MOGAD patient autoantibodies induce complement, phagocytosis, and cellular cytotoxicity.

Myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease (MOGAD) is an inflammatory demyelinating CNS condition characterized by the presence of MOG autoantibodies. We sought to investigate whether human MOG autoantibodies are capable of mediating damage to MOG-expressing cells through multiple mechanisms. We developed high-throughput assays to measure complement activity (CA), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and antibody-dependent cellular cytotoxicity (ADCC) of live MOG-expressing cells. MOGAD patient sera effectively mediate all of these effector functions. Our collective analyses reveal that (a) cytotoxicity is not incumbent on MOG autoantibody quantity alone; (b) engagement of effector functions by MOGAD patient serum is bimodal, with some sera exhibiting cytotoxic capacity while others did not; (c) the magnitude of CDC and ADCP is elevated closer to relapse, while MOG-IgG binding is not; and (d) all IgG subclasses can damage MOG-expressing cells. Histopathology from a representative MOGAD case revealed congruence between lesion histology and serum CDC and ADCP, and we identified NK cells, mediators of ADCC, in the cerebrospinal fluid of relapsing patients with MOGAD. Thus, MOGAD-derived autoantibodies are cytotoxic to MOG-expressing cells through multiple mechanisms, and assays quantifying CDC and ADCP may prove to be effective tools for predicting risk of future relapses.

The Immune System and Idiopathic Nephrotic Syndrome.

Idiopathic nephrotic syndrome often responds to immunosuppressive treatment. Nevertheless, this syndrome-and the drugs used to treat it-remain important causes of patient morbidity. Idiopathic nephrotic syndrome is usually caused by minimal change disease or FSGS, diseases that primarily affect the podocytes. In spite of decades of research, the underlying causes of both diseases remain incompletely understood. There is, however, a large body of observational and experimental data linking the immune system with both minimal change disease and FSGS, including associations with systemic infections and hematologic malignancies. Perhaps most compellingly, many different immunomodulatory drugs are effective for treating idiopathic nephrotic syndrome, including biologic agents that have well-defined immune targets. In fact, the unexpected efficacy of targeted therapeutic agents has provided important new insights into the pathogenesis of these diseases. Given the large number of drugs that are available to deplete or block specific cells and molecules within the immune system, a better understanding of the immunologic causes of idiopathic nephrotic syndrome may lead to better diagnostic and therapeutic approaches.

Extracellular vesicles derived from patients with antibody-mediated rejection induce tubular senescence and endothelial to mesenchymal transition in renal cells.

Extracellular vesicles (EV) are emerging mediators in several diseases. However, their role in the pathophysiology of antibody-mediated allograft rejection (AMR) has been poorly investigated. Here, we investigated the role of EV isolated from AMR patients in inducing tubular senescence and endothelial to mesenchymal transition (EndMT) and analyzed their miRNA expression profile. By multiplex bead flow cytometry, we characterized the immunophenotype of plasma AMR-derived EV and found a prevalent platelet and endothelial cell origin. In vitro, AMR-derived EV induced tubular senescence by upregulating SA-ß Gal and CDKN1A mRNA. Furthermore, AMR-derived EV induced EndMT. The occurrence of tubular senescence and EndMT was confirmed by analysis of renal biopsies from the same AMR patients. Moreover, AMR-derived EV induced C3 gene upregulation and CFH downregulation in tubular epithelial cells, with C4d deposition on endothelial cells. Interestingly, RNase-mediated digestion of EV cargo completely abrogated tubular senescence and EndMT. By microarray analysis, miR-604, miR-515-3p, miR-let-7d-5p, and miR-590-3p were significantly upregulated in EV from AMR group compared with transplant controls, whereas miR-24-3p and miR-29a-3p were downregulated. Therefore, EV-associated miRNA could act as active player in AMR pathogenesis, unraveling potential mechanisms of accelerated graft senescence, complement activation and early fibrosis that might lead to new therapeutic intervention.

Minimal Change Disease Is Associated With Endothelial Glycocalyx Degradation and Endothelial Activation.

