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Assessment of corneal endothelial cell changes in patients with type 2 diabetes mellitus
*Corresponding author: Mudra Kamlesh Bhatt, Department of Ophthalmology, GMERS Medical College and Hospital, Vadodara, Gujarat, India. mudrabhatt1@gmail.com
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Received: ,
Accepted: ,
How to cite this article: Bhatt MK, Radadia HC, Patel HA, Sapre AA, Patel HN. Assessment of corneal endothelial cell changes in patients with type 2 diabetes mellitus. Global J Cataract Surg Res Ophthalmol. 2025;4:122-7. doi: 10.25259/GJCSRO_1_2025
Abstract
Objectives:
The aim is to assess and quantify the changes in corneal endothelial cells amongst patients with type 2 diabetes mellitus attending a tertiary care hospital and compare them with a non-diabetic control group.
Materials and Methods:
It is a prospective analytical study carried out in the ophthalmology outpatient department, in patients attending a tertiary care hospital in western Gujarat. After taking ethical committee approval and informed consent from the patient, the corneal endothelial cells were measured using a specular microscope. Specular microscopy was done in the control group as well as the diabetic group. The t-test was used to compare the mean values of corneal endothelial cell parameters such as cell density (CD), percentage of hexagonal cells, and coefficient of variation in cell size between two independent groups: Diabetic and non-diabetic patients. The test was performed at a 95% confidence level, with P < 0.05 indicating statistical significance.
Results:
The study was carried out in a total of 276 patients, of which 138 belonged to the diabetic group and 138 belonged to the non-diabetic group. Diabetic patients showed a lower endothelial CD (2300 ± 350 cells/mm2) compared to non-diabetic patients (2700 ± 320 cells/mm2), with a highly significant difference (P < 0.001). Diabetic patients exhibited a higher coefficient of variance (38.5 ± 5.8%) compared to non-diabetic patients (32.7 ± 4.9%), indicating greater variability in cell size and shape in diabetics. This difference was statistically significant (P < 0.001). Diabetic patients had a lower percentage of hexagonal cells (50.2 ± 5.1%) compared to non-diabetic patients (58.6 ± 4.7%), indicating less regular cell morphology in diabetics. This difference was statistically significant (P < 0.001).
Conclusion:
Study revealed that diabetic patients exhibit marked reduction in CD, increased coefficient of variance and decreased percentage of hexagonal cells as compared to non-diabetics.
Keywords
Diabetes mellitus type 2
Endothelial cell density
Hexagonal cells
non-contact specular microscope
INTRODUCTION
Corneal endothelium is a single layer of cells that plays an important role in maintaining the optical transparency of the cornea through Na+/K+ adenosine triphosphatase (ATPase) pump activity. Corneal endothelial cell has the highest density at birth, around 6000 cell/mm2, which decreases by approximately 26% in 1st and further 26% in the next 11 years. Hence, with increasing age, the number of cells are reduced by about 2400–3000 cell/mm2.[1,2] The endothelial cells are hexagonal, 20 μm. The number of endothelial cells decreases with age at a rate of 0.3–0.6%/year. Cornea loses its clarity when the endothelial cell densities (ECD) reach 400–700 cells/mm2 below which corneal oedema occurs.[3] Endothelial cells are best evaluated by non-contact specular microscopy.
Diabetes mellitus is a metabolic disease; chronic hyperglycaemia may lead to micro- and macrovascular disorders. High glucose level leads to an increased activity of aldose reductase, causing sorbitol build-up and sorbitol acts as an osmotic agent and leading to swelling of the endothelial cell. Furthermore, diabetes mellitus reduces Na+/K+ ATPase activity of the corneal endothelium.[4,5] Polymegathism and pleomorphism of corneal endothelial cells seem to be positively associated with type 2 diabetes mellitus (T2DM), which indicates cellular stress and may contribute to compromised corneal function (Shenoy et al. 2009).[6] Furthermore, the ECD is often reduced in diabetics, reflecting a gradual loss of cells due to chronic metabolic stress.
The purpose of this study is to assess changes in corneal endothelial cell morphology and cell density (CD) in diabetic patients and non-diabetic patients aged>40 years.
Normal values amongst the healthy population above the age of 40 years are as follows:[3]
CD cells/mm2 = 2623 ± 187
Coefficient of variation (CV)% = 29 ± 3
Percentage of hexagonal cells = 69 ± 5.
