Lupus nephritis, one of the most serious manifestations of systemic lupus erythematosus (SLE), has both a heterogeneous clinical and pathological presentation. For example, proliferative nephritis identifies a more aggressive disease class that requires immunosuppression. However, the current classification system relies on the static appearance of histopathological morphology which does not capture differences in the inflammatory response. Therefore, a biomarker grounded in the disease biology is needed to understand the molecular heterogeneity of lupus nephritis and identify immunologic mechanism and pathways. Here, we analyzed the patterns of 1000 urine protein biomarkers in 30 patients with active lupus nephritis. We found that patients stratify over a chemokine gradient inducible by interferon-gamma. Higher values identified patients with proliferative lupus nephritis. After integrating the urine proteomics with the single-cell transcriptomics of kidney biopsies, it was observed that the urinary chemokines defining the gradient were predominantly produced by infiltrating CD8 T cells, along with natural killer and myeloid cells. The urine chemokine gradient significantly correlated with the number of kidney-infiltrating CD8 cells. These findings suggest that urine proteomics can capture the complex biology of the kidney in lupus nephritis. Patient-specific pathways may be noninvasively tracked in the urine in real time, enabling diagnosis and personalized treatment.
Andrea Fava, Jill P. Buyon, Chandra Mohan, Ting Zhang, H. Michael Belmont, Peter Izmirly, Robert Clancy, Jose Monroy Trujillo, Derek M. Fine, Yuji Zhang, Laurence Magder, Deepak A. Rao, Arnon Arazi, Celine C. Berthier, Anne Davidson, Betty Diamond, Nir Hacohen, David Wofsy, William Apruzzese, The Accelerating Medicines Partnership, Soumya Raychaudhuri, Michelle Petri
Autosomal co-dominant genetic disorder alpha-1 antitrypsin (AAT) deficiency (AATD) causes pulmonary and liver disease. Individuals homozygous for the mutant Z allele accumulate polymers of Z-AAT protein in hepatocytes, where AAT is primarily produced. This accumulation causes endoplasmic reticulum (ER) stress, oxidative stress, damage to mitochondria and inflammation, leading to fibrosis, cirrhosis and hepatocellular carcinoma. The magnitude of AAT reduction and duration of response from first generation intravenously-administered RNA interference (RNAi) therapeutic ARC-AAT and then with next generation subcutaneously-administered ARO-AAT were assessed by measuring AAT protein in serum of the PiZ transgenic mouse model and human volunteers. The impact of Z-AAT reduction by RNAi on liver disease phenotypes was evaluated in PiZ mice by measuring polymeric Z-AAT in the liver; expression of genes associated with fibrosis, autophagy, apoptosis and redox regulation; inflammation; Z-AAT globule parameters; and tumor formation. Ultrastructure of the ER, mitochondria and autophagosomes in hepatocytes were evaluated by electron microscopy. In mice, sustained RNAi treatment reduced hepatic Z-AAT polymer, restored ER and mitochondrial health, normalized expression of disease-associated genes, reduced inflammation and prevented tumor formation. RNAi therapy holds promise for the treatment of patients with AATD-associated liver disease. ARO-AAT is currently in Phase 2/3 clinical trials.
