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Abstract
BACKGROUND. Urine proteomics may provide mechanistic insights on why patients experience a higher risk of kidney function decline after hospitalization. METHDOS. In 174 patients with and without acute kidney injury (AKI) from the Assessment, Serial Evaluation, and Subsequent Sequelae in AKI (ASSESS-AKI) cohort, we used Olink to profile 2783 urine proteins collected at 3 months post-hospitalization and determined their association with estimated glomerular filtration rate (eGFR) decline during median [IQR] of 5.1[4.0-6.0] years follow-up. In four independent cohorts including the Kidney Precision Medicine Project (KPMP), we determined if proteins were differentially expressed with AKI. We used weighted correlation network analysis to determine proteins’ cellular enrichment in the kidney transcriptome (single-cell and spatial transcriptomics) in patients with AKI receiving research kidney biopsy. RESULTS. We identified 387 and 10 proteins associated with faster and slower eGFR decline, respectively, most of which were differentially expressed in patients at the time of AKI. Among these proteins, 283 (71%) were expressed by kidney cells in participants with AKI from KPMP. The expression formed 3 clusters enriched in the proximal tubule, degenerative tubule and myeloid cells, and stromal cells, and correlated with histopathological features of AKI, such as tubular injury, interstitial inflammation, and fibrosis, respectively. CONCLUSION. Urinary proteins reflecting degenerative tubular injury, inflammation, and fibrosis are associated with eGFR decline in recently hospitalized patients. FUNDING. The Kidney Precision Medicine Project (KPMP) is supported by the National Institute of Diabetes and Digestive Kidney Diseases (NIDDK) through the following grantsU01DK133081, U01DK133091, U01DK133092, U01DK133093, U01DK133095, U01DK133097, U01DK114866, U01DK114908, U01DK133090, U01DK133113, U01DK133766, U01DK133768, U01DK114907, U01DK114920, U01DK114923, U01DK114933, U24DK114886, UH3DK114926, UH3DK114861, UH3DK114915, and UH3DK114937 We gratefully acknowledge the essential contributions of our patient participants and support of the American public though their tax dollars. SM is supported by NIDDK Grant K23DK128358.
Authors
Yumeng Wen, Steven Menez, Heather Thiessen Philbrook, Dennis Moledina, Steven G. Coca, Jiashu Xue, James Kaufman, Vernon Chinchillil, Paul L. Kimmel, T. Alp Ikizler, Chi-yuan Hsu, Tanika Kelly, Ana Ricardo, Jonathan Himmelfarb, Chirag R. Parikh
Abstract
FOXP3+ Treg cells are critical for immune tolerance. Genetic deletion of the Forkhead domain containing proteins of the FOXP-subfamily member FOXP1 from Tregs results in impaired function associated with reduced CD25 expression and IL-2 signaling, but to date the only other FOXP family member expressed in Tregs, FOXP4, has been minimally studied. To investigate the potential functional interactions among FOXP family members in Treg cells, we specifically deleted Foxp1, Foxp4 or both in FOXP3+ committed Treg cells in mice. Our findings show that mice with combined, but not individual, deficiency in FOXP1 and FOXP4 exhibit lymphoproliferation, inflammation, autoimmunity, and early lethality. The combined absence of FOXP1 and FOXP4 in Tregs results in an activated/effector-like phenotype with compromised suppressive function in peripheral lymphoid organs, an enhanced germinal center response and proinflammatory cytokine production. We further show that FOXP1 and FOXP4 bind to Il2ra promoter regions to regulate CD25 expression in Tregs. Through pairwise comparison among mouse strains with Treg specific deletion of Foxp1, Foxp4 or both, our findings indicate a non-redundant but insufficient role of FOXP4 in Treg cell function.
