The CXCR4 chemokine receptor regulates migration and homing of cancer cells to specific metastatic sites. Determination of the CXCR4 receptor status will provide predictive information for disease ...prognosis and possible therapeutic intervention. However, previous attempts to localize CXCR4 using poorly characterized mouse monoclonal or rabbit polyclonal antibodies have produced predominant nuclear and occasional cytoplasmic staining but did not result in the identification of bona fide cell surface receptors.
In the present study, we extensively characterized the novel rabbit monoclonal anti-CXCR4 antibody (clone UMB-2) using transfected cells and tissues from CXCR4-deficient mice. Specificity of UMB-2 was demonstrated by cell surface staining of CXCR4-transfected cells; translocation of CXCR4 immunostaining after agonist exposure; detection of a broad band migrating at M(r) 38,000-43,000 in Western blots of homogenates from CXCR4-expressing cells; selective detection of the receptor in tissues from CXCR4+/+ but not from CXCR4-/- mice; and abolition of tissue immunostaining by preadsorption of UMB-2 with its immunizing peptide. In formalin-fixed, paraffin-embedded human tumor tissues, UMB-2 yielded highly effective plasma membrane staining of a subpopulation of tumor cells, which were often heterogeneously distributed throughout the tumor. A comparative analysis of the mouse monoclonal antibody 12G5 and other frequently used commercially available antibodies revealed that none of these was able to detect CXCR4 under otherwise identical conditions.
Thus, the rabbit monoclonal antibody UMB-2 may prove of great value in the assessment of the CXCR4 receptor status in a variety of human tumors during routine histopathological examination.
Members of the SLC4 bicarbonate transporter family are involved in solute transport and pH homeostasis. Here we report that disrupting the Slc4a10 gene, which encodes the Na⁺-coupled Cl⁻-HCO₃⁻ ...exchanger Slc4a10 (NCBE), drastically reduces brain ventricle volume and protects against fatal epileptic seizures in mice. In choroid plexus epithelial cells, Slc4a10 localizes to the basolateral membrane. These cells displayed a diminished recovery from an acid load in KO mice. Slc4a10 also was expressed in neurons. Within the hippocampus, the Slc4a10 protein was abundant in CA3 pyramidal cells. In the CA3 area, propionate-induced intracellular acidification and attenuation of 4-aminopyridine-induced network activity were prolonged in KO mice. Our data indicate that Slc4a10 is involved in the control of neuronal pH and excitability and may contribute to the secretion of cerebrospinal fluid. Hence, Slc4a10 is a promising pharmacological target for the therapy of epilepsy or elevated intracranial pressure.
Objective: A computer-enhanced instrumentation system was used in 148 patients to minimize access in cardiac surgical procedures. Methods: The da Vinci telemanipulation system (Intuitive Surgical, ...Mountain View, Calif) provides a high-resolution 3-dimensional videoscopic image and allows remote, tremor-free, and scaled control of endoscopic surgical instruments with 6 degrees of freedom. By April 2000, the system had been used in 131 patients for coronary artery bypass grafting and 17 patients for mitral valve repair. In the coronary bypass group, the system was used in one of three ways: (1) to take down the internal thoracic artery followed by a minimally invasive direct coronary bypass procedure (n = 81); (2) to perform the anastomosis between the internal thoracic artery and the left anterior descending coronary artery in standard-sternotomy coronary bypass (n = 15); or (3) for total endoscopic coronary artery bypass grafting to anastomose the left internal thoracic artery to the left anterior descending on the arrested heart (n = 27) or the beating heart (n = 8). In 17 patients with nonischemic mitral valve insufficiency the mitral valve was repaired. Closed-chest cardiopulmonary bypass with cardioplegic arrest (Port-Access technique; Heartport, Inc, Redwood City, Calif) was used for arrested-heart total endoscopic coronary bypass and mitral valve repair. Results: The da Vinci system allows for precise tissue handling and enables the endoscopic performance of cardiac surgical tasks that require a high degree of dexterity (coronary anastomosis, mitral valve repair). No technical mishaps have occurred. The internal thoracic artery was successfully taken down in 79 of 81 patients in the group undergoing minimally invasive coronary bypass and, after a steep learning curve, is currently performed in less than 40 minutes. The postoperative patency rate is 96.3%. Total endoscopic coronary bypass was completed in 22 of 27 cases with 95.4% patency as demonstrated by angiography at 3 months' follow-up. Closed-chest endoscopic beating-heart bypass grafting was successfully performed in 2 out of 8 patients with the use of a new endoscopic stabilizer. In the group having mitral valve repair, primary endoscopic computer-enhanced repair was successfully completed in 14 of 17 patients; three others had to be changed to a standard endoscopic technique, including 1 who required valve replacement. At 3 months' follow-up, 1 additional patient underwent early reoperation for recurrent mitral insufficiency. Overall early and late mortality in this cohort of 148 patients was 2.0% and was not related to the use of the system. Conclusion: In conclusion, computer-enhanced endoscopic cardiac surgery can be performed safely in selected patients. Internal thoracic artery takedown is now routinely performed with good results. Total endoscopic coronary bypass is feasible on the arrested heart but does not offer a major benefit over the minimally invasive direct approach because cardiopulmonary bypass is still required. The early clinical experience with closed-chest beating-heart bypass grafting outlines the limitations of this approach despite some procedural success. (J Thorac Cardiovasc Surg 2001;121:842-53)
The somatostatin receptor subtypes 1–5 (sst
1–sst
5) exhibit different intracellular trafficking and endosomal sorting after agonist exposure. The internalization of the somatostatin receptor ...subtypes sst
2, sst
3 and sst
5 occurs to a much higher extent after somatostatin exposure than of sst
1 or sst
4. After endocytosis, sst
2 and sst
5 recycle to the plasma membrane, whereas sst
3 is predominantly down-regulated. This review will focus on the molecular mechanisms of the differential intracellular trafficking of sst
2, sst
3 and sst
5, and discusses our current knowledge on somatostatin receptor interacting proteins.
