High phosphate and calcium induce osteoblastic phenotype switching and calcification of corneal epithelial cells in a Runx2-dependent and synergistic manner; a possible mechanism of chronic kidney disease-associated corneal calcification.

Patients with advanced chronic kidney disease (CKD) have elevated circulating calcium × phosphate product levels and exhibit soft tissue calcification. Besides the cardiovascular system, calcification is commonly observed in the cornea in CKD patients on hemodialysis. Cardiovascular calcification is a cell-mediated, highly regulated process, and we hypothesized that a similar regulatory mechanism is implicated in corneal calcification with the involvement of corneal epithelial cells (CECs). We established a mouse model of CKD-associated corneal calcification by inducing CKD in DBA/2J mice with an adenine and high phosphate diet. CKD was associated with aorta and corneal calcification as detected by OsteoSense staining and corneal Ca measurement (1.67-fold elevation, p < 0.001). In vitro, excess phosphate and Ca induced human CEC calcification in a dose-dependent and synergistic manner, without any influence on cell viability. High phosphate and Ca-containing osteogenic medium (OM; 2.5 mmol/L excess phosphate and 0.6 mmol/L excess Ca over control) increased the protein expression of Runx2 and induced its nuclear translocation. OM increased the expression of the bone-specific Ca-binding protein osteocalcin (130-fold increase, p < 0.001). Silencing of Runx2 attenuated OM-induced CEC calcification. Immunohistology revealed upregulation of Runx2 and overlapping between the Runx2 and the Alizarin red positive areas of calcification in the cornea of CKD mice. This work sheds light on the mechanism of CKD-induced corneal calcification and provides tools to test calcification inhibitors for the prevention of this detrimental process.

Ponatinib Induces a Procoagulant Phenotype in Human Coronary Endothelial Cells via Inducing Apoptosis.

BCR-ABL tyrosine kinase inhibitors (TKIs) are effective drugs in the treatment of patients with chronic myeloid leukemia. However, based on clinical studies, ponatinib was associated with the development of thrombotic complications. Since endothelial cells (ECs) regulate blood coagulation, their abnormal phenotype may play a role in the development of thrombotic events. We here aimed to investigate the effect of ponatinib on the procoagulant activity of cultured endothelial cells in vitro. Human coronary artery endothelial cells (HCAECs) were incubated with 50, 150, and 1000 nM of ponatinib. Subsequently, phosphatidylserine (PS) exposure and endothelial microvesicles (EMVs) were measured by flow cytometry. In addition, EC- and EMV-dependent thrombin generation was analyzed. To investigate pro-apoptotic effects of ponatinib, the level of Bax and Bcl-xL proteins were studied using Western blot and F3, THBD, and VCAM1 mRNAs were quantified by qPCR. Therapeutic concentrations of ponatinib significantly increased PS expression on ECs and the amount of EMVs which significantly shortened the time parameters of thrombin generation. In addition, these changes were associated with an increased ratio of Bax and Bcl-xL proteins in the presence of the decreased THBD mRNA level. Overall, ponatinib enhances the procoagulant activity of ECs via inducing apoptosis, which may contribute to thrombotic events.

Inflammation, Sepsis, and the Coagulation System.

Sepsis has been a major health problem for centuries and it is still the leading cause of hospital deaths. Several studies in the past decades have identified numerous biochemical abnormalities in severe cases, and many of these studies provide evidence of the perturbation of the hemostatic system. This can result in complications, such as disseminated intravascular coagulation that can lead to multiorgan failure. Nevertheless, large clinical studies have demonstrated that the simple approach of inhibiting the coagulation processes by any means fails to provide significant improvement in the survival of septic patients. A cause of this failure could be the fact that in sepsis the major clinical problems result not primarily from the presence of the infective agent or enhanced coagulation but from the complex dysregulated systemic host response to pathogens. If this overt reaction is not fully deciphered, appropriate interference is highly unlikely and any improvement by conventional therapeutic interventions would be limited. Cellular activation in sepsis can be targeted by novel approaches like inhibition of the heterotypic cellular interactions of blood cells by targeting surface receptors or posttranscriptional control of the hemostatic system by noncoding ribonucleic acid (RNA) molecules. Stable RNA molecules can affect the expression of several proteins. Thus, it can be anticipated that modulation of microRNA production would result in a multitude of effects that may be beneficial in septic cases. Here, we highlight some of the recent diagnostic possibilities and potential novel routes of the dysregulated host response.

