**2. Acute Pyelonephritis (APN) in children and genetic susceptibility to APN**

Urinary tract infections (UTIs) are common among children of all ages including infants. UTI is defined as a penetration of microorganisms, mainly *E. coli*, into the tissue of urina‐ ry tract, which is marked by significant bacteriuria (>105 bacteria per 1ml of urine) [1]. UTIs are classified into three categories: upper UTI- acute pyelonephritis, lower UTIacute cystitis and asymptomatic bacteriuria (ABU). The upper UTI, or acute pyelonephri‐ tis (APN), represents bacterial infection of renal parenchyma, which may cause various inflammatory lesions. Post-infectious renal scarring is the most serious complication fol‐ lowing APN in children, with an estimated incidence of 10-65% [1-2]. Vesico-ureteral re‐ flux (VUR) may also play an important role in renal damage [3]. VUR is suggested to be a weak predictor of permanent renal damage in children hospitalized with UTI [4] but it is also known that the grade of VUR positively correlates with likelihood of renal scarring [5]. Extensive renal scarring leads to renal insufficiency and hypertension [6-7]. Early diag‐ nosis of APN and follow up to identify renal scarring after the first APN are thus very important. The primary distinction of APN is based on clinical manifestations and indirect laboratory testing of inflammatory markers such as C-reactive protein (CRP) serum levels, peripheral white blood cells' (WBC) count etc. However, these tests are unreliable in acute phase of pyelonephritis. The 99mTc-dimercaptosuccinic acid (DMSA) scintigraphy is a gold‐ en standard method for detection of acute renal inflammatory lesions specific for diagno‐ sis of APN as well as for the follow-up detection of renal cortical scars [8-9]. Detection of permanent renal parenchymal defects following APN is ultimate for long-term prognosis of kidney function. The incidence of renal defects correlates inversely with the time inter‐ val between pyelonephritis and the scintigraphic study and stabilizes 4-6 months follow‐ ing acute disease. DMSA scintigraphy is based on binding of 99mTc-DMSA to renal parenchyma cells and therefore provides means of distinguishing APN from lower UTI and evaluating persistent DMSA uptake defects after the initial infection in children [8- 10]. Given that DMSA exposes the patients to radiation, this procedure is not regularly used to diagnose APN.

### **2.1. Bacterial virulence and uroepithelial contact**

cytokines. Changes in genes' expression as well as presence of certain alleles associated with disease phenotype support the hypothesis that genetic factors could modify susceptibility to

Several mechanical forces including urine flow and voiding, mucus shedding, and epithelial cell sloughing are important in minimizing UTI incidence. Bacterial adherence to the epithe‐ lium triggers defense responses. One of these is innate immunity response, which is impor‐ tant for uropathogenic *E. coli* recognition and immunomobilization. The innate immunity response is mediated by toll-like receptors (TLR4, TLR5, TLR11), adhesion molecules (E-se‐ lectin, ICAM-1, PECAM-1) and secreted factors such as cytokines (TNF-alpha, IL-1beta, IL-6, G-CSF, IL-17) and chemokines (CXCL1, CXCL2, CXCL3, CXCL8, CCL4). These molecules have been detected in mammalian bladder upon infection. Therefore, polymorphisms in genes coding for these molecules have been recognized as genetic susceptibility factors for UTIs. Neutrophils are the most abundant early responders to infection, while the anti‐ gen(Ag)-presenting macrophages, dendritic cells and innate-like lymphocytes (such as gam‐

The cytokine response is essential for antibacterial defense of the urinary tract. Interleu‐ kin-8 (IL-8) is a potent chemoattractant responsible for neutrophil infiltration into the urinary tract. It was reported that neutrophils of children with recurrent pyelonephritis had lower expression of IL-8 receptor (CXCR1) than neutrophils of healthy controls. In‐ terleukin-6 (IL-6) is one of the pro-inflammatory cytokines, which stimulates production of all acute-phase proteins thus contributing to acute-phase response and systemic in‐ flammation. High serum or urine levels of IL-6 have been found in children with UTIs, particularly in children with APN compared to those with lower UTI. It was also indi‐ cated that urine and serum levels of cytokines could be observed as markers of renal damage as well as tools for monitoring the development and outcome of APN. Urine analysis has been extensively used by clinicians to diagnose various renal diseases. Ad‐ vances in technology of molecular biology enable analyses of genes' expression levels in urine sediment. These expression studies have a potential to improve the diagnosis of APN by detecting urinary gene expression profiles, which are specific for patients with

Besides susceptibility to UTIs, of great therapeutic importance is susceptibility to post-infec‐ tious renal damage in APN patients. This kidney damage susceptibility has the genetic com‐ ponent. Among the candidate genes are those coding for molecules like growth factors (TGF-β1, VEGF), which play important roles in processes characteristic for the tissue dam‐ age and scarring such as cell proliferation and accumulation of extracellular matrix. Angio‐ tensin II, main effector of the renin-angiotensin system, is also considered a growth factor

Here we will review the roles of candidate genes' polymorphisms and expression in sus‐ ceptibility to APN and post-infectious renal scarring, in order to summarize the existing re‐

involved in all phenomena of renal tissue damaging and scar formation.

sults and point out to further possible directions for research in this field.

acute pyelonephritis and post-infectious renal damage.

138 Recent Advances in the Field of Urinary Tract Infections

ma delta T cells) have been implicated in the UTI host defense.

APN.

Uropathogenic *Escherichia coli* is the most common causative agent (80%) of uncomplicated UTIs although other enteric organisms have been identified as well [11]. After colonization of the urethra and ascent to the bladder, bacteria bind to glycosphingolipid and glycopro‐ tein receptors on the urinary tract epithelium and penetrate into tissue of urinary tract [12]. They express a number of virulence determinants that contribute to successful colonization of the urinary tract [13]. Many pathogenic microorganisms use host cell surface oligosac‐ charides including glycosphingolipids (GSLs) as receptors to attach to uroepithelial cells. The attachment of *E. coli* is mediated trough expression of flagellin and ascending of *E. coli* to the upper urinary tract and dissemination of bacteria within the host are enabled through a flagellum-mediated motility [14-16]. These actions, along with the lipid A moiety of lipo‐ polysaccharide (endotoxin), have been shown to enhance activation of the host inflammato‐ ry response. Cytokines mediate this response [7,17-19]. Neutrophils are the first cells that migrate to the uroepithelium in the event of UTI and they are crucial in control of infection at early time points [20].

thelial lining that they cross to reach peripheral tissues. There is increasing evidence that the fate of neutrophils outside the vascular system is governed by specialized molecular interac‐ tions distinct from those in blood vessels [3] but these aspects have received less attention and are not well understood. Mucosal pathogens trigger a rapid neutrophil response [4-5], given that neutrophils are crucial effectors of the host defense [6-7]. The mucosal neutrophil response initiates when bacteria stimulate the epithelial cells to secrete chemokines [5,42] and to increase their chemokine receptors' expression. Neutrophils respond to so-formed chemotactic gradient, leave the bloodstream, travel through the submucosa and reach the basal side of the epithelial barrier, which they cross into the lumen [3,42]. Attention has been focused on molecular interactions of neutrophils with endothelial cells during the extravasa‐ tion process [1-2], because this is the key to subsequent pathology and tissue destruction.

