A Case of C3 Nephritis With a Rare Variant of the CFHR5 Gene.

C3 nephropathy is a renal disease caused by the aberrant activation of the alternative complement pathway. The long-term renal prognosis of C3 nephropathy is generally poor, and elucidation of its pathogenesis is clinically important. Genetic abnormalities within complement genes, encompassing autoantibodies targeting complement components and complement factor H-related proteins (CFHRs), can lead to abnormal complement activation. CFHR5 is one of the best-known responsible genes for C3 nephritis. Moreover, the renal prognosis can vary depending on the specific type of genetic mutation. Here, we report the case of a young woman with C3 nephritis and a heterozygous rare variant, P453S, in CFHR5. The P453S variant, characterized by amino acid substitutions with a low allele frequency, was located in the region essential for CFHR5 protein function, and multiple in silico analyses were done suggesting the pathological significance of P453S. The renal function of our patient remains stable. The P453S variant might contribute to the suppression of the CFHR5 protein's function, resulting in gradual complement progression and a favorable renal prognosis.

Treatment for patients with autosomal dominant polycystic kidney disease in the chronic kidney disease without kidney replacement therapy in real-world clinical practice: a descriptive retrospective cohort study.

BACKGROUND: In real-world clinical practice, treatments selected for patients with autosomal dominant polycystic kidney disease (ADPKD) in the chronic kidney disease (CKD) without kidney replacement therapy (KRT) have not been reported. This study investigated the oral treatments used in these patients and the changes in their use in recent years. Additionally, we studied the factors affecting tolvaptan dose reduction or discontinuation. METHODS: This retrospective cohort study was conducted using the medical records of 160 hospitals in Japan. Patients with ADPKD or polycystic kidney disease registered on the database between January 2014 and December 2020 were selected. Changes in prescription proportions over time were assessed using the Cochran-Armitage test. We focused on patients prescribed with >15 mg of tolvaptan daily to identify the factors related to its dose reduction or discontinuation and used Multivariate Cox regression analysis to evaluate them. RESULTS: Tolvaptan use in patients with ADPKD in the CKD without KRT stage has increased. As of 2020, 25% of patients were treated with tolvaptan. Overall, 3639 patients with ADPKD were enrolled in the database, of whom 156 were treated with tolvaptan. Of these, 64 patients (41%) reduced or discontinued tolvaptan during the observation period. The presence of an estimated glomerular filtration rate <60 mL/min/1.73 m2 at the beginning of the treatment was associated with a higher risk of tolvaptan dose reduction or discontinuation. CONCLUSION: The proportion of patients with ADPKD treated with high-dose tolvaptan is increasing. However, patients with late-stage CKD tended to reduce or discontinue tolvaptan.

Low serum concentrations of bevacizumab and nivolumab owing to excessive urinary loss in patients with proteinuria: a case series.

PURPOSE: Proteinuria can cause interindividual variability in the pharmacokinetics of therapeutic antibodies and may affect therapeutic efficacy. Here, we measured the serum and urinary concentrations of bevacizumab (BV) and nivolumab (NIVO) in patients with proteinuria and reported a case series of these patients. METHODS: Thirty-two cancer patients who received BV every 3 weeks or NIVO every 2 weeks between November 2020 and September 2021 at Kyoto University Hospital were enrolled in this study. The serum and urinary concentrations of BV and NIVO were measured using liquid chromatography-tandem mass spectrometry. RESULTS: We divided the BV-treated patients and the NIVO-treated patients into two groups based on the urine protein-creatinine ratio (UPCR): UPCR 1 g/g or higher (BV, n = 9; NIVO, n = 3) and UPCR less than 1 g/g (BV, n = 14; NIVO, n = 6). Serum concentrations of the therapeutic antibodies adjusted by their doses were significantly lower in both BV- and NIVO-treated patients with UPCR 1 g/g or higher compared to those with less than 1 g/g. In patients with UPCR 1 g/g or higher, urinary concentrations of the therapeutic antibodies adjusted by their serum concentrations and urinary creatinine concentrations tended to increase. CONCLUSION: This case-series study suggests a possibility of reduction in serum concentrations of BV and NIVO in patients with proteinuria by urinary excretion of these drugs.

