RESUMO
BACKGROUND: C3 Glomerulopathy (C3G) is a rare glomerular disease caused by dysregulation of the complement pathway. Based on its pathophysiology, treatment with the monoclonal antibody eculizumab targeting complement C5 may be a therapeutic option. Due to the rarity of the disease, observational data on the clinical response to eculizumab treatment is scarce. METHODS: Fourteen patients (8 female, 57%) treated for C3 glomerulopathy at the medical center of the University of Freiburg between 2013 and 2022 were included. Subjects underwent biopsy before enrollment. Histopathology, clinical data, and response to eculizumab treatment were analyzed. Key parameters to determine the primary outcome were changes of estimated glomerular filtration rate (eGFR) over time. Positive outcome was defined as > 30% increase, stable outcome as ±30%, negative outcome as decrease > 30% of eGFR. RESULTS: Eleven patients (78.8%) were treated with eculizumab, three received standard of care (SoC, 27.2%). Median follow-up time was 68 months (IQR: 45-98 months). Median eculizumab treatment duration was 10 months (IQR 5-46 months). After eculizumab treatment, five patients showed a stable outcome, six patients showed a negative outcome. Among patients receiving SoC, one patient showed a stable outcome, two patients showed a negative outcome. CONCLUSIONS: The benefit of eculizumab in chronic progressive C3 glomerulopathy is limited.
Assuntos
Inativadores do Complemento , Glomerulonefrite Membranoproliferativa , Feminino , Humanos , Complemento C3/análise , Glomerulonefrite Membranoproliferativa/tratamento farmacológico , Proteinúria/etiologia , Estudos Retrospectivos , Resultado do Tratamento , Inativadores do Complemento/uso terapêutico , MasculinoRESUMO
The primary cilium is a sensory organelle at the cell surface with integral functions in cell signaling. It contains a microtubular axoneme that is rooted in the basal body (BB) and serves as a scaffold for the movement of intraflagellar transport (IFT) particles by Kinesin-2 along the cilium. Ift88, a member of the anterograde moving IFT-B1 complex, as well as the Kinesin-2 subunit Kif3a are required for cilia formation. To facilitate signaling, the cilium restricts the access of molecules to its membrane ("ciliary gate"). This is thought to be mediated by cytoskeletal barriers ("subciliary domains") originating from the BB subdistal/distal appendages, the periciliary membrane compartment (PCMC) as well as the transition fibers and zone (TF/TZ). The PCMC is a poorly characterized membrane domain surrounding the ciliary base with exclusion of certain apical membrane proteins. Here we describe that Ift88, but not Kinesin-2, is required for the establishment of the PCMC in MDCK cells. Likewise, in C. elegans mutants of the Ift88 ortholog osm-5 fail to establish the PCMC, while Kinesin-2 deficient osm-3 mutants form PCMCs normally. Furthermore, disruption of IFT-B1 into two subcomplexes, while disrupting ciliogenesis, does not interfere with PCMC formation. Our findings suggest that cilia are not a prerequisite for the formation of the PCMC, and that separate machineries with partially overlapping functions are required for the establishment of each.
Assuntos
Membrana Celular/metabolismo , Cílios/metabolismo , Células Epiteliais/metabolismo , Cinesinas/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Animais , Corpos Basais/metabolismo , Caenorhabditis elegans/citologia , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Citoesqueleto/metabolismo , Cães , Células Madin Darby de Rim Canino , Microscopia de Fluorescência , Proteínas do Tecido Nervoso/metabolismo , Transdução de SinaisRESUMO
The transcription factor NRF2 plays a key role in the protection against environmental stress and maintaining cellular homeostasis. The acetyltransferase p300 is a known component of the NRF2 transcriptional complex and promotes its transcriptional activity. In this study we describe a novel mechanism by which p300 facilitates NRF2 activity. p300 physically interacts with NRF2 and interferes with NRF2-KEAP1 complex formation. In particular, p300 increases NRF2 protein abundance and stability, thereby promoting NRF2 nuclear localization. Notably, the acetyltransferase activity of p300 was indispensable for the stabilizing effects towards NRF2. Furthermore, overexpression of p300 protected HEK293T cells from oxidative stress and increased viability. Together our study uncovers a link between p300 and control of NRF2-KEAP1 signaling via regulation of NRF2 stability and this may act as a novel checkpoint on the adaptation to oxidative stress.
