Right Dorsolateral Frontal Lobe N-Acetyl Aspartate and Myoinositol Concentration Estimation in Type 2 Diabetes with Magnetic Resonance Spectroscopy

Globally 387 million people are suffering with diabetes out of which 90% of the cases are type 2 diabetic in nature, it is expected that 592 million people are going to suffer with diabetes by 2035 [1]. Type 2 diabetes mellitus is the result of non-responsiveness by the peripheral tissues to the secreted insulin or by the decreased secretion of insulin from the β cells of pancreas and the aetiological factor for type 2 diabetes is mainly obesity [2]. Reduced frontal lobe volumes are observed in obese type 2 adolescents [3]. Type 2 diabetes affects the functioning of central nervous system (cognition) by altering the brain metabolite concentrations [4]. Regular yogasana and pranayama enhances the functioning of central nervous system (cognition) [5]. Magnetic resonance spectroscopy (MRS) enables the identification and quantification of metabolites in given sample [6]. As frontal lobe is involved in executive functions and working memory, we are interested in finding, how the N-acetyl aspartate (NAA) & myoinositol (mI) concentrations are altered in the right dorsolateral frontal lobe of type 2 diabetic subjects and also to observe the effects of yogasana and pranayama on these metabolite concentrations. Change in the metabolite concentrations in the right dorsolateral frontal lobe will affect the memory [7,8]. Glycosilated haemoglobin (HbA1c) concentration is related to the development of complications and it reflects the mean plasma glucose levels over the last 2-3 months and has a minimum effect from the drugs that are influencing the glucose metabolism [9]. Absence or decreased concentration of N-acetyl aspartate (NAA) is a sign of neuronal loss or degradation [6,10] and it might reflect intrinsic imbalance between excitatory versus inhibitory neurons [11].

Increased myoinositol (mI) concentration is observed with proliferation of glial cells or with increased glial-cell size, as found in inflammation [6]. Blood glucose levels will be elevated in type 2 diabetes [12]. In type 2 diabetic subjects, increased blood glucose levels will also increase the brain glucose levels and increased brain glucose levels will alter the brain metabolite concentrations [13]. The study is aimed to observe the HbA1c levels, NAA and mI concentrations in right dorsolateral frontal lobe in type 2 diabetic subjects who did the yogasana and pranayama and to compare the same with diabetic controls.

Materials and Methods

The present study was approved by the Institutional Ethical Committee.

Inclusion criteria: Type 2 diabetes patients, both the sex are included, minimum duration of diabetes is 2 years, age between 35-65 years were included in the study.

Exclusion criteria: Smokers, alcohol consumers, any neuro-psychological disorders, recent history of major surgeries & those who are on insulin treatment were excluded. Sixty eight type 2 diabetic subjects were recruited in this study after taking the written informed consent. Thirty four type 2 diabetic test group subjects did the yogasana and pranayama for 6 months, 6 days in a week, for a period of 45-60 minutes daily under the supervision of a qualified yoga expert in Yogi Vemana Yoga Research Institute in Hyderabad. Age and sex matched 34 type 2 diabetic subjects, who are not on any specific exercise regimen were included in the study as control group. Test group subjects did a set of yogasana followed by a set of pranayama which are listed in detail in [Table/Fig-1,2] respectively. Both the control and test group subjects are on oral hypoglycaemic agents. HbA1c levels were estimated by using the Bio-Rad D-10™ haemoglobinA1c program, and it is intended for the percent determination of haemoglobin A1c in human whole blood. It is based on high-performance liquid chromatography (HPLC). Samples will be injected in to the analytical cartridge after diluting them on D-10, programmed buffer gradient of increasing ionic strength will be delivered to the cartridge by the D-10 and here the haemoglobins are separated. The D-10 software and the exponentially modified Gaussian (EMG) algorithm will do the rest in quantifying the HbA1c levels [14]. The D-10 Haemoglobin A1c program is for use only with the Bio-Rad D-10 Haemoglobin Testing System [14]. Magnetic Resonance Spectroscopy was performed in the Department of Radiology in MediCiti Institute of Medical Sciences. Metabolite concentrations in right dorsolateral frontal lobe were estimated with 1.5 Tesla Phillips magnetic resonance imaging machine.

[Table/Fig-1]:

List of Yogasana

S. No	Name of Yogasana	Duration
1	Dhanurasana	1/2 minute to one minute for the pose being maintained, adding 1/2 minute per week
2	Naukasana	2 - 4 turn of each, the pose being maintained for ten seconds adding one turn each, every fortnight
3	Arthamasthendrasana	¼ minute to one minute for each side, adding ¼ minute per week
4	Bhujangasana	2 - 4 turn of each, the pose being maintained for ten seconds adding one turn each, every fortnight
5	Shavaasana / Makarasana	3 turn of each, the pose being maintained for 30 seconds

[Table/Fig-2]:

List of Pranayama

S. No	Name of Pranayama	Duration
1	Anuloma-viloma	2-5 minutes
2	Surya anuloma-viloma	5 minutes
3	Chandra anuloma-viloma	5 minutes
4	Nadishuddi pranayama	10 minutes

Acquisition parameters

Right dorsolateral frontal lobe NAA and mI concentrations were estimated with Magnetic Resonance Spectroscopy (MRS) using a single voxel technique. Single voxel point resolved spectroscopy (PRESS) sequence was used for volume of interest (VOI) localization (TR/TE =1800 msec/36msec; NSA 96; Spectral band width 1000; scan time 3 min 25 sec) [6]. Based on the axial T2-weighted image, a voxel (20×20×15 mm) was positioned in the frontal white matter, avoiding the cortex and lateral ventricle. MOIST was used for water suppression in spectroscopy sequences and spectral correction was done [6]. Only peaks of NAA and mI were estimated but not the peaks of other metabolites like choline, glutamate, glutamine, acetylcholine etc.

