European Journal of Cardiovascular Medicine

Diabetes mellitus is common endocrine disorder with metabolic abnormalities in this era. Many microvascular and macrovascular complications are seen in long term. In India its prevalence is on the rise.  Diabetic nephropathy occurs in as many as 30% of insulin dependent diabetes mellitus patients and 25% of non-insulin dependent diabetes mellitus patients (1). In Diabetic nephropathy there is progressive and continuous deterioration in glomerular function that results in end stage renal disease. Diabetic is now most common cause of end stage renal disease (2). In early stage of diabetic nephropathy there is microalbuminuria (30-299 mg/24 hr urinary excretion of albumin) called incipient diabetic nephropathy. (2).  That can be detectable only by use of sensitive assay for urinary albumin (3,4,5). At this stage urine is negative for macro albumin and renal function is normal. Development of overt nephropathy take 10-15 years from incipient nephropathy (3,4). In this stage progression to end stage renal disease can be prevented. Microalbuminuria can be prevented by strict control of blood sugar level, insulin treatment, dietary protein restriction, and control of hypertension by ACE inhibitors and Beta-blocker.  Microalbuminuria is a warning sign, if proper precaution and therapeutic intervention not taken can lead to irreversible renal damage. Association of Microalbuminuria with other microvascular complications e.g. retinopathy, neuropathy, and ischemic heart disease.  Microalbuminuria can be used as a marker for widespread microvascular damage in a patient of diabetes mellitus. (6,7,8,9)

The aim of this study was to study the occurrence of microalbuminuria in patients with diabetes mellitus and also to find out it its association with the duration of diabetes mellitus and the microvascular complications and macrovascular complications of diabetes mellitus.

AIMS AND OBJECTIVES

1. Screening of microalbuminuria in patients with diabetes mellitus.
2. To correlate microalbuminuria with the duration of diabetes mellitus and association with other microvascular and macrovascular complications of diabetes mellitus.

Study design: Observational cross-sectional study.

Study setting: The study was conducted at tertiary care centre on patients who attend the medicine OPD and hospitalised in the medicine ward. Total 82 cases were included in the study by random sample selection method. Patients were selected as per their history of diabetes and considered diabetic based on WHO ⁽²⁾ criteria for diagnosis of diabetes mellitus.

Exclusion criteria:

1. Patients with macroalbuminuria
2. Patients with congestive cardiac failure, urinary tract infection. 
3. Ketonuria
4. Pregnant patients 
5. Patients with overt diabetic nephropathy.

STUDY PROTOCOL: In our study total 82 cases were enrolled after fulfilling the inclusion criteria, 82 cases were subjected to detailed history, physical examination, characteristics age, sex, age of onset and duration of diabetes. In All patients details regarding the presenting complaints were noted. Total duration of diabetes, along with treatment were noted. The family history regarding diabetes was taken. A complete clinical examination was carried out in all patient along with diabetic related complications like retinopathy, neuropathy, diabetic foot and ischemic heart disease are noted.

Hypertension was said to be present when there was a history of hypertension or the systolic blood pressure was recorded greater than 160mm of hg and/or diastolic pressure greater than 90 mm of hg on 3 consecutive occasions. Ischemic heart disease was recorded to be present in the presence of suggestive history of angina or myocardial infarction with electrocardiographic evidence. Peripheral neuropathy was judged to be present if there was historical evidence of neuropathic pain, numbness or tingling sensation in the extremities and or absence of ankle jerks along with diminished vibratory threshold or pin prick sensation in hands or feet on examination. Fundus examination was done in all patients for evidence of diabetic retinopathy. Retinopathy was said to be present when there was evidence of microaneurysm, soft or hard exudates and haemorrhages. Neovascularity was considered as evidence for proliferative retinopathy. Peripheral vascular disease was considered to be present with history of amputations and /or absent of one or more peripheral pulses and /or presence of gangrenous foot. (6)

In all patients the following investigations were done. 

• Microalbuminuria was estimated by Micral test in all the cases. 
• Fasting Blood sugar and Postprandial blood sugar
• Glycosylated haemoglobin
• Blood urea and serum creatinine
• Fasting lipid profile
• Urine routine and culture
• Electrocardiogram
• Ultrasonography of the abdomen,
• echocardiogram and
• chest x-raywas done in selected cases only. 

Estimation of Microalbuminuria by Micral test: (10)

Micral test, an immunological rapid dip stick semi qualitative technique for detection of microalbuminuria, was used for estimation of microalbuminuria. 

Micral test components: 1 test strip contains monoclonal antibodies against human albumin (immunoglobulin G) labelled with colloid gold 2.2mg, fixed albumin 7.7 mg.

TEST PRINCIPLE:

There is a serial arrangement of several reagent pads, which are in fluid communication by a reaction controlling chroma. This step combines one step handling with a complex chemistry. 

