Atlas: Global estimates of the prevalence of diabetes for 2011 and 2030. Diabetes Res Clin Pract 2011;94:311-21.
3. Hoshino T, Yamada K, Masuoka K, Tsuboi I, Itoh K, Nonaka K, et al. Elevated adenosine deaminase activity in the serum of patients with diabetes mellitus. Diabetes Res Clin Pract. 1994;25(2):97–102.
4. Warrier AC, Rao NY, Kulpati DS, Mishra TK, Kabi BC. Evaluation of adenosine deaminase activity and lipid peroxidation level in diabetes mellitus. Indian Journal of Clinical Biochemistry 1995;10(1): 9 -13.
5. Suhail Ahmed Almani et al, Serum Adenosine Deaminase Level in Patients with Type 2 Diabetes Mellitus, Indo Am. J. P. Sci, 2017; 4(07).
6. Kundu D, Sen S, Paul A, Chatterjee A, Sarkar P, Chakrabarti I. Diagnostic value of serum adenosine deaminase in Type 2 diabetic patients. Int. J. Med. Res. Rev. 2019;7(1):1-7.
7. Kaur A, Kukeraja S, Singh T, Sandhu IS, Singh G. Effect of adenosine deaminase activity on lipid profile in patients of type 2 diabetes mellitus. Group. 2016;100(30):30.
8. Ray D, Kundu D, Choudhury DG et al. Relation of elevated serum adenosine deaminase levels to glycated hemoglobin and serum uric acid in type 2 diabetes mellitus. International Journal of Medical Research and Review July, 2016/ Vol. 4/Issue 7.
9. Niraula A, Thapa S, Kunwar S, Lamsal M, Baral N, Maskey R. Adenosine deaminase activity in type 2 diabetes mellitus: does it have any role? BMC endocrine disorders. 2018 Dec;18(1):1-5.
10. Singh S, Suri A, Sinha M, Fiza B. A study on the correlation of adenosine deaminase and glycated haemoglobin in the patients of type 2 diabetes mellitus. International Journal of Medical and Biomedical Studies. 2019;3(12):43-50.
11. Aishwarya R. To Study the Correlation of Serum Adenosine Deaminase Levels with Hba1c in Patients of Type 2 Diabetes Mellitus. J Assoc. Physicians India. 2022 Apr;70(4):11-12.