Assuntos
Regulação da Expressão Gênica , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Fator 2 Relacionado a NF-E2/genética , Fatores de Transcrição de p300-CBP/genética , Adaptação Fisiológica , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Peróxido de Hidrogênio/farmacologia , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo , Ligação Proteica , Estabilidade Proteica , Transporte Proteico , Transdução de Sinais , Transcrição Gênica , Fatores de Transcrição de p300-CBP/deficiênciaRESUMO
BACKGROUND: Critically ill coronavirus disease 2019 (COVID-19) patients have a high risk of acute kidney injury (AKI) that requires renal replacement therapy (RRT). A state of hypercoagulability reduces circuit life spans. To maintain circuit patency and therapeutic efficiency, an optimized anticoagulation strategy is needed. This study investigates whether alternative anticoagulation strategies for RRT during COVID-19 are superior to administration of unfractionated heparin (UFH). METHODS: Retrospective cohort study on 71 critically ill COVID-19 patients (≥18 years), admitted to intensive care units at a tertiary health care facility in the southwestern part of Germany between February 26 and May 21, 2020. We collected data on the disease course, AKI, RRT, and thromboembolic events. Four different anticoagulatory regimens were administered. Anticoagulation during continuous veno-venous hemodialysis (CVVHD) was performed with UFH or citrate. Anticoagulation during sustained low-efficiency daily dialysis (SLEDD) was performed with UFH, argatroban, or low molecular weight heparin (LMWH). Primary outcome is the effect of the anticoagulation regimen on mean treatment times of RRT. RESULTS: In patients receiving CVVHD, mean treatment time in the UFH group was 21.3 h (SEM: ±5.6 h), in the citrate group 45.6 h (SEM: ±2.7 h). Citrate anticoagulation significantly prolonged treatment times by 24.4 h (P = .001). In patients receiving SLEDD, mean treatment time with UFH was 8.1 h (SEM: ±1.3 h), with argatroban 8.0 h (SEM: ±0.9 h), and with LMWH 11.8 h (SEM: ±0.5 h). LMWH significantly prolonged treatment times by 3.7 h (P = .008) and 3.8 h (P = .002), respectively. CONCLUSIONS: UFH fails to prevent early clotting events in the dialysis circuit during COVID-19. For patients, who do not require effective systemic anticoagulation, regional citrate dialysis is the most effective strategy. For patients, who require effective systemic anticoagulation, the usage of LMWH results in the longest circuit life spans. The proposed anticoagulatory strategies are safe, can easily be monitored, and allow an individualized treatment.
Assuntos
Injúria Renal Aguda/terapia , Anticoagulantes/administração & dosagem , Betacoronavirus , Infecções por Coronavirus/epidemiologia , Pneumonia Viral/epidemiologia , Terapia de Substituição Renal/métodos , Injúria Renal Aguda/sangue , Injúria Renal Aguda/epidemiologia , Adulto , Idoso , Arginina/análogos & derivados , Coagulação Sanguínea , COVID-19 , Ácido Cítrico/administração & dosagem , Comorbidade , Infecções por Coronavirus/sangue , Cuidados Críticos , Estado Terminal , Falha de Equipamento , Feminino , Alemanha/epidemiologia , Heparina/administração & dosagem , Heparina de Baixo Peso Molecular/administração & dosagem , Humanos , Masculino , Pessoa de Meia-Idade , Pandemias , Ácidos Pipecólicos/administração & dosagem , Pneumonia Viral/sangue , Terapia de Substituição Renal/instrumentação , Estudos Retrospectivos , SARS-CoV-2 , Sulfonamidas , Centros de Atenção TerciáriaRESUMO
INTRODUCTION: Hemangioblastomas are rare, histologically benign, highly vascularized tumors of the brain, the spinal cord, and the retina, occurring sporadically or associated with the autosomal dominant inherited von Hippel-Lindau (VHL) disease. Children or adults with VHL disease have one of > 300 known germline mutations of the VHL gene located on chromosome 3. They are prone to develop hemangioblastomas, extremely rarely starting at age 6, rarely at age 12-18, and, typically and almost all, as adults. There is a plethora of VHL-associated tumors and cysts, mainly in the kidney, pancreas, adrenals, reproductive organs, and central nervous system. Due to a lack of causal treatment, alleviation of symptoms and prevention of permanent neurological deficits as well as malignant transformation are the main task. Paucity of data and the nonlinear course of tumor progression make management of pediatric VHL patients with hemangioblastomas challenging. METHODS: The Freiburg surveillance protocol was developed by combining data from the literature and our experience of examinations of > 300 VHL patients per year at our university VHL center. RESULTS: Key recommendations are to start screening of patients at risk by funduscopy with dilated pupils for retinal tumors with admission to school and with MRI of the brain and spinal cord at age 14, then continue biannually until age 18, with emergency MRI in case of neurological symptoms. Indication for surgery