Protective role for C3aR in experimental chronic pyelonephritis.

Emerging evidence suggest that C3aR plays important roles in homeostasis, host defense and disease. Although it is known that C3aR is protective in several models of acute bacterial infections, the role for C3aR in chronic infection is largely unknown. Here we show that C3aR is protective in experimental chronic pyelonephritis. Global C3aR deficient (C3ar-/- ) mice had higher renal bacterial load, more pronounced renal histological lesions, increased renal apoptotic cell accumulation, tissue inflammation and extracellular matrix deposition following renal infection with uropathogenic E. coli (UPEC) strain IH11128, compared to WT control mice. Myeloid C3aR deficient (Lyz2-C3ar-/- ) mice exhibited a similar disease phenotype to global C3ar-/- mice. Pharmacological treatment with a C3aR agonist reduced disease severity in experimental chronic pyelonephritis. Furthermore, macrophages of C3ar-/- mice exhibited impaired ability to phagocytose UPEC. Our data clearly demonstrate a protective role for C3aR against experimental chronic pyelonephritis, macrophage C3aR plays a major role in the protection, and C3aR is necessary for phagocytosis of UPEC by macrophages. Our observation that C3aR agonist curtailed the pathology suggests a therapeutic potential for activation of C3aR in chronic infection.

Human neutrophil peptides 1-3 protect the murine urinary tract from uropathogenic Escherichia coli challenge.

Antimicrobial peptides (AMPs) are critical to the protection of the urinary tract of humans and other animals from pathogenic microbial invasion. AMPs rapidly destroy pathogens by disrupting microbial membranes and/or augmenting or inhibiting the host immune system through a variety of signaling pathways. We have previously demonstrated that alpha-defensins 1-3 (DEFA1A3) are AMPs expressed in the epithelial cells of the human kidney collecting duct in response to uropathogens. We also demonstrated that DNA copy number variations in the DEFA1A3 locus are associated with UTI and pyelonephritis risk. Because DEFA1A3 is not expressed in mice, we utilized human DEFA1A3 gene transgenic mice (DEFA4/4) to further elucidate the biological relevance of this locus in the murine urinary tract. We demonstrate that the kidney transcriptional and translational expression pattern is similar in humans and the human gene transgenic mouse upon uropathogenic Escherichia coli (UPEC) stimulus in vitro and in vivo. We also demonstrate transgenic human DEFA4/4 gene mice are protected from UTI and pyelonephritis under various UPEC challenges. This study serves as the foundation to start the exploration of manipulating the DEFA1A3 locus and alpha-defensins 1-3 expression as a potential therapeutic target for UTIs and other infectious diseases.

TcpC inhibits neutrophil extracellular trap formation by enhancing ubiquitination mediated degradation of peptidylarginine deiminase 4.

TcpC is a multifunctional virulence factor of uropathogenic E. coli (UPEC). Neutrophil extracellular trap formation (NETosis) is a crucial anti-infection mechanism of neutrophils. Here we show the influence of TcpC on NETosis and related mechanisms. We show NETosis in the context of a pyelonephritis mouse model induced by TcpC-secreting wild-type E. coli CFT073 (CFT073wt) and LPS-induced in vitro NETosis with CFT073wt or recombinant TcpC (rTcpC)-treated neutrophils are inhibited. rTcpC enters neutrophils through caveolin-mediated endocytosis and inhibits LPS-induced production of ROS, proinflammatory cytokines and protein but not mRNA levels of peptidylarginine deiminase 4 (PAD4). rTcpC treatment enhances PAD4 ubiquitination and accumulation in proteasomes. Moreover, in vitro ubiquitination kit analyses show that TcpC is a PAD4-targetd E3 ubiquitin-ligase. These data suggest that TcpC inhibits NETosis primarily by serving as an E3 ligase that promotes degradation of PAD4. Our findings provide a novel mechanism underlying TcpC-mediated innate immune evasion.

TcpC inhibits toll-like receptor signaling pathway by serving as an E3 ubiquitin ligase that promotes degradation of myeloid differentiation factor 88.

