Renal Medulla Research Articles

Computational Modeling of Substrate-Dependent Mitochondrial Respiration and Bioenergetics in the Heart and Kidney Cortex and Outer Medulla.

Integrated computational modeling provides a mechanistic and quantitative framework to characterize alterations in mitochondrial respiration and bioenergetics in response to different metabolic substrates in-silico. These alterations play critical roles in the pathogenesis of diseases affecting metabolically active organs such as heart and kidney. Therefore, the present study aimed to develop and validate thermodynamically constrained integrated computational models of mitochondrial respiration and bioenergetics in the heart and kidney cortex and outer medulla (OM). The models incorporated the kinetics of major biochemical reactions and transport processes as well as regulatory mechanisms in the mitochondria of these tissues. Intrinsic model parameters such as Michaelis-Menten constants were fixed at previously estimated values, while extrinsic model parameters such as maximal reaction and transport velocities were estimated separately for each tissue. This was achieved by fitting the model solutions to our recently published respirometry data measured in isolated rat heart and kidney cortex and OM mitochondria utilizing various NADH- and FADH2-linked metabolic substrates. The models were validated by predicting additional respirometry and bioenergetics data, which were not used for estimating the extrinsic model parameters. The models were able to predict tissue-specific and substrate-dependent mitochondrial emergent metabolic system properties such as redox states, enzyme and transporter fluxes, metabolite concentrations, membrane potential, and respiratory control index under diverse physiological and pathological conditions. The models were also able to quantitatively characterize differential regulations of NADH- and FADH2-linked metabolic pathways, which contribute differently toward regulations of oxidative phosphorylation and ATP synthesis in the heart and kidney cortex and OM mitochondria.

Low-dose lipopolysaccharide inducing continuous and obvious increase in urinary protein in hyperglycemic rats and the underlying mechanism.

Proteinuria is an important hallmark of diabetic nephropathy models, however it takes a long time for the proteinuria and is not stable. Therefore, low-dose lipopolysaccharide (LPS) was investigated in this work to induce rapid and stable proteinuria in hyperglycemic rats and the underlying mechanism was studied. Hyperglycemia rats was induced by high-fat feeding combined with intraperitoneal injection of streptozotocin (STZ). After 21 days, the model rats received a subinjury dose of 0.8 mg / kg LPS intraperitoneally (i.p.). We detected related biochemical indexes at different time periods after LPS injection and examined the expression of glomerular podocyte-associated proteins. Simultaneously, we measured expression of inflammatory factors, apoptotic proteins and albumin (ALB) in the renal cortex and renal medulla, respectively. PAS (Periodic Acid Schiff) staining was used to observe renal pathology. After LPS injection, urinary microalbumin (umALB) increased significantly and lasted longer. The expression of Nephrin, Podocin and necroptosis factor kappa B (NF-κB) in rennal cortex and Interleukin 18 (IL-18), Caspase-1, NF-κB and ALB in the renal medulla was significantly changed. Pathologically, the glomerular basement membrane was observed to be significantly thickened, the renal tubules were dilated, and the epithelial cells fell off in a circle. LPS promoted the continuous increase in urinary microalbumin in hyperglycemic rats, which was related to the damage to the glomerular basement membrane and renal tubular epithelial cells and to the inflammatory reaction in the kidney involved in NF-κB signaling, and this pathological damage can help to establish a stable model of diabetic nephropathy with increased proteinuria.

Their last will and testament: dying immune cells protect the urinary system with extracellular DNA traps

Like most epithelial organs, the bladder and kidney can be directly accessed by bacteria evolved for invasion. Epithelia and immune cells attempt to stymie this infection with biophysical and chemical mechanisms. Goldspink etal. connected the Na+ gradient in the kidney medulla with an immune defense mounted by dead cells (namely, the explosive death of neutrophils and macrophages), resulting in extracellular DNA traps. The pathway from Na+ concentration to immune death is depicted.

Development of a multimodal kidney age prediction based on automatic segmentation CT image in patients with normal renal function.