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141

The interindividual differences in frequency and severity of UTIs exist and they are con‐ sistent with a genetic predisposition among disease-prone individuals. Structural defects such as congenital anomalies of kidney and urinary tract as well as social and environ‐ mental factors influence disease susceptibility [1-2]. There have been many attempts to identify the host factors that predispose to UTIs, especially to acute pyelonephritis (APN). Recurrent APN occurs within a small group of highly susceptible individuals, some of whom develop progressive renal scarring and therefore may need dialysis or kidney

In an attempt to characterize the critical mechanisms and candidate genes for APN suscepti‐ bility, the "knockout" mice were investigated. It has been shown that the innate immunity

Large interindividual differences both in frequency and severity of UTIs are consistent with genetic predisposition among disease-prone individuals although inherited defects in mech‐ anisms of defense against UTIs have not been identified. Since the experimental studies sug‐ gested that susceptibility to clinical APN was genetically controlled and that the disease severity might vary with the expression levels of specific host response molecules, the next

Phagocytosis of microorganisms and their intracellular degradation by the tissue macro‐ phages represent stimuli for synthesis of proinflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-α) [43]. IL-1 and TNF-α induce expression of the adhesion molecules on the surface of endothelial cells, which bind the circulating leuko‐ cytes and allow their recruitment into the tissue. IL-1 and TNF-α also stimulate cells of the infected tissue to produce other mediators of inflammation such as cytokine interleu‐ kin-6 (IL-6) and chemokines, which regulate leukocytes' functions as well as their transen‐ dothelial migration into the inflammatory tissue [43]. Chemokines, particularly interleukin-8 (IL-8), are released from stimulated endothelial cells and macrophages. They

response genes strongly influence susceptibility to UTIs, particularly APN [5-7].

step was to investigate the susceptibility candidate genes in human population.

**2.5. Genetic susceptibility to APN and renal scarring following APN**

transplantation [2-4].

*2.5.1. Cytokines*

### **2.2. Renal scarring following APN**

Bacterial infection of renal parenchyma during APN represents the major cause of acquired renal damage in children. The inflammatory changes associated with acute pyelonephritis are reversible but in some cases they result in renal defects. The percentage of children with renal scarring detected six months after the first APN is similar in the recent studies [21-23] and is in agreement with the results of European meta-analysis study of post-pyelonephritic renal scarring incidence [24]. Post-infectious renal scarring, as the most serious complication following APN, appears in 10-65% of children [8,25]. This renal damage can lead to hyper‐ tension and chronic renal failure [26-28].

The actual etiology of renal scarring remains controversial. The risk factors supposed to be associated with renal scars are: presence of vesico-ureteral reflux (VUR), delay in adequate antibiotic treatment, presence of recurrent UTI, bacterial virulence, host defense factors, host inflammatory and immunologic reactions and genetic susceptibility.

### **2.3. Vesico-Ureteral Reflux (VUR) as a risk factor for renal scarring**

VUR is classically considered a risk factor for development of renal scars. The theory that reflux might play an important role in renal damage was proposed by Hodson and Edvards in 1960 [29]. Ransley and Risdon showed that scarring occurred only when urinary infection was present in association with VUR and intrarenal reflux [30]. Later it was suggested that VUR was a weak predictor of permanent renal damage in children hospitalized with UTI [31]. However, development of scars occurred even in absence of VUR, so there has been a debate for many years over the role of VUR in children who developed renal scars following UTI [31-35]. There is also a debate whether the grade of VUR positively correlates with like‐ lihood of renal scarring or not [36] and whether the age represents a risk factor for scars' formation [37]. Gleeson and Gordon reported that there was a significant correlation be‐ tween detection of a scarred kidney on DMSA scan and presence of VUR in children who were less than one year old [37]. In children aged over one year there was a poor correlation with renal scarring, so they suggested that the young growing kidney might be more vul‐ nerable to insults. However, others [25,38-39] did not confirm that younger children were at greater risk for development of renal sequelae following pyelonephritis. Moreover, in some studies [40] children aged over one year had a higher frequency of renal scarring in compar‐ ison to infants.

### **2.4. Host inflammatory response and kidney damage following APN**

Roberts et al. have suggested that the acute inflammatory response causing the eradication of bacteria could be responsible for the early pyelonephritic damage of renal tissue and sub‐ sequent renal scarring [41]. Neutrophils migrate between tissue compartments and exert their effector functions at different sites. They circulate in blood and interact with the endo‐ thelial lining that they cross to reach peripheral tissues. There is increasing evidence that the fate of neutrophils outside the vascular system is governed by specialized molecular interac‐ tions distinct from those in blood vessels [3] but these aspects have received less attention and are not well understood. Mucosal pathogens trigger a rapid neutrophil response [4-5], given that neutrophils are crucial effectors of the host defense [6-7]. The mucosal neutrophil response initiates when bacteria stimulate the epithelial cells to secrete chemokines [5,42] and to increase their chemokine receptors' expression. Neutrophils respond to so-formed chemotactic gradient, leave the bloodstream, travel through the submucosa and reach the basal side of the epithelial barrier, which they cross into the lumen [3,42]. Attention has been focused on molecular interactions of neutrophils with endothelial cells during the extravasa‐ tion process [1-2], because this is the key to subsequent pathology and tissue destruction.

### **2.5. Genetic susceptibility to APN and renal scarring following APN**

The interindividual differences in frequency and severity of UTIs exist and they are con‐ sistent with a genetic predisposition among disease-prone individuals. Structural defects such as congenital anomalies of kidney and urinary tract as well as social and environ‐ mental factors influence disease susceptibility [1-2]. There have been many attempts to identify the host factors that predispose to UTIs, especially to acute pyelonephritis (APN). Recurrent APN occurs within a small group of highly susceptible individuals, some of whom develop progressive renal scarring and therefore may need dialysis or kidney transplantation [2-4].

In an attempt to characterize the critical mechanisms and candidate genes for APN suscepti‐ bility, the "knockout" mice were investigated. It has been shown that the innate immunity response genes strongly influence susceptibility to UTIs, particularly APN [5-7].

