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1.
Bone ; 154: 116202, 2021 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-34534708

RESUMO

Contemporary intravenous iron formulations allow administration of high doses of elemental iron and enable correction of total iron deficit in one or two infusions. An important but underappreciated complication of certain formulations is hypophosphatemia caused by increased secretion of the phosphaturic hormone, fibroblast growth factor 23 (FGF23). The pathophysiology of FGF23-induced hypophosphatemia due to certain intravenous iron formulations has been recently investigated in prospective clinical trials. To reach the correct diagnosis, clinicians must recognize the typical clinical manifestations of intravenous iron-induced hypophosphatemia and identify a specific pattern of biochemical changes (hyperphosphaturic hypophosphatemia triggered by high FGF23 that causes low 1,25 (OH)2 vitamin D, hypocalcemia and secondary hyperparathyroidism). Physicians and patients should be aware of hypophosphatemia as a common complication of intravenous iron therapy and monitor serum phosphate concentrations in patients receiving repeated doses of specific intravenous iron formulations. Symptoms of hypophosphatemia are associated with severity and duration. Persistent hypophosphatemia can occur with iron therapy and can cause debilitating diseases including myopathy, osteomalacia and fractures. This review summarizes the current understanding of the iron-phosphate axis as well as complications of intravenous iron-induced hypophosphatemia.

2.
Cell Syst ; 12(8): 780-794.e7, 2021 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-34139154

RESUMO

COVID-19 is highly variable in its clinical presentation, ranging from asymptomatic infection to severe organ damage and death. We characterized the time-dependent progression of the disease in 139 COVID-19 inpatients by measuring 86 accredited diagnostic parameters, such as blood cell counts and enzyme activities, as well as untargeted plasma proteomes at 687 sampling points. We report an initial spike in a systemic inflammatory response, which is gradually alleviated and followed by a protein signature indicative of tissue repair, metabolic reconstitution, and immunomodulation. We identify prognostic marker signatures for devising risk-adapted treatment strategies and use machine learning to classify therapeutic needs. We show that the machine learning models based on the proteome are transferable to an independent cohort. Our study presents a map linking routinely used clinical diagnostic parameters to plasma proteomes and their dynamics in an infectious disease.


Assuntos
Biomarcadores/análise , COVID-19/patologia , Progressão da Doença , Proteoma/fisiologia , Fatores Etários , Contagem de Células Sanguíneas , Gasometria , Ativação Enzimática , Humanos , Inflamação/patologia , Aprendizado de Máquina , Prognóstico , Proteômica , SARS-CoV-2/imunologia
3.
Cells ; 10(2)2021 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-33498986

RESUMO

We explore the status of quiescence, stemness and adipogenic differentiation capacity in adipose stem/progenitor cells (ASCs) ex vivo, immediately after isolation from human subcutaneous white adipose tissue, by sorting the stromal vascular fraction into cell-surface DLK1+/CD34-, DLK1+/CD34dim and DLK1-/CD34+ cells. We demonstrate that DLK1-/CD34+ cells, the only population exhibiting proliferative and adipogenic capacity, express ex vivo the bonafide quiescence markers p21Cip1, p27Kip1 and p57Kip2 but neither proliferation markers nor the senescence marker p16Ink4a. The pluripotency markers NANOG, SOX2 and OCT4 are barely detectable in ex vivo ASCs while the somatic stemness factors, c-MYC and KLF4 and the early adipogenic factor C/EBPß are highly expressed. Further sorting of ASCs into DLK1-/CD34+/CD24- and DLK1-/CD34+/CD24+ fractions shows that KLF4 and c-MYC are higher expressed in DLK1-/CD34+/CD24+ cells correlating with higher colony formation capacity and considerably lower adipogenic activity. Proliferation capacity is similar in both populations. Next, we show that ASCs routinely isolated by plastic-adherence are DLK1-/CD34+/CD24+. Intriguingly, CD24 knock-down in these cells reduces proliferation and adipogenesis. In conclusion, DLK1-/CD34+ ASCs in human sWAT exist in a quiescent state, express high levels of somatic stemness factors and the early adipogenic transcription factor C/EBPß but senescence and pluripotency markers are barely detectable. Moreover, our data indicate that CD24 is necessary for adequate ASC proliferation and adipogenesis and that stemness is higher and adipogenic capacity lower in DLK1-/CD34+/CD24+ relative to DLK1-/CD34+/CD24- subpopulations.