For decades, description of renal function has been of interest to clinicians and researchers. Serum creatinine (Scr) and estimated glomerular filtration rate (eGFR) are familiar but also limited in many circumstances. Meanwhile, the physiological volumes of the kidney cortex and medulla are presumed to change with age and have been proven to change with decreasing kidney function. We recruited 182 patients with normal Scr levels between October 2021 and February 2022 in Peking Union Medical College Hospital (PUMCH) with demographic and clinical data. A 3D U-Net architecture is used for both cortex and medullary separation, and volume calculation. In addition, we included patients with the same inclusion criteria but with diabetes (PUMCH-DM test set) and diabetic nephropathy (PUMCH-DN test set) for internal comparison to verify the possible clinical value of "kidney age" (K-AGE). The PUMCH training set included 146 participants with a mean age of 47.5±7.4 years and mean Scr 63.5±12.3μmol/L. The PUMCH test set included 36 participants with a mean age of 47.1±7.9 years and mean Scr 66.9±13.0μmol/L. The multimodal method predicted K-AGE approximately close to the patient's actual physiological age, with 92% prediction within the 95% confidential interval. The mean absolute error increases with disease progression (PUMCH 5.00, PUMCH-DM 6.99, PUMCH-DN 9.32). We established a machine learning model for predicting the K-AGE, which offered the possibility of evaluating the whole kidney health in normal kidney aging and in disease conditions.

A better explanation of countercurrent multiplication in the formation of the corticopapillary osmotic gradient in the outer medulla.

Countercurrent multiplication (CCM) is widely accepted as the mechanism for the generation of the corticopapillary osmotic gradient in the outer medulla of mammalian kidneys. However, several issues in the literature cause the current explanations of CCM to be inefficient and incomplete. As a result, it is challenging to clearly explain CCM in physiology education. The goal of this article is to share a modified version of CCM with more understandable explanation in the hopes of motivating peer discussion, further improvement, and future research. To reach this goal, the logical processes leading to CCM are first analyzed, which results in a set of formulas that serve as the principles governing CCM. Next, the cessation of CCM is addressed to provide a complete picture of the modified version of CCM. Throughout these two steps, the issues mentioned above are identified and addressed so that how the modified version of CCM eliminates these issues becomes clear. The formulas mentioned above are provided in the Tables S1, S2, and S3 (all Supplemental material is available in the Supplemental Excel File at https://doi.org/10.6084/m9.figshare.23515614) to explain how the interstitial and intrathick ascending limb osmotic concentration (OC) values used in the figures in this article are simulated and how alternative OC values can be generated from Tables S1 and S2 to illustrate CCM.NEW & NOTEWORTHY Countercurrent multiplication is widely accepted as the mechanism for the generation of the corticopapillary osmotic gradient in the outer medulla of mammalian kidneys, but the current explanations of it in textbooks and the literature are inefficient and incomplete, leading to confusion for students. This article shares a modified version of countercurrent multiplication with more understandable explanation as a way of motivating peer discussion, further improvement, and future research.

2D Shear wave elastography for evaluation of renal cortex and medulla: stiffness values in healthy children.

To evaluate renal cortical and medullary stiffness using 2D Shear-wave elastography (SWE) in healthy children. In this IRB approved prospective study, we measured the stiffness of cortex and medulla of children (4months-17years) at the upper pole, mid pole, and lower pole in bilateral kidneys. The median (IQR) values of renal cortex in <1year age group was 8.7 (5.7-11.7) kPa for right and 8.7 (4.2-14.1) kPa for the left side. For 1-5years age group, it was 7.3 (5.3-10) kPa for the right and 8.9 (6-12.3) kPa for the left side. For >5years, it was 7.4 (5.3-11.2) kPa for the right and 9.6 (6.2-12.7) kPa for the left side. The median (IQR) values of renal medulla in <1year age group was 7.1 (5.1-12.5) kPa for right and 6.8 (4-10.6) kPa for the left side. For 1-5years age group, it was 7.2 (4.9-9.7) kPa for the right and 6.9 (5.6-9.9) kPa for the left side. For >5years, it was 6.8 (5.1-9.6) kPa for the right and 7 (5-10.2) kPa for the left side. The differences in the elasticity values amongst these groups were statistically insignificant (p>0.05). There was a significant correlation between SWE values of cortex and medulla of right kidney (ρ=0.64) and of left kidney (ρ=0.61), respectively. SWE values of renal cortical and medullary stiffness in healthy children do not correlate with age. There is a significant correlation between SWE values of cortex and medulla of the kidneys in healthy children.

