Heterogeneity of Fibroblasts in Healthy and Diseased Kidneys

Chronic kidney disease (CKD) is a worldwide health problem affecting 9.1% of the world’s population. The treatments to prevent the progression of CKD remain limited, however. Resident fibroblasts in the kidneys play crucial roles in the pathological conditions commonly recognized in CKD, such as renal fibrosis, renal anemia, and peritubular capillary loss. Fibroblasts in the kidney provide structural backbone by producing extracellular matrix proteins and produce erythropoietin for normal hematopoiesis under physiological conditions. In the diseased condition, however, fibroblasts differentiate into myofibroblasts that produce excessive extracellular matrix proteins at the cost of the inherent erythropoietin-producing abilities, resulting in renal fibrosis and renal anemia. Pericytes, which are mesenchymal cells that enwrap peritubular capillaries and highly overlap with resident fibroblasts, detach from peritubular capillary walls in response to kidney injury, resulting in peritubular capillary loss and tissue hypoxia. Several reports have demonstrated the beneficial roles of fibroblasts in the regeneration of renal tubules Renal fibroblasts also have the potential to differentiate into a proinflammatory state, producing various cytokines and chemokines and prolonging inflammation by forming tertiary lymphoid tissues, functional lymphoid aggregates, in some pathological conditions. In this article, we describe the heterogenous functions of renal fibroblasts under healthy and diseased conditions.

Caspase-3 dependent peritubular capillary dysfunction is pivotal for transition from acute to chronic kidney disease after acute ischemia-reperfusion injury

Ischemia-reperfusion injury (IRI) is a major risk factor for chronic renal failure. Caspase-3, an effector responsible for apoptosis execution, is activated within peritubular capillary (PTC) in the early stage of IRI-induced acute kidney injury (AKI). Recently, we showed that caspase-3-dependent microvascular rarefaction plays a key role in fibrosis development after mild renal IRI. Here, we further characterize the role of caspase-3 in microvascular dysfunction and progressive renal failure in both mild and severe AKI, by performing unilateral renal artery clamping for 30/60 minutes with contralateral nephrectomy in wild-type (C57BL/6) or caspase-3-/- mice. In both forms of AKI, caspase-3-/- mice showed better long-term outcomes in spite of worse initial tubular injury. After 3 weeks, they showed reduced PTC injury, decreased PTC collagen deposition and α-SMA expression, and lower tubular injury scores when compared to wild-type animals. Caspase-3-/- mice with severe IRI also showed better preservation of long-term renal function. Intra-vital imaging and micro Computed Tomography (microCT) revealed preserved PTCs permeability and better terminal capillary density in caspase-3-/- mice. Collectively, these results demonstrate the pivotal importance of caspase-3 in regulating long-term renal function after IRI and establish the predominant role of PTCs dysfunction as a major contributor to progressive renal dysfunction.

Quantitative assessment of inflammatory infiltrates in kidney transplant biopsies using multiplex tyramide signal amplification and deep learning

Delayed graft function (DGF) is a strong risk factor for development of interstitial fibrosis and tubular atrophy (IFTA) in kidney transplants. Quantitative assessment of inflammatory infiltrates in kidney biopsies of DGF patients can reveal predictive markers for IFTA development. In this study, we combined multiplex tyramide signal amplification (mTSA) and convolutional neural networks (CNNs) to assess the inflammatory microenvironment in kidney biopsies of DGF patients (n = 22) taken at 6 weeks post-transplantation. Patients were stratified for IFTA development (<10% versus ≥10%) from 6 weeks to 6 months post-transplantation, based on histopathological assessment by three kidney pathologists. One mTSA panel was developed for visualization of capillaries, T- and B-lymphocytes and macrophages and a second mTSA panel for T-helper cell and macrophage subsets. The slides were multi spectrally imaged and custom-made python scripts enabled conversion to artificial brightfield whole-slide images (WSI). We used an existing CNN for the detection of lymphocytes with cytoplasmatic staining patterns in immunohistochemistry and developed two new CNNs for the detection of macrophages and nuclear-stained lymphocytes. F1-scores were 0.77 (nuclear-stained lymphocytes), 0.81 (cytoplasmatic-stained lymphocytes), and 0.82 (macrophages) on a test set of artificial brightfield WSI. The CNNs were used to detect inflammatory cells, after which we assessed the peritubular capillary extent, cell density, cell ratios, and cell distance in the two patient groups. In this cohort, distance of macrophages to other immune cells and peritubular capillary extent did not vary significantly at 6 weeks post-transplantation between patient groups. CD163+ cell density was higher in patients with ≥10% IFTA development 6 months post-transplantation (p < 0.05). CD3+CD8−/CD3+CD8+ ratios were higher in patients with <10% IFTA development (p < 0.05). We observed a high correlation between CD163+ and CD4+GATA3+ cell density (R = 0.74, p < 0.001). Our study demonstrates that CNNs can be used to leverage reliable, quantitative results from mTSA-stained, multi spectrally imaged slides of kidney transplant biopsies.