Authors
Dachuan Dong, Vishal J. Sindhava, Ananthakrishnan Ganesan, Martin S. Naradikian, Tom L. Stephen, Andrew Frisch, Kristen M. Valentine, Elizabeth Buza, Karla R. Wiehagen, Michael P. Cancro, Edward E. Morrisey, Haley Tucker, Katrina K. Hoyer, Purvesh Khatri, Jonathan S. Maltzman
Abstract
BACKGROUND. Glucagon-like peptide-2 (GLP-2) analogs are used clinically to enhance nutrient absorption in patients with short bowel syndrome (SBS); however, the precise mechanism remains unclear. To address this, the study aimed to clarify the dynamics of intestinal epithelial cells and immune cells in patients with SBS treated with GLP-2 analogs. METHODS. Five male patients diagnosed with SBS, all of whom received treatment with the GLP-2 analog teduglutide, were included in the study. We conducted longitudinal single-cell RNA sequencing (scRNA-seq) analysis of intestinal tissue from SBS patients over a year, integrating microbiome composition analysis. RESULTS. After treatment, the alpha diversity of the gut microbiome increased, indicating a more varied microbial environment. ScRNA-seq analysis revealed a reduction of T helper 2 cells and an increase in regulatory T (Treg) cells, suggesting a shift towards an immunoregulatory intestinal environment. Additionally, nutrient-absorbing enterocyte-Top2 and middle clusters expanded, enhancing the absorption capacity, whereas major histocompatibility complex class I/II-expressing enterocyte-Top1 cells declined, potentially modulating immune responses. CONCLUSION. The study findings indicate that GLP-2 analogs reshape intestinal immunity and microbiota, fostering a less inflammatory environment while promoting nutrient uptake efficiency. These insights offer a deeper understanding of the role of GLP-2 analogs in gut adaptation and provide a foundation for refining clinical strategies for SBS treatment. FUNDING. This work was supported by Sakaguchi Memorial Foundation, Grants-in-Aid from the Japanese Society for the Promotion of Science (JSPS) (21K18272, 23H03665, 23H02899, 23K27590, 25K22627, 23K08037), JST FOREST(21457195), and the Takeda Japan Medical Office Funded Research Grant 2022.
Authors
Yumi Kudo, Kentaro Miyamoto, Shohei Suzuki, Akihiko Chida, Anna Tojo, Mai Hasegawa, Arina Shigehara, Ikuko Koya, Yoshinari Ando, Masayasu Sato, Aya Kondo, Tomoko Kumagai, Harunori Deguchi, Yoshiki Sugiyama, Yoko Ito, Koji Shirosaki, Satoko Yamagishi, Yutaro Maeda, Hiroki Kanamori, Motohiro Kano, Mototoshi Kato, Hanako Tsujikawa, Yusuke Yoshimatsu, Kaoru Takabayashi, Koji Okabayashi, Takanori Kanai, Naoki Hosoe, Motohiko Kato, Jonathan Moody, Chung-Chau Hon, Tatsuo Kuroda, Yohei Yamada, Akihiro Fujino, Tomohisa Sujino
Abstract
Pathological cardiac remodeling is associated with the reactivation of fetal genes, yet the extent of the heart’s fetal gene program and its impact on proteome compositions remain incompletely understood. Here, using a new proteome-wide protein ratio quantification strategy with mass spectrometry, we identify pervasive isoform usage shifts in fetal and postnatal mouse hearts, involving 145 pairs of highly homologous paralogs and alternative splicing-derived isoform proteins. Proteome-wide ratio comparisons readily rediscover hallmark fetal gene signatures in muscle contraction and glucose metabolism pathways, while revealing novel isoform usage in mitochondrial and gene expression proteins, including PPA1/PPA2, ANT1/ANT2, and PCBP1/PCBP2 switches. Paralogs with differential fetal usage tend to be evolutionarily recent, consistent with functional diversification. Alternative splicing adds another rich source of fetal isoform usage differences, involving PKM M1/M2, GLS-1 KGA/GAC, PDLIM5 long/short, and other spliceoforms. When comparing absolute protein proportions, we observe a partial reversion toward fetal gene usage in pathological hearts. In summary, we present a ratiometric catalog of paralogs and spliceoform pairs in the cardiac fetal gene program. More generally, the results demonstrate the potential of applying the proteome-wide ratio test concept to discover new regulatory modalities beyond differential gene expression.