Assuntos
Adipogenia , Tecido Adiposo Branco/citologia , Antígenos CD34/metabolismo , Antígeno CD24/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Ciclo Celular , Proteínas de Membrana/metabolismo , Células-Tronco/citologia , Adipogenia/genética , Biomarcadores/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Ciclo Celular/genética , Proliferação de Células , Células Cultivadas , Feminino , Regulação da Expressão Gênica , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Masculino , Proteínas Proto-Oncogênicas c-myc/metabolismo , RNA Interferente Pequeno/metabolismo , Células-Tronco/metabolismo , Células Estromais/metabolismo , Gordura Subcutânea/citologia
4.
Adipocyte ; 9(1): 626-635, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-33070670

RESUMO

The CRISPR/Cas9 system is a powerful tool to generate a specific loss-of-function phenotype by gene knockout (KO). However, this approach is challenging in primary human cells. In this technical report, we present a reliable protocol to achieve a functional KO in the genome of human adipose stem/progenitor cells (ASCs). Using Sprouty1 (SPRY1) as a model target gene for a CRISPR/Cas9 mediated KO, we particularize the procedure including the selection of the CRISPR/Cas9 target sequences and the employment of appropriate lentiviral vectors to obtain a functional gene KO. The efficiency of CRISPR/Cas9 to mutate the SPRY1 gene is determined by a PCR-based mutation detection assay and sequence analysis. Effects on mRNA and protein levels are studied by RT-qPCR and Western blotting. In addition, we demonstrate that CRISPR/Cas9 mediated SPRY1 KO and gene silencing by shRNA are similarly effective to deplete the Sprouty1 protein and to inhibit adipogenic differentiation. In summary, we show a reliable approach to achieve a gene KO in human ASCs, which could also apply to other primary cell types. Abbreviations: ASC: Adipogenic Stem/Progenitor Cell; Cas: CRISPR-associated system; CRISPR: Clustered Regularly Interspaced Palindromic Repeat; gDNA: Genomic DNA; GOI: Gene of interest; gRNA: Guide RNA; NHEJ: Non-homologous end joining; Indel: Insertion/Deletion; PAM: Protospacer adjacent motif; sWAT: Subcutaneous white adipose tissue; TIDE: Tracking of indels by decomposition.

5.
Hepatol Commun ; 4(9): 1239-1241, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32923830
7.
Mol Aspects Med ; 75: 100862, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32444112

RESUMO

Intravenous infusions of iron have evolved from a poorly effective and dangerous intervention to a safe cornerstone in the treatment of iron deficiency. Modern iron formulations are composite nanoparticles composed of carbohydrate ferric oxy-hydroxides. Iron dextran, iron derisomaltose (formely known as iron isomaltoside 1000), ferric carboxymaltose, ferrumoxytol, iron sucrose and sodium ferric gluconate can be infused at different doses and allow correction of total iron deficit with single or repeated doses in 1-2 weeks depending on the specific formulation. All iron preparations are associated with a risk of severe infusion reactions. In recent prospective clinical trials, the risk of moderate to severe infusion reactions was comparable among all modern preparations affecting <1% of patients. Hence, intravenous iron therapy is reserved for iron deficiency anemia patients with intolerance or unresponsiveness of oral iron. As per European drug label, intravenous iron may also be preferred when rapid correction of the iron deficit is required. In patients with inflammation, iron-deficiency should also be suspected as anemia cause when transferrin saturation is low because serum ferritin can be spuriously normal. The main treatment target for i.v. iron is an improvement of the quality of life, for which hemoglobin is a surrogate marker. An emerging complication affecting 50-74% of patients treated with ferric carboxymaltose in prospective clinical trials is hypophosphatemia - or more accurately the 6H syndrome (hyperphosphaturic hypophosphatemia triggered by high fibroblast growth factor 23 that causes hypovitaminosis D, hypocalcemia and secondary hyperparathyroidism). These biochemical changes can cause severe and potentially irreversible clinical complications, such a bone pain, osteomalacia and fractures. Individual selection of the appropriate iron therapy and evaluation of treatment response are mandatory to safely deliver improved outcome through intravenous iron therapies.

8.
J Gerontol A Biol Sci Med Sci ; 75(12): 2308-2319, 2020 11 13.
Artigo em Inglês | MEDLINE | ID: mdl-32304210

RESUMO

The role of Ras-Mitogen-activated protein kinase (MAPK) signaling in cellular aging is not precisely understood. Recently, we identified Sprouty1 (SPRY1) as a weight-loss target gene in human adipose stem/progenitor cells (ASCs) and showed that Sprouty1 is important for proper regulation of adipogenesis. In the present study, we show that loss-of-function of Sprouty1 by CRISPR/Cas9-mediated genome editing in human ASCs leads to hyper-activation of MAPK signaling and a senescence phenotype. Sprouty1 knockout ASCs undergo an irreversible cell cycle arrest, become enlarged and stain positive for senescence-associated ß-galactosidase. Sprouty1 down-regulation leads to DNA double strand breaks, a considerably increased number of senescence-associated heterochromatin foci and induction of p53 and p21Cip1. In addition, we detect an increase of hypo-phosphorylated Retinoblastoma (Rb) protein in SPRY1 knockout ASCs. p16Ink4A is not induced. Moreover, we show that Sprouty1 knockout leads to induction of a senescence-associated secretory phenotype as indicated by the activation of the transcription factors NFκB and C/EBPß and a significant increase in mRNA expression and secretion of interleukin-8 (IL-8) and CXCL1/GROα. Finally, we demonstrate that adipogenesis is abrogated in senescent SPRY1 knockout ASCs. In conclusion, this study reveals a novel mechanism showing the importance of Sprouty1 for the prevention of senescence and the maintenance of the proliferation and differentiation capacity of human ASCs.


Assuntos
Tecido Adiposo/citologia , Senescência Celular/genética , Proteínas de Membrana/genética , Fosfoproteínas/genética , Células-Tronco/citologia , Adipogenia/genética , Diferenciação Celular/genética , Proliferação de Células/genética , Células Cultivadas , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Humanos , Mutação com Perda de Função , Fenótipo , Transdução de Sinais , beta-Galactosidase/metabolismo
9.
EBioMedicine ; 46: 387-398, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31327694

RESUMO

BACKGROUND: The bone marrow (BM) is a major reservoir of resting memory T cells and long-lived plasma cells, capable of providing protection against recurrent infections. Whether the age-related accumulation of adipose tissue in the BM affects the functionality and maintenance of memory cells is not well understood. METHODS: For the first time, we compare human femur marrow adipose tissue (fMAT) and subcutaneous white adipose tissue of the thigh (tsWAT) obtained from the same donors. Therefore, we used microarrays for comparative global gene expression analysis, and employed assays to analyse parameters of adipocyte biology, inflammation and oxidative stress. FINDINGS: We show that fMAT adipocytes differ significantly from tsWAT adipocytes regarding specific gene expression profiles including inflammatory responses and adipogenesis/adipocyte phenotype. Concomitant with considerably lower levels of CD36, a membrane-associated protein important for long-chain fatty acid uptake that is used as maturation marker, fMAT adipocytes are smaller and contain less triglycerides. fMAT adipocytes secrete similar levels of adiponectin and leptin as tsWAT adipocytes, and express increased levels of pro-inflammatory molecules concomitant with an elevated generation of reactive oxygen species (ROS) and impaired function of plasma cells in the BM. INTERPRETATION: Our findings suggest that fMAT is a unique type of adipose tissue containing small adipocytes with lower CD36 protein and triglyceride levels than tsWAT but high adipokine secretion. Moreover, fMAT adipocytes secrete high levels of pro-inflammatory cytokines, contributing to inflammation and impairment of plasma cell function in the BM, suggesting that fMAT has more immune regulatory functions than tsWAT.


Assuntos
Adipócitos/imunologia , Adipócitos/metabolismo , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Imunomodulação , Idoso , Biomarcadores , Antígenos CD36/metabolismo , Citocinas/metabolismo , Feminino , Citometria de Fluxo , Imunofluorescência , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo
10.
Cell Death Dis ; 10(6): 411, 2019 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-31138786

