A single-center, retrospective analysis to compare measurement of fibrinogen using the TEG6 analyzer to the Clauss measurement in children undergoing heart surgery.

BACKGROUND: Newer generation viscoelastic tests, TEG6s, offer point-of-care hemostatic therapy in adult patients. However, their efficacy in estimating fibrinogen levels in pediatric patients undergoing cardiac surgery is not well established. AIMS: This study evaluates TEG6s for estimating fibrinogen levels in pediatric cardiac surgery patients and its predictive capability for post-bypass hypofibrinogenemia. METHODS: A single-center, retrospective study on pediatric patients (under 18 years) who underwent cardiac surgery with cardiopulmonary bypass from August 2020 and November 2022. Blood samples for estimated whole blood functional fibrinogen level via TEG6s (Haemonetics Inc.) and concurrent laboratory-measured plasma fibrinogen via von Clauss assay were collected at pre- and post-cardiopulmonary bypass. RESULTS: Paired data for TEG6s estimated functional fibrinogen levels and plasma fibrinogen were analyzed for 432 pediatric patients pre-bypass. It was observed that functional fibrinogen consistently overestimated plasma fibrinogen across all age groups with a mean difference of 138 mg/dL (95% confidence interval [CI]: 128-149 mg/dL). This positive bias in the pre-bypass data was confirmed by Bland-Altman analysis. Post-bypass, functional fibrinogen estimates were comparable to plasma fibrinogen in all patient groups with a mean difference of -6 mg/dL (95% CI: -20-8 mg/dL) except for neonates, where functional fibrinogen levels underestimated plasma fibrinogen with a mean difference of -38 mg/dL (95% CI: -64 to -12 mg/dL). The predictive accuracy of functional fibrinogen for detecting post-bypass hypofibrinogenemia (plasma fibrinogen ≤250 mg/dL) demonstrated overall fair accuracy in all patients, indicated by an area under the curve of 0.73 (95% CI: 0.65-0.80) and good accuracy among infants, with an area under the curve of 0.80 (95% CI: 0.70-0.90). Similar performance was observed in predicting critical post-bypass hypofibrinogenemia (plasma fibrinogen ≤200 mg/dL). Based on these analyses, optimal cutoffs for predicting post-bypass hypofibrinogenemia were established as a functional fibrinogen level ≤270 mg/dL and MAFF ≤15 mm. CONCLUSION: This study demonstrates that whole blood functional fibrinogen, as estimated by TEG6s, tends to overestimate baseline plasma fibrinogen levels in pediatric age groups but aligns more accurately post-cardiopulmonary bypass, particularly in neonates and infants, suggesting its potential as a point-of-care tool in pediatric cardiac surgery. However, the variability in TEG6s performance before and after bypass highlights the need for careful interpretation of its results in clinical decision-making. Despite its contributions to understanding TEG6s in pediatric cardiac surgery, the study's design and inherent biases warrant cautious application of these findings in clinical settings.

Modified technique for endobronchial blocker placement in pediatric patients undergoing thoracic surgery.

BACKGROUND: One lung ventilation (OLV) in small children can be achieved using an Arndt endobronchial blocker (AEBB), but it presents challenges. OLV during thoracic procedures provides better surgical conditions and postoperative outcomes. AIM: To report a novel technique to improve placement and repositioning of an extraluminal AEBB for OLV. MATERIAL AND METHODS: We describe how an angled wire is successfully used for extraluminal AEBB placement in pediatric thoracic procedures. DISCUSSION: Since 2017, we have successfully used this technic in over 50 infants and toddlers and overcome challenges of the classic OLV in this age group. CONCLUSIONS: The described technique allows for fast, safe, and reliable OLV while maintaining the ability to reposition the AEBB.

Intraarterial papaverine for relief of catheter-induced peripheral arterial vasospasm during pediatric cardiac surgery: A randomized double-blind controlled trial.