Comparison of Different Vascular Biomarkers for Predicting In-Hospital Mortality in Severe SARS-CoV-2 Infection.

Severe SARS-CoV-2 elicits a hyper-inflammatory response that results in intravascular inflammation with endothelial injury, which contributes to increased mortality in COVID-19. To predict the outcome of severe SARS-CoV-2 infection, we analyzed the baseline level of different biomarkers of vascular disorders in COVID-19 subjects upon intensive care unit (ICU) admission and prior to any vaccination. A total of 70 severe COVID-19 patients (37 survivors and 33 non-survivors) were included with 16 age- and sex-matched controls. Vascular dysfunction was monitored via soluble VCAM-1, E-selectin, ACE2 and Lp-PLA2, while abnormal platelet activation was evaluated by soluble P-selectin and CD40L in parallel. These results were correlated with routine laboratory parameters and disease outcomes. Among these parameters, VCAM-1 and ACE2 showed significantly higher serum levels in COVID-19 patients with early death vs. convalescent subjects. VCAM-1 was significantly correlated with the Horowitz index (r = 0.3115) and IL-6 (r = 0.4599), while ACE2 was related to E-selectin (r = 0.4143) and CD40L (r = 0.2948). Lp-PLA2 was altered in none of these COVID-19 subcohorts and showed no relationship with the other parameters. Finally, the pre-treatment level of VCAM-1 (≥1420 ng/mL) and ACE2 activity (≥45.2 µU/mL) predicted a larger risk for mortality (Log-Rank p = 0.0031 and p = 0.0117, respectively). Vascular dysfunction with endothelial cell activation is linked to lethal COVID-19, and highly elevated soluble VCAM-1 and ACE2 at admission to ICU may predict unfavorable outcomes.

Distinct subsets of anti-pulmonary autoantibodies correlate with disease severity and survival in severe COVID-19 patients.

Autoantibodies targeting the lung tissue were identified in severe COVID-19 patients in this retrospective study. Fifty-three percent of 104 patients developed anti-pulmonary antibodies, the majority of which were IgM class, suggesting that they developed upon infection with SARS-CoV-2. Anti-pulmonary antibodies correlated with worse pulmonary function and a higher risk of multiorgan failure that was further aggravated if 3 or more autoantibody clones were simultaneously present (multi-producers). Multi-producer patients were older than the patients with less or no autoantibodies. One of the identified autoantibodies (targeting a pulmonary protein of ~ 50 kDa) associated with worse clinical outcomes, including mortality. In summary, severe COVID-19 is associated with the development of lung-specific autoantibodies, which may worsen the clinical outcome. Tissue proteome-wide tests, such as the ones applied here, can be used to detect autoimmunity in the post-COVID state to identify the cause of symptoms and to reveal a new target for treatment.

Elevated level of serum human epididymis protein 4 (HE4) predicts disease severity and mortality in COVID-19 pneumonia.