TcpC is a virulence factor of uropathogenic E. coli (UPEC). It was found that TIR domain of TcpC impedes TLR signaling by direct association with MyD88. It has been a long-standing question whether bacterial pathogens have evolved a mechanism to manipulate MyD88 degradation by ubiquitin-proteasome pathway. Here, we show that TcpC is a MyD88-targeted E3 ubiquitin ligase. Kidney macrophages from mice with pyelonephritis induced by TcpC-secreting UPEC showed significantly decreased MyD88 protein levels. Recombinant TcpC (rTcpC) dose-dependently inhibited protein but not mRNA levels of MyD88 in macrophages. Moreover, rTcpC significantly promoted MyD88 ubiquitination and accumulation in proteasomes in macrophages. Cys12 and Trp106 in TcpC are crucial amino acids in maintaining its E3 activity. Therefore, TcpC blocks TLR signaling pathway by degradation of MyD88 through ubiquitin-proteasome system. Our findings provide not only a novel biochemical mechanism underlying TcpC-medicated immune evasion, but also the first example that bacterial pathogens inhibit MyD88-mediated signaling pathway by virulence factors that function as E3 ubiquitin ligase.

Kidney dendritic cells: fundamental biology and functional roles in health and disease.

Dendritic cells (DCs) are chief inducers of adaptive immunity and regulate local inflammatory responses across the body. Together with macrophages, the other main type of mononuclear phagocyte, DCs constitute the most abundant component of the intrarenal immune system. This network of functionally specialized immune cells constantly surveys its microenvironment for signs of injury or infection, which trigger the initiation of an immune response. In the healthy kidney, DCs coordinate effective immune responses, for example, by recruiting neutrophils for bacterial clearance in pyelonephritis. The pro-inflammatory actions of DCs can, however, also contribute to tissue damage in various types of acute kidney injury and chronic glomerulonephritis, as DCs recruit and activate effector T cells, which release toxic mediators and maintain tubulointerstitial immune infiltrates. These actions are counterbalanced by DC subsets that promote the activation and maintenance of regulatory T cells to support resolution of the immune response and allow kidney repair. Several studies have investigated the multiple roles for DCs in kidney homeostasis and disease, but it has become clear that current tools and subset markers are not sufficient to accurately distinguish DCs from macrophages. Multidimensional transcriptomic analysis studies promise to improve mononuclear phagocyte classification and provide a clearer view of DC ontogeny and subsets.

Preventive Effect of L-Carnitine on Scar Formation During Acute Pyelonephritis: A Randomized Placebo-Controlled Trial.

BACKGROUND: Urinary tract infection and pyelonephritis are clinical problems that frequently occur in children. Several factors are responsible for renal tissue injury, morbidity, and renal scarring after pyelonephritis. The aim of this study was to evaluate the preventive effect of L-carnitine on renal scarring in acute pyelonephritis. METHODS: A randomized double-blind clinical trial was conducted on 65 children aged 6 months to 10 years. Patients were randomized into 2 groups to receive 7-day treatment with only antibiotics without L-carnitine (control group; n = 32) and 7-day treatment with L-carnitine (case group; n = 33) during the acute phase of infection. Technetium-99m-labeled dimercaptosuccinic acid (DMSA) scintigraphy was performed for all children during the acute phase (in 2-7 days of hospitalization) and late phase. P-value less than 0.05 was statistically significant. RESULTS: We recruited 65 participants in the study: 32 children in control group and 33 children in case group. Three children in the control group and 2 children in the case group refused to perform the second DMSA scan. Overall, data analysis at the end of the study was done on 60 patients. Age distribution of girl patients with upper urinary infection was 6.5% in girl children aged between 6 months and 12 months, 41.1% aged between 1 and 5 years, 33.3% aged between 5 and 10 years, respectively. There was no significant difference between 2 groups in age and sex. There was no significant difference between 2 groups in systolic blood pressure, diastolic blood pressure, the lab data including urine white blood cells and serum erythrocyte sedimentation rate, and antibiogram profiles. Voiding dysfunction was detected in 10% of the participants. The baseline DMSA was not significantly difference in 2 groups, but worsening of kidney lesions was significantly higher in control group after 6 months (P = 0.012). CONCLUSION: Our study showed that L-carnitine significantly decreased renal scarring because of acute pyelonephritis.

