Hybrid Interventions in Congenital Heart Disease.

Hybrid interventions in congenital heart disease (CHD) embody the inherent collaboration between congenital interventional cardiology and cardiothoracic surgery. Hybrid approaches to complex and common lesions provide the opportunity to circumvent the limitations of patient size, vascular access, severity of illness, and anatomy that would otherwise be prohibitive to surgical and percutaneous techniques alone. This review describes several important hybrid approaches to interventions in CHD.

Proteomic snapshot of saliva samples predicts new pathways implicated in SARS-CoV-2 pathogenesis.

BACKGROUND: Information on the microbiome's human pathways and active members that can affect SARS-CoV-2 susceptibility and pathogenesis in the salivary proteome is very scarce. Here, we studied a unique collection of samples harvested from April to June 2020 from unvaccinated patients. METHODS: We compared 10 infected and hospitalized patients with severe (n = 5) and moderate (n = 5) coronavirus disease (COVID-19) with 10 uninfected individuals, including non-COVID-19 but susceptible individuals (n = 5) and non-COVID-19 and nonsusceptible healthcare workers with repeated high-risk exposures (n = 5). RESULTS: By performing high-throughput proteomic profiling in saliva samples, we detected 226 unique differentially expressed (DE) human proteins between groups (q-value ≤ 0.05) out of 3376 unambiguously identified proteins (false discovery rate ≤ 1%). Major differences were observed between the non-COVID-19 and nonsusceptible groups. Bioinformatics analysis of DE proteins revealed human proteomic signatures related to inflammatory responses, central cellular processes, and antiviral activity associated with the saliva of SARS-CoV-2-infected patients (p-value ≤ 0.0004). Discriminatory biomarker signatures from human saliva include cystatins, protective molecules present in the oral cavity, calprotectins, involved in cell cycle progression, and histones, related to nucleosome functions. The expression levels of two human proteins related to protein transport in the cytoplasm, DYNC1 (p-value, 0.0021) and MAPRE1 (p-value, 0.047), correlated with angiotensin-converting enzyme 2 (ACE2) plasma activity. Finally, the proteomes of microorganisms present in the saliva samples showed 4 main microbial functional features related to ribosome functioning that were overrepresented in the infected group. CONCLUSION: Our study explores potential candidates involved in pathways implicated in SARS-CoV-2 susceptibility, although further studies in larger cohorts will be necessary.

Evaluation of tissue-engineered human acellular vessels as a Blalock-Taussig-Thomas shunt in a juvenile primate model.

Objectives: Palliative treatment of cyanotic congenital heart disease (CCHD) uses systemic-to-pulmonary conduits, often a modified Blalock-Taussig-Thomas shunt (mBTTs). Expanded polytetrafluoroethylene (ePTFE) mBTTs have associated risks for thrombosis and infection. The Human Acellular Vessel (HAV) (Humacyte, Inc) is a decellularized tissue-engineered blood vessel currently in clinical trials in adults for vascular trauma, peripheral artery disease, and end-stage renal disease requiring hemodialysis. In addition to restoring blood flow, the engineered HAV demonstrates the capacity for host cellular remodeling into native-like vasculature. Here we report preclinical evaluation of a small-diameter (3.5 mm) HAV as a mBTTs in a non-human primate model. Methods: We implanted 3.5 mm HAVs as right subclavian artery to pulmonary artery mBTTs in non-immunosuppressed juvenile rhesus macaques (n = 5). HAV patency, structure, and blood flow were assessed by postoperative imaging from 1 week to 6 months. Histology of HAVs and surrounding tissues was performed. Results: Surgical procedures were well tolerated, with satisfactory anastomoses, showing feasibility of using the 3.5 mm HAV as a mBTTs. All macaques had some immunological reactivity to the human extracellular matrix, as expected in this xenogeneic model. HAV mBTTs remained patent for up to 6 months in animals, exhibiting mild immunoreactivity. Two macaques displaying more severe immunoreactivity to the human HAV material developed midgraft dilatation without bleeding or rupture. HAV repopulation by host cells expressing smooth muscle and endothelial markers was observed in all animals. Conclusions: These findings may support use of 3.5 mm HAVs as mBTTs in CCHD and potentially other pediatric vascular indications.

