Shear‐aggregated fibronectin with anti‐adhesive properties

Biomaterials based on proteins, such as fibronectin, have the potential to guide cell and tissue behaviour during healing as a function of their unique mechanical and bioactive properties. Fibronectin has been reported as a scaffold for attachment of fibroblasts and subsequent deposition of collagen. We have recently developed a derivative process of shear‐aggregated fibronectin that prevents cell attachment without causing cell death. This has potential applications in clinical situations where adhesions form across gliding surfaces and cause loss of function, e.g. peritoneal or flexor tendon adhesions. This in vitro study tested this derivative fibronectin biomaterial and its effects on aggressive adhesion‐forming cells, using rabbit flexor tendon synovial fibroblasts. We investigated degradation of the novel biomaterial, and attachment of fibroblasts to glass coated with the biomaterial, relative to fibroblast attachment to uncoated and fibronectin‐coated glass. We assessed infiltration of the derivative fibronectin biomaterial by fibroblasts and cytotoxicity of the biomaterial to fibroblasts. The interaction between fibroblasts and the derivative fibronectin biomaterial was visualized using time‐lapse photography. The derivative fibronectin biomaterial dissolved by 88% of its mass by 3 weeks. Fibroblast attachment to the novel biomaterial was significantly reduced at 6 h. After 24 h of exposure to the novel biomaterial, fibroblasts did not migrate into it, there was no cell death and no attachment was seen using time‐lapse. This novel derivative fibronectin biomaterial combines inhibition of fibroblast attachment with barrier effects and has suitable mechanical properties for surgical use in preventing adhesions in vivo. Copyright © 2010 John Wiley & Sons, Ltd.     

PET imaging of prostate tumors with 18F‐Al‐NOTA‐MATBBN

Overexpression of the gastrin‐releasing peptide receptor (GRPR) in prostate cancer provides a promising target for detection the disease. MATBBN is a new bombesin analog originating from the GRPR antagonists with a hydrophilic linker. In this study NOTA‐conjugated MATBBN was labeled by the Al¹⁸F method and the potential of ¹⁸F‐Al‐NOTA‐MATBBN for prostate tumor PET imaging was also evaluated. NOTA‐MATBBN was radiolabeled with ¹⁸F using Al¹⁸F complexes. Partition coefficient, in vitro stability and GRPR binding affinity were also determined. PET studies were performed with ¹⁸F‐Al‐NOTA‐MATBBN in PC‐3 tumor‐bearing mice. ¹⁸F‐Al‐NOTA‐MATBBN can be produced within 30 min with a decay‐corrected yield of 62.5 ± 2.1% and a radiochemical purity of >98%. The logP octanol–water value for the Al¹⁸F‐labeled BBN analog was ?2.40 ± 0.07 and the radiotracer was stable in phosphate‐buffered saline and human serum for 2 h. The IC₅₀ values of displacement for the ¹⁸F‐Al‐NOTA‐MATBBN with MATBBN was 126.9 ± 2.75 nm . The PC‐3 tumors were clearly visible with high contrast after injection of the labeled peptide. At 60 min post‐injection, the tumor uptakes for ¹⁸F‐Al‐NOTA‐MATBBN and ¹⁸F‐FDG were 4.59 ± 0.43 and 1.98 ± 0.35% injected dose/g, and tumor to muscle uptake radios for two tracers were 6.77 ± 1.10 and 1.78 ± 0.32, respectively. Dynamic PET revealed that ¹⁸F‐Al‐NOTA‐MATBBN was excreted mainly through the kidneys. GRPR‐binding specificity was also demonstrated by reduced tumor uptake of ¹⁸F‐Al‐NOTA‐MATBBN after coinjection with excess unlabeled MATBBN peptide at 1 h post‐injection. NOTA‐ MATBBN could be labeled rapidly with ¹⁸F using one step method. ¹⁸F‐Al‐NOTA‐MATBBN may be a promising PET imaging agent for prostate cancer. Copyright © 2014 John Wiley & Sons, Ltd.     

