A physiologically relevant culture platform for long-term studies of in vitro gingival tissue.

There is a clinical need to understand the etiologies of periodontitis, considering the growing socio-economic impact of the disease. Despite recent advances in oral tissue engineering, experimental approaches have failed to develop a physiologically relevant gingival model that combines tissue organization with salivary flow dynamics and stimulation of the shedding and non-shedding oral surfaces. Herein, we develop a dynamic gingival tissue model composed of a silk scaffold, replicating the cyto-architecture and oxygen profile of the human gingiva, along with a saliva-mimicking medium that reflected the ionic composition, viscosity, and non-Newtonian behavior of human saliva. The construct was cultured in a custom designed bioreactor, in which force profiles on the gingival epithelium were modulated through analysis of inlet position, velocity and vorticity to replicate the physiological shear stress of salivary flow. The gingival bioreactor supported the long-term in vivo features of the gingiva and improved the integrity of the epithelial barrier, critical against the invasion of pathogenic bacteria. Furthermore, the challenge of the gingival tissue with P. gingivalis lipopolysaccharide, as an in vitro surrogate for microbial interactions, indicated a greater stability of the dynamic model in maintaining tissue homeostasis and, thus, its applicability in long-term studies. The model will be integrated into future studies with the human subgingival microbiome to investigate host-pathogen and host-commensal interactions. STATEMENT OF SIGNIFICANCE: The major societal impact of human microbiome had reverberated up to the establishment of the Common Fund's Human Microbiome Project, that has the intent of studying the role of microbial communities in human health and diseases, including periodontitis, atopic dermatitis, or asthma and inflammatory bowel disease. In addition, these chronic diseases are emergent drivers of global socioeconomic status. Not only common oral diseases have been shown to be directly correlated with several systemic conditions, but they are differentially impacting some racial/ethnic and socioeconomic groups. To address this growing social disparity, the development of in vitro gingival model would provide a time and cost-effective experimental platform, able to mimic the spectrum of periodontal disease presentation, for the identification of predictive biomarkers for early-stage diagnosis.

Multisystemic inflammatory syndrome related to COVID-19, with latent tuberculosis in bone marrow, and satisfactory response to tocilizumab, in a 7-year-old boy.

Paediatric inflammatory multisystem syndrome temporally associated with COVID-19 (PIMS-TS) or multisystem inflammatory syndrome in children (MIS-C) is a new acute-onset systemic inflammatory disease, which mainly affects children. Latent tuberculosis infection (LTBI) is characterized by the presence of immune sensitization to Mycobacterium tuberculosis (MTB) in the absence of any clinical or radiological evidence of active disease. We present a child with MIS-C related to COVID-19, with latent TB in the bone marrow, and satisfactory response to tocilizumab. It is important to pay attention in the investigation of TB cases in countries with a high prevalence of tuberculosis, especially when opting for immunusuppression.

Mapping of brain tissue hematocrit in glioma and acute stroke using a dual autoradiography approach.

Hematocrit (Hct) determines the ability of blood to carry oxygen. While changes in systemic Hct are known to impact stroke or tumor control, changes in local (tissue) Hct (tHct) induced by these diseases have however received little attention. In this study, we evaluate tHct in acute stroke and in glioma models using a new approach to map tHct across the brain, a dual isotope autoradiography, based on injections of 125I-labeled albumin and 99mTc-lalbeled red blood cells in the same animal. For validation purpose, tHct was mapped in the rat brain (i) under physiological conditions, (ii) following erythropoietin injection, and (iii) following hemodilution. Then, tHct was then mapped in stroke (middle cerebral artery occlusion) and tumor models (9LGS and C6). The mean tHct values observed in healthy brains (tHct = 29 ± 1.3%), were modified as expected by erythropoietin (tHct = 36.7 ± 2.6%) and hemodilution (tHct = 24.2 ± 2.4%). Using the proposed method, we observed a local reduction, spatially heterogeneous, in tHct following acute stroke (tHct = 19.5 ± 2.5%) and in both glioma models (9LGS: tHct = 18.5 ± 2.3%, C6: tHct = 16.1 ± 1.2%). This reduction and this heterogeneity in tHct observed in stroke and glioma raises methodological issues in perfusion imaging techniques where tHct is generally overlooked and could impact therapeutic strategies.