Pediatric extracorporeal cardiopulmonary resuscitation for yew cardiotoxicity.

INTRODUCTION: English yew is an evergreen conifer frequently planted in household gardens and, when ingested in large doses, results in severe cardiotoxicity characterized by difficult to control ventricular arrhythmias with high rates of mortality. CASE REPORT: A previously healthy teenage female presented as an out-of-hospital cardiac arrest with refractory ventricular arrhythmias and severe biventricular dysfunction. Due to rapid deterioration in her clinical status, she was cannulated onto venoarterial extracorporeal membrane oxygenation (ECMO) which resulted in rapid normalization of her rhythm and ventricular function. DISCUSSION: Our case highlights the importance of keeping a broad differential diagnosis when considering etiologies of ventricular arrhythmias in the pediatric population. The final diagnosis was not made until after discharge and implantable cardiac defibrillator (ICD) placement. CONCLUSION: The delayed diagnosis of this intentional English yew ingestion ultimately resulted subsequent ICD removal. Early ECMO activation in cases of English yew toxicity can be essential for patient survival.

Outcomes After Hybrid Palliation for Infants With Critical Left Heart Obstruction.

BACKGROUND: Hybrid palliation (bilateral pulmonary artery banding with or without ductal stenting) is an initial management strategy for infants with critical left heart obstruction and serves as palliation until subsequent operations are pursued. OBJECTIVES: This study sought to determine patient characteristics and factors associated with subsequent outcomes for infants who underwent hybrid palliation. METHODS: From 2005 to 2019, 214 of 1,236 prospectively enrolled infants within the Congenital Heart Surgeons' Society's critical left heart obstruction cohort underwent hybrid palliation across 24 institutions. Multivariable hazard modeling with competing risk methodology was performed to determine risk and factors associated with outcomes of biventricular repair, Fontan procedure, transplantation, or death. RESULTS: Preoperative comorbidities (eg, prematurity, low birth weight, genetic syndrome) were identified in 70% of infants (150 of 214). Median follow-up was 7 years, ranging up to 17 years. Overall 12-year survival was 55%. At 5 years after hybrid palliation, 9% had biventricular repair, 36% had Fontan procedure, 12% had transplantation, 35% died without surgical endpoints, and 8% were alive without an endpoint. Factors associated with transplantation were absence of ductal stent, older age, absent interatrial communication, smaller aortic root size, larger tricuspid valve area z-score, and larger left ventricular volume. Factors associated with death were low birth weight, concomitant genetic syndrome, cardiopulmonary bypass use during hybrid palliation, moderate to severe tricuspid valve regurgitation, and smaller ascending aortic size. CONCLUSIONS: Mortality remains high after hybrid palliation for infants with critical left heart obstruction. Nonetheless, hybrid palliation may facilitate biventricular repair for some infants and for others may serve as stabilization for intended functional univentricular palliation or primary transplantation.

A ROTEM-guided algorithm aimed to reduce blood product utilization during neonatal and infant cardiac surgery.