The corneal endothelium, in particular, is vital for maintaining corneal clarity through its pump and barrier functions, which prevent corneal swelling and opacity. In T2DM patients, significant changes occur in endothelial cells including alterations in CD, size, shape and function which potentially impairs vision.
Aim
The aim of the study is to assess and quantify the changes in corneal endothelial cells amongst patients with T2DM attending a tertiary care hospital and to compare these findings with a non-diabetic control group to identify specific patterns or alterations associated with T2DM.
Objectives
The objectives of the study are (1) to evaluate the corneal ECD in T2DM patients: Measure and compare the ECD in patients with T2DM and a healthy control group to identify any significant differences indicating endothelial cell loss or deterioration in the diabetic cohort. (2) To Assess Corneal Endothelial Cell Morphology: Examine changes in cell morphology, including cell size (polymegathism) and shape (pleomorphism) variations in T2DM patients compared to controls, using specular microscopy or other appropriate imaging techniques. (3) To investigate the correlation between Duration and Control of Diabetes with Corneal Endothelial Changes: Analyse the relationship between the duration and control of T2DM and the extent of endothelial cell changes, to determine if longer disease duration or poorer glycaemic control is associated with more significant corneal alterations.
MATERIALS AND METHODS
Study setting
The present study is a hospital-based study done in patients visiting the ophthalmology outpatient department (OPD) at a tertiary care centre.
Study design
This is a prospective analytical study.
Sample size
At 95% confidence level and 80% of power, assuming that the proportion of control exposure was 0.01 and the hypothetical proportion of case exposure was 0.08, with 8.60 was assumed to be an odd ratio that required a sample size of 138 per group. The total sample size required for the study is 276, as per calculations by StatCalc (www.openEpi.com) using Kelsey’s formula.
Inclusion criteria
Age: Patients aged 40 years and above. This age group is selected to focus on the adult population commonly affected by T2DM and its related complications.
The patient has diabetes.
Diagnosis of T2DM: Patients must have a confirmed diagnosis of T2DM, as verified by medical records or a diagnosis by a healthcare provider.
Duration of diabetes: Patients with a known history of diabetes mellitus type 2 for more than 2 years. This criterion is set to assess the long-term effects of diabetes on the corneal endothelial cells.
Informed consent: Patients who are willing and able to provide informed consent to participate in the study.
Exclusion criteria
Denied consent
Age <40 years
Type 1 diabetic patients
Diabetic patients on insulin
Blood glucose level <200 mg/dL
History of ocular surgery – Patients who have undergone ocular surgery such as cataract extraction, corneal transplantations, refractive surgery, which can alter corneal endothelial cell parameters
Other ocular infection/pathology – Patients with other ocular diseases (e.g., glaucoma, severe dry eye, and retinal diseases) that could affect the corneal endothelial cell counts
Patients having other systemic illnesses such as hypertension, cardiovascular disease, tuberculosis, and autoimmune conditions.
Ethical considerations
The study protocol received approval from the institutional ethics committee (IEC) before commencement. The IEC thoroughly reviewed our research proposal, ensuring that it complied with ethical guidelines and safeguarded the rights and well-being of participants.
We obtained informed consent from all participants involved in the study. This process entailed providing comprehensive information about the study’s objectives, procedures, potential risks and benefits. Participants were informed that their involvement was voluntary and that they had the right to withdraw from the study at any point without any repercussions. We ensured that participants fully understood the information provided and had the opportunity to ask questions before signing the consent form.
To protect the confidentiality of participants, we implemented stringent data protection measures. Personal identifiers were removed from data records to maintain anonymity.
Procedure
OPD patients with satisfied the inclusion criteria were included in the study
Written informed consent was obtained from the patients
Complete ocular examination including visual acuity, best-corrected visual acuity, slit-lamp examination, fundus examination and applanation tonometry were done in each case before hand to rule out any other ocular disease or condition
The specular microscopy procedure was meticulously followed to ensure accurate assessment of corneal endothelial cell morphology. The equipment used was a non-contact specular microscope.