Effects of ROS pathway inhibitors and NADH and FADH2 linked substrates on mitochondrial bioenergetics and ROS emission in the heart and kidney cortex and outer medulla

Mitochondria are major sources of reactive oxygen species (ROS), which play important roles in both physiological and pathological processes. However, the specific contributions of different ROS production and scavenging components in the mitochondria of metabolically active tissues such as heart and kidney cortex and outer medulla (OM) are not well understood. Therefore, the goal of this study was to determine contributions of different ROS production and scavenging components and provide detailed comparisons of mitochondrial respiration, bioenergetics, ROS emission between the heart and kidney cortex and OM using tissues obtained from the same Sprague-Dawley rat under identical conditions and perturbations. Specifically, data were obtained using both NADH-linked substrate pyruvate + malate and FADH2-linked substrate succinate followed by additions of inhibitors of different components of the electron transport chain (ETC) and oxidative phosphorylation (OxPhos) and other ROS production and scavenging systems. Currently, there is limited data available for the mitochondria of kidney cortex and OM, the two major energy-consuming tissues in the body only next to the heart, and scarce quantitative information on the interplay between mitochondrial ROS production and scavenging systems in the three tissues. The findings from this study demonstrate significant differences in mitochondrial respiratory and bioenergetic functions and ROS emission among the three tissues. The results quantify the rates of ROS production from different complexes of the ETC, identify the complexes responsible for variations in mitochondrial membrane depolarization and regulations of ROS production, and quantify the contributions of ROS scavenging enzymes towards overall mitochondrial ROS emission. These findings advance our fundamental knowledge of tissue-specific and substrate-dependent mitochondrial respiratory and bioenergetic functions and ROS emission. This is important given the critical role that excess ROS production, oxidative stress, and mitochondrial dysfunction in the heart and kidney cortex and OM play in the pathogenesis of cardiovascular and renal diseases, including salt-sensitive hypertension.

Pathological Characterization of Spontaneous AA Amyloidosis in Microminipigs.

The minipig has been used as a non-rodent species in nonclinical toxicology studies, but little is known about amyloid A (AA) amyloidosis in this species. Among domestic pigs, reports of AA amyloidosis have been limited to animals with mutations in the N-terminal residue of serum AA (SAA), which is thought to be a primary etiological factor. In this study, we histologically examined 26 microminipigs aged 0.6 to 10 years and observed amyloid deposition in one 0.6-year-old and six 5-year-old or older microminipigs. The amyloid deposits were identified as AA based on mass spectrometry (MS) and immunohistochemistry (IHC). The 0.6-year-old microminipig showed severe deposition in the renal cortex and spleen, whereas 5-year-old or older animals had severe deposition in the renal medulla. MS and IHC detected serum amyloid P-component (SAP) in amyloid deposits in older animals but not in a 0.6-year-old animals. Based on the proteomic analysis and gene sequencing, amino acid mutations of SAA, previously found in domestic pigs, were not involved in the pathogenesis of AA amyloidosis in microminipigs. This study demonstrates that microminipigs with wild-type SAA develop AA amyloidosis and presents the possibility that differences in the environment surrounding amyloid, such as SAP, may influence differences in the pathological phenotype.

Reaction of Kidney Medullary Laminin to Prolonged Dehydration of Rats

Laminin is the main water-soluble protein in the basement membrane of epithelial tissue. The content of laminin in the renal parenchyma was studied in rats that were on an alternative water diet for 60 h with excessive water intake or a complete lack of drinking water in the food. It has been shown that the osmotic concentration of urine, which develops as a result of water deprivation, is accompanied by a quantitative change in the composition of laminin. It has been established that the amount of laminin beta chain increases in the kidney medulla of dehydrated rats. This effect is observed only in WAG rats with normally expressed vasopressin gene and is absent in mutant Brattleboro rats unable to synthesize the hormone. The increase in the level of beta-laminin also does not extend to the cortical substance. Based on the original and new data, it is assumed that laminin, as a key regulatory element in the basement membrane of the tubular epithelium, participates in the adaptive reaction of the concentrating system of the kidney to conditions of prolonged dehydration. Hyperhydration does not affect the level of laminin in the renal parenchyma.