BACKGROUND: Neonates and infants undergoing cardiac surgery tend to receive high volumes of blood products. The use of rotational thromboelastometry (ROTEM®) has been shown to reduce the administration of blood products in adults after cardiac surgery. We sought to develop a targeted administration of blood products based on ROTEM® to reduce blood product utilization during and after neonatal and infant cardiac surgery. METHODS: We conducted a retrospective review of data from a single center for neonates and infants undergoing congenital cardiac surgery using cardiopulmonary bypass (CPB) from September 2018-April 2019 (control group). Then, using a ROTEM® algorithm, we collected data prospectively between April-November 2021 (ROTEM group). Data collected included age, weight, gender, procedure, STAT score, CPB time, aortic cross-clamp time, volume, and type of blood products administered in the operating room and cardiothoracic intensive care unit (CTICU). In addition, ROTEM® data, coagulation profile in CTICU, chest tube output at 6 and 24 hours, use of factors concentrate, and thromboembolic complications were recorded. RESULTS: The final cohort of patients included 28 patients in the control group and 40 patients in the ROTEM group. The cohort included neonates and infants undergoing the following procedures: arterial switch, aortic arch augmentation, Norwood procedure, and comprehensive stage II procedure. There were no differences in the demographics or procedure complexity between the two groups. Patients in the ROTEM® group received fewer platelets (36 ± 12 vs. 49 ± 27 mL/kg, p 0.028) and cryoprecipitate (8 ± 3 vs. 15 ± 10 mL/kg, p 0.001) intraoperatively when compared to the control group. CONCLUSION: The utilization of ROTEM® may have contributed to a significant reduction in some blood product administration during cardiac surgery for infants and neonates. ROTEM® data may play a role in reducing blood product administration in neonatal and infant cardiac surgery.

Patch Materials for Pulmonary Artery Arterioplasty and Right Ventricular Outflow Tract Augmentation: A Review.

Patch augmentation of the right ventricular outflow tract (RVOT) and pulmonary artery (PA) arterioplasty are relatively common procedures in the surgical treatment of patients with congenital heart disease. To date, several patch materials have been applied with no agreed upon clinical standard. Each patch type has unique performance characteristics, cost, and availability. There are limited data describing the various advantages and disadvantages of different patch materials. We performed a review of studies describing the clinical performance of various RVOT and PA patch materials and found a limited but growing body of literature. Short-term clinical performance has been reported for a multitude of patch types, but comparisons are limited by inconsistent study design and scarce histologic data. Standard clinical criteria for assessment of patch efficacy and criteria for intervention need to be applied across patch types. The field is progressing with improvements in outcomes due to newer patch technologies focused on reducing antigenicity and promoting neotissue formation which may have the ability to grow, remodel, and repair.

Proactive risk mitigation for cardiac arrest prevention in high-risk patients with congenital heart disease.

Objective: The prevalence of postoperative cardiac arrest (CA) increases with cardiothoracic surgical case complexity and is associated with a 40% to 50% mortality. Despite having a low overall surgical mortality rate at our center, our postoperative CA rates were higher than expected, with an observed-to-expected ratio of 2.6. Utilizing quality improvement methodology, we evaluated the influence of proactive risk mitigation on postprocedure CA in a high-risk cohort of pediatric cardiac patients. Methods: This single-center study utilized the Institute for Healthcare Improvement model. We created and implemented our Proactive Mitigation to Decrease Serious Adverse Events program in July 2020, prospectively enrolling preidentified high-risk patients. Enrolled patients underwent scheduled multidisciplinary reviews via virtual platform at 2 periprocedural time points with discussion of patient-specific risks and the subsequent development of proactive risk mitigation plans. Primary outcome measures were derived from the Pediatric Cardiac Critical Care Consortium national registry and included rate of postprocedure CA within 7 days and an institution-specific observed-to-expected ratio for postoperative CA. Results: Our baseline median number of high-risk cases between postprocedure CAs was 3. Following project initiation, median high-risk cases between events increased to 7. Our observed-to-expected ratio for postoperative CA decreased from 2.56 during the 12 months before Proactive Mitigation to Decrease Serious Adverse Events program implementation to 1.01 during the 12 months after Proactive Mitigation to Decrease Serious Adverse Events program implementation, and hospital length of stay decreased by ∼10 days. Conclusions: Implementation of periprocedure-related proactive risk mitigation strategies in high-risk pediatric cardiac patients led to improvement in postprocedure CA with a 133% increase in high-risk cases between events.

Preemptive stenting of the left pulmonary artery during comprehensive stage 2 procedure does not influence Fontan candidacy.