Objective: The overexpression of somatostatin receptor 2 (sst2) in neuroendocrine tumors is the molecular basis for diagnostic and therapeutic application of the stable somatostatin analog ...octreotide. Recent evidence has shown that the immunocytochemical evaluation of sst2A status is of value for predicting response to octreotide therapy and disease prognosis. However, due to the lack of monoclonal and limited availability of specific polyclonal anti-sst2A antibodies, only very few patients can currently benefit from in vitro sst2 evaluation.
Methods: In the present study, we extensively characterized the novel rabbit monoclonal anti-sst2A antibody (clone UMB-1) using tissues from sst2-deficient mice and their wild-type littermates. UMB-1 was then subjected to a comparative study of immunohistochemistry on a series of histological specimens from formalin-fixed, paraffin-embedded human tumors and adjacent normal tissues.
Results: Immunoprecipitation experiments unequivocally demonstrated that UMB-1 selectively detected its cognate sst2A and did not cross-react with other proteins present in crude tissue homogenates. The UMB-1 monoclonal antibody, when compared with currently available polyclonal antisera, yielded several times more effective immunohistochemical staining of fixed-embedded tissues with a predominance of plasma membrane staining and very low cytoplasmic signal even without heat-based antigen retrieval. In addition, dual immunofluorescence revealed for the first time that the sst2A is present on not only gastrin-containing but also ghrelin-containing cells in human gastric mucosa.
Conclusion: Thus, the rabbit monoclonal antibody UMB-1 may prove of great value in the assessment of sst2A status in human neuroendocrine tumors during routine histopathological examination.
Pasireotide (SOM230) is currently under clinical evaluation as a successor compound to octreotide for the treatment of acromegaly, Cushing's disease, and carcinoid tumors. Whereas octreotide acts ...primarily via the sst(2A) somatostatin receptor, pasireotide was designed to exhibit octreotide-like sst(2A) activity combined with enhanced binding to other somatostatin receptor subtypes. In the present study, we used phophosite-specific antibodies to examine agonist-induced phosphorylation of the rat sst(2A) receptor. We show that somatostatin and octreotide stimulate the complete phosphorylation of a cluster of four threonine residues within the cytoplasmic (353)TTETQRT(359) motif in a variety of cultured cell lines in vitro as well as in intact animals in vivo. This phosphorylation was mediated by G protein-coupled receptor kinases (GRK) 2 and 3 and followed by rapid cointernalization of the receptor and ss-arrestin into the same endocytic vesicles. In contrast, pasireotide failed to promote substantial phosphorylation and internalization of the rat sst(2A) receptor. In the presence of octreotide or SS-14, SOM230 showed partial agonist behavior, inhibiting phosphorylation, and internalization of sst(2A). Upon overexpression of GRK2 or GRK3, pasireotide stimulated selective phosphorylation of Thr356 and Thr359 but not of Thr353 or Thr354 within the (353)TTETQRT(359) motif. Pasireotide-mediated phosphorylation led to the formation of relatively unstable beta-arrestin-sst(2A) complexes that dissociated at or near the plasma membrane. Thus, octreotide and pasireotide are equally active in inducing classical G protein-dependent signaling via the sst(2A) somatostatin receptor. Yet, we find that they promote strikingly different patterns of sst(2A) receptor phosphorylation and, hence, stimulate functionally distinct pools of beta-arrestin.
In 2007, the use of aprotinin was temporarily suspended following several publications raising serious safety concerns.(1.2.3) However, Canadian and European Health Authorities concluded that these ...studies suffered from major flaws and lifted the suspension in 2013.(4,5)
The reintroduction of aprotinin in 2016 was accompanied by a mandatory registry Nordic Aprotinin Patient Registry (NAPaR) in order to monitor the pattern of use, the effect of risk minimization measures, and overall safety.
Objective of this study was to analyse the use of aprotinin in Belgium.
