Diabetic nephropathy (DN) represents one of the most serious microvascular complications of diabetes mellitus and is responsible for nearly 44% of cases of kidney failure requiring long-term hemodialysis [1]. The global prevalence of type 2 diabetes mellitus (T2DM) continues to increase at an alarming rate, affecting approximately 382 million people in 2013, with projections suggesting a rise to nearly 592 million by 2035. This growing burden is expected to be accompanied by a substantial increase in diabetes-related complications, particularly diabetic nephropathy [2].

The development of DN is complex and multifactorial, involving a combination of genetic susceptibility and environmental influences. Persistent hyperglycemia leads to significant metabolic, biochemical, and hemodynamic disturbances within renal tissue. Key pathological mechanisms include glomerular hyperfiltration, elevated intraglomerular pressure, oxidative stress mediated by reactive oxygen species, accumulation of advanced glycation end products, and mechanical stress induced by hypertension [3]. In addition, hyperglycemia enhances the synthesis of angiotensin II, which promotes inflammation, fibrosis, and hemodynamic alterations within the kidney.

Inflammatory signaling pathways play a central role in the progression of diabetic kidney disease. Several mediators, including nuclear factor kappa B (NF-κB), pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-1 (IL-1), toll-like receptor-4 (TLR-4), adiponectin, and nuclear hormone receptors, have been implicated in renal injury associated with diabetes. Nuclear receptors are increasingly recognized for their regulatory effects on oxidative stress, inflammation, and fibrotic processes. Among these, the vitamin D receptor (VDR) has gained particular attention due to its involvement in multiple inflammatory and metabolic pathways [4–10].

Despite extensive public health initiatives, vitamin D deficiency and insufficiency remain highly prevalent across populations worldwide [11,12]. Individuals with T2DM are at an increased risk of vitamin D deficiency, and emerging evidence suggests that inadequate vitamin D levels may contribute significantly to the development and progression of chronic kidney disease (CKD), including diabetic kidney disease (DKD). In CKD, renal impairment limits the conversion of vitamin D into its active form, 1,25-dihydroxyvitamin D [1,25(OH)₂D], which requires renal activation. At the same time, the kidney serves as a major target organ for vitamin D, with abundant expression of the vitamin D receptor throughout renal tissue [13–15].

Reduced synthesis of calcitriol and disturbances in calcium, phosphate, and vitamin D metabolism are key contributors to metabolic bone disease in patients with CKD. Additionally, urinary loss of vitamin D and vitamin D-binding protein (VDBP) has been documented in DKD, leading to a progressive decline in circulating vitamin D levels and worsening clinical outcomes. Diabetic kidney disease accounts for nearly half of all cases of end-stage kidney disease requiring renal replacement therapy in developed countries and significantly increases cardiovascular morbidity and mortality, particularly among patients undergoing hemodialysis [16].

Vitamin D is a fat-soluble hormone with physiological roles extending well beyond mineral metabolism. Although vitamin D deficiency was initially linked to rickets, it is now known to influence a wide range of biological processes through both genomic and non-genomic mechanisms, regulating the expression of approximately 2,000 genes. Low vitamin D levels have been associated with increased risks of both type 1 and type 2 diabetes, autoimmune disorders, allergic conditions, adverse pregnancy outcomes, and infections. A meta-analysis published in 2017 reported a significant association between vitamin D deficiency and an increased incidence of respiratory tract infections [17].

The active metabolite of vitamin D, 1,25-dihydroxyvitamin D₃, exerts its effects through the vitamin D receptor, a nuclear transcription factor expressed in multiple renal structures, including the juxtaglomerular apparatus, proximal and distal tubules, collecting ducts, podocytes, and glomerular epithelial cells [18]. Experimental studies have demonstrated that diabetic VDR knockout mice exhibit increased activation of the renin–angiotensin–aldosterone system, resulting in severe renal damage characterized by early albuminuria, glomerulosclerosis, and interstitial fibrosis [19,20].