Objective: Pulmonary artery reconstruction during comprehensive stage 2 (CS2) procedure can be challenging. Since 2017, we have employed preemptive left pulmonary artery (LPA) stenting. We hypothesized that LPA stenting promotes adequate growth and without compromising Fontan candidacy. Herewith, we report our midterm results. Methods: From 2002 to 2020, 159 patients underwent CS2. Patients were divided as follows: no stent (n = 122; Group 1) and perioperative LPA stent (n = 37; Group 2). Group 2 was subdivided according to unplanned stent (n = 17; Group 2a) or preemptive stent (n = 20; Group 2b). Relevant perioperative data was reviewed. Nonparametric statistics were utilized. Results: Median age and weight at surgery and hospital length of stay after CS2 did not differ between groups. Median cardiopulmonary bypass and crossclamp times were significantly greater in Group 1 (265 vs 243 minutes [P = .021] and 46 vs 26 minutes [P = .008]). In-hospital mortality was similar between Groups 1 and 2 (9.0% vs 18.9%, respectively [P = .1348]). Group 2b demonstrated a superior survival compared to Group 2a (P = .0335) but not Group 1 (P > .9999). Preemptive stenting significantly increased median hilar LPA diameter at CS2 exit angiogram compared with no stenting (P < .0001). Groups 2a and 2b significantly increased the pre-Fontan diameter of the hilar LPA when compared with Group 1 (6.1 and 6.8 vs 5.7 mm, respectively [P < .0001]). A further 120 patients underwent Fontan operation (75%). Median follow-up for Groups 1 and 2 were 7.4 and 3.0 years, respectively. Conclusions: Perioperative LPA stenting during CS2 does not adversely affect pulmonary growth. Preemptive stenting seems advantageous for LPA growth in preparation for Fontan completion.

Enzyme adaptation to habitat thermal legacy shapes the thermal plasticity of marine microbiomes.

Microbial communities respond to temperature with physiological adaptation and compositional turnover. Whether thermal selection of enzymes explains marine microbiome plasticity in response to temperature remains unresolved. By quantifying the thermal behaviour of seven functionally-independent enzyme classes (esterase, extradiol dioxygenase, phosphatase, beta-galactosidase, nuclease, transaminase, and aldo-keto reductase) in native proteomes of marine sediment microbiomes from the Irish Sea to the southern Red Sea, we record a significant effect of the mean annual temperature (MAT) on enzyme response in all cases. Activity and stability profiles of 228 esterases and 5 extradiol dioxygenases from sediment and seawater across 70 locations worldwide validate this thermal pattern. Modelling the esterase phase transition temperature as a measure of structural flexibility confirms the observed relationship with MAT. Furthermore, when considering temperature variability in sites with non-significantly different MATs, the broadest range of enzyme thermal behaviour and the highest growth plasticity of the enriched heterotrophic bacteria occur in samples with the widest annual thermal variability. These results indicate that temperature-driven enzyme selection shapes microbiome thermal plasticity and that thermal variability finely tunes such processes and should be considered alongside MAT in forecasting microbial community thermal response.

Norwood operation versus comprehensive stage II after bilateral pulmonary artery banding palliation for infants with critical left heart obstruction.

OBJECTIVE: To determine patient characteristics and outcomes after Norwood versus comprehensive stage II (COMPSII) for infants with critical left heart obstruction who had prior hybrid palliation (bilateral pulmonary artery banding ± ductal stent). METHODS: From 23 Congenital Heart Surgeons' Society institutions (2005-2020), 138 infants underwent hybrid palliation followed by either Norwood (n = 73, 53%) or COMPSII (n = 65). Baseline characteristics were compared between Norwood and COMPSII groups. Parametric hazard model with competing risk methodology was used to determine risk and factors associated with outcomes of Fontan, transplantation, or death. RESULTS: Infants who underwent Norwood versus COMPSII had a higher prevalence of prematurity (26% vs 14%, P = .08), lower birth weight (median 2.8 vs 3.2 kg, P < .01) and less frequent ductal stenting (37% vs 99%; P < .01). Norwood was performed at a median age of 44 days and median weight of 3.5 kg, versus COMPSII at 162 days and 6.0 kg (both P < .01). Median follow-up was 6.5 years. At 5 years after Norwood and COMPSII, respectively; 50% versus 68% had Fontan (P = .16), 3% versus 5% had transplantation (P = .70), 40% versus 15% died (P = .10), and 7% versus 11% are alive without transition, respectively. For factors associated with either mortality or Fontan, only preoperative mechanical ventilation occurred more frequently in the Norwood group. CONCLUSIONS: Higher prevalence of prematurity, lower birth weight, and other patient-related characteristics in the Norwood versus COMPSII groups may influence differences in outcomes that were not statistically significant for this limited risk-adjusted cohort. The clinical decision regarding Norwood versus COMPSII after initial hybrid palliation remains challenging.

