Allosteric modulators of solute carrier function: a theoretical framework.

Large-scale drug screening is currently the basis for the identification of new chemical entities. This is a rather laborious approach, because a large number of compounds must be tested to cover the chemical space in an unbiased fashion. However, the structures of targetable proteins have become increasingly available. Thus, a new era has arguably been ushered in with the advent of methods, which allow for structure-based docking campaigns (i.e., virtual screens). Solute carriers (SLCs) are among the most promising drug targets. This claim is substantiated by the fact that a large fraction of the 400 solute carrier genes is associated with human diseases. The ability to dock large ligand libraries into selected structures of solute carriers has set the stage for rational drug design. In the present study, we show that these structure-based approaches can be refined by taking into account how solute carriers operate. We specifically address the feasibility of targeting solute carriers with allosteric modulators, because their actions differ fundamentally from those of ligands, which bind to the substrate binding site. For the pertinent analysis we used transition state theory in conjunction with the linear free energy relationship (LFER). These provide the theoretical framework to understand how allosteric modulators affect solute carrier function.

The selectivity filter of the voltage-gated sodium channel is involved in channel activation.

Amino acids located in the outer vestibule of the voltage-gated Na+ channel determine the permeation properties of the channel. Recently, residues lining the outer pore have also been implicated in channel gating. The domain (D) IV P-loop residue alanine 1529 forms a part of the putative selectivity filter of the adult rat skeletal muscle (mu1) Na+ channel. Here we report that replacement of alanine 1529 by aspartic acid enhances entry to an ultra-slow inactivated state. Ultra-slow inactivation is characterized by recovery time constants on the order of approximately 100 s from prolonged depolarizations and by the fact that entry to this state can be reduced by binding to the pore of a mutant mu-conotoxin GIIIA, suggesting that ultra-slow inactivation may reflect a structural rearrangement of the outer vestibule. The voltage dependence of ultra-slow inactivation in DIV-A1529D is U-shaped, with a local maximum near -60 mV, whereas activation is maximal only above -20 mV. Furthermore, a train of brief depolarizations produces more ultra-slow inactivation than a single maintained depolarization of the same duration. These data suggest that ultra-slow inactivation emanates from "partially activated" closed states and that the P-loop in DIV may undergo a conformational change during channel activation, which is accentuated by DIV-A1529D.

Inflammatory mediators in relation to the development of multiple organ failure in patients after severe blunt trauma.

OBJECTIVE: To evaluate the posttraumatic course of several inflammatory mediators or markers (complement components C3, C3a, terminal complement complex, thromboxane B2, C-reactive protein, elastase, and neopterin) in relation to the development of multiple organ failure and mortality. DESIGN: Prospective study of a selected patient group. SETTING: Surgical intensive care units in three European trauma hospitals. PATIENTS: Patients (n = 56) with severe blunt trauma (Injury Severity Score of > or = 33). INTERVENTIONS: Arterial blood samples were sequentially obtained. MEASUREMENTS AND MAIN RESULTS: Nonsurvivors (n = 8) had significantly higher circulating C3a and elastase concentrations on the first postinjury day, compared with survivors (n = 48). No differences between these groups were found for terminal complement complex, thromboxane B2, C-reactive protein, and the neopterin/creatinine ratio. Five patients died before day 5. Eighteen patients developed multiple organ failure, which was diagnosed from day 5 onward, leaving 33 patients without multiple organ failure. The patients with subsequent multiple organ failure showed significantly higher mean circulating concentrations of C3a (914 +/- 190 [SEM] ng/mL), terminal complement complex (57 +/- 17 U/mL), and thromboxane B2 (275 +/- 37 pg/mL) at the first postinjury day than the patients without multiple organ failure (566 +/- 110 ng/mL, 27 +/- 2 U/mL, and 169 +/- 14 pg/mL, respectively). In patients with multiple organ failure, elastase concentrations were significantly higher on days 2, 3, 4, and 5 postinjury. Neopterin/creatinine ratios, on the other hand, were significantly higher in patients with multiple organ failure when the multiple organ failure had already become established (on days 8 and 10). CONCLUSION: In multiple trauma patients, excessive triggering of the inflammatory cascade-as expressed by complement activation and stimulation of neutrophils producing elastase--plays an important and early role in the development of multiple organ failure.

Scoring systems and blood lactate concentrations in relation to the development of adult respiratory distress syndrome and multiple organ failure in severely traumatized patients.

