Macrophage polarization in kidney transplant patients.

BACKGROUND: Macrophages can oscillate between two functionally distinct states: proinflammatory M1 and anti-inflammatory M2. Classically- activated M1 macrophages produce proinflammatory cytokines (TNF-α, IFN-Æ´, and IL-6), which ares associated with graft dysfunction/rejections. In contrast, alternatively-activated macrophages M2 produce anti-inflammatory cytokines (IL-10) that are involved in host defense, tissue repair/remodeling, debris scavenging, and immune regulation, thereby helps to improve long-term graft survival. METHODS: In this study, we have identified graft dysfunction or rejection by biopsies using immunohistochemistry. Flow cytometry was used to detect M1 (CD163+, CD206+, and CD200R+) and M2 (CD86+, CD80+, and CD68+) macrophages. Enzyme-linked immunosorbent assay (ELISA) was used to measure a panel of cytokines. RESULTS: Histological analysis of the kidney transplants (n = 30) was used to distinguish those with acute/chronic allograft rejection (n = 15) from those with stable kidney function (n = 15). Flow cytometry results showed that patients with graft rejection exhibited macrophages with decreased expression (33.28%) of M2 macrophage markers (CD163+, CD206+, and CD200R+) and reduced production of IL-10 (as detected using ELISA). However, 71.33% of the macrophages were found to have M1 markers (CD86+, CD80+, and CD68+; p = 0.002) and produced proinflammatory cytokines (TNF-α, IFN-Æ´, and IL-6) by ELISA (p = 0.001) when compared with the healthy control group. In contrast, stable kidney transplants had 65.58% M2 and 27.66% M1 macrophages (p = 0.03) and produced IL-10. These findings suggest that M1 macrophages dominate in kidney grafts with dysfunction or rejection, whereas M2 macrophages dominate in kidney grafts with stable function. CONCLUSION: Our observations implicate a major shift towards M2 macrophages in stable kidney transplants, which are markedly downregulated in patients with graft dysfunction or rejection. In contrast, an increased frequency of M1 macrophages remained dominant in the pathophysiology of kidney transplants undergoing active dysfunction or rejection.

Revisiting Vasculopathy in Lupus Nephritis: A Renal Biopsy Evaluation Study.

Introduction: The classification of lupus nephritis (LN) on biopsy is essentially focused on morphologic changes in glomeruli. Renal vascular lesions are not addressed in detail in current classifications and are often overlooked. We aimed to determine the prevalence of vascular lesions in LN on biopsies and to compare these with biopsies not showing the vasculopathies. Methods: A total of 740 renal biopsies of LN were analysedfor presence of vasculopathies from January 2013 to June 2019. Of these, 527 (71.2%) biopsies showed vascular lesions (vascular group), which were further categorized into known five subtypes according to morphology and immunofluorescence (IF) findings. Remaining 213 (28.8%) biopsies constituted non-vascular group. Clinical, demographic and laboratory parameters were compared between these two groups. Results: The mean age was 27.95 ± 9.8 years and 27.0 ± 9.4 years in the vascular and non-vascular groups respectively with higher M:F (1:2 > 1:7) in vascular group. Majority of vasculopathies (257, 48.7%) were found in biopsies with class IV LN. Haematuria (69.8% vs. 20.1%), proteinuria (100% vs. 62%), anemia (48.3% vs. 3.60%) and hypertension (39.8% vs. 8.46%) were common in group I. Uncomplicated vascular immune deposits (426; 80.8%) were the most common vasculopathy and true vasculitis (4;0.8%) was least common. Activity and chronicity indices (7.35 ± 3 and 2.45 ± 1.5, respectively) were significantly higher in the vascular group. Activity index was highest in uncomplicated vascular immune deposits (7.45 ± 2.8) and chronicity index was highest in non-specific sclerotic vascular lesions (2.7 ± 1.6). Conclusion: Vascular involvement is common in LN. Uncomplicated vascular immune deposits were common vasculopathies whereas true vasculitis was least common. The morphology and IF both need to be carefully screened for the diagnosis of vasculopathies.

Correlation between the clinical remission and histological remission in repeat biopsy findings of quiescent proliferative lupus nephritis.