Large interindividual differences both in frequency and severity of UTIs are consistent with genetic predisposition among disease-prone individuals although inherited defects in mech‐ anisms of defense against UTIs have not been identified. Since the experimental studies sug‐ gested that susceptibility to clinical APN was genetically controlled and that the disease severity might vary with the expression levels of specific host response molecules, the next step was to investigate the susceptibility candidate genes in human population.

### *2.5.1. Cytokines*

migrate to the uroepithelium in the event of UTI and they are crucial in control of infection

Bacterial infection of renal parenchyma during APN represents the major cause of acquired renal damage in children. The inflammatory changes associated with acute pyelonephritis are reversible but in some cases they result in renal defects. The percentage of children with renal scarring detected six months after the first APN is similar in the recent studies [21-23] and is in agreement with the results of European meta-analysis study of post-pyelonephritic renal scarring incidence [24]. Post-infectious renal scarring, as the most serious complication following APN, appears in 10-65% of children [8,25]. This renal damage can lead to hyper‐

The actual etiology of renal scarring remains controversial. The risk factors supposed to be associated with renal scars are: presence of vesico-ureteral reflux (VUR), delay in adequate antibiotic treatment, presence of recurrent UTI, bacterial virulence, host defense factors, host

VUR is classically considered a risk factor for development of renal scars. The theory that reflux might play an important role in renal damage was proposed by Hodson and Edvards in 1960 [29]. Ransley and Risdon showed that scarring occurred only when urinary infection was present in association with VUR and intrarenal reflux [30]. Later it was suggested that VUR was a weak predictor of permanent renal damage in children hospitalized with UTI [31]. However, development of scars occurred even in absence of VUR, so there has been a debate for many years over the role of VUR in children who developed renal scars following UTI [31-35]. There is also a debate whether the grade of VUR positively correlates with like‐ lihood of renal scarring or not [36] and whether the age represents a risk factor for scars' formation [37]. Gleeson and Gordon reported that there was a significant correlation be‐ tween detection of a scarred kidney on DMSA scan and presence of VUR in children who were less than one year old [37]. In children aged over one year there was a poor correlation with renal scarring, so they suggested that the young growing kidney might be more vul‐ nerable to insults. However, others [25,38-39] did not confirm that younger children were at greater risk for development of renal sequelae following pyelonephritis. Moreover, in some studies [40] children aged over one year had a higher frequency of renal scarring in compar‐

inflammatory and immunologic reactions and genetic susceptibility.

**2.3. Vesico-Ureteral Reflux (VUR) as a risk factor for renal scarring**

**2.4. Host inflammatory response and kidney damage following APN**

Roberts et al. have suggested that the acute inflammatory response causing the eradication of bacteria could be responsible for the early pyelonephritic damage of renal tissue and sub‐ sequent renal scarring [41]. Neutrophils migrate between tissue compartments and exert their effector functions at different sites. They circulate in blood and interact with the endo‐

at early time points [20].

ison to infants.

**2.2. Renal scarring following APN**

140 Recent Advances in the Field of Urinary Tract Infections

tension and chronic renal failure [26-28].

Phagocytosis of microorganisms and their intracellular degradation by the tissue macro‐ phages represent stimuli for synthesis of proinflammatory cytokines, interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF-α) [43]. IL-1 and TNF-α induce expression of the adhesion molecules on the surface of endothelial cells, which bind the circulating leuko‐ cytes and allow their recruitment into the tissue. IL-1 and TNF-α also stimulate cells of the infected tissue to produce other mediators of inflammation such as cytokine interleu‐ kin-6 (IL-6) and chemokines, which regulate leukocytes' functions as well as their transen‐ dothelial migration into the inflammatory tissue [43]. Chemokines, particularly interleukin-8 (IL-8), are released from stimulated endothelial cells and macrophages. They act as chemoattractants stimulating the chemotaxis of neutrophils and neutrophil adhe‐ sion to stimulated endothelium [43,44].

dren included in this study have been followed from their first episode of APN and adults had a history of APN in childhood. Kidney status was defined by DMSA scan. The UTI-as‐ sociated CXCR1 variant, +217 C/G, has been shown to reduce RUNX1 binding to the puta‐ tive intronic binding site. Furthermore, transfection experiments showed that transcription level of the mutant allele was lower, suggesting that +217 C/G polymorphism could reduce

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143

The other study investigated polymorphisms of IL-8 gene, -251 A/T and +2767 A/G, and a polymorphism +2608 G/C of IL-8 receptor gene (CXCR1) in children with the first episode of upper UTI and APN documented by DMSA [18]. There were no statistically significant dif‐ ferences in genotype and allele frequencies of the IL-8 and CXCR1 polymorphisms between the UTI population and controls [18]. After comparison of genotype frequencies between DMSA positive children (with definite APN) and DMSA negative children, there were no significant differences between these two groups. Still, IL-8 -251 TT genotype was signifi‐ cantly more frequent in DMSA negative children suggesting that a carriage of A allele repre‐ sents susceptibility factor for APN. By exclusion of patients with VUR, the genotype frequencies between DMSA positive and DMSA negative children were significantly differ‐ ent for IL-8 gene polymorphisms, -251 A/T and +2767 A/G. Again, -251 TT homozygotes were more frequent in DMSA negative children. These results, overall, suggest that IL-8 -251 A allele is significantly associated with presence of DMSA documented pyelonephritis [18]. Experimental data also showed that -251 A allele was associated with increase in IL-8 pro‐ duction in lipopolysaccharide stimulated whole blood [42]. The CXCR1 gene polymorphism

Polymorphisms of genes coding for MCP-1 (monocyte chemotactic protein-1)/CCL2 and RANTES (Regulated upon Activation, Normal T-cell Expressed, and Secreted)/CCL5 and their receptors, CCR2 and CCR5 respectively, have been associated with the upper UTI. On‐ ly RANTES -403 G allele was significantly associated with risk for UTI, irrespectively of

Initial steps of the inflammatory response are mediated by adhesion of inflammatory cells (leukocytes) to vascular endothelial cells [58-59]. Inflammatory cells then exit the circulation and infiltrate the surrounding tissue. This process is mediated by E-selectin, intercellular ad‐ hesion molecule-1 (ICAM-1) and platelet endothelial cell adhesion molecule-1 (PECAM-1) through the sequential steps of rolling, strong adhesion and diapedesis, respectively [59-60]. Data from ICAM-1 knockout animals showed that these animals had elevated neutrophil

In the group of children with a proven history of UTI, E-selectin, ICAM-1, PECAM-1 and CD11b gene polymorphisms were investigated. There were no significant differences in al‐ lele frequencies between patients and controls for any of the investigated polymorphisms

and lymphocyte counts and decreased neutrophil influx to the site of infection [61].

CXCR1 transcription [56].

+2608 G/C was not associated with APN [18].

*2.5.1.4. MCP-1/CCL2 and RANTES/CCL5*

VUR [57].