Introduction: Minimal change disease (MCD) is considered a podocyte disorder triggered by unknown circulating factors. Here, we hypothesized that the endothelial cell (EC) is also involved in MCD. Methods: We studied 45 children with idiopathic nephrotic syndrome (44 had steroid sensitive nephrotic syndrome [SSNS], and 12 had biopsy-proven MCD), 21 adults with MCD, and 38 healthy controls (30 children, 8 adults). In circulation, we measured products of endothelial glycocalyx (EG) degradation (syndecan-1, heparan sulfate [HS] fragments), HS proteoglycan cleaving enzymes (matrix metalloprotease-2 [MMP-2], heparanase activity), and markers of endothelial activation (von Willebrand factor [vWF], thrombomodulin) by enzyme-linked immunosorbent assay (ELISA) and mass spectrometry. In human kidney tissue, we assessed glomerular EC (GEnC) activation by immunofluorescence of caveolin-1 (n = 11 MCD, n = 5 controls). In vitro, we cultured immortalized human GEnC with sera from control subjects and patients with MCD/SSNS sera in relapse (n = 5 per group) and performed Western blotting of thrombomodulin of cell lysates as surrogate marker of endothelial activation. Results: In circulation, median concentrations of all endothelial markers were higher in patients with active disease compared with controls and remained high in some patients during remission. In the MCD glomerulus, caveolin-1 expression was higher, in an endothelial-specific pattern, compared with controls. In cultured human GEnC, sera from children with MCD/SSNS in relapse increased thrombomodulin expression compared with control sera. Conclusion: Our data show that alterations involving the systemic and glomerular endothelium are nearly universal in patients with MCD and SSNS, and that GEnC can be directly activated by circulating factors present in the MCD/SSNS sera during relapse.

Upregulation of complement proteins in lung cancer cells mediates tumor progression.

Introduction: In vivo, cancer cells respond to signals from the tumor microenvironment resulting in changes in expression of proteins that promote tumor progression and suppress anti-tumor immunity. This study employed an orthotopic immunocompetent model of lung cancer to define pathways that are altered in cancer cells recovered from tumors compared to cells grown in culture. Methods: Studies used four murine cell lines implanted into the lungs of syngeneic mice. Cancer cells were recovered using FACS, and transcriptional changes compared to cells grown in culture were determined by RNA-seq. Results: Changes in interferon response, antigen presentation and cytokine signaling were observed in all tumors. In addition, we observed induction of the complement pathway. We previously demonstrated that activation of complement is critical for tumor progression in this model. Complement can play both a pro-tumorigenic role through production of anaphylatoxins, and an anti-tumorigenic role by promoting complement-mediated cell killing of cancer cells. While complement proteins are produced by the liver, expression of complement proteins by cancer cells has been described. Silencing cancer cell-specific C3 inhibited tumor growth In vivo. We hypothesized that induction of complement regulatory proteins was critical for blocking the anti-tumor effects of complement activation. Silencing complement regulatory proteins also inhibited tumor growth, with different regulatory proteins acting in a cell-specific manner. Discussion: Based on these data we propose that localized induction of complement in cancer cells is a common feature of lung tumors that promotes tumor progression, with induction of complement regulatory proteins protecting cells from complement mediated-cell killing.

Detection of pro angiogenic and inflammatory biomarkers in patients with CKD.

Cardiovascular disease (CVD) is the most common cause of death in patients with native and post-transplant chronic kidney disease (CKD). To identify new biomarkers of vascular injury and inflammation, we analyzed the proteome of plasma and circulating extracellular vesicles (EVs) in native and post-transplant CKD patients utilizing an aptamer-based assay. Proteins of angiogenesis were significantly higher in native and post-transplant CKD patients versus healthy controls. Ingenuity pathway analysis (IPA) indicated Ephrin receptor signaling, serine biosynthesis, and transforming growth factor-ß as the top pathways activated in both CKD groups. Pro-inflammatory proteins were significantly higher only in the EVs of native CKD patients. IPA indicated acute phase response signaling, insulin-like growth factor-1, tumor necrosis factor-α, and interleukin-6 pathway activation. These data indicate that pathways of angiogenesis and inflammation are activated in CKD patients' plasma and EVs, respectively. The pathways common in both native and post-transplant CKD may signal similar mechanisms of CVD.