Interpretable machine learning-based individual analysis of acute kidney injury in immune checkpoint inhibitor therapy.

BACKGROUND: Acute kidney injury (AKI) is a critical complication of immune checkpoint inhibitor therapy. Since the etiology of AKI in patients undergoing cancer therapy varies, clarifying underlying causes in individual cases is critical for optimal cancer treatment. Although it is essential to individually analyze immune checkpoint inhibitor-treated patients for underlying pathologies for each AKI episode, these analyses have not been realized. Herein, we aimed to individually clarify the underlying causes of AKI in immune checkpoint inhibitor-treated patients using a new clustering approach with Shapley Additive exPlanations (SHAP). METHODS: We developed a gradient-boosting decision tree-based machine learning model continuously predicting AKI within 7 days, using the medical records of 616 immune checkpoint inhibitor-treated patients. The temporal changes in individual predictive reasoning in AKI prediction models represented the key features contributing to each AKI prediction and clustered AKI patients based on the features with high predictive contribution quantified in time series by SHAP. We searched for common clinical backgrounds of AKI patients in each cluster, compared with annotation by three nephrologists. RESULTS: One hundred and twelve patients (18.2%) had at least one AKI episode. They were clustered per the key feature, and their SHAP value patterns, and the nephrologists assessed the clusters' clinical relevance. Receiver operating characteristic analysis revealed that the area under the curve was 0.880. Patients with AKI were categorized into four clusters with significant prognostic differences (p = 0.010). The leading causes of AKI for each cluster, such as hypovolemia, drug-related, and cancer cachexia, were all clinically interpretable, which conventional approaches cannot obtain. CONCLUSION: Our results suggest that the clustering method of individual predictive reasoning in machine learning models can be applied to infer clinically critical factors for developing each episode of AKI among patients with multiple AKI risk factors, such as immune checkpoint inhibitor-treated patients.

Effect of multidisciplinary care on diabetic kidney disease: a retrospective cohort study.

BACKGROUND: Diabetic kidney disease (DKD) is the most common disease among patients requiring dialysis for the first time in Japan. Multidisciplinary care (MDC) may prevent the progression of kidney failure. However, the effectiveness and timing of MDC to preserve kidney function in patients with DKD is unclear. Therefore, the aim of this study was to investigate whether MDC for patients with DKD affects the preservation of kidney function as well as the timing of MDC in clinical practice. METHODS: In this retrospective cohort study, we identified patients with type 2 diabetes mellitus and DKD from April 2012 to January 2020 using a nationwide Japanese healthcare record database. The fee code for medical guidance to prevent dialysis in patients with diabetes was used to distinguish between the MDC and non-MDC groups. The primary outcome was a 40% decline in the estimated glomerular filtration rate, and secondary outcomes were death, hospitalization, permanent dialysis, kidney failure with replacement therapy, and emergency temporary catheterization. Propensity score matching was performed, and Kaplan-Meier and multivariable Cox regression analyses were performed. RESULTS: Overall, 9,804 eligible patients met the inclusion criteria, of whom 5,614 were matched for the main analysis: 1,039 in the MDC group, and 4,575 in the non-MDC group. The primary outcome did not differ between the groups (hazard ratio: 1.18, [95% confidence interval: 0.99-1.41], P = 0.07). The groups also did not differ in terms of the secondary outcomes. Most patients with DKD received their first MDC guidance within 1 month of diagnosis, but most received guidance only once per year. CONCLUSIONS: Although we could not demonstrate the effectiveness of MDC on kidney function in patients with DKD, we clarified the characteristics of such patients assigned the fee code for medical guidance to prevent dialysis related to diabetes.

Improved Renal Function in Initial Treatment Improves Patient Survival, Renal Outcomes, and Glucocorticoid-Related Complications in IgG4-Related Kidney Disease in Japan.