Early assessment of acute kidney injury in severe acute pancreatitis with multimodal DWI: an animal model.

To evaluate the feasibility of multimodal diffusion-weighted imaging (DWI) for detecting the occurrence and severity of acute kidney injury (AKI) caused by severe acute pancreatitis (SAP) in rats. SAP was induced in thirty rats by the retrograde injection of 5.0% sodium taurocholate through the biliopancreatic duct. Six rats underwent MRI of the kidneys 24h before and 2, 4, 6, and 8h after this AKI model was generated. Conventional and functional MRI sequences were used, including intravoxel incoherent motion imaging (IVIM), diffusion tensor imaging (DTI), and diffusion kurtosis imaging (DTI). The main DWI parameters and histological results were analyzed. The fast apparent diffusion coefficient (ADC) of the renal cortex was significantly reduced at 2h, as was the fractional anisotropy (FA) value of the renal cortex on DTI. The mean kurtosis (MK) values for the renal cortex and medulla gradually increased after model generation. The renal histopathological score was negatively correlated with the medullary slow ADC, fast ADC, and perfusion scores for both the renal cortex and medulla, as were the ADC and FA values of the renal medulla in DTI, whereas the MK values of the cortex and medulla were positively correlated (r = 0.733, 0.812). Thus, the cortical fast ADC, medullary MK, FADTI, and slow ADC were optimal parameters for diagnosing AKI. Of these parameters, cortical fast ADC had the highest diagnostic efficacy (AUC = 0.950). The fast ADC of the renal cortex is the core indicator of early AKI, and the medullary MK value might serve as a sensitive biomarker for grading renal injury in SAP rats. The multimodal parameters of renal IVIM, DTI, and DKI are potential beneficial for the early diagnosis and severity grading of renal injury in SAP patients. • The multimodal parameters of renal DWI, including IVIM, DTI, and DKI, may be valuable for the noninvasive detection of early AKI and the severity grading of renal injury in SAP rats. • Cortical fast ADC, medullary MK, FA, and slow ADC are optimal parameters for early diagnosis of AKI, and cortical fast ADC has the highest diagnostic efficacy. • Medullary fast ADC, MK, and FA as well as cortical MK are useful for predicting the severity grade of AKI, and the renal medullary MK value exhibits the strongest correlation with pathological scores.

Metabolic Responses of Normal Rat Kidneys to a High Salt Intake.

In this study, novel methods were developed, which allowed continuous (24/7) measurement of arterial blood pressure and renal blood flow in freely moving rats and the intermittent collection of arterial and renal venous blood to estimate kidney metabolic fluxes of O2 and metabolites. Specifically, the study determined the effects of a high salt (HS; 4.0% NaCl) diet upon whole kidney O2 consumption and arterial and renal venous plasma metabolomic profiles of normal Sprague-Dawley rats. A separate group of rats was studied to determine changes in the cortex and outer medulla tissue metabolomic and mRNAseq profiles before and following the switch from a 0.4% to 4.0% NaCl diet. In addition, targeted mRNA expression analysis of cortical segments was performed. Significant changes in the metabolomic and transcriptomic profiles occurred with feeding of the HS diet. A progressive increase of kidney O2 consumption was found despite a reduction in expression of most of the mRNA encoding enzymes of TCA cycle. A novel finding was the increased expression of glycolysis-related genes in Cx and isolated proximal tubular segments in response to an HS diet, consistent with increased release of pyruvate and lactate from the kidney to the renal venous blood. Data suggests that aerobic glycolysis (eg, Warburg effect) may contribute to energy production under these circumstances. The study provides evidence that kidney metabolism responds to an HS diet enabling enhanced energy production while protecting from oxidative stress and injury. Metabolomic and transcriptomic analysis of kidneys of Sprague-Dawley rats fed a high salt diet.

Evaluation of the parenchymal distribution of renal steatosis in chronic kidney disease using chemical shift magnetic resonance imaging.