remains personalized and should be approved by an experienced VHL board, but we regard neurological symptoms, rapid tumor growth, or critically large tumor/cyst sizes as the key indications to remove hemangioblastomas. Since repeated surgery on hemangioblastomas in VHL patients is not rare, modern neurosurgical techniques should encompass microsurgery, neuronavigation, intraoperative neuromonitoring, fluorescein dye-based intraoperative angiography, intraoperative ultrasound, and minimally invasive approaches, preceded in selected cases by endovascular embolization. Highly specialized neurosurgeons are able to achieve a very low risk of permanent morbidity for the removal of hemangioblastomas from the cerebellum and spinal cord. Small retinal tumors of the peripheral retina can be treated by laser coagulation, larger tumors by cryocoagulation or brachytherapy. CONCLUSION: We consider management at experienced VHL centers mandatory and careful surveillance and monitoring of asymptomatic lesions are required to prevent unnecessary operations and minimize morbidity.
Assuntos
Hemangioblastoma , Neoplasias da Medula Espinal , Doença de von Hippel-Lindau , Adolescente , Adulto , Criança , Patrimônio Genético , Hemangioblastoma/diagnóstico por imagem , Hemangioblastoma/genética , Hemangioblastoma/cirurgia , Humanos , Procedimentos Neurocirúrgicos , Neoplasias da Medula Espinal/diagnóstico por imagem , Neoplasias da Medula Espinal/genética , Neoplasias da Medula Espinal/cirurgia , Doença de von Hippel-Lindau/genética , Doença de von Hippel-Lindau/cirurgiaRESUMO
BACKGROUND: C3 glomerulopathy (C3G) is a rare, but severe glomerular disease with grim prognosis. The complex pathogenesis is just unfolding, and involves acquired as well as inherited dysregulation of the alternative pathway of the complement cascade. Currently, there is no established therapy. Treatment with the C5 complement inhibitor eculizumab may be a therapeutic option. However, due to rarity of the disease, parameters predicting treatment response remain largely unknown. METHODS: Seven patients with C3G (five with C3 glomerulonephritis and two with dense deposit disease) were treated with eculizumab. Subjects underwent biopsy before enrollment. The histopathology, clinical data, and response to eculizumab treatment were analyzed. The key parameters to determine outcome were changes of serum creatinine and urinary protein over time. RESULTS: After treatment with eculizumab, four subjects showed significantly improved or stable renal function and urinary protein. A positive response occurred between 2 weeks and 6 months after therapy initiation. One subject (with allograft recurrent C3 glomerulonephritis) initially showed a positive response, but relapsed when eculizumab was discontinued, and did not respond after re-initiation of treatment. Two subjects showed impaired renal function and increasing urinary protein despite therapy with eculizumab. CONCLUSIONS: Eculizumab may be a therapeutic option for a subset of C3G patients. The response to eculizumab is heterogeneous, and early as well as continuous treatment may be necessary to prevent disease progression. These findings emphasize the need for studies identifying genetic and functional complement abnormalities that may help to guide eculizumab treatment and predict response.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Complemento C3/metabolismo , Glomerulonefrite/sangue , Glomerulonefrite/tratamento farmacológico , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Resultado do Tratamento , Adulto JovemRESUMO
Despite its apparent simplicity, the nematode Caenorhabditis elegans has a high rating as a model in molecular and developmental biology and biomedical research. C. elegans has no excretory system comparable with the mammalian kidney but many of the genes and molecular pathways involved in human kidney diseases are conserved in C. elegans. The plethora of genetic, molecular and imaging tools available in C. elegans has enabled major discoveries in renal research and advanced our understanding of the pathogenesis of genetic kidney diseases. In particular, studies in C. elegans have pioneered the fundamental role of cilia for cystic kidney diseases. In addition, proteins of the glomerular filtration barrier and podocytes are critical for cell recognition, assembly of functional neuronal circuits, mechanosensation and signal transduction in C. elegans. C. elegans has also proved tremendously valuable for aging research and the Von Hippel-Lindau tumor suppressor gene has been shown to modulate lifespan in the nematode. Further, studies of the excretory canal, membrane transport and ion channel function in C. elegans have provided insights into mechanisms of tubulogenesis and cellular homeostasis. This review recounts the way that C. elegans can be used to investigate various aspects of genetic and molecular nephrology. This model system opens up an exciting and new area of study of renal development and diseases.