Innate immunity and urinary tract infection.

Urinary tract infections are a severe public health problem. The emergence and spread of antimicrobial resistance among uropathogens threaten to further compromise the quality of life and health of people who develop acute and recurrent upper and lower urinary tract infections. The host defense mechanisms that prevent invasive bacterial infection are not entirely delineated. However, recent evidence suggests that versatile innate immune defenses play a key role in shielding the urinary tract from invading uropathogens. Over the last decade, considerable advances have been made in defining the innate mechanisms that maintain immune homeostasis in the kidney and urinary tract. When these innate defenses are compromised or dysregulated, pathogen susceptibility increases. The objective of this review is to provide an overview of how basic science discoveries are elucidating essential innate host defenses in the kidney and urinary tract. In doing so, we highlight how these findings may ultimately translate into the clinic as new biomarkers or therapies for urinary tract infection.

The pathogenesis and management of renal scarring in children with vesicoureteric reflux and pyelonephritis.

Bacterial urinary tract infections (UTIs) are one of the most common reasons for children to be admitted to hospital. Bacteria infect and invade the bladder (the lower urinary tract) and if the infection disseminates to the upper urinary tract, significant inflammation in the kidneys may arise. Inflammation is a double-edged sword: it is needed to clear bacteria, but if excessive, kidney tissue is injured. During injury, nephrons are destroyed and replaced with deposition of extracellular matrix and a renal scar. In this review, we explore the pathogenesis of UTIs and discuss the risk factors that result in dissemination of bladder infection to the kidneys. Three major risk factors predispose to kidney infections: the presence of vesicoureteric reflux, the presence of bladder and bowel dysfunction, and defects in the ability of the host immune response to clear bacteria. In this review, we will discuss these factors, their relationship to renal scarring, and potential treatments that might be beneficial to prevent renal scar formation in children.

Using single-cell technologies to map the human immune system - implications for nephrology.

Advances in single-cell technologies are transforming our understanding of cellular identity. For instance, the application of single-cell RNA sequencing and mass cytometry technologies to the study of immune cell populations in blood, secondary lymphoid organs and the renal tract is helping researchers to map the complex immune landscape within the kidney, define cell ontogeny and understand the relationship of kidney-resident immune cells with their circulating counterparts. These studies also provide insights into the interactions of immune cell populations with neighbouring epithelial and endothelial cells in health, and across a range of kidney diseases and cancer. These data have translational potential and will aid the identification of drug targets and enable better prediction of off-target effects. The application of single-cell technologies to clinical renal biopsy samples, or even cells within urine, will improve diagnostic accuracy and assist with personalized prognostication for patients with various kidney diseases. A comparison of immune cell types in peripheral blood and secondary lymphoid organs in healthy individuals and in patients with systemic autoimmune diseases that affect the kidney will also help to unravel the mechanisms that underpin the breakdown in self-tolerance and propagation of autoimmune responses. Together, these immune cell atlases have the potential to transform nephrology.

[Immune disturbances in acute pyelonephritis. Part I].

In the part I of a literature review of modern national and foreign publications dedicated to the problem of immune disorders in acute pyelonephritis, information on the role of innate and adaptive immunity in the pathogenesis of acute pyelonephritis is summarized and systematized. The relationship between the pathogens and immune system, in particular the ability of pathogens to overcome the protective immune mechanisms are discussed. The role of neutrophilic granulocytes as well as cellular immunity in the pathogenesis of acute pyelonephritis is shown. The changes in level of pro- and anti-inflammatory cytokines are separately highlighted. In conclusion, perspective directions for the studies in the area of immune disorders in acute pyelonephritis are proposed.

Sex differences in IL-17 contribute to chronicity in male versus female urinary tract infection.

