Embolization of Perforated Coronary Artery with a Fragment of Balloon Catheter (Cut Balloon Technique)-Multicenter Study.

BACKGROUND: Iatrogenic distal coronary artery perforation can be a life-threatening complication. While there are different dedicated devices for the embolization of distal perforations, there are scarce data about the embolization using the fragmented balloon catheter, the so-called cut balloon technique (CBT). METHODS: We included consecutive patients with distal coronary perforations treated with CBT in four cardiac centers between 2017 and 2023. Clinical, angiographic and procedural characteristics as well as in-hospital outcomes were recorded. RESULTS: Twenty-six patients (68% men, mean age: 71 ± 10.6 years) with 25 distal coronary perforations and one septal collateral perforation were included. Eleven patients (42%) had elective percutaneous coronary intervention, while fifteen patients (58%) were treated for acute coronary syndrome. The site of perforation was most frequently distributed in the left anterior descending artery (40%), followed by the circumflex artery (28%) and right coronary artery (24%). The diameter of balloons for CBT ranged from 1.5 to 4.0 mm, with most balloons (76%) being either 2.0 or 2.5 mm in diameter. Most balloons (88%) were previously used for lesion predilatation. The numbers of cut balloons needed to seal the perforation were 1, 2 and ≥3 in 48%, 20% and 32% of cases, respectively. The in-hospital prognosis was favorable, with cardiac tamponade requiring pericardiocentesis in only four (16%) patients. Neither emergency surgery nor cardiac death occurred. CONCLUSIONS: CBT is a safe, efficient and easy-to-implement technique for the embolization of coronary perforations. Most distal coronary perforations can be sealed with one or two fragments of cut balloons, obviating the need for additional devices.

Long-term effects of hydrocyclone operation on activated sludge morphology and full-scale secondary settling tank wet-weather operation in long sludge age WWTP.

This paper presents the study concerning long-term effects of a full scale hydrocyclone unit implemented in a continuous flow long sludge age system, on sedimentation, treatment efficiency and sludge morphology. The research concentrates on identifying the mechanisms of sludge behaviour within the system. The gravimetric selection of activated sludge via a hydrocyclone is a recent development for enhancing sludge separation, where heavier flocs are retained in the system, and lighter ones are discarded as waste sludge. The effects of implementing hydroclyclones were analysed with the use of SEM imagining and fractal dimensioning through the frequent assessment of sludge settling capabilities, effluent quality, and floc properties. Over the course of 60 weeks of hydrocyclone operation, sedimentation efficiency varied significantly. Sludge volume index values of 40 mL/g, achieved during the warm season, were not sustained when the temperature decreased and an overgrowth of filamentous bacteria occurred. Good settling efficiency was also observed in batch tests, where settling velocity of experimental sludge was app. 1 m/h higher than for the reference train at the same concentrations. This was confirmed during wet weather, as the experimental train sustained safe sludge blanket height in secondary clarifiers. SEM imaging and fractal dimension analysis revealed that the underflow that returned to the system had a more compact and spherical shape, which led to an increased content of granule-like particles in the reactor. The presence of flocs with a diameter exceeding 900 µm in the underflow, which is not observed in the feed, indicated agglomeration within the hydrocyclone. This is contradictory to most of the literature data coming from laboratory experiments. This phenomenon was attributed to differences in the size and geometry of the used hydrocyclones, and the potential process mechanism was presented.

Imaging of Clear Cell Renal Carcinoma with Immune Checkpoint Targeting Aptamer-Based Probe.