*2.5.2. Adhesion molecules*

Elevated serum levels of the proinflammatory cytokines- TNF-α, IL-1 and IL-6, have been measured in children with UTIs, with a significantly greater increase in children with APN than in those with lower UTI [10].

### *2.5.1.1. TNF-α*

A single nucleotide polymorphism (SNP) -308 A/G in the promoter sequence of TNF-α gene is located at the binding site of the transcription factor activating protein-2 [45]. It has been suggested that TNF -308 A allele is related to a higher production of TNF-α [46]. No differ‐ ences have been demonstrated in TNF-α -308 A/G genotype frequencies between infants with UTI (with and without renal scarring) and controls [47].

### *2.5.1.2. IL-6*

Interleukin-6 (IL-6) is one of the pro-inflammatory cytokines, which stimulates production of all acute-phase proteins thus contributing to acute-phase response and systemic inflam‐ mation [10]. IL-6 is synthesized by several types of cells, in response to various antigens such as bacterial pathogens [48-49]. There are studies suggesting that IL-6 could be synthe‐ sized by uroepithelial or renal tubular cells [49-50]. High serum or urine levels of IL-6 have been detected in children with UTIs, particularly in those with APN in comparison to those with lower UTI [51-52]. It was indicated that urine and serum levels of this cytokine could be observed as markers of renal damage as well as tools for monitoring the development and outcome of APN [22,51-52].

Regulation of IL-6 expression is mainly accomplished at transcription level. A SNP -174 G/C, in the promoter region of IL-6 gene, is located 11 bp upstream from cis-regulatory element (CRE) and reported to influence the level of IL-6 expression in healthy individuals [53]. Moreover, -174 G/C polymorphism was found to be associated with circulating IL-6 levels and course of certain inflammatory diseases [54-55]. This polymorphism was investigated in association with APN and renal scarring in our study [21]. The genotype distributions and allele frequencies were not significantly different between the two investigated patients' groups, with APN and lower UTI. We concluded that IL-6 -174 G/C polymorphism was not a susceptibility factor for APN. This polymorphism was neither recognized as a risk factor for renal scarring in patients with the first APN [21]. Still, we detected a significant increase in white blood cells' count in APN children with CC genotype compared to those with wild type, GG, genotype.

### *2.5.1.3. IL-8 and CXCR1*

The IL-8 receptor, CXCR1, was identified a candidate gene for acute pyelonephritis when mIL-8Rh mutant mice developed APN with severe tissue damage [6-7]. After sequencing that covered the entire CXCR1 gene two genetic variants, +217 C/G and +2608 G/C, were found to be susceptibility factors for APN in both children and adults [56]. Infants and chil‐ dren included in this study have been followed from their first episode of APN and adults had a history of APN in childhood. Kidney status was defined by DMSA scan. The UTI-as‐ sociated CXCR1 variant, +217 C/G, has been shown to reduce RUNX1 binding to the puta‐ tive intronic binding site. Furthermore, transfection experiments showed that transcription level of the mutant allele was lower, suggesting that +217 C/G polymorphism could reduce CXCR1 transcription [56].

The other study investigated polymorphisms of IL-8 gene, -251 A/T and +2767 A/G, and a polymorphism +2608 G/C of IL-8 receptor gene (CXCR1) in children with the first episode of upper UTI and APN documented by DMSA [18]. There were no statistically significant dif‐ ferences in genotype and allele frequencies of the IL-8 and CXCR1 polymorphisms between the UTI population and controls [18]. After comparison of genotype frequencies between DMSA positive children (with definite APN) and DMSA negative children, there were no significant differences between these two groups. Still, IL-8 -251 TT genotype was signifi‐ cantly more frequent in DMSA negative children suggesting that a carriage of A allele repre‐ sents susceptibility factor for APN. By exclusion of patients with VUR, the genotype frequencies between DMSA positive and DMSA negative children were significantly differ‐ ent for IL-8 gene polymorphisms, -251 A/T and +2767 A/G. Again, -251 TT homozygotes were more frequent in DMSA negative children. These results, overall, suggest that IL-8 -251 A allele is significantly associated with presence of DMSA documented pyelonephritis [18]. Experimental data also showed that -251 A allele was associated with increase in IL-8 pro‐ duction in lipopolysaccharide stimulated whole blood [42]. The CXCR1 gene polymorphism +2608 G/C was not associated with APN [18].

### *2.5.1.4. MCP-1/CCL2 and RANTES/CCL5*

Polymorphisms of genes coding for MCP-1 (monocyte chemotactic protein-1)/CCL2 and RANTES (Regulated upon Activation, Normal T-cell Expressed, and Secreted)/CCL5 and their receptors, CCR2 and CCR5 respectively, have been associated with the upper UTI. On‐ ly RANTES -403 G allele was significantly associated with risk for UTI, irrespectively of VUR [57].

### *2.5.2. Adhesion molecules*

act as chemoattractants stimulating the chemotaxis of neutrophils and neutrophil adhe‐

Elevated serum levels of the proinflammatory cytokines- TNF-α, IL-1 and IL-6, have been measured in children with UTIs, with a significantly greater increase in children with APN

A single nucleotide polymorphism (SNP) -308 A/G in the promoter sequence of TNF-α gene is located at the binding site of the transcription factor activating protein-2 [45]. It has been suggested that TNF -308 A allele is related to a higher production of TNF-α [46]. No differ‐ ences have been demonstrated in TNF-α -308 A/G genotype frequencies between infants

Interleukin-6 (IL-6) is one of the pro-inflammatory cytokines, which stimulates production of all acute-phase proteins thus contributing to acute-phase response and systemic inflam‐ mation [10]. IL-6 is synthesized by several types of cells, in response to various antigens such as bacterial pathogens [48-49]. There are studies suggesting that IL-6 could be synthe‐ sized by uroepithelial or renal tubular cells [49-50]. High serum or urine levels of IL-6 have been detected in children with UTIs, particularly in those with APN in comparison to those with lower UTI [51-52]. It was indicated that urine and serum levels of this cytokine could be observed as markers of renal damage as well as tools for monitoring the development

Regulation of IL-6 expression is mainly accomplished at transcription level. A SNP -174 G/C, in the promoter region of IL-6 gene, is located 11 bp upstream from cis-regulatory element (CRE) and reported to influence the level of IL-6 expression in healthy individuals [53]. Moreover, -174 G/C polymorphism was found to be associated with circulating IL-6 levels and course of certain inflammatory diseases [54-55]. This polymorphism was investigated in association with APN and renal scarring in our study [21]. The genotype distributions and allele frequencies were not significantly different between the two investigated patients' groups, with APN and lower UTI. We concluded that IL-6 -174 G/C polymorphism was not a susceptibility factor for APN. This polymorphism was neither recognized as a risk factor for renal scarring in patients with the first APN [21]. Still, we detected a significant increase in white blood cells' count in APN children with CC genotype compared to those with wild

The IL-8 receptor, CXCR1, was identified a candidate gene for acute pyelonephritis when mIL-8Rh mutant mice developed APN with severe tissue damage [6-7]. After sequencing that covered the entire CXCR1 gene two genetic variants, +217 C/G and +2608 G/C, were found to be susceptibility factors for APN in both children and adults [56]. Infants and chil‐

sion to stimulated endothelium [43,44].