The single reaction steps are as follows: 

• Urine of the sample is transported through the wick fleece to the buffer fleece, where acidic urine is adjusted to proper 61 
• Upon entering the conjugatefleece, the antigen – antibody reaction takes place Albumin of the sample is specifically bound to a soluble conjugate of antibodies and marker enzyme resulting in an antigen – conjugate complex
• The excess antibodies are bound to immobilized albumin on the capture matrix and removed from the sample in this way. 
• Only the complex of conjugate with sample-albumin reaches the substrate pad. Here thecolour reaction takes place, the marker enzyme B-Galactosidase cleaves off the purple dye chlorophenol red from the yellow substrate (chlorophenol red galactoside) in a kinetic reaction. The intensity of the colour produced is proportional to the albumin concentration in the urine. 

SPECIMEN COLLECTION: 

All patients were afebrile and were kept at rest during the collection of urine. Urine of the patient was first tested for albumin by albustic method. Patients who were negative for albumin by the albustic method were only included in this study. First morning mid-stream urine sample that was collected in a sterile container was used for determining microalbuminuria. Test strip was immersed in urine such that fluid level was between the two black bars provided on the strips. Strip was withdrawn after 5 seconds. Strip was placed horizontally across the urine vessel and colour change in the test zone was compared with colour scale after one minute. Sensitivity of the kit is 0.4ng/ml and measuring range is 0.8 to 10ng/ml. 

Microalbuminuria was graded as follows: (11)

Mild (20-50mg/L)- + 

Moderate (50-100mg/L) - ++ 

Severe (100-300mg/L)- +++ 

Depending on the colour change in the strip. 

Severity of diabetes was graded based on the HbA1c levels, as follows: 

Mild - < 7%

Moderate- 7-7.5%

Severe- > 7.5%

Blood urea, serum creatinine and lipid profile were estimated in all cases. 

STATISTICAL ANALYSIS:

Data was analysed using SPSS 22 version software. Categorical data was represented in the form of Frequencies and proportions. Chi-square test was used to find the significance of proportion of incidence of microalbuminuria between various levels of study and used as test of significance for qualitative data. Continuous data was represented as mean and standard deviation (12)

Statistical software: MS Excel, SPSS version 22 (IBM SPSS Statistics, Somers NY, USA) was used to analyse data. (13)

This study was conducted at tertiary care hospital in Medicine Out-Patient or In-Patient Department among 82 patients. Male preponderance of 48 (58.5%) as compared to 34 (41.5%) females. Mean age was 54.03 ± 9.6 years. The mean duration of diabetes since diagnosis was 7.271+_ 4.71 years among the male patients and 7.309 +_4.91 years among the female patients. 48 patients were negative for microalbuminuria., 34 patients have Microalbuminuria, Incidence of Microalbuminuria is 3.08 times more likely for the age group > 50 years of age as compared to the age group. Duration of Diabetes is an important factor for development of Diabetic nephropathy. The number patients with the duration of diabetes since detection less than 5 years were 45 and among them 2 were positive for microalbuminuria. The number patients with the duration of diabetes since detection 5 years and 10 years were 18 and among them 15 were positive for microalbuminuria. The number patients with the duration of diabetes since detection 10 years and 15 years were 12 and among them 10 were positive for microalbuminuria. All the patients i.e. 7 with duration more than 15 years were positive for Microalbuminuria   Control of Diabetes also determine development of nephropathy in study 8 patients had HbA1c values less than 6.5% and among them 2 were positive for microalbuminuria. 14 patients had HbA1c values between 6.5% and 7.0%, among them 3 were positive for microalbuminuria. 16 patients had HbA1c values between 7.0% and 7.5%, among them 7 were positive for microalbuminuria. 44 patients had a HbA1c values more than 7.5%, among them 22 were positive for microalbuminuria albuminuria. Association of other complications of Diabtes with Microalbuminuria also determine in study which shows 13 patients had retinopathy; among them 8 patients were positive for microalbuminuria. 39 patients had peripheral neuropathy; among them 21 patients were positive for microalbuminuria. 6 patients had peripheral vascular disease; among them 4 patients were positive for microalbuminuria. 27 patients had ischemic heart disease (IHD), among them 16 patients were positive for microalbuminuria. 36 patients had hypertension; among them 31 patients were positive for microalbuminuria.

Table 1: - No of patients with microalbuminuria

Table no-2 Showing various parameters.