Assuntos
Caenorhabditis elegans , Modelos Animais de Doenças , Doenças Genéticas Inatas , Doenças Renais Císticas , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Doenças Genéticas Inatas/genética , Doenças Genéticas Inatas/metabolismo , Humanos , Doenças Renais Císticas/genética , Doenças Renais Císticas/metabolismoRESUMO
The Nrf family of transcription factors is critical for stress defense and detoxification. In Caenorhabditis elegans, the Nrf protein ortholog SKN-1 mediates this conserved stress response and promotes longevity. Moreover, SKN-1 is well known for its essential functions during C. elegans embryogenesis. SKN-1 is maternally deployed and initiates a signaling network specifying development of the endoderm and mesoderm. In this study, we identify the conserved Notch ligand OSM-11 as a novel regulator of SKN-1. We find that genetic inactivation of osm-11 re-establishes development of the pharynx and intestine in skn-1 deficient embryos and thereby rescues embryonic lethality associated with loss of skn-1 function. Inactivation of other DSL- and DOS-motif Notch ligands does not prevent skn-1 embryonic lethality. In addition, we show that inactivation of osm-11 in adult worms robustly enhances lifespan and promotes resistance to environmental stress. SKN-1 is required for increased longevity and heat and oxidative stress resistance but not hyperosmotic stress conferred by osm-11. OSM-11 prevents the nuclear accumulation of SKN-1 and represses the transcriptional activation of SKN-1 target genes for cellular detoxification. Our findings indicate that OSM-11 antagonizes SKN-1 during embryonic development and reveal a highly context-specific relationship between OSM-11 and SKN-1 in promoting stress resistance and longevity.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/embriologia , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Longevidade/fisiologia , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Envelhecimento/metabolismo , Animais , Animais Geneticamente Modificados , Caenorhabditis elegans/crescimento & desenvolvimento , Longevidade/genética , Estresse Oxidativo/fisiologia , Interferência de RNARESUMO
In Caenorhabditis elegans (C. elegans), the promotion of longevity by the transcription factor DAF-16 requires reduced insulin/IGF receptor (IIR) signaling or the ablation of the germline, although the reason for the negative impact of germ cells is unknown. FOXO/DAF-16 activity inhibits germline proliferation in both daf-2 mutants and gld-1 tumors. In contrast to its function as a germline tumor suppressor, we now provide evidence that somatic DAF-16 in the presence of IIR signaling can also result in tumorigenic activity, which counteracts robust lifespan extension. In contrast to the cell-autonomous IIR signaling, which is required for larval germline proliferation, activation of DAF-16 in the hypodermis results in hyperplasia of the germline and disruption of the surrounding basement membrane. SHC-1 adaptor protein and AKT-1 kinase antagonize, whereas AKT-2 and SGK-1 kinases promote, this cell-nonautonomous DAF-16 function. Our data suggest that a functional balance of DAF-16 activities in different tissues determines longevity and reveals a novel, cell-nonautonomous role of FOXO/DAF-16 to affect stem cells.
Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Transformação Celular Neoplásica/metabolismo , Longevidade/genética , Transdução de Sinais/genética , Células-Tronco/metabolismo , Fatores de Transcrição/metabolismo , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proliferação de Células , Transformação Celular Neoplásica/genética , Fatores de Transcrição Forkhead , Regulação da Expressão Gênica no Desenvolvimento , Células Germinativas/citologia , Células Germinativas/metabolismo , Humanos , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Somatomedina/genética , Receptores de Somatomedina/metabolismo , Proteínas Adaptadoras da Sinalização Shc/genética , Proteínas Adaptadoras da Sinalização Shc/metabolismo , Células-Tronco/citologia , Fatores de Transcrição/genéticaRESUMO
The evolutionarily conserved target of rapamycin (TOR) kinase controls fundamental metabolic processes to support cell and tissue growth. TOR functions within the context of two distinct complexes, TORC1 and TORC2. TORC2, with its specific component Rictor, has been recently implicated in aging and regulation of growth and metabolism. Here, we identify rict-1/Rictor as a regulator of embryonic development in C. elegans. The transcription factor skn-1 establishes development of the mesendoderm in embryos, and is required for cellular homeostasis and longevity in adults. Loss of maternal skn-1 function leads to mis-specification of the mesendodermal precursor and failure to form intestine and pharynx. We found that genetic inactivation of rict-1 suppressed skn-1-associated lethality by restoring mesendodermal specification in skn-1 deficient embryos. Inactivation of other TORC2 but not TORC1 components also partially rescued skn-1 embryonic lethality. The SGK-1 kinase mediated these functions downstream of rict-1/TORC2, as a sgk-1 gain-of-function mutant suppressed the rict-1 mutant phenotype. These data indicate that TORC2 and SGK-1 antagonize SKN-1 during embryonic development.
Assuntos
Proteínas de Caenorhabditis elegans/antagonistas & inibidores , Caenorhabditis elegans/embriologia , Proteínas de Ligação a DNA/antagonistas & inibidores , Embrião não Mamífero/citologia , Endoderma/embriologia , Mesoderma/embriologia , Complexos Multiproteicos/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição/antagonistas & inibidores , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Genes Letais , Alvo Mecanístico do Complexo 2 de RapamicinaRESUMO
Nephropathia epidemica, a zoonosis caused by Hantavirus infection (most commonly subtype Puumala) is associated with flu-like symptoms and acute kidney failure. Kidney manifestations are characterized predominantly by tubulointerstitial nephritis, hemorrhage into medullary tissues, interstitial edema, and tubular cell necrosis. Kidney failure is accompanied by proteinuria, and in some cases, nephrotic-range proteinuria may occur. However, the cellular mechanisms of proteinuria remain to be elucidated. We describe a Hantavirus (Puumala) infection in a 27-year-old man with acute kidney failure and severe and rapidly reversible proteinuria. Light microscopy of a kidney biopsy specimen showed only minor changes of glomeruli. However, transmission electron microscopy revealed podocyte foot-process effacement. Immunofluorescence staining of the slit diaphragm protein podocin and the tight junction protein ZO-1 revealed a partial mislocalization of these proteins. Together, these findings highlight that Hantavirus infection may perturb podocyte integrity, resulting in glomerular proteinuria. These alterations of podocytes and consequently the glomerular filtration barrier may be transient and resolve within weeks.
