European Journal of Cardiovascular Medicine

Type 2 diabetes mellitus, also known as non-insulin dependent diabetes mellitus or adult-onset diabetes, is characterized by elevated blood glucose levels due to insulin resistance and relative insulin deficiency. Its incidence is rapidly increasing, with a projected doubling by 2030 [1,2].

The autonomic nervous system (ANS) innervates nearly all organ systems and plays a crucial role in maintaining homeostasis, regulating visceral activities, modulating the body's responses to environmental changes, stress, and exercise, and assisting the endocrine system in regulating various functions. Diabetic neuropathy refers to nerve dysfunction associated with diabetes mellitus (DM), including autonomic neuropathy, that affects gastrointestinal, cardiovascular sudomotor, genitourinary, and metabolic systems. Cardiac autonomic neuropathy specifically results from autonomic nerve fiber damage to the heart and blood vessels, leading to altered heart rate (HR) control and vascular dynamics [3-5].

Cardiovascular autonomic function tests (CAFTs) are noninvasive tools extensively validated in clinical trials for evaluating baroreceptor reflex and identifying individuals at risk of cardiac complications due to DM. These tests, including HR and blood pressure (BP) responses to standing, deep breathing, and isometric handgrip, aid in early intervention to prevent morbidity and mortality in DM patients [6,7]. This study aimed to compare heart rate variability and classical AFTs in individuals with type 2 DM and healthy volunteers

Subjects aged 30 to 60 years of both genders diagnosed with T2DM for a duration of three years were included in the study (n = 89). The control group comprised normal healthy volunteers within the same age group (n = 89). Diabetic cases were recruited from the OPDs based on the inclusion criteria, while controls were selected from the medical OPD among non-diabetic healthy volunteers. Detailed information regarding present and treatment histories was obtained from all participants.

Patients using medications known to affect autonomic functions (such as Beta-Blockers, anticholinergics, antidepressants, and antipsychotics), those with psychiatric illnesses impacting autonomic functions (anxiety, depression, psychosis), uncontrolled long-term hypertension despite medication for over five years, individuals with Hypothyroidism/Hyperthyroidism, recent history of fever, meningitis, or encephalitis affecting CNS function or structure, alcohol/substance abusers, individuals with cardiac pacemakers or other implanted/external electrical devices, and pregnant or lactating females were excluded from the study.

HRV Analysis and AFTs:

Time-domain analysis involved applying descriptive statistical tools to quantify variations in RR intervals and compute parameters like Standard Deviation of RR intervals (SDNN), Root mean square of successive differences (RMSSD), Number of successive NN intervals varying by more than 50 ms (NN50), and Percentage of NN50 counts (pnn50). Frequency-domain analysis utilized the Fast Fourier Technique (FFT) to transform R-R intervals into waves, analyzing components such as very low frequency (VLF), low frequency (LF), and high frequency (HF). LF normalized unit (LF nu) and HF normalized unit (HF nu) were calculated, and LF/HF Ratio indicated sympathovagal balance and sympathetic modulation. Expiration-inspiration ratio, Valsalva ratio, Heart rate rise after hand grip and DBP rise after hand Grip were also recorded.

Biochemical Analysis:

Fasting and postprandial blood samples were collected for plasma glucose estimation by the hexokinase method, and HBA1C was measured using high-performance liquid chromatography (HPLC). Prediabetes and diabetes were diagnosed based on specific criteria established by the American Diabetes Association (ADA).

Statistical Analysis:

Data analysis was performed using IBM SPSS (Version 20.0). Since the data were not normally distributed, descriptive statistics like median, quartiles for continuous variables, and frequency/percentages for categorical variables were employed. Non-parametric tests were used to analyze differences between subgroups, with a significance level set at P<0.05.

The anthropometric parameters of the study participants are summarized in Table 1. The mean age of the cases was 49.05 ± 7.54 years, compared to 48.70 ± 5.36 years for the controls, with no statistically significant difference between the groups. Similarly, there was no significant difference in the mean height, weight, BMI or the waist-hip ratio. Thus, both groups were similar in terms of anthropometric variables.

Table 1: Anthropometric parameters among study groups

Table 2 displays the HRV time domain parameters for cases and controls. The mean SDNN was significantly lower in cases (30.18 ± 3.79 ms) compared to controls (36.65 ± 4.92 ms). Similarly, the RMSSD was significantly lower in cases (23.25 ± 3.66) than in controls (29.36 ± 3.12) with a P value of <0.05.