Introduction: We aimed to clarify long-term renal prognosis, complications of malignancy, glucocorticoid (GC) toxicity, and mortality in immunoglobulin G4 (IgG4)-related kidney disease (IgG4-RKD). Methods: Reviewing the medical records of 95 patients with IgG4-RKD, we investigated clinical and pathologic features at baseline, the course of renal function, complications of malignancy, GC toxicity, and mortality during follow-up (median 71 months). The standardized incidence ratio (SIR) of malignancy and standardized mortality ratio were calculated using national statistics. Factors related to outcomes were assessed by Cox regression analyses. Results: At diagnosis, the median estimated glomerular infiltration rate (eGFR) was 46 ml/min per 1.73 m2. GC achieved initial improvement. Additional renal function recovery within 3-months of initial treatment occurred in patients with highly elevated serum IgG and IgG4 levels and hypocomplementemia. During follow-up, 68%, 17%, and 3% of the patients had chronic kidney disease (CKD), >30% eGFR decline, and end-stage renal disease (ESRD), respectively. Age-adjusted and sex-adjusted Cox regression analyses indicated that eGFR (hazard ratio [HR], 0.71) and extensive fibrosis (HR, 2.58) at treatment initiation had a significant impact on the time to CKD. Ten patients died, and the standardized mortality ratio was 0.94. The SIR of malignancy was 1.52. The incidence rate (IR) of severe infection was 1.80/100 person-years. Cox regression analyses showed that the best eGFR within 3 months after treatment initiation were associated with lower mortality (HR 0.67) and fewer severe infections (HR 0.63). Conclusion: This study suggests that more renal function recovery through early treatment initiation may improve patient survival, renal outcomes, and some GC-related complications in IgG4-RKD.

Proteinuria frequency and subsequent renal dysfunction in bevacizumab-treated patients: a single center, retrospective, observational study.

BACKGROUND: Proteinuria is a common adverse event observed during treatment with antivascular endothelial growth factor (VEGF) antibodies. Proteinuria is a risk factor for renal dysfunction and cardiovascular complications in patients with chronic kidney disease. However, the association between anti-VEGF antibody-induced proteinuria and renal dysfunction or cardiovascular complications remains unclear. METHODS: This retrospective, observational study included patients with cancer that were treated with bevacizumab (BV) at Kyoto University Hospital (Kyoto, Japan) between January 2006 and March 2018. Adverse event rates were compared between patients who developed qualitative ≥ 2 + proteinuria and those who developed < 1 + proteinuria. Adverse events were defined as renal dysfunction (i.e., ≥ 57% decrease in the eGFR, compared to the rate at the initial treatment) and hospitalization due to BV-associated cardiovascular complications and other adverse events. RESULTS: In total, 734 patients were included in this analysis. Renal dysfunction was more common in patients with ≥ 2 + proteinuria than in those with < 1 + proteinuria (13/199, 6.5% vs. 12/535, 2.3%). Seven of these 13 patients with ≥ 2 + proteinuria had transient reversible renal dysfunction. Only four (2.0%) patients had BV-associated renal dysfunction. Of the 734 patients, six patients, 16 patients, and 13 patients were hospitalized because of the adverse events of cardiovascular complications, thromboembolisms, and cerebrovascular complications, respectively. No relationship was observed between these adverse events and proteinuria. CONCLUSION: BV treatment-induced proteinuria was not associated with renal dysfunction or other adverse events. Continuing BV with caution is a possible treatment option, even after proteinuria develops, in patients with cancer and a limited prognosis.

Beyond 2D: A scalable and highly sensitive method for a comprehensive 3D analysis of kidney biopsy tissue.

