ABSTRACT-Introduction: Importance of measurement of glycated hemoglobin (HbA1c) has been recommended for the diagnosis of diabetes and pre-diabetes. However, various epidemiological studies conducted different parts of the universe have shown significant discordance between HbA1c and glucose-based tests. Glycated hemoglobin (HbA1c) is assumed to be the gold standard for monitoring glycemic control in patients with diabetes mellitus disorder. The Glycated hemoglobin (HbA1c) assay provided an accurate, precise measure of chronic glycemic levels, and associates with the risk of diabetes complications.
Materials and Methods:This is a cross sectional prospective study. A total of 868 individuals attended to the medicine outpatient clinic at Lord Buddha Koshi Medical College, Saharsa, Bihar between Jan 2016 to Dec 2016 were selected for the study after screening a large cohort visited OPD. The results of FPG, OGTT, and HbA1c for 868 individual were analyzed as well as all grouped as diabetic patients, glucose intolerant (pre-diabetes) patients, and non-diabetic patients according to new ADA criteria for the diagnosis of diabetes.
Results: Diagnostic sensitivity of all diabetic criteria were 80.33% for A1c; 75% for OGTT and only 41.87% for FPG respectively.
Conclusion: The proposed A1c diagnostic criteria have greater diagnostic than FPG and 2-h OGTT regarding a diagnosis of diabetes mellitus disorder.
Key-words- Glycated Hemoglobin, Fasting Plasma Glucose, Oral glucose tolerances test (OGTT), Diabetes Mellitus, and Pre- diabetes

INTRODUCTION- Glycated hemoglobin (HbA1c) has been recommended for the diagnosis of diabetes and pre-diabetes these days. However, various epidemiological studies conducted different parts of the universe have shown significant discordance between HbA1c and glucose-based tests. Some factors that could influence agreement between HbA1c and the oral glucose tolerance test (OGTT), body weight has not been fully evaluated.
Glycated hemoglobin (HbA1c) is assumed to be the gold standard for monitoring glycemic control in patients with diabetes mellitus.
The Glycated hemoglobin (HbA1c) evaluates an accurate, accurate measure of chronic glycemic levels, and associates with the risk of diabetes complications.
The purpose & utility of this test has been extended to diagnose and screen for diabetes mellitus with the endorsement of several international diabetes societies and the World Health Organization. In 2010, the International Expert Committee and the American Diabetes Association postulated diagnostic criteria for diabetes and prediabetes based on HbA1c levels. These are HbA1c
6.5% (
48 mmol/mol) to diagnose diabetes mellitus and between 5.7–6.4% (39–46 mmol/mol) for prediabetes [1]. Since the recommendation of the International Expert Committee in 2009 to use HbA1c test to diagnose diabetes [3], the American Diabetes Association (ADA), the Endocrine Society [5], the World Health Organization [6] and most scientists in different countries all over the world have endorsed using HbA1c to diagnose diabetes.
Epidemiological studies have shown significant between HbA1c and glucose-based tests for defining diabetes and pre-diabetes. The diagnosis of diabetes, HbA1c showed 24% sensitivity and 99% specificity [2]. These levels of sensitivity and specificity were replicated in several other Research Article (Open access)studies [3–7], all suggesting the poor agreement between HbA1c, fasting plasma glucose (FPG) and 2-h plasma glucose (2 h PG).
therefore more , diagnostic agreement of HbA1c criteria with the fasting and 2 h glucose-based criteria for pre-diabetes was also questioned [8-9], and might be different across ethnic groups and populations[10], thus suggesting that the diagnostic performance of HbA1c will depend also on the target population. In the study by Mann [8], for example, pre-diabetes by the HbA1c criterion showed 27% sensitivity and 93% specificity, with 61% positive predictive value, a result confirmed by Heinaza [9], everywhere a threshold of HbA1c 5.7% again showed low sensitivity (24%) with high specificity (91%), whereas HbA1c of 5.5% gave the highest combination of specificity (76%) and sensitivity (46%).
Obesity is one of the most important risk factors for diabetes and impaired glucose regulation [11], It might be postulated that in obese subjects, at increased risk for glucose abnormalities, the efficacy of HbA1c could be higher than in normal weight people, and therefore of increased clinical utility. One current study has shown a modest increased risk of prediabetes linked with obesity [12]. Some exist our knowledge; no studies have explored the impact of different grades of obesity (class I–III) on the efficacy of HbA1c to diagnose diabetes and prediabetes. Last decades some studied were showed that the large population of patients that are discordantly categorized by HbA1c or OGTT; their phenotypic characterization needs to be assessed, in order to identify those parameters that could be of help in the choice of the most appropriate diagnostic tests.
Finally, in last decades studied [13] so far has analyzed that the association between HbA1c and plasma glucose values for the diagnosis of prediabetes, showing again poor agreement between HbA1c and FPG.
Past study HbA1c can be used as a dual marker of hyperglycemia and dyslipidemia in type 2 diabetes mellitus [14]. Improving glycemic control can substantially reduce the risk of cardiovascular events in diabetics [15]. Cholesterol, saturated fats and excessive amounts of sodium have been identified as factors of high blood pressure and Cardiovascular disease [16]. Other factors play a similarly important role, if not more, in the pathogenesis of diabetic complications, oxidative stress plays a significant role in diabetes and its complications [17]. The alteration function of endothelium along with antioxidant/pro-oxidant imbalance in hypertension can lead to detrimental consequences and long-term adverse effects of atherosclerosis and cardiovascular disease [18]. Past study shown between antioxidant nutrient intake and decrease in the development of diabetic complications [19]. Vitamin C may be helpful in decreasing blood glucose type 2 diabetes and thus reducing the risk of complications [20]. Hence, the aims of the current study were to evaluate the impact of HbA1c criteria to diagnose diabetes and pre-diabetes in two large cohorts of participants undergoing OGTT. Then, we aimed to investigate whether differences exist between obesity classes I–III with respect to the relationship of HbA1c and blood glucose. Finally, we examined who had a diagnosis of prediabetes with the OGTT, but had a normal HbA1c, comparing them with those that were concordant with both tests, consider to might most appropriate diagnostic test.