NAPaR is a non-interventional Post-Authorisation Safety Study (PASS), endorsed by the Pharmacovigilance Risk Assessment Committee of the European Medicine Agency.
Between October 2018 and August 2020, six Belgian NAPaR centers included 694 patients median age 70 (range 13-90y); 62% male; median EuroSCORE II 4.6 (range 0.5-93.4). 25% of the patients had an EuroSCOREII > 10.
57% of the patients were under an active antiplatelet therapy and 17% were treated with novel oral anti-coagulants. Preoperative renal impairment was present in 2/3 of patients.
Only 4% of the patients received aprotinin on-label, i.e. for isolated coronary artery bypass grafting (CABG) on cardiopulmonary bypass (CPB).
In 96% of the patients, aprotinin was used off-label, mainly for redo surgery (32%), CABG + valve surgery (29%), aortic surgery (20%) and multiple valve surgery (17%). One third of the procedures were urgent (23%) or emergent (9%). Median bypass time was 128min (range 13-600min).
Median hospital stay was 11 days (range 3-151 days).
Main reported indications for the use of aprotinin were active dual antiplatelet therapy (68% in iCABG), redo surgery (16%) and expected prolonged CPB time (55%).
In 83% of the cases, aprotinin was administered at the Half Hammersmith dose (1 Mio KIU loading- and priming-dose + 250.000 KIU/h); most patients received the appropriate test dose (98%), none developed anaphylactic reactions.
Median perioperative blood loss was 400ml (range 40 -10050 ml), 44% of the patients had RBC transfusion and 6% underwent reoperation for bleeding or tamponade.
Overall mortality at discharge was 5%, i.e. within the range or lower than predicted by the EuroSCORE II. 2% of the patients had a postoperative stroke. Postoperative AKI as defined by a serum creatinine increase of 0,3mg/dL or more within 48 hours after surgery was present in 13% of cases. Most of these patients (76%) suffered from preoperative moderate or severe renal impairment including dialysis. Notably, in patients with postoperative AKI median CPB time was significantly higher than in patients without AKI: 172min (range 56-402) vs 125min (range 13-600min) (p<0,0001).
▪
In the Belgian centers, aprotinin was mainly used off-label and predominantly administered to older patients with significant comorbidities undergoing high-risk procedures. The observed safety outcomes are in line with published data within this high risk patient population.
The data from the Belgian NAPaR suggest that aprotinin has an excellent safety profile in adult cardiac surgery.
OBJECTIVES
To evaluate the performance and safety of an adjustable semi-rigid annuloplasty ring for mitral regurgitation (MR) in a multicentre study.
METHODS
Between March 2010 and December 2011, 30 ...subjects underwent mitral valve (MV) repair using the Cardinal adjustable annuloplasty ring. This device is a semi-rigid ring allowing postimplantation size adjustment, under beating-heart conditions, to optimize leaflet coaptation under echocardiographic guidance. Coaptation length was determined before and after adjustment by transoesophageal echocardiography.
RESULTS
The study enrolled 21 (70%) male and 9 (30%) female subjects with a mean age of 64 years. The approach was conventional midline sternotomy or mini-invasive right thoracotomy. Leaflet resection was done in 17 subjects, and chordal repair was used in 13. Concomitant procedures included coronary artery bypass grafting in 2 (7%) subjects, atrial ablation in 4 (13%) and tricuspid repair in 4 (13%). There was 1 (3%) early death unrelated to the study device. Intraoperative ring adjustment was performed in 24 of the 30 subjects. Residual MR was detected prior to adjustment in 6 subjects (4 mild and 2 moderate MR). Following adjustment, 5 subjects had no MR and 1 had trace MR. After adjustment, mean coaptation length improved from 7 ± 3 to 10 ± 3 mm (P < 0.0001). All patients who completed 1-year follow-up had less-than-mild MR, with the exception of 1 patient with ring dehiscence (and resultant 2+ MR) and 1 functional MR patient who developed recurrent 2+ MR due to persistent leaflet tethering.
CONCLUSIONS
MV repair with the Cardinal adjustable annuloplasty ring is a reliable technique that enables the adjustment of the ring diameter on a beating heart under echocardiographic control. Such technology allows the optimization of leaflet coaptation, providing minimal residual MR and durable repair.
A new poly(2-alkyl-2-oxazoline) having a branched side chain, namely, 2-(3-ethylheptyl)-2-oxazoline (EHOx), was synthesized. The microwave-assisted homopolymerization as well as the copolymerization ...with 2-ethyl-2-oxazoline (EtOx) are reported, whereby the determination of the reactivity ratios indicates the occurrence of a random polymerization. Further studies were performed regarding the thermal and the surface properties. p(EHOx) showed a glass transition of −6 °C, whereas no melting temperature could be detected, representing the amorphous poly(2-oxazoline) with the lowest reported T g to date. Furthermore, the systematic random copolymerization with EtOx in the range from 0 to 100% EHOx revealed a linear dependence of T g with composition as well as a complex dependence of the surface energy on the composition showing two plateau regimes.
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