BACKGROUND: Maintaining the patency of peripheral arterial lines in pediatric patients during surgery can be challenging due to multiple factors, and catheter-related arterial vasospasm is a potentially modifiable cause. Papaverine, a potent vasodilator, improves arterial line patency when used as a continuous infusion in the pediatric intensive care setting, but this method is not convenient during surgery. AIM: Extrapolating from the benefit seen in the intensive care unit, the authors hypothesize that a small-volume intraarterial bolus of papaverine immediately after arterial line placement will reduce vasospasm-related arterial line malfunction. METHODS: This was a prospective, randomized, double-blind study. Patients less than 17 years of age undergoing cardiac surgery were enrolled. Patients were randomized into the heparin or papaverine groups. Immediately after arterial line insertion, an intraarterial bolus of heparin (2 units/ml, 1 ml) or papaverine (0.12 mg/ml, 1 ml) was administered (T1, Figure 1). An optimal waveform was defined as the ease of aspirating a standardized blood sample within 30 s, absence of cavitation when sampling, absence of color change at the catheter site during injection, and presence of a dicrotic notch. The primary outcome evaluated was the presence of an optimal arterial waveform at 5 min after the first randomized dose (T1 + 5 min). The secondary outcomes were the presence of optimal arterial waveform an hour after the first dose and the ability of papaverine to rescue suboptimal waveforms. RESULTS: A total of 100 patients were enrolled in the study. Twelve patients were excluded from the analysis. Complete datasets after randomization were available in 88 patients (heparin group, n = 46; papaverine group, n = 42). At baseline, groups were similar for age, weight, arterial vessel size, and arterial line patency. At T1 + 5 min, an improvement in the waveform characteristics was observed in the papaverine group (heparin,39% [8/46] vs. papaverine, 64% [27/42]; p = .02; odds ratio, 2.8; 95% CI, 1.2 to 6.6, Figure 3, Table 2). At the end of 1 h, both groups showed continued improvement in arterial line patency. After the second dose, a higher number of patients in the heparin group had suboptimal waveforms and were treated with papaverine (heparin,37% [17/46] vs. papaverine,17% [7/42], p = .05). Patients in the heparin group treated with papaverine showed significant improvement in patency (13/17 vs. 3/7, p = .01). No serious adverse events were reported. CONCLUSIONS: In pediatric patients, papaverine injection immediately after peripheral arterial catheter placement was associated with relief of vasospasm and improved initial arterial line patency. Further, papaverine can be used as a rescue to improve and maintain arterial line patency.

Anesthetic management of catheter-based patent ductus arteriosus closure in neonates weighing <3 kg: A Retrospective Observational Study.

BACKGROUND: The patent ductus arteriosus is a cardiac lesion commonly found in premature neonates. Though surgical closure via thoracotomy is the most definitive treatment option, it is associated with significant morbidity. New catheter-based closure options offer a potentially safer alternative treatment, even in premature neonates. However, no literature reports the anesthetic techniques, challenges, and risks associated with this procedure in this population. AIM: This study documents the anesthetic challenges and potential complications associated with the management of catheter-based closure of the ductus arteriosus in neonates under 3 kg. METHODS: This single-center, retrospective study examined patients who underwent catheter-based ductus arteriosus closure between August 2015 and February 2019. A clinical protocol for anesthetic management of these patients was utilized throughout the study period. Clinical outcomes considered were new hemodynamic instability or vasoactive medication requirements, hypothermia, prolonged intubation (>3 days postoperatively), postprocedure acute kidney injury, perioperative red blood cell transfusion, and accidental extubation. RESULTS: Seventy-six neonates underwent 78 procedures. No patient developed perioperative hemodynamic instability, vasoactive medication requirements, or acute kidney injury. Four patients (5%) required red blood cell transfusion, two (3%) became hypothermic, and one (1%) was accidentally extubated. Closure was achieved in 73 patients (96%) on the first attempt. However, 17 patients (40%) required prolonged periods of mechanical ventilation following the procedure. CONCLUSION: Despite multiple clinical and logistical challenges, anesthetic risk associated with catheter-based PDA closure in small neonates can be effectively managed through standardized and multidisciplinary care.

Transfusing Platelets During Bypass Rewarming in Neonates Improves Postoperative Outcomes: A Randomized Controlled Trial.

BACKGROUND: In neonates, transfusion of platelets after hemodilution from cardiopulmonary bypass (CPB) has been standard. We hypothesize that platelet administration during the rewarming phase before termination of CPB would reduce coagulopathy, enhance hemostasis, reduce transfusion, and improve postoperative outcomes after neonatal cardiac surgery. METHODS: A prospective, randomized trial was performed in 46 neonates. Controls received platelets only at the end of bypass with other blood products to assist in hemostasis. The treatment group received 10 mL/kg of platelets during the rewarming phase of bypass after cross-clamp release. After protamine, transfusion and perioperative management protocols were identical and constant among groups. RESULTS: Two neonates in each group were excluded secondary to postoperative need for extracorporeal support. Controls (n = 21) and treatment patients (n = 21) were similar in age, weight, case complexity, associated syndromes, single ventricle status, and CPB times. Compared to controls, the treatment group required 40% less postbypass blood products (58 ± 29 vs 103 ± 80 mL/kg, P = .04), and case completion time after protamine administration was 28 minutes faster (P = .016). The treatment group required fewer postoperative mediastinal explorations for bleeding (P = .045) and had a lower fluid balance (P = .04). The treatment group had shorter mechanical ventilation (P = .016) and length of intensive care unit times (P = .033). There were no 30-day mortalities in either group. CONCLUSION: Platelet transfusion during the rewarming phase of neonatal cardiac surgery was associated with reduced bleeding and improved postoperative outcomes, compared to platelets given after coming off bypass. Further studies are necessary to understand mechanisms and benefits of this strategy.