BACKGROUND: The optimal duration of maintenance therapy is controversial in proliferative lupus nephritis (LN). Discordance between clinical parameters of renal remission and histological findings has made repeat biopsy a compulsory tool to confirm the histological remission (HR), but the timing is debatable. Aim of this study was to find the correlation of sustained complete clinical remission (CR) in sever lupus nephritis with histological findings on repeat kidney biopsy and appropriate duration of treatment in maintenance phase after achieving complete clinical remission. METHODS: Repeated kidney biopsy (biopsy 2) was performed on patients of biopsy proven (biopsy 1) proliferative LN who had been in CR for at least 2-years. The clinical and histologic findings of these groups (biopsy 1 and biopsy 2) were compared. Total 29 patients were included for the final analysis. Group 2 was further divided as per the duration of sustained CR (>48 months & <48 months). Regression analysis were used to find predictors of the HR. RESULTS: Average time taken to achieve CR was 9(range 2-24) months. Average duration of follow up and maintenance therapy was 68 ± 17.8 and 62.5 ± 14.2 months respectively. In the repeat kidney biopsy, HR was observed in 93.1% patients. Immunofluorescence study (IF) was normal in 72% of the patients. Normal light microscopy (LM) findings were observed in 58% patients. Transformation from proliferative to nonproliferative LN was noted in 82% cases. Other than the duration of CR on maintenance therapy and blood pressure, rest of the variables failed to predict HR. In sustained remission for more than 48-months group, 100% patients achieved HR whereas only 84% in 24-48-months group. CONCLUSION: Sustained CR on maintenance immunosuppressive therapy for more than 48-months duration predicts HR in repeat kidney biopsy findings in quiescent proliferative LN. Hence the minimum duration of maintenance therapy in proliferative LN should be at least for another 48 months after achieving CR.

Endothelial-to-mesenchymal transition compromises vascular integrity to induce Myc-mediated metabolic reprogramming in kidney fibrosis.

Endothelial-to-mesenchymal transition (EndMT) is a cellular transdifferentiation program in which endothelial cells partially lose their endothelial identity and acquire mesenchymal-like features. Renal capillary endothelial cells can undergo EndMT in association with persistent damage of the renal parenchyma. The functional consequence(s) of EndMT in kidney fibrosis remains unexplored. Here, we studied the effect of Twist or Snail deficiency in endothelial cells on EndMT in kidney fibrosis. Conditional deletion of Twist1 (which encodes Twist) or Snai1 (which encodes Snail) in VE-cadherin+ or Tie1+ endothelial cells inhibited the emergence of EndMT and improved kidney fibrosis in two different kidney injury/fibrosis mouse models. Suppression of EndMT limited peritubular vascular leakage, reduced tissue hypoxia, and preserved tubular epithelial health and function. Hypoxia, which was exacerbated by EndMT, resulted in increased Myc abundance in tubular epithelial cells, enhanced glycolysis, and suppression of fatty acid oxidation. Pharmacological suppression or epithelial-specific genetic ablation of Myc in tubular epithelial cells ameliorated fibrosis and restored renal parenchymal function and metabolic homeostasis. Together, these findings demonstrate a functional role for EndMT in the response to kidney capillary endothelial injury and highlight the contribution of endothelial-epithelial cross-talk in the development of kidney fibrosis with a potential for therapeutic intervention.

Anti-glomerular basement membrane disease: A clinicomorphological study of 16 cases.

INTRODUCTION: Antiglomerular basement membrane disease manifests as rapidly progressive glomerulonephritis and alveolar hemorrhage. It encompasses 10-15% of crescentic glomerulonephritis and is associated with poor outcome. In this study, we have elaborated on the clinical details, morphological features, and outcome of anti-GBM glomerulonephritis. MATERIALS AND METHODS: All the consecutive biopsy-proven cases of anti-GBM glomerulonephritis over a period of 4½ years were analyzed, retrospectively. RESULTS: Sixteen cases were diagnosed as anti-GBM glomerulonephritis during the study period. Twelve patients presented with rapidly progressive renal failure of which four patients required hemodialysis at the time of presentation. Goodpasture's syndrome was noted in two patients. Thirteen cases were positive for circulating anti-GBM antibodies and two patients showed double positivity for both anti-GBM antibodies and ANCA. Fifteen biopsies revealed crescentic glomerulonephritis with linear deposition of IgG along the glomerular basement membrane in all the 16 cases. CONCLUSION: Renal biopsy analysis is important in the diagnosis of Anti GBM nephritis. Morphology is an important predictor of disease progression.

Isolated Vascular Lesions (IVL) in Early Allograft Biopsies: A Case Series.