142 Recent Advances in the Field of Urinary Tract Infections

with UTI (with and without renal scarring) and controls [47].

than in those with lower UTI [10].

and outcome of APN [22,51-52].

type, GG, genotype.

*2.5.1.3. IL-8 and CXCR1*

*2.5.1.1. TNF-α*

*2.5.1.2. IL-6*

Initial steps of the inflammatory response are mediated by adhesion of inflammatory cells (leukocytes) to vascular endothelial cells [58-59]. Inflammatory cells then exit the circulation and infiltrate the surrounding tissue. This process is mediated by E-selectin, intercellular ad‐ hesion molecule-1 (ICAM-1) and platelet endothelial cell adhesion molecule-1 (PECAM-1) through the sequential steps of rolling, strong adhesion and diapedesis, respectively [59-60]. Data from ICAM-1 knockout animals showed that these animals had elevated neutrophil and lymphocyte counts and decreased neutrophil influx to the site of infection [61].

In the group of children with a proven history of UTI, E-selectin, ICAM-1, PECAM-1 and CD11b gene polymorphisms were investigated. There were no significant differences in al‐ lele frequencies between patients and controls for any of the investigated polymorphisms [62]. Still, A allele of ICAM-1 exon 4 (G/A) polymorphism had significantly lower frequency in patients who developed renal scars following UTI compared with the patients without scars. This suggested that the A allele might be a protective factor for renal scarring follow‐ ing UTI. It is possible that protective effect of the A allele on development of renal scars fol‐ lowing UTI may be a result of decreased binding of neutrophils and other inflammatory cells to ICAM-1, resulting in reduced leukocyte infiltration and reduced tissue damage [62].

*2.5.4. Vascular Endothelial Growth Factor (VEGF) and Transforming Growth Factor-beta1*

Vascular endothelial growth factor (VEGF) is a potent mitogen that enhances angiogene‐ sis, microvascular permeability and proliferation of vascular endothelial cells [74]. Expres‐ sion of VEGF has been detected in glomerular podocytes, epithelial cells of distal tubules and collecting ducts of kidney. Neovascularization in scarred kidney tissue and significant increase of urine VEGF levels associated with increased severity of renal scarring have been reported [75-76]. Transforming growth factor-beta1 (TGF-β1) appears to be one of the key factors in process of tissue repair. It is involved in regulation of cell proliferation, differentiation, extracellular matrix production and immune response [77]. Studies sug‐ gested that TGF-β1 could be involved in pathogenesis of congenital obstructive uropathies

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Several polymorphisms in VEGF and TGF-β1 genes have been linked to overproduction of these proteins as well as predisposition to progressive renal disease [79-81]. The VEGF -460 T/C polymorphism and TGF-β1 polymorphisms, -800 G/A and -509 C/T, have been associat‐ ed with UTI and VUR in children [82]. VEGF -460 CC genotype was significantly more fre‐ quent in children with UTI and VUR than in controls [82]. Presence of VEGF -460 C allele increased basal VEGF promoter activity by 71% compared to the wild-type sequence [83]. Both UTI and VUR groups showed a significant increase in frequencies of TGF-β1 -800 GG and -509 CC genotypes in comparison to controls [82]. Cotton et al. [81] observed a correla‐ tion between -800 GA genotype and low TGF-β1 production *in vitro* suggesting a protective role against renal scarring. However, there was no correlation between TGF-β1 -509 geno‐

In the study of Yim et al. [82], the UTI group was subdivided into two subgroups according to presence of renal scars. Significantly increased frequency of TGF-β1 +869 CC genotype was found in the subgroup of patients positive for renal scarring [82]. A study of nephrop‐ athy resulted in an association of the +869 CC genotype with heavy proteinuria and a higher

Angiotensin II (Ang II), a powerful effector peptide of the renin-angiotensin system (RAS), is now considered a growth factor that plays active roles in all phenomena characteristic for renal tissue damage such as: proliferation of cells, accumulation of extracellular matrix and mononuclear cells' recruitment [19,85-87]. Since local kidney and interstitial fluid levels of Ang II are higher than the circulating [87], a blockade of Ang II actions may provide protec‐

Angiotensin I-converting enzyme (ACE), representing a target for ACE inhibitors (ACEI), is the key enzyme of RAS system. An insertion/deletion (I/D) polymorphism resulting from the presence/absence of a 287 bp *Alu* sequence has been identified in intron 16 of ACE gene [88-89]. This polymorphic variation in ACE gene correlates with levels of both circulating

*(TGF-β1)*

and renal scarring [78].

types and protein production [81].

score of mesangial cell proliferation [84].

*2.5.5. Angiotensin I-Converting Enzyme (ACE)*

tion against functional and structural kidney deterioration.

### *2.5.3. Toll-Like Receptors (TLRs)*

Toll-like receptors' (TLRs) genes are among the most commonly studied in association with UTIs. Toll-like receptors are critical sensors of microbial attack and effectors of the TLR-de‐ pendent innate defense, which enables host to eliminate pathogens [63-65]. TLRs are located on the cell surface or within organelles, like phagosomes, and are involved in detection of microbial ligands such as flagellin (TLR5), lipopolysaccharide (TLR4) and bacterial lipopep‐ tides (TLR1/2/6) [66].

Given that TLRs play a crucial role in the innate immune defense, their structures and func‐ tions are tightly regulated [67]. Numerous attempts have been made to identify TLRs struc‐ tural genes' variations, which might be related to diseases. Structural gene polymorphisms are relatively rare [68] and their contributions to human diseases still remain unclear.

Recently, TLR4 gene promoter region has been shown considerably more variable than pre‐ viously known [69]. It's been suggested that few genotype patterns might reflect selection for low-responder variants in the primary asymptomatic bacteriuria group, which might protect against severe UTI [69]. Previously, low TLR4 expression had mostly been attributed to tolerance and not to genetic variation affecting TLR4 expression. Recently, the first study proposing impact of TLR4 promoter genetic variants on TLR4 expression has been publish‐ ed [69]. The authors have shown that single and multiple SNPs mostly suppressed TLR4 promoter activity *in vitro*, especially in response to *E. coli* infection. They have also observed that TLR4 promoter sequence variations could influence clinical presentation of UTI.