Acute Kidney Injury, Microvascular Rarefaction, and Estimated Glomerular Filtration Rate in Kidney Transplant Recipients

Background and objectivesAnimal studies suggest that microvascular rarefaction is a key factor in the acute kidney disease to CKD transition. Hence, delayed graft function appears as a unique human model of AKI to further explore the role of microvascular rarefaction in kidney transplant recipients. Here, we assessed whether delayed graft function is associated with peritubular capillary loss and evaluated the association between this loss and long-term kidney graft function.Design, setting, participants, & measurementsThis observational, retrospective cohort study included 61 participants who experienced delayed graft function and 130 who had immediate graft function. We used linear regression models to evaluate associations between delayed graft function and peritubular capillary density expressed as the percentage of efficient cortical area occupied by peritubular capillaries in pre- and post-transplant graft biopsies. eGFRs 1 and 3 years post-transplant were secondary outcomes.ResultsPost-transplant biopsies were performed at a median of 113 days (interquartile range, 101–128) after transplantation. Peritubular capillary density went from 15.4% to 11.5% in patients with delayed graft function (median change, −3.7%; interquartile range, −6.6% to −0.8%) and from 19.7% to 15.1% in those with immediate graft function (median change, −4.5%; interquartile range, −8.0% to −0.8%). Although the unadjusted change in peritubular capillary density was similar between patients with and without delayed graft function, delayed graft function was associated with more peritubular capillary loss in the multivariable analysis (adjusted difference in change, −2.9%; 95% confidence interval, −4.0 to −1.8). Pretransplant peritubular capillary density and change in peritubular capillary density were associated with eGFR 1 and 3 years post-transplantation.ConclusionsPerioperative AKI is associated with lower density in peritubular capillaries before transplantation and with loss of peritubular capillaries following transplantation. Lower peritubular capillary density is linked to lower long-term eGFR.

Evaluation of Renal Fibrosis by Mapping Histology and Magnetic Resonance Imaging

<b><i>Background:</i></b> Renal fibrosis is a key driver of progression in chronic kidney disease (CKD). Recent advances in diagnostic imaging techniques have shown promising results for the noninvasive assessment of renal fibrosis. However, the specificity and accuracy of these techniques are controversial because they indirectly assess renal fibrosis. This limits fibrosis assessment by imaging in CKD for clinical practice. To validate magnetic resonance imaging (MRI) assessment for fibrosis, we derived representative models by mapping histology-proven renal fibrosis and imaging in CKD. <b><i>Methods:</i></b> Ninety-seven adult Chinese CKD participants with histology were studied. The kidney cortex interstitial extracellular matrix volume was calculated by the Aperio ScanScope system using Masson’s trichrome slices. The kidney cortex microcirculation was quantitatively assessed by peritubular capillary density using CD34 staining. The imaging techniques included intravoxel incoherent motion diffusion-weighted imaging and magnetic resonance elastography (MRE) imaging. Relevant analyses were performed to evaluate the correlations between MRI parameters and histology variables. Multiple linear regression models were used to describe the relationships between a response variable and other variables. The best-fit lines, which minimize the sum of squared residuals of the multiple linear regression models, were generated. <b><i>Results:</i></b> MRE values were negatively associated with the interstitial extracellular matrix volume (<i>Rho</i> = −0.397, <i>p</i> &#x3c; 0.001). The best mapping model of extracellular matrix volume with the MRE value and estimated glomerular filtration rate (eGFR) we obtained was as follows: Interstitial extracellular matrix volume = 218.504 – 14.651 × In(MRE) – 18.499 × In(eGFR). DWI-fraction values were positively associated with peritubular capillary density (<i>Rho</i> = 0.472, <i>p</i> &#x3c; 0.001). The best mapping model of peritubular capillary density with DWI-fraction value and eGFR was as follows: Peritubular capillaries density = 17.914 + 9.403 × (DWI – fraction) + 0.112 × (eGFR). <b><i>Conclusions:</i></b> The study provides histological evidence to support that MRI can effectively evaluate fibrosis in the kidney. These findings picture the graphs of the mapping model from imaging and eGFR into fibrosis, which has significant value for clinical implementation.