Immune checkpoint targeting immunotherapy has revolutionized the treatment of certain cancers in the recent years. Determination of the status of immune checkpoint expression in particular cancers may assist decision making. Here, we describe the development of a single-stranded aptamer-based molecular probe specifically recognizing human PD-L1. Target engaging aptamers are selected by iterative enrichment from a random ssDNA pool and the binding is characterized biochemically. Specificity and dose dependence is demonstrated in vitro in the cell culture using human kidney tumor cells (786-0), human melanoma cells (WM115 and WM266.4) and human glioblastoma LN18 cancer cells. The utility of the probe in vivo is demonstrated using two mouse tumor models, where we show that the probe exhibits excellent potential in imaging. We postulate that further development of the probe may allow universal imaging of different types of tumors depending on their PD-L1 status, which may find utility in cancer diagnosis.

Can nitrifiers from the sidestream deammonification process be a remedy for the N-overload of the mainstream reactor?

The combination of sidestream deammonification and bioaugmentation of the mainstream reactor using ammonia oxidizers from partial nitritation (PN) was not achieved before. This novel solution not only enables the efficient sidestream nitrogen removal, but also improves mainstream resistance to stress situations such as biomass washout or nitrogen overload. This feature is important for wastewater treatment plants (WWTPs) equipped with reject water deammonification as its implementation leads to lower nitrifier mass in the mainstream reactor and therefore diminish ability to cope with rapid increase in the loading rate (i.e. due to sidestream process failure). The proposed approach presents the use of the excess sludge from a modified PN process to boost the mainstream nitrification in unfavourable conditions. In a long-term laboratory experiment, the operation of an existing WWTP at low temperature was simulated in two reactors using real wastewater fluxes. One of them was augmented with the excess sludge from a PN reactor that treats reject water containing 20% of the WWTP N-load. The treatment efficiency in both reactors was tested under different nitrogen loading rates, as well as in the case of the of biomass loss. The bioaugmentation intensity was set according to the actual nitrogen load balance of the modelled WWTP, resulting in a daily seed volume only equal to 0.28% of the reactors' influent. Two incidents were simulated, where the nitrogen load increased by about 24.5% and 34%. In both cases, the nitrification efficiency in the non-augmented reactor dropped by about 45%, while the augmented reactor maintained efficient ammonium removal. The bioaugmentation effect was also noticeable during biomass washout - only in the non-augmented reactor nitrification was insufficient for over 60 days. These results undoubtedly show the possibility of combining two different approaches for sidestream nitrogen removal into one technology demonstrating the advantages of both component solutions.

Impact of the thermal drying of sludge on the nitrogen mass balance of a WWTP, and GHG emissions with classical and novel treatment approach - A full-scale case study.

This is the first study that identifies nitrogen loss and drier liquor parameters in a full scale sludge drying facility. Obtained data enabled proposing novel treatment based on deammonification process that allows not only nitrogen removal from drier liquor but also from gravity thickeners supernatant, the stream that is considered too cold for deammonification. The novel treatment approach is compared with nitrification/denitrification in activated sludge process in terms of greenhouse gas emissions. The nitrogen loss during drying was calculated based on two independent methods using full scale data from a long-term measurement campaign. According to results, 9.8-11.2% of nitrogen from dewatered sludge, or 4.9-5.5 g N/kg of TS, was vaporized during drying and then was captured through the purification of process air in drier liquor. Overall, over 40 tN/a from drier liquor and 77 tN/a from gravity thickeners supernatant is recirculated to the activated sludge process and results in the additional emission of 670 426 kg CO2e/a when nitrification/denitrification is applied. That can be lowered by 398 858 kgCO2e/a when the novel approach is considered.

Determination of the major factor responsible for soluble organic matter release during nitrite/free nitrous acid pre-treatment of waste activated sludge.