Deep Tracheal Extubation Using Dexmedetomidine in Children With Congenital Heart Disease Undergoing Cardiac Catheterization: Advantages and Complications.

Objective. Deep tracheal extubation using dexmedetomidine is safe and provides smooth recovery in children with congenital heart disease undergoing cardiac catheterization. Design. Single-institution, retrospective study of prospectively collected data. Participants. All patients aged between 1 month and 5 years who underwent general endotracheal anesthesia for diagnostic and interventional cardiac catheterizations in the cardiac catheterization suite from January 2015 (change in standard operating procedure) through October 2016 (approval of institutional review board for study). Measurement and Main Results. One hundred and eighty-nine patients (81%) of the 232 patients who underwent cardiac catheterization during the study period were noted to undergo deep tracheal extubation. Cyanotic heart disease was present in 87 patients (46%), history of prematurity in 51 (27%), and pulmonary hypertension in 26 (14%) patients. A documented smooth recovery in the postoperative care unit (PACU) requiring no additional analgesics or sedatives was observed in 91% of the patients. The majority of patients required no airway support after deep extubation (n = 140, 74%, P = .136). The presence of pulmonary hypertension (odds ratio = 4.45, P = .035) and presence of a cough on the day of the procedure (odds ratio = 7.10, P = .03) were significantly associated with the use of oxygen or use of oral airway for greater than 20 minutes in the PACU. After extubation, there were no reported events of aspiration, the use of noninvasive positive pressure ventilation, reintubation, heart block, or systemic hypotension requiring treatment or cardiac arrest. Conclusions. Deep extubation using dexmedetomidine in infants and toddlers after cardiac catheterization is feasible and enables smooth postoperative recovery with minimal adverse effects.

Triglyceride alters lysosomal cholesterol ester metabolism in cholesteryl ester-laden macrophage foam cells.

In late-stage atherosclerosis, much of the cholesterol in macrophage foam cells resides within enlarged lysosomes. Similarly, human macrophages incubated in vitro with modified LDLs contain significant amounts of lysosomal free cholesterol and cholesteryl ester (CE), which disrupts lysosomal function similar to macrophages in atherosclerotic lesions. The lysosomal cholesterol cannot be removed, even in the presence of strong efflux promoters. Thus, efflux of sterol is prevented. In the artery wall, foam cells interact with triglyceride-rich particles (TRPs) in addition to modified LDLs. Little is known about how TRP metabolism affects macrophage cholesterol. Therefore, we explored the effect of TRP on intracellular CE metabolism. Triglyceride (TG), delivered to lysosomes in TRP, reduced CE accumulation by 50%. Increased TG levels within the cell, particularly within lysosomes, correlated with reductions in CE content. The volume of cholesterol-engorged lysosomes decreased after TRP treatment, indicating cholesterol was cleared. Lysosomal TG also reduced the cholesterol-induced inhibition of lysosomal acidification allowing lysosomes to remain active. Enhanced degradation and clearance of CE may be explained by movement of cholesterol out of the lysosome to sites where it is effluxed. Thus, our results show that introduction of TG into CE-laden foam cells influences CE metabolism and, potentially, atherogenesis.-Ullery-Ricewick, J. C., B. E. Cox, E. E. Griffin, and W. G. Jerome. Triglyceride alters lysosomal cholesterol ester metabolism in cholesteryl ester-laden macrophage foam cells.

Benzimidazole- and benzoxazole-based inhibitors of Rho kinase.

Inhibitors of Rho kinase have been developed based on two distinct scaffolds, benzimidazoles, and benzoxazoles. SAR studies and efforts to optimize the initial lead compounds are described. Novel selective inhibitors of ROCK-II with excellent potency in both enzyme and cell-based assays were obtained. These inhibitors possess good microsomal stability, low cytochrome P-450 inhibitions and good oral bioavailability.

Chroman-3-amides as potent Rho kinase inhibitors.