Table 2: HRV Time domain parameters in cases and controls

As shown in Table 3, the frequency domain parameters of HRV also indicated significant differences between the groups. The mean VLF (very low frequency) power was higher in cases compared to controls. LF (low frequency) power was significantly higher in cases than in controls, and HF (high frequency) power was higher in cases compared to controls. The LF/HF ratio was significantly elevated in cases relative to controls with a P value of <0.05.

Table 3: HRV Frequency domain parameters in cases and controls

Table 4 presents the results of the autonomic function tests (AFTs). The expiration-inspiration ratio was significantly lower in cases than in controls. The Valsalva ratio was also significantly reduced in cases compared to controls. Additionally, the heart rate rise after hand grip was significantly lower in cases than in controls. The diastolic blood pressure (DBP) rise after hand grip was markedly lower in cases compared to controls. Overall, the data indicated significant impairments in HRV time and frequency domain parameters, as well as in autonomic function tests, among cases compared to controls.

Table 4: Comparison of AFTs cases and controls

(HRV) derived from short-term analysis, was significantly reduced in patients with long-term type 2 diabetes mellitus (LT2DM) compared to controls. This reduction was more pronounced in DM patients with longer disease duration, consistent with previous findings showing decreased overall HRV in these patients [8]. Similarly, mean RR interval, root mean square of successive differences (RMSSD), and variance were notably decreased in LT2DM patients, indicating marked impairment of parasympathetic nerve function with minor involvement of the sympathetic component, in agreement with prior studies [9,10]. However, these values were comparable to controls in the recently diagnosed group, suggesting minimal effects in these patients compared to previous reports, potentially due to differences in patient selection criteria [11]. Duration of DM has been linked to the degree of autonomic dysfunction in numerous studies [12].

Mean heart rate (HR), reflecting the interplay between sympathetic and parasympathetic modulation, showed a significant increase in LT2DM compared to controls and recently diagnosed DM, consistent with earlier observations [13]. This rise in HR may be attributed to decreased parasympathetic function, as noted in previous studies [14], indicating increased sympathetic nerve activity in LT2DM patients.

The ratio of SDNN to RMSSD, considered a marker of autonomic balance, was significantly elevated in the LT2DM group, indicating autonomic imbalance characterized by sympathetic dominance. Notably, prior studies examining LT2DM or recently diagnosed type 2 diabetes mellitus (RT2DM) did not exclude the presence of cardiac autonomic neuropathy (CAN) based on the Ewing battery test [15].

The pathogenesis of CAN remains unclear, although hyperglycemia, hyperinsulinemia, or insulin resistance-induced neuronal damage are areas of focus for researchers [16]. Early-stage CAN or subclinical CAN may initially affect the vagus nerve, leading to sympathetic overactivity manifested by resting tachycardia, exercise intolerance, and abnormal Ewing battery test results. Subsequent stages may involve sympathetic denervation and orthostatic hypotension [17,18].

Overall, reduced HRV in both DM groups, even without clinically evident autonomic neuropathy, indicates early-stage autonomic dysfunction. Greater impairment of the parasympathetic component leads to autonomic imbalance characterized by sympathetic overactivity, particularly evident in LT2DM. Findings in RT2DM suggest the onset of autonomic dysfunction early in diabetes, consistent with previous studies suggesting cardiac autonomic dysfunction at or shortly after diagnosis of type 2 diabetes [19,20]. However, the progression from subclinical to clinical CAN or development of abnormal Ewing battery test results remains uncertain. Subclinical CAN may manifest with reduced HRV detectable through HRV analysis, underscoring the importance of short-term HRV in detecting early autonomic dysfunction.

The study's findings revealed the presence of cardiac autonomic dysfunction in patients with T2DM who did not exhibit clinically detectable cardiac autonomic neuropathy (CAN) based on conventional autonomic tests. This dysfunction was characterized by a predominant impairment in parasympathetic nerve function and increased sympathetic activity. Furthermore, the study indicated that subtle cardiac autonomic dysfunction also occurs in patients with recently diagnosed type 2 diabetes mellitus who do not yet show signs of CAN. Therefore, the use of short-term heart rate variability (HRV) testing was highlighted as an essential technique for detecting the subclinical stages of CAN in individuals with T2DM.

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