Sex-based differences influence incidence and outcome of infectious disease. Women have a significantly greater incidence of urinary tract infection (UTI) than men, yet, conversely, male UTI is more persistent with greater associated morbidity. Mechanisms underlying these sex-based differences are unknown, in part due to a lack of experimental models. We optimized a model to transurethrally infect male mice and directly compared UTI in both sexes. Although both sexes were initially equally colonized by uropathogenic E. coli, only male and testosterone-treated female mice remained chronically infected for up to 4 weeks. Female mice had more robust innate responses, including higher IL-17 expression, and increased γδ T cells and group 3 innate lymphoid cells in the bladder following infection. Accordingly, neutralizing IL-17 abolished resolution in female mice, identifying a cytokine pathway necessary for bacterial clearance. Our findings support the concept that sex-based responses to UTI contribute to impaired innate immunity in males and provide a rationale for non-antibiotic-based immune targeting to improve the response to UTI.

Endocan as a marker of microvascular inflammation in kidney transplant recipients.

Endocan is a water-soluble proteoglycan exclusively secreted by vascular endothelium. Endocan levels may be elevated in kidney transplant recipients experiencing antibody-mediated rejection (ABMR), which is characterized by vascular inflammation in transplanted kidney. We evaluated the clinical relevance of endocan as markers of microvascular inflammation in patients who underwent kidney transplantation. Plasma and urinary endocan levels were measured in 203 kidney transplant recipients and were compared across different etiologies of allograft dysfunction and various pathologic scores. Both plasma and urinary endocan levels were significantly higher in patients with acute ABMR than those in patients with normal pathology, acute tubular necrosis (ATN), acute pyelonephritis, BK virus associated nephropathy (BKVN), and T-cell mediated rejection (TCMR). Patients with chronic active ABMR also exhibited significantly higher plasma and urinary endocan levels than patients with long-term graft survival. Scores of glomerulitis and peritubular capillaritis, which are typical features of microvascular inflammation, were significantly elevated in patients with higher plasma and/or urinary endocan levels. Furthermore, plasma and urinary endocan levels could effectively discriminate ABMR from ATN, BKVN, and TCMR. Finally, patients exhibiting high urinary and plasma endocan levels in acute ABMR group showed significantly worse renal survival. Altogether, plasma and urinary endocan levels may serve as potential markers of microvascular inflammation in kidney transplant recipients.

Emphysematous pyelonephritis in renal allograft related to antibody-mediated rejection: A case report and literature review.

Emphysematous pyelonephritis (EPN) is a rare condition which can rapidly progress to sepsis and multiple organ failure with high mortality. We experienced a rare case of EPN in a renal allograft related to antibody-mediated rejection (AMR). The patient received a deceased donor kidney transplant due to end-stage renal disease secondary to diabetes mellitus. Cross-match test was negative but she had remote history of anti-HLA-A2 antibody corresponding with the donor HLA. Surgery concluded without any major events. Anti-thymoglobulin was given perioperatively for induction. She was compliant with her immunosuppressive medications making urine of 2 L/d with serum creatinine of 1.9 mg/dL at discharge on post-operative day (POD) 6. She did well until POD 14 when she presented to the clinic with features of sepsis, pain over the transplanted kidney area and decline in urine volume with elevated serum creatinine. CT revealed extensive gas throughout the transplanted kidney. Renal scan revealed non-functional transplant kidney with no arterial flow. Based on these findings, a decision to perform transplant nephrectomy was made. At laparotomy, the kidney was completely necrotic. Pathology showed non-viable kidney parenchyma with the tubules lacking neutrophilic casts suggestive of ischemic necrosis. Donor-specific antibody (DSA) returned positive with high intensity anti-HLA-A2 antibody. This is the first case of early EPN in allograft considered to have occurred as a result of thrombotic ischemia secondary to AMR. This case suggests consideration of perioperative anti-B-cell and/or anti-plasma cell therapies for historical DSA and strict post-operative follow-up in immunologically high-risk recipients to detect early signs of rejection and avoid deleterious outcomes.

Cytotoxic Necrotizing Factor 1 Downregulates CD36 Transcription in Macrophages to Induce Inflammation During Acute Urinary Tract Infections.

Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) induce cystitis, pyelonephritis, and can cause kidney scarring and failure if inflammation is not under control. The detailed effects of cytotoxic necrotizing factor 1 (CNF1), the key UPEC toxin, on the pathogenicity of UPEC remain unclear. CD36 is an important scavenger receptor, responsible for pathogen and apoptotic cell clearance, and plays an essential role in host immune defense and homeostasis. Regulation of CD36 by bacterial toxins has not been reported. In this study, using a pyelonephritis mouse model, CNF1 was observed to contribute to increasing neutrophils and bacterial titers in infected bladder and kidney tissues, resulting in severe inflammation and tissue damage. CD36 expression in macrophages was found to be decreased by CNF1 in vitro and in vivo. We demonstrated that CNF1 attenuated CD36 transcription by decreasing expressions of its upstream transcription factors LXRß and C/EBPα and their recruitment to the CD36 promotor. In addition, Cdc42 was found to be involved in CNF1-mediated downregulation of LXRß. Our study investigated the pathogenesis of cnf1-carrying UPEC, which affected host innate immune defenses and homeostasis through regulation of CD36 in macrophages during acute UTIs.

Therapeutic effects of 5,2'-dibromo-2,4',5'-trihydroxydiphenylmethanone (LM49) in an experimental rat model of acute pyelonephritis by immunomodulation and anti-inflammation.

Antibiotics are still the primary therapy for acute pyelonephritis (APN); rarely, natural polyphenols are also used. LM49 is a novel marine bromophenol derivative displaying strong anti-inflammatory effects. We investigated the therapeutic efficacy of LM49 in an experimental rat model of APN. The model was established by injecting 0.5 mL Escherichia coli (ATCC 25922, 108 CFU/mL) into the urinary bladders of Sprague Dawley rats. This model showed increased kidney viscera indices and renal bacterial growth scores, as well as pathological changes in kidneys. We also performed a broth microdilution antimicrobial susceptibility test of the E. coli strain. Both norfloxacin and LM49 treatment reduced kidney viscera indices and decreased microbial counts in urine cultures and kidney homogenates in APN rats. However, in vitro experiments showed that LM49 did not directly inhibit bacteria. Rather, LM49 treatment inhibited inflammatory cell infiltration or abscess and improved tissue lesions in the renal medullary junction, renal pelvis, and calyx, and high-dose LM49 treatment inhibited the production of inflammatory interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) in serum. CD4+ T cells were higher in the LM49 groups treated with high, medium, and low doses than in the model group, whereas only high-dose LM49 treatment increased the number of CD8+ T cells, as compared with that in the model group. However, LM49 treatment did not influence hematological parameters. Our results show that LM49 therapeutic effects in an APN animal model may be achieved by regulating immune responses and inhibiting inflammatory mediators, suggesting it as a candidate APN treatment.

O-Acetylation is essential for functional antibody generation against Staphylococcus aureus capsular polysaccharide.

Staphylococcus aureus produces an antiphagocytic polysaccharide capsule to evade neutrophil-mediated killing. Many vaccines against encapsulated bacterial pathogens require generation of functional anti-capsular antibodies to mediate protection against infection and disease. Here it is shown that the generation of such antibody responses to S. aureus in vivo and in vitro requires the presence of O-acetyl modifications on the capsular polysaccharides. O-acetylation of S. aureus capsular polysaccharide therefore should be monitored carefully during vaccine development and production. This finding may provide additional insight into the previous failure of a S. aureus capsular polysaccharide conjugate vaccine.

[The nephroprotective potential of peloid therapy used for the rehabilitation of the patients presenting with chronic pyelonephritis]. / Nefroprotektivnyi potentsial peloidoterapii v reabilitatsii bol'nykh khronicheskim pielonefritom.