MATERIALS AND METHODS- This is a cross sectional prospective study. A total of 868 individuals attended to the medicine outpatient clinic between Jan 2016 to Dec 2016 were selected for the study after screening a large cohort visited OPD. The results of FPG, OGTT and HbA1c for 868 individual were analyzed and all grouped as diabetic patients, glucose intolerant (pre-diabetes) patients and non-diabetic patients according to new ADA criteria for the diagnosis of diabetes.
Inclusion Criterion: Only those with concurrent FPG, OGTT and A1c results and diabetes mellitus suspicion were included. OGTT is routinely obtained in our hospital, if there is a suspicion of diabetes mellitus.
Exclusion Criterion: Diabetic subjects and patients, who had been using drugs associated with the development of diabetes, were excluded.
Study group consisted of 348 males (40%) and 520 females (60%). Mean age of the subjects were 57.3 ± 18.6 yr. FPG, OGTT, A1c levels of subjects were measured by the help of central laboratory & department of Biochemistry. All individuals subjects (n =868) were grouped as diabetic patients, glucose intolerant (pre-diabetes) patients, and non-diabetic patients according to new ADA criteria for the diagnosis of diabetes disorder. The current diagnostic criteria proposed by American Diabetes Association (ADA) for diabetes are A1c
6.5%, FPG
126 mg/dl (7.0 mmol/l), 2nd h plasma glucose
200 mg/dl (11.1 mmol/l) during the OGTT in the patients with classic symptoms of hyperglycemia or hyperglycemic crisis, a random plasma glucose
200 mg/dl (11.1 mmol/l). IFG was defined as a FPG with 100 mg/dl (5.6 mmol/l) - 125 mg/dl (6.9 mmol/l). IGT was defined as 2-h glucose with 140 mg/dl (7.8 mmol/l)-199 mg/dl (11.0 mmol/l) or A1c values between 5.7% and 6.4%.
Performance of FPG, OGTT, and A1c tests were done with the following steps:

1. FPG: After 12 h fasting period, blood samples were drawn by standard phlebotomy into regular blood (serum) test-tubes between 8:00 AM and 10:00 AM and serum glucose level was measured by an enzymatic method (hexokinase).
2. OGTT: All subjects were informed to take at least 150 g of carbohydrate each day for atleast 3 days before this test. After 12 h fasting period, 75 g of glucose were given to each individual to ingest in the form of a cool drink. Blood samples were taken by standard phlebotomy into regular blood (serum) test tubes at time 0 and 120 min by a health- care provider.
3. HbA1C: Blood samples were obtained by standard phlebotomy into ethylene diamine tetra acetic acid-containing tubes by Nephelometry method was used in the analysis of HbA1c.
   

STATISTICAL ANALYSIS- All results were shown as a mean ± standard deviation. The p values were based on the two- side tests with a cut off for statistical significance of 0.05. The Chi-square test, Kolmogorov- Smirnov test, and Analysis of co-variance test were used to evaluate values. All statistical analyses were performed with SPSS of version 20.0.

