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In this study the authors examined the sequences of the ribosomal 18S rRNA of Drosophila and man and 16 mRNA sequences coding for different members of the family of the mammalian formyl peptide receptors (FPRs). The positions in the sequences of all >or=7-base oligonucleotide identities occurring in at least one of the 18S rRNAs and one of the FPR mRNAs were recorded. On the basis of the positional data, the Drosophila 18S-FPR and human 18S-FPR distances (in nucleotides) were determined for each identity. Then the actual frequency distribution of the distances (grouped into 200-unit classes) was derived. The theoretical frequency distribution of distances was also calculated under the assumption of non-relatedness between the 18S and FPR sequences. Comparison between the theoretical and the actual distributions showed that at class -500 (range from - 400 to - 600) of the 18S-FPR values the actual frequency was significantly (p < 0.01) higher than the theoretical frequency, in both Drosophila and man, suggesting that the second section of the FPR genes (approximately from nucleotide 400 to the end of sequence) may be structurally related to the first section of the ribosomal 18S genes (approximately nucelotides 1-650). The authors advance the hypothesis that the two families of genes may have used common ancestral raw genetic materials in the building of the extant sequences.

Formyl peptides (FPs) released by some bacteria are powerful chemoattractants and activators of granulocytes, monocytes, and macrophages, acting through the members of a subfamily of specific seven-transmembrane G-protein-coupled formyl peptide receptors (FPRs), which are expressed only in mammals. Upon stimulation, granulocytes chemotactically move towards sites of maximal FP concentration, and release different bactericidal lytic enzymes and reactive oxygen species (ROI). In some instances, such as ischemia/reperfusion, the proinflammatory mediators released by the injured tissues and the intestinal bacteria and endotoxins, which may permeate across the damaged mucosal barrier, prime the inflowing granulocytes for an enhanced ROI production, resulting in severe damage to the host tissues. In this investigation 16 representative FPR and FPR-like mRNAs were selected to study the pattern of mutation/conservation of the individual nucleotides (nt) in the coding sequences. Mutations occur in 56.7%, 46.4%, and 87.5 % of cases in the first, second, and third nt, respectively, of the coding triplets. A probabilistic analysis demonstrated a significant nonrandom linkage between mutations in the first and second nt. Furthermore, the triplets that are variously double-mutated in the first two nt code, on average, for more hydrophobic amino acids (AA) in the transmembrane segments and more hydrophilic AA in the external and intracytoplasmic segments, thus preserving the general structure of the receptor. The authors hypothesize that when in one of the first two nt a mutation leading to a nonfunctioning protein product occurred, the mutated gene was eventually eliminated; however, a second mutation occurring in the other previously unmutated nt may have led to a protein product that is compatible with functional activity, although mutated in one (noncritical) AA. Such double mutations effecting a "functional repair" have thus survived and are retained among the extant sequences. Moreover, the combined mutation of all three nt in coding triplets occurs with a significantly higher than random frequency and this finding may be interpreted in a similar way.

Comparisons between the sequences of insect and vertebrate 18S rRNAs and the sequences of mammalian formyl peptide and some vertebrate chemokine receptor mRNAs demonstrated non-random structural similarities between these two groups of RNAs. It has been proposed that sections of the more ancient and conserved rRNA genes could have participated in the building of these more recent genes involved in immune responses. Here we analyze the sequence architecture of the 18S rRNA in insects (Drosophila simulans) and vertebrates (man), in terms of similarities between selected segments within the individual molecules. The insect and vertebrate 18S rRNAs are basically similar, but show specific insertions/deletions and base changes. In spite of these differences, in both sequences a significantly higher-than-expected (by random occurrence) number of 7-or-more-base oligonucleotide repeats was observed between segments roughly corresponding to nt 350-1050 and nt 1150-1850, with mutual between-repeats distances comprised in the range 700-900 nt. Based on this result we performed a multialignment of segments 317-1035 of Drosophila, 360-1005 of man, 1096-1864 of Drosophila, and 1066-1736 of man, the first two segments covering the region of first occurrence of the repeats and the last two the region of recurrences. At both ends of these segments the four sequences could be aligned with relatively minor gaps and the number of base identities in all four sequences was significantly higher than expected by random coincidences. These results support the hypothesis that an ancestral gene structure, composed of a chain of about 700 nt, duplicated to form a two-unit tandem repeat which still represents the most substantial part of the 18S rRNA molecule in extant insects and vertebrates.

