Safety, Efficacy and Outcome of Rotational Thrombectomy assisted Endovascular Revascularisation of the Superior Mesenteric Artery in Acute Thromboembolic Mesenteric Ischaemia.

PURPOSE: To evaluate the efficacy and safety of percutaneous rotational thrombectomy-assisted endovascular revascularization of acute thromboembolic superior mesenteric artery occlusions in acute mesenteric ischemia. MATERIALS AND METHODS: Fifteen cases of percutaneous rotational thrombectomy- assisted (Rotarex S, BD, USA) revascularization were retrospectively analyzed. The etiology was embolic in 40 % of cases and thrombotic in 60 %. A "Thrombectomy in Visceral Ischemia" (TIVI) 5-point score determined vessel patency at presentation, after percutaneous rotational thrombectomy, and after adjunctive technologies. TIVI 3 indicated nearly complete revascularization (minimal residual side branch thrombus). TIVI 4 indicated complete revascularization. Technical success was defined as successful device application and a final TIVI score of 3/4 after adjunctive technologies. Safety and outcome were also analyzed. RESULTS: Device application via femoral access was feasible in 100 % of cases and improved flow in 86.7 % of cases (1â€Š× TIVI 0→1, 11â€Š× TIVI 0→2, 1â€Š× TIVI 1→2). There was no change in 13.3 % of cases (2â€Š× TIVI 2→2). Additional devices resulted in further flow improvement in 93.3 % of cases (8â€Š× TIVI 3, 6â€Š× TIVI 4). One recanalization failed (TIVI 2→2→2). After adjunctive technologies (10â€Š× manual aspiration, 11â€Š× angioplasty, 9â€Š× stenting), the technical success rate was 93.3 %. The mean procedure time was 40.5(±â€Š14) minutes for embolism and 72.1(±â€Š20) minutes for thrombosis. There was one device-related major complication (catheter tip fracture) resulting in a device-related safety rate of 93.3 %. The overall major complication rate was 20 %. Surgical exploration (13 ×), bowel resection (9 ×) and Fogarty embolectomy/bypass (3 ×) were also performed. The 30-day mortality rate was 40 %. CONCLUSION: Percutaneous rotational thrombectomy is an effective adjunct for rapid endovascular recanalization of acute thromboembolic superior mesenteric artery occlusions with an acceptable rate of major procedural complications. KEY POINTS: · Percutaneous rotational thrombectomy-assisted superior mesenteric artery revascularization in acute occlusive mesenteric ischemia is feasible and effective.. · Percutaneous rotational thrombectomy facilitates rapid flow restoration in native and stented superior mesenteric artery segments.. · Brachial access should be considered in the case of steep take-off angles of the superior mesenteric artery..

Training for endovascular therapy of acute arterial disease and procedure-related complication: An extracorporeally-perfused human cadaver model study.

PURPOSE: The aim of this study was to evaluate the usability of a recently developed extracorporeally-perfused cadaver model for training the angiographic management of acute arterial diseases and periprocedural complications that may occur during endovascular therapy of the lower extremity arterial runoff. MATERIALS AND METHODS: Continuous extracorporeal perfusion was established in three fresh-frozen body donors via inguinal and infragenicular access. Using digital subtraction angiography for guidance, both arterial embolization (e.g., embolization using coils, vascular plugs, particles, and liquid embolic agents) and endovascular recanalization procedures (e.g., manual aspiration or balloon-assisted embolectomy) as well as various embolism protection devices were tested. Furthermore, the management of complications during percutaneous transluminal angioplasty, such as vessel dissection and rupture, were exercised by implantation of endovascular dissection repair system or covered stents. Interventions were performed by two board-certified interventional radiologists and one resident with only limited angiographic experience. RESULTS: Stable extracorporeal perfusion was successfully established on both thighs of all three body donors. Digital subtraction angiography could be performed reliably and resulted in realistic artery depiction. The model allowed for repeatable training of endovascular recanalization and arterial embolization procedures with typical tactile feedback in a controlled environment. Furthermore, the handling of more complex angiographic devices could be exercised. Whereas procedural success was be ascertained for most endovascular interventions, thrombectomies procedures were not feasible in some cases due to the lack of inherent coagulation. CONCLUSION: The presented perfusion model is suitable for practicing time-critical endovascular interventions in the lower extremity runoff under realistic but controlled conditions.