Influence of vitamin D treatment on functional expression of drug disposition pathways in human kidney proximal tubule cells during simulated uremia.

Effects of cholecalciferol (VitD3) and calcitriol (1,25-VitD3), on the expression and function of major vitamin D metabolizing enzymes (cytochrome P450 [CYP]2R1, CYP24A1) and select drug transport pathways (ABCB1/P-gp, SLCO4C1/OATP4C1) were evaluated in human kidney proximal tubule epithelial cells (hPTECs) under normal and uraemic serum conditions.hPTECs were incubated with 10% normal or uraemic serum for 24 h followed by treatment with 2% ethanol vehicle, or 100 and 240 nM doses of VitD3, or 1,25-VitD3 for 6 days. The effects of treatment on mRNA and protein expression and functional activity of select CYP enzymes and transporters were assessedUnder uraemic serum, treatment with 1,25-VitD3 resulted in increased mRNA but decreased protein expression of CYP2R1. Activity of CYP2R1 was not influenced by serum or VitD analogues. CYP24A1 expression was increased with 1,25-VitD3 under normal as well as uraemic serum, although to a lesser extent. ABCB1/P-gp mRNA expression increased under normal and uraemic serum, with exposure to 1,25-VitD3. SLCO4C1/OATP4C1 exhibited increased mRNA but decreased protein expression, under uraemic serum + 1,25-VitD3. Functional assessments of transport showed no changes regardless of exposure to serum or 1,25-VitD3.Key findings indicate that uraemic serum and VitD treatment led to differential effects on the functional expression of CYPs and transporters in hPTECs.

Rhabdomyolysis and complement-once again, epithelial cells take center stage.

Rhabdomyolysis is frequently associated with kidney injury. Unfortunately, there are no specific treatments for this condition, and patient care primarily consists of supportive measures. In this edition of Kidney International, Boudhabhay et al. demonstrate that myoglobin released from injured skeletal muscle cells triggers tubulointerstitial complement activation. In a mouse model of the disease, interventions that scavenged free heme or that prevented complement activation ameliorated kidney injury, raising the possibility that these strategies may be effective treatments for the condition.

Complement and Cancer-A Dysfunctional Relationship?

Although it was long believed that the complement system helps the body to identify and remove transformed cells, it is now clear that complement activation contributes to carcinogenesis and can also help tumors to escape immune-elimination. Complement is activated by several different mechanisms in various types of cancer, and complement activation fragments have multiple different downstream effects on cancer cells and throughout the tumor microenvironment. Thus, the role of complement activation in tumor biology may vary among different types of cancer and over time within a single tumor. In multiple different pre-clinical models, however, complement activation has been shown to recruit immunosuppressive myeloid cells into the tumor microenvironment. These cells, in turn, suppress anti-tumor T cell immunity, enabling the tumor to grow. Based on extensive pre-clinical work, therapeutic complement inhibitors hold great promise as a new class of immunotherapy. A greater understanding of the role of complement in tumor biology will improve our ability to identify those patients most likely to benefit from this treatment and to rationally combine complement inhibitors with other cancer therapies.

Complement factor H-deficient mice develop spontaneous hepatic tumors.