The spatial organization of various cell populations is critical for the major physiological and pathological processes in the kidneys. Most evaluation of these processes typically comes from a conventional 2D tissue cross-section, visualizing a limited amount of cell organization. Therefore, the 2D analysis of kidney biopsy introduces selection bias. The 2D analysis potentially omits key pathological findings outside a 1- to 10-µm thin-sectioned area and lacks information on tissue organization, especially in a particular irregular structure such as crescentic glomeruli. In this study, we introduce an easy-to-use and scalable method for obtaining high-quality images of molecules of interest in a large tissue volume, enabling a comprehensive evaluation of the 3D organization and cellular composition of kidney tissue, especially the glomerular structure. We show that CUBIC and ScaleS clearing protocols could allow a 3D analysis of the kidney tissues in human and animal models of kidney disease. We also demonstrate that the paraffin-embedded human biopsy specimens previously examined via 2D evaluation could be applicable to 3D analysis, showing a potential utilization of this method in kidney biopsy tissue collected in the past. In summary, the 3D analysis of kidney biopsy provides a more comprehensive analysis and a minimized selection bias than 2D tissue analysis. Additionally, this method enables a quantitative evaluation of particular kidney structures and their surrounding tissues, with the potential utilization from basic science investigation to applied diagnostics in nephrology.

Pharmacokinetics of GS-441524, the active metabolite of remdesivir, in patients receiving continuous renal replacement therapy: A case series.

Remdesivir plays a key role in the treatment of coronavirus disease in 2019 (COVID-19). Haemodialysis is sometimes required for hospitalised patients with COVID-19, and patients undergoing haemodialysis are at an increased risk of severe COVID-19. In the present study, we report the serum concentrations of GS-441524, the active metabolite of remdesivir, in four patients undergoing continuous renal replacement therapy (CRRT). Patient 1, a male aged 70s, received a loading dose of 200 mg remdesivir on day 1, followed by 100 mg remdesivir from day 2, according to the package insert as in non-haemodialysis patients. The mean trough serum concentration of GS-441524 was 783.5 ng/mL, which was approximately 7-fold higher than the mean for patients with an estimated glomerular filtration rate (eGFR) ≥ 60 mL/min. Patients 2-4 received a loading dose of 200 mg remdesivir on day 1, followed by 100 mg once every 2 days from day 2. The mean trough serum concentrations of GS-441524 were 302.2 ng/mL, 585.8 ng/mL and 677.3 ng/mL, respectively. These were 3 to 6-fold higher than the mean for patients with eGFR ≥60 mL/min. The target doses for patients 1, 2, 3, and 4 receiving CRRT were 13.6 mL/kg/h, 6.0-12.5 mL/kg/h, 20.1 mL/kg/h, and 15.1 mL/kg/h, respectively, using a polysulphone membrane. The package insert dose of remdesivir is an overdose for CRRT patients with a target dose of 10-20 mL/kg/h. In low-intensity CRRT, as in Japan, it may be necessary to extend the interval between the doses of remdesivir.

Removal rate of 5-fluorouracil and its metabolites in patients on hemodialysis: a report of two cases of colorectal cancer patients with end-stage renal failure.

PURPOSE: Hyperammonemia is a serious adverse effect of 5-fluorouracil (5FU) administration. Hemodialysis can be used for its management, but detailed data on the concentrations and removal rate of 5FU and its metabolites during hemodialysis remain unclear. Here, we present two cases of hemodialysis patients with end-stage renal disease who received concurrent 5FU infusion. METHODS: Blood samples were collected from the hemodialysis circuit before and after the dialyzer during day 2 hemodialysis sessions, and from the internal shunt just before and after day 4 hemodialysis sessions. The serum levels of 5FU and its metabolites-α-fluoro-ß-alanine (FBAL) and monofluoroacetate (FA)-were measured using liquid chromatography-tandem mass spectrometry. RESULTS: Seven sets of blood samples were collected for case 1; the removal rates (mean ± standard deviation) of 5FU and FBAL by the dialyzer were 81.2 ± 23.2% and 96.1 ± 8.6%, respectively (p < 0.001). Three sets of blood samples were collected for case 2; the removal rates of 5FU and FBAL were 81.7 ± 3.9% and 94.8 ± 2.7%, respectively (p = 0.03). Twenty-seven sets of blood samples were collected for case 1; reductions in blood FBAL and FA levels were 49.3 ± 8.8% (p < 0.001) and 64.2 ± 30.3% (p = 0.04), respectively. Bayesian estimation yielded similar results. Three sets of blood samples were collected for case 2; reductions in the blood FBAL and FA levels were 49.9 ± 6.9% and 50.6 ± 33.0%, respectively. CONCLUSION: In this study, 5FU and its metabolite FBAL were directly removed from the blood by approximately 90% during hemodialysis, and the blood levels of FBAL and FA were reduced by approximately 50% with a single hemodialysis session.