Surgical outcomes of infective endocarditis in pediatrics: Moving the needle to a contemporary, multidisciplinary approach.

BACKGROUND: Infective endocarditis (IE) is an uncommon disease in children that, when present, is accompanied by significant morbidity and mortality. The presence of congenital heart disease often complicates management. The aim of the present study is to describe the characteristics and outcomes of children undergoing surgery for IE. METHODS: A retrospective chart review from 2004 to 2020 was conducted to identify consecutive patients younger than age 20 years with IE undergoing surgery. RESULTS: A total of 94 patients with IE were identified, of whom 47 underwent surgery at a median age of 16.7 years. Thirty-one patients (65.95%) had congenital heart disease. Vegetation and embolic phenomena occurred in 41 and 29 patients (87.23% and 61.7%), respectively, with the brain as most common location (57.1%). Native valve involvement had a greater tendency to embolize (P < .001). Staphylococcus spp was the most common organism (49%). The mitral valve was the most affected (31.9%). Seven (14.9%) patients had multivalvar involvement and valve replacement was the most common procedure performed (37 patients; 78.7%). There were 3 operative deaths (6.4%). Median length of hospital stay was 21 days. Risk factors for prolonged hospital stay were time to surgery in days (P < .001) and native valvar involvement (P = .05). Five patients (10.6%) had postoperative recurrent IE. Survival at 1 and 5 years was 93.6% and 89.4%, respectively. CONCLUSIONS: Children with IE can undergo surgery with acceptable results. The morbidity, but not mortality, is driven by embolic complications. Staphylococcus spp and native valve involvement are significant risk factors. VIDEO ABSTRACT.

Quality Report: Postoperative Guideline Implementation Reduces Length of Stay after Fontan Procedure.

Introduction: Patients following the Fontan procedure have a physiology that results in prolonged pleural effusion, often delaying hospital discharge. The hospital length of stay (LOS) of patients following the Fontan procedure at our institution was significantly longer than the Society of Thoracic Surgery benchmark. This quality improvement project aimed to decrease hospital LOS in patients following the Fontan procedure from a baseline of 23 days to 7 days by January 1, 2021, and sustain indefinitely. Methods: We implemented standardized postoperative clinical practice guidelines in April 2020. We designed guidelines using previously published protocols. Key features included an ambulatory PleurX drain (BD, Franklin Lakes, N.J.), diuresis with fluid restriction, and pulmonary vasodilation with supplemental oxygen and sildenafil. All patients were discharged from the hospital with a PleurX drain in place. We compared clinical outcome variables before and after guideline implementation. As a balancing measure, we tracked 30-day readmissions. Results: One hundred seven patients underwent the Fontan procedure before guideline implementation from January 2015 to January 2020, with an average hospital LOS of 23 days. Postguideline implementation, 35 patients underwent the Fontan procedure from April 2020 to July 2022, with an average hospital LOS of 8 days in 2020, which further improved to an average hospital LOS of 7 days. There was no change in 30-day readmission after guideline implementation (24% pre versus 23% post; P = 0.86). Conclusion: Implementing clinical practice guidelines for patients following the Fontan procedure led to an over 50% reduction in hospital LOS without increasing 30-day readmission.

Impact of Maternal-Fetal Environment on Outcomes Following the Hybrid Procedure in the Single Ventricle Population.