Concentration‐independent MRI of pH with a dendrimer‐based pH‐responsive nanoprobe

The measurement of extracellular pH (pH_e) has significant clinical value for pathological diagnoses and for monitoring the effects of pH‐altering therapies. One of the major problems of measuring pH_e with a relaxation‐based MRI contrast agent is that the longitudinal relaxivity depends on both pH and the concentration of the agent, requiring the use of a second pH‐unresponsive agent to measure the concentration. Here we tested the feasibility of measuring pH with a relaxation‐based dendritic MRI contrast agent in a concentration‐independent manner at clinically relevant field strengths. The transverse and longitudinal relaxation times in solutions of the contrast agent (GdDOTA‐4AmP)₄₄‐G5, a G5–PAMAM dendrimer‐based MRI contrast agent in water, were measured at 3 T and 7 T magnetic field strengths as a function of pH. At 3 T, longitudinal relaxivity (r₁) increased from 7.91 to 9.65 mM^?1 s^?1 (on a per Gd³⁺ basis) on changing pH from 8.84 to 6.35. At 7 T, r₁ relaxivity showed pH response, albeit at lower mean values; transverse relaxivity (r₂) remained independent of pH and magnetic field strengths. The longitudinal relaxivity of (GdDOTA‐4AmP)₄₄‐G5 exhibited a strong and reversible pH dependence. The ratio of relaxation rates R₂/R₁ also showed a linear relationship in a pH‐responsive manner, and this pH response was independent of the absolute concentration of (GdDOTA‐4AmP)₄₄‐G5 agent. Importantly, the nanoprobe (GdDOTA‐4AmP)₄₄‐G5 shows pH response in the range commonly found in the microenvironment of solid tumors. Copyright © 2015 John Wiley & Sons, Ltd.     

Heparin‐induced non‐necrotizing skin lesions: rarely associated with heparin‐induced thrombocytopenia

See also Warkentin TE, Linkins L‐A. Non‐necrotizing heparin‐induced skin lesions and the 4T’s score. This issue, pp 1483–. Summary. Background: Recently, there has been an increasing number of reports regarding adverse skin reactions to subcutaneous heparin administration. Case series have implied that heparin‐induced skin lesions are predominantly associated with life‐threatening heparin‐induced thrombocytopenia (HIT) in at least 22% of patients. Skin lesions, therefore, have been included in clinical scores for HIT. Objectives: To determine the association of heparin‐induced skin lesions with HIT. This would have a pivotal impact on further anticoagulatory management in patients with heparin‐induced skin lesions. Patients/Methods: In our observational cohort study, 87 consecutive patients with heparin‐induced skin lesions (85 occurring during low‐molecular‐weight heparin administration) were evaluated using a standardized internal protocol, including HIT diagnostics (heparin‐platelet factor 4‐ELISA, heparin‐induced platelet activation assay), platelet count monitoring, clinical/sonographical screening for thrombosis, skin allergy testing and, if necessary, histology. Results: None of the observed heparin‐induced skin lesions was due to HIT; all lesions were caused by delayed‐type IV‐hypersensitivity reactions (DTH) instead. Even the cutaneous reaction in one patient with concomitant HIT could be classified histologically as DTH reaction, amounting to an association of heparin‐induced skin lesions and HIT in 1.2% (1/87; 95% confidence interval, 0.00–0.06). Conclusion: Heparin‐induced skin lesions associated with use of low‐molecular‐weight heparins do not seem to be strongly associated with a systemic immunologic reaction in terms of HIT and might rather be due to DTH reactions than due to microvascular thrombosis. Hence, we propose refining existing pretest probability scores for HIT, unless underlying causes have been clarified.     

Imaging integrin alpha‐v‐beta‐3 expression in tumors with an 18F‐labeled dimeric RGD peptide

