The molecular mechanism of cardiac injury in SARS-CoV-2 infection: Focus on mitochondrial dysfunction.

BACKGROUND: Coronavirus disease 2019(COVID-19) caused a large number of infections worldwide. Although some patients recovered from the disease, some of the other problems that accompanied it, such as cardiac injury, could affect the patient's subsequent quality of life and prognosis. OBJECTIVES: To clarify the molecular mechanism of cardiac injury in SARS-CoV-2 Infection. METHODS: The RNA-Seq dataset (GSE184715) comparing expression profiling of Mock human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and SARS-CoV-2-infected hiPSC-CMs was downloaded from Gene Expression Omnibus (GEO). Differentially expressed genes(DEGs) were performed by the R software. Degs were analyzed by enrichment analysis to clarify the affected pathways. Hub genes were screened out by a PPI network constructed from Degs. Finally, Connectivity Map was used to screen for the treatment of COVID-19 induced cardiac injury. RESULTS: 2705 differentially expressed genes were identified. Enrichment analysis confirmed that mitochondrial dysfunction was caused by SARS-CoV-2, meanwhile, cardiac muscle contraction was suppressed and NF-κB was activated. Based on the PPI network, 15 hub genes were identified. These 15 down-regulated hub genes were mainly involved in the reduced activity of complexes in the mitochondrial respiratory chain associated with mitochondrial dysfunction. Moreover, 5 candidate drugs were identified to treat cardiac injury. CONCLUSION: In conclusion, SARS-CoV-2 infection of cardiomyocytes causes mitochondrial dysfunction, including reduced mitochondrial respiratory chain complex activity and decreased ATP synthesis, leading to cardiomyocyte apoptosis, while the activated NF-κB also induced cytokine storms, ultimately resulting in cardiac injury.

Kinetic changes in serum procalcitonin predict persistent acute kidney injury in critical patients.

PURPOSE: Persistent acute kidney injury (AKI) has been shown to be closely associated with poor prognosis in critical patients. Recent studies have shown that procalcitonin (PCT) is valuable for the early prediction of AKI in critically patients. Our aim was to determine whether PCT and its kinetic changes could predict the occurrence of persistent AKI in critical patients. METHODS: This is a prospective observational study. The definition of AKI was based on the Kidney Disease: Improving Global Outcomes criteria. Persistent AKI was defined as renal function that does not return to baseline serum creatinine levels within 48 h. Blood samples were obtained at the onset of AKI and two subsequent days of hospital stay. 24-h PCT change (ΔPCT-24 h) was defined as 24 h PCT minus baseline PCT (day 0). RESULTS: A total of 91 critical patients with AKI were included in this study. The persistent AKI group had a stepwise increase in PCT concentration. ΔPCT-24 h was higher in the persistent AKI group (p < .01). Logistic regression analysis showed that ΔPCT-24 h (p = .04) was independent predictors of persistent AKI. The receiver operating characteristic curves showed that area under the curve of ΔPCT-24 h was 0.84 (p < .01), and the cut-off value for PCT to predict persistent AKI was 0.56 ng/ml. CONCLUSION: Our study showed that the observation of kinetic changes in PCT is more significant for the early prediction of persistent AKI than the index of PCT at a single time point. ΔPCT-24 h is a good predictor of persistent AKI in critical patients.

Combination of hepcidin with neutrophil gelatinase-associated lipocalin for prediction of the development of sepsis-induced acute kidney injury.