Renal steatosis is an abnormal accumulation of fat in the kidney and may cause chronic kidney disease (CKD) or CKD progression. This pilot study aimed to evaluate the quantitative measurability of the parenchymal distribution of lipid deposition in the renal cortex and medulla using chemical shift magnetic resonance imaging (MRI) and investigate its relationship with clinical stages in CKD patients. The study groups included CKD patients with diabetes (CKD-d) (n = 42), CKD patients without diabetes (CKD-nd) (n = 31) and control subjects (n = 15), all of whom underwent a 1.5T MRI of the abdomen using the Dixon two-point method. The fat fraction (FF) values in the renal cortex and medulla were calculated from measurements made on Dixon sequences, and then compared between the groups. The cortical FF value was higher than the medullary FF value in control (0.057 (0.053-0.064) compared to 0.045 (0.039-0.052)), CKD-nd (0.066 (0.059-0.071) compared to 0.063 (0.054-0.071)), and CKD-d (0.081 (0.071-0.091) compared to 0.069 (0.061-0.077)) groups (all p < 0.001). The CKD-d group cortical FF values were higher than those of the CKD-nd group (p < 0.001). The FF values began increasing at CKD stages 2 and 3, and reached statistical significance at stages 4 and 5 in CKD patients (p < 0.001). Renal parenchymal lipid deposition can be quantified separately in the cortex and medulla using chemical shift MRI. Fat accumulation occurred in cortical and medullary parenchyma in CKD patients, though predominantly in the cortex. This accumulation increased proportionally with the disease stage.

Claudin-19 localizes to the thick ascending limb where its expression is required for junctional claudin-16 localization.

The kidney is critical for mineral homeostasis. Calcium and magnesium reabsorption in the renal thick ascending limb (TAL) involves claudin-16 (CLDN16) and claudin-19 (CLDN19) and pathogenic variants in either gene lead to familial hypomagnesemia with hypercalciuria and nephrocalcinosis (FHHNC) with severe calcium and magnesium wasting. While both CLDN16 and CLDN19 localize to the TAL, varying expression patterns in the renal tubule have been reported using different antibodies. We, therefore, studied the localization of CLDN19 in the kidneys of wild-type and Cldn19-deleted mice using three anti-CLDN19 antibodies and examined the role of Cldn19 deletion on CLDN16 and CLDN10 localization. We find that CLDN19 localizes to basolateral membrane domains of the medullary and cortical TAL but only to the tight junction of TALs in the outer stripe of outer medulla and cortex, where it colocalizes with CLDN16. Furthermore, in TALs from Cldn19-deleted mice, CLDN16 is expressed in basolateral membrane domains but not at the tight junction. In contrast, Cldn19 ablation does not change CLDN10 localization. These findings directly implicate CLDN19 in regulating permeability in the TAL by allowing junctional insertion of CLDN16 and may explain the shared renal phenotypic characteristics in FHHNC patients.

NBCe1-B/C-knockout mice exhibit an impaired respiratory response and an enhanced renal response to metabolic acidosis.

The sodium-bicarbonate cotransporter (NBCe1) has three primary variants: NBCe1-A, -B and -C. NBCe1-A is expressed in renal proximal tubules in the cortical labyrinth, where it is essential for reclaiming filtered bicarbonate, such that NBCe1-A knockout mice are congenitally acidemic. NBCe1-B and -C variants are expressed in chemosensitive regions of the brainstem, while NBCe1-B is also expressed in renal proximal tubules located in the outer medulla. Although mice lacking NBCe1-B/C (KOb/c) exhibit a normal plasma pH at baseline, the distribution of NBCe1-B/C indicates that these variants could play a role in both the rapid respiratory and slower renal responses to metabolic acidosis (MAc). Therefore, in this study we used an integrative physiologic approach to investigate the response of KOb/c mice to MAc. By means of unanesthetized whole-body plethysmography and blood-gas analysis, we demonstrate that the respiratory response to MAc (increase in minute volume, decrease in pCO2) is impaired in KOb/c mice leading to a greater severity of acidemia after 1day of MAc. Despite this respiratory impairment, the recovery of plasma pH after 3-days of MAc remained intact in KOb/c mice. Using data gathered from mice housed in metabolic cages we demonstrate a greater elevation of renal ammonium excretion and greater downregulation of the ammonia recycling enzyme glutamine synthetase in KOb/c mice on day 2 of MAc, consistent with greater renal acid-excretion. We conclude that KOb/c mice are ultimately able to defend plasma pH during MAc, but that the integrated response is disturbed such that the burden of work shifts from the respiratory system to the kidneys, delaying the recovery of pH.