Assuntos
Infecções por Hantavirus/complicações , Infecções por Hantavirus/patologia , Podócitos/patologia , Proteinúria/complicações , Proteinúria/patologia , Adulto , Humanos , Masculino , Índice de Gravidade de DoençaRESUMO
The von Hippel-Lindau tumor suppressor gene (VHL) is mutated in up to 90% of clear cell renal cell carcinoma (ccRCC) cases, thus playing a key role in ccRCC pathogenesis. ccRCC can be classified as a metabolic disease in which alterations in fatty acid metabolism facilitate cancer cell proliferation. Enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase (EHHADH) is an enzyme involved in peroxisomal fatty acid degradation. It is primarily expressed in renal proximal tubule cells, presumably the origin of ccRCC. Although EHHADH is still a relatively unexplored gene, it is known to be differentially expressed in several tumors. In this study, analysis of several databases revealed that EHHADH expression is downregulated in ccRCC samples compared to healthy kidney samples. Moreover, cell culture experiments were performed to investigate the relationship between EHHADH and VHL at the gene and protein level. qPCR and Western blot analyses using the human ccRCC cell line RCC4 revealed that EHHADH is expressed in a VHL-dependent manner. RCC4 cells reconstituted with VHL show significantly higher EHHADH mRNA and protein levels than VHL-deficient RCC4 control cells. These results indicate that the downregulation of EHHADH in ccRCC reported may be due to the loss of VHL function. This study is the first to molecularly characterize EHHADH, a key enzyme in peroxisomal ß-oxidation, in relation to VHL, suggesting a potential pathogenic interaction that is worthy of further investigation.
RESUMO
To maximize the impact of precision medicine approaches, it is critical to identify genetic variants underlying disease and to accurately quantify their functional effects. A gene exemplifying the challenge of variant interpretation is the von Hippel-Lindautumor suppressor (VHL). VHL encodes an E3 ubiquitin ligase that regulates the cellular response to hypoxia. Germline pathogenic variants in VHL predispose patients to tumors including clear cell renal cell carcinoma (ccRCC) and pheochromocytoma, and somatic VHL mutations are frequently observed in sporadic renal cancer. Here we optimize and apply saturation genome editing to assay nearly all possible single-nucleotide variants (SNVs) across VHL's coding sequence. To delineate mechanisms, we quantify mRNA dosage effects and compare functional effects in isogenic cell lines. Function scores for 2,268 VHL SNVs identify a core set of pathogenic alleles driving ccRCC with perfect accuracy, inform differential risk across tumor types and reveal new mechanisms by which variants impact function. These results have immediate utility for classifying VHL variants encountered clinically and illustrate how precise functional measurements can resolve pleiotropic and dosage-dependent genotype-phenotype relationships across complete genes.
Assuntos
Alelos , Carcinoma de Células Renais , Edição de Genes , Neoplasias Renais , Polimorfismo de Nucleotídeo Único , Proteína Supressora de Tumor Von Hippel-Lindau , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Humanos , Edição de Genes/métodos , Carcinoma de Células Renais/genética , Neoplasias Renais/genética , Linhagem Celular Tumoral , Predisposição Genética para Doença , MutaçãoRESUMO
Clear cell renal cell carcinoma (ccRCC) is the most common histological subtype of renal cancer, and inactivation of the VHL tumor suppressor gene is found in almost all cases of hereditary and sporadic ccRCCs. CcRCC is associated with the reprogramming of fatty acid metabolism, and stearoyl-CoA desaturases (SCDs) are the main enzymes controlling fatty acid composition in cells. In this study, we report that mRNA and protein expression of the stearoyl-CoA desaturase SCD5 is downregulated in VHL-deficient cell lines. Similarly, in C. elegans vhl-1 mutants, FAT-7/SCD5 activity is repressed, supporting an evolutionary conservation. SCD5 regulation by VHL depends on HIF, and loss of SCD5 promotes cell proliferation and a metabolic shift towards ceramide production. In summary, we identify a novel regulatory function of VHL in relation to SCD5 and fatty acid metabolism, and propose a new mechanism of how loss of VHL may contribute to ccRCC tumor formation and progression.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Animais , Humanos , Carcinoma de Células Renais/metabolismo , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Neoplasias Renais/patologia , Proliferação de Células/genética , Homeostase , Lipídeos , Estearoil-CoA Dessaturase/genéticaRESUMO
Clear cell renal cell carcinoma (ccRCC) accounts for ~75% of kidney cancers. The biallelic inactivation of the von Hippel-Lindau tumor suppressor gene (VHL) is the truncal driver mutation of most cases of ccRCC. Cancer cells are metabolically reprogrammed and excrete modified nucleosides in larger amounts due to their increased RNA turnover. Modified nucleosides occur in RNAs and cannot be recycled by salvage pathways. Their potential as biomarkers has been demonstrated for breast or pancreatic cancer. To assess their suitability as biomarkers in ccRCC, we used an established murine ccRCC model, harboring Vhl, Trp53 and Rb1 (VPR) knockouts. Cell culture media of this ccRCC model and primary murine proximal tubular epithelial cells (PECs) were investigated by HPLC coupled to triple-quadrupole mass spectrometry using multiple-reaction monitoring. VPR cell lines were significantly distinguishable from PEC cell lines and excreted higher amounts of modified nucleosides such as pseudouridine, 5-methylcytidine or 2'-O-methylcytidine. The method's reliability was confirmed in serum-starved VPR cells. RNA-sequencing revealed the upregulation of specific enzymes responsible for the formation of those modified nucleosides in the ccRCC model. These enzymes included Nsun2, Nsun5, Pus1, Pus7, Naf1 and Fbl. In this study, we identified potential biomarkers for ccRCC for validation in clinical trials.