Authors
Yu Han, Shaonil Binti, Sara A. Wennersten, Boomathi Pandi, Dominic C.M. Ng, Edward Lau, Maggie P.Y. Lam
Abstract
Anemia is a common and disabling complication of chronic kidney disease (CKD). Current therapies can be burdensome, and full correction of anemia is limited by cardiovascular side effects. New approaches that may offer additional therapeutic options are needed. We explored the anti-anemic effects of erythroferrone, an erythroid hormone that induces iron mobilization by suppressing the master iron-regulatory hormone hepcidin. In a preclinical murine model of adenine-induced CKD, transgenic augmentation of erythroferrone mobilized iron, increased hemoglobin concentrations by approximately 2 g/dl, and modestly improved renal function without affecting systemic or renal inflammation, fibrosis, or markers of mineral metabolism. This study supports the concept that therapeutic augmentation of erythroferrone is a promising approach for alleviating CKD-associated anemia.
Authors
Brian Czaya, Joseph D. Olivera, Moya Zhang, Amber Lundin, Christian D. Castro Andrade, Grace Jung, Mark R. Hanudel, Elizabeta Nemeth, Tomas Ganz
Abstract
Regulatory T cells (Tregs) are essential for peripheral tolerance and depend on TCR and IL-2R signaling for their homeostasis and function. In mice, IL-2-dependent BLIMP-1 contributes to Treg homeostasis. BLIMP-1 is a major transcriptional hub in human Tregs, but its mechanisms of action remain undefined. Here, using CRISPR/Cas9 ablation, we show that BLIMP-1 limits human Treg proliferation, but supports IL-10, CTLA4, several immune checkpoints, including CEACAM1, and Treg functional activity. BLIMP-1 restrains Treg expansion to IL-2 by downregulating CD25 and IL-2R signaling, and by enhancing CEACAM1 expression, which in turn inhibits responsiveness to CD3/CD28 signaling and activation of mTOR. Prolonged IL-2R signaling optimizes BLIMP-1 expression, supporting chromosomal opening of CEACAM1 to increased CEACAM1 expression through STAT5- and BLIMP-1-driven enhancers. Correspondingly, CEACAM1 is highly induced on Tregs from autoimmune patients undergoing low-dose IL-2 therapy, and these Tregs showed reduced proliferation. A humanized mouse model of xenogeneic graft versus host disease demonstrates that BLIMP-1 normally promotes, while CEACAM1 restrains, Treg suppressive activity. Collectively, our findings reveal that BLIMP-1 and CEACAM1 function in an IL-2-dependent feedback loop to restrain Treg proliferation and affect suppressive function. CEACAM1 also acts as a highly selective biomarker of IL-2R signaling in human T cells.
Authors
Ying Ding, Aixin Yu, Milos Vujanac, Sabrina N. Copsel, Alejandro Moro, Luis Nivelo, Molly Dalzell, Nicolas Tchitchek, Michelle Rosenzwajg, Alejandro V. Villarino, Robert B. Levy, David Klatzmann, Thomas R. Malek
Abstract
Bronchopulmonary dysplasia (BPD), a prevalent and chronic lung disease affecting premature newborns, results in vascular rarefaction and alveolar simplification. Although the vasculature has been recognized as a main player in this disease, the recently found capillary heterogeneity and cellular dynamics of endothelial subpopulations in BPD remain unclear. Here, we show Cap2 cells are damaged during neonatal hyperoxic injury, leading to their replacement by Cap1 cells which, in turn, significantly decline. Single cell RNA-seq identifies the activation of numerous p53 target genes in endothelial cells (ECs), including Cdkn1a (p21). While global deletion of p53 results in worsened vasculature, endothelial-specific deletion of p53 reverses the vascular phenotype and improves alveolar simplification during hyperoxia. This recovery is associated with the emergence of a transitional EC state, enriched for oxidative stress response genes and growth factors. Notably, this transitional EC gene signature is conserved in an aberrant capillary population identified in human BPD with pulmonary hypertension, underscoring the biological and clinical relevance of our findings. These results reveal a key role for p53 in maintaining endothelial lineage fidelity during pulmonary capillary repair following hyperoxic injury and highlight the critical contribution of the endothelium to BPD pathogenesis.