Christine I. Wooddell, Keith Blomenkamp, Ryan M. Peterson, Vladimir M. Subbotin, Christian Schwabe, James Hamilton, Qili Chu, Dawn R. Christianson, Julia O. Hegge, John Kolbe, Holly L. Hamilton, Maria F. Branca-Afrazi, Bruce D. Given, David L. Lewis, Edward Gane, Steven B. Kanner, Jeffrey H. Teckman
The ability of Mycobacterium tuberculosis to form serpentine cords is intrinsically related to its virulence, but specifically how M. tuberculosis cording contributes to pathogenesis remains obscure. We show that several M. tuberculosis clinical isolates form intracellular cords in primary human lymphatic endothelial cells (hLEC) in vitro and also in the lymph nodes of patients with tuberculosis. We identified via RNA-seq a transcriptional programme that activates, in infected-hLECs, cell-survival and cytosolic surveillance of pathogens pathways. Consistent with this, cytosolic access is required for intracellular M. tuberculosis cording. Mycobacteria lacking ESX-1 type VII secretion system or PDIM expression, which fail to access to the cytosol, are indeed unable to cords within hLECs. Finally, we show that M. tuberculosis cording is a size-dependent mechanism used by the pathogen to avoid its recognition by cytosolic sensors and evade either resting or IFN-γ-induced hLEC immunity. These results explain the long-standing association between M. tuberculosis cording and virulence and how virulent mycobacteria use intracellular cording as strategy to successfully adapt and persist in the lymphatic tracts.
Thomas R. Lerner, Christophe J. Queval, Rachel PJ Lai, Matthew Robert Geoffrey Russell, Antony Fearns, Daniel J. Greenwood, Lucy Collinson, Robert J. Wilkinson, Maximiliano G. Gutierrez
ABSTRACTWith the effectiveness of antimicrobials declining as antimicrobial resistance continues to threaten public health, we must look to alternative strategies for the treatment of infections. In this study, we investigated an innovative ‘drug-free’ dual-wavelength irradiation approach that combines two wavelengths of light, 460 nm and 405 nm, against methicillin resistant Staphylococcus aureus (MRSA). MRSA was initially irradiated with 460 nm light (90-360 J/cm2) and subsequently irradiated with aliquots of 405 nm light (54-324 J/cm2). For in vivo studies, mouse skin was abraded and infected with approximately 107 CFU of MRSA and incubated for 3 hours before irradiating with 460 nm (360 J/cm2) and 405 nm (342 J/cm2). Naïve mouse skin was also irradiated to investigate apoptosis. We found that staphyloxanthin, the carotenoid pigment in MRSA cells, promoted resistance to the antimicrobial effects of 405 nm light. In addition, we found that the photolytic effect of 460 nm light on staphyloxanthin attenuated resistance of MRSA to 405 nm light inactivation. Irradiation of 460 nm alone did not elicit any antimicrobial effect on MRSA. In a proof-of-principle mouse skin abrasion infection model, we observed significant inactivation of MRSA by the dual-wavelength irradiation approach. However, when either wavelength of light was administered alone, no significant decrease in bacterial viability was observed. Moreover, exposure of the dual-wavelength irradiation to naïve mouse skin did not result in any visible apoptosis.In conclusion, dual-wavelength irradiation strategy may offer an innovative, effective and safe approach for the treatment of skin infections caused by MRSA.
Leon G. Leanse, Xueping Sharon Goh, Ji-Xin Cheng, David Hooper, Tianhong Dai
There is limited understanding of the role of host metabolism in the pathophysiology of human tuberculosis (TB). Using high resolution metabolomics with an unbiased approach to metabolic pathway analysis, we discovered that the tryptophan pathway is highly regulated throughout the spectrum of TB infection and disease. This regulation is characterized by increased catabolism of tryptophan to kynurenine, which was evident not only in active TB disease, but also in latent TB infection (LTBI). Further, we found that tryptophan catabolism is reversed with effective treatment of both active TB disease and LTBI in a manner commensurate with bacterial clearance. Persons with active TB and LTBI also exhibit increased expression of indoleamine 2,3-dioxygenase-1 (IDO-1), suggesting IDO-1 mediates observed increases in tryptophan catabolism. Together, these data indicate IDO-1-mediated tryptophan catabolism is highly preserved in the human response to Mycobacterium tuberculosis and could be a target for biomarker development as well as host-directed therapies.
