chronic kidney disease ( ckd ) is associated with a number of comorbidities and prognosis is poor , because many patients experience progression to end - stage renal disease . also , in the majority of patients disease progression may be altered due to more or less suitable immunosuppressive protocol treatments and therapeutic approaches . the mechanisms of injury underlying progression are blurred , but traditional opinion pointed to an association between decline in renal function with the degree of proteinuria and histological findings of glomerulosclerosis and interstitial fibrosis [ 2 , 3 ] . kidney injury molecule-1 ( kim-1 ) is a new specific biomarker of proximal tubule injury that can be measured both in urine and kidney tissue samples . it is an apoptotic - cell phagocytosis and scavenger receptor that is most highly upregulated in proximal tubular epithelium in acute and chronic kidney injury . also , much attention has been paid to its possible pathophysiological role in modulation of tubular damage and repair [ 57 ] . besides a close relationship between tissue kim-1 expression and urinary kim-1 concentration , our previous six - month prospective pilot study showed that kim-1 expression in tissue correlated better with tin features and renal function in different chronic kidney diseases than with urinary kim-1 concentration . therefore , that investigation was the basis of this long - term retrospective study , where only kim-1 tissue expression as a predictor of long - term kidney function was examined . the aim of this retrospective investigation was to evaluate ( 1 ) possible associations between tissue kim-1 expression and tubulointerstitial ( tin ) inflammation , atrophy , and fibrosis in different chronic kidney diseases and ( 2 ) possible associations between tissue kim-1 expression and some demographic and laboratory parameters as well as kidney function and proteinuria at the time of biopsy and 6 , 12 , 24 , and 36 months later . the retrospective study included 60 patients ( 27 men ) of mean age 34.42  12.15 years ( range 1859 years ) , who were hospitalized in the clinical center of serbia for kidney biopsy from 2006 to 2009 . indications for kidney biopsy were nephrotic syndrome , pathologic proteinuria without nephrotic syndrome , and abnormal urinary sediment ( erythrocyturia and leukocyturia ) in several samples . pathohistological analysis revealed minimal change glomerulonephritis ( mcgn ) in two patients , non - iga gn in nine , iga gn in six , membranous and membranoproliferative gn in seven patients each , focal glomerulosclerosis in eleven , lupus nephritis in ten , and crescentic gn in eight patients . after diagnosis , the participants were treated according to established protocols for each type of gn with immunological and nonimmunological therapy ( nonsalt diets , statins , angiotensin - converting - enzyme - ace inhibitors , and angiotensin receptor type 1 blockers atb ) . the institutional review committee approved our study protocol therebys following local biomedical research regulations ( irb no . a complete blood count ; serum urea and creatinine , albumin ; lipid status , and urine sediment were determined in blood and urine samples collected 1 day before kidney biopsy as well proteinuria / day . only samples with a sterile urine culture were processed . urine sediments with more than 3 rbc / hpf or 5 wbc / hpf were defined as clinically significant erythrocyturia or leukocyturia . estimated glomerular filtration rate ( egfr ) was calculated with a shortened version of the modification of diet in renal disease ( mdrd ; patients 18 years ) . alteration in tin were assessed after routine hematoxylin - eosin staining by a qualified nephropathologist who was not familiar with the preset histopathologic or clinical diagnosis , nor the identity of patients . inflammation activity was then scored numerically ( 0 : no inflammation , 1 : slightly marked inflammation , 2 : moderate inflammation , and 3 : strong inflammation ) as well as atrophy and fibrosis advancement ( 0 : no atrophy and no fibrosis , 1 : slightly marked atrophy and fibrosis , 2 : moderate atrophy and fibrosis , 3 : very pronounced atrophy and fibrosis ) . after deparaffinization tissue samples were rehydrated and stained immunohistochemically for kim-1 using the tissue kim-1/tim-1 kit ( r & d systems inc . , minneapolis , mn , usa ) with contrasting hematoxylin and eosin staining . a tubule was considered to be kim-1 positive if it contained at least one kim-1 positive cell regardless of whether the kim-1 expression was cytoplasmic or apical . kim-1 staining was scored semiquantitatively by estimating the percentage of cortical tubules expressing kim-1 per field ( the complete biopsy area was scored , with a minimum of five fields ; in controls 30 fields were scored ) . unaffected