Inhibition of Rho kinase (ROCK) is an attractive strategy for the treatment of diseases such as hypertension, glaucoma, and cancer. Here we report chroman-3-amides as highly potent ROCK inhibitors with sufficient kinase selectivity, excellent cell activity, good microsomal stability, and desirable pharmacokinetic properties for study as potential therapeutic agents.

Discovery of substituted 4-(pyrazol-4-yl)-phenylbenzodioxane-2-carboxamides as potent and highly selective Rho kinase (ROCK-II) inhibitors.

The identification of a new class of potent and selective ROCK-II inhibitors is presented. Compound 5 (SR-3677) had an IC 50 of approximately 3 nM in enzyme and cell based assays and had an off-target hit rate of 1.4% against 353 kinases, and inhibited only 3 out of 70 nonkinase enzymes and receptors. Pharmacology studies showed that 5 was efficacious in both, increasing ex vivo aqueous humor outflow in porcine eyes and inhibiting myosin light chain phosphorylation.

Detection of myosin light chain phosphorylation--a cell-based assay for screening Rho-kinase inhibitors.

Here, we describe the first example of a cell-based myosin light chain phosphorylation assay in 96-well format that allows for the rapid screening of novel Rho-kinase inhibitors. We obtained IC(50) values for the prototypic Rho-kinase inhibitors Y-27632 (1.2+/-0.05microM) and Fasudil (3.7+/-1.2microM) that were similar to those previously published utilizing electrophoresis-based methodologies. H-1152P, a Fasudil analog showed an IC(50) value of 77+/-30nM. Data derived from a set of 21 novel Rho-kinase inhibitors correlate with those generated by a well-established cell-based phenotypic Rho-kinase inhibition assay (R(2)=0.744). These results show that imaging technology measuring changes in myosin light chain phosphorylation can be used to rapidly generate quantitative IC(50) values and to screen a larger set of small molecule Rho-kinase inhibitors and suggests that this approach can be broadly applied to other cell lines and signaling pathways.

Lysosomal cholesterol accumulation inhibits subsequent hydrolysis of lipoprotein cholesteryl ester.

Human macrophages incubated for prolonged periods with mildly oxidized LDL (oxLDL) or cholesteryl ester-rich lipid dispersions (DISP) accumulate free and esterified cholesterol within large, swollen lysosomes similar to those in foam cells of atherosclerosis. The cholesteryl ester (CE) accumulation is, in part, the result of inhibition of lysosomal hydrolysis due to increased lysosomal pH mediated by excessive lysosomal free cholesterol (FC). To determine if the inhibition of hydrolysis was long lived and further define the extent of the lysosomal defect, we incubated THP-1 macrophages with oxLDL or DISP to produce lysosome sterol engorgement and then chased with acetylated LDL (acLDL). Unlike oxLDL or DISP, CE from acLDL normally is hydrolyzed rapidly. Three days of incubation with oxLDL or DISP produced an excess of CE in lipid-engorged lysosomes, indicative of inhibition. After prolonged oxLDL or DISP pretreatment, subsequent hydrolysis of acLDL CE was inhibited. Coincident with the inhibition, the lipid-engorged lysosomes failed to maintain an acidic pH during both the initial pretreatment and subsequent acLDL incubation. This indicates that the alterations in lysosomes were general, long lived, and affected subsequent lipoprotein metabolism. This same phenomenon, occurring within atherosclerotic foam cells, could significantly affect lesion progression.

Structure-activity relationships, and drug metabolism and pharmacokinetic properties for indazole piperazine and indazole piperidine inhibitors of ROCK-II.

ROCK has been implicated in many diseases ranging from glaucoma to spinal cord injury and is therefore an important target for therapeutic intervention. In this study, we have designed a series of 1-(4-(1H-indazol-5-yl)piperazin-1-yl)-2-hydroxy(or 2-amino) analogs and a series of 1-(4-(1H-indazol-5-yl amino)piperidin-1-yl)-2-hydroxy(or 2-amino) inhibitors of ROCK-II. SR-1459 has IC(50)=13nM versus ROCK-II while the IC(50)s for SR-715 and SR-899 are 80nM and 100nM, respectively. Many of these inhibitors, especially the 2-amino substituted analogs for both series, are modest/potent CYP3A4 inhibitors as well. However, a few of these inhibitors (SR-715 and SR-899) show strong selectivity for ROCK-II over CYP3A4, but the overall potency of the 2-amino analogs (SR-1459) on CYP3A4 and the high clearance and volume of distribution of these compounds makes the in vivo utility of these analogs undesirable.

Effects of cellular cholesterol loading on macrophage foam cell lysosome acidification.