Hawn et al. [70] suggested that TLR4 Asp299Gly polymorphism was associated with protec‐ tion from recurrent UTI, but not pyelonephritis. Furthermore, they showed that TLR5 +1174 C/T polymorphism was associated with an increased risk of recurrent UTI but not pyelo‐ nephritis, while a polymorphism in TLR1, +1805 G/T, was associated with protection from pyelonephritis in women [70]. The study that included children with recurrent UTI did not reveal a significant difference in Asp299Gly genotypes of TLR4 between children with UTI and control group [71].

Another study showed a relationship between the carrier status of HSPA1B (heat shock 70 kDa protein 1B) +1267 G and TLR4 +896 G alleles and development of recurrent UTI in childhood, independently on other urinary tract abnormalities [72]. Study in adults revealed that TLR4 +896 G allele had higher prevalence in UTI patients than in controls, and that TLR4 expression in monocytes was significantly lower in chronic UTI patients than in APN patients or healthy controls [73].

### *2.5.4. Vascular Endothelial Growth Factor (VEGF) and Transforming Growth Factor-beta1 (TGF-β1)*

Vascular endothelial growth factor (VEGF) is a potent mitogen that enhances angiogene‐ sis, microvascular permeability and proliferation of vascular endothelial cells [74]. Expres‐ sion of VEGF has been detected in glomerular podocytes, epithelial cells of distal tubules and collecting ducts of kidney. Neovascularization in scarred kidney tissue and significant increase of urine VEGF levels associated with increased severity of renal scarring have been reported [75-76]. Transforming growth factor-beta1 (TGF-β1) appears to be one of the key factors in process of tissue repair. It is involved in regulation of cell proliferation, differentiation, extracellular matrix production and immune response [77]. Studies sug‐ gested that TGF-β1 could be involved in pathogenesis of congenital obstructive uropathies and renal scarring [78].

Several polymorphisms in VEGF and TGF-β1 genes have been linked to overproduction of these proteins as well as predisposition to progressive renal disease [79-81]. The VEGF -460 T/C polymorphism and TGF-β1 polymorphisms, -800 G/A and -509 C/T, have been associat‐ ed with UTI and VUR in children [82]. VEGF -460 CC genotype was significantly more fre‐ quent in children with UTI and VUR than in controls [82]. Presence of VEGF -460 C allele increased basal VEGF promoter activity by 71% compared to the wild-type sequence [83]. Both UTI and VUR groups showed a significant increase in frequencies of TGF-β1 -800 GG and -509 CC genotypes in comparison to controls [82]. Cotton et al. [81] observed a correla‐ tion between -800 GA genotype and low TGF-β1 production *in vitro* suggesting a protective role against renal scarring. However, there was no correlation between TGF-β1 -509 geno‐ types and protein production [81].

In the study of Yim et al. [82], the UTI group was subdivided into two subgroups according to presence of renal scars. Significantly increased frequency of TGF-β1 +869 CC genotype was found in the subgroup of patients positive for renal scarring [82]. A study of nephrop‐ athy resulted in an association of the +869 CC genotype with heavy proteinuria and a higher score of mesangial cell proliferation [84].

### *2.5.5. Angiotensin I-Converting Enzyme (ACE)*

[62]. Still, A allele of ICAM-1 exon 4 (G/A) polymorphism had significantly lower frequency in patients who developed renal scars following UTI compared with the patients without scars. This suggested that the A allele might be a protective factor for renal scarring follow‐ ing UTI. It is possible that protective effect of the A allele on development of renal scars fol‐ lowing UTI may be a result of decreased binding of neutrophils and other inflammatory cells to ICAM-1, resulting in reduced leukocyte infiltration and reduced tissue damage [62].

Toll-like receptors' (TLRs) genes are among the most commonly studied in association with UTIs. Toll-like receptors are critical sensors of microbial attack and effectors of the TLR-de‐ pendent innate defense, which enables host to eliminate pathogens [63-65]. TLRs are located on the cell surface or within organelles, like phagosomes, and are involved in detection of microbial ligands such as flagellin (TLR5), lipopolysaccharide (TLR4) and bacterial lipopep‐

Given that TLRs play a crucial role in the innate immune defense, their structures and func‐ tions are tightly regulated [67]. Numerous attempts have been made to identify TLRs struc‐ tural genes' variations, which might be related to diseases. Structural gene polymorphisms

Recently, TLR4 gene promoter region has been shown considerably more variable than pre‐ viously known [69]. It's been suggested that few genotype patterns might reflect selection for low-responder variants in the primary asymptomatic bacteriuria group, which might protect against severe UTI [69]. Previously, low TLR4 expression had mostly been attributed to tolerance and not to genetic variation affecting TLR4 expression. Recently, the first study proposing impact of TLR4 promoter genetic variants on TLR4 expression has been publish‐ ed [69]. The authors have shown that single and multiple SNPs mostly suppressed TLR4 promoter activity *in vitro*, especially in response to *E. coli* infection. They have also observed

are relatively rare [68] and their contributions to human diseases still remain unclear.

that TLR4 promoter sequence variations could influence clinical presentation of UTI.

Hawn et al. [70] suggested that TLR4 Asp299Gly polymorphism was associated with protec‐ tion from recurrent UTI, but not pyelonephritis. Furthermore, they showed that TLR5 +1174 C/T polymorphism was associated with an increased risk of recurrent UTI but not pyelo‐ nephritis, while a polymorphism in TLR1, +1805 G/T, was associated with protection from pyelonephritis in women [70]. The study that included children with recurrent UTI did not reveal a significant difference in Asp299Gly genotypes of TLR4 between children with UTI

Another study showed a relationship between the carrier status of HSPA1B (heat shock 70 kDa protein 1B) +1267 G and TLR4 +896 G alleles and development of recurrent UTI in childhood, independently on other urinary tract abnormalities [72]. Study in adults revealed that TLR4 +896 G allele had higher prevalence in UTI patients than in controls, and that TLR4 expression in monocytes was significantly lower in chronic UTI patients than in APN

*2.5.3. Toll-Like Receptors (TLRs)*

144 Recent Advances in the Field of Urinary Tract Infections

tides (TLR1/2/6) [66].

and control group [71].

patients or healthy controls [73].

Angiotensin II (Ang II), a powerful effector peptide of the renin-angiotensin system (RAS), is now considered a growth factor that plays active roles in all phenomena characteristic for renal tissue damage such as: proliferation of cells, accumulation of extracellular matrix and mononuclear cells' recruitment [19,85-87]. Since local kidney and interstitial fluid levels of Ang II are higher than the circulating [87], a blockade of Ang II actions may provide protec‐ tion against functional and structural kidney deterioration.