Hepatocellular carcinoma (HCC) is difficult to detect, carries a poor prognosis, and is one of few cancers with an increasing yearly incidence. Molecular defects in complement factor H (CFH), a critical regulatory protein of the complement alternative pathway (AP), are typically associated with inflammatory diseases of the eye and kidney. Little is known regarding the role of CFH in controlling complement activation within the liver. While studying aging CFH-deficient (fH-/-) mice, we observed spontaneous hepatic tumor formation in more than 50% of aged fH-/- males. Examination of fH-/- livers (3-24 months) for evidence of complement-mediated inflammation revealed widespread deposition of complement-activation fragments throughout the sinusoids, elevated transaminase levels, increased hepatic CD8+ and F4/80+ cells, overexpression of hepatic mRNA associated with inflammatory signaling pathways, steatosis, and increased collagen deposition. Immunostaining of human HCC biopsies revealed extensive deposition of complement fragments within the tumors. Investigating the Cancer Genome Atlas also revealed that increased CFH mRNA expression is associated with improved survival in patients with HCC, whereas mutations are associated with worse survival. These results indicate that CFH is critical for controlling complement activation in the liver, and in its absence, AP activation leads to chronic inflammation and promotes hepatic carcinogenesis.

Immunoglobulin-driven Complement Activation Regulates Proinflammatory Remodeling in Pulmonary Hypertension.

Rationale: Pulmonary hypertension (PH) is a life-threatening cardiopulmonary disorder in which inflammation and immunity have emerged as critical early pathogenic elements. Although proinflammatory processes in PH and pulmonary arterial hypertension (PAH) are the focus of extensive investigation, the initiating mechanisms remain elusive.Objectives: We tested whether activation of the complement cascade is critical in regulating proinflammatory and pro-proliferative processes in the initiation of experimental hypoxic PH and can serve as a prognostic biomarker of outcome in human PAH.Methods: We used immunostaining of lung tissues from experimental PH models and patients with PAH, analyses of genetic murine models lacking specific complement components or circulating immunoglobulins, cultured human pulmonary adventitial fibroblasts, and network medicine analysis of a biomarker risk panel from plasma of patients with PAH.Measurements and Main Results: Pulmonary perivascular-specific activation of the complement cascade was identified as a consistent critical determinant of PH and PAH in experimental animal models and humans. In experimental hypoxic PH, proinflammatory and pro-proliferative responses were dependent on complement (alternative pathway and component 5), and immunoglobulins, particularly IgG, were critical for activation of the complement cascade. We identified Csf2/GM-CSF as a primary complement-dependent inflammatory mediator. Furthermore, using network medicine analysis of a biomarker risk panel from plasma of patients with PAH, we demonstrated that complement signaling can serve as a prognostic factor for clinical outcome in PAH.Conclusions: This study establishes immunoglobulin-driven dysregulated complement activation as a critical pathobiological mechanism regulating proinflammatory and pro-proliferative processes in the initiation of experimental hypoxic PH and demonstrates complement signaling as a critical determinant of clinical outcome in PAH.

Complement fragments are biomarkers of antibody-mediated endothelial injury.

Antibody-mediated rejection (AbMR) adversely affects long-term graft survival in kidney transplantation. Currently, the diagnosis of AbMR requires a kidney biopsy, and detection of complement C4d deposition in the allograft is one of the diagnostic criteria. Complement activation also generates several soluble fragments which could potentially provide non-invasive biomarkers of the process. Furthermore, microvesicles released into the plasma from injured cells can serve as biomarkers of vascular injury. To explore whether soluble complement fragments or complement fragments bound to endothelial microvesicles can be used to non-invasively detect AbMR, we developed an in vitro model in which human endothelial cells were exposed to anti-HLA antibodies and complement sufficient serum. We found that complement fragments C4a and sC5b-9 were increased in the supernatants of cells exposed to complement-sufficient serum compared to cells treated complement-deficient serum. Furthermore, complement activation on the cell surface was associated with the release of microvesicles bearing C4 and C3 fragments. We next measured these analytes in plasma from kidney transplant recipients with biopsy-proven acute AbMR (n = 9) and compared the results with those from transplant recipients who also had impaired allograft function but who did not have AbMR (n = 30). Consistent with the in vitro results, complement fragments C4a and Ba were increased in plasma from patients with AbMR compared to control subjects (P < 0.001 and P < 0.01, respectively). Endothelial microvesicle counts were not increased in patients with AbMR, however, and the number of microvesicles with C4 and C3 bound to the surface was actually lower compared to control subjects (both P < 0.05). Our results suggest that plasma complement activation fragments may be useful as non-invasive biomarkers of antibody-mediated complement activation within the allograft. Complement-opsonized endothelial microvesicles are decreased in patients with AbMR, possibly due to enhanced clearance of microvesicles opsonized with C3 and C4 fragments.