BACKGROUND: We retrospectively analyzed serum level of human epididymis protein 4 (HE4) as a pulmonary inflammatory biomarker in patients with COVID-19 pneumonia in association with disease severity and outcome. METHODS: Ninety-nine (40 critically ill, 40 severe and 19 mild) COVID-19 patients and as controls 25 age- and sex-matched non-COVID-19 bacterial sepsis subjects were included. Serum HE4 was measured by an immunoassay (Architect® i1000SR, Abbott) in the baseline samples of all study participants obtained at intensive care unit (ICU) admission or during outpatient clinic visit and follow-up sera were available in case of 30 COVID-19 subjects with life-threating conditions. Associations were studied between serum HE4, routinely available laboratory parameters, clinical characteristics, and disease progression. RESULTS: Baseline HE4 level was significantly higher (P < 0.0001) in critically ill (524.7 [300.1-1153.0] pmol/L) than severe COVID-19 subjects (157.4 [85.2-336.9] pmol/L) and in mild SARS-CoV-2 infection (46.7 [39.1-57.2] pmol/L). Similarly increased HE4 concentrations were found in bacterial sepsis (1118.0 [418.3-1953.0] pmol/L, P = 0.056) compared to critically ill COVID-19 individuals. Serum HE4 levels significantly correlated with age, SOFA-score, inflammation-dependent biomarkers, and the degree of lung manifestation evaluated by chest CT examination in ICU COVID-19 individuals. Based on ROC-AUC curve analysis, baseline HE4 independently indicated the severity of COVID-19 with an AUC value of 0.816 (95% CI [0.723-0.908]; P < 0.0001), while binary logistic regression test found HE4 as an independent prognostic parameter for death (OR: 10.618 [2.331-48.354]; P = 0.002). Furthermore, COVID-19 non-survivors showed much higher baseline HE4 levels without a substantial change under treatment vs. survivors (P < 0.0001). Finally, pre-treatment HE4 level of ≥ 331.7 pmol/L effectively predicted a larger risk for mortality (Log-Rank P < 0.0001) due to severe COVID-19 pneumonia. CONCLUSION: Elevated serum HE4 level at ICU admission highly correlates with COVID-19 severity and predicts disease outcome.

Technically Challenging Percutaneous Interventions of Chronic Total Occlusions Are Associated with Enhanced Platelet Activation.

Percutaneous coronary intervention (PCI) is a frequently performed treatment option for recanalization in patients with chronic total occlusion (CTO). As CTO-PCIs are often complicated and challenging for interventionalists, the stressful and damaging nature of the procedure can be remarkable, thus platelets can be easily activated. Our aim was to investigate the effect of CTO-PCI on platelet activation and the expression of selected circulating microRNAs (miR) of platelet and endothelium origin after CTO-PCI. In this study, 50 subjects after CTO-PCI were enrolled. Blood samples were obtained before PCI, at 2 days and 3-6 months after the procedure to measure the degree of platelet activation and the level of plasma miR-223, miR-181b, and miR-126. Patients were divided based on the characteristics of the intervention. Patients with higher Japanese CTO scores and longer duration of PCI showed significantly elevated platelet P-selectin positivity (p = 0.004 and p = 0.013, respectively) 2 days after the procedure compared to pre-PCI and increased concentration of soluble P-selectin 3-6 months after the intervention (higher Japanese CTO score: p = 0.028 and longer duration of PCI: p = 0.023) compared to baseline values. Shorter total stent length caused a significantly lower miR-181b expression at 3-6 months after the intervention (p = 0.031), while no difference was observed in miR-223 and miR-126. One stent thrombosis occurred during the follow-up period. Although these technically challenging CTO-PCIs may cause enhanced platelet activation right after the intervention and long-term endothelial cell dysfunction, these interventions are not associated with more adverse clinical events.

Effect of anti-SARS-CoV-2 BNT162b2 mRNA vaccination on thrombin generation in children with inflammatory bowel disease.