Angiotensin I-converting enzyme (ACE), representing a target for ACE inhibitors (ACEI), is the key enzyme of RAS system. An insertion/deletion (I/D) polymorphism resulting from the presence/absence of a 287 bp *Alu* sequence has been identified in intron 16 of ACE gene [88-89]. This polymorphic variation in ACE gene correlates with levels of both circulating [88] and tissue-localized ACE [90] and DD genotype is found to be associated with the high‐ est ACE levels.

combining gene-environment and probably gene-gene interactions. To date, association with UTIs has been studied through a candidate gene approach. Among the candidate genes are those that code for soluble mediators, receptors and adhesion molecules included in reg‐ ulation of the host response upon UTI. There are two basic approaches in these genetic stud‐ ies. The first approach, which is less common, involves investigation of the candidate genes' effects on UTI susceptibility only (case-control study design). The second, more common ap‐ proach, is based on investigation of susceptibility to renal scarring as the most serious com‐ plication of UTI (case study design). The second approach has a greater clinical potential since the primary aim of such research is to identify the patients susceptible to progressive

Genetic Factors Underlying Susceptibility to Acute Pyelonephritis and Post-infectious Renal Damage

http://dx.doi.org/10.5772/52852

147

The field investigating genetic susceptibility to UTIs in humans started only a decade ago as we can see from the reviewed articles. The first evidence that susceptibility to APN as well as asymptomatic bacteriuria (ABU) could be inherited came only a few years ago [69,103]. As a consequence, limited number of studies has been performed. Most of these studies in‐ cluded children only, some of the rest included both children and adults, while others in‐ cluded adults only. This review is focused on studies in children. Besides the number of studies, another limitation of research in this field is the number of participants included in these studies. The majority of studies reviewed here had about 100 patients, only few had up to 250. In further analysis of study design we must notice dividing of patients' group in subgroups- those with APN and those with lower UTI, and further dividing of the APN subgroup according to presence/absence of renal scarring. These limitations must be mini‐ mized in order to improve the statistical power of studies. Nevertheless, the results so far support the hypothesis of genetic impact on susceptibility to UTI/APN and give a good rea‐

Among genes mostly investigated in susceptibility to UTIs are the cytokine family genes. Results suggest that IL-8 -251 A allele represents a risk factor for APN, after exclusion of pa‐ tients with VUR [18]. Single base changes in IL-8 receptor (CXCR1) gene, +217 C/G and +2608 G/C, are associated with a risk for APN in both children and adults [56]. RANTES/

The effects of TLRs genes' polymorphisms are hard to summarize due to a large number of genes and polymorphisms in this family and a small number of homogenous studies. It is proposed that genetic variants in TLR4 promoter influence TLR4 expression as well as clini‐ cal presentation of UTI [69]. Probably the most intereting, recent findings are the results of the group from Lundt University. Children with ABU express less TLR4 than APN prone children or controls but do not carry structural gene mutations explaining this phenotype. They recently defined the eight TLR4 promoter sequence variants, forming 19 haplotypes and 29 genotype patterns. The ABU-associated genotypes reduced TLR4 expression and the response to infection [56, 69]. Host susceptibility to common infections like UTI may thus be strongly influenced by single gene modifications affecting the innate immune response. For example, genetic alterations that reduce TLR4 function are associated with ABU, while poly‐ morphisms reducing IRF3 or CXCR1 expression are associated with acute pyelonephritis and an increased risk for renal scarring [104]. The TLR1 +1805 G/T polymorphism is shown

CCL5 -403 G allele is susceptibility factor for UTI, irrespectively of VUR [57].

kidney damage, which often results in end-stage renal disease.

son for further research.

Gene polymorphisms of the RAS, especially I/D polymorphism of ACE gene, were associat‐ ed with development of renal scarring in patients with congenital urological abnormalities [91-95]. It was concluded that the DD genotype could be a genetic susceptibility factor con‐ tributing to renal parenchymal damage. In our previous study [96], we found a difference in ACE I/D genotypes' distribution in patients with bladder dysfunction according to pres‐ ence/absence of renal scarring. Although the two groups of patients (with and without scar‐ ring) did not differ by conventional risk factors, significant increase of D allele frequency was present in patients with renal scarring [96]. Only few studies investigated effect of ACE I/D polymorphism on renal scarring following APN. In Korean children frequencies of ACE I/D genotypes and alleles were not different between renal scar-positive and scar-negative groups, irrespectively of VUR [97]. Another study in Greek population did not confirm a correlation between ACE DD genotype and renal scar formation in children with UTIs [98].

### *2.5.6. Meta-analysis of genetic susceptibility factors for renal scar formation following UTI*

Recently, meta-analysis of candidate gene polymorphisms as genetic susceptibility factors for renal scars' formation following UTI has been performed [99]. After systematic analysis of previously published data, the authors made strict inclusion criteria for this meta-analy‐ sis. From 523 original citations they identified only 18 articles that met the inclusion criteria. The results of meta-analysis showed that, according to recessive model of inheritance, ACE I/D polymorphism was a significant risk factor for renal scarring although with a high de‐ gree of between-study variability. According to dominant model, the T allele of TGF-β1 -509 C/T polymorphism was related to increased susceptibility for renal scarring, again with a high degree of between-study variability [99].

The risk for renal scarring occurred in ACE DD genotype carriers [99]. This genotype was correlated with increased expression of renal ACE [100] and thus with increased production of Ang II. Ang II is a mediator of progressive renal failure [101] and it may induce expres‐ sion of TGF-β1, which is involved in pathogenesis of renal scarring [78,102]. There was no correlation found between TGF-β1 -509 genotypes and TGF-β1 protein production [81]. Considering the results of current meta-analysis [99] and known effects of ACE I/D poly‐ morphism [100], it is of interest to study the effects of TGF-β1 -509 C/T gene polymorphism on gene expression and TGF-β1 levels in renal tissue of UTI (APN) patients having renal scars.

## **3. Conclusions**

The symptoms of urinary tract infections (UTIs) depend on localization of infection and magnitude of the host response to bacteria. There are many risk factors for UTIs such as gen‐ der, VUR, environmental and socio-economical factors and, as it is proposed and reviewed here, genetic risk factors. Hence, UTIs represent a classical example of multifactorial disease combining gene-environment and probably gene-gene interactions. To date, association with UTIs has been studied through a candidate gene approach. Among the candidate genes are those that code for soluble mediators, receptors and adhesion molecules included in reg‐ ulation of the host response upon UTI. There are two basic approaches in these genetic stud‐ ies. The first approach, which is less common, involves investigation of the candidate genes' effects on UTI susceptibility only (case-control study design). The second, more common ap‐ proach, is based on investigation of susceptibility to renal scarring as the most serious com‐ plication of UTI (case study design). The second approach has a greater clinical potential since the primary aim of such research is to identify the patients susceptible to progressive kidney damage, which often results in end-stage renal disease.