Assuntos
Carcinoma de Células Renais , Neoplasias Renais , Humanos , Animais , Camundongos , Carcinoma de Células Renais/patologia , Nucleosídeos/uso terapêutico , Reprodutibilidade dos Testes , Transcriptoma , Proteína Supressora de Tumor Von Hippel-Lindau/genética , Neoplasias Renais/patologia , RNA/uso terapêuticoRESUMO
Monoclonal antibodies (mAbs) directed against the spike of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are effective therapeutic options to combat infections in high-risk patients. Here, we report the adaptation of SARS-CoV-2 to the mAb cocktail REGN-COV in a kidney transplant patient with hypogammaglobulinemia. Following mAb treatment, the patient did not clear the infection. During viral persistence, SARS-CoV-2 acquired three novel spike mutations. Neutralization and mouse protection analyses demonstrate a complete viral escape from REGN-COV at the expense of ACE-2 binding. Final clearance of the virus occurred upon reduction of the immunosuppressive regimen and total IgG substitution. Serology suggests that the development of highly neutralizing IgM rather than IgG substitution aids clearance. Our findings emphasise that selection pressure by mAbs on SARS-CoV-2 can lead to development of escape variants in immunocompromised patients. Thus, modification of immunosuppressive therapy, if possible, might be preferable to control and clearance of the viral infection.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Camundongos , Anticorpos Antivirais , Testes de Neutralização , Anticorpos Neutralizantes , Hospedeiro Imunocomprometido , Imunoglobulina G , Glicoproteína da Espícula de CoronavírusRESUMO
Mutations of the immunoglobulin superfamily proteins nephrin and Neph1 lead to congenital nephrotic syndrome in humans or mice. Neph proteins are three closely related molecules that are evolutionarily conserved and mediate cell recognition. Their importance for morphogenetic processes including the formation of the kidney filtration barrier in vertebrates and synaptogenesis in Caenorhabditis elegans has recently been uncovered. However, the individual morphogenetic function of mammalian Neph1-3 isoforms remained elusive. We demonstrate now that the Neph/nephrin family proteins can form cell-cell adhesion modules across species. Expression of all three mammalian Neph isoforms partially rescued mutant C. elegans lacking their Neph homolog syg-1 and restored synapse formation, suggesting a functional redundancy between the three isoforms. Strikingly, the rescue of defective synaptic connectivity was prevented by deletion of the highly conserved cytoplasmic PSD95/Dlg/ZO-1-binding motif of SYG-1/Neph proteins, indicating the critical role of this intracellular signaling motif for SYG-1/Neph-dependent morphogenetic events. To determine the significance of Neph isoform redundancy for vertebrate kidney development, we analyzed the expression pattern and the functional role of Neph proteins in zebrafish. In situ hybridizations identified zNeph1 and zNeph2 as glomerular proteins. Morpholino knockdown of either zNeph1 or zNeph2 resulted in loss of slit diaphragms and leakiness of the glomerular filtration barrier. This is the first report utilizing C. elegans to study mammalian Neph/nephrin protein function and to demonstrate a functional overlap of Neph1-3 proteins. Furthermore, we identify Neph2 as a novel critical regulator of glomerular function, indicating that both Neph1 and Neph2 are required for glomerular maintenance and development.
Assuntos
Imunoglobulinas/fisiologia , Rim/crescimento & desenvolvimento , Proteínas de Membrana/fisiologia , Neurônios/fisiologia , Sequência de Aminoácidos , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/crescimento & desenvolvimento , Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/classificação , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/fisiologia , Adesão Celular , Células HeLa , Humanos , Imunoglobulinas/classificação , Imunoglobulinas/genética , Proteínas de Membrana/classificação , Proteínas de Membrana/genética , Camundongos , Modelos Animais , Morfogênese/genética , Domínios PDZ , FilogeniaRESUMO
Von Hippel-Lindau disease (VHL) is a hereditary disorder associated with malignant tumors including clear cell renal cell carcinoma (ccRCC). Partial nephrectomy is complicated by multilocular tumor occurrence and a high recurrence rate. The aim of this study was to evaluate the potential of stereotactic body radiotherapy (SBRT) as an alternative treatment approach in VHL patients with multiple ccRCC. Patients with VHL and a diagnosis of ccRCC were enrolled. SBRT was conducted using five fractions of 10 Gy or eight fractions of 7.5 Gy. The primary endpoint was local control (LC). Secondary endpoints included alteration of renal function and adverse events. Seven patients with a total of eight treated lesions were enrolled. Median age was 44 years. Five patients exhibited multiple bilateral kidney cysts in addition to ccRCC. Three patients underwent at least one partial nephrectomy in the past. After a median follow-up of 43 months, 2-year LC was 100%, while 2-year CSS, 2-year PFS and 2-year OS was 100%, 85.7% and 85.7%, respectively. SBRT was very well tolerated with no acute or chronic toxicities grade ≥ 2. Mean estimated glomerular filtration rate (eGFR) at baseline was 83.7 ± 13.0 mL/min/1.73 m2, which decreased to 76.6 ± 8.0 mL/min/1.73 m2 after 1 year. Although the sample size was small, SBRT resulted in an excellent LC rate and was very well tolerated with preservation of kidney function in patients with multiple renal lesions and cysts.
RESUMO
Hypertensive disorders occur in up to 10â% of pregnancies and increase both maternal and fetal morbidity and mortality. The most important differential diagnoses comprise pre-existing chronic hypertension, pregnancy-associated hypertension, and preeclampsia with simultaneous proteinuria. Antihypertensive therapy during pregnancy should be initiated when blood pressure is 150-160/100-110âmmHg. With regard to an earlier initiation of therapy, the data situation is not clear. Pre-eclampsia is defined as new or pre-existing elevated blood pressure ≥â140/90âmmHg in pregnancy with at least one new organ manifestation, usually proteinuria ≥â300âmg/day or ≥â30âmg/mmol in the protein-creatinine ratio. Thrombotic microangiopathies TTP and aHUS are altogether rare but potentially life-threatening diseases that should be clarified in case of severe or atypical courses.
Assuntos
Hipertensão Induzida pela Gravidez , Adulto , Anti-Hipertensivos/uso terapêutico , Pressão Sanguínea/fisiologia , Feminino , Humanos , GravidezRESUMO
Inactivation of the tumor suppressor von Hippel-Lindau (VHL) gene is a key event in hereditary and sporadic clear cell renal cell carcinomas (ccRCC). The mechanistic target of rapamycin (mTOR) signaling pathway is a fundamental regulator of cell growth and proliferation, and hyperactivation of mTOR signaling is a common finding in VHL-dependent ccRCC. Deregulation of mTOR signaling correlates with tumor progression and poor outcome in patients with ccRCC. Here, we report that the regulatory-associated protein of mTOR (RAPTOR) is strikingly repressed by VHL. VHL interacts with RAPTOR and increases RAPTOR degradation by ubiquitination, thereby inhibiting mTORC1 signaling. Consistent with hyperactivation of mTORC1 signaling in VHL-deficient ccRCC, we observed that loss of vhl-1 function in C. elegans increased mTORC1 activity, supporting an evolutionary conserved mechanism. Our work reveals important new mechanistic insight into deregulation of mTORC1 signaling in ccRCC and links VHL directly to the control of RAPTOR/mTORC1. This may represent a novel mechanism whereby loss of VHL affects organ integrity and tumor behavior.