Soluble chemical oxygen demand (SCOD) release by free nitrous acid (FNA)/NO2 system is usually called "FNA disintegration", despite lack of evidence that FNA is the main agent responsible for organic matter breakdown. The aim of this study was to investigate whether FNA or NO2 is the primary disintegration factor of thickened secondary sludge in a wide spectrum of process parameters (T = 48 h, 0-2280 mg NO2-N/L, pH 3.2-6.4 and FNA between 0 and 47.4 mg HNO2-N/L). Statistical analysis based on multiple regression and the Akaike Information Criterion showed that NO2, not FNA, is a main disintegrating factor leading to SCOD release (p = 0.005206 and 0.00009 respectively) and that the FNA concentration is without statistical significance (p = 0.800234 and 0.328099 respectively). These findings are important as understanding key factors is essential for productive future research and technology development. Moreover, these findings give doubts about the role of FNA in its other applications such as inhibition of nitrification.

Urine nitrification robustness for application in space: Effect of high salinity and the response to extreme free ammonia concentrations.

Urine nitrification is one of the possibilities for the future recovery of water and elements for soilless crop production in space systems. The start-up of artificial urine nitrification was conducted for over 85 days in a sequencing batch reactor (SBR). Two free ammonia (FA) incidents occurred, which gave the opportunity to demonstrate the impressive ability of nitrifiers to resist temporary inhibition by FA without long lasting adverse effects. The failures led to very high FA concentrations of 280 and 84 gN-NH3/m3, respectively. Sludge was exposed to FA for 19 and 27 h. It was possible to restore nitrification with simple remedy actions (dilution and pH restoration). No inhibitory effects on the nitrification rate were seen after implementation of the remedy actions and the nitrification rate increased considerably (up to 300% of the pre-failure value) due to decrease in salinity. After a few days of normal operation and salt concentration, the nitrification rate returned to the pre-failure values in both cases.

Balloon Fragment Technique Used to Close Distal Coronary Vessel Perforation.

AIMS: Coronary artery perforation is a rare but potentially serious complication of percutaneous coronary intervention (PCI). Clinical manifestation and prognosis of patients depend on the severity of the complication, the occurrence of tamponade, and the methods of treatment. A special type of perforation is caused by damage to the distal segment of the coronary vessel by guidewire. This type of perforation is mostly connected with subacute clinical presentation. The treatment can be difficult, especially when prolonged balloon inflation is not efficient. The aim of the article is to present a balloon fragment technique as a treatment method for distal vessel perforation. METHODS: The method of treating distal perforation, which we presented at EuroPCR 2019, involves embolization of a perforated vessel with a cut part of balloon catheter. Six patients are included in this registry; only 1 truncated proximal balloon fragment was enough to effectively embolize the perforated vessel in 4 patients, while 2 balloons were required in 1 patient and 3 balloons were required in 1 patient. CONCLUSION: Compared with other methods, this technique is an easy, cheap, and effective treatment for distal vessel perforation. The described method is available in every catheterization laboratory and does not require expensive equipment or exceptional operator experience.

Myeloperoxidase level and inflammatory markers and lipid and lipoprotein parameters in stable coronary artery disease.