Based on functional and histopathological correlations: is diffusion kurtosis imaging valuable for noninvasive assessment of renal damage in early-stage of chronic kidney disease?

To evaluate the potential of 3T magnetic resonance diffusion kurtosis imaging (DKI) in assessing the renal damage in early-stage of chronic kidney disease (CKD) patients with normal or slightly changed functional index, using histopathology as reference standard. 49 CKD patients and 18 healthy volunteers were recruited in this study. CKD patients were divided into two groups based on estimated glomerular filtration rate (eGFR): Study group I (eGFR ≥ 90ml/min/1.73m2 [n = 20]) and Study group II (eGFR < 90ml/min/1.73m2 [n = 29]). DKI was performed in all participants. The DKI parameters (mean kurtosis [MK], mean diffusivity [MD], fractional anisotropy [FA]) of renal cortex and medulla were measured. The differences of parenchymal MD, MK and FA values among the different groups were compared. The correlations between DKI parameters and clinicopathological characteristics were assessed. Diagnostic performance of DKI to assess renal damage in early-stage of CKD was analyzed. The cortex MD and MK showed significant difference among three groups (P < 0.05): trend of cortex MD: Study group II < Study group I < control group; trend of cortex MK: control group < Study group I < Study group II. The cortex MD and MK and medulla FA were correlated with eGFR and Interstitial fibrosis/Tubular atrophy score (0.3 < r < 0.5). Cortex MD and MK yielded an AUC of 0.752 for differentiating healthy volunteers from CKD patients with eGFR ≥ 90ml/min/1.73 m2. DKI shows potential in non-invasive and multi-parameter quantitative assessment of renal damage in early-stage of CKD patients and provide additional information for changes in renal function and histopathology.

Prediction of potato dry matter content by FT-NIR spectroscopy: Impact of tuber tissue on model performance

Potato, an important cash crop, is critically affected by supply chain losses that require attention of both research and industry to ensure sustainable production (SDG 12.3). Within potato supply chain management, dry matter (DM) determination is a key factor affecting the harvest quality, storability, food losses, designated user and tuber prices. Even though rapid methods like near-infrared spectroscopy (NIR) enable operators to carry out fast, reliable in situ measurements, the prediction results are often unsatisfactory due to inherent variability of tuber's composition. Thus, this study aims to evaluate Fourier Transformed-NIR based DM prediction in different potato tissues: i) periderm, ii) cortex, iii) outer medulla and iv) inner medulla of 12 different cultivars. The DM prediction models developed using standard chemometrics approaches (i.e., partial least squares, PLS; and interval-PLS, iPLS) showed that features selection by iPLS allowed for better performances. Among the evaluated tissues, the inner medulla had more precise and reliable measurements with RMSE and R2 prediction values of 0.93% and 0.90, respectively. This information can be used to develop more advanced systems inclusive of portable tools and advanced technologies for in situ evaluation to support operators and ensure better pre- and post-harvest management of potatoes.