Background: Inflammatory bowel disease (IBD) including Crohn's disease (CD) and ulcerative colitis (UC), are associated with higher thrombotic risk and enhanced thrombin generation (TG) in adults. Despite encouraging data reporting vaccine safety and low IBD flare rates in adults with IBD, vaccine hesitancy was demonstrated to be high in families of children with IBD. We aimed to find out whether TG is increased in children with IBD as compared to healthy controls and whether TG parameters show significant changes following SARS-CoV-2 mRNA vaccination. Patients and methods: In this observational case-control study, 38 children with IBD (CD:18, UC: 20) aged 12-18 years and 62 healthy age-and sex-matched children were enrolled. Blood was collected before the first dose and 2-6 weeks after the second dose of BNT162b2 (Pfizer-BioNTech) mRNA vaccine dose. Blood cell counts, fibrinogen, inflammatory markers (hsCRP, ferritin), anti-SARS-CoV-2 antibody levels were investigated, TG assay was carried-out using platelet-poor plasma. Detailed clinical parameters including disease activity scores (PUCAI, PCDAI) were registered pre-and post- vaccination. A guided questionnaire was used to collect data on adverse reactions (AEs) post- vaccination. Results: Baseline TG parameters did not differ between patients and controls. Endogenous thrombin potential showed a significant positive correlation with markers of inflammation and with PCDAI. Inflammatory parameters and TG did not increase in patients and controls post-vaccination. Vaccination significantly increased antibody levels in all three investigated groups, but post-vaccination anti-SARS-CoV-2 S IgG/IgM levels were below the 5th percentile value of healthy children in more than one third of patients. Those receiving TNFα inhibitor therapy presented significantly lower SARS-CoV-2 S IgG/IgM levels as compared to patients on other immunosuppressive regimens. Systemic AEs did not differ between patients and controls while lower rate of local symptoms was found post-vaccination in children with IBD. Only 2 IBD flares were detected 2-6 weeks after the second dose of vaccination. Conclusion: Our study is the first to support the safety and efficacy of anti-SARS-CoV-2 BNT162b2 vaccination in children with IBD with detailed pre-and post-vaccination laboratory data including TG. Results of this study may further increase confidence and reduce vaccine hesitancy in caretakers of pediatric IBD patients.

Shikonin Causes an Apoptotic Effect on Human Kidney Cancer Cells through Ras/MAPK and PI3K/AKT Pathways.

(1) Background: Shikonin, the main ingredient in Chinese herbal medicine, is described as a novel angiogenesis inhibitor, and its anticancer effects have already been studied. Shikonin and its derivatives induce apoptosis and suppress metastasis, which further enhance the effectiveness of chemotherapy. However, their mechanism of function has not been completely elucidated on human renal cancer cells. (2) Methods: In our study, CAKI-2 and A-498 cells were treated with increasing concentrations (2.5-40 µM) of shikonin, when colony formation ability and cytotoxic activity were tested. The changes in the expression of the main targets of apoptotic pathways were measured by RT-qPCR and Western blot. The intracellular levels of miR-21 and miR-155 were quantified by RT-qPCR. (3) Results: Shikonin exerted a dose-dependent effect on the proliferation of the cell lines examined. In 5 µM concentration of shikonin in vitro elevated caspase-3 and -7 levels, the proteins of the Ras/MAPK and PI3K/AKT pathways were activated. However, no significant changes were detected in the miR-21 and miR-155 expressions. (4) Conclusions: Our findings indicated that shikonin causes apoptosis of renal cancer cells by activating the Ras/MAPK and PI3K/AKT pathways. These effects of shikonin on renal cancer cells may bear important potential therapeutic implications for the treatment of renal cancer.

Hypoxia-inducible factor activation promotes osteogenic transition of valve interstitial cells and accelerates aortic valve calcification in a mice model of chronic kidney disease.

Introduction: Valve calcification (VC) is a widespread complication in chronic kidney disease (CKD) patients. VC is an active process with the involvement of in situ osteogenic transition of valve interstitial cells (VICs). VC is accompanied by the activation of hypoxia inducible factor (HIF) pathway, but the role of HIF activation in the calcification process remains undiscovered. Methods and result: Using in vitro and in vivo approaches we addressed the role of HIF activation in osteogenic transition of VICs and CKD-associated VC. Elevation of osteogenic (Runx2, Sox9) and HIF activation markers (HIF-1α and HIF-2α) and VC occurred in adenine-induced CKD mice. High phosphate (Pi) induced upregulation of osteogenic (Runx2, alkaline-phosphatase, Sox9, osteocalcin) and hypoxia markers (HIF-1α, HIF-2α, Glut-1), and calcification in VICs. Down-regulation of HIF-1α and HIF-2α inhibited, whereas further activation of HIF pathway by hypoxic exposure (1% O2) or hypoxia mimetics [desferrioxamine, CoCl2, Daprodustat (DPD)] promoted Pi-induced calcification of VICs. Pi augmented the formation of reactive oxygen species (ROS) and decreased viability of VICs, whose effects were further exacerbated by hypoxia. N-acetyl cysteine inhibited Pi-induced ROS production, cell death and calcification under both normoxic and hypoxic conditions. DPD treatment corrected anemia but promoted aortic VC in the CKD mice model. Discussion: HIF activation plays a fundamental role in Pi-induced osteogenic transition of VICs and CKD-induced VC. The cellular mechanism involves stabilization of HIF-1α and HIF-2α, increased ROS production and cell death. Targeting the HIF pathways may thus be investigated as a therapeutic approach to attenuate aortic VC.