Clinical and population-based studies further support the association between vitamin D deficiency and diabetic nephropathy. Data from the Third National Health and Nutrition Examination Survey (NHANES III) showed an inverse relationship between serum 25-hydroxyvitamin D [25(OH)D] concentrations and albuminuria [21]. Subsequent analyses confirmed an independent association between low vitamin D levels and diabetic nephropathy among individuals with diabetes [22]. Similarly, the Australian Diabetes, Obesity and Lifestyle (AusDiab) study demonstrated a strong correlation between reduced estimated glomerular filtration rate (eGFR) and vitamin D insufficiency [23]. Collectively, these findings suggest that vitamin D deficiency may play an important role in the onset and progression of diabetic nephropathy and that maintaining adequate vitamin D levels may help slow the progression of chronic kidney disease.

MATERIALS AND METHODS

AIMS AND OBJECTIVES

• To study the 25-hydroxy vitamin D levels in type 2 diabetes mellitus with and without nephropathy.
• To correlate serum vitamin D levels in patients with type 2 diabetes mellitus with and without nephropathy.

Study Design

This was a hospital-based cross-sectional comparative study aimed at evaluating serum 25-hydroxyvitamin D [25(OH)D] levels in patients with type 2 diabetes mellitus (T2DM) with and without diabetic nephropathy (DN).

Study Setting

The study was conducted in the Department of General Medicine at Hassan Institute of Medical Sciences (HIMS), Hassan, Karnataka.

Study Duration

The study was carried out over one year, from July  2024 to June 2025.

Study Population

Patients attending the outpatient and inpatient departments of General Medicine at HIMS, Hassan, who were diagnosed with T2DM, were considered for the study.

Sample Size

The sample size was calculated based on previous studies reporting differences in serum 25(OH)D levels between T2DM patients with and without nephropathy. Using the formula for comparison of two means, with 95% confidence interval (Zα/2 = 1.96), 80% power (Zβ = 0.84), pooled standard deviation (σ) from prior studi, and expected mean difference (d), the minimum required sample size was 54 per group. Accounting for a 10% non-response rate, the final sample size was rounded to 120 participants, divided as follows:

INCLUSION CRITERIA

Inclusion Criteria

• Patients aged 18 years and above
• Diagnosed cases of Type 2 Diabetes Mellitus
• Duration of diabetes 5 years or more
• Patients willing to participate and give informed consent
• Patients divided into two groups:
  • Type 2 diabetes patients with diabetic nephropathy
  • Type 2 diabetes patients without diabetic nephropathy
• Patients not taking vitamin D supplements for the last 3 months
• Patients on regular follow-up for diabetes treatment

EXCLUSION CRITERIA

1. Subjects below 18 years
2. Type I diabetes mellitus
3. Patients with a clinical history of renal disease from causes other than diabetes
4. Hypertension
5. Cancer
6. Autoimmune disorders
7. Recent liver disorder
8. Patients on supplementation of vitamin D and calcium
9. Pregnant women

Data Collection

After obtaining written informed consent, a detailed clinical history was recorded, including duration of diabetes, treatment history, and comorbidities. Physical examination was performed for all participants.

Laboratory Investigations

Venous blood samples were collected under aseptic conditions for the following tests:

• Fasting and postprandial blood glucose
• HbA1c
• Serum creatinine
• Serum 25-hydroxyvitamin D [25(OH)D]

Urine samples were collected for urinary albumin estimation to assess nephropathy.

Diabetic nephropathy was diagnosed based on persistent albuminuria (≥30 mg/g creatinine) and/or eGFR <60 mL/min/1.73 m² on at least two occasions.

Serum 25(OH)D levels were measured using a standardized immunoassay method.

Ethical Considerations

The study protocol was approved by the Institutional Ethics Committee of HIMS, Hassan. Participant confidentiality was strictly maintained throughout the study.

Statistical Analysis

Data were entered in Microsoft Excel and analyzed using SPSS software version 21.

