SERUM AMYLASE IN PATIENTS OF CHRONIC KIDNEY DISEASE STAGE THREE TO STAGE FIVE Affiliation

Chronic Kidney Disease (CKD) is a progressive loss in renal func on over period of many months or years. As compared to the past decades, the number of kidney diseases leading to end CKD is increasing in Nepal. The disease is associated with the decreased glomerular filtra on rate (GFR). There is decline in nephron func on and number generally quan tated as reduc on in glomerular filtra on rate. As the GFR declines, there is accumula on of metabolic end products excreted by Kidney. Chronic kidney disease is iden fied by blood tests, crea nine and urea are two such substances rou nely measured. Serum amylase is a pancrea c diges ve enzyme that normally acts extracellular to cleave starch into smaller carbohydrate groups and, finally, into monosaccharide's. It is produced by 40-45% from the pancreas and (45%) reabsorbed by tubular cells. Eleva ons in serum total amylase among pa ents with CKD is due to impaired renal clearance and seen mostly when the crea nine clearance is below 50 ml/min. Several studies have been reported on this but there are no studies that have been done so far in Nepalese context. Objec ves This study is designed to correlate serum amylase with CKD stage three to stage five in pa ents of chronic renal disease irrespec ve of hemodialysis and prevalence of risk factors of CKD and different factors that may affect the level of serum amylase in pa ents presen ng to Bir Hospital Nephrology department, Nepal.


INTRODUCTION
Kidneys are paired intra-abdominal organs with func onal unit being nephron.Each nephron is made up of a glomerulus, which comprises four main cell types: endothelial cells; visceral epithelial cells; parietal epithelial 1 cells and mesangial cells.
The ultra filtra on rate (glomerular filtra on rate; GFR) varies with age and sex but is approximately 120-130 2 mL/min per 1.73 m surface area in adults.GFR is defined as the amount of plasma being filtered by the glomeruli per minute.The normal GFR in Indians is approximately 75 to 79 2 2 mL/min/1.73m .
In healthy subjects serum urea and crea nine do not rise above the normal range un l GFR drops by 50-60%.Renal failure is a condi on where kidneys fail to adequately filter waste products from the blood.Two main forms are acute kidney injury, which is o en reversible with adequate treatment, and chronic kidney disease, which is o en not 2 reversible.
Chronic Kidney Disease (CKD) encompasses a spectrum of different pathophysiology processes associated with abnormal kidney func on and a progressive decline in GFR over a period of 3 months.It is es mated by calcula ng crea nine clearance using different formulas, commonest Amylase is a diges ve enzyme that normally acts extracellular to cleave starch into smaller carbohydrate groups and, finally, into monosaccharides.It is produced by 40-45% from the pancreas and 55-60% from the salivary glands.The other loca ons of amylase include fallopian tubes and cyst fluid, testes, lungs, thyroid, tonsils, breast milk, sweat, tears, and some malignant neoplasm.They contribute li le to the serum amylase level.The normal range of Serum amylase is <80 Units/liter (U/L).Amylase has a molecular weight of 50,000 Kilo Dalton and mostly 4 (45%) reabsorbed by tubular cells.
Twenty percent of pancrea c enzymes are excreted by the kidney.Thus pa ents with end stage renal disease have elevated levels of serum pancrea c enzymes.Humans who have had a nephrectomy or have renal insufficiency have average serum amylase levels 50% higher than healthy individuals.Therefore, the kidneys can be assumed to play a major role in amylase metabolism.Mostly serum amylase is raised in acute pancrea s but its rela on to renal failure pa ents is also seen.Serum amylase levels are increased in pa ents with renal insufficiency due to decreased excre on of the enzyme.The levels are not as high as pancrea s and 5 there is no rela on with degree of uremia.
A study done by Bindu C.M. et al...In Bangalore revealed that "Eleva ons in serum total amylase among pa ents with CKD or ESRD is due to impaired renal clearance and seen 5 mostly when the crea nine clearance is below 50 ml/min.
A various studies done in various parts of the world showed varied studies but as such this type of study has not found to be done in our country.The purpose of this study was to correlate serum amylase levels in pa ents of CKD stage three to stage five regardless of hemodialysis in Bir Hospital, Nephrology department.

METHODOLOGY
The study was a cross-sec onal, observa onal, descrip ve, hospital based carried out among 126 pa ents above age 15 who got admi ed and were visi ng Bir Hospital Nephrology Outpa ent department (OPD) irrespec ve of hemodialysis from March 2014 to March 2015, under regular medicine for CKD with clinical, biochemical and sonographic evidence were included in the study a er obtaining informed consent.All the pa ents mee ng the criteria were included in the study both from OPD and inpa ent.Ethical approval was taken from the Bir Hospital Ins tu onal Review Board (IRB) before star ng the research.
Pa ents with increased serum amylase if occurred due to other than renal cause as acute Pancrea s, Mumps, Intes nal Obstruc on, Pep c Ulcer, Cancer, were excluded.The results were analyzed using SPSS (Sta s cal Package for Social Science) version 11 and Microso Excel using correla on coefficient.
A serum or heparinized plasma free from hemolysis was drawn under asep c precau ons from these subjects in vacuum tube via venipuncture.The blood was then allowed to clot, centrifuged and serum was used to perform biochemical analysis on the same day.Plasma samples that had been an coagulated with citrate or oxalate and EDTA were avoided as they chelate calcium and chloride which were required as cofactors in reac on resul ng in false low levels.α -Amylase ac vity in sample was stable for 7 days at 15-25 c.Method used in our lab was CNPG (2-chloro-4nitrophenol-β-1-4 galactopyranosylmaltotrioside) substrate for determina on of α-amylase ac vity.The normal range used at 37 c is up to 80U/l.