Jeffrey M. Collins, Amnah Siddiqa, Dean P. Jones, Ken Liu, Russell R. Kempker, Azhar Nizam, N. Sarita Shah, Nazir Ismail, Samuel G. Ouma, Nestani Tukvadze, Shuzhao Li, Cheryl L. Day, Jyothi Rengarajan, James C. M. Brust, Neel R. Gandhi, Joel D. Ernst, Henry M. Blumberg, Thomas R. Ziegler
Tumor-Associated Macrophages (TAMs) contribute to the maintenance of a strong immunosuppressive environment, supporting tumor progression and resistance to treatment. To date, the mechanisms that drive acquisition of these immunosuppressive features are still poorly defined. Heme oxygenase-1 (HO-1) is the rate-limiting enzyme that catabolizes free heme. It displays important cytoprotective, anti-inflammatory and antioxidant properties. A growing body of evidence suggests that HO-1 may also promote tumor development. Herein, we show that HO-1 is highly expressed in monocytic cells in the tumor microenvironment (TME) once they differentiate into TAMs. Deletion of HO-1 in the myeloid compartment enhances the beneficial effects of a therapeutic antitumor vaccine by restoring CD8 T-cell proliferation and cytotoxicity. We further show that induction of HO-1 plays a major role on monocyte education by tumor cells by modulating their transcriptional and epigenetic programs. These results identify HO-1 as a valuable therapeutic target to reprogram the TME and synergize with current cancer therapies to facilitate antitumoral response.
Emmanuelle Alaluf, Benoît Vokaer, Aurélie Detavernier, Abdulkader Azouz, Marion Splittgerber, Alice Carrette, Louis Boon, Frédérick Libert, Miguel P. Soares, Alain Le Moine, Stanislas Goriely
HIV infection is associated with an increase in the proportion of activated CD8 memory T cells (Tmem) that express CX3CR1, but how these cells are generated and maintained in vivo is unclear. We demonstrate that increased CX3CR1 expression on CD8 Tmem in people living with HIV (PLWH) is dependent on coinfection with human cytomegalovirus (CMV), and CX3CR1+ CD8 Tmem are enriched for a putatively immunosenescent CD57+CD28– phenotype. The cytokine IL-15 promotes the phenotype, survival, and proliferation of CX3CR1+CD57+ CD8 Tmem in vitro, whereas TCR stimulation leads to their death. IL-15-driven survival is dependent on STAT5 and Bcl-2 activity, and IL-15-induced proliferation requires STAT5 and mTORC1. Thus, we identify mechanistic pathways that could explain how “inflammescent” CX3CR1+CD57+ CD8 Tmem dominate the overall memory T cell pool in CMV-seropositive PLWH and that support reevaluation of immune senescence as a nonproliferative dead-end.
Stephen R. Morris, Bonnie Chen, Joseph C. Mudd, Soumya Panigrahi, Carey L. Shive, Scott F. Sieg, Cheryl M. Cameron, David A. Zidar, Nicholas T. Funderburg, Souheil-Antoine Younes, Benigno Rodriguez, Sara Gianella, Michael M. Lederman, Michael L. Freeman
The Wnt/beta-catenin signaling pathway plays an important role in renal development and is re-expressed in the injured kidney and other organs. Beta-catenin signaling is protective in acute kidney injury (AKI) through actions on the proximal tubule, but the current dogma is that Wnt/beta-catenin signaling promotes fibrosis and development of chronic kidney disease (CKD). As the role of proximal tubular beta-catenin signaling in CKD remains unclear, we genetically stabilized (i.e. activated) beta-catenin specifically in murine proximal tubules. Mice with increased tubular beta-catenin signaling were protected in two different murine models of AKI to CKD progression. Oxidative stress, a common feature of CKD, reduced the conventional TCF/LEF-dependent beta-catenin signaling and augmented FoxO3-dependent activity in proximal tubule cells in vitro and in vivo. The protective effect of proximal tubular beta-catenin in renal injury required the presence of FoxO3 in vivo. Furthermore, we identified cystathionine gamma-lyase (CSE) as a novel transcriptional target of beta-catenin/FoxO3 interactions in the proximal tubule. Thus, our studies overturn the conventional dogma about beta-catenin signaling and CKD by showing a protective effect of proximal tubule beta-catenin in CKD and identified a new transcriptional target of beta-catenin/FoxO3 signaling that has therapeutic potential for CKD.