Amino acid catabolite markers for early prognostication of pneumonia in patients with COVID-19.

Effective early-stage markers for predicting which patients are at risk of developing SARS-CoV-2 infection have not been fully investigated. Here, we performed comprehensive serum metabolome analysis of a total of 83 patients from two cohorts to determine that the acceleration of amino acid catabolism within 5 days from disease onset correlated with future disease severity. Increased levels of de-aminated amino acid catabolites involved in the de novo nucleotide synthesis pathway were identified as early prognostic markers that correlated with the initial viral load. We further employed mice models of SARS-CoV2-MA10 and influenza infection to demonstrate that such de-amination of amino acids and de novo synthesis of nucleotides were associated with the abnormal proliferation of airway and vascular tissue cells in the lungs during the early stages of infection. Consequently, it can be concluded that lung parenchymal tissue remodeling in the early stages of respiratory viral infections induces systemic metabolic remodeling and that the associated key amino acid catabolites are valid predictors for excessive inflammatory response in later disease stages.

Morphomics via next-generation electron microscopy.

The living body is composed of innumerable fine and complex structures. Although these structures have been studied in the past, a vast amount of information pertaining to them still remains unknown. When attempting to observe these ultra-structures, the use of electron microscopy (EM) has become indispensable. However, conventional EM settings are limited to a narrow tissue area, which can bias observations. Recently, new trends in EM research have emerged that provide coverage of far broader, nano-scale fields of view for two-dimensional wide areas and three-dimensional large volumes. Moreover, cutting-edge bioimage informatics conducted via deep learning has accelerated the quantification of complex morphological bioimages. Taken together, these technological and analytical advances have led to the comprehensive acquisition and quantification of cellular morphology, which now arises as a new omics science termed 'morphomics'.

Preemptive Intravenous Human Immunoglobulin G Suppresses BK Polyomavirus Replication and Spread of Infection In Vitro.

BK polyomavirus (BKPyV) infection causes various diseases in immunocompromised patients. Cells from human lung and kidney were infected with BKPyV and treated with commercially available intravenous immunoglobulin G (IVIG). Its effects on BKPyV replication and spread of infection were investigated, focusing on administration timing. IVIG treatment 3 hours after infection suppressed BKPyV replication assessed by real-time PCR and expression of the viral capsid protein 1 and large T-antigen. IVIG effectively reduced the number of BKPyV-infected cells 2 weeks after infection in an antibody titer-dependent manner. Virus release in the culture supernatants was not influenced by IVIG treatment 6-80 hours and 3-9 days after infection. Collectively, IVIG did not affect viral release from infected cells but inhibited the spread of infection by neutralizing the released virus and blocking the new infected cell formation, indicating greater efficacy in early localized infection. BKPyV replication resumed in IVIG-treated cultures at 7 days after IVIG removal. Early prophylactic administration of IVIG is expected to reduce the growth and spread of BKPyV infection, resulting in the reduction of infected cell lesions and prevention of BKPyV-associated diseases.

Manipulation of components of the renin angiotensin system in renal proximal tubules fails to alter atherosclerosis in hypercholesterolemic mice.