Urine complement activation fragments are increased in patients with kidney injury after cardiac surgery.

Experiments in mouse models have shown that the complement cascade is activated within the kidney after ischemia-reperfusion and that complement activation contributes to tubular injury in this setting. Less is known, however, about complement activation in human kidneys after ischemia or whether complement activation in the tubulointerstitium can be detected by measurement of complement fragments in the urine. We hypothesized that urine biomarkers of complement activation would rapidly increase in patients who develop ischemic acute kidney injury, signaling complement activation within the kidney. We confirmed that the alternative pathway of complement is activated in the kidneys of mice after ischemia-reperfusion, and we found that levels of factor B fragments (generated during alternative pathway activation) rapidly increase in the urine. We next performed a case-control study in which we measured complement fragments in human urine samples from patients undergoing cardiac surgery using ELISAs. The level of Ba increased after cardiac surgery and was significantly higher in patients who developed acute kidney injury. The increase in Ba also correlated with magnitude of the subsequent rise in serum creatinine and with the need for hemodialysis during the hospitalization. These findings demonstrate that the alternative pathway of complement is activated in patients who develop acute kidney injury after cardiac surgery and that increases in the level of urine Ba may be a predictive and functional biomarker of severe kidney injury.

Cyclophilin D knockout protects the mouse kidney against cyclosporin A-induced oxidative stress.

Mitochondrial dysfunction and oxidative stress have been implicated in cyclosporin A (CsA)-induced nephrotoxicity. CsA interacts with cyclophilin D (CypD), an essential component of the mitochondrial permeability transition pore and regulator of cell death processes. Controversial reports have suggested that CypD deletion may or may not protect cells against oxidative stress-induced cell death. In the present study, we treated wild-type (WT) mice and mice lacking CypD [peptidylprolyl isomerase F knockout (Ppif-/-) mice] with CsA to test the role and contribution of CypD to the widely described CsA-induced renal toxicity and oxidative stress. Our results showed an increase in the levels of several known uremic toxins as well as the oxidative stress markers PGF2α and 8-isoprostane in CsA-treated WT animals but not in Ppif-/- animals. Similarly, a decline in S-adenosylmethionine and the resulting methylation potential indicative of DNA hypomethylation were observed only in CsA-treated WT mice. This confirms previous reports of the protective effects of CypD deletion on the mouse kidney mediated through a stronger resistance of these animals to oxidative stress and DNA methylation damage. However, a negative effect of CsA on the glycolysis and overall energy metabolism in Ppif-/- mice also indicated that additional, CypD-parallel pathways are involved in the toxic effects of CsA on the kidney. In summary, CsA-mediated induction of oxidative stress is associated with CypD, with CypD deletion providing a protective effect, whereas the reduction of energy production observed upon CsA exposure did not depend on the animals' CypD status.

SPECT/CT Imaging of Mycobacterium tuberculosis Infection with [125I]anti-C3d mAb.

