A. Gomez

Documento «Sevilla» de Consenso sobre Alternativas a la Transfusión de Sangre Alogénica

El Documento de Consenso sobre Alternativas a la Transfusion de Sangre Alogenica (ATSA) ha sido elaborado por un panel de expertos pertenecientes a 5 sociedades cientificas. Han participado y patrocinado las sociedades espanolas de Anestesiologia (SEDAR), Medicina Intensiva (SEMICYUC), Hematologia y Hemoterapia (AEHH), Transfusion sanguinea (SETS) y Trombosis y Hemostasia (SETH). Las alternativas a la transfusion se han clasificado en farmacologicas y no farmacologicas, con un total de 4 modulos y 12 topicos. La disminucion de las transfusiones de sangre alogenica y/o el numero de pacientes transfundidos fue la principal variable objetivo. El grado de cumplimiento de este objetivo, para cada ATSA, se llevo a cabo siguiendo la metodologia Delphi, que clasifica el grado de recomendacion desde «A» (apoyado por estudios controlados) hasta «E» (estudios no controlados y opinion de expertos). Los expertos concluyeron que la mayor parte de las indicaciones de las ATSA se sustentan en grados de recomendacion medios y bajos, «C», «D» o «E», precisandose nuevos estudios controlados.

Characterization of muscarinic receptor subtypes in human tissues

The affinities of selective, pirenzepine and AF-DX 116, and classical, N-methylscopolamine and atropine, muscarinic cholinergic receptor antagonists were investigated in displacement binding experiments with [3H]Pirenzepine and [3H]N-methylscopolamine in membranes from human autoptic tissues (forebrain, cerebellum, atria, ventricle and submaxillary salivary glands). Affinity estimates of N-methylscopolamine and atropine indicated a non-selective profile. Pirenzepine showed differentiation between the M1 neuronal receptor of the forebrain and the receptors in other tissues while AF-DX 116 clearly discriminated between muscarinic receptors of heart and glands. The results in human tissues confirm the previously described selectivity profiles of pirenzepine and AF-DX 116 in rat tissues. These findings thus reveal the presence also in man of three distinct muscarinic receptor subtypes: the neuronal M1, the cardiac M2 and the glandular M3.

Muscarinic receptor subtypes in human and rat colon smooth muscle

Muscarinic receptor subtypes in human and rat colon smooth muscle homogenates were characterized with [3H]N-methylscopolamine ([3H]NMS) by ligand binding studies. [3H]NMS saturation experiments show the existence of a homogeneous population of non-interacting binding sites with similar affinity (KD values of 1.38 +/- 0.20 nM in human colon smooth muscle and 1.48 +/- 0.47 nM in rat colon smooth muscle) and with Hill slopes close to unity in both samples of tissue. However, a significant (P less than 0.01) increase in muscarinic receptor density (Bmax) is found in human colon (29.9 +/- 2.9 fmol/mg protein) compared with rat colon (17.2 +/- 1.5 fmol/mg protein). Inhibition of [3H]NMS binding by non-labelled compounds shows the following order in human colon: atropine greater than AF-DX 116 greater than pirenzepine. Whereas in rat colon the rank order obtained is atropine greater than pirenzepine greater than AF-DX 116. Atropine and pirenzepine bind to a homogeneous population of binding sites, although pirenzepine shows higher affinity to bind to the sites present in rat colon (Ki = 1.08 +/- 0.08 microM) than those in human colon (Ki = 1.74 +/- 0.02 microM) (P less than 0.05). Similarly, IC50 values obtained in AF-DX 116 competition experiments were significantly different (P less than 0.01) in human colon (IC50 = 1.69 +/- 0.37 microM) than in rat colon (IC50 = 3.78 +/- 0.75 microM). Unlike atropine and pirenzepine, the inhibition of [3H]NMS binding by AF-DX 116 did not yield a simple mass-action binding curve (nH less than 1, P less than 0.01) suggesting the presence of more than one subtype of muscarinic receptor in both species. Computer analysis of these curves with a two binding site model suggests the presence of two populations of receptor. The apparent Ki1 value for the high affinity binding site is 0.49 +/- 0.07 microM for human colon smooth muscle and 0.33 +/- 0.05 microM for rat colon smooth muscle. The apparent Ki2 for the low affinity binding site is 8.01 +/- 1.0 microM for human samples and 6.07 +/- 1.1 microM for rat samples. These values are close enough to suggest that the first subtype of muscarinic receptor may be considered cardiac (M2) and the second subtype glandular (M3). The relative densities of the receptor subtypes are significantly different for both species. Human colon samples show the major densities of subtype M2, 22.62 +/- 1.11 fmol/mg protein, this represents 75.66 +/- 3.73% of the total receptors.(ABSTRACT TRUNCATED AT 400 WORDS)