Biodistribution and preliminary toxicity studies of nanoparticles made of Biotransesterified ß-cyclodextrins and PEGylated phospholipids.

BACKGROUND: The modification of ß-cyclodextrins (ßCDs) by grafting alkyl chains on the primary and/or secondary face yields derivatives (ßCD-C10) able to self-organize under nanoprecipitating conditions into nanoparticles (ßCD-C10-NP) potentially useful for drug delivery. The co-nanoprecipitation of ßCD-C10 with polyethylene glycol (PEG) chains yields PEGylated NPs (ßCD-C10-PEG-NP) with potentially improved stealthiness. The objectives of the present study were to characterize the in vivo biodistribution of ßCD-C10-PEG-NP with PEG chain length of 2000 and 5000Da using nuclear imaging, and to preliminarily evaluate the in vivo acute and extended acute toxicity of the most suitable system. RESEARCH DESIGN AND METHODS: The in vivo and ex vivo biodistribution features of naked and decorated nanoparticles were investigated over time following intravenous injection of 125I-radiolabeled nanoparticles to mice. The potential toxicity of PEGylated ßCD-C10 nanosuspensions was evaluated in a preliminary in vivo toxicity study involving blood assays and tissue histology following repeated intraperitoneal injections of nanoparticles to healthy mice. RESULTS: The results indicated that ßCD-C10-PEG5000-NP presented increased stealthiness with decreased in vivo elimination and increased blood kinetics without inducing blood, kidney, spleen, and liver acute and extended acute toxicity. CONCLUSIONS: ßCD-C10-PEG5000-NPs are stealth and safe systems with potential for drug delivery.

New diketopiperazines as vectors for peptide protection and brain delivery: Synthesis and biological evaluation.

New strategies allowing the transfer of molecules, especially peptides, through the blood-brain barriers are a major pharmacological challenge for the treatment of brain diseases. The present study aims at evaluating in vivo the cerebral bioavailability of carrier systems, based on small and functionalizable 2,5-diketopiperazine (DKP) motifs. We studied 2 different cyclo(Lys-Lys) DKP scaffolds alone and a cyclo(Lys-Gly) DKP carrier bearing as peptide model, the tau protein hexapeptide VQIVYK sequence. The different carrier systems were synthesized and radiolabeled using one of the free domains. The stability, biodistribution, and ability to cross blood-brain barrier were investigated in vivo in mice for 99m Tc-DKP scaffolds, 99m Tc-HVQIVYK peptide alone, and 99m Tc-DKP-VQIVYK. 125 I-labelled bovine serum albumin was used as negative control for brain uptake. Both radiolabeled DKPs scaffolds and 99m Tc-DKP-VQIVYK showed a high stability, while peptide 99m Tc-HVQIVYK alone was quickly degraded in vivo. The presence of 99m Tc-DKPs scaffolds and 99m Tc-DKP-VQIVYK was observed in the ventricular and subarachnoid spaces and to a lower extent in the brain parenchyma up to 45 minutes post-injection in mice. This work highlights the potentiality of DKP scaffolds as vectors to transport peptides into the brain by limiting proteolysis and favoring cerebral bioavailability.

Response of colonic motility to dopaminergic stimulation is subverted in rats with nigrostriatal lesion: relevance to gastrointestinal dysfunctions in Parkinson's disease.

BACKGROUND: Constipation is extremely common in patients with Parkinson's disease (PD) and has been described in PD animal models. In this study, we investigated whether a PD-like degeneration of dopaminergic neurons of the substantia nigra can influence peristalsis in colonic segments of rats by impacting on enteric dopaminergic transmission. METHODS: Male, Sprague-Dawley rats received a unilateral injection of neurotoxin 6-hydroxydopamine (6-OHDA), or saline, into the medial-forebrain-bundle. Peristaltic activity was recorded in isolated colonic segments, in baseline conditions and following exposure to combinations of D2 receptor (DRD2) agonist sumanirole and antagonist L-741626. Dopamine levels and DRD2 expression were assessed in the ileum and colon of animals. We also investigated the involvement of the dorsal motor nucleus of the vagus (DMV) - a potential relay station between central dopaminergic denervation and gastrointestinal (GI) dysfunction - by analyzing cytochrome c oxidase activity and FosB/DeltaFosB expression in DMV neurons. KEY RESULTS: We observed profound alterations in the response of colonic segments of 6-OHDA lesioned animals to DRD2 stimulation. In fact, the inhibition of colonic peristalsis elicited by sumanirole in control rats was absent in 6-OHDA-lesioned animals. These animals also showed reduced DRD2 expression in the colon, along with elevation of dopamine levels. No significant changes were detected within the DMV. CONCLUSIONS & INFERENCES: Our results demonstrate that selective lesion of the nigrostriatal dopaminergic pathway subverts the physiological response of the colon to dopaminergic stimulation, opening new perspectives in the comprehension and treatment of GI dysfunctions associated with PD.

Anionic fibroin-derived polypeptides accelerate MSC osteoblastic differentiation in a three-dimensional osteoid-like dense collagen niche.

Incorporation of anionic fibroin derived polypeptides into dense collagen gels provided a dynamic, three-dimensional, tissue-equivalent matrix together with biochemical cues that resembled the role of the bone morphogenic growth factors commonly used to promote osteogenic differentiation of mesenchymal stem cells.

Peripheral expression of key regulatory kinases in Alzheimer's disease and Parkinson's disease.

Alteration of key regulatory kinases may cause aberrant protein phosphorylation and aggregation in Alzheimer's disease (AD) and Parkinson's disease (PD). In this study, we investigated expression and phosphorylation status of glycogen synthase kinase 3 (GSK-3), protein kinase B (Akt) and tau protein in peripheral blood lymphocytes of 20 AD, 25 PD patients and 20 healthy controls. GSK-3 was increased in AD and PD patients. In these latter, GSK-3 levels were positively correlated with daily L-Dopa intake. Phosphorylated Akt expression was augmented in both groups; total Akt levels were increased only in AD patients and were positively correlated with disease duration and severity. Total and phosphorylated tau were increased only in AD, with phospho-tau levels being positively correlated with levels of total tau, Akt, and disease duration. No correlations between protein levels and clinical variables were found in PD patients. Investigation of peripheral changes in the expression of specific kinases may, therefore, lead to the development of innovative biomarkers of neurodegeneration, particularly for AD.

Pre-clinical and clinical evaluation of nuclear tracers for the molecular imaging of vulnerable atherosclerosis: an overview.

Cardiovascular diseases (CVD) are the leading cause of mortality worldwide. Despite major advances in the treatment of CVD, a high proportion of CVD victims die suddenly while being apparently healthy, the great majority of these accidents being due to the rupture or erosion of a vulnerable coronary atherosclerotic plaque. A non-invasive imaging methodology allowing the early detection of vulnerable atherosclerotic plaques in selected individuals prior to the occurrence of any symptom would therefore be of great public health benefit. Nuclear imaging could allow the identification of vulnerable patients by non-invasive in vivo scintigraphic imaging following administration of a radiolabeled tracer. The purpose of this review is to provide an overview of radiotracers that have been recently evaluated for the detection of vulnerable plaques together with the biological rationale that initiated their development. Radiotracers targeted at the inflammatory process seem particularly relevant and promising. Recently, macrophage targeting allowed the experimental in vivo detection of atherosclerosis using either SPECT or PET. A few tracers have also been evaluated clinically. Targeting of apoptosis and macrophage metabolism both allowed the imaging of vulnerable plaques in carotid vessels of patients. However, nuclear imaging of vulnerable plaques at the level of coronary arteries remains challenging, mostly because of their small size and their vicinity with unbound circulating tracer. The experimental and pilot clinical studies reviewed in the present paper represent a fundamental step prior to the evaluation of the efficacy of any selected tracer for the early, non-invasive detection of vulnerable patients.