Photon-Counting Detector CT for Femoral Stent Imaging in an Extracorporeally Perfused Human Cadaveric Model.

BACKGROUND AND AIMS: This study aims to compare the performance of first-generation dual-source photon-counting detector computed tomography (PCD-CT) to third-generation dual-source energy-integrating detector (EID-CT) regarding stent imaging in the femoral arterial runoff. METHODS: Continuous extracorporeal perfusion was established in 1 human cadaver using an inguinal and infragenicular access and peristaltic pump. Seven peripheral stents were implanted into both superior femoral arteries by means of percutaneous angioplasty. Radiation dose-equivalent CT angiographies (high-/medium-/low-dose: 10/5/3 mGy) with constant tube voltage of 120 kVp, matching iterative reconstruction algorithm levels, and convolution kernels were used both with PCD-CT and EID-CT. In-stent lumen visibility, luminal and in-stent attenuation as well as contrast-to-noise ratio (CNR) were assessed via region of interest and diameter measurements. Results were compared using analyses of variance and regression analyses. RESULTS: Maximum in-stent lumen visibility achieved with PCD-CT was 94.48% ± 2.62%. The PCD-CT protocol with the lowest lumen visibility (BV40: 78.93% ± 4.67%) performed equal to the EID-CT protocol with the best lumen visibility (BV59: 79.49% ± 2.64%, P > 0.999). Photon-counting detector CT yielded superior CNR compared with EID-CT regardless of kernel and dose level ( P < 0.001). Maximum CNR was 48.8 ± 17.4 in PCD-CT versus 31.28 ± 5.7 in EID-CT (both BV40, high-dose). The theoretical dose reduction potential of PCD-CT over EID-CT was established at 88% (BV40), 83% (BV48/49), and 73% (BV59/60), respectively. In-stent attenuation was not significantly different from luminal attenuation outside stents in any protocol. CONCLUSIONS: With superior lumen visibility and CNR, PCD-CT allowed for noticeable dose reduction over EID-CT while maintaining image quality in a continuously perfused human cadaveric model.

Comparison of ultrahigh and standard resolution photon-counting CT angiography of the femoral arteries in a continuously perfused in vitro model.

BACKGROUND: With the emergence of photon-counting CT, ultrahigh-resolution (UHR) imaging can be performed without dose penalty. This study aims to directly compare the image quality of UHR and standard resolution (SR) scan mode in femoral artery angiographies. METHODS: After establishing continuous extracorporeal perfusion in four fresh-frozen cadaveric specimens, photon-counting CT angiographies were performed with a radiation dose of 5 mGy and tube voltage of 120 kV in both SR and UHR mode. Images were reconstructed with dedicated convolution kernels (soft: Body-vascular (Bv)48; sharp: Bv60; ultrasharp: Bv76). Six radiologists evaluated the image quality by means of a pairwise forced-choice comparison tool. Kendall's concordance coefficient (W) was calculated to quantify interrater agreement. Image quality was further assessed by measuring intraluminal attenuation and image noise as well as by calculating signal-to-noise ratio (SNR) and contrast-to-noise ratios (CNR). RESULTS: UHR yielded lower noise than SR for identical reconstructions with kernels ≥ Bv60 (p < 0.001). UHR scans exhibited lower intraluminal attenuation compared to SR (Bv60: 406.4 ± 25.1 versus 418.1 ± 30.1 HU; p < 0.001). Irrespective of scan mode, SNR and CNR decreased while noise increased with sharper kernels but UHR scans were objectively superior to SR nonetheless (Bv60: SNR 25.9 ± 6.4 versus 20.9 ± 5.3; CNR 22.7 ± 5.8 versus 18.4 ± 4.8; p < 0.001). Notably, UHR scans were preferred in subjective assessment when images were reconstructed with the ultrasharp Bv76 kernel, whereas SR was rated superior for Bv60. Interrater agreement was high (W = 0.935). CONCLUSIONS: Combinations of UHR scan mode and ultrasharp convolution kernel are able to exploit the full image quality potential in photon-counting CT angiography of the femoral arteries. RELEVANCE STATEMENT: The UHR scan mode offers improved image quality and may increase diagnostic accuracy in CT angiography of the peripheral arterial runoff when optimized reconstruction parameters are chosen. KEY POINTS: • UHR photon-counting CT improves image quality in combination with ultrasharp convolution kernels. • UHR datasets display lower image noise compared with identically reconstructed standard resolution scans. • Scans in UHR mode show decreased intraluminal attenuation compared with standard resolution imaging.