Impact of Postoperative Unwashed Shed Blood Retrieved after Total Knee Arthroplasty on Endotoxin-stimulated Tumor Necrosis Factor α Release In Vitro

Background:Allogeneic or autologous blood seems to have an immunosuppressive effect that is largely attributable to storage-dependent factors. However, transfusion of postoperative unwashed shed blood (USB) after elective total knee replacement does not undergo storage. Therefore, the authors explored the effects of USB on the mitogen-driven cytokine synthesis by the patient’s peripheral blood mononuclear cells. Methods:Perioperative blood samples were obtained from 12 total knee replacement patients with and 5 without reinfusion of leukoreduced USB, and from USB reinfusion line, before and after leukoreduction. Venous blood obtained at 4–6 postoperative hours was coincubated with USB. Endotoxin-stimulated release of tumor necrosis factor &agr; and interleukin 10 was measured after 24 h of culture by solid-phase enzyme-labeled chemiluminescent immunometric assay. Results:Coincubation of postoperative venous blood with USB, USB cells, or USB plasma resulted in a significant depression of tumor necrosis factor-&agr; synthesis, without significant effects on interleukin-10 synthesis. However, no differences were observed for endotoxin-stimulated cytokine release in perioperative blood samples from patients receiving or not receiving USB. Conclusion:These data suggest that USB seemed to contain an antiinflammatory agent. However, at the actual retransfusion rate, USB does not seem to further enhance the immunosuppression that follows knee replacement surgery.

Characterization of muscarinic receptors in human submandibular salivary glands

We studied the binding of the muscarinic antagonist [3H]N-methyl-scopolamine [( 3H]NMS) in order to characterize muscarinic receptors located in human submandibular salivary glands obtained from intrasurgical biopsy. [3H]NMS bound with a Kd value of 1.56 nM to a single class of muscarinic receptors (Bmax 37.3 fmol/mg protein) since pirenzepine exhibited a homogeneous binding profile.

Circadian rhythm in muscarinic receptor subtypes in rat forebrain

The binding of different ligands to muscarinic receptors in the central nervous system is regulated by several factors. Among these are the administration of drugs, disease, ontogeny or aging. Studies carried out in rat brains have demonstrated changes in the density of the muscarinic receptors at different times of the day. These changes might be related to variations in the circadian rhythms. In this work we have studied the binding of the [3H]-N-methyl-escopolamine, the agonist carbachol and the antagonist pirenzepine to muscarinic receptors in rat forebrains at 10.00, 14.00, 18.00, 22.00, 02.00 and 06.00 hr. We have observed changes in the density of muscarinic receptors but not changes in affinity to the radioligand. The Bmax values obtained by saturation studies were maximum at 14.00 hr and minimum at 02.00 hr (P less than 0.05 Mann-Whitneys test). Inhibition studies in the presence of the non-selective agonist carbachol and the selective antagonist pirenzepine, at the same time-points, did not show statistically significant changes in the Bmax values. These data indicate that changes in the Bmax values are only observed in the total population of muscarinic receptors and are not due to modifications in the subtypes of muscarinic receptors nor to the different affinity states of agonist binding.