BACKGROUND: Myeloperoxidase (MPO) impairing endothelial functions. We investigated whether increasing concentration of myeloperoxidase (MPO) and inflammatory markers induce progression and incident acute coronary syndrome (ACS) in stable coronary artery disease (SCAD) patients. Therefore, the concentration of MPO, lipids, lipoproteins (apo(apolipoprotein) AI, apoB, lipoprotein associated phospholipase A2 (LpPLA2) level), inflammatory markers (high sensitivity C-reactive protein (hsCRP), tumor necrosis factor-α (TNF-α), interleukine-6 (IL-6) concentration) were examined. METHODS: This study concerned 67 SCAD patients divided into groups: all patients, patients with MPO < 200 ng/ml, MPO 200-300 ng/ml, MPO > 300 ng/ml concentration and 15 controls. ApoAI, apoB and hsCRP levels were examined using the immunonephelometric method, and MPO, LpPLA2, IL-6, TNF-α concentration was performed by using Quantikine ELISA kit R&D Systems. RESULTS: In the all patients, and in group with MPO 200-300 ng/ml TC, LDL-C, nonHDL-C, LpPLA2 concentration and TC/HDL-C, LDL-C/HDL-C ratios were insignificant, and significantly higher concentration of TG, apoB, MPO, inflammatory markers and TG/HDL-C, MPO/apoAI, MPO/HDL-C ratios but HDL-C, apoAI level and HDL-C/apoAI ratio were significantly reduced. In the group of patients with MPO < 200 ng/ml, level of TC, LDL-C, nonHDL-C, apoAI, apoAII, LpPLA2 and MPO and LDL-C/HDL-C ratio were in-significant, HDL-C was decreased but apoB, TG, inflammatory markers, apoB/apoAI, TG/HDL-C, MPO/apoAI, MPO/HDL-C ratio were significantly increased. In the group of patients with MPO > 300 ng/ml concentration of TC, LDL-C, nonHDL-C, apoAII, LpPLA2 and LDL-C/HDL-C ratios were not significant, but HDL-C and apoAI concentrations were significantly decreased. The concentrations of TG, apoB, MPO and inflammatory markers and TG/HDL-C, MPO/apoAI, MPO/HDL-C ratios were significantly increased compared to the controls. The apoAI concentration was significantly decreased and the concentration of MPO and hsCRP as well as MPO/apoAI and MPO/HDL-C ratios were significantly higher as compared to the group of patients with MPO < 200 ng/ml. Spearman's correlation test showed a positive correlation between MPO concentration and MPO/apoAI and MPO/HDL-C ratios in all patients and MPO < 200 ng/ml, MPO 200-300 ng/ml. The patients with MPO > 300 ng/ml showed a positive correlation between the concentration of MPO and the level of hsCRP and IL-6, and a negative correlation between MPO/apoAI ratio and the concentration of HDL-C, apoAI and apoAII. CONCLUSION: The results suggest that moderate dyslipidemia and dyslipoproteinemia deepening of inflammation, and inflammation slowly induce increase MPO concentration which decrease apoAI and HDL-C level and disturb HDLs function. The increasing MPO level and MPO/HDL-C, MPO/apoAI ratios can differentiate the SCAD patients at the risk of acute coronary syndrome (ACAD) and stroke.

Identification of small-molecule inhibitors of USP2a.

USP2a is a deubiquitinating protease that rescues its target proteins from destruction by the proteasome by reversing the process of protein ubiquitination. USP2a shows oncogenic properties in vivo and has been found to be a specific activator of cyclin D1. Many types of cancers are addicted to cyclin D1 expression. Targeting USP2a is a promising strategy for cancer therapy but little progress has been made in the field of inhibition of USP2a. Using NMR-based fragment screening and biophysical binding assays, we have discovered small molecules that bind to USP2a. Iterations of fragment combination and structure-driven design identified two 5-(2-thienyl)-3-isoxazoles as the inhibitors of the USP2a-ubiquitin protein-protein interaction. The affinity of these molecules for the catalytic domain of USP2a parallels their ability to interfere with USP2a binding to ubiquitin in vitro. Altogether, our results establish the 5-(2-thienyl)-3-isoxazole pharmacophore as an attractive starting point for lead optimization.

Lithocholic Acid Hydroxyamide Destabilizes Cyclin D1 and Induces G0/G1 Arrest by Inhibiting Deubiquitinase USP2a.

USP2a is a deubiquitinase responsible for stabilization of cyclin D1, a crucial regulator of cell-cycle progression and a proto-oncoprotein overexpressed in numerous cancer types. Here we report that lithocholic acid (LCA) derivatives are inhibitors of USP proteins, including USP2a. The most potent LCA derivative, LCA hydroxyamide (LCAHA), inhibits USP2a, leading to a significant Akt/GSK3ß-independent destabilization of cyclin D1, but does not change the expression of p27. This leads to the defects in cell-cycle progression. As a result, LCAHA inhibits the growth of cyclin D1-expressing, but not cyclin D1-negative cells, independently of the p53 status. We show that LCA derivatives may be considered as future therapeutics for the treatment of cyclin D1-addicted p53-expressing and p53-defective cancer types.