The specular microscopy used is Topcon SP-1p. It is a noninvasive photographic technique that enables the detailed visualisation and quantitative analysis of the cornea. This technology operates on the principle of specular reflection, where light directed onto the cornea is reflected back from the interface between the corneal endothelium and aqueous humour. The reflected light is then captured to produce high contrast image of the endothelial cell, allowing for the assessment of CD, morphology and distribution.[7]
The patients were informed about the procedure and their consent was obtained. They were then seated comfortably with their chin and forehead resting on the device’s support. The procedure began with the alignment of the microscope. It was adjusted to focus on the corneal endothelium, ensuring that the central corneal zone was properly imaged. To minimise artefacts and obtain the best quality images, patients were instructed to keep their eyes open and to avoid blinking during the image capture process. The captured images were then analysed for parameters such as CD, pleomorphism and polymegathism. We have included findings of the right eye of 138 cases and 138 controls.
Statistical analysis
To compare corneal endothelial cell parameters between diabetic and non-diabetic patients, we employed several comparative analysis techniques, including the t-test and analysis of variance (ANOVA).
The t-test was utilised to compare the mean values of corneal endothelial cell parameters, such as CD, CV in cell size and percentage of hexagonal cells, between two independent groups: Diabetic and non-diabetic patients. We performed the test at a 95% confidence level, with a P < 0.05 indicating statistical significance.
For comparisons involving more than two groups, ANOVA was used. This was particularly relevant when comparing subgroups within the diabetic population, categorised by the severity or duration of diabetes. The null hypothesis for ANOVA posited that all group means were equal. A significant F-statistic (P < 0.05) indicated that at least one group mean was different, prompting post hoc tests to identify the specific groups with significant differences.
The data analysis was carried out using two primary software tools: the Statistical Package for the Social Sciences and R.
RESULTS
Table 1 summarises demographic characteristics of diabetic and non-diabetic patients (n = 138 each). Diabetic patients were older (59.2 ± 10.3 years) compared to non-diabetic patients (56.7 ± 9.8 years), though the difference was not statistically significant (P = 0.13). Gender distribution showed similar proportions between groups. Significant differences were observed in blood glucose levels.
| Characteristic | Diabetic patients (n=138) | Non-diabetic patients (n=138) | P-value |
|---|---|---|---|
| Age (years) | 59.2±10.3 | 56.7±9.8 | 0.13 |
| Gender | - | - | 0.45 |
| Male | 78 (56.5%) | 80 (58.0%) | - |
| Female | 60 (43.5%) | 58 (42.0%) | - |
| Duration of diabetes (years) | 12.3+6.4 | N/A | N/A |
| Blood glucose levels | - | - | - |
| Fasting (mg/dL) | 158.4±20.1 | 92.5±10.4 | <0.0001 |
| Post-prandial (mg/dL) | 226.7±30.2 | 117.3±15.5 | <0.0001 |
Note: For gender chi-square test is used and for age and sugar levels t-test is used. P < 0.05 is significant. Coefficient of variance, cell density and hexagonality is expressed as Mean+-Standard deviation. N/A: Not applicable
Table 2 summarises that diabetic patients showed a lower CD (2300 ± 350 cells/mm2) compared to non-diabetic patients (2700 ± 320 cells/mm2), with a highly significant P-value (P < 0.0001).
| Characteristic | Diabetic (n=138) | Non- diabetics (n=138) | P-value |
|---|---|---|---|
| Cell density (cells/mm2) | 2300±350 | 2700±320 | <0.001 |
| Coefficient of variation (%) | 38.5±5.8 | 32.7±4.9 | <0.001 |
| Hexagonal cells (%) | 50.2±5.1 | 58.6±4.7 | <0.001 |
Note: t-test was used for comparison of endothelial cell parameters. P<0.05 is significant, Coefficient of variance, cell density and hexagonality is expressed as Mean+-Standard deviation
Diabetic patients exhibited a higher CV (38.5 ± 5.8%) compared to non-diabetic patients (32.7 ± 4.9%), indicating greater variability in cell size and shape in diabetics. This difference was statistically significant (P < 0.0001), highlighting potential structural alterations in corneal endothelial cells associated with diabetes.
Diabetic patients had a lower percentage of hexagonal cells (50.2 ± 5.1%) compared to non-diabetic patients (58.6 ± 4.7%), indicating less regular cell morphology in diabetics. This difference was statistically significant (P < 0.0001).
For diabetic patients, CD was compared for three groups: Duration <5 years (2328 ± 296 cells/mm2), 5–10 years (2294 ± 312 cells/mm2) and >10 years (2160 ± 336 cells/mm2). All differences were highly significant (P = 0.04), indicating a consistent decline in CD associated with longer duration of diabetes [Figure 1].