[88] and tissue-localized ACE [90] and DD genotype is found to be associated with the high‐

Gene polymorphisms of the RAS, especially I/D polymorphism of ACE gene, were associat‐ ed with development of renal scarring in patients with congenital urological abnormalities [91-95]. It was concluded that the DD genotype could be a genetic susceptibility factor con‐ tributing to renal parenchymal damage. In our previous study [96], we found a difference in ACE I/D genotypes' distribution in patients with bladder dysfunction according to pres‐ ence/absence of renal scarring. Although the two groups of patients (with and without scar‐ ring) did not differ by conventional risk factors, significant increase of D allele frequency was present in patients with renal scarring [96]. Only few studies investigated effect of ACE I/D polymorphism on renal scarring following APN. In Korean children frequencies of ACE I/D genotypes and alleles were not different between renal scar-positive and scar-negative groups, irrespectively of VUR [97]. Another study in Greek population did not confirm a correlation between ACE DD genotype and renal scar formation in children with UTIs [98].

*2.5.6. Meta-analysis of genetic susceptibility factors for renal scar formation following UTI*

high degree of between-study variability [99].

Recently, meta-analysis of candidate gene polymorphisms as genetic susceptibility factors for renal scars' formation following UTI has been performed [99]. After systematic analysis of previously published data, the authors made strict inclusion criteria for this meta-analy‐ sis. From 523 original citations they identified only 18 articles that met the inclusion criteria. The results of meta-analysis showed that, according to recessive model of inheritance, ACE I/D polymorphism was a significant risk factor for renal scarring although with a high de‐ gree of between-study variability. According to dominant model, the T allele of TGF-β1 -509 C/T polymorphism was related to increased susceptibility for renal scarring, again with a

The risk for renal scarring occurred in ACE DD genotype carriers [99]. This genotype was correlated with increased expression of renal ACE [100] and thus with increased production of Ang II. Ang II is a mediator of progressive renal failure [101] and it may induce expres‐ sion of TGF-β1, which is involved in pathogenesis of renal scarring [78,102]. There was no correlation found between TGF-β1 -509 genotypes and TGF-β1 protein production [81]. Considering the results of current meta-analysis [99] and known effects of ACE I/D poly‐ morphism [100], it is of interest to study the effects of TGF-β1 -509 C/T gene polymorphism on gene expression and TGF-β1 levels in renal tissue of UTI (APN) patients having renal

The symptoms of urinary tract infections (UTIs) depend on localization of infection and magnitude of the host response to bacteria. There are many risk factors for UTIs such as gen‐ der, VUR, environmental and socio-economical factors and, as it is proposed and reviewed here, genetic risk factors. Hence, UTIs represent a classical example of multifactorial disease

est ACE levels.

146 Recent Advances in the Field of Urinary Tract Infections

scars.

**3. Conclusions**

The field investigating genetic susceptibility to UTIs in humans started only a decade ago as we can see from the reviewed articles. The first evidence that susceptibility to APN as well as asymptomatic bacteriuria (ABU) could be inherited came only a few years ago [69,103]. As a consequence, limited number of studies has been performed. Most of these studies in‐ cluded children only, some of the rest included both children and adults, while others in‐ cluded adults only. This review is focused on studies in children. Besides the number of studies, another limitation of research in this field is the number of participants included in these studies. The majority of studies reviewed here had about 100 patients, only few had up to 250. In further analysis of study design we must notice dividing of patients' group in subgroups- those with APN and those with lower UTI, and further dividing of the APN subgroup according to presence/absence of renal scarring. These limitations must be mini‐ mized in order to improve the statistical power of studies. Nevertheless, the results so far support the hypothesis of genetic impact on susceptibility to UTI/APN and give a good rea‐ son for further research.

Among genes mostly investigated in susceptibility to UTIs are the cytokine family genes. Results suggest that IL-8 -251 A allele represents a risk factor for APN, after exclusion of pa‐ tients with VUR [18]. Single base changes in IL-8 receptor (CXCR1) gene, +217 C/G and +2608 G/C, are associated with a risk for APN in both children and adults [56]. RANTES/ CCL5 -403 G allele is susceptibility factor for UTI, irrespectively of VUR [57].

The effects of TLRs genes' polymorphisms are hard to summarize due to a large number of genes and polymorphisms in this family and a small number of homogenous studies. It is proposed that genetic variants in TLR4 promoter influence TLR4 expression as well as clini‐ cal presentation of UTI [69]. Probably the most intereting, recent findings are the results of the group from Lundt University. Children with ABU express less TLR4 than APN prone children or controls but do not carry structural gene mutations explaining this phenotype. They recently defined the eight TLR4 promoter sequence variants, forming 19 haplotypes and 29 genotype patterns. The ABU-associated genotypes reduced TLR4 expression and the response to infection [56, 69]. Host susceptibility to common infections like UTI may thus be strongly influenced by single gene modifications affecting the innate immune response. For example, genetic alterations that reduce TLR4 function are associated with ABU, while poly‐ morphisms reducing IRF3 or CXCR1 expression are associated with acute pyelonephritis and an increased risk for renal scarring [104]. The TLR1 +1805 G/T polymorphism is shown to protect against pyelonephritis in women [70]. It seems plausible to personalize the diag‐ nosis and therapy of APN and ABU in the future, by combining information on bacterial virulence and the host response

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plement 1975;13 1-26.

plement 1978;14 1-38.

To assess the genetic basis of renal scarring following UTI, many risk factors have to be ana‐ lyzed. As we discussed, the heterogeneity between studies is large. Therefore, we may only point out to results of recent meta-analysis. The meta-analysis reveals that ACE I/D and TGF-β1 -509 C/T polymorphisms are risk factors for development of renal scars following UTI [99].

The most recent results strongly suggest the innate immunity as possible key factor for ge‐ netic susceptibility to APN and renal scarring after infection, although on murine models in which certan genes are functionaly similar to humans. It was shown that acute pyelonephri‐ tis and renal scarring are caused by dysfunctional innate immunity in mCxcr2 heterozygous mice [105].

Up to date no genome-wide association study has been done to use new approach for dis‐ covering novel genetic susceptibility factors for UTIs on a large number of individuals. New approaches to risk assasment and therapy shoud be encouraged and it is time for UTI to combine molecular medicine and social and behaviral factors. It is certain that some children are protected from APN and others prone to severe and recurent infections. Also, some of the gene polymorphisms are differentialy presented in those groups of children., The final aim is to identify the patients genetically susceptible to renal scarring and to discover and enable novel strategies in management of recurrent UTIs in order to prevent further renal damage especially in susceptible children. Until then, the prophylactic antibiotic treatment to prevent recurrent UTIs remains of limited usefulness [106].