Pancreatic Ischemia-Reperfusion Injury Following Endovascular Treatment of Symptomatic Celiac and Superior Mesenteric Artery Stenosis.

CLINICAL IMPACT: With endovascular therapy becoming the first-line treatment for symptomatic chronic mesenteric ischemia, acute pancreatitis within the context of abdominal ischemia-reperfusion injury may be seen more often in cross-sectional imaging following this kind of interventions and should therefore be kept in mind by the reading physician.

Photon-Counting Versus Energy-Integrating Detector CT Angiography of the Lower Extremity in a Human Cadaveric Model With Continuous Extracorporeal Perfusion.

OBJECTIVES: Detailed visualization of the arterial runoff is mandatory for the assessment of peripheral arterial occlusive disease. This study aims to compare the performance of a first-generation photon-counting detector computed tomography (PCD-CT) to a third-generation energy-integrating detector CT (EID-CT). MATERIALS AND METHODS: Computed tomography angiographies of 8 upper leg arterial runoffs were performed on human cadaveric models with continuous extracorporeal perfusion. For both PCD-CT and EID-CT, radiation dose-equivalent 120 kVp acquisition protocols (low-/medium-/high-dose: CTDI Vol = 3/5/10 mGy) were used. All scans were performed with standard collimation (PCD-CT: 144 × 0.4 mm; EID-CT: 96 × 0.6 mm), a pitch factor of 0.4, and a gantry rotation time of 1.0 second. Reformatting of data included the use of comparable vascular kernels (Bv 48/49), a slice thickness and increment of 1.0 mm, and a field of view of 150 × 150 mm. Eight radiologists evaluated image quality independently using a browser-based pairwise forced-choice comparison setup. Kendall concordance coefficient ( W ) was calculated to estimate interrater agreement. Signal-to-noise ratio and contrast-to-noise ratio (CNR) were compared based on 1-way analyses of variance and linear regression analysis. RESULTS: Low-dose PCD-CT achieved superior signal-to-noise ratio/CNR values compared with high-dose EID-CT ( P < 0.001). Linear regression analysis suggested that an EID-CT scan with a CTDI Vol of at least 15.5 mGy was required to match the CNR value of low-dose PCD-CT. Intraluminal contrast attenuation was higher in PCD-CT than EID-CT, irrespective of dose level (415.0 ± 31.9 HU vs 329.2 ± 29.4 HU; P < 0.001). Subjective image quality of low-dose PCD-CT was considered superior to high-dose EID-CT ( P < 0.001). Interrater agreement was high ( W = 0.989). CONCLUSIONS: Using cadaveric models with continuous extracorporeal perfusion allows for intraindividual image quality comparisons between PCD-CT and EID-CT on variable dose levels. With superior luminal contrast attenuation and denoising in angiographies of the peripheral arterial runoff, PCD-CT displayed potential for radiation saving of up to 83% compared with EID-CT.

Continuous extracorporeal femoral perfusion model for intravascular ultrasound, computed tomography and digital subtraction angiography.