Morphometric study of the interproximal unit in the esthetic region to correlate anatomic variables affecting the aspect of soft tissue embrasure space.

BACKGROUND: The presence of a normal papilla is crucial to avoid the unpleasant esthetic defects that are of major concern to periodontists, restorative dentists, and patients. During the course of progressive periodontitis and following periodontal treatment, it is not uncommon to have a partial loss of the interdental papilla. This loss can lead to an unesthetic gingival appearance. This study evaluated different anatomic variables in an effort to determine their role in the papillary appearance of maxillary incisors. METHODS: A total of 178 interdental embrasures in 58 patients were selected randomly for examination. For each patient, a digital photograph and a modified periapical radiograph of the interdental embrasure of the four maxillary incisors were taken by using a special metric device fixed to a centrator as a reference marker. Clinical and radiographic data were obtained for the distance from the contact point to the alveolar crest and for the interradicular distance. We used a classification system with regard to peri-implant soft tissue based on esthetic assessments related to the space between reference lines through the highest gingival curvature of the crown-tooth margin and the contact point. RESULTS: In the group of interdental sites with an interradicular distance of less than approximately 2.4 mm, an increase in the distance between the contact point and the bone crest corresponded to a marked increase in the interdental black triangle's dimensions and, therefore, a less esthetic smile. In particular, when the interradicular distance was >2.4 mm, we statistically estimated that the other anatomic variable considered, the distance from the contact point to the alveolar crest, lost its influence on whether the interdental papilla would be present or absent. CONCLUSION: The interradicular distance and the distance between the contact point and the alveolar crest have independent and combined effects on the presence or absence of the interdental papilla.

Lercanidipine, enalapril and their combination in the treatment of elderly hypertensive patients: placebo-controlled, randomized, crossover study with four ABPM.

This double-blind, placebo-controlled, four-way balanced design crossover study included hypertensive patients aged 60-85 years with mean office-measured sitting systolic blood pressure (SBP) 160-179 mm Hg and daytime SBP > or =135 mm Hg. After a 2-week run-in period, during which previous medications were discontinued, each patient received the following four treatments in randomized order for 4 weeks each: lercanidipine 10 mg (L), enalapril 20 mg (E), lercanidipine 10 mg plus enalapril 20 mg (L/E) and placebo (P). At the end of each treatment period, office trough blood pressure (BP) was measured and a 24-h Ambulatory Blood Pressure Monitoring (ABPM) was performed. Seventy-five patients (mean age 66 years, office BP 168/92 mm Hg, daytime SBP 151 mm Hg) were randomized and 62 completed the study with four valid post-baseline ABPMs. The administration of P, L, E and L/E was associated with a mean 24-h SBP of 144, 137, 133 and 127 mm Hg, respectively. All active treatments significantly reduced the mean 24-h SBP in comparison with placebo, but L/E was significantly more effective than L and E alone. Similarly, office SBP was significantly more reduced with L/E (-16.9 mm Hg) than with L (-5.0 mm Hg) or E (-5.9 mm Hg). A BP <140/90 mm Hg was recorded in 18% of patients with L, 19% with E and 45% with L/E. Two patients on P and two on L/E were withdrawn from the study due to adverse events. In conclusion, combination therapy with L/E has additive antihypertensive effects on both ambulatory and office BP in elderly patients and is well tolerated.

Molecular imaging of vascular cell adhesion molecule-1 expression in experimental atherosclerotic plaques with radiolabelled B2702-p.

PURPOSE: VCAM-1 plays a major role in the chronic inflammatory processes present in vulnerable atherosclerotic plaques. The residues 75-84 (B2702-p) and 84-75/75-84 (B2702-rp) of the major histocompatibility complex-1 (MHC-1) molecule B2702 were previously shown to bind specifically to VCAM-1. We hypothesised that radiolabelled B2702-p and B2702-rp might have potential for the molecular imaging of vascular cell adhesion molecule-1 (VCAM-1) expression in atherosclerotic plaques. METHODS: Preliminary biodistribution studies indicated that 125I-B2702-rp was unsuitable for in vivo imaging owing to extremely high lung uptake. 123I- or 99mTc-labelled B2702-p was injected intravenously to Watanabe heritable hyperlipidaemic rabbits (WHHL, n=6) and control animals (n=6). After 180 min, aortas were harvested for ex vivo autoradiographic imaging, gamma-well counting, VCAM-1 immunohistology and Sudan IV lipid staining. RESULTS: Robust VCAM-1 immunostaining was observed in Sudan IV-positive and to a lesser extent in Sudan IV-negative areas of WHHL animals, whereas no expression was detected in control animals. Significant 2.9-fold and 1.9-fold increases in 123I-B2702-p and 99mTc-B2702-p aortic-to-blood ratios, respectively, were observed between WHHL and control animals (p<0.05). Tracer uptake on ex vivo images co-localised with atherosclerotic plaques. Image quantification indicated a graded increase in 123I-B2702-p and 99mTc-B2702-p activities from control to Sudan IV-negative and to Sudan IV-positive areas, consistent with the observed pattern of VCAM-1 expression. Sudan IV-positive to control area tracer activity ratios were 17.0+/-9.0 and 5.9+/-1.8 for 123I-B2702-p and 99mTc-B2702-p, respectively. CONCLUSION: Radiolabelled B2702-p is a potentially useful radiotracer for the molecular imaging of VCAM-1 in atherosclerosis.

Assessment of non-reperfused and reperfused myocardial infarction using diffusible or deposited radiolabelled perfusion imaging agents.

PURPOSE: Incomplete microvascular reperfusion is often observed in patients undergoing thrombolytic therapy or angioplasty for acute myocardial infarction and has important prognostic implications. We compared the myocardial uptake of diffusible ((201)Tl) and deposited ((99m)TcN-NOET) perfusion imaging agents in the setting of experimental infarction. METHODS: Rats were subjected to permanent coronary occlusion (OCC, n=10) or to 45-min occlusion and reperfusion (REP, n=17). Seven days later, the tracers were co-injected and the animals were euthanised 15 min (all ten rats in the OCC group and 12 rats in the REP group) or 120 min (five rats from the REP group, euthanised at this time point to evaluate any redistribution of the tracers: REP-RED group) afterwards. Infarct size determination and (99m)TcN-NOET/(201)Tl ex vivo imaging were performed. Regional flow and tissue oedema were quantified using radioactive microspheres and (99m)Tc-DTPA, respectively. RESULTS: (99m)TcN-NOET and (201)Tl defect magnitudes were similar in OCC animals (0.11+/-0.01 vs 0.13+/-0.01). In REP animals, (201)Tl defect magnitude (0.25+/-0.02) was significantly lower than the magnitude of (99m)TcN-NOET and flow defects (0.14+/-0.03 and 0.17+/-0.01, respectively; p<0.05), despite the lack of (201)Tl redistribution (REP-RED animals). (99m)Tc-DTPA indicated the presence of oedema in the reperfused area. Blood distribution studies showed that, unlike (99m)TcN-NOET, (201)Tl plasma activity was mostly unbound to plasma proteins. CONCLUSION: (99m)TcN-NOET and (201)Tl delineated the non-viable area in chronic non-reperfused and reperfused myocardial infarction. The significantly decreased (201)Tl defect in reperfused infarction was likely due to partial diffusion of the tracer from the plasma into the oedema present in the infarcted area. Deposited perfusion tracers might be better suited than diffusible agents for the assessment of regional flow following reperfusion of myocardial infarction.

Triage: experiencia en un servicio de urgencias pediátricas / Triage: experience in a pediatric emergency service

Identificar rápidamente a los pacientes con urgencias o condiciones que ponen en peligro la vida, determinar el área de tratamiento más apropiado, disminuir la congestión en las áreas de la sala de urgencias, proveer los mecanismos para la evaluación permanente de los pacientes, proveer información a los pacientes y sus familiares en relacióna los cuidados y al tiempo de espera y aportar información exacta sobre la precisión del trabajo en urgencias.

Triage: experiencia en un servicio de Urgencias Pediátricas / Triage: experience in a pediatric emergency service

Objetivo: Evaluar el resultado de la aplicacón de un sistmea triage en el servicio de urgencias pediátricas del Hospital Central del Instituto de Previsión Social (HCIPS). Diseño y Metodología: Estudio observacional descriptivo, propectivo, sobre 1122 niños que consultaron en julio de 2005, de 07:00 a 16:00 horas de lunes a viernes. Los datos asientan en el sistema informático y fueron tabulados mediante el programa Microsoff Excel. Se elaboró un listado de patologías y signos y/o síntomas a considerarse como urgente, por gravedad y riesgo de compromiso vital, número de pacientes y frecuencia de su presentación en este servicio. Se denominó prioridad uno (P1) a las que requería atención inmediata y prioridad dos (P2) al resto. Se marcó como tiempo de corte 30 minutos para evaluar la capacidad de atención a los pacientes de mayor gravedad. Resultados: Total de pacientes clasificados como P1: 272/1122 (24 porcientos), P2: 850/1122 (76 porcientos). Mediana de tiempo de espera fue para P1: 10 min, p25-p75 (5-13min) tiempo máximo para la atención fue 65 min. Para P2 la mediana fue 65 min. p25-p75 (20-139min) tiempo máximo para la atención 300 min; la proporción atendida dentro de los primeros 30 minutos fue P1: 79 porcientos; P2: 36 porcientos. Pacientes hospitalizados según prioridad: P1: 25/272 (9 porcientos), ninguno del grupo de P2. Conclusión: La implementación de un sistema de triage, en el servicio de urgencias pediátricas del HCIPS, permitió conocer que solo una cuarta parte de los pacientes que acuden a la consulta fueron clasificados como urgencias, lo que hace presumir, por la similitud de los motivos de consulta, que gran mayoría de los mismos, representan a aquellos que no pudieron ser absorbidos por los consultorios externos. Demostró que más de las tres cuartas partes de las urgencias fueron atendidas dentro de los treinta minutos de su adminsión

Assessment of insulin sensitivity in vivo in control and diabetic mice with a radioactive tracer of glucose transport: [125I]-6-deoxy-6-iodo-D-glucose.

BACKGROUND: Impairment of insulin-stimulated glucose transport is a characteristic of type 2 diabetes. A radioactive glucose analogue has been synthesized: [(125)I]-6-deoxy-6-iodo-D-glucose. Its biological behaviour in vitro is similar to that of 3-O-methyl-D-glucose, the reference tracer of glucose transport. The aim of the present study was to determine the ability of [(125)I]-6-deoxy-6-iodo-D-glucose to evaluate variations in glucose transport in vivo. METHODS: Biodistributions of [(125)I]-6-deoxy-6-iodo-D-glucose were performed with or without exogenous insulin (iv injection of 1.5 IU/kg) in db/+ non-diabetic control mice and in db/db type 2 diabetic mice, exhibiting a severe insulin resistance characterized by a lack of increase in glucose uptake in response to insulin. RESULTS: In db/+ mice, insulin increased [(125)I]-6-deoxy-6-iodo-D-glucose transport by 30% in most insulin-sensitive tissues (heart, diaphragm and skeletal muscle, p < 0.05) and had no effect in other organs. In db/db mice, [(125)I]-6-deoxy-6-iodo-D-glucose transport in these organs was not modified by insulin. CONCLUSION: [(125)I]-6-deoxy-6-iodo-D-glucose is able to trace in vivo an increase in glucose transport with insulin in non-diabetic mice and a defect of glucose transport in type 2 diabetic mice. It is the first time that an iodinated analogue of glucose has shown such promising results after in vivo injection. The use of this tracer to assess glucose transport in vivo in humans via nuclear imaging warrants further investigation.