Treatment of infants with hypoplastic left heart syndrome (HLHS) remains challenging, and those affected remain with significant risks for mortality and morbidity throughout their lifetimes. The maternal-fetal environment (MFE) has been shown to affect outcomes for infants with HLHS after the Norwood procedure. The hybrid procedure, comprised of both catheterization and surgical components, is a less invasive option for initial intervention compared to the Norwood procedure. It is unknown how the MFE impacts outcomes following the hybrid procedure. This is a single-center, retrospective study of infants born with HLHS who underwent hybrid palliation from January 2009 to August 2021. Predictor variables analyzed included fetal, maternal, and postnatal factors. The primary outcome was mortality prior to Stage II palliation. We studied a 144-subject cohort. There was a statistically significant difference in mortality prior to stage II palliation in infants with prematurity, small for gestational age, and aortic atresia subtype (p < 0.001, p = 0.009, and p = 0.008, respectively). There was no difference in mortality associated with maternal diabetes, hypertension, obesity, smoking or illicit drug use, or advanced maternal age. State and national area deprivation index scores were associated with increased risk of mortality in the entire cohort, such that infants born in areas with higher deprivation had a higher incidence of mortality. Several markers of an impaired MFE, including prematurity, small for gestational age, and higher deprivation index scores, are associated with mortality following hybrid palliation. Individual maternal comorbidities were not associated with higher mortality. The MFE may be a target for prenatal counseling and future interventions to improve pregnancy and neonatal outcomes in this population.

Computational analysis of serum-derived extracellular vesicle miRNAs in juvenile sheep model of single stage Fontan procedure.

Patients with single ventricle heart defects requires a series of staged open-heart procedures, termed Fontan palliation. However, while lifesaving, these operations are associated with significant morbidity and early mortality. The attendant complications are thought to arise in response to the abnormal hemodynamics induced by Fontan palliation, although the pathophysiology underlying these physicochemical changes in cardiovascular and other organs remain unknown. Here, we investigated the microRNA (miRNA) content in serum and serum-derived extracellular vesicles (EVs) by sequencing small RNAs from a physiologically relevant sheep model of the Fontan operation. The differential expression analysis identified the enriched miRNA clusters in (1) serum vs. serum-derived EVs and (2) pre-Fontan EVs vs. post-Fontan EVs. Metascape analysis showed that the overexpressed subset of EV miRNAs by Fontan procedure target liver-specific cells, underscoring a potentially important pathway involved in the liver dysfunction that occurs as a consequence of Fontan palliation. We also found that post-Fontan EV miRNAs were associated with senescence and cell death, whereas pre-Fontan EV miRNAs were associated with stem cell maintenance and epithelial-to-mesenchymal transition. This study shows great potential to identify novel circulating EV biomarkers from Fontan sheep serum that may be used for the diagnosis, prognosis, and therapeutics for patients that have undergone Fontan palliation.

Fecal Metabolome and Bacterial Composition in Severe Obesity: Impact of Diet and Bariatric Surgery.

The aim of this study was to monitor the impact of a preoperative low-calorie diet and bariatric surgery on the bacterial gut microbiota composition and functionality in severe obesity and to compare sleeve gastrectomy (SG) versus Roux-en-Y gastric bypass (RYGB). The study also aimed to incorporate big data analysis for the omics results and machine learning by a Lasso-based analysis to detect the potential markers for excess weight loss. Forty patients who underwent bariatric surgery were recruited (14 underwent SG, and 26 underwent RYGB). Each participant contributed 4 fecal samples (baseline, post-diet, 1 month after surgery and 3 months after surgery). The bacterial composition was determined by 16S rDNA massive sequencing using MiSeq (Illumina). Metabolic signatures associated to fecal concentrations of short-chain fatty acids, amino acids, biogenic amines, gamma-aminobutyric acid and ammonium were determined by gas and liquid chromatography. Orange 3 software was employed to correlate the variables, and a Lasso analysis was employed to predict the weight loss at the baseline samples. A correlation between Bacillota (formerly Firmicutes) abundance and excess weight was observed only for the highest body mass indexes. The low-calorie diet had little impact on composition and targeted metabolic activity. RYGB had a deeper impact on bacterial composition and putrefactive metabolism than SG, although the excess weight loss was comparable in the two groups. Significantly higher ammonium concentrations were detected in the feces of the RYGB group. We detected individual signatures of composition and functionality, rather than a gut microbiota characteristic of severe obesity, with opposing tendencies for almost all measured variables in the two surgical approaches. The gut microbiota of the baseline samples was not useful for predicting excess weight loss after the bariatric process.