RESUMO

The differentiation of adipose stem/progenitor cells (ASCs) into adipocytes contributes to adipose tissue expansion in obesity. This process is regulated by numerous signalling pathways including MAPK signalling. In the present study, we show that weight loss (WL) interventions induce upregulation of Sprouty1 (SPRY1), a negative regulator of MAPK signalling, in human ASCs and elucidate the role of the Sprouty1/MAPK interaction for adipogenic differentiation. We found that the Sprouty1 protein levels are low in proliferating ASCs, increasing in density arrested ASCs at the onset of adipogenic differentiation and decreasing in the course of adipogenesis. Knock-down (KD) of Sprouty1 by RNA interference led to elevated MAPK activity and reduced expression of the early adipogenic transcription factor CCAAT/enhancer-binding protein ß (C/EBP ß), concomitant with an abrogation of adipogenesis. Intriguingly, co-treatment of Sprouty1 KD ASCs with differentiation medium and the pharmacological MEK inhibitor U0126 blunted ERK phosphorylation; however, failed to rescue adipogenic differentiation. Thus, the effects of the Sprouty1 KD are not reversed by inhibiting MAPK signalling although the inhibition of MAPK signalling by U0126 did not prevent adipogenic differentiation in wild type ASCs. In conclusion, we show that Sprouty1 is induced after WL in ASCs of formerly obese people acting as a negative regulator of MAPK signalling, which is necessary to properly trigger adipogenesis at early stages by a C/EBP ß dependent mechanism.


Assuntos
Adipogenia/genética , Tecido Adiposo/metabolismo , Proteínas de Membrana/metabolismo , Fosfoproteínas/metabolismo , Células-Tronco/metabolismo , Perda de Peso/genética , Adipócitos/metabolismo , Adipogenia/efeitos dos fármacos , Tecido Adiposo/efeitos dos fármacos , Adolescente , Adulto , Butadienos/farmacologia , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Células Cultivadas , Feminino , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Nitrilas/farmacologia , Obesidade/metabolismo , Fosfoproteínas/genética , Células-Tronco/efeitos dos fármacos , Perda de Peso/efeitos dos fármacos , Adulto Jovem
11.
J Pediatr Surg ; 52(10): 1583-1590, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28499711

RESUMO

BACKGROUND: Measurements in chest wall deformities are typically conducted using a thorax caliper or a CT scan of the chest wall. This paper focuses on the possible correlation between these two methods to validate the reliability of the thorax caliper, minimize radiation exposure, and limit the usage of expensive imaging techniques. METHODS: We evaluated 95 consecutive patients (77 pectus excavatum (PE), 17 pectus carinatum (PC), 1 mixed deformity) who received surgical correction of the anterior chest wall. The results of the external chest wall measurements and the CT-based measurements were statistically compared. RESULTS: A significant correlation between the two measurements was observed in PE and PC at the highest point of the deformation. The strongest correlation was noted in PE. We also noted a correlation between the transverse diameter of the external measurement and the inner thoracic diameter of the CT scan but not for the sagittal diameters in the upper parts of the sternum. CONCLUSIONS: Thorax caliper measurements are suitable for determining the sagittal thoracic diameter at the maximum level of the deformity and the transverse diameter with an accuracy comparable to that of CT measurements. Since these values key, the thorax caliper is reliable for monitoring and documenting chest wall malformations. LEVEL OF EVIDENCE: Study of diagnostic test. Testing previously developed diagnostic criteria in a consecutive series of patients and a universally "gold" standard-Level I.


Assuntos
Tórax em Funil/diagnóstico por imagem , Tórax em Funil/patologia , Parede Torácica/anormalidades , Parede Torácica/diagnóstico por imagem , Adolescente , Criança , Testes Diagnósticos de Rotina , Fixadores Externos , Feminino , Tórax em Funil/cirurgia , Humanos , Masculino , Reprodutibilidade dos Testes , Esterno/diagnóstico por imagem , Parede Torácica/patologia , Parede Torácica/cirurgia , Tomografia Computadorizada por Raios X/métodos
12.
Tuberc Res Treat ; 2012: 768723, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22567275

RESUMO

Interferon gamma release assays (IGRAs) are in vitro immunologic diagnostic tests used to identify Mycobacterium tuberculosis infection. They cannot differentiate between latent and active infections. The cutoff suggested by the manufacturer is 0.35 IU/mL for latent tuberculosis. As IGRA tests were recently approved for the differential diagnosis of active tuberculosis, we assessed the diagnostic accuracy of the latest generation IGRA for detection of active tuberculosis in a low-incidence area in Germany. Our consecutive case series includes 61 HIV negative, Mycobacterium tuberculosis culture positive patients, as well as 234 control patients. The retrospective analysis was performed over a period of two years. In 11/61 patients with active tuberculosis (18.0%) the test result was <0.35 IU/mL, resulting in a sensitivity of 0.82. We recommend establishing a new cut-off value for the differential diagnosis of active tuberculosis assessed by prospective clinical studies and in various regions with high and low prevalence of tuberculosis.

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