- Endothelial cell density (Y axis) by duration of diabetes. (X axis), Endothelial cell density is expressed as Mean+-Standard deviation.
Diabetic patients showed higher coefficient of variance values with increased duration of diabetes: Duration <5 years (36.2 ± 5.0%), 5–10 years (38.0 ± 5.3%) and >10 years (40.3 ± 5.8%). These differences were statistically significant (P = 0.004), indicating greater variability in cell size and shape with longer duration of diabetes [Figure 2].
- Coefficient of variance (Y axis) by duration of diabetes. (X axis), Coefficient of variance is expressed as Mean+-Standard deviation.
With increased duration of diabetes, we reported a decline in hexagonal cells: Diabetes duration <5 years (53.4 ± 5.2%), 5–10 years (50.1 ± 5.0%) and >10 years (47.2 ± 5.5%). These differences were statistically significant (P < 0.001), indicating reduced cell regularity with longer diabetes duration [Figure 3]. Specular biomicroscopy image shown in Figure 4, illustrates reduced cell density, increased pleomorphism and reduced hexagonality in diabetic patients.
- Hexagonality (Y axis) by duration of diabetes. (X axis), Hexagonality is expressed as Mean+-Standard deviation.
- Comparison of corneal endothelial cell parameters of (a) diabetic and (b) non-diabetic.
DISCUSSION
The study included 138 diabetic and 138 non-diabetic patients, with comparable age and gender distributions, ensuring a balanced comparison. Diabetic patients had significantly higher fasting and postprandial blood glucose (PP2BS) levels. These baseline differences are crucial as they provide a comprehensive context for interpreting corneal endothelial changes.
Glycaemic control, as measured by fasting blood glucose levels and PP2BS levels, had a significant impact on corneal endothelial cell parameters. These findings emphasise the importance of maintaining optimal glycaemic control to preserve corneal endothelial structure and function. The observed corneal endothelial changes in diabetic patients can be attributed to several factors, including chronic hyperglycaemia, oxidative stress and microvascular damage.
Hyperglycaemia can lead to the accumulation of advanced glycation end-products (AGEs), which can disrupt cellular function and induce apoptosis in endothelial cells. There is excessive accumulation of sorbitol in the endothelium, leading to osmotic cell damage. With corneal endothelium cells decreasing, the remaining neighbouring endothelial cells would expand and migrate. Meanwhile, diabetes inhibits the activity of Na+/K+-ATPase of the corneal endothelium, leading to disruption of its structure.[8] The reduction in corneal endothelial cell adhesions occurs due to direct adhesive protein modifications of Descemet’s membrane by AGEs. In addition, oxidative stress resulting from chronic inflammation and metabolic dysregulation in diabetes can further exacerbate endothelial cell damage.
Regular screening for corneal endothelial changes in diabetic patients can facilitate early detection of ocular complications and prompt intervention to prevent vision loss. Emphasising glycaemic control, managing intraocular pressure and monitoring for diabetic retinopathy (DR) are critical strategies in preserving corneal endothelial health in diabetic individuals.