TRPV1 Contributes to Modulate the Nitric Oxide Pathway and Oxidative Stress in the Isolated and Perfused Rat Heart during Ischemia and Reperfusion.

The transient vanilloid receptor potential type 1 (TRPV1) regulates neuronal and vascular functions mediated by nitric oxide (NO) and by the calcitonin gene-related peptide (CGRP). Here, we study the participation of TRPV1 in the regulation of myocardial injury caused by ischemia-reperfusion and in the control of NO, tetrahydrobiopterin (BH4), the cGMP pathway, CGRP, total antioxidant capacity (TAC), malondialdehyde (MDA) and phosphodiesterase-3 (PDE-3). Isolated hearts of Wistar rats perfused according to the Langendorff technique were used to study the effects of an agonist of TRPV1, capsaicin (CS), an antagonist, capsazepine (CZ), and their combination CZ+CS. The hearts were subjected to three conditions: (1) control, (2) ischemia and (3) ischemia-reperfusion. We determined cardiac mechanical activity and the levels of NO, cGMP, BH4, CGRP, TAC, MDA and PDE-3 in ventricular tissue after administration of CS, CZ and CZ+CS. Western blots were used to study the expressions of eNOS, iNOS and phosphorylated NOS (pNOS). Structural changes were determined by histological evaluation. CS prevented damage caused by ischemia-reperfusion by improving cardiac mechanical activity and elevating the levels of NO, cGMP, BH4, TAC and CGRP. TRPV1 and iNOS expression were increased under ischemic conditions, while eNOS and pNOS were not modified. We conclude that the activation of TRPV1 constitutes a therapeutic possibility to counteract the damage caused by ischemia and reperfusion by regulating the NO pathway through CGRP.

The bone-degrading enzyme machinery: From multi-component understanding to the treatment of residues from the meat industry.

Many microorganisms feed on the tissue and recalcitrant bone materials from dead animals, however little is known about the collaborative effort and characteristics of their enzymes. In this study, microbial metagenomes from symbionts of the marine bone-dwelling worm Osedax mucofloris, and from microbial biofilms growing on experimentally deployed bone surfaces were screened for specialized bone-degrading enzymes. A total of 2,043 taxonomically (closest match within 40 phyla) and functionally (1 proteolytic and 9 glycohydrolytic activities) diverse and non-redundant sequences (median pairwise identity of 23.6%) encoding such enzymes were retrieved. The taxonomic assignation and the median identity of 72.2% to homologous proteins reflect microbial and functional novelty associated to a specialized bone-degrading marine community. Binning suggests that only one generalist hosting all ten targeted activities, working in synergy with multiple specialists hosting a few or individual activities. Collagenases were the most abundant enzyme class, representing 48% of the total hits. A total of 47 diverse enzymes, representing 8 hydrolytic activities, were produced in Escherichia coli, whereof 13 were soluble and active. The biochemical analyses revealed a wide range of optimal pH (4.0-7.0), optimal temperature (5-65 °C), and of accepted substrates, specific to each microbial enzyme. This versatility may contribute to a high environmental plasticity of bone-degrading marine consortia that can be confronted to diverse habitats and bone materials. Through bone-meal degradation tests, we further demonstrated that some of these enzymes, particularly those from Flavobacteriaceae and Marinifilaceae, may be an asset for development of new value chains in the biorefinery industry.