BACKGROUND AND AIMS: The early prediction of the development of acute kidney injury (AKI) in critically ill patients with sepsis would facilitate early effective intervention. Recently, interest has focused on the biomarkers for AKI-linked iron metabolism. This study aimed to assess the early predictive values of hepcidin, neutrophil gelatinase-associated lipocalin (NGAL), and their combination for secondary AKI in patients with sepsis. MATERIALS AND METHODS: A prospective cohort study was performed in septic patients. Serum and urine hepcidin, and urine NGAL were analyzed at admission. The primary outcome measure was occurrence of sepsis-induced AKI based on 2011 Kidney Disease: Improving Global Outcomes (KDIGO) criteria during the first week of ICU stay. RESULTS: Of the 90 patients analyzed finally in the study, 44 (48.9%) patients developed AKI. Patients with AKI occurrence were more likely than those without AKI to have higher serum hepcidin and urine NGAL levels at admission (P < 0.01). Higher concentrations of these biomarkers were each independent predictor of the development of AKI in critically septic patients within the first week of their ICU stay. Serum hepcidin and urine NGAL (AUROC 0.787, 95% CI 0.688 to 0.8660 and AUROC 0.729, 95% CI 0.625 to 0.818, respectively) were comparable predictive indicators of AKI occurrence (P = 0.43 for DeLong's test). Combining both biomarkers increased the AUROC to 0.828(95% CI 0.733 to 0.899), and this performance was statistically significantly better than urine NGAL alone (P = 0.03 for DeLong's test). CONCLUSION: Serum hepcidin measured at admission predicts the development of AKI similarly to urine NGAL. However, serum hepcidin adds significant accuracy to this prediction in combination with urine NGAL alone and has a good predictive value in patients with sepsis. Larger studies are needed to validate and explain these findings.

Mortality and serum hepcidin are associated with persistent and transient acute kidney injury in septic patients.

PURPOSE: Acute kidney injury (AKI) is a common complication of sepsis and has high mortality. The 2017 Acute Disease Quality Initiative (AQDI) workgroup proposed new definitions for AKI - transient AKI and persistent AKI; however, very little is known about the effect of transient and persistent septic AKI on short-term mortality among critically ill patients with sepsis. The purpose of this study was to assess the impact of persistent AKI on mortality and to evaluate whether serum hepcidin can predict the occurrence of persistent AKI in critically ill patients with sepsis. MATERIALS AND METHODS: This prospective observational study was performed in a general hospital mixed surgical-medical ICU in Pudong, China. Consecutive adults with sepsis admitted to the ICU with absence of chronic kidney disease, renal transplant, and AKI were included. AKI was defined according to the KDIGO criteria and classified as transient (< 48-hour duration) or persistent (48-hour duration). Blood samples were obtained within 6 hours from when AKI was diagnosed. RESULTS: A total of 90 patients with sepsis or septic shock were included in the analysis. 44 (48.89%) patients developed AKI during ICU stay: 20 (45.45%) had transient and 24 (54.55%) had persistent AKI. Persistent AKI has a higher mortality than transient AKI (66.7 vs. 30.0%, p = 0.002). Persistent AKI and sequential organ failure assessment (SOFA) scores were an independent predictor of 60-day mortality. Patients with persistent AKI had higher concentrations of serum creatinine (SCr) and hepcidin than transient AKI patients when AKI was diagnosed. Logistic regression indicated that serum hepcidin was an independent predictor of persistent AKI in septic patients, with a fairly predictive value (AUC 0.71, 95% CI: 0.47 - 0.87; p = 0.02). CONCLUSION: Persistent AKI was associated with increased 60-day mortality compared with transient AKI in septic patients. The serum hepcidin levels measured when AKI was diagnosed have a fair predictive value to predict the occurrence of persistent AKI in septic patients.

Characterization of progression-related alternative splicing events in testicular germ cell tumors.

Accumulating evidence supports the significance of aberrant alternative splicing (AS) events in cancer; however, genome-wide profiling of progression-free survival (PFS)-related AS events in testicular germ cell tumors (TGCT) has not been reported. Here, we analyzed high-throughput RNA-sequencing data and percent-spliced-in values for 150 patients with TGCT. Using univariate and multivariate Cox regression analysis and a least absolute shrinkage and selection operator method, we identified the top 15 AS events most closely associated with disease progression. A risk-associated AS score (ASS) for the 15 AS events was calculated for each patient. ASS, pathological stage, and T stage were significantly associated with disease progression by univariate analysis, but only ASS and pathological stage remained significant by multivariate analysis. The ability of these variables to predict 5-year progression was assessed using receiver operating characteristic curve analysis. ASS had stronger predictive value than a combination of age, pathological stage, and T stage (area under the curve = 0.899 and 0.715, respectively). Furthermore, Kaplan-Meier analysis of patients with low and high ASS demonstrated that high ASS was associated with significantly worse PFS than low ASS (P = 1.46 × 10-7). We also analyzed the biological functions of the PFS-related AS-related genes and found enrichment in pathways associated with DNA repair and modification. Finally, we identified a regulatory network of splicing factors with expression levels that correlated significantly with AS events in TGCT. Collectively, this study identifies a novel method for risk stratification of patients and provides insight into the molecular events underlying TGCT.