Structural basis for small molecule targeting of the programmed death ligand 1 (PD-L1).

Targeting the PD-1/PD-L1 immunologic checkpoint with monoclonal antibodies has provided unprecedented results in cancer treatment in the recent years. Development of chemical inhibitors for this pathway lags the antibody development because of insufficient structural information. The first nonpeptidic chemical inhibitors that target the PD-1/PD-L1 interaction have only been recently disclosed by Bristol-Myers Squibb. Here, we show that these small-molecule compounds bind directly to PD-L1 and that they potently block PD-1 binding. Structural studies reveal a dimeric protein complex with a single small molecule which stabilizes the dimer thus occluding the PD-1 interaction surface of PD-L1s. The small-molecule interaction "hot spots" on PD-L1 surfaces suggest approaches for the PD-1/PD-L1 antagonist drug discovery.

Rap1b in smooth muscle and endothelium is required for maintenance of vascular tone and normal blood pressure.

OBJECTIVE: Small GTPase Ras-related protein 1 (Rap1b) controls several basic cellular phenomena, and its deletion in mice leads to several cardiovascular defects, including impaired adhesion of blood cells and defective angiogenesis. We found that Rap1b(-/-) mice develop cardiac hypertrophy and hypertension. Therefore, we examined the function of Rap1b in regulation of blood pressure. APPROACH AND RESULTS: Rap1b(-/-) mice developed cardiac hypertrophy and elevated blood pressure, but maintained a normal heart rate. Correcting elevated blood pressure with losartan, an angiotensin II type 1 receptor antagonist, alleviated cardiac hypertrophy in Rap1b(-/-) mice, suggesting a possibility that cardiac hypertrophy develops secondary to hypertension. The indices of renal function and plasma renin activity were normal in Rap1b(-/-) mice. Ex vivo, we examined whether the effect of Rap1b deletion on smooth muscle-mediated vessel contraction and endothelium-dependent vessel dilation, 2 major mechanisms controlling basal vascular tone, was the basis for the hypertension. We found increased contractility on stimulation with a thromboxane analog or angiotensin II or phenylephrine along with increased inhibitory phosphorylation of myosin phosphatase under basal conditions consistent with elevated basal tone and the observed hypertension. Cyclic adenosine monophosphate-dependent relaxation in response to Rap1 activator, Epac, was decreased in vessels from Rap1b(-/-) mice. Defective endothelial release of dilatory nitric oxide in response to elevated blood flow leads to hypertension. We found that nitric oxide-dependent vasodilation was significantly inhibited in Rap1b-deficient vessels. CONCLUSIONS: This is the first report to indicate that Rap1b in both smooth muscle and endothelium plays a key role in maintaining blood pressure by controlling normal vascular tone.

Fluorescence linked enzyme chemoproteomic strategy for discovery of a potent and selective DAPK1 and ZIPK inhibitor.

DAPK1 and ZIPK (also called DAPK3) are closely related serine/threonine protein kinases that regulate programmed cell death and phosphorylation of non-muscle and smooth muscle myosin. We have developed a fluorescence linked enzyme chemoproteomic strategy (FLECS) for the rapid identification of inhibitors for any element of the purinome and identified a selective pyrazolo[3,4-d]pyrimidinone (HS38) that inhibits DAPK1 and ZIPK in an ATP-competitive manner at nanomolar concentrations. In cellular studies, HS38 decreased RLC20 phosphorylation. In ex vivo studies, HS38 decreased contractile force generated in mouse aorta, rabbit ileum, and calyculin A stimulated arterial muscle by decreasing RLC20 and MYPT1 phosphorylation. The inhibitor also promoted relaxation in Ca(2+)-sensitized vessels. A close structural analogue (HS43) with 5-fold lower affinity for ZIPK produced no effect on cells or tissues. These findings are consistent with a mechanism of action wherein HS38 specifically targets ZIPK in smooth muscle. The discovery of HS38 provides a lead scaffold for the development of therapeutic agents for smooth muscle related disorders and a chemical means to probe the function of DAPK1 and ZIPK across species.