Authors
Lisandra Vila Ellis, Jonathan D. Bywaters, Amanda Ceas, Yun Liu, Jennifer M.S. Sucre, Jichao Chen
Abstract
Biallelic variants in SCN1B, encoding the voltage-gated sodium channel β1/β1B subunits, are linked to DEE52, a developmental and epileptic encephalopathy with a high risk of Sudden Unexpected Death in Epilepsy (SUDEP). DEE52 patients present clinically with Dravet syndrome or the more severe early infantile DEE. SCN1B is expressed in brain and heart in humans and in mice. Thus, we have proposed that, in addition to generalized seizures, cardiac arrhythmia may play a role in SUDEP. Mice with homozygous expression of the DEE52 variant Scn1b-c.265C>T, predicting p.R89C, have spontaneous and hyperthermia-induced generalized seizures and SUDEP. Here we conducted cardiac characterization of Scn1b-c.265C>T mice and studied induced pluripotent stem cell cardiomyocytes (iPSC-CMs) derived from two SCN1B-c.265C>T DEE52 patients. Scn1bC89/C89 mouse CMs showed increased transient outward potassium current (Ito) density and heart sections revealed ventricular fibrosis. Scn1bC89/C89 mice were susceptible to pacing-induced cardiac arrhythmias. Patient-derived iPSC-CMs with biallelic SCN1B-c.265C>T variant expression showed increased sodium current (INa), late INaL, and Ito current densities. We conclude that, while mouse and human cardiac AP waveforms have critical differences, increased Ito is common to both models of DEE52. Overall, our data suggest that electrical and structural substrates may lead to arrhythmias and contribute to SUDEP in DEE52.
Authors
Roberto Ramos-Mondragon, Shuyun Wang, Nnamdi Edokobi, Qinghua Liu, Xiaotan Qiao, Maya Shih, Louis T. Dang, Yao-Chang Tsan, Katalin Štěrbová, Adam S. Helms, Sarah Weckhuysen, Luis F. Lopez-Santiago, Jack M. Parent, Lori L. Isom
Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutations in PKD1 or PKD2, which encode polycystin-1 (PC1) and polycystin-2 (PC2) respectively. These proteins are thought to form a signaling complex that can flux cations including calcium. One of the earliest symptoms in ADPKD is a decline in the concentrating ability of the kidneys, occurring prior to cyst formation. We reasoned that hyperosmolality stimulates the polycystin complex, and that the loss of this function impairs water reabsorption. We found that hyperosmolality resulted in the phosphorylation of a microtubule associated protein 4 (MAP4) in a PC1-dependent manner which then elicited ER-localized PC2 calcium signals. ER-localized PC2 hyperosmotic calcium signals were required for trafficking of the water channel aquaporin (AQP2). Pre-cystic PC1-KO and PC2-KO murine kidneys had cytosolic localized AQP2, and diluted urine compared to their respective controls. Kidney tissue sections from ADPKD patients showed decreased AQP2 apical membrane localization in cystic and non-cystic tubules. Our study demonstrates that osmolality is a physiological stimulus of the polycystin complex, and loss of polycystin osmosensing results in impaired water reabsorption via AQP2. This likely contributes to the declined concentrating ability of the kidneys and high circulating vasopressin levels in ADPKD patients.