Background and objective: Whole body manipulation of the renin-angiotensin system (RAS) consistently exerts profound effects on experimental atherosclerosis development. A deficit in the literature has been a lack of attention to the effects of sex. Also, based on data with gene-deleted mice, the site of RAS activity that influences lesion formation is at an unknown distant location. Since angiotensin (AngII) concentrations are high in kidney and the major components of the RAS are present in renal proximal tubule cells (PTCs), this study evaluated the role of the RAS in PTCs in atherosclerosis development. Methods and results: Mice with an LDL receptor -/- background were fed Western diet to induce hypercholesterolemia and atherosclerosis. We first demonstrated the role of AT1 receptor antagonism on atherosclerosis in both sexes. Losartan, an AngII type 1 (AT1) receptor blocker, had greater blood pressure-lowering effects in females than males, but equivalent effects between sexes in reducing atherosclerotic lesion size. To determine the roles of renal AT1a receptor and angiotensin-converting enzyme (ACE), either component was deleted in PTCs after weaning using a tamoxifen-inducible Cre expressed under the control of an Ndrg1 promoter. Despite profound deletion of AT1a receptor or ACE in PTCs, the absence of either protein did not influence development of atherosclerosis in either sex. Conversely, mice expressing human angiotensinogen and renin in PTCs or expressing human angiotensinogen in liver but human renin in PTCs did not change atherosclerotic lesion size in male mice. Conclusion: Whole-body AT1R inhibition reduced atherosclerosis equivalently in both male and female mice; however, PTC-specific manipulation of the RAS components had no effects on hypercholesterolemia-induced atherosclerosis.

A Rare Case of Drug Reaction with Eosinophilia and Systemic Symptoms Syndrome Leading to Hemodialysis and Multiple Infections: A Case Report and Literature Review.

Drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome is a severe drug eruption that causes multiple organ damage. The renal impairment in these patients usually improves with immunosuppressants, but subsequent infections often develop. We herein report a rare case of DRESS syndrome leading to hemodialysis and multiple infections with Pneumocystis pneumonia, cytomegalovirus and Aspergillus despite the administration of low-dose prednisolone. We also present a literature review of cases requiring dialysis after DRESS syndrome. In patients with chronic kidney disease, it is important to be alert for not only the development of DRESS syndrome but also subsequent infections.

Matrix metalloproteinase-10 deficiency has protective effects against peritoneal inflammation and fibrosis via transcription factor NFκΒ pathway inhibition.

One of the most common causes of discontinued peritoneal dialysis is impaired peritoneal function. However, its molecular mechanisms remain unclear. Previously, by microarray analysis of mouse peritoneum, we showed that MMP (matrix metalloproteinase)-10 expression is significantly increased in mice with peritoneal fibrosis, but its function remains unknown. Chlorhexidine gluconate (CG) was intraperitoneally injected to wild-type and MMP-10 knockout mice to induce fibrosis to elucidate the role of MMP-10 on peritoneal injury. We also examined function of peritoneal macrophages and mesothelial cells obtained from wild-type and MMP-10 knockout mice, MMP-10-overexpressing macrophage-like RAW 264.7 cells and MeT-5A mesothelial cells, investigated MMP-10 expression on peritoneal biopsy specimens, and the association between serum proMMP-10 and peritoneal solute transfer rates determined by peritoneal equilibration test on patients. MMP-10 was expressed in cells positive for WT1, a mesothelial marker, and for MAC-2, a macrophage marker, in the thickened peritoneum of both mice and patients. Serum proMMP-10 levels were well correlated with peritoneal solute transfer rates. Peritoneal fibrosis, inflammation, and high peritoneal solute transfer rates induced by CG were all ameliorated by MMP-10 deletion, with reduction of CD31-positive vessels and VEGF-A-positive cells. Expression of inflammatory mediators and phosphorylation of NFκΒ subunit p65 at S536 were suppressed in both MMP-10 knockout macrophages and mesothelial cells in response to lipopolysaccharide stimulation. Overexpression of MMP-10 in RAW 264.7 and MeT-5A cells upregulated pro-inflammatory cytokines with phosphorylation of NFκΒ subunit p65. Thus, our results suggest that inflammatory responses induced by MMP-10 are mediated through the NFκΒ pathway, and that systemic deletion of MMP-10 ameliorates peritoneal inflammation and fibrosis caused by NFκΒ activation of peritoneal macrophages and mesothelial cells.

Chapter 2:indications and dosing of anticancer drug therapy in patients with impaired kidney function, from clinical practice guidelines for the management of kidney injury during anticancer drug therapy 2022.