PURPOSE: Diagnosis and therapeutic monitoring of chronic bacterial infection requires methods to detect and localize sites of infection accurately. Complement C3 activation fragments are generated and covalently bound to selective bacterial pathogens during the immune response and can serve as biomarkers of ongoing bacterial infection. We have developed several probes for detecting tissue-bound C3 deposits, including a monoclonal antibody (mAb 3d29) that recognizes the tissue-bound terminal processing fragments iC3b and C3d but does not recognize native circulating C3 or tissue-bound C3b. PROCEDURES: To determine whether mAb 3d29 could be used to detect chronic Mycobacterium tuberculosis infection non-invasively, aerosol-infected female C3HeB/FeJ mice were injected with [125I]3d29 mAb and either imaged using single-photon emission computed tomography (SPECT)/X-ray computed tomography (CT) imaging at 24 and 48 h after radiotracer injection or being subjected to biodistribution analysis. RESULTS: Discrete lesions were detected by SPECT/CT imaging in the lungs and spleens of infected mice, consistent with the location of granulomas in the infected animals as detected by CT. Low-level signal was seen in the spleens of uninfected mice and no signal was seen in the lungs of healthy mice. Immunofluorescence microscopy revealed that 3d29 in the lungs of infected mice co-localized with aggregates of macrophages (detected with anti-CD68 antibodies). 3d29 was detected in the cytoplasm of macrophages, consistent with the location of internalized M. tuberculosis. 3d29 was also present within alveolar epithelial cells, indicating that it detected M. tuberculosis phagocytosed by other CD68-positive cells. Healthy controls showed very little retention of fluorescent or radiolabeled antibody across tissues. Radiolabeled 3d29 compared with radiolabeled isotype control showed a 3.5:1 ratio of increased uptake in infected lungs, indicating specific uptake by 3d29. CONCLUSION: 3d29 can be used to detect and localize areas of infection with M. tuberculosis non-invasively by 24 h after radiotracer injection and with high contrast.

The relationship between complement C3 expression and the MUC5B genotype in pulmonary fibrosis.

The common gain-of-function MUC5B promoter variant ( rs35705950 ) is the strongest risk factor for the development of idiopathic pulmonary fibrosis (IPF). While the role of complement in IPF is controversial, both MUC5B and the complement system play a role in lung host defense. The aim of this study was to evaluate the relationship between complement component 3 (C3) and MUC5B in patients with IPF and in bleomycin-induced lung injury in mice. To do this, we evaluated C3 gene expression in whole lung tissue from 300 subjects with IPF and 175 healthy controls. Expression of C3 was higher in IPF than healthy controls {1.40-fold increase [95% confidence interval (CI) 1.31-1.50]; P < 0.0001} and even greater among IPF subjects with the highest-risk IPF MUC5B promoter genotype [TT vs. GG = 1.59-fold (95% CI 1.15-2.20); P < 0.05; TT vs. GT = 1.66-fold (95% CI 1.20-2.30); P < 0.05]. Among subjects with IPF, C3 expression was significantly higher in the lung tissue without microscopic honeycombing than in the lung tissue with microscopic honeycombing [1.40-fold increase (95% CI 1.23- 1.59); P < 0.01]. In mice, while bleomycin exposure increased Muc5b protein expression, C3-deficient mice were protected from bleomycin-induced lung injury. In aggregate, our findings indicate that the MUC5B promoter variant is associated with higher C3 expression and suggest that the complement system may contribute to the pathogenesis of IPF.

Complement Activation via a C3a Receptor Pathway Alters CD4+ T Lymphocytes and Mediates Lung Cancer Progression.

The complement cascade is a part of the innate immune system that acts primarily to remove pathogens and injured cells. However, complement activation is also peculiarly associated with tumor progression. Here we report mechanistic insights into this association in multiple immunocompetent orthotopic models of lung cancer. After tumor engraftment, we observed systemic activation of the complement cascade as reflected by elevated levels of the key regulator C3a. Notably, growth of primary tumors and metastases was both strongly inhibited in C3-deficient mice (C3-/- mice), with tumors undetectable in many subjects. Growth inhibition was associated with increased numbers of IFNγ+/TNFα+/IL10+ CD4+ and CD8+ T cells. Immunodepletion of CD4+ but not CD8+ T cells in tumor-bearing subjects reversed the inhibitory effects of C3 deletion. Similarly, antagonists of the C3a or C5a receptors inhibited tumor growth. Investigations using multiple tumor cell lines in the orthotopic model suggested the involvement of a C3/C3 receptor autocrine signaling loop in regulating tumor growth. Overall, our findings offer functional evidence that complement activation serves as a critical immunomodulator in lung cancer progression, acting to drive immune escape via a C3/C5-dependent pathway.Significance: This provocative study suggests that inhibiting complement activation may heighten immunotherapeutic responses in lung cancer, offering findings with immediate implications, given the existing clinical availability of complement antagonists. Cancer Res; 78(1); 143-56. ©2017 AACR.