• Continuous variables were expressed as mean ± standard deviation (SD)
• Categorical variables were expressed as percentages
• Comparisons between groups were performed using Student’s t-test for continuous variables and Chi-square test for categorical variables
• A p-value of <0.05 was considered statistically significant

RESULTS

A total of 120 patients with type 2 diabetes mellitus (T2DM) were included in the study, with 60 patients in the diabetic nephropathy (DN) group and 60 patients in the non-diabetic nephropathy (non-DN) group. The mean age of participants was 57.2 ± 9.3 years in the DN group and 55.6 ± 8.7 years in the non-DN group. There was no statistically significant difference in age or gender distribution between the groups (p > 0.05). The duration of diabetes was significantly longer in the DN group compared to the non-DN group (12.4 ± 4.8 years vs 10.1 ± 3.9 years, p = 0.01), suggesting that longer diabetes duration may contribute to nephropathy development .

Table 1: Comparison of Biochemical Parameters Between DN and Non-DN Groups

Table 2: Comparison of eGFR and Vitamin D Levels Within DN Group

Interpretation: Within the DN group, lower eGFR was associated with significantly lower vitamin D levels, suggesting a link between declining renal function and vitamin D deficiency.

Table 3: Correlation Between Serum 25(OH)D and UACR in DN Group

Table 4: Correlation Between Serum 25(OH)D and Serum Creatinine in DN Group

DISCUSSION

In the present study, the age distribution showed that 13% of participants were between 41–50 years, 43% were 51–60 years, 42% were 61–70 years, and 2% were above 70 years. The mean age was 58.82 ± 6.07 years. These findings are consistent with the study by Zomorodian SA et al., where the mean age was 58.75 ± 9.37 years [24]. Similar observations were reported by Balla et al., where the mean age in the non-nephropathy and nephropathy groups was 52.8 ± 6.1 years and 53.2 ± 6.7 years, respectively [25]. Xie S et al. reported a mean age of 55.2 ± 10.3 years [26], while Zhao et al. observed a mean age of 59 years in their study population [27]. Azeem HA et al. also found the majority of participants aged between 50–70 years [28]. These findings suggest that the majority of T2DM patients developing nephropathy fall within the middle-to-late adult age group, reflecting the cumulative effect of long-standing diabetes on renal function.

In the current study, gender distribution was equal, with 50% males and 50% females. This differs from the study by Zomorodian SA et al., which reported 38.5% males and 61.5% females [24], and from Balla et al., who reported female predominance in both nephropathy and non-nephropathy groups [25]. Xie S et al. reported a male predominance (68.9% males) [26], and Zhao et al. also observed more males than females [27]. In contrast, Kumar S et al. found a slightly higher proportion of females (54%) [29]. These variations across studies may reflect demographic differences in study populations or regional gender prevalence in diabetes.

The duration of diabetes in this study ranged from 5–10 years, with a mean of 8.01 ± 1.18 years. In DN patients, the mean duration was longer than in non-nephropathic patients. Zomorodian SA et al. reported a mean diabetes duration of 10.99 ± 8.03 years [24], while Balla et al. reported a mean duration of 13.6 ± 4.69 years in nephropathic patients and 10.6 ± 4.18 years in non-nephropathic patients [25]. Xie S et al. found an average duration of 7.7 ± 6.0 years [26], and Zhao et al. reported 11.27 ± 7.93 years [27]. The results confirm that longer duration of diabetes is a significant risk factor for the development of diabetic nephropathy.

Regarding comorbidities, 40% of participants had hypertension, with 44% in the DN group and 36% in the non-DN group. Zomorodian SA et al. reported 71.2% hypertensive participants [24], while Balla et al. and Xie S et al. reported hypertension in 50–60% of participants [25,26]. Zhao et al. observed higher SBP and DBP in DN patients compared to non-DN patients [27]. These findings suggest that hypertension remains a key contributing factor in the progression of diabetic nephropathy.