Mesenchymal stem cell exosomes reverse acute lung injury through Nrf-2/ARE and NF-κB signaling pathways.

Acute lung injury (ALI) is associated with histopathological diffuse alveolar damage. The potential role of mesenchymal stem cells (MSCs) in the treatment of various clinical disorders have been widely documented, such as those for ALI. Recent evidence has demonstrated that exosomes from endothelial progenitor cells can improve outcomes of the lipopolysaccharide (LPS)-induced ALI. However, there has been no research on the potential role of MSC-exosomes in the treatment of sepsis-induced ALI, which is worth further exploration. Thus, the objective of our study was to identify whether the MSC-exosomes could reverse ALI. The ALI model induced by LPS was established in this study. MTT assay was performed to test cell proliferation. Expression of inflammatory factors (TNF-α, IL-6, and IL-10) in the LPS-treated type II alveolar epithelial cells (AECs) (MLE-12) was detected by ELISA. After co-culture of MSC-exosomes with LPS-treated MLE-12 cells, we found that the cell proliferation of MLE-12 cells gradually increased. Furthermore, we selected five of the Nrf-2/ARE- and NF-κB signaling pathway-related genes to explore if MSC-exosomes could reverse LPS-induced ALI through Nrf-2/ARE and NF-κB signaling pathways. QRT-PCR and western blot experiment results showed that the expression of these five genes were significantly regulated after stimulation with high-concentration LPS and exosome intervention. Taken together, these findings highlighted the fact that MSC-exosomes could reverse ALI through the Nrf-2/ARE and NF-κB signaling pathways. The MSC-exosome may be the potential future therapeutic strategy for the treatment of ALI.

The protective effect of ticagrelor on renal function in a mouse model of sepsis-induced acute kidney injury.

Platelets are traditionally considered to be essential components of primary hemostasis. Recent investigations have revealed that platelets can be activated in patients with sepsis and are implicated in the development of sepsis and sepsis-induced-acute kidney injury (SAKI). In the present study, ticagrelor was used to induce a mouse model of SAKI by cecal ligation and puncture. It was found that ticagrelor could inhibit platelet activity, decrease the levels of interleukin-1ß and serum creatinine, reduce infiltration of neutrophils in renal tissue, and attenuate cell apoptosis in the kidney. The results suggested that ticagrelor could protect renal function by inhibiting inflammation, recruitment of neutrophils into the kidney, and cell apoptosis in renal tissue. Thus, the findings might provide new strategies for preventing SAKI.

[Value of Hepcidin as a diagnostic biomarker of sepsis in critically ill adults].

OBJECTIVE: To investigate the diagnostic value of Hepcidin as a sepsis biomarker in critically ill adults. METHODS: An observational study was conducted. The patients with suspected or proven infection admitted to intensive care unit (ICU) of Zhoupu Hospital Affiliated to Shanghai University of Medicine and Health Sciences from March 2016 to November 2017 were enrolled. According to the third international consensus definitions for sepsis and septic shock (Sepsis-3), the patients were divided into non-sepsis group and sepsis group, and the septic patients were subdivided into general sepsis subgroup and septic shock subgroup according to the severity of disease. The differences in serum Hepcidin, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), procalcitonin (PCT), C-reactive protein (CRP), white blood cell (WBC), neutrophil granulocytes (NEUT) and lactic acid (Lac) within 1 hour after ICU admission between non-sepsis and sepsis groups and among the sepsis subgroups were compared. The acute physiology and chronic health evaluation II (APACHE II) within 24 hours after ICU admission and sequential organ failure score (SOFA) were recorded, and the mortality rate was followed up for 28 days. Receiver operation characteristic curve (ROC) was used to evaluate and compare the diagnostic value of Hepcidin and PCT, CRP, WBC for sepsis. Logistic regression model was used to estimate the association between Hepcidin and sepsis. Spearman correlation analysis was used to analyze the correlation between Hepcidin and other parameters of sepsis patients. RESULTS: A total of 183 patients were enrolled, 93 in the non-sepsis group and 90 in the sepsis group (48 with general sepsis and 42 with septic shock). (1) The levels of Hepcidin, IL-6, TNF-α, PCT, Lac in serum, and APACHE II and SOFA scores in the sepsis group were significantly higher than those in the non-sepsis group. ROC analysis showed that the area under the ROC curve (AUC) of Hepcidin and PCT for sepsis diagnosis were 0.865 [95% confidence interval (95%CI) = 0.807-0.911] and 0.848 (95%CI = 0.788-0.897), respectively, without statistical significance (Z = 0.443, P = 0.657). Furthermore, the AUC of Hepcidin for sepsis diagnosis was significantly higher than that of the conventional biomarkers CRP and WBC [AUC was 0.530 (95%CI = 0.455-0.604) and 0.527 (95%CI = 0.452-0.601), respectively] with statistical significance (both P < 0.01). When Hepcidin > 54.00 µg/L, its sensitivity for sepsis diagnosis was 95.56%, specificity was 66.67%, positive and negative predictive value was 73.51% and 93.94%, respectively. Parallel test was conducted for combination of Hepcidin and PCT, which showed that the AUC was 0.885, and the sensitivity and negative predictive value was significantly improved to 98.96% and 98.36%, respectively. Logistic regression analysis demonstrated that after adjusted for PCT, Hepcidin > 54.00 µg/L was also associated with sepsis independently, with odds ratio (OR) of 1.011 (95%CI = 1.008-1.015, P < 0.001), indicating that Hepcidin and PCT were not completely overlapped in the diagnosis of sepsis. (2) With the increase in infection severity, serum Hepcidin, PCT, IL-6, TNF-α, Lac, APACHE II, SOFA score and 28-day mortality all showed an increasing trend in patients. There was a significantly positive correlation between Hepcidin and IL-6, TNF-α, PCT, APACHE II, and SOFA in the sepsis patients (r value was 0.526, 0.449, 0.591, 0.359, and 0.374, respectively, all P < 0.01), but no correlation was found between Hepcidin and Lac (r = 1.104, P > 0.05). CONCLUSIONS: Serum Hepcidin is a useful biomarker for the diagnosis of sepsis, and it is correlated to the severity of the sepsis. The combination of Hepcidin and PCT can improve the accuracy of diagnosis of sepsis. CLINICAL TRIAL REGISTRATION: China Clinical Trial Registration Center, ChiCTR-DDD-16008522.

Cluster secondary ion mass spectrometry imaging of interfacial reactions of TiO2 microspheres embedded in ionic liquids.

RATIONALE: Our goal is to develop protocols for the elucidation of the identity and structure of reaction products embedded in a reaction medium. Results should find significance in a variety of disciplines ranging from the study of biological cells and tissues, to the steps associated with the functionalization of nanoparticles. METHODS: We utilize cluster secondary ion mass spectrometry (cluster-SIMS) to acquire three-dimensional (3D) information about 5-30 µm TiO2 microspheres imbedded into an ionic liquid. The method allows molecular depth profiling with submicron spatial resolution and depth profiling with a resolution of several tens of nanometers. The ionic liquid matrix enshrouds the spheres, allowing them to be introduced into the vacuum environment of the mass spectrometer. RESULTS: The results provide 3D chemical information about these microspheres as they are synthesized by interfacial sol-gel reactions. We show that with 40 keV C60 (+) , it is possible to erode through the reaction medium and map the distribution of those embedded TiO2 microspheres. Moreover, we demonstrate that it is possible to monitor surface modification of the particles and, via ion beam drilling, elucidate their internal structure. CONCLUSIONS: Using cluster-SIMS imaging, we are able to elucidate the identity and structure of reaction products embedded in a reaction medium, a problem of long-standing interest for materials characterization. With this strategy, we have provided a new approach that may be especially useful for the characterization of biological tissue and cells within the vacuum confines of the mass spectrometer. Copyright © 2015 John Wiley & Sons, Ltd.

Relative ion yields in mammalian cell components using C60 SIMS.

Time of flight secondary ion mass spectrometry has been used to better understand the influence of molecular environment on the relative ion yields of membrane lipid molecules found in high abundance in a model mammalian cell line, RAW264.7. Control lipid mixtures were prepared to simulate lipid-lipid interactions in the inner and outer leaflet of cell membranes. Compared with its pure film, the molecular ion yields of 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine are suppressed when mixed with 2-dipalmitoyl-sn-glycero-3-phosphocholine. In the mixture, proton competition between 1,2-dioleoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, and 2-dipalmitoyl-sn-glycero-3-phosphocholine led to lower ionization efficiency. The possible mechanism for ion suppression was also investigated with 1H and 13C nuclear magnetic resonance spectroscopy. The formation of a hydroxyl bond in lipid mixtures confirms the mechanism involving proton exchange with the surrounding environment. Similar effects were observed for lipid mixtures mimicking the composition of the inner leaflet of cell membranes. The secondary molecular ion yield of 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine was observed to be enhanced in the presence of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine.

Depth profiling of metal overlayers on organic substrates with cluster SIMS.

Molecular depth profiling of organic thin films by erosion with energetic cluster ion beams is a unique aspect of secondary ion mass spectrometry (SIMS) experiments. Although depth profiles of complex multilayer organic structures can be acquired with little damage accumulation and with depth resolution of <10 nm using either C60(+) or Arx(+) with x = 500-5000, hybrid materials consisting of both organic and inorganic layers often yield poor results. To unravel the factors that lead to this difficulty, we developed a model system composed of a thin gold layer of 1.4 to 3.5 nm deposited either on top of or sandwiched within a cholesterol thin film matrix which is several hundred nanometers thick. For these systems, the results show that by erosion with a 40 keV C60(+) beam, reliable depth profiles can always be acquired as indicated by the presence of a steady state molecular ion signal. During the erosion process, however, gold atoms from the gold overlayer are implanted into the cholesterol matrix beneath it, resulting in a reduced sputter yield, an increase in the amount of cholesterol fragmentation and an increase in the thickness of the cluster ion-induced altered layer. The results also show that the effects of the metal film on the organic substrate are independent of the gold film thickness once the film thickness exceeds 1.4 nm. In general, this model study provides mechanistic insight into the depth profiling of heterogeneous thin film structures and offers a possible path for improving the quality of the depth profiles by employing low energy atomic ion sputtering in the region of the metal layer.

C60-ToF SIMS imaging of frozen hydrated HeLa cells.

Sample preparation continues to be a major challenge for secondary ion mass spectrometry studies of biological materials. Maintaining the native hydrated state of the material is important for preserving both chemical and spatial information. Here, we discuss a method which combines a sample wash and dry protocol discussed by Berman et al1 (1) followed by plunge freezing in liquid ethane for a frozen-hydrated analysis of mammalian cells (HeLa). This method allows for the removal of the growth media and maintains the hydrated state of the cells so that they can be prepared frozen-hydrated without the need for a freeze-fracture device. The cells, which were grown on silicon, have been successfully re-grown after the cleaning procedure, confirming that a significant portion of the cells remain undamaged during the wash and dry. Results from preliminary SIMS measurements show that is it possible to detect a large variety of bio-molecular signals, including intact lipids from the plasma membrane in the mass range of 700-900 Da from single cells, with little external water interference at the surface.

Preliminary investigations on a new disposable potentiometric biosensor for uric acid.

In this paper, uricase, catalase, and electron mediator were coimmobilized on the surface of the tin oxide (SnO2)/indium tin oxide (ITO) glass, to develop a disposable potentiometric uric acid biosensor. The SnO2/ITO glass was employed as a pH sensor, fabricated by sputtering SnO2 thin films on the ITO glass. 3-Glycidyloxypropyltrimethoxysilane (GPTS) was utilized to immobilize uricase, catalase and the electron mediator (ferrocenecarboxylic acid, FcA) on the sensing window. The experimental results reveal that the optimal weight ratio of uricase, FcA to catalase (CAT) is 4:1:2. The sensor responds linearly between 2 mg/dl and 7 mg/dl at pH 7.5, in 20 mM of test solution, with a correlation coefficient of 0.99213. Accordingly, no significant interference was observed when interfering substances, glucose, urea and ascorbic acid, were added to the uric acid solution. Moreover, the recorded voltage was relatively constant during the first 28 days of measurement. Consequently, a potentiometric uric acid biosensor was realized with the advantages of low cost and simple fabrication.