Authors
Karla M. Márquez-Nogueras, Ryne M. Knutila, Virdjinija Vuchkovska, Charlie Yang, Patricia Outeda, Darren P. Wallace, Ivana Y. Kuo
Abstract
Radiation-induced lymphopenia (RIL) remains a challenging side effect of radiation therapy, often associated with poor prognosis and reduced overall survival. Although CD8+ T cells are highly radiosensitive, the dynamics of quantitative and qualitative changes to the CD8 T cell pool following exposure to high doses of ionizing radiation (IR) remains understudied. Herein, we sought to determine the long-term impact of sublethal whole body irradiation (WBI) on antigen (Ag)-inexperienced CD8 T cell pool, comprised of naïve (TN) and virtual memory (TVM) CD8+ T cells. We show that although both TN and TVM cells gradually regenerate after WBI-induced loss, TN recovery only occurs through de novo thymic production. Despite the numerical restoration, the subset and phenotypic composition of post-recovery Ag-inexperienced CD8+ T cells do not qualitatively recapitulate the pre-WBI state. Specifically, the frequency of TVM cells is increased, especially during the early stages of recovery. Within the TN subset, a lasting overrepresentation of Ly6C+CD122+ cells and an altered TCR clonotype diversity are also observed. Overall, our data highlight the dynamic changes to the Ag-inexperienced CD8+ T cell pool upon recovery from RIL.
Authors
Mohammad Heidarian, Shravan K. Kannan, Whitney Swanson, Thomas S. Griffith, John T. Harty, Vladimir P. Badovinac
Abstract
Infections with non-tuberculous mycobacterium (NTM) are on the rise. Here, we investigated an uncommon NTM infection, by M. haemophilum (Mh, n = 3), from a shared geographic location in the USA. All patients had underlying immunosuppressive conditions or treatments. We identified that all these individuals had a non-synonymous mutation in GATA2 gene, which was absent in Healthy controls (HCs, n = 4) from the same geographic area (Missouri, USA). Whole blood from these individuals had attenuated cytokine responses to Mh stimulation for IL1β, IL-6, IL-8, MIP-1α and β, but not to Phytohemagglutinin (PHA) or another NTM, M. abscessus. Impaired whole blood transcriptional responses in individuals with GATA2 mutation, included heightened Ras-homolog (Rho) guanosine triphosphate hydrolases (GTPase) and lowered Transforming growth factor (TGF)-β responses among others. Our results highlight that comparatively, M. abscessus and Mh elicit differential immune responses in humans, and we identify a 23-gene signature that distinguishes host response to Mh and M. abscessus and show that in vitro GATA2 siRNA knockdown indeed attenuates cytokine responses to Mh. Thus, we provide new evidence that links GATA2 mutation and immune dysfunction in individuals with compromised immunity to Mh infection in humans and outline host factors associated with the immune response of this clinically relevant NTM.
Authors
Ananya Gupta, Shail B. Mehta, Abhimanyu A, Bruce A. Rosa, John Martin, Mushtaq Ahmed, Shyamala Thirunavukkarasu, Farheen Fatma, Gaya Amarsinghe, Makedonka Mitreva, Thomas C. Bailey, David B. Clifford, Shabaana A. Khader
Abstract
Transforming growth factor beta (TGF-β) signaling is the master modulator of renal fibrosis. However, targeting drugs have failed to prevent the progression of chronic kidney disease (CKD) in clinical trials due to the extensive biological regulation of TGF-β signaling. It is necessary to investigate the precise downstream mechanisms of TGF-β signaling that regulate renal fibrosis. In this study, we found that PR-domain containing 16 (PRDM16) expression in human renal tubular epithelial cells was markedly reduced by TGF-β. Mechanistically, activated Smad3 induced by TGF-β interacted with the cofactor H-Ras and bound to the promoter of PRDM16, downregulating its transcription. Tubular-specific knockout of PRDM16 promoted renal fibrosis in models of unilateral ureteral occlusion (UUO) and unilateral ischemia-reperfusion injury (UIRI) by exacerbating mitochondrial dysfunction. In vitro, PRDM16 blocked TGF-β-induced mitochondrial injury and lipid deposition by upregulating Peroxisome Proliferator-Activated Receptor Gamma Coactivator-1α (PGC-1α). Delivery of the exogenous PRDM16 gene preserved renal function and ameliorated the progression of renal fibrosis by protecting mitochondrial function. We report PRDM16 as a novel downstream target of TGF-β signaling that attenuates renal fibrosis by safeguarding tubular mitochondrial function.