OBJECTIVES: We developed a novel human cadaveric perfusion model with continuous extracorporeal femoral perfusion suitable for performing intra-individual comparison studies, training of interventional procedures and preclinical testing of endovascular devices. Objective of this study was to introduce the techniques and evaluate the feasibility for realistic computed tomography angiography (CTA), digital subtraction angiography (DSA) including vascular interventions, and intravascular ultrasound (IVUS). METHODS: The establishment of the extracorporeal perfusion was attempted using one formalin-fixed and five fresh-frozen human cadavers. In all specimens, the common femoral and popliteal arteries were prepared, introducer sheaths inserted, and perfusion established by a peristaltic pump. Subsequently, we performed CTA and bilateral DSA in five cadavers and IVUS on both legs of four donors. Examination time without unintentional interruption was measured both with and without non-contrast planning CT. Percutaneous transluminal angioplasty and stenting was performed by two interventional radiologists on nine extremities (five donors) using a broad spectrum of different intravascular devices. RESULTS: The perfusion of the upper leg arteries was successfully established in all fresh-frozen but not in the formalin-fixed cadaver. The experimental setup generated a stable circulation in each procedure (ten upper legs) for a period of more than six hours. Images acquired with CT, DSA and IVUS offered a realistic impression and enabled the sufficient visualization of all examined vessel segments. Arterial cannulating, percutaneous transluminal angioplasty as well as stent deployment were feasible in a way that is comparable to a vascular intervention in vivo. The perfusion model allowed for introduction and testing of previously not used devices. CONCLUSIONS: The continuous femoral perfusion model can be established with moderate effort, works stable, and is utilizable for medical imaging of the peripheral arterial system using CTA, DSA and IVUS. Therefore, it appears suitable for research studies, developing skills in interventional procedures and testing of new or unfamiliar vascular devices.

Awakening the Sleeping Carboxylase Function of Enzymes: Engineering the Natural CO2-Binding Potential of Reductases.

Developing new carbon dioxide (CO2) fixing enzymes is a prerequisite to create new biocatalysts for diverse applications in chemistry, biotechnology and synthetic biology. Here we used bioinformatics to identify a "sleeping carboxylase function" in the superfamily of medium-chain dehydrogenases/reductases (MDR), i.e. enzymes that possess a low carboxylation side activity next to their original enzyme reaction. We show that propionyl-CoA synthase from Erythrobacter sp. NAP1, as well as an acrylyl-CoA reductase from Nitrosopumilus maritimus possess carboxylation yields of 3 ± 1 and 4.5 ± 0.9%. We use rational design to engineer these enzymes further into carboxylases by increasing interactions of the proteins with CO2 and suppressing diffusion of water to the active site. The engineered carboxylases show improved CO2-binding and kinetic parameters comparable to naturally existing CO2-fixing enzymes. Our results provide a strategy to develop novel CO2-fixing enzymes and shed light on the emergence of natural carboxylases during evolution.

The multicatalytic compartment of propionyl-CoA synthase sequesters a toxic metabolite.

Cells must cope with toxic or reactive intermediates formed during metabolism. One coping strategy is to sequester reactions that produce such intermediates within specialized compartments or tunnels connecting different active sites. Here, we show that propionyl-CoA synthase (PCS), an ∼ 400-kDa homodimer, three-domain fusion protein and the key enzyme of the 3-hydroxypropionate bi-cycle for CO2 fixation, sequesters its reactive intermediate acrylyl-CoA. Structural analysis showed that PCS forms a multicatalytic reaction chamber. Kinetic analysis suggested that access to the reaction chamber and catalysis are synchronized by interdomain communication. The reaction chamber of PCS features three active sites and has a volume of only 33 nm3. As one of the smallest multireaction chambers described in biology, PCS may inspire the engineering of a new class of dynamically regulated nanoreactors.

A Chemo-Enzymatic Road Map to the Synthesis of CoA Esters.

Coenzyme A (CoA) is a ubiquitous cofactor present in every known organism. The thioesters of CoA are core intermediates in many metabolic processes, such as the citric acid cycle, fatty acid biosynthesis and secondary metabolism, including polyketide biosynthesis. Synthesis of CoA-thioesters is vital for the study of CoA-dependent enzymes and pathways, but also as standards for metabolomics studies. In this work we systematically tested five chemo-enzymatic methods for the synthesis of the three most abundant acyl-CoA thioester classes in biology; saturated acyl-CoAs, α,ß-unsaturated acyl-CoAs (i.e., enoyl-CoA derivatives), and α-carboxylated acyl-CoAs (i.e., malonyl-CoA derivatives). Additionally we report on the substrate promiscuity of three newly described acyl-CoA dehydrogenases that allow the simple conversion of acyl-CoAs into enoyl-CoAs. With these five methods, we synthesized 26 different CoA-thioesters with a yield of 40% or higher. The CoA esters produced range from short- to long-chain, include branched and α,ß-unsaturated representatives as well as other functional groups. Based on our results we provide a general guideline to the optimal synthesis method of a given CoA-thioester in respect to its functional group(s) and the commercial availability of the precursor molecule. The proposed synthetic routes can be performed in small scale and do not require special chemical equipment, making them convenient also for biological laboratories.