RESULTS AND DISCUSSION- The present study showed the poor agreement between HbA1c, FPG, and 2-h glucose post OGTT for the diagnosis of Diabetes Mellitus and Pre-diabetes. This investigation compared between FPG and HbA1C rapid tests in identifying diabetes and pre-diabetes when used in a screening strategy among 35–74 year old persons.
HbA1c, therefore, could reproduce a mishmash of the patho-physiological defects underlying IFG and IGT over time. In fact, we examined the highest concordance with HbA1c when the two conditions of IFG + IGT were present together.
According to new ADA criteria; we were determined 480 diabetic patients among the 868 individuals (55.3%). However, 96 diabetic patients (20.0%) met all ADA criteria. All results are shown in Table 1 & Table 2. 388 diabetic patients (80.83%) were diagnosed by A1c alone, 360 diabetic patients (75%) with 2-h OGTT alone, and 201 (41.87 %) diabetic patients were diagnosed with FPG alone (Table 2). Differences between FPG versus 2-h OGTT, FPG versus As1c and OGTT versus A1c were statistically significant (p <0.05, p <0.05 and p<0.05, respectively). Diagnostic sensitivity of all diabetic criteria was found 80.33% for As1c; 75% for OGTT, and only 41.87% for FPG respectively.

Table 1: Frequency of individuals and distribution of mean FPG, 2-h OGTT and HbA1c values of the diabetic patients and non-diabetic individuals according to age groups

Age Grup	Gender		Total	Test	Non Diabetic Patients	Diabetic Patients	All individuals
	Male	Female
45 yrs	80	101	181	FPG	112±32	148±61	96±12
				2-HOGTT	149±82	232±112	139±31
				HbA1c	6.11±1.37	7.98±1.72	5.01±0.88
45-60 yrs	154	206	366	FPG	115±41	138±51	91±19
				2-HOGTT	149±71	232±89	132±21
				HbA1c	5.81±1.41	7.08±1.02	6.01±0.28
60 & Above	122	199	321	FPG	118±30	158±68	86±19
				2-HOGTT	165±80	252±119	131±39
				HbA1c	5.31±1.27	8.98±1.62	5.31±0.81

FPG= Fasting Pasma glucose, OGTT= Oral glucose tolerance test, HbA1c = glycated hemoglobin

Table 2: Pre-diabetes and diabetes frequencies

S. No	Diagonostic criterion	Positive	Negatiave	Total	p -value
1	FPG 2-hOGTT HbA1c IFG	201 360 388 304	279 120 92 176	480 480 480 480	p<0.05
2	IGT 2-h OGTT IGTHbA1c IFG	86 123 154	394 357 326	480 480 480	p<0.05
3	IGT 2-h OGTT IGT HbA1c	170 401	698 467	868 868	p<0.05

1. Results according to new ADA Criterion, 2. Frequencies of imparied fasting glucose & imparied glucose tolerence (2-h OGTT & HbA1c),
3. Frequency of impaired fasting glucose & imparied glucose tolerence (2-h OGTT & A1c) among diabetic patients
FPG= Fasting plasma glucose, OGTT= Oral glucose tolerance test, IFG = Impaired fasting glucose, IGT = Impaired glucose tolerance

IFG and glucose intolerance utility:According to new ADA criteria, of the 868 subjects tested, 1094 (60.3%) were classified as having IFG, 154 (32.08%) as having IGT following OGTT and 401 (46.01%) as having IGT by A1c. In terms of a diagnostic ratio of glucose intolerance; the difference between A1c and OGTT was statistically significant (p< 0.05) [Table 2 & Table 3].

Table 3: Distribution of all diabetic patients according to FPG, 2-H OGTT and HbA1c

	Fasting PlasmaGlucose mg/dL		Total	2-h OGTT mg/dL			Total
	<126		>126	<140	140-200	>200
A1C <6.5	336	154	490	313	217	98	628
A1C <6.5	267	111	378	101	88	51	240
Total	603	265	868	530	305	149	868

CONCLUSION- By using A1C as a marker of diabetes criterion would re-classify the diabetes diagnosis. It is suggested that clinicians and health systems understand the differences and similarities by using A1c or FPG and 2-h OGTT in the diagnosis of diabetes mellitus and pre-diabetes. The proposed A1c diagnostic criteria have greater diagnostic than FPG and 2-h OGTT regarding the diagnosis of diabetes mellitus.

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How to cite this article: Jha NK: Role of Glycated Hemoglobin in the Diagnosis of Diabetes Mellitus and Pre-diabetes. Int. J. Life. Sci. Scienti. Res., 2017; 3(2): 882-886. DOI:10.21276/ijlssr. 2017.3.2.1 Source of Financial Support: Nil, Conflict of interest: Nil