Evaluation of the Diagnostic Performance of Two Automated SARS-CoV-2 Neutralization Immunoassays following Two Doses of mRNA, Adenoviral Vector, and Inactivated Whole-Virus Vaccinations in COVID-19 Naïve Subjects.

BACKGROUND: Limited data are available on humoral responses determined by automated neutralization tests following the administration of the three different types of COVID-19 vaccinations. Thus, we here evaluated anti-SARS-CoV-2 neutralizing antibody titers via two different neutralization assays in comparison to total spike antibody levels. METHODS: Healthy participants (n = 150) were enrolled into three subgroups who were tested 41 (22-65) days after their second dose of mRNA (BNT162b2/mRNA-1273), adenoviral vector (ChAdOx1/Gam-COVID-Vac) and inactivated whole-virus (BBIBP-CorV) vaccines, with no history or serologic evidence of prior SARS-CoV-2 infection. Neutralizing antibody (N-Ab) titers were analyzed on a Snibe Maglumi® 800 instrument and a Medcaptain Immu F6® Analyzer in parallel to anti-SARS-CoV-2 S total antibody (S-Ab) levels (Roche Elecsys® e602). RESULTS: Subjects who were administered mRNA vaccines demonstrated significantly higher SARS-CoV-2 N-Ab and S-Ab levels compared to those who received adenoviral vector and inactivated whole-virus vaccinations (p < 0.0001). N-Ab titers determined by the two methods correlated with each other (r = 0.9608; p < 0.0001) and S-Ab levels (r = 0.9432 and r = 0.9324; p < 0.0001, respectively). Based on N-Ab values, a new optimal threshold of Roche S-Ab was calculated (166 BAU/mL) for discrimination of seropositivity showing an AUC value of 0.975 (p < 0.0001). Low post-vaccination N-Ab levels (median value of 0.25 µg/mL or 7.28 AU/mL) were measured in those participants (n = 8) who were infected by SARS-CoV-2 within 6 months after immunizations. CONCLUSION: Both SARS-CoV-2 N-Ab automated assays are effective to evaluate humoral responses after various COVID-19 vaccines.

Karyotype differentiation in the Nothobranchiusugandensis species group (Teleostei, Cyprinodontiformes), seasonal fishes from the east African inland plateau, in the context of phylogeny and biogeography.

The karyotype differentiation of the twelve known members of the Nothobranchiusugandensis Wildekamp, 1994 species group is reviewed and the karyotype composition of seven of its species is described herein for the first time using a conventional cytogenetic protocol. Changes in the architecture of eukaryotic genomes often have a major impact on processes underlying reproductive isolation, adaptation and diversification. African annual killifishes of the genus Nothobranchius Peters, 1868 (Teleostei: Nothobranchiidae), which are adapted to an extreme environment of ephemeral wetland pools in African savannahs, feature extensive karyotype evolution in small, isolated populations and thus are suitable models for studying the interplay between karyotype change and species evolution. The present investigation reveals a highly conserved diploid chromosome number (2n = 36) but a variable number of chromosomal arms (46-64) among members of the N.ugandensis species group, implying a significant role of pericentric inversions and/or other types of centromeric shift in the karyotype evolution of the group. When superimposed onto a phylogenetic tree based on molecular analyses of two mitochondrial genes the cytogenetic characteristics did not show any correlation with the phylogenetic relationships within the lineage. While karyotypes of many other Nothobranchius spp. studied to date diversified mainly via chromosome fusions and fissions, the N.ugandensis species group maintains stable 2n and the karyotype differentiation seems to be constrained to intrachromosomal rearrangements. Possible reasons for this difference in the trajectory of karyotype differentiation are discussed. While genetic drift seems to be a major factor in the fixation of chromosome rearrangements in Nothobranchius, future studies are needed to assess the impact of predicted multiple inversions on the genome evolution and species diversification within the N.ugandensis species group.