Macrophages incubated with mildly oxidized low density lipoprotein (OxLDL), aggregated low density lipoprotein (AggLDL), or cholesteryl ester-rich lipid dispersions (DISPs) accumulate cholesterol in lysosomes followed by an inhibition of lysosomal cholesteryl ester (CE) hydrolysis. The variety of cholesterol-containing particles producing inhibition of hydrolysis suggests that inhibition may relate to general changes in lysosomes. Lysosome pH is a key mediator of activity and thus is a potential mechanism for lipid-induced inhibition. We investigated the effects of cholesterol accumulation on THP-1 macrophage lysosome pH. Treatment with OxLDL, AggLDL, and DISPs resulted in inhibition of the lysosome's ability to maintain an active pH and concomitant decreases in CE hydrolysis. Consistent with an overall disruption of lysosome function, exposure to OxLDL or AggLDL reduced lysosomal apolipoprotein B degradation. The lysosomal cholesterol sequestration and inactivation are not observed in cholesterol-equivalent cells loaded using acetylated low density lipoprotein (AcLDL). However, AcLDL-derived cholesterol in the presence of progesterone (to block cholesterol egression from lysosomes) inhibited lysosome acidification. Lysosome inhibition was not attributable to a decrease in the overall levels of vacuolar ATPase. However, augmentation of membrane cholesterol in isolated lysosomes inhibited vacuolar ATPase-dependent pumping of H+ ions into lysosomes. These data indicate that lysosomal cholesterol accumulation alters lysosomes in ways that could exacerbate foam cell formation and influence atherosclerotic lesion development.

Severely altered cholesterol homeostasis in macrophages lacking apoE and SR-BI.

Mice deficient in scavenger receptor class B type I (SR-BI) and apolipoprotein E (apoE) [double knockout (DKO) mice] develop dyslipidemia, accelerated atherosclerosis, and myocardial infarction, and die prematurely. We examined effects of apoE and SR-BI deficiency on macrophage cholesterol homeostasis. DKO macrophages had increased total cholesterol (TC) stores (220-380 microg/mg protein) compared with apoE-/- cells (40 microg/mg), showed significant lysosomal lipid engorgement, and increased their TC by 34% after exposure to HDL. DKO macrophages from apoE-/- mice reconstituted with DKO bone marrow showed less cholesterol accumulation (89 microg/mg), suggesting that the dyslipidemia of DKO mice explains part of the cellular cholesterol defect. However, analyses of DKO and apoE-/- macrophages from transplanted apoE-/- mice revealed a role for macrophage SR-BI, inasmuch as the TC in DKO macrophages increased by 10% in the presence of HDL, whereas apoE-/- macrophage TC decreased by 33%. After incubation with HDL, the free cholesterol (FC) increased by 29% in DKO macrophages, and decreased by 8% in apoE-/- cells, and only DKO cells had FC in large peri-nuclear pools. Similar trends were observed with apoA-I as an acceptor. Thus, the abnormal cholesterol homeostasis of DKO macrophages is due to the plasma lipid environment of DKO mice and to altered trafficking of macrophage cholesterol. Both factors are likely to contribute to the accelerated atherosclerosis in DKO mice.

Aggregated LDL and lipid dispersions induce lysosomal cholesteryl ester accumulation in macrophage foam cells.

Macrophage foam cells in atherosclerotic lesions accumulate substantial cholesterol stores within large, swollen lysosomes. Previous studies with mildly oxidized low density lipoprotein (OxLDL)-treated THP-1 macrophages suggest an initial buildup of free cholesterol (FC), followed by an inhibition of lysosomal cholesteryl ester (CE) hydrolysis and a subsequent lysosomal accumulation of unhydrolyzed lipoprotein CE. We examined whether other potential sources of cholesterol found within atherosclerotic lesions could also induce similar lysosomal accumulation. Biochemical analysis combined with microscopic analysis showed that treatment of THP-1 macrophages with aggregated low density lipoprotein (AggLDL) or CE-rich lipid dispersions (DISP) produced a similar lysosomal accumulation of both FC and CE. Co-treatment with an ACAT inhibitor, CP113,818, confirmed that the CE accumulation was primarily the result of the inhibition of lysosomal CE hydrolysis. The rate of unhydrolyzed CE buildup was more rapid with DISP than with AggLDL. However, with both treatments, FC appeared to accumulate in lysosomes before the inhibition in hydrolysis and CE accumulation, a sequence shared with mildly OxLDL. Thus, lysosomal accumulation of FC and CE can be attributable to more general mechanisms than just the inhibition of hydrolysis by oxidized lipids.