Lifestyle factors were also assessed. In this study, 12% were smokers, and 13% consumed alcohol. These results differ from Adisen E et al., who reported 50.3% smokers and 3.3% alcohol users [31], and Wu X et al., who found a combined prevalence of alcohol consumption and smoking that varied by gender [32]. Patra J et al. reported tobacco use and alcohol consumption in 50% and 37% of men, respectively [33]. This suggests regional variations in lifestyle behaviors that may influence diabetic complications.

The glycemic status showed that mean FBS and HbA1c levels were higher in DN patients (FBS: 196.58 ± 66.4 mg/dL; HbA1c: 8.1 ± 1.2%) compared to non-DN patients (FBS: 156.74 ± 16.4 mg/dL; HbA1c: 7.6 ± 1.1%). Balla et al. reported similar findings, with higher FBS in DN patients (206.4 ± 32.9 mg/dL) compared to non-DN patients (182.9 ± 29.13 mg/dL) [25]. Zhao et al. also observed elevated FPG and HbA1c in nephropathic patients [27]. These results indicate that poor glycemic control is strongly associated with nephropathy development.

Renal function parameters showed significantly higher serum creatinine and UACR in DN patients (2.49 ± 1.079 mg/dL and 341.19 ± 143.28 mg/g) compared to non-DN patients (0.6 ± 0.1 mg/dL and normal UACR). Microalbuminuria was observed in 40% and macroalbuminuria in 60% of DN patients. These findings are consistent with Zomorodian SA et al., who reported 33% microalbuminuria [24], Balla et al. [25], Zhao et al. [27], and Chen HM et al., who observed elevated proteinuria and creatinine in DN patients [34].

The eGFR distribution among DN patients revealed 40% with eGFR <60 mL/min/1.73 m², 40% <30 mL/min/1.73 m², and 20% <15 mL/min/1.73 m², while all non-DN patients had normal eGFR. These findings align with Chen HM et al. and Fiesha T et al., demonstrating reduced renal function in DN patients [34,35].

A key finding of this study was the significantly lower vitamin D levels in DN patients (14.91 ± 9.37 ng/mL) compared to non-DN patients (33.86 ± 13.62 ng/mL). In the DN group, 95.1% had vitamin D deficiency. Zomorodian SA et al. also reported lower 25(OH)D in DN patients (18.31 ± 8.12 ng/mL vs. 29.99 ± 13.69 ng/mL) [24]. Similar observations were reported by Xie S et al., Zhao et al., Azeem HA et al., Kumar S et al., and Kafeshani M et al., highlighting the high prevalence of hypovitaminosis D in diabetic nephropathy [26–30,36,37].

Analysis within the DN group revealed that patients with lower eGFR had significantly lower vitamin D levels, suggesting a link between impaired renal function and decreased vitamin D metabolism. This is consistent with studies showing that renal impairment reduces the conversion of vitamin D to its active form [39–41].

Correlation analysis demonstrated a strong negative association between serum 25(OH)D and both UACR (r = -0.91, p < 0.001) and serum creatinine (r = -0.85, p < 0.001). This indicates that lower vitamin D levels are associated with increased albuminuria and reduced renal function. Similar correlations were reported by Zomorodian SA et al., Balla et al., and Zhao et al. [24,25,27].

Overall, the study highlights that vitamin D deficiency is highly prevalent in DN patients and correlates with worsening renal function, higher albuminuria, and longer diabetes duration. The findings suggest that monitoring and correcting vitamin D deficiency may play an important role in preventing or slowing the progression of diabetic nephropathy.

CONCLUSION

The present study demonstrates that patients with type 2 diabetes mellitus and diabetic nephropathy have significantly lower vitamin D levels compared to those without nephropathy. Vitamin D deficiency correlates strongly with reduced eGFR, elevated serum creatinine, and higher albuminuria, indicating its potential role in the pathogenesis and progression of diabetic kidney disease. Early detection and management of vitamin D deficiency may help in slowing the progression of nephropathy and improving renal outcomes in T2DM patients.

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