Mitochondrial cyclophilin D ablation is associated with the activation of Akt/p70S6K pathway in the mouse kidney.

The mitochondrial matrix protein cyclophilin D (CypD) is an essential component of the mitochondrial permeability transition pore (MPTP). Here we characterized the effects of CypD ablation on bioenergetics in the kidney. CypD loss triggers a metabolic shift in Ppif-/- male and female mouse kidneys towards glycolysis and Krebs cycle activity. The shift is accompanied by increased glucose consumption and a transcriptional upregulation of effectors of glucose metabolism in the kidney. These included activation of Akt, AMPK (only in males) and p70S6K kinases. Gender specific differences between the Ppif-/- male and female mouse kidneys were observed including activation of pro-surviving ERK1/2 kinase and inhibited expression of pro-apoptotic and pro-fibrotic JNK and TGFß1 proteins in Ppif-/- females. They also showed the highest expression of phosphorylated-ERK1/2 and Akt S473 proteins of all four investigated animal groups. Furthermore, Ppif-/- females showed higher lactate concentrations and ATP/ADP-ratios in the kidney than males. These metabolic and transcriptional modifications could provide an additional level of protection to Ppif-/- females. In summary, loss of mitochondrial CypD results in a shift in bioenergetics and in activation of glucose-metabolism regulating Akt/AMPK/p70S6 kinase pathways that is expected to affect the capability of Ppif-/- mice kidneys to react to stimuli and injury.

Never make assumptions: the complicated role of complement in urinary tract infections.

Complement activation can cause tissue inflammation and injury, and complement-inhibitory drugs are effective treatments for several inflammatory diseases. The complement cascade is part of the body's defense against bacteria and other pathogens, however, and a major concern regarding inhibition of this system is that it may increase the risk for infection. Now, a study by Choudhry et al. demonstrates that blockade of signaling at one of the C5a receptors (C5a receptor 1 [C5aR1]) reduces renal fibrosis in a mouse model of urinary tract infection with Escherichia coli. Surprisingly, C5aR1 blockade was also associated with faster clearance of the infection. The results of this study demonstrate that C5a-a highly proinflammatory molecule-reduces bacterial killing by macrophages. Other recent studies have also shown that C5a impairs the elimination of tumor cells by the immune system. These data indicate that complement inhibition may have some unexpected benefits. These results also demonstrate, however, that the complement cascade probably has physiologic functions that have yet to be discovered.

γδ T cells protect against LPS-induced lung injury.

γδ T lymphocytes are a unique T cell population with important anti-inflammatory capabilities. Their role in acute lung injury, however, is poorly understood but may provide significant insight into lung-protective mechanisms occurring after injury. In a murine model of lung injury, wild-type C57BL/6 and TCRδ(-/-) mice were exposed to Escherichia coli LPS, followed by analysis of γδ T cell and macrophage subsets. In the absence of γδ T cells, TCRδ(-/-) mice developed increased inflammation and alveolar-capillary leak compared with wild-type C57BL/6 mice after LPS exposure that correlated with expansion of distinct macrophage populations. Classically activated M1 macrophages were increased in the lung of TCRδ(-/-) mice at d 1, 4, and 7 after LPS exposure that peaked at d 4 and persisted at d 7 compared with wild-type animals. In response to LPS, Vγ1 and Vγ7 γδ T cells were expanded in the lung and expressed IL-4. Coculture experiments showed decreased expression of TNF-α by resident alveolar macrophages in the presence of γδ T cells that was reversed in the presence of an anti-IL-4-blocking antibody. Treatment of mice with rIL4 resulted in reduced numbers of M1 macrophages, inflammation, and alveolar-capillary leak. Therefore, one mechanism by which Vγ1 and Vγ7 γδ T cells protect against LPS-induced lung injury is through IL-4 expression, which decreases TNF-α production by resident alveolar macrophages, thus reducing accumulation of M1 macrophages, inflammation, and alveolar-capillary leak.