[The role of classic serum tumor markers in the diagnosis and treatment monitoring of solid tumors in adults]. / A klasszikus szérumtumormarkerek helye a felnottkori daganatok kivizsgálásában és a terápia monitorizálásában.

Malignancies are considered as leading cause of death in parallel to cardio- and cerebrovascular diseases and their incidence is still growing from year to year. Early detection and monitoring of cancers after complex therapeutic interventions are essential for the survival of patients. In these aspects, beside radiological investigations, some lab tests play a key role, namely the tumor markers. These mostly protein-based mediators are produced in a large quantity by either cancer cells or the human body itself in response to the development of tumor. Measurement of tumor markers is usually assessed in serum samples, however, to locally detect an early malignant event, other body fluids, such as ascites, cerebrospinal fluid, or pleural effusion sample can also be analyzed. Due to the potential effects of other non-malignant conditions on the serum level of a tumor marker, the entire clinical status of investigated person needs to be considered for the correct interpretation of these results. In this review article, we summarized some important characteristics of the most widely used tumor markers.

Human epididymis protein 4 (HE4) plasma concentration inversely correlates with the improvement of cystic fibrosis lung disease in p.Phe508del-CFTR homozygous cases treated with the CFTR modulator lumacaftor/ivacaftor combination.

BACKGROUND: We previously documented that elevated HE4 plasma concentration decreased in people with CF (pwCF) bearing the p.Gly551Asp-CFTR variant in response to CFTR modulator (CFTRm) ivacaftor (IVA), and this level was inversely correlated with the FEV1% predicted values (ppFEV1). Although the effectiveness of lumacaftor (LUM)/IVA in pwCF homozygous for the p.Phe508del-CFTR variant has been evaluated, plasma biomarkers were not used to monitor treatment efficacy thus far. METHODS: Plasma HE4 concentration was examined in 68 pwCF drawn from the PROSPECT study who were homozygous for the p.Phe508del-CFTR variant before treatment and at 1, 3, 6 and 12 months after administration of LUM/IVA therapy. Plasma HE4 was correlated with ppFEV1 using their absolute and delta values. The discriminatory power of delta HE4 was evaluated for the detection of lung function improvements based on ROC-AUC analysis and multiple regression test. RESULTS: HE4 plasma concentration was significantly reduced below baseline following LUM/IVA administration during the entire study period. The mean change of ppFEV1 was 2.6% (95% CI, 0.6 to 4.5) by 6 months of therapy in this sub-cohort. A significant inverse correlation between delta values of HE4 and ppFEV1 was observed especially in children with CF (r=-0.7053; p<0.0001). Delta HE4 predicted a 2.6% mean change in ppFEV1 (AUC: 0.7898 [95% CI 0.6823-0.8972]; P < 0.0001) at a cut-off value of -10.7 pmol/L. Moreover, delta HE4 independently represented the likelihood of being a responder with ≥ 5% delta ppFEV1 at 6 months (OR: 0.89, 95% CI: 0.82-0.95; P = 0.001). CONCLUSIONS: Plasma HE4 level negatively correlates with lung function improvement assessed by ppFEV1 in pwCF undergoing LUM/IVA CFTRm treatment.

Evaluation of the Efficacy of BBIBP-CorV Inactivated Vaccine Combined with BNT62b2 mRNA Booster Vaccine.