Screening and Engineering the Synthetic Potential of Carboxylating Reductases from Central Metabolism and Polyketide Biosynthesis.

Carboxylating enoyl-thioester reductases (ECRs) are a recently discovered class of enzymes. They catalyze the highly efficient addition of CO2 to the double bond of α,ß-unsaturated CoA-thioesters and serve two biological functions. In primary metabolism of many bacteria they produce ethylmalonyl-CoA during assimilation of the central metabolite acetyl-CoA. In secondary metabolism they provide distinct α-carboxyl-acyl-thioesters to vary the backbone of numerous polyketide natural products. Different ECRs were systematically assessed with a diverse library of potential substrates. We identified three active site residues that distinguish ECRs restricted to C4 and C5-enoyl-CoAs from highly promiscuous ECRs and successfully engineered a selected ECR as proof-of-principle. This study defines the molecular basis of ECR reactivity, allowing for predicting and manipulating a key reaction in natural product diversification.

Direct evidence for a covalent ene adduct intermediate in NAD(P)H-dependent enzymes.

The pyridine nucleotides NADH and NADPH (NAD(P)H) are ubiquitous redox coenzymes that are present in all living cells. Although about 16% of all characterized enzymes use pyridine nucleotides as hydride donors or acceptors during catalysis, a detailed understanding of how the hydride is transferred between NAD(P)H and the corresponding substrate is lacking for many enzymes. Here we present evidence for a new mechanism that operates during enzymatic hydride transfers using crotonyl-CoA carboxylase/reductase (Ccr) as a case study. We observed a covalent ene intermediate between NADPH and the substrate, crotonyl-CoA, using NMR, high-resolution MS and stopped-flow spectroscopy. Preparation of the ene intermediate further allowed direct access to the catalytic cycle of other NADPH-dependent enzymes-including those from type II fatty acid biosynthesis-in an unprecedented way, suggesting that formation of NAD(P)H ene intermediates is a more general principle in catalysis.

Ultrastructural analysis reveals cAMP-dependent enhancement of microvascular endothelial barrier functions via Rac1-mediated reorganization of intercellular junctions.

Evidence exists that cAMP stabilizes the endothelial barrier, in part via activation of the small GTPase Rac1. However, despite the high medical relevance of this signaling pathway, the mechanistic effects on intercellular contacts on the ultrastructural level are largely unknown. In microvascular endothelial cell monolayers, in which increased cAMP strengthened barrier properties, similar to intact microvessels in vivo, both forskolin and rolipram (F/R) to increase cAMP and 8-(4-chlorophenylthio)-2'-O-methyladenosine-3',5'-cyclic monophosphorothioate (O-Me-cAMP) to stimulate exchange protein directly activated by cAMP/Ras proximate-1 (EPac/Rap 1) signaling enhanced transendothelial electrical resistance and induced activation of Rac1. Concurrently, augmented immunofluorescence intensity and linearization of signals at cell borders were observed for intercellular junction proteins VE-cadherin and claudin 5. Ultrastructural analysis of the intercellular contact zone architecture documented that exposure to F/R or O-Me-cAMP led to a significant increase in the proportion of contact sites displaying complex interdigitations of cell borders, in which membranes of neighboring cells were closely apposed over comparatively long distances; in addition, they were stabilized by numerous intercellular junctions. Interference with Rac1 activation by NSC-23766 completely abolished both barrier stabilization and contact zone reorganization in response to O-Me-cAMP, whereas F/R-mediated Rac1 activation and barrier enhancement were not affected by NSC-23766. In parallel experiments using macrovascular endothelium, increased cAMP failed to induce Rac1 activation, barrier enhancement, and contact zone reorganization. These results indicate that, in microvascular endothelium, Rac1-mediated alterations in contact zone architecture contribute to cAMP-induced barrier stabilization.