The cAMP-responsive Rap1 guanine nucleotide exchange factor, Epac, induces smooth muscle relaxation by down-regulation of RhoA activity.

Agonist activation of the small GTPase, RhoA, and its effector Rho kinase leads to down-regulation of smooth muscle (SM) myosin light chain phosphatase activity, an increase in myosin light chain (RLC(20)) phosphorylation and force. Cyclic nucleotides can reverse this process. We report a new mechanism of cAMP-mediated relaxation through Epac, a GTP exchange factor for the small GTPase Rap1 resulting in an increase in Rap1 activity and suppression of RhoA activity. An Epac-selective cAMP analog, 8-pCPT-2'-O-Me-cAMP ("007"), significantly reduced agonist-induced contractile force, RLC(20), and myosin light chain phosphatase phosphorylation in both intact and permeabilized vascular, gut, and airway SMs independently of PKA and PKG. The vasodilator PGI(2) analog, cicaprost, increased Rap1 activity and decreased RhoA activity in intact SMs. Forskolin, phosphodiesterase inhibitor isobutylmethylxanthine, and isoproterenol also significantly increased Rap1-GTP in rat aortic SM cells. The PKA inhibitor H89 was without effect on the 007-induced increase in Rap1-GTP. Lysophosphatidic acid-induced RhoA activity was reduced by treatment with 007 in WT but not Rap1B null fibroblasts, consistent with Epac signaling through Rap1B to down-regulate RhoA activity. Isoproterenol-induced increase in Rap1 activity was inhibited by silencing Epac1 in rat aortic SM cells. Evidence is presented that cooperative cAMP activation of PKA and Epac contribute to relaxation of SM. Our findings demonstrate a cAMP-mediated signaling mechanism whereby activation of Epac results in a PKA-independent, Rap1-dependent Ca(2+) desensitization of force in SM through down-regulation of RhoA activity. Cyclic AMP inhibition of RhoA is mediated through activation of both Epac and PKA.

The actin associated protein palladin is important for the early smooth muscle cell differentiation.

Palladin, an actin associated protein, plays a significant role in regulating cell adhesion and cell motility. Palladin is important for development, as knockdown in mice is embryonic lethal, yet its role in the development of the vasculature is unknown. We have shown that palladin is essential for the expression of smooth muscle cells (SMC) marker genes and force development in response to agonist stimulation in palladin deficient SMCs. The goal of the study was to determine the molecular mechanisms underlying palladin's ability to regulate the expression of SMC marker genes. Results showed that palladin expression was rapidly induced in an A404 cell line upon retinoic acid (RA) induced differentiation. Suppression of palladin expression with siRNAs inhibited the expression of RA induced SMC differentiation genes, SM α-actin (SMA) and SM22, whereas over-expression of palladin induced SMC gene expression. Chromatin immunoprecipitation assays provided evidence that palladin bound to SMC genes, whereas co-immunoprecipitation assays also showed binding of palladin to myocardin related transcription factors (MRTFs). Endogenous palladin was imaged in the nucleus, increased with leptomycin treatment and the carboxyl-termini of palladin co-localized with MRTFs in the nucleus. Results support a model wherein palladin contributes to SMC differentiation through regulation of CArG-SRF-MRTF dependent transcription of SMC marker genes and as previously published, also through actin dynamics. Finally, in E11.5 palladin null mouse embryos, the expression of SMA and SM22 mRNA and protein is decreased in the vessel wall. Taken together, our findings suggest that palladin plays a key role in the differentiation of SMCs in the developing vasculature.