Table -3: Duration of DM with microalbuminuria

Table-4:  Association of Complications with the incidence of Microalbuminuria

Diabetes mellitus is a metabolic disorder in which on long term generalised endothelium dysfunction occur. The severity of the dysfunction depends on the age of the patient and duration of the diabetes. Generalized endothelial dysfunction manifested clinically in several forms). Microalbuminuria marks the onset of endothelial dysfunction related to the kidney (14,15).  Microalbuminuria is a warning for imminent nephropathy. In diabetic patients with microalbuminuria not only have ongoing progressive nephropathy but are also likely to have retinopathy, nephropathy and cardiovascular problems including coronary artery disease and hypertension (16). In this observational cross-sectional study 82 patients attending medicine OPD and/or hospitalized in medicine wards of tertiary care patients with diabetes mellitus were studied. In study there is mean age was 54.03 ± 9.6 years and male preponderance of 48 (58.5%) as compared to 34 (41.5%) of females with male: female ratio of 1.4:1. In a similar study by Aneesah A. AlFehaid et all the mean age of the patients was 52.01 years, Male patients constituted 50.2% while females were 49.8% (17). In this study among randomly selected patients for microalbuminuria is 41.46%. Among various other studies the prevalence of microalbuminuria ranges from 25% to 35% (17,18,19,20)

An increase in the percentage of microalbuminuria in study can be due large number of elderly patients more than 50 years, and rural population with poor diabetic control. In study microalbuminuria occurs more commonly in diabetic subjects who are more than 50 years of age. In study microalbuminuria tended to be 2.54 times more common in the age group of above 50 years as compared to the age group of less than 50 years. In a similar study by A Varghese, R Deepa, M Rema, V Mohan et all shows similar pattern. (21) Poor values of HbA1c are known to be associated with increasing incidence of microalbuminuria. In study 22 patients who had a normal HbA1c (< 7.0%) in whom 5 patients had microalbuminuria, whereas with HbA1c values more than 7, 29 out of 60 (nearly 50%) had microalbuminuria. It is seen from the above result that even small increments of HbA1c more than 7.0% result in almost doubling of the incidence of microalbuminuria. In this cross-sectional study, these findings raise concern regarding the association between poor glycaemic control and microalbuminuria in a rural setting. In a similar study by Muhammad Bilal Habib and Noreen Sher Akbar shown same association of hb1ac and microalbuminuria (22) The incidence of microalbuminuria is significantly associated with the presence of retinopathy, peripheral neuropathy, ischemic heart disease, hypertension. Peripheral neuropathy and hypertension have the most significant association with microalbuminuria. In our study 13 patients had retinopathy, among them 8 patients were positive for microalbuminuria. 39 patients had peripheral neuropathy; among them 21 patients were positive for microalbuminuria. 6 patients had peripheral vascular disease; among them 4 patients were positive for microalbuminuria. 27 patients had ischemic heart disease (IHD), among them 16 patients were positive for microalbuminuria. 36 patients had hypertension; among them 31 patients were positive for microalbuminuria. In a similar study by Aneesah A. AlFehaid et all(23)⁾, Bhoomika P Dadhania*, Amit H Aravat, Gauravi A Dhruva et all. (24)

We studied 82 diabetes patients for detection of microalbuminuria through the dipstick method.

The incidence of microalbuminuria is estimated to be 41.46 % in this study.

Microalbuminuria shows a direct relationship with increasing age of patients and increasing duration of diabetes mellitus since diagnosis.

A HbA1c value above 7% is associated with 50% or higher incidence of microalbuminuria. Microalbuminuria is associated with poor glycaemic controls.

Incidence of microalbuminuria is significantly associated with presence of hypertension, neuropathy, ischemic heart disease, retinopathy and high body mass index.

Recommendations

Due to increasing modernization, we come across lots of patients with diabetes but these patients have lack of awareness regarding complication, that lead to life threatening condition of patients. So, any patients visited to us with diabetic we should do a detail examination and counselling of these patients regarding probable complication at time of visit and ask to follow them regularly and screen for microalbuminuria in early stage so that we can treat it or herald complication. In patients who have microalbuminuria a detailed cardiovascular examination is necessary in early stage. Hypertension must be treated. Left ventricular hypertrophy and function should be assessed byechocardiographic study at the stage of microalbuminuria and thereafter every 6 –12 months. Effective antihypertensive therapy can reverse left ventricular hypertrophy. If further evaluation needed then do electrocardiography, stress testing, coronary angiography and Holter monitoring. In Diabetes patients are prone to silent MI. so any ischemic heart disease should be treated aggressively.

Peripheral vascular disease must be assessed and treated as necessary. Doppler flow studies and arteriography may be useful to assess the severity of the disease. 

Testing vibration perception threshold and thermal discrimination may identify the risk of neuropathic ulceration. The test should be repeated regularly as sensation may become impaired later, during the course of nephropathy. Autonomic dysfunction is very common in nephropathicpatients. The important manifestations are postural hypotension and incomplete bladder emptying which predisposes to urinary tract infection. 

Microalbuminuria is frequently associated with retinopathy. In diabetes patients on screening fundoscopy should be done at time of diagnosis and should be repeat after a particular interval. Early and regular ophthalmic review and prompt treatment is necessary to prevent blindness.

Area of conflict- None

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24. STUDY OF MICROALBUMINURIA IN DIABETES TYPE 2 PATIENTS AS A MARKER OF MORBIDITY (A STUDY OF 100 CASES IN RAJKOT CITY) Bhoomika P Dadhania*, Amit H Aravat, Gauravi A Dhruva *Department of Pathology, P. D. U. Medical College, Rajkot, Gujarat, India 763.