parts of kidneys from patients with renal cell carcinoma were used as the negative control . the 6- , 12- , 24- , and 36-month followup for all patients included measurement of serum creatinine concentration , egfr , and proteinuria . data are presented as mean 1 standard deviation ( sd ) and the range of values . the one - sample kolmogorov - smirnov test was used to check for normal distribution of the variables . relationships between variables were estimated using pearson 's rank and spearmen 's correlation tests as appropriate . independent predictors of renal function 6 , 12 , 24 , and 36 months after the kidney biopsy were identified by stepwise multivariant regression analysis . table 2 presents the demographic , clinical , and laboratory data for the patients at the time of kidney biopsy . tissue kim-1 expression was significantly induced in all kidney biopsies except in one patient with mcgn . tissue kim-1 had a typically apical localization , but it could also be found in the cytoplasm with no apparent affinity for the apical membrane of tubulocytes . kim-1 positive tubules were located mainly in the tin regions affected by inflammation or fibrosis but were not observed in regions with end - stage fibrosis . kim-1 univariate analysis showed significant positive associations between tissue kim-1 expression and age ( r = 0.313 ; p = 0.015 ) , tin inflammation ( r = 0.4563 ; p = 0.004 ) , tin fibrosis ( r = 0.317 ; p = 0.021 ) , and hemoglobin ( r = 0.440 ; p = 0.001 ) as well as negative associations with albumin concentration ( r = 0.376 ; p = 0.011 ) and egfr ( r = 0.572 ; p < 0.001 ) at the time of biopsy and 6 ( r = 0.442 ; p = 0.002 ) , 24 ( r = 0.398 ; p = 0.012 ) , and 36 ( r = 0.412 ; p = 0.015 ) months later . tissue kim-1 expression correlated significantly only with proteinuria / day 6 months after biopsy ( r = 0.394 ; p = 0.026 ) but not with proteinuria at the time of biopsy and 12 , 24 , and 36 months later . table 3 gives the correlation coefficients between egfr at the time of biopsy and 6 , 12 , 24 , and 36 months later and the examined variables . multivariant stepwise regression analysis showed that the best predictor of kidney function at the time of biopsy ( egfr 0 ) was tin inflammation ( p < 0.001 ) , as well as of egfr12 ( p = 0.015 ) , egfr 24 ( p = 0.039 ) , and egfr 36 ( p = 0.012 ) . however tissue kim-1 expression was the best predictor of egfr 6 ( p = 0.016 ) along with tin inflammation ( p = 0.016 ) ( table 4 ) . figure 2 presents the linear correlation between tin inflammation ( p < 0.001 ) , tissue kim-1 expression ( p < 0.001 ) , and egfr 6 . in our previous 6-month prospective study , we found higher urine kim-1 concentrations in twenty chronic renal patients than in control subjects and a significant positive correlation between urine kim-1 level and kim-1 expression in tissue . although , urine kim-1 content reflected tissue kim-1 expression , tissue kim-1 correlated better with tin inflammation and fibrosis as well as with kidney function at the time of biopsy and 3 and 6 months later . therefore , we retrospectively examined sixty patients with different kidney diseases in order to evaluate the importance of tissue kim-1 expression in predicting kidney function in the ensuing 36 months after kidney biopsy . in the present study tissue in addition , kim-1 positive tubules were located in parts of the cortex that were imbued with inflammatory infiltrate and/or fibrotic changes but not in completely atrophic areas . it is well known that the expression of kim-1 in tubules is associated with inflammation and tin damage . double - labeling studies have shown simultaneous expression of kim-1 and markers of prefibrotic changes , repair , and chemotaxis [ 10 , 11 ] . the study of van timmeren et al .   also demonstrated a connection between kim-1 expression in tissue , interstitial fibrosis , and macrophage accumulation . .   first showed that tubular epithelial cells expressing kim-1 on their surface could act as phagocytes for apoptotic and necrotic renal epithelial cells . kim-1 is expressed predominantly in renal tubules , which , judging by the simultaneous expression of markers of dedifferentiation , are currently under the influence of agents that cause injury , but is absent from completely atrophic tubules . controversy remains about the function of kim-1 : is it actively regulating the inflammatory process ? or is its expression just a response to damage , attempted recovery , and/or repair ? . the present study revealed a significant negative correlation between tissue kim-1 expression and all estimated gfr at the time of kidney biopsy and 6 , 24 , and 36 months later . however multivariate analysis pointed to tin inflammation , but neither tin fibrosis nor proteinuria , as the best predictor of kidney function at biopsy and 6 , 12 , 24 , and 36 months afterwards . however , along with tin inflammation , tissue kim-1 expression was the best predictor of kidney function 6 months after biopsy . interstitial infiltration of inflammatory cells can be seen in a variety of immune and nonimmune kidney diseases and is thought to play a significant role in tin damage and fibrosis . upon exposure to high protein concentrations , renal tubule cells produce a host of chemokines , vasoactive mediators , and adhesion molecules , which may contribute to interstitial fibrosis . protocol immunosuppressive therapy and treatment with ace inhibitors and at blockers for chronic renal patients with different chronic immune and nonimmune - mediated kidney disorders proven by biopsy have the strongest effects during the first 6 months of therapy . the severity of pretreatment tin damage predicts a dulled response to renoprotective intervention , with a worse long - term renal outcome . therefore , we can speculate that patients with less interstitial inflammation at the time of kidney biopsy will have a more favorable response to treatment and better kidney function afterwards . also , the finding that kim-1 was the strongest predictor of kidney function 6 months after biopsy along with tin inflammation may support the hypothesis about its potential role in the development of interstitial fibrosis and an unsatisfactory treatment response . association between higher urinary kim-1 levels and poorer kidney function was found in some human studies in native [ 10 , 14 ] and transplanted kidneys   but data on human tissue kim-1 expression are scarce . zhang et al .   found that kim-1 expression in transplant biopsies is a sensitive measure of cell injury . in addition , they showed that more intense kim-1 staining predicts a better graft outcome over the ensuing 18 months and speculated that its level of expression may be an indicator of graft function recovery . it is well known that proteinuria is an ominous biomarker of progressive kidney disease   and our study revealed that only proteinuria 6 months after kidney biopsy correlated negatively with kidney function 24 and 36 months after the biopsies and positively with kim-1 expression in tissue . earlier investigations [ 10 , 14 , 17 ] have yielded conflicting data regarding the relationship between urinary kim-1 level or tissue kim-1 expression and proteinuria . one explanation may be that proteinuria is not always accompanied by tin damage and a progressive decline in renal function [ 18 , 19 ] . data regarding the selectivity of proteinuria were not available for the purposes of this study . the present study exposed a strong association between kim-1 tissue staining and tin inflammation and fibrosis but not with tin atrophy . although multivariate analysis pointed to tin inflammation , as the best predictor of kidney function at biopsy and 6 , 12 , 24 , and 36 months later , tissue kim-1 expression is one of the best predictor of kidney function 6 months after biopsy , the time when treatment effects are the strongest . therefore , we can speculate that kim-1 has a potential role in the development of interstitial fibrosis and poor treatment responses . the major limitation of this investigation is the relatively small number of patients examined but it could be the basis for a larger prospective follow - up study for evaluation of urinary kim-1 concentration and tissue kim-1 expression in patients with various chronic and acute renal diseases .
objectives . retrospective study was designed to examine the importance of tissue kidney injury molecule-1 ( kim-1 ) expression in predicting kidney function in sixty patients ( 27 males ) aged 34.15  12.23 years with different kidney diseases over three years after kidney biopsy . 
 materials and methods . 
 tissue kim-1 expression was determined immunohistochemically and kim-1 staining was scored semiquantitatively , as well as tubulointerstitialis ( tin ) , inflammation , atrophy , and fibrosis . kidney function ( mdrd formula ) and 
 proteinuria / day were evaluated at the time of biopsy ( gfr0 ) and 6 , 12 , 24 , and 36 months later . 
 results . significantly positive correlations between tissue kim-1 expression and age ( r = 0.313 ) , tin inflammation ( r = 0.456 ) , fibrosis ( r = 0.317 ) , and 
 proteinuria at 6 months ( r = 0.394 ) as well as negative correlations with gfr0 ( r = 0.572 ) , gfr6 ( r = 0.442 ) , gfr24 ( r = 0.398 ) , and gfr36 ( r = 0.412 ) were found . meanwhile , tin inflammation was the best predictor of all measured kidney functions during three years , while tissue kim-1 expression ( p = 0.016 ) was a predictor only at 6 months after biopsy . 
 conclusion . 
 tissue kim-1 expression significantly predicts kidney function solely at 6 months after biopsy , when the effects of immune and nonimmune treatments are the strongest .