#6073 MAGNETIC RESONANCE RADIOMIC ANALYSIS FOR THE PREDICTION OF GRAFT FAILURE IN PATIENTS WITH KIDNEY TRANSPLANTATION

Abstract Background and Aims Prediction of future graft failure with non-invasive investigations is a relevant objective for transplant clinicians. We previously demonstrated a promising discrimination capacity for interstitial fibrosis / tubular atrophy (IFTA) &amp;gt;50% in kidney biopsy of a machine learning (ML) based magnetic resonance imaging (MRI) radiomic algorithm. Aim of the present study is to evaluate accuracy of MRI radiomic-based ML algorithms in predicting future graft failure. Method Single-center retrospective observational cohort study on a previously characterized cohort of kidney transplant recipients who underwent graft biopsy and graft MRI imaging within 6 months from biopsy, both on clinical indication, at the “Azienda Ospedaliero-Universitaria di Modena”, Italy, from 1/1/2012 to 1/3/2021. The primary outcome was to identify the best combination of MRI radiomic features for the prediction of graft failure during follow-up. Segmentation of renal parenchyma, cortex and medulla on MRI sequences was performed using the 3DSlicer software. Radiomic features were then extracted using an in-house software based on pyradiomics applying Wavelet and Gaussian filters. LASSO algorithm was employed to select correlated features with outcome and summarize them in a radiomic signature. Using radiomic signature alone and then merged with meaningful clinical data we trained ML-algorithms using 70% of cases for training/validation, with a 10-fold internal cross-validation, and 30% for model testing. Model performance was assessed using AUC with a 95% confidence interval (CI). Results Seventy coupled tests (63 patients) were included and randomly subdivided into a training/validation cohort (n = 50) and a test cohort (n = 20). Median follow-up was 24.73 months (interquartile range 13.64-46.57). Radiomic model had an AUC of 0.88 (95%_CI 0.70-0.97) in training and 0.57 (95%_CI 0.23-0.86) in test cohort. Radiomic-clinical mixed model had an AUC of 0.90 (95%_CI 0.73-0.98) in training and 0.66 (95%_CI 0.33-0.88) in test cohort. Conclusion We produced an MRI radiomic-based ML algorithm with good prediction ability for future graft failure in patients with kidney transplant. Given the limited number of enrolled patients, validation in larger (and ideally prospective) cohorts is required to confirm our findings. Comparison of radiomic-based ML with histological parameters (i.e., IFTA) for the prediction of graft failure in our cohort is currently under assessment.

#5276 DEVELOPMENT OF A MULTIMODAL “KIDNEY AGE” PREDICTION BASED ON AUTOMATIC SEGMENTATION CT IMAGE IN PATIENTS WITH NORMAL RENAL FUNCTION

Abstract Background and Aims Renal function, for decades, has been of interest to clinicians and researchers to describe. For example, serum creatinine (Scr) and estimated glomerulus filtration rate (eGFR) is familiar but also limited in many circumstances. Meanwhile, the physiological volumes of the kidney cortex and medulla are presumed to change with age and have been proven to change with decreasing kidney function. Method We recruited 182 patients with normal Scr levels and contrasted CT images between Oct. 2021 and Feb 2022 in Peking Union Medical College Hospital (PUMCH) with their demographic and clinical data. The automatic segmentation method was modified from U-NET and used for both cortex and medullary separation and their volume calculation, respectively. We combined the kidney volume and clinical data as multimodal features of the machine learning model. All the data were separated into a training dataset (PUMCH training set, 80%) and a test dataset (PUMCH test set, 20%). Besides, we included patients with the same inclusion criteria but with diabetes (PUMCH-DM test set) and diabetic nephropathy (PUMCH-DN test set) for internal comparison to verify the possible clinical value of “KIDNEY AGE” (K-AGE). Data from DongZhiMen Hospital (DZMH test set) was used as separate external validation sets to evaluate model generalizability. Results The PUMCH training set included 146 participants with a mean age of 47.5±7.4 years. 58.9% were female, and the mean Scr is 63.5±12.3 μmol/L. The PUMCH test set included 36 participants with a mean age of 47.1±7.9 years, 52.7% were female, and the mean Scr was 66.9±13.0 μmol/L. For segmented kidneys, volume differences between non-contrast-enhanced CT and non-contrast-enhanced CT in the left and right kidneys were not evident. The multimodal neural network predicted: K-AGE approximately close to the patient's actual physiological age, with 92% prediction within the 95% confidential interval (Figure 1). We also compared the K-AGE prediction in PUMCH, PUMCH-DM, and PUMCH-DN test sets by applying U-NET-nonCon segmentation algorithm in non-contrast CT images. The mean absolute error increases along with the disease process (control group 5.00, diabetes group 6.99, diabetic nephropathy group 9.32) (Figure 2). Conclusion We established a machine learning model for predicting the K-AGE of normal Scr patients.