Authors
Qian Yuan, Ben Tang, Yuting Zhu, Chao Wan, Yaru Xie, Yajuan Xie, Cheng Wan, Hua Su, Youhua Liu, Chun Zhang
Abstract
Spondyloarthritis (SpA) is an inflammatory arthritis of the spine and joints associated with intestinal inflammation, in which it is hypothesized that innate immune exposure to entero-invasive species is followed by self/bacterial peptide presentation. However, the mechanisms underlying loss of tolerance to gut bacteria in genetically at-risk individuals are unclear. Curdlan (β-1,3-glucan, dectin-1 ligand)-treated ZAP-70W163C (SKG) mice develop autoimmune arthritis and ileitis associated with Gram-negative faecal dysbiosis. Using gnotobiotic mice, we show that curdlan-treated SKG mice mono-associated with Parabacteroides goldsteinii or Lactobacillus murinus developed ileitis, arthritis and enthesitis, while BALB/c mice were tolerant. Gnotobiotic SKG ileum upregulated Il23a and ER stress genes and lost goblet cells. Whereas bacterial DNA co-localised with neutrophils and inflammatory macrophages in SKG lamina propria, peri-articular bone marrow, entheses and spleen, in BALB/c bacterial DNA co-localised with resident macrophages in lamina propria and spleen. Human psoriatic-arthritis synovial tissue also contained cell-associated peri-vascular bacterial DNA. Curdlan-treated SKG spleen/bone marrow macrophages transferred severe arthritis and expanded Th17 cells in naïve SKG recipients, while BALB/c or germ free-SKG macrophages transferred mild arthritis and regulated Th17 cells. Thus, bacterial DNA and myeloid cells in the gut and their subsequent traffic regulate or enforce T cell pathogenicity in SpA.
Authors
Benjamin Cai, Rabina Giri, Amy J. Cameron, M. Arifur Rahman, Annabelle Small, Christopher Altmann, Yenkai Lim, Linda M. Rehaume, Mark Morrison, Mihir D. Wechalekar, Jakob Begun, Anne-Sophie Bergot, Ranjeny Thomas
Abstract
Reprogramming autoreactive CD4⁺ effector T (Teff) cells into immunosuppressive regulatory T (Treg) cells represents a promising strategy for treating established autoimmune diseases. However, the stability and function of such reprogrammed Tregs under inflammatory conditions remain unclear. Here, we show that epigenetic activation of core Treg identity genes in Teff cells yields lineage-stable Effector T cell Reprogrammed Tregs (ER-Tregs). A single adoptive transfer of ER-Tregs not only prevents autoimmune neuroinflammation in mice when given before disease onset but also arrests its progression when administered after onset. Compared to Foxp3 overexpressing Teff cells, induced Tregs from naïve precursors, and endogenous Tregs, ER Tregs provide superior protection against autoimmune neuroinflammation. This enhanced efficacy stems from their inherited autoantigen specificity and selectively preserved effector cell transcriptional programs, which together bolster their fitness in inflammatory environments and enhance their suppressive capacity. Our results establish epigenetic reprogramming of autoreactive Teff cells as an effective approach to generate potent, stable Tregs for the treatment of refractory autoimmune conditions.