Formyl peptides released from Gram-negative bacteria ligate a group of specific mammalian receptors, expressed mainly on granulocytes, monocytes, and macrophages. Receptor ligation activates different transduction cascades, eventually leading to the release of reactive oxygen species and other bactericidal chemical species, and the activation of the actin cytoskeleton with extension of lamellipodia and migration toward the sites of maximal formyl peptide concentration. In vitro, under conditions of nongradient formyl peptide concentrations, lamellipodia form all around the cell contour (chemokinesis). In granulocytes challenged under these conditions with N-formyl-methionyl-leucyl-phenylalanine, (i) the power spectrum of the contour of activated cells shows a peak at a specific periodicity, indicating that the lamellipodial extension is not completely random but stochastically conforms to a deterministic scheme, and (ii) the morphological response (percent of cells exhibiting chemokinesis) tends to reach a maximum at certain drug concentrations, then declining at higher concentrations. Accordingly, the logarithm of the drug concentration-polarizing effect curve is bell-shaped. Herein we illustrate theoretical models for the simulation of these two components of the chemokinetic responses. We show that the main traits of the general morphology and arrangement of lamellipodia may be simulated by an algorithm that starting from a situation of random distribution of active receptors on the cell membrane, encompasses in the successive calculation cycles both a local autocatalytic enhancement of the actin polymerization and a relative inhibition of the actin polymerization at some distance from the more active polymerization foci. In addition, a drug log concentration-polarizing effect bell-shaped curve may be simulated by assuming that the N-formyl-methionyl-leucyl-phenylalanine, while binding with high affinity to the specific receptor, is also able to bind to another lower affinity receptor that may effect depolarizing actions or, more generally, metabolic blocking effects. Under these conditions, at low drug concentrations the polarizing effect brought about by the ligation of the specific receptor is largely predominant. However, as the drug concentration increases and the specific receptors approach saturation, the inhibitory effects become more and more powerful and the net polarizing effect is reduced.

Formyl peptides are oligopeptides released by Gram-negative bacteria. So far, specific formyl peptide receptors (FPRs) have been described in mammals only. FPRs are seven-transmembrane G-coupled molecules and make up a relatively homogeneous group, although exhibiting different levels of affinity for the ligands. We examined the patterns of conservation/mutation within the FPR group of genes, as studied in 16 mRNAs from different species. Following alignment of the coding sections, those nucleotides identical in at least 15 sequences were assigned a "conservation index" 2; those with 8-14 identities an index 1; those with less than 8 identities an index zero. The cumulative average conservation index was 1.36. The autocorrelation function and the power spectrum of the whole series of indexes demonstrated a 3-unit periodicity. This periodicity is explained by the fact that the average conservation indexes of the first, second and third nucleotides of the coding triplets were 1.46, 1.55 (both above the mean), and 1.06 (below the mean), respectively, so that correlations at lag 3 tend to be all positive. In mRNAs, regardless of the position in the coding triplets, T is significantly more frequently conserved (average index = 1.60) than A, C, and G (1.21 - 1.38). In the nucleotides with conservation index 1 or zero, we recorded the two more frequently represented bases. In 35% of mRNA nucleotides the two more frequently represented bases were C and T; in 28% of cases the two more frequently represented bases were A and G; other couples occurred with lower frequencies. Both mutations may arise following C methylation with subsequent transformation into T (by deamination), either in the template or the coding DNA strand. Thus, we hypothesized that in FPR mRNAs there is an evolutionary trend of transformation from G to A and from C to T, the latter being the more stable of the bases.

1 The present study aimed to evaluate the role of kappa-opioid receptors at two peripheral sites, the vas deferens and the proximal colon, in kappa-opioid receptor knockout mice. We investigated the role of the kappa-opioid receptor in the vas deferens twitch response and in the colonic "off-contraction", a rebound contractile response which follows the inhibitory response to low frequencies stimulation (10, 20, 30 Hz) and which has been suggested to "locally" reproduce the contractile component of the peristaltic reflex. 2 Transmural stimulation of the vas deferens at lower frequencies (10 Hz, 10 V, 1 ms pulse trains lasting 0.5 s) evoked a contractile response that was significantly higher in the preparations from knockout mice because of lack of kappa-opioid receptors than in wild type mice. A selective kappa-opioid receptor agonist, U-50,488H, induced a dose-dependent inhibition of the electrically stimulated contraction in vas deferens. The percentages of reduction of the twitch response were significantly lower in knockout mice than in wild type mice after treatment with U-50,488H. The reduction of twitch response caused by U-50,488H was not reversed by administration of nor-binaltorphimine (nor-BNI) (5 x 10-6 m), a selective kappa-opioid receptor antagonist, in preparations from both knockout mice and wild type mice. U-50,488H has no effect on postsynaptic adrenergic receptors, as its administration did not affect the direct contractile response to noradrenaline. 3 Transmural stimulation (5 Hz, 20 V, 2 ms pulse trains lasting 30 s) induced inhibition of spontaneous activity of colonic strips during the period of stimulation, followed by an "off-contraction" after the cessation of stimulation. The statistical evaluation of the "off-contraction" responses between the two strains showed no significant difference. The off-contraction, measured in specimens from knockout mice, was inhibited concentration-dependently by U-50,488H (P < 0.01) and significantly less than from wild type mice. 4 The effect of U-50,488H was not reversed by administration of nor-BNI (5 x 10-6 m), either in preparations from knockