RESUMO
OBJECTIVE: To determine whether microRNAs (miRs) are elevated in the plasma of individuals with the inherited peripheral neuropathy Charcot-Marie-Tooth disease type 1A (CMT1A), miR profiling was employed to compare control and CMT1A plasma. METHODS: We performed a screen of CMT1A and control plasma samples to identify miRs that are elevated in CMT1A using next-generation sequencing, followed by validation of selected miRs by quantitative PCR, and correlation with protein biomarkers and clinical data: Rasch-modified CMT Examination and Neuropathy Scores, ulnar compound muscle action potentials, and motor nerve conduction velocities. RESULTS: After an initial pilot screen, a broader screen confirmed elevated levels of several muscle-associated miRNAs (miR1, -133a, -133b, and -206, known as myomiRs) along with a set of miRs that are highly expressed in Schwann cells of peripheral nerve. Comparison to other candidate biomarkers for CMT1A (e.g., neurofilament light) measured on the same sample set shows a comparable elevation of several miRs (e.g., miR133a, -206, -223) and ability to discriminate cases from controls. Neurofilament light levels were most highly correlated with miR133a. In addition, the putative Schwann cell miRs (e.g., miR223, -199a, -328, -409, -431) correlate with the recently described transmembrane protease serine 5 (TMPRSS5) protein biomarker that is most highly expressed in Schwann cells and also elevated in CMT1A plasma. CONCLUSIONS: These studies identify a set of miRs that are candidate biomarkers for clinical trials in CMT1A. Some of the miRs may reflect Schwann cell processes that underlie the pathogenesis of the disease. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that a set of plasma miRs are elevated in patients with CMT1A.
Assuntos
Doença de Charcot-Marie-Tooth/diagnóstico , Doença de Charcot-Marie-Tooth/genética , MicroRNAs/análise , MicroRNAs/genética , Potenciais de Ação , Adulto , Envelhecimento , Biomarcadores/análise , Biologia Computacional , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Neurônios Motores , Músculo Esquelético/fisiopatologia , Condução Nervosa , Proteínas de Neurofilamentos/química , Nervos Periféricos/metabolismo , Reprodutibilidade dos Testes , Células de Schwann/metabolismo , Nervo Ulnar/fisiopatologiaRESUMO
Receptor interacting protein kinase 1 (RIPK1) mediates cell death and inflammatory signaling and is increased in multiple sclerosis (MS) brain samples. Here, we investigate the role of glial RIPK1 kinase activity in mediating MS pathogenesis. We demonstrate RIPK1 levels correlate with MS disease progression. We find microglia are susceptible to RIPK1-mediated cell death and identify an inflammatory gene signature that may contribute to the neuroinflammatory milieu in MS patients. We uncover a distinct role for RIPK1 in astrocytes in regulating inflammatory signaling in the absence of cell death and confirm RIPK1-kinase-dependent regulation in human glia. Using a murine MS model, we show RIPK1 inhibition attenuates disease progression and suppresses deleterious signaling in astrocytes and microglia. Our results suggest RIPK1 kinase activation in microglia and astrocytes induces a detrimental neuroinflammatory program that contributes to the neurodegenerative environment in progressive MS.
Assuntos
Microglia/metabolismo , Esclerose Múltipla/genética , Doenças Neuroinflamatórias/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Humanos , Camundongos , Esclerose Múltipla/patologia , Transdução de SinaisRESUMO
OBJECTIVE: Development of biomarkers for Charcot-Marie-Tooth (CMT) disease is critical for implementing effective clinical trials. The most common form of CMT, type 1A, is caused by a genomic duplication surrounding the PMP22 gene. A recent report (Neurology 2018;90:e518-3524) showed elevation of neurofilament light (NfL) in plasma of CMT1A disease patients, which correlated with disease severity. However, no plasma/serum biomarker has been identified that is specific to Schwann cells, the most directly affected cells in CMT1A. METHODS: We used the Olink immuno PCR platform to profile CMT1A patient (n = 47, 2 cohorts) and normal control plasma (n = 41, two cohorts) on five different Olink panels to screen 398 unique proteins. RESULTS: The TMPRSS5 protein (Transmembrane protease serine 5) was elevated 2.07-fold (P = <0.0001) in two independent cohorts of CMT1A samples relative to controls. TMPRSS5 is most highly expressed in Schwann cells of peripheral nerve. Consistent with early myelination deficits in CMT1A, TMPRSS5 was not significantly correlated with disease score (CMTES-R, CMTNS-R), nerve conduction velocities (Ulnar CMAP, Ulnar MNCV), or with age. TMPRSS5 was not significantly elevated in smaller sample sets from patients with CMT2A, CMT2E, CMT1B, or CMT1X. The Olink immuno PCR assays confirmed elevated levels of NfL (average 1.58-fold, P < 0.0001), which correlated with CMT1A patient disease score. INTERPRETATION: These data identify the first Schwann cell-specific protein that is elevated in plasma of CMT1A patients, and may provide a disease marker and a potentially treatment-responsive biomarker with good disease specificity for clinical trials.
Assuntos
Doença de Charcot-Marie-Tooth/sangue , Doença de Charcot-Marie-Tooth/diagnóstico , Proteínas de Membrana/sangue , Proteínas Mitocondriais/sangue , Células de Schwann , Serina Endopeptidases/sangue , Adulto , Animais , Biomarcadores/sangue , Células Cultivadas , Doença de Charcot-Marie-Tooth/fisiopatologia , Estudos de Coortes , Diagnóstico Diferencial , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Condução Nervosa/fisiologia , Reação em Cadeia da Polimerase , RatosRESUMO
MicroRNA-21 (miR-21) contributes to the pathogenesis of fibrogenic diseases in multiple organs, including the kidneys, potentially by silencing metabolic pathways that are critical for cellular ATP generation, ROS production, and inflammatory signaling. Here, we developed highly specific oligonucleotides that distribute to the kidney and inhibit miR-21 function when administered subcutaneously and evaluated the therapeutic potential of these anti-miR-21 oligonucleotides in chronic kidney disease. In a murine model of Alport nephropathy, miR-21 silencing did not produce any adverse effects and resulted in substantially milder kidney disease, with minimal albuminuria and dysfunction, compared with vehicle-treated mice. miR-21 silencing dramatically improved survival of Alport mice and reduced histological end points, including glomerulosclerosis, interstitial fibrosis, tubular injury, and inflammation. Anti-miR-21 enhanced PPARα/retinoid X receptor (PPARα/RXR) activity and downstream signaling pathways in glomerular, tubular, and interstitial cells. Moreover, miR-21 silencing enhanced mitochondrial function, which reduced mitochondrial ROS production and thus preserved tubular functions. Inhibition of miR-21 was protective against TGF-ß-induced fibrogenesis and inflammation in glomerular and interstitial cells, likely as the result of enhanced PPARα/RXR activity and improved mitochondrial function. Together, these results demonstrate that inhibition of miR-21 represents a potential therapeutic strategy for chronic kidney diseases including Alport nephropathy.
Assuntos
MicroRNAs/genética , Nefrite Hereditária/terapia , Oligorribonucleotídeos Antissenso/genética , Animais , Autoantígenos/genética , Colágeno Tipo IV/deficiência , Colágeno Tipo IV/genética , Progressão da Doença , Fibrose/metabolismo , Rim/metabolismo , Rim/patologia , Redes e Vias Metabólicas/genética , Camundongos da Linhagem 129 , MicroRNAs/metabolismo , Nefrite Hereditária/metabolismo , Nefrite Hereditária/patologia , Espécies Reativas de Oxigênio/metabolismo , Transcriptoma , Regulação para CimaRESUMO
Altered bone turnover is a key pathologic feature of chronic kidney disease-mineral and bone disorder (CKD-MBD). Expression of TGF-ß1, a known regulator of bone turnover, is increased in bone biopsies from individuals with CKD. Similarly, TGF-ß1 mRNA and downstream signaling is increased in bones from jck mice, a model of high-turnover renal osteodystrophy. A neutralizing anti-TGF-ß antibody (1D11) was used to explore TGF-ß's role in renal osteodystrophy. 1D11 administration to jck significantly attenuated elevated serum osteocalcin and type I collagen C-telopeptides. Histomorphometric analysis indicated that 1D11 administration increased bone volume and suppressed the elevated bone turnover in a dose-dependent manner. These effects were associated with reductions in osteoblast and osteoclast surface areas. Micro-computed tomography (µCT) confirmed the observed increase in trabecular bone volume and demonstrated improvements in trabecular architecture and increased cortical thickness. 1D11 administration was associated with significant reductions in expression of osteoblast marker genes (Runx2, alkaline phosphatase, osteocalcin) and the osteoclast marker gene, Trap5. Importantly, in this model, 1D11 did not improve kidney function or reduce serum parathyroid hormone (PTH) levels, indicating that 1D11 effects on bone are independent of changes in renal or parathyroid function. 1D11 also significantly attenuated high-turnover bone disease in the adenine-induced uremic rat model. Antibody administration was associated with a reduction in pSMAD2/SMAD2 in bone but not bone marrow as assessed by quantitative immunoblot analysis. Immunostaining revealed pSMAD staining in osteoblasts and osteocytes but not osteoclasts, suggesting 1D11 effects on osteoclasts may be indirect. Immunoblot and whole genome mRNA expression analysis confirmed our previous observation that repression of Wnt/ß-catenin expression in bone is correlated with increased osteoclast activity in jck mice and bone biopsies from CKD patients. Furthermore, our data suggest that elevated TGF-ß may contribute to the pathogenesis of high-turnover disease partially through inhibition of ß-catenin signaling.