In 56 patients with multiple trauma with ISSs > or = 33 we prospectively collected data of seven scoring systems (ISS, TS, TRISS, GCS, PTS, APACHE II, SSS) and sequentially determined blood lactate concentrations. These data were analyzed in relation to the patients later developing adult respiratory distress syndrome (ARDS) and multiple organ failure (MOF). Twenty-two patients developed ARDS, and 18 developed MOF. Of the mentioned scoring systems only ISS, PTS, and SSS were predictive of subsequent ARDS, and only ISS and SSS were predictive of subsequent MOF. Lactate concentrations at days 2, 3, and 4 were significantly different between patients with and without subsequent ARDS, MOF, or both. Surprisingly, APACHE II scores did not correlate with subsequent ARDS or MOF, nor did they show any significant relation with lactate concentrations at any time. By stepwise regression analysis ISS, SSS, and lactate level at day 3 were the most significant variables toward the development of ARDS and MOF. It is concluded that scoring systems directly grading the severity of groups of trauma patients have predictive value for late and remote complications such as ARDS and MOF, whereas scoring systems that grade the physiologic response to trauma--although clearly related to mortality--have no such predictive value.

[Catheter epidural analgesia in serial rib fractures]. / Katheterepiduralanalgesie bei Serienrippenfrakturen.

According to our results, permanent epidural anaesthesia was significantly superior to systemic opioid treatment in patients with serial rib fractures. The main advantages were not only continuous pain relief despite the fact that the nonepidural control group required more than twice the dosage of morphine derivatives; also, the respiratory and pain-related recovery time was reduced. Another advantage was the selective effect (due to the local application) on respiratory pain and therefore on respiration as a whole. Deep breathing and expectoration were easier, so that the use of respirators and other artificial breathing aids could be avoided or at least reduced in duration in some cases. This makes the method particularly suitable for use in the management of polytraumatized patients. The standard dose was a mixture of 3.3 mg morphine and 37.5 mg bupivacaine (= 1/3 ampoule morphine + 15 ml Carbostesin 0.25%) every 12 h. When morphine was temporary contraindicated (frequently the final diagnosis in the case of an "acute abdomen" delayed the administration of morphine) the use of bupivacaine alone provided a satisfactory result for a certain time (we never observed tachyphylaxis). Additional systemic pain relievers were only necessary when the patient was suffering from pain caused by other injuries beyond the area of effectiveness of the epidural catheter (the only obvious disadvantage of the local application technique). On the other hand, epidural anaesthesia enabled us to treat a patient's lower-leg fracture by interlocking nailing, while adding only 0.01 mg fentanyl (= 2 ml Fentanyl Janssen) and 1.2 mg flunitrazepam (Rohypnol).

Plasma fibronectin, albumin, IgM and total protein during cardiopulmonary bypass.

Plasma concentrations of fibronectin, albumin, total protein and IgM were measured in 14 male patients undergoing aorto-coronary bypass operations. Fibronectin and IgM concentrations fell to 55% of the preoperative values 5 minutes after start of the extracorporeal circulation, and the same percentages were encountered 5 minutes after the termination of bypass. The concentrations had recovered to 75% of the preoperative values by the end of the operation. The plasma concentration of albumin was 68 +/- 9% of the preoperative value after 5 minutes of bypass, 66 +/- 7% at the end of bypass, and 83 +/- 9% of the end of operation (significantly different from fibronectin and IgM, p less than 0.05; Wilcoxon's test for paired differences). No correlation was found between the duration of extracorporeal circulation and the post-bypass concentrations of any protein (Kendall correlation). It is concluded that the fall of fibronectin concentrations during cardiopulmonary bypass can be sufficiently explained by dilution, and that a specific consumption of fibronectin does not occur. The less marked decreases of albumin and total protein were probably due to infusion of plasma-protein solution, which is poor in fibronectin and IgM.

[Perioperative cefamandole prevention in aortocoronary bypass operations: the course of serum levels in extracorporeal circulation]. / Perioperative Cefamandolprophylaxe bei aortocoronaren Bypass-Operationen: Serumkonzentrationsverlauf während der extrakorporalen Zirkulation.

Antibiotic prophylaxis with 2 g Cefamandole at induction of anaesthesia was performed in 12 male patients undergoing aortocoronary bypass surgery. Caused by hemodilution, there was a marked decrease of serum concentration at the beginning of extracorporeal circulation, from 110.96 +/- 40.29 mcg/ml to 70.89 +/- 34.65 mcg/ml within 10 min. During extracorporeal circulation, elimination was as fast as before and after perfusion. 240 min after application, mean serum concentrations of 16.80 +/- 9.32 mcg/ml were measured. Failure of antibiotic prophylaxis in operations exceeding 4 h might be due to unadaequate antibiotic concentrations, beyond the minimal inhibitory concentration for the pathogens, reported to cause infections after cardiac operations.