The spectrum of C4d deposition in renal biopsies of lupus nephritis patients

Objectives This study aims to determine the prevalence and localization of complement factor C4d in renal biopsies of lupus nephritis (LN) patients, as well as its association with the clinico-pathological features of the disease. Especially, the correlation between arteriolar C4d deposition and renal microvascular lesions (RVL) was further analyzed. Methods A total of 325 biopsy-proven lupus nephritis patients were enrolled and their clinico-pathological data were collected. C4d staining in renal biopsies was performed by immunohistochemistry. The association between C4d deposition and the clinico-pathological features was further analyzed. Results C4d deposition was present in most of renal specimens (98.8%) in our cohort. They were localized in the glomeruli (98.2%), tubular basement membrane (TBM) (43.7%), arterioles (31.4%) and peritubular capillary (33.8%), respectively. TBM C4d staining was closely related to the disease activity (SLEDAI) and NIH pathological activity and chronicity indices (P < 0.01). Patients with arteriolar C4d deposition were more likely to develop RVL (91.2%) in comparison to those negative (78.0%; P = 0.004), especially with two or more types of RVL (P < 0.001). During an average follow-up of 55.8 months, the presence of arteriolar C4d was related to worse renal outcomes (HR: 2.074, 95% CI 1.056–4.075, P = 0.034). Co-deposition of arteriolar C4d and C3c was an independent risk factor (HR: 2.539, 95% CI 1.130–5.705, P = 0.024) for predicting renal outcomes by the multivariate stepwise Cox hazard analysis Conclusions C4d deposition was common in renal tissues of lupus nephritis patients. TBM C4d deposition was related to the disease activity and arteriolar C4d deposition was associated with RVL and worse renal outcomes.

Shen-Qi-Wan Protects the Renal Peritubular Capillary From Adenine-mediated Damage by Upregulating Aquaporin 1

Background：Shen-Qi-Wan (SQW), a commonly used prescription against chronic kidney disease (CKD) in Traditional Chinese Medicine (TCM), has a nephroprotective action in adenine-induced kidney injury. However, the mechanism of SQW in renal injury remains unclear.Methods: We evaluated the nephroprotective action of SQW in adenine-induced kidney injury and investigated its mechanism in vitro studies.Results: SQW supplementation could alleviate the pathological makers for CKD, ameliorate dysfunction of Hypothalamic-Pituitary-Adrenal (HPA) axis and renal function loss caused by adenine. Alternatively, SQW administration showed an ameliorating effect from the toxicity and alleviated the injury of capillaries around renal tubules instigated by adenine through increasing AQP1 mRNA and protein level. In vitro experiments, SQW medicated serum enhanced the migration and lumen formation ability of HMEC-1 cells, and significantly increased AQP1 protein level. Moreover, AQP1 knockdown efficiently inhibited migration and lumen formation ability in HMEC-1 cells which could be reversed by SQW medicated serum.Trial registration : This is an animal and cell experiment, trial registration is not necessary.Conclusion: These results suggested that SQW attenuated peritubular capillary injury in adenine induced CKD model rats through boosting angiogenesis in endothelial cell and AQP1 may be a potential target of SQW for treating renal injury.

Peritubular Capillary Rarefaction: An Underappreciated Regulator of CKD Progression

Peritubular capillary (PTC) rarefaction is commonly detected in chronic kidney disease (CKD) such as hypertensive nephrosclerosis and diabetic nephropathy. Moreover, PTC rarefaction prominently correlates with impaired kidney function and predicts the future development of end-stage renal disease in patients with CKD. However, it is still underappreciated that PTC rarefaction is a pivotal regulator of CKD progression, primarily because the molecular mechanisms of PTC rarefaction have not been well-elucidated. In addition to the established mechanisms (reduced proangiogenic factors and increased anti-angiogenic factors), recent studies discovered significant contribution of the following elements to PTC loss: (1) prompt susceptibility of PTC to injury, (2) impaired proliferation of PTC, (3) apoptosis/senescence of PTC, and (4) pericyte detachment from PTC. Mainly based on the recent and novel findings in basic research and clinical study, this review describes the roles of the above-mentioned elements in PTC loss and focuses on the major factors regulating PTC angiogenesis, the assessment of PTC rarefaction and its surrogate markers, and an overview of the possible therapeutic agents to mitigate PTC rarefaction during CKD progression. PTC rarefaction is not only a prominent histological characteristic of CKD but also a central driving force of CKD progression.