This review article presents the data on the mechanism of action of peloid therapy from the perspective of its defibrosing effect, the structure and functions of the extracellular matrix under the normal and pathological conditions. In addition, role of this treatment modality in the progression of tubulointerstitial fibrosis which determines the severity and prognosis of chronic pyelonephritis is considered. The researchers are currently carry out the extensive studies aimed at the search of the methods for the primary and secondary prevention of chronic pyelonephritis. A wide range of pharmacotherapeutic modalities are currently used for this purpose. Moreover the development of long-acting anti-relapse medications is currently underway along with the further improvement of high-tech reconstructive surgical methods for the intervention on the organs of the urinary system. At the same time, the nephroprotective potential of the natural physical factors, such as therapeutic muds, e.g. peloids, remains poorly explored even though their well apparent thermophysical properties, unique organic and mineral composition, and saturation with biologically active compounds are well known long ago. The systemic response to peloid therapy manifests itself as the changes in the metabolism of the intercellular matrix and collagen of the connective tissue associated with the alterations in the process of fibrogenesis and the development of tubulointerstitial disorders. The direct and indirect influence of peloids on the connective tissue is possible. The indirect effects are attributable to the peloid impact on the antioxidant status, immunity, hormonal regulation, and metabolic processes. These findings suggest the necessity of the relevant experimental and clinical studies for the evaluation of the influence of peloid therapy on the structure and metabolism of the connective tissue in the kidneys including dynamics of the markers of inflammation, proliferation and fibrogenesis, and the hormonal status of the patients suffering from chronic pyelonephritis based on the application of the modern technologies in accordance with the requirements of evidence-based medicine.

Role of antioxidant defence, renal toxicity markers and inflammatory cascade in disease progression of acute pyelonephritis in experimental rat model.

Urinary tract infections are the most common bacterial infections affecting millions of people each year worldwide. The animal model provides an excellent and suitable system for studying cystitis and pyelonephritis caused by Escherichia coli and other uropathogens. Using this established model, we evaluate the role of antioxidant defence system, renal injury markers, and blood parameters in the diseases progression during Escherichia coli infection on 0th day, 12h and 7th day. The antioxidant enzymes like SOD, CAT, GSH, GPx, GR levels were evaluated. The blood parameters like AST, ALT, ALP, Total protein, BUN, creatinine level were estimated in infection model. The relative organ weights, anti microbial status of kidney, CRP, WBC count were done for the evaluation of inflammatory response associated with the infection. The oxidative stress marker like MDA was also evaluated. Histopathological analysis of renal tissue provides direct vision to tissue damage. The antioxidant status of renal tissue was decreased during the 7th day of infection. Likewise, renal toxicity markers were significantly increased during bacterial infection. The inflammatory markers like CRP, WBC count and oxidative stress marker like MDA were significantly increased by the infection on 7th day. The histopathology of renal tissue also reveals the inflammation and tissue damage associated with acute pyelonephritis.

Inflammation drives renal scarring in experimental pyelonephritis.

Acquired renal scarring occurs in a subset of patients following febrile urinary tract infections and is associated with hypertension, proteinuria, and chronic kidney disease. Limited knowledge of histopathology, immune cell recruitment, and gene expression changes during pyelonephritis restricts the development of therapies to limit renal scarring. Here, we address this knowledge gap using immunocompetent mice with vesicoureteral reflux. Transurethral inoculation of uropathogenic Escherichia coli in C3H/HeOuJ mice leads to renal mucosal injury, tubulointerstitial nephritis, and cortical fibrosis. The extent of fibrosis correlates most significantly with inflammation at 7 and 28 days postinfection. The recruitment of neutrophils and inflammatory macrophages to infected kidneys is proportional to renal bacterial burden. Transcriptome analysis reveals molecular signatures associated with renal ischemia-reperfusion injury, immune cell chemotaxis, and leukocyte activation. This murine model recapitulates the cardinal histopathological features observed in humans with acquired renal scarring following pyelonephritis. The integration of histopathology, quantification of cellular immune influx, and unbiased transcriptional profiling begins to define potential mechanisms of tissue injury during pyelonephritis in the context of an intact immune response. The clear relationship between inflammatory cell recruitment and fibrosis supports the hypothesis that acquired renal scarring arises as a consequence of excessive host inflammation and suggests that immunomodulatory therapies should be investigated to reduce renal scarring in patients with pyelonephritis.