Stellor Nlandu-Khodo, Yosuke Osaki, Lauren Scarfe, Hai-chun Yang, Melanie Phillips-Mignemi, Jane Tonello, Kenyi Saito-Diaz, Surekha Neelisetty, Alla V. Ivanova, Tessa Huffstater, Robert S. McMahon, Makoto M. Taketo, Mark deCaestecker, Balakuntalam S. Kasinath, Raymond C. Harris, Ethan Lee, Leslie Gewin
Insulin receptor signaling is crucial for white adipose tissue (WAT) function. Consequently, lack of insulin receptor (IR) in WAT results in a diabetes-like phenotype. Yet, causes for IR downregulation in WAT of diabetic patients are not well understood. By using multiple mouse models of obesity and insulin resistance, we identify a common downregulation of the IR with a reduction of mRNA expression of the selenoproteins Txnrd3, Sephs2, and Gpx3. Consistently, GPX3 is also decreased in adipose tissue of insulin resistant and obese patients. Inducing Gpx3 expression via selenite treatment enhances IR expression via activation of the transcription factor Sp1 in 3T3-L1 preadipocytes and improves adipocyte differentiation and function. Feeding mice a selenium-enriched high-fat diet alleviates diet-induced insulin resistance with increased insulin sensitivity, decreased tissue inflammation and elevated IR expression in WAT. Again, IR expression correlates positively with Gpx3 expression, a phenotype which is also conserved in humans. Consequently, decreasing GPx3 using siRNA technique reduces IR expression in 3T3-L1 preadipocytes and insulin sensitivity. Overall our data identify GPx3 as a novel regulator of IR expression and insulin sensitivity in adipose tissue.
Robert Hauffe, Vanessa Stein, Chantal Chudoba, Tanina Flore, Michaela Rath, Katrin Ritter, Mareike Schell, Kristina Wardelmann, Stefanie Deubel, Johannes F. Kopp, Maria Schwarz, Kai Kappert, Matthias Blüher, Tanja Schwerdtle, Anna P. Kipp, Andre Kleinridders
Loss of melanocytes is the pathological hallmark of vitiligo, a chronic inflammatory skin depigmenting disorder induced by exaggerated immune response, including autoreactive CD8 T cells producing high levels of type-1 cytokines. However, the interplay between this inflammatory response and melanocyte disappearance remains to be fully characterized. Here, we demonstrate that vitiligo skin contains a significant proportion of suprabasal melanocytes, associated with disruption of E-cadherin expression, a major protein involved in melanocyte adhesion. This phenomenon is also observed in lesional psoriatic skin. Importantly, apoptotic melanocytes were mainly observed once cells were detached from the basal layer of the epidermis, suggesting that additional mechanism(s) could be involved in melanocyte loss. The type-1 cytokines IFNg and TNFa induce melanocyte detachment through E-cadherin disruption, and the release of its soluble form, partly due to the matrix metalloproteinase MMP-9. MMP-9, whose levels are increased in vitiligo skin and patients’ sera, is produced by keratinocytes in response to IFNg and TNFa. Inhibition of MMP-9 or the JAK/STAT signaling pathway prevents melanocyte detachment in vitro and in vivo. Therefore, stabilization of melanocytes in the basal layer of the epidermis by preventing E-cadherin disruption appears promising to prevent the depigmentation occurring in vitiligo and during chronic skin inflammation.
Nesrine Boukhedouni, Christina Martins, Anne-Sophie Darrigade, Claire Drullion, Jérôme Rambert, Christine Barrault, Julien Garnier, Clement Jacquemin, Denis Thiolat, Fabienne Lucchese, Franck Morel, Khaled Ezzedine, Alain TAIEB, François-Xavier Bernard, Julien Seneschal, Katia Boniface
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