Authors
Tyler R. Colson, James J. Cameron, Hayley I. Muendlein, Mei-An Nolan, Jamie L. Leiriao, James H. Kim, Alexander N. Poltorak, Xudong Li
Abstract
Authors
Veethika Pandey, Heike R. Doeppler, Ligia I. Bastea, Alicia K. Fleming Martinez, Barath Shreeder, Brandy H. Edenfield, Keith L. Knutson, DeLisa Fairweather, Peter Storz
Abstract
Glycogen storage disease type Ia (GSD Ia) is caused by a deficiency of glucose-6-phosphatase (G6Pase) in the liver leading to lethal hypoglycemia. Gene therapy with adeno-associated virus (AAV) vectors encoding G6Pase fails to stably treat GSD Ia early in life. We evaluated genome editing in 12 day-old infant mice with GSD Ia using two AAV vectors, one containing Cas9 from Streptococcus pyogenes and a second Donor vector that expresses a guide RNA and a G6PC transgene. Gene therapy with the Donor vector only was compared with genome editing using both Donor and CRISPR vectors. Treatment with genome editing (total vector dose 0.2 to 2E+13 vector genomes/kg) and bezafibrate (to stimulate autophagy) was efficacious as assessed by hypoglycemia prevention and the frequency of transgene integration, which correlated with improved survival. This therapy achieved 5.9% chromosomal transgene integration through homology directed repair, which surpassed a threshold to prevent long-term hepatic complications. No integration was detected in absence of the CRISPR vector. Importantly for safety, CRISPR vector genomes were depleted, and no intact, integrated CRISPR genomes were detected by long-read sequencing. Thus, genome editing warrants further development as a potentially stable treatment for human infants with GSD Ia.
Authors
Benjamin Arnson, Ekaterina Ilich, Troy von Beck, Songtao Li, Elizabeth D. Brooks, Dorothy Gheorghiu, Gordon He, Matthew Weinrub, Sze Ying Chan, Hye-Ri Kang, David Courtney, Jeffrey Everitt, Bryan R. Cullen, Dwight D. Koeberl
Low‐intensity pulsed ultrasound stimulation to treat renal fibrosis through inhibiting tubular IL-1R
Abstract
Low-intensity pulsed ultrasound stimulation (LIPUS) has become increasingly appreciated for its therapeutic effect on kidney diseases. However, its role and biological mechanism in treating chronic kidney disease (CKD) remain poorly defined. Here, we revealed that LIPUS was applied in a safe range with an intensity of 25-315 mW/cm2. Daily LIPUS at an intensity of 315 mW/cm2 ameliorated ischemia-reperfusion (IR)-induced tubular injury and renal fibrosis, accompanied by the remarkable downregulation of IL-1R. Transcriptome sequencing showed that LIPUS significantly down-regulated IL-1R and its downstream genes in IL-1β-stimulated IR-injured mice. LIPUS effectively reversed IL-1β-induced tubular injury and reduced the production of profibrotic cytokines by down-regulating IL-1R in vivo and in vitro. Renal proximal tubule-specific Il1r1 knockout mice exhibited milder renal tubular injury and fibrosis after IR injury. However, LIPUS did not ameliorate IR injury in proximal tubule-specific Il1r1 knockout mice. Collectively, daily LIPUS at an intensity of 315 mW/cm2 relieves IR-induced tubular injury and fibrosis, potentially through down-regulating tubular IL-1R.