This case series includes five patients diagnosed as isolated vascular lesion (IVL) on allograft biopsy in an early post-transplant period. These patients presented with graft dysfunction. The biopsies satisfied the criteria for IVL as laid down by Banff 2009. Four of these patients were treated with corticosteroids and other anti rejection measures. C4d and DSA were negative in all. The patients showed good response to treatment with stable graft function at the longest follow-up of one year. We have also reviewed the literature about IVL as a specific entity. There are differences between the molecular and clinical data of IVL. It is difficult to differentiate whether IVL is a rejection or non-rejection process. This study aims to highlight the importance of a rare entity.

Revisiting renal amyloidosis with clinicopathological characteristics, grading, and scoring: A single-institutional experience.

INTRODUCTION: Kidney involvement is a major cause of mortality in systemic amyloidosis. Glomerulus is the most common site of deposition in renal amyloidosis, and nephrotic syndrome is the most common presentation. Distinction between AA and AL is done using immunofluorescence (IF) and immunohistochemistry (IHC). Renal biopsy helps in diagnosis and also predicting the clinical course by applying scoring and grading to the biopsy findings. MATERIALS AND METHODS: The study includes all cases of biopsy-proven renal amyloidosis from January 2008 to May 2017. Light microscopic analysis; Congo red with polarization; IF; IHC for Amyloid A, kappa, and lambda; and bone marrow evaluation were done. Classification of glomerular amyloid deposition and scoring and grading are done as per the guidelines of Sen S et al. RESULTS: There are 40 cases of biopsy-proven renal amyloidosis with 12 primary and 23 secondary cases. Mean age at presentation was 42.5 years. Edema was the most common presenting feature. Secondary amyloidosis cases were predominant. Tuberculosis was the most common secondary cause. Multiple myeloma was detected in four primary cases. Grading of renal biopsy features showed a good correlation with the class of glomerular involvement. CONCLUSION: Clinical history, IF, and IHC are essential in amyloid typing. Grading helps provide a subtle guide regarding the severity of disease in the background of a wide range of morphological features and biochemical values. Typing of amyloid is also essential for choosing the appropriate treatment.

Origin and function of myofibroblasts in kidney fibrosis.

Myofibroblasts are associated with organ fibrosis, but their precise origin and functional role remain unknown. We used multiple genetically engineered mice to track, fate map and ablate cells to determine the source and function of myofibroblasts in kidney fibrosis. Through this comprehensive analysis, we identified that the total pool of myofibroblasts is split, with 50% arising from local resident fibroblasts through proliferation. The nonproliferating myofibroblasts derive through differentiation from bone marrow (35%), the endothelial-to-mesenchymal transition program (10%) and the epithelial-to-mesenchymal transition program (5%). Specific deletion of Tgfbr2 in α-smooth muscle actin (αSMA)(+) cells revealed the importance of this pathway in the recruitment of myofibroblasts through differentiation. Using genetic mouse models and a fate-mapping strategy, we determined that vascular pericytes probably do not contribute to the emergence of myofibroblasts or fibrosis. Our data suggest that targeting diverse pathways is required to substantially inhibit the composite accumulation of myofibroblasts in kidney fibrosis.

Activin-like kinase 3 is important for kidney regeneration and reversal of fibrosis.

Molecules associated with the transforming growth factor ß (TGF-ß) superfamily, such as bone morphogenic proteins (BMPs) and TGF-ß, are key regulators of inflammation, apoptosis and cellular transitions. Here we show that the BMP receptor activin-like kinase 3 (Alk3) is elevated early in diseased kidneys after injury. We also found that its deletion in the tubular epithelium leads to enhanced TGF-ß1-Smad family member 3 (Smad3) signaling, epithelial damage and fibrosis, suggesting a protective role for Alk3-mediated signaling in the kidney. A structure-function analysis of the BMP-Alk3-BMP receptor, type 2 (BMPR2) ligand-receptor complex, along with synthetic organic chemistry, led us to construct a library of small peptide agonists of BMP signaling that function through the Alk3 receptor. One such peptide agonist, THR-123, suppressed inflammation, apoptosis and the epithelial-to-mesenchymal transition program and reversed established fibrosis in five mouse models of acute and chronic renal injury. THR-123 acts specifically through Alk3 signaling, as mice with a targeted deletion for Alk3 in their tubular epithelium did not respond to therapy with THR-123. Combining THR-123 and the angiotensin-converting enzyme inhibitor captopril had an additive therapeutic benefit in controlling renal fibrosis. Our studies show that BMP signaling agonists constitute a new line of therapeutic agents with potential utility in the clinic to induce regeneration, repair and reverse established fibrosis.