This comprehensive review discusses the dosing strategies of cancer treatment drugs for patients with impaired kidney function, specifically those with chronic kidney disease (CKD), undergoing hemodialysis, and kidney transplant recipients. CKD patients often necessitate dose adjustments of chemotherapeutic agents, e.g., platinum preparations, pyrimidine fluoride antimetabolites, antifolate agents, molecularly targeted agents, and bone-modifying agents, to prevent drug accumulation and toxicity due to diminished renal clearance of the administered drugs and their metabolites. In hemodialysis patients, factors such as drug removal from hemodialysis and altered pharmacokinetics demand careful optimization of anticancer drug therapy, including dose adjustment and timing of administration. While free cisplatin is removed by hemodialysis, most of the tissue- and protein-bound cisplatin remains in the body and rebound cisplatin elevations are observed after hemodialysis. It is not recommended hemodialysis for drug removal, regardless of timing. Kidney transplant patients encounter unique challenges in cancer treatment, as maintaining the balance between reduction of immunosuppression, switching to mTOR inhibitors, and considering potential drug interactions with chemotherapeutic agents and immunosuppressants are crucial for preventing graft rejection and achieving optimal oncologic outcomes. The review underscores the importance of personalized, patient-centric approaches to anticancer drug therapy in patients with impaired kidney function.

Tertiary Lymphoid Tissues Are Microenvironments with Intensive Interactions between Immune Cells and Proinflammatory Parenchymal Cells in Aged Kidneys.

SIGNIFICANCE STATEMENT: Ectopic lymphoid structures called tertiary lymphoid tissues (TLTs) develop in several kidney diseases and are associated with poor renal prognosis. However, the mechanisms underlying TLT expansion and their effect on renal regeneration remain unclear. The authors report that single-nucleus RNA sequencing and validation experiments demonstrate that TLTs potentially amplify inflammation in aged injured kidneys. Lymphocytes within TLTs promote proinflammatory phenotypes of the surrounding proximal tubules and fibroblasts within the TLTs via proinflammatory cytokine production. These proinflammatory parenchymal cells then interact with immune cells by chemokine or cytokine production. Such cell-cell interactions potentially increase inflammation, expand TLTs, and exacerbate kidney injury. These findings help illuminate renal TLT pathology and suggest potential therapeutic targets. BACKGROUND: Ectopic lymphoid structures called tertiary lymphoid tissues (TLTs) develop in several kidney diseases and are associated with poor renal prognosis. However, the mechanisms that expand TLTs and underlie exacerbation of kidney injury remain unclear. METHODS: We performed single-nucleus RNA sequencing (snRNA-seq) on aged mouse kidneys with TLTs after ischemia-reperfusion injury. The results were validated using immunostaining, in situ hybridization of murine and human kidneys, and in vitro experiments. RESULTS: Using snRNA-seq, we identified proinflammatory and profibrotic Vcam1+ injured proximal tubules (PTs) with NF κ B and IFN-inducible transcription factor activation. VCAM1 + PTs were preferentially localized around TLTs and drove inflammation and fibrosis via the production of multiple chemokines or cytokines. Lymphocytes within TLTs expressed Tnf and Ifng at high levels, which synergistically upregulated VCAM1 and chemokine expression in cultured PT cells. In addition, snRNA-seq also identified proinflammatory and profibrotic fibroblasts, which resided within and outside TLTs, respectively. Proinflammatory fibroblasts exhibited STAT1 activation and various chemokine or cytokine production, including CXCL9/CXCL10 and B cell-activating factor, contributing to lymphocyte recruitment and survival. IFN γ upregulated the expression of these molecules in cultured fibroblasts in a STAT1-dependent manner, indicating potential bidirectional interactions between IFN γ -producing CXCR3 + T cells and proinflammatory fibroblasts within TLTs. The cellular and molecular components described in this study were confirmed in human kidneys with TLTs. CONCLUSIONS: These findings suggest that TLTs potentially amplify inflammation by providing a microenvironment that allows intense interactions between renal parenchymal and immune cells. These interactions may serve as novel therapeutic targets in kidney diseases involving TLT formation.