Integrin α_vβ₃ receptors are expressed on activated endothelial cells during neovascularization to maintain tumor growth. Many radiolabeled probes utilize the tight and specific association between the arginine–glycine–aspartatic acid (RGD) peptide and integrin α_vβ₃, but one main obstacle for any clinical application of these probes is the laborious multistep radiosynthesis of ¹⁸F. In this study, the dimeric RGD peptide, E‐[c(RGDfK)]₂, was conjugated with NODAGA and radiolabeled with ¹⁸F in a simple one‐pot process with a radiolabeling yield of 20%, the whole process lasting only 45 min. NODAGA‐E‐[c(RGDfK)]₂ labeled with ¹⁸F at a specific activity of 1.8 MBq nmol^?1 and a radiochemical purity of 100% could be achieved. The logP value of ¹⁸F‐labeled NODAGA‐E‐[c(RGDfK)]₂ was ?4.26 ± 0.02. In biodistribution studies, ¹⁸F‐NODAGA‐E‐[c(RGDfK)]₂ cleared rapidly from the blood with 0.03 ± 0.01 percentage injected dose per gram (%ID g^?1) in the blood at 2 h p.i., mainly via the kidneys, and showed good in vivo stability. Tumor uptake of ¹⁸F‐NODAGA‐E‐[c(RGDfK)]₂ (3.44 ± 0.20 %ID g^?1, 2 h p.i.) was significantly lower than that of reference compounds ⁶⁸Ga‐labeled NODAGA‐E‐[c(RGDfK)]₂ (6.26 ± 0.76 %ID g^?1; p <0.001) and ¹¹¹In‐labeled NODAGA‐E‐[c(RGDfK)]₂ (4.99 ± 0.64 %ID g^?1; p < 0.01). Co‐injection of an excess of unlabeled NODAGA‐E‐[c(RGDfK)]₂ along with ¹⁸F‐NODAGA‐E‐[c(RGDfK)]₂ resulted in significantly reduced radioactivity concentrations in the tumor (0.85 ± 0.13 %ID g^?1). The α_vβ₃ integrin‐expressing SK‐RC‐52 tumor could be successfully visualized by microPET with ¹⁸F‐labeled NODAGA‐E‐[c(RGDfK)]₂. In conclusion, NODAGA‐E‐[c(RGDfK)]₂ could be labeled rapidly with ¹⁸F using a direct aqueous, one‐pot method and it accumulated specifically in α_vβ₃ integrin‐expressing SK‐RC‐52 tumors, allowing for visualization by microPET. Copyright © 2013 John Wiley & Sons, Ltd.     

First‐in‐man fully leadless transvenous CRT‐P with a transseptal implant of WISE‐CRT® system and Micra® PM

We describe the world's first fully leadless cardiac resynchronization therapy pacing implant with transseptal approach, in a patient with pacemaker dependency, atrial fibrillation, ischemic hypokinetic cardiomyopathy, and a history of pocket infections. After lead extraction, we implanted a MicraTranscathether Pacing System® (Medtronic, Minneapolis, MN, USA) as the sole right ventricular pacemaker, and the WISECRT system (EBR Systems, Sunnyvale, CA, USA) to provide biventricular pacing. We performed the WISECRT implant procedure using the transseptal approach given the presence of a prosthetic aortic mechanical valve, achieving satisfactory periprocedural results.     

Interaction of fibrin with VE‐cadherin and anti‐inflammatory effect of fibrin‐derived fragments

Summary. Background: The interaction of the fibrin βN‐domain with VE‐cadherin on endothelial cells is implicated in transendothelial migration of leukocytes, and the β15–42 fragment representing part of this domain has been shown to inhibit this process. However, our previous study revealed that only a dimeric (β15–66)₂ fragment, corresponding to the full‐length βN‐domain and mimicking its dimeric arrangement in fibrin, bound to VE‐cadherin. Objective: To test our hypothesis that dimerization of β15–42‐containing fragments increases their affinity for VE‐cadherin and ability to inhibit transendothelial migration of leukocytes. Methods: Interaction of β15–42‐containing fragments with VE‐cadherin was characterized by ELISA and surface plasmon resonance. The inhibitory effect of such fragments was tested in vitro with a leukocyte transendothelial migration assay and in vivo with mouse models of peritonitis and myocardial ischemia–reperfusion injury. Results: First, we prepared the monomeric β15–42 and β15–64 fragments and their dimeric forms, (β15–44)₂ and (β15–66)₂, and studied their interaction with the fibrin‐binding domain of VE‐cadherin, VE‐cad(3). The experiments revealed that both dimeric fragments bound to VE‐cad(3) with high affinity, whereas the affinities of β15–42 and β15–64 were significantly lower. Next, we tested the ability of these fragments to inhibit leukocyte transmigration in vitro and infiltration into the inflamed peritoneum in vivo, and found that the inhibitory effects of the dimers on these processes were also superior. Furthermore, (β15–44)₂ significantly reduced myocardial injury induced by ischemia–reperfusion. Conclusion: The results confirm our hypotheses and indicate that (β15‐66)₂ and (β15‐44)₂, which exhibited much higher affinity for VE‐cadherin, are highly effective in suppressing inflammation by inhibiting leukocyte transmigration.