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Cell-free DNA (cfDNA) is used in clinical research to identify biomarkers for diagnosis of and follow-up on cancer. Here, we propose a fast and innovative approach using traditional housekeeping genes as cfDNA targets in a copy number analysis. We focus on the application of highly sensitive technology such as digital PCR (dPCR) to differentiate breast cancer (BC) patients and controls by quantifying regions of PUM1 and RPPH1 (RNase P) in plasma samples.
We conducted a case-control study with 82 BC patients and 82 healthy women. read more cfDNA was isolated from plasma using magnetic beads and quantified by spectrophotometry to estimate total cfDNA. Then, both PUM1 and RPPH1 genes were specifically quantified by dPCR. Data analysis was calibrated using a reference genomic DNA in different concentrations.
We found RNaseP and PUM1 values were correlated in the patient group (intraclass correlation coefficient [ICC]=0.842), but they did not have any correlation in healthy women (ICC=0.519). In dPCR quantification, PUM1 showed the capacity to distinguish early-stage patients and controls with good specificity (98.67%) and sensitivity (100%). Conversely, RNaseP had lower cfDNA levels in triple-negative BC patients than luminal subtypes (p<0.025 for both), confirming their utility for patient classification.
We propose the PUM1 gene as a cfDNA marker for early diagnosis of BC and RNaseP as a cfDNA marker related to hormonal status and subtype classification in BC. Further studies with larger sample sizes are warranted.
We propose the PUM1 gene as a cfDNA marker for early diagnosis of BC and RNase P as a cfDNA marker related to hormonal status and subtype classification in BC. Further studies with larger sample sizes are warranted.Cisplatin (Cis) is one of the most potent and effective broad-spectrum antitumor drugs, but its use is limited due to nephrotoxicity. The current study investigated the renoprotective effect of umbelliferone (UMB) on Cis-induced nephrotoxicity in rats. Renal injury was induced by a single injection of Cis (7 mg/kg, ip). Our results exhibited that the injection of Cis significantly disrupted renal function biomarkers as well as KIM-1 expression. The expressions of TNF-α, IL-1β, NF-kB-p65, and IKKβ were elevated along with downregulation of IkBα expression. Also, Cis disrupted cellular oxidant/antioxidant balance through the reduction of glutathione (GSH), glutathione-S-transferase (GST), and superoxide dismutase (SOD) levels and elevation of malondialdehyde (MDA) content. On the contrary, the levels of renal function biomarkers, cytokines, NF-kB-p65, IkBα, IKKβ, and oxidant/antioxidant status have been improved after UMB treatment. Mechanistically, rats administered Cis only exhibited a significant decrease in NRF2 and cytoglobin expressions as well as the CREB, SIRT1, FOXO-3, and PPAR-γ genes. Treatment with UMB significantly upregulated NRF2 and cytoglobin proteins, as well as effectively increased the expression of CREB, SIRT1, FOXO-3, PPAR-γ, and NRF2 genes. Histopathological findings strongly supported our biochemical results, as evidenced by attenuation of renal hemorrhage, cast diffusion, and inflammatory cell infiltration. Interestingly, UMB significantly enhanced Cis cytotoxicity in both HL-60 and HeLa cells in a dose-dependent manner. Together, our results demonstrated that UMB can protect against Cis-induced nephrotoxicity in normal rats along with the enhancement of its in vitro antitumor activity. These findings suggested that UMB could be used as a potential adjuvant therapy in Cis chemotherapeutic protocols.
It is important to prepare 'hypoimmunogenic' or 'universal' human pluripotent stem cells (hPSCs) with gene-editing technology by knocking out or in immune-related genes, because only a few hypoimmunogenic or universal hPSC lines would be sufficient to store for their off-the-shelf use. However, these hypoimmunogenic or universal hPSCs prepared previously were all genetically edited, which makes laborious processes to check and evaluate no abnormal gene editing of hPSCs.
Universal human-induced pluripotent stem cells (hiPSCs) were generated without gene editing, which were reprogrammed from foetal stem cells (human amniotic fluid stem cells) with mixing 2-5 allogenic donors but not with single donor. We evaluated human leucocyte antigen (HLA)-expressing class Ia and class II of our hiPSCs and their differentiated cells into embryoid bodies, cardiomyocytes and mesenchymal stem cells. We further evaluated immunogenic response of transient universal hiPSCs with allogenic mononuclear cells from survival rate a.To determine the potential effect of a donation after cardiac death active program on the number of organ donors in a Italian Pediatric Intensive Care Unit (PICU). We conducted a retrospective study of all deaths in PICU of an academic Children Hospital between 2012 and 2020, tracing the organ donation activity. Patients were categorized as brain deaths, deaths despite maximal resuscitation, and deaths after withdrawal or limitation of life support. Patient demographics, premortem physiology, end-of-life circumstances, and functional warm ischemia time were recorded. Eligible donors after cardiac death were identified by the absence of medical contraindication and functional warm ischemia time less then 60 minutes. Of 124 deaths that occurred during the study period, 34 met criteria for brain death, 23 were potential donors, and 13 became actual donors. Of the remaining 90 patients that met criteria for cardiac death, 66 died despite maximal resuscitation, 24 died after withdrawal or limitation of care and between them 13 were identified as theoretically eligible DCD donors. Of these, 5 patients had a functional warm ischemia time of less then 1 hour and were potential candidates for DCD of 10 kidneys and 2 lungs. Even if few children could have been eligible for DCD in the study period, an active program could have been able to increase the number of potential organ donors by 20% in the last eight years at our institution. DCD deserves to be explored in Italy as a new option for children.
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