Influence of calcium channel inhibitors on the myocardial uptake and retention of technetium 99m N-NOET, a new myocardial perfusion imaging agent: a study on isolated perfused rat hearts.

BACKGROUND: Technetium 99m N-NOET is a new myocardial perfusion imaging agent currently in phase III clinical trials in Europe. In vitro studies on newborn rat cardiomyocytes have shown that calcium inhibitors, such as verapamil or diltiazem, inhibit its cellular uptake by 40%. To determine whether such a specificity exists ex vivo, we studied the effect of verapamil, diltiazem, and nifedipine on the myocardial uptake and retention of Tc-99m N-NOET in isolated perfused rat hearts. METHODS AND RESULTS: After a 15-minute baseline period, rat hearts were perfused with 0.5 micromol/L verapamil (n = 6), 0.75 micromol/L diltiazem (n = 6), or 0.1 micromol/L nifedipine (n = 6) for 10 minutes before the injection of a bolus (40 microCi/250 microL) of the tracer. Control hearts were perfused with either 1.5 mmol/L calcium (same concentration as in the treated groups; n = 7) or 0.75 mmol/L calcium (same contractility as in the treated groups; n = 6). Myocardial activity of Tc-99m N-NOET was monitored for 30 minutes. The functional parameters of the hearts were recorded throughout the experiments. Calcium inhibitors induced a 40% to 55% decrease in maximal first derivative of left ventricular pressure (dP/dt) (0.0001

Cellular uptake mechanisms of 99mTcN-NOET in cardiomyocytes from newborn rats: calcium channel interaction.

BACKGROUND: Bis[N-ethoxy,N-ethyl(dithiocarbamato)]nitrido Tc (V) (TcN-NOET) is a new technetium complex proposed as a tracer of myocardial perfusion. However, its cellular uptake mechanisms are unknown, although membrane localization on rat heart preparations and preferential binding to polymorphonuclear neutrophils (PMNs) have been reported. Because of the central role of calcium in PMN actions, a relationship was hypothesized between this ion flux and TcN-NOET cellular uptake. METHODS AND RESULTS: The mechanisms of cellular uptake of TcN-NOET were investigated in newborn rat cardiomyocytes by study of the effect of calcium channel modulators on tracer binding. Nifedipine had no effect on tracer uptake at 1 minute. However, verapamil 0.1 micromol/L and diltiazem 0.5 micromol/L induced a 40% decrease in uptake. Conversely, Bay K 8644 0.25 micromol/L increased TcN-NOET uptake by 73%. Alterations in other membrane ion transports failed to modify tracer uptake, indicating the specificity of the relationship between TcN-NOET uptake and calcium channels. Kinetic studies indicated that cellular net accumulation of the tracer was slow (t1/2=28.5 minutes) and retention was prolonged (84% of initial activity retained after 120 minutes of washout). The energy dependence of TcN-NOET uptake was investigated after 60 minutes of metabolic inhibition by iodoacetic acid plus rotenone. The ATP decrease was not associated with reduction in tracer uptake at 1 minute (114.9+/-21.9% of control, P=NS). CONCLUSIONS: The decrease in uptake observed with verapamil and diltiazem, the increase with Bay K 8644, and the lack of effect with nifedipine suggest that TcN-NOET binds to L-type calcium channels in the open configuration, without entering cardiomyocytes. The kinetics of TcN-NOET accumulation and retention are slow, and the mechanism for cellular uptake is not energy-dependent. From a clinical point of view, the effect of concurrent treatment by calcium inhibitors on myocardial binding of TcN-NOET should be taken into account.