The effect of neurotrophic factors on morphology, TRPV1 expression and capsaicin responses of cultured human DRG sensory neurons.

We have studied the effect of key neurotrophic factors (NTFs) on morphology, levels of the vanilloid receptor-1 (TRPV1) and responses to capsaicin in adult human sensory neurons in vitro. Avulsed dorsal root ganglia (DRG, n = 5) were cultured with or without a combination of nerve growth factor (NGF), glial cell (line)-derived growth factor (GDNF) and neurotrophin3 (NT3) for 5 days. In the absence of NTFs, the diameter of neurons ranged from 20 to 100 microm (mean 42 +/- 4 microm). Adding NTFs caused a significant increase in neuronal sizes, up to 120 microm (mean diameter 62 +/- 5 microm, P < 0.01, t-test), an overall 35% increase of TRPV1-positive neurons (P < 0.003), and notably of large TRPV1-positive neurons > 80 microm (P < 0.05). Responses to capsaicin were significantly enhanced with calcium ratiometry (P < 0.0001). Short duration (1h) exposure of dissociated sensory neurons to NTFs increased numbers of TRPV1-positive neurons, but not of TRPV3, Nav 1.8 and IK1 and the morphological size-distribution remained similar to intact post-mortem DRG neurons. NTFs thus increase size, elevate TRPV1 levels and enhance capsaicin responses in cultured human DRG neurons; these changes may relate to pathophysiology in disease states, and provide an in vitro model to study novel analgesics.

Identification of amino acid residues responsible for the alpha5 subunit binding selectivity of L-655,708, a benzodiazepine binding site ligand at the GABA(A) receptor.

L-655,708 is a ligand for the benzodiazepine site of the gamma-aminobutyric acid type A (GABA(A)) receptor that exhibits a 100-fold higher affinity for alpha5-containing receptors compared with alpha1-containing receptors. Molecular biology approaches have been used to determine which residues in the alpha5 subunit are responsible for this selectivity. Two amino acids have been identified, alpha5Thr208 and alpha5Ile215, each of which individually confer approximately 10-fold binding selectivity for the ligand and which together account for the 100-fold higher affinity of this ligand at alpha5-containing receptors. L-655,708 is a partial inverse agonist at the GABA(A) receptor which exhibited no functional selectivity between alpha1- and alpha5-containing receptors and showed no change in efficacy at receptors containing alpha1 subunits where amino acids at both of the sites had been altered to their alpha5 counterparts (alpha1Ser205-Thr,Val212-Ile). In addition to determining the binding selectivity of L-655,708, these amino acid residues also influence the binding affinities of a number of other benzodiazepine (BZ) site ligands. They are thus important elements of the BZ site of the GABA(A) receptor, and further delineate a region just N-terminal to the first transmembrane domain of the receptor alpha subunit that contributes to this binding site.

Serum neopterin and soluble interleukin-2 receptor for prediction of a shock state in gram-negative sepsis.

PURPOSE: This study aimed to investigate the predictive value of neopterin and soluble interleukin-2 (IL-2) receptor for shock occurrence in gram-negative sepsis. METHODS: We examined 57 patients admitted to an intensive care unit with gram-negative sepsis diagnosed according to preestablished criteria. Blood samples were collected every 24 hours and neopterin and soluble IL-2 receptor were measured by using commercially available test kits. To judge the predictive significance of these analyses the Cox proportional hazards regression model was used. RESULTS: Both neopterin (P < .05) and soluble IL-2 receptor (P < .01) were identified as significant predictors of a shock state, but the prognostic strength of neopterin exceeded that of soluble IL-2 receptor. To further assess if other factors could interfere with the predictive significance of both compounds, we also investigated other clinical and laboratory variables but these candidate predictors did not contribute any additional significant predictive information. CONCLUSION: The measurement of serum neopterin and soluble IL-2 receptor concentrations has predictability for identifying patients with gram-negative sepsis at risk for progression toward the syndrome of septic shock.

[Neopterin and interleukin 2 soluble receptors as biochemical markers of cellular immune response to surgical trauma]. / Neopterina e recettore solubile dell'interleuchina 2 come indicatori biochimici della risposta immunitaria cellulare al trauma chirurgico.

In 67 patients submitted to surgical procedures serum neopterin (NPT) and Interleukin 2 soluble receptors (IL2R) were evaluated at the end of the operation as well as 24, 48, 72 hours later. Thirty seven of the subjects (Group B) had undergone minor surgery (average time of operation: 40 +/- 10 min.), thirty (Group A) had undergone major surgery (average time: 180 +/- 30 min.). The results showed elevated NPT and IL2R levels in the latter cases and, in particular, 48 and 72 h after surgery. Neopterin levels were positively correlated with IL2R (r = 0.548 p < 0.01). These data suggest an activation of the cellular immune response which parallels the magnitude and length of surgical trauma. Thus NPT and IL2R levels could represent biochemical markers of postoperative disorders of the immune homeostasis.