Statistical Analysis

The results were analysed by using Med Calc Statistical Software version 12.7.8 (Med Calc Software bvba, Ostend, Belgium; http://www.medcalc.org; 2014). Unpaired student’s t-test was used in finding the metabolite concentration between test and control groups. A p-value <0.05 was considered as statistically significant.

Results

[Table/Fig-3] shows the HbA1c levels in control and test group subjects and they are 7.7 ± 1.84 and 6.02 ± 0.46 respectively. The p-value for HbA1c is <0.001 which is highly significant statistically. [Table/Fig-4] shows the NAA concentrations in the right dorsolateral frontal lobe of control and test group, and they are 1.44 ± 0.15 and 1.54 ± 0.19 respectively. The p-value for NAA is 0.02. [Table/Fig-5] shows the mI concentrations in the right dorsolateral frontal lobe of control and test group, and they are 0.61 ± 0.22 and 0.47 ± 0.24 respectively. The p-value for mI is 0.01. [Table/Fig-6] is the graphical representation of Magnetic Resonance Spectroscopy (MRS) of right dorsolateral frontal lobe in a given subject, showing the peaks of the different metabolites.

[Table/Fig-3]:

HbA1c levels in control and test group subjects

[Table/Fig-4]:

N-acetyl aspartate concentration in right dorsolateral frontal lobe in control and test group

[Table/Fig-5]:

Myoinositol concentration in right dorsolateral frontal lobe in control and test group

[Table/Fig-6]:

Right dorsolateral frontal lobe MRS in a subject

Discussion

High HbA1c levels and low NAA concentration are observed in controls when compared to yogasana and pranayama group that are reflected in [Table/Fig-3,4] respectively. Low concentration of NAA in control group is because of decreased neuronal viability [6,11]. It indicates that neuronal loss is the cause for low concentration of NAA in type 2 diabetes, this neuronal loss is the result of osmolyte imbalance and this is due to accumulation of advanced glycosilated end products [6]. Myoionositol concentration is more in controls than the yogasana and pranayama group as shown in [Table/Fig-5]. Increased mI concentration is an indication of myelin degradation, proliferation of glial cells or with increased glial cell size i.e., inflammation [15]. Increased HbA1c levels in controls will increase the right dorsolateral frontal lobe glucose levels [13]. High glucose levels in right dorsolateral frontal lobe will increase the myoinositol concentration in type 2 diabetic subjects [16–18] and decreases the N-acetyl aspartate (NAA) concentration [13,16]. Myoinositol (mI) concentration is more in diabetic controls than the yogasana and pranayama group and this is either because of increased intra neuronal glucose which have converted in to inositol 1-phosphate which in turn converted in to mI [19] or because of increased intracellular osmotic pressure which have led the increase in mI levels [6]. Where as in a study by Sinha et al., there is no significant change in the mI concentration in right frontal lobe although increased right frontal lobe glucose was observed [13]. Further studies are required in analysing this contradicting variation in frontal lobe myoinositol (mI) in type 2 diabetes. In test group yogasana and pranayama normalized or near normalized the HbA1c levels either by increasing the peripheral uptake of glucose or by increasing insulin secretion by beta cells of islets of Langerhas [20]. Increased uptake of peripheral glucose or increased insulin secretion in test group is because yogasana and pranayama stimulate the parasympathetic nervous system resulting in normal HbA1c levels. Normal or near normal plasma HbA1c levels might have kept the right dorsolateral frontal lobe glucose levels in control. Normal right dorsolateral frontal lobe glucose levels maintained the near normal N-acetyl aspartate (NAA) and myoinositol (mI) concentrations in test group subjects. Thus yogasana and pranayama are helpful in stabilizing the plasma HbA1c levels [20–33] and stable HbA1c levels will keep the N-acetyl aspartate (NAA) and myoinositol (mI) concentrations near normal values in right dorsolateral frontal lobe. Outcome of this study is, yogasana and pranyama should be advised to type 2 diabetic patients for maintaining the normal functioning of the right frontal lobe by keeping the normal N-acetyl aspartate (NAA) and myoinositol (mI) concentrations.

Study limitations and Future Prospects

We could not measure the type and quantity of food the subjects were taking. Measuring the stress levels also help in better understanding the role of yogasana and pranayama on HbA1c levels. Difficult to record the number of hyperglycaemic and hypoglycaemic attacks per day in diabetic subjects for assessing the effects of the same on brain metabolites. All the brain metabolite concentrations should be explored in multiple lobes of the brain in type 2 diabetes. By knowing the degree of variations of these metabolites in different lobes of the brain with HbA1c levels one can take the prophylactic measures.

Conclusion

In test group, yogasana and pranayama reduced the HbA1c levels to near upper limit of the normal values and this near normal HbA1c level is restricted the wide variations of right dorsolateral frontal lobe N-acetyl aspartate (NAA) and myoinositol (mI) concentrations, which are seen in diabetic controls.

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