Authors
Zhimin Huang, Jiaxin Dong, Ziqi Fu, Li Li, Simeng liu, Lin Wu, Honglei Guo, Ao Bian, Kang Liu, Wei Sun, Changying Xing, Steven D. Crowley, Jiafa Ren, Xiangqing Kong, Huijuan Mao
Abstract
Autophagy is a recycling pathway in which damaged proteins, protein aggregates, and organelles are delivered to lysosomes for degradation. Autophagy insufficiency is thought to contribute to osteoporosis. Accordingly, autophagy elimination from the osteoblast lineage reduces bone formation and bone mass. However, whether increasing autophagy would benefit bone health is unknown. Here, we increased expression of endogenous transcription factor EB gene (Tfeb) in osteoblast lineage cells in vivo via CRISPR activation (TfebCRa mice). Elevated Tfeb stimulated autophagy and lysosomal biogenesis in osteoblasts. TfebCRa mice displayed a robust increase in femoral and vertebral cortical thickness at 4.5 months of age. Increases in cortical thickness was due to increased periosteal bone formation. Tfeb elevation also increased femoral trabecular bone volume. These changes increased bone strength of TfebCRa mice. Female TfebCRa mice displayed a progressive increase in bone mass and at 12 months of age had high cortical thickness and trabecular bone volume. Increased vertebral trabecular bone volume was due to elevated bone formation. Osteoblastic cultures showed that Tfeb elevation increased proliferation and mineral deposition. Overall, these results demonstrate TFEB-driven stimulation of autophagy in osteoblast lineage cells is associated with increased bone formation and strength and may represent an effective approach to combat osteoporosis.
Authors
Alicen James, James A. Hendrixson, Ilham Kadhim, Adriana Marques-Carvalho, Jacob Laster, Julie Crawford, Jeff Thostenson, Visanu Wanchai, Amy Y. Sato, Intawat Nookaew, Jinhu Xiong, Maria Almeida, Melda Onal
Abstract
The tumor microenvironment (TME) significantly impacts cancer progression, yet traditional animal models do not fully recapitulate the situation in humans. To address this, we developed tumor-derived precision lung slices (TD-PCLS), an ex vivo platform for studying the lung TME and evaluating therapies. TD-PCLS, viable for 8 to 10 days, preserve the heterogeneity and metabolic activity of primary tumors, as confirmed by seahorse analysis. Using multispectral FACS and phenocycler multiplex imaging, we spatially profiled TME components and cancer cell functionality. Additionally, TD-PCLS revealed patient-specific responses to chemo- and immunotherapies. To complement TD-PCLS, we established tumor-cell-seeded PCLS (TCS-PCLS) by introducing tumor and immune cells into healthy lung slices. This model highlighted macrophage-tumor interactions as critical for tumor cell proliferation, migration, and immune modulation. Together, these platforms provide a robust tool for lung cancer research, enabling precision medicine and advancing therapeutic discovery.
Authors
Siavash Mansouri, Annika Karger, Clemens Ruppert, Marc A. Schneider, Andreas Weigert, Rajender Nandigama, Blerina Aliraj, Lisa Strotmann, Anoop V. Cherian, Diethard Pruefer, Peter Dorfmuller, Ludger Fink, Ibrahim Alkoudmani, Stefan Gattenlöhner, Bastian Eul, Andre Althoff, Peter Kleine, Hauke Winter, Andreas Guenther, Ardeschir Ghofrani, Soni S. Pullamsetti, Friedrich Grimminger, Werner Seeger, Rajkumar Savai
Abstract
Secondary lymphedema is characterized by fibrosis and impaired lymphatic function. Although TGF-β is a key regulator of fibrosis in this disease, the cellular mechanisms regulating this process remain unknown. Epithelial–mesenchymal transition (EMT), a mechanism by which TGF-β induces fibrosis in other skin diseases, is characterized by loss of epithelial cell markers and cellular polarity, upregulation of fibrotic gene expression, and gain of migratory capacity. Using clinical lymphedema biopsy specimens and animal models, we show that keratinocytes in the basal layer of the epidermis undergo EMT in lymphedematous skin, migrate into the dermis, and contribute to dermal fibrosis. In vitro studies using cultured primary human keratinocytes treated with lymphatic fluid from the affected limbs of patients with secondary lymphedema resulted in a TGF-β–mediated increased expression of EMT markers. We show for the first time that EMT is activated by TGF-β in secondary lymphedema and that this process plays an important role in regulating skin fibrosis in this disease.
Authors
Hyeung Ju Park, Jinyeon Shin, Ananta Sarker, Mark G. Klang, Elyn Riedel, Michelle Coriddi, Joseph H. Dayan, Sarit Pal, Babak J. Mehrara, Raghu P. Kataru