Diabetic patients exhibited a significantly lower CD (2300 ± 350 cells/mm2) compared to non-diabetic patients (2700 ± 320 cells/mm2, P < 0.0001). This result is similar with Schultz et al. 1984, observed a reduction in endothelial cell density in diabetic patients, suggesting that prolonged exposure to high blood glucose levels could cause endothelial cell dysfunction and death.[9] This result is consistent with the findings of Lee et al. in 2006,[10] who reported that CD in diabetics is less (2577.2±27.3 cell/mm2) than in control group (2699.9 ± 38.7 cell/mm2), decrease in endothelial CD in diabetic patients, attributing it to chronic hyperglycaemia leading to endothelial cell loss due to metabolic stress and apoptosis. This observation is supported by the work of Inoue et al. 2002 who observed that CD in diabetic patients (2493 ± 330) is less than control group (2599 ± 278).[11]
The CV of corneal endothelial cells was higher in diabetic patients (38.5 ± 5.8%) compared to non-diabetic patients (32.7 ± 4.9%, P < 0.0001), indicating greater variability in cell shape and size. This result is consistent with the findings of Lee et al. in 2006[10], and the CV in cell area was significantly higher in the diabetic group (38.2 ± 0.4) than in the control group (35.2 ± 0.6). This observation is supported by the work of Inoue et al. (2002). CV in cell area was significantly higher in the diabetic group (37.2 ± 6.0) than in the control group (35.4 ± 5.0) (P = 023, t-test).[11] Moreover, Sudhir et al. (2012),56 reported similar findings, linking the increased CV to oxidative stress and glycation end-products that compromise cell integrity and function.[12]
The percentage of hexagonal cells, a marker of endothelial cell health, was lower in diabetic patients (50.2 ± 5.1%) compared to non-diabetic patients (58.6 ± 4.7%, P < 0.0001). This finding aligns with studies by Lee et al. in 2006[10], reductions in hexagonal cells in diabetic patients (52.7 ± 0.6) than in the control group (60.6 ± 0.9). These results are in line with the findings of Saini et al. (1996)[13] who reported that longer duration of diabetes exacerbates endothelial cell loss and morphological changes. McCarey et al. (2008),[14] which observed a reduction in hexagonal cells in diabetic patients, indicating a loss of regular cell morphology due to metabolic dysregulation and chronic hyperglycaemia. Inoue et al. in 2002 found that hexagonality is lower in diabetic as compared to non-diabetic.[11] The decrease in hexagonal cells in our study, corroborated by these studies, suggests that diabetes affects the structural integrity of corneal endothelial cells, leading to reduced cellular health.
Our study further highlighted that the duration of diabetes correlated with greater reductions in CD, increased CV and decreased hexagonal cells.
A study shows that maintaining optimal blood sugar levels through proper diabetes management can significantly slow down or even prevent further damage to the corneal endothelium. Corneal endothelial damage cannot be reversed as the endothelial cells have limited regenerative capacity.[15,16]
Cataract surgery is a well-known risk factor for endothelial cell loss. Studies show that even healthy patients experience a decrease in endothelial cell count and an increase in polymegathism and pleomorphism in the first 3 months postoperatively. Cataract surgery in diabetic patients results in greater endothelial cell loss as well as a decrease in hexagonal cell count. Low endothelial cell counts significantly increase the risk for corneal oedema and low vision post-surgery.[17]
Limitations
Cross-sectional nature
As a prospective analytical (cross-sectional) study, it captures data at one point in time. This restricts to finding out the relationships between diabetes duration, glycaemic control and endothelial cell changes.
Glycaemic control assessment
Long-term glycaemic control markers such as HbA1c were not included, which could have offered a better correlation with endothelial changes.
Non-inclusion of systemic comorbidities
Patients with systemic illnesses such as hypertension and cardiovascular disease were excluded. While this reduces confounding, it also limits the understanding of real-world diabetic populations, where such comorbidities are common.
Lack of longitudinal follow-up
The study does not evaluate long-term progression or changes in corneal endothelial parameters over time, which could provide more insight into the cumulative impact of diabetes on the cornea.
CONCLUSION
Our findings reveal that diabetic patients exhibit marked alterations in corneal endothelial cell parameters, including reduced CD, increased CV and decreased percentage of hexagonal cells, compared to non-diabetic individuals. These changes are indicative of compromised corneal endothelial cell structure and function due to chronic hyperglycaemia. The reduction in corneal endothelial CD observed in diabetic patients aligns with previous studies, suggesting that prolonged exposure to high blood glucose levels leads to endothelial cell loss and increased cell variability. The higher CV and lower hexagonal cells in diabetic patients further underscore the detrimental effects of diabetes on corneal endothelial cell morphology. These structural alterations can be attributed to oxidative stress and the accumulation of AGEs, which disrupt cellular integrity and function. Our study also highlights the impact of diabetes duration on corneal endothelial health. Longer durations of diabetes are associated with greater reductions in CD, increased CV and decreased hexagonal cells, indicating cumulative damage over time.
Ethical approval:
The research/study was approved by the Institutional Review Board at GMERS Medical College and Hospital, Gotri, Vadodara, number IHEC/23/OUT/SRPG042, dated 02 January, 2023.
Declaration of patient consent:
The authors certify that they have obtained all appropriate patient consent.
Conflicts of interest:
There are no conflicts of interest.
Use of artificial intelligence (AI)-assisted technology for manuscript preparation:
The authors confirm that there was no use of artificial intelligence (AI)-assisted technology for assisting in the writing or editing of the manuscript and no images were manipulated using AI.
Financial support and sponsorship: Nil.
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