RESULTS
126 cases of CKD stage three to five under maintenance hemodialysis or not were included in the present study done in nephrology unit of Bir Hospital, Kathmandu.The age ranged between 15years to 78 years among which 57.14% were males and 42.86% were females with highest percentage belonging to the age group of 36-55 years (table 1)   M. F. Mahomoodally study tends to show that serum α-amylase had a poor correla on with both urea and crea nine in pre-6 dialysis and post-dialysis samples.Amita Yadav et al conducted the study to iden fy whether amylase can be used as an early serum marker of hypertension which showed a sta s cally significant change in the levels of serum amylase with significant (p-value 0.017).Some also proposed that hypertension may increase capillary pressure and acute eleva on in systemic perfusion pressure 7 accelera ng hyperfiltra on, leading to damage to kidney.
Present study showed that serum amylase was elevated in pa ents with CKD V more than 161U/l normal being less than 80 U/l.The mean serum amylase level was 231.28.In CKD III and CKD IV the range was mostly below 80 U/l that is mostly normal.The highest serum amylase mean value present in CKD III and CKD IV were 210 U/l and 243.71U/l respec vely.Study done by JB Keogh et al demonstrates that rise of both salivary and pancrea c amylase in the serum of pa ents with chronic renal insufficiency compared with controls.This increase was approximately two fold that was less than the corresponding rise in serum crea nine that was three fold.It would appear that in terminal disease associated with renal failure a high amylase/crea nine clearance ra o need not indicate 8 pancrea c disease.
Bindu et al study noted, pa ents with advanced CKD had higher level of serum amylase (mean 94.4+86.2SD) in comparison to ESRD pa ents (Mean84.2+22.7SD).It could be due to clearance of amylase during dialysis and malnutri on associated with dialysis.It was found that 60% of pa ents had increased serum amylase ac vity and in 10 pa ents it was more than twice upper limit of normal.In the same study quoted that Bastani et al study compared 22 peritoneal dialysis pa ents with 43 hemodialysis pa ent and 22 non dialyzed chronic renal pa ents, mean total amylase ac vity was similar and above the upper limit of normal in all 3 groups it was abnormal in 75% and above twice upper limit of normal in 24% of all pa ents.
Dardamanis et alshowed that CRF pa ents with or without renal replacement treatment have increased values of total amylase as well as of isoamylases, thus confirming the results of previous studies.A significant number of pa ents had serum amylases more than three mes the upper normal 9 limits without exhibi ng any clinical sign of acute pancrea s.
In this study table 4 depicts that correla on between CKD Stage III was very weak and nega ve correla on with normal range but highly insignificant.The Correla on between CKD IV and Serum amylase level 80-100 U/L was significant at 10 percent but weak of 0.189.It was found that the CKD V had nega ve rela on at same Serum Amylase of 80-120.CKD V was highly significant but nega ve at more than 161 Serum Amylase.
As shown in figure 1, it indicates that correla on of eGFR against Serum Amylase was significant at normal range <80 U/l, 80-120 and 121-140.It was noteworthy to men on that CKD stage in pa ents significantly affect in three Serum Amalyse level.

CONCLUSION
Our inves ga ons done among 126 par cipants indicate that there was a certain amount of reten on of amylase in uraemia due to decreased excre on of amylase by the kidneys.Serum total amylase was found to be elevated in chronic kidney disease pa ents.Serum amylase as a diagnos c tool in recognizing acute pancrea s leads to false posi ve results in chronic kidney disease pa ents.Hence interpreta on of elevated amylase in chronic kidney disease pa ents has to be supported by other laboratory and clinical evidence.As the sample size of our study was small, further studies are required to establish correla on of serum amylase with chronic kidney disease.

RECOMMENDATIONS
Pa ents found to have hyperamylasemia should also be considered for having CKD

LIMITATION OF THE STUDY
Sample Size of this study may not be enough to consider establishing the exact correla on of serum amylase with CKD.
equa on= (140-age×wt./Scr×72) (if Female=0.85)It has been es mated from popula on survey data that at least 6% of the adult popula on in the United States has CKD stages I and II.Nearly 4.5% of the U.S. popula on are 3 es mated to have stages III and Stage IV CKD .

DISCUSSION
The total number of pa ents included in this study was 126 which were equally divided into 42 pa ents in all three groups of CKD Stage three to stage five.Age varied from 15 to 78 years with maximum number of people, falling in the group 46 years to 55 years.There were 57.1% male and 42.8% female under CKD stage three to stage five.

Table 1 :
-Age and Sex distribu on of the pa ents.

Table 1 :
-Age and Sex distribu on of the pa ents.As shown in table 2 serum amylase was in the higher range >161 IU/l among CKD stage V pa ents with mean value of 231.28 and significant p-value.The pa ents with normal serum amylase in CKD V with mean of 57.6 might be due to pa ents undergoing hemodialysis.In CKD stage III serum amylase was maximum at range of 80-100 with mean of

Table 2 :
Serum Amylase in pa ents with CKD Stage Three to Stage Five.

Table 3 :
Correla on of Serum Amylase in pa ents with CKD Stage Three to Five.
Where r is Pearson correla on.