#4947 LMO7 IN URINE AS A SPECIFIC MARKER OF COMPROMISED RENAL EPITHELIAL INTEGRITY

Abstract Background and Aims Renal epithelial barrier (REB) are the physical isolator as well as physiological interface mediating body fluid and electrolytes homeostasis. Maintaining epithelial integrity is tightly related to REB biological function. LMO7 (LIM domain only 7) is a protein associated with the cell–cell junctional complex and is predominantly distributed in renal tubular epithelial cells. Previously, we reported that the LMO7 is associated with the maintenance of renal epithelial integrity, and that LMO7 releases to conditioned medium when the epithelial integrity was lost. In this study, we attempt to characterize roles of LMO7 and its functional interacting protein in maintaining epithelial integrity, and relevance of the LMO7 released from plasma membrane and the loss of tubular epithelial integrity. Method Renal ischemia-reperfusion injury (IRI) in mice as a model of acute kidney injury (AKI) were settled as a platform to investigate the the roles of LMO7 and its functional interacting protein in the pathological progression of compromised tubular epithelial integrity. Additionally, renal epithelial cells cultured in hypoxia as the pathogenic condition equivalent to AKI were also employed to investigate the role of LMO7. We also search the STRING database with LMO7 as query to find out its functional interacting proteins. Pathological pattern associated with LMO7 and its functional interacting protein in mice suffering from AKI as well as loss of tubular epithelial integrity were examined by immunofluorescent visualization. The LMO7 in urine as well as in conditioned medium derived from medium of hypoxia culture were quantified by Western blot analysis. Results Renal damage in IRI mice showed loss of epithelial integrity by histochemical staining, presence of LMO7 in urine, and high serum creatinine on the third day post IRI. Under the periodic acid-schiff (PAS) staining, loss of epithelial integrity is visible in proximal tubules, Henle's loops, and collecting ducts at the third day after reperfusion, and brush border impairment is observed in the outer stripe of the outer medulla. Most severe damages in renal tubules are present in Henle's loops where the epithelia express abundant LMO7. In the AKI mice, the LMO7 in renal tubules was apparently lower in Henle's loops. Additionally, the VANGL1, a LMO7 interacting protein participated in epithelial integrity and planar cell polarity in kidney, is also absent in Henle's loop of AKI mice. Hypoxia and glucose deprivation in cultured renal epithelial cells resulted in loss of epithelial integrity, and down-regulation of LMO7 and VANGL1. LMO7 was also identified in conditioned medium in which cells exposed to hypoxia but not normoxia. Additionally, the LMO7 was detected in urine of some patients with chronic kidney disease. Conclusion Presence of LMO7 in urine has been promising in experimental models holding compromised epithelial integrity. These approaches identified the role of LMO7 in maintaining epithelial integrity and that LMO7 being released into urine after renal epithelial damage as a novel marker for loss of REB.

Detection of Distinct Distributions of Acetaminophen and Acetaminophen-Cysteine in Kidneys up to 10 μm Resolution and Identification of a Novel Acetaminophen Metabolite Using an AP-MALDI Imaging Mass Microscope.

Drug distribution studies in tissue are crucial for understanding the pharmacokinetics and potential toxicity of drugs. Recently, matrix-assisted laser desorption ionization-mass spectrometry imaging (MALDI-MSI) has gained attention for drug distribution studies due to its high sensitivity, label-free nature, and ability to distinguish between parent drugs, their metabolites, and endogenous molecules. Despite these advantages, achieving high spatial resolution in drug imaging is challenging. Importantly, many drugs and metabolites are rarely detectable by conventional vacuum MALDI-MSI because of their poor ionization efficiency. It has been reported that acetaminophen (APAP) and one of its major metabolites, APAP-Cysteine (APAP-CYS), cannot be detected by vacuum MALDI-MSI without derivatization. In this context, we showed the distribution of both APAP and APAP-CYS in kidneys at high spatial resolution (25 and 10 μm) by employing an atmospheric pressure-MALDI imaging mass microscope without derivatization. APAP was highly accumulated in the renal pelvis 1 h after drug administration, while APAP-CYS exhibited characteristic distributions in the outer medulla and renal pelvis at both 30 min and 1 h after administration. Interestingly, cluster-like distributions of APAP and APAP-CYS were observed in the renal pelvis at 10 μm spatial resolution. Additionally, a novel APAP metabolite, tentatively coined as APAP-butyl sulfate (APAP-BS), was identified in the kidney, brain, and liver by combining MSI and tandem MSI. For the first time, our study revealed differential distributions of APAP, APAP-CYS (in kidneys), and APAP-BS (in kidney, brain, and liver) and is believed to enhance the understanding of the pharmacokinetics and potential nephrotoxicity of this drug.