OBJECTIVES: In this prospective study, SARS-CoV-2 spike protein specific total immunoglobulin (Ig) levels were analyzed before and after BNT162 b2 mRNA booster vaccination in individuals previously administered with two doses of BBIBP-CorV vaccine in comparison to immunized participants with three doses of BNT162 b2 vaccination. METHODS: Sixty-one Caucasian volunteers (39 females, 22 males) vaccinated by BBIBP-CorV were included (mean age: 63.9 years). Sixty-one patients (41 females, 20 males) as controls were vaccinated with BNT162b2 (mean age: 59.9 years). Both groups received the third booster BNT162b2 vaccine. Total anti-SARS-CoV-2 S1-RBD Ig levels were measured by an immunoassay (Roche Diagnostics) and their calculated ratios after/before booster dose were compared between the two groups. RESULTS: At baseline, significantly lower anti-SARS-CoV-2 S1-RBD total antibody levels were determined after initial immunization by two doses of inactivated BBIBP-CorV compared to BNT62b2 mRNA vaccine (p < 0.001). After BNT162b2 boosters, similarly high total Ig levels were detected in both the heterologous (27,195 [15,604-42,754] BAU/mL, p < 0.001) and the homologous booster cohort (24,492 [13,779-42,671] BAU/mL, p < 0.001) compared to baseline. Hence, the ratio of after/before total Ig levels was significantly higher with heterologous vs homologous immunization (p < 0.001). CONCLUSION: To address the concept that basic BBIBP-CorV vaccination is not as effective as BNT162b, we analyzed the effect of heterologous vaccination with BNT162b2. Our results suggest that BNT162b2 can successfully boost the effects of two-dose BBIBP-CorV vaccination.

A mobile setup for simultaneous and in situ neutron reflectivity, infrared spectroscopy, and ellipsometry studies.

Neutron reflectivity at the solid/liquid interface offers unique opportunities for resolving the structure-function relationships of interfacial layers in soft matter science. It is a non-destructive technique for detailed analysis of layered structures on molecular length scales, providing thickness, density, roughness, and composition of individual layers or components of adsorbed films. However, there are also some well-known limitations of this method, such as the lack of chemical information, the difficulties in determining large layer thicknesses, and the limited time resolution. We have addressed these shortcomings by designing and implementing a portable sample environment for in situ characterization at neutron reflectometry beamlines, integrating infrared spectroscopy under attenuated total reflection for determination of molecular entities and their conformation, and spectroscopic ellipsometry for rapid and independent measurement of layer thicknesses and refractive indices. The utility of this combined setup is demonstrated by two projects investigating (a) pH-dependent swelling of polyelectrolyte layers and (b) the impact of nanoparticles on lipid membranes to identify potential mechanisms of nanotoxicity.

Investigation of Circulating MicroRNA Levels in Antibody-Mediated Rejection After Kidney Transplantation.

BACKGROUND: One of the most important possible complications determining long-term graft survival after kidney transplant is antibody-mediated rejection (ABMR). The criterion standard approach to recognize ABMR is currently the kidney biopsy with histopathologic analysis. However, this test has limitations because of difficulties in timing of sampling, the evaluability of histology because of the questionable representativeness of specimens, and the limited number of this intervention. Hence, new reliable, noninvasive biomarkers are required to detect the development of ABMR in time. METHODS: In this study, we analyzed the clinical data of 45 kidney transplant patients (mean age of 44.51 years, 20 male and 25 female subjects). These participants were recruited into 5 subcohorts based on their clinical status, histologic findings, and level of donor-specific anti-HLA antibodies. Circulating microRNAs (miR-21, miR-181b, miR-146a, miR-223, miR-155, miR-150) in plasma samples were quantified by quantitative polymerase chain reaction and their levels were correlated with the clinical characteristics in different subgroups. RESULTS: The relative expression of plasma miR-155 (P = .0003), miR-223 (P = .0316), and miR-21 (P = .0147) were significantly higher in patients who had subsequent histology-approved ABMR with donor-specific anti-HLA antibody positivity (n = 10) than in the "triple negative" group (n = 21), and miR-155 showed the highest sensitivity (90%) and specificity (81%) to indicate ABMR development based on receiver operating characteristic analysis. CONCLUSIONS: According to our preliminary data, plasma miR-155, miR-21, and miR-223 can indicate the development of ABMR after kidney transplant in correlation with classic clinical parameters. However, future studies with larger number of participants are necessary to further evaluate the diagnostic properties of blood miRNAs in prediction of this life-threatening condition.

Structure of Self-Initiated Photopolymerized Films: A Comparison of Models.

Self-initiated photografting and photopolymerization (SI-PGP) uses UV illumination to graft polymers to surfaces without additional photoinitiators using the monomers as initiators, "inimers". A wider use of this method is obstructed by a lack of understanding of the resulting, presumably heterogeneous, polymer structure and of the parallel degradation under continuous UV illumination. We have used neutron reflectometry to investigate the structure of hydrated SI-PGP-prepared poly(HEMA-co-PEG10MA) (poly(2-hydroxyethyl methacrylate-co-(ethylene glycol)10 methacrylate)) films and compared parabolic, sigmoidal, and Gaussian models for the polymer volume fraction distributions. Results from fitting these models to the data suggest that either model can be used to approximate the volume fraction profile to similar accuracy. In addition, a second layer of deuterated poly(methacrylic acid) (poly(dMAA)) was grafted over the existing poly(HEMA-co-PEG10MA) layer, and the resulting double-grafted films were also studied by neutron reflectometry to shed light on the UV-polymerization process and the inevitable UV-induced degradation which competes with the grafting.

The majority of severe COVID-19 patients develop anti-cardiac autoantibodies.

Severe cases of COVID-19 are characterized by an inflammatory burst, which is accompanied by multiorgan failure. The elderly population has higher risk for severe or fatal outcome for COVID-19. Inflammatory mediators facilitate the immune system to combat viral infection by producing antibodies against viral antigens. Several studies reported that the pro-inflammatory state and tissue damage in COVID-19 also promotes autoimmunity by autoantibody generation. We hypothesized that a subset of these autoantibodies targets cardiac antigens. Here we aimed to detect anti-cardiac autoantibodies in severe COVID-19 patients during hospitalization. For this purpose, 104 COVID-19 patients were recruited, while 40 heart failure patients with dilated cardiomyopathy and 20 patients with severe aortic stenosis served as controls. Patients were tested for anti-cardiac autoantibodies, using human heart homogenate as a bait. Follow-up samples were available in 29 COVID-19 patients. Anti-cardiac autoantibodies were detected in 68% (71 out of 104) of severe COVID-19 patients. Overall, 39% of COVID-19 patients had anti-cardiac IgG autoantibodies, while 51% had anti-cardiac autoantibodies of IgM isotype. Both IgG and IgM anti-cardiac autoantibodies were observed in 22% of cases, and multiple cardiac antigens were targeted in 38% of COVID-19 patients. These anti-cardiac autoantibodies targeted a diverse set of myocardial proteins, without apparent selectivity. As controls, heart failure patients (with dilated cardiomyopathy) had similar occurrence of IgG (45%, p = 0.57) autoantibodies, while significantly lower occurrence of IgM autoantibodies (30%, p = 0.03). Patients with advanced aortic stenosis had significantly lower number of both IgG (11%, p = 0.03) and IgM (10%, p < 0.01) type anti-cardiac autoantibodies than that in COVID-19 patients. Furthermore, we detected changes in the anti-cardiac autoantibody profile in 7 COVID-19 patients during hospital treatment. Surprisingly, the presence of these anti-cardiac autoantibodies did not affect the clinical outcome and the prevalence of the autoantibodies did not differ between the elderly (over 65 years) and the patients younger than 65 years of age. Our results demonstrate that the majority of hospitalized COVID-19 patients produce novel anti-cardiac IgM autoantibodies. COVID-19 also reactivates resident IgG autoantibodies. These autoantibodies may promote autoimmune reactions, which can complicate post-COVID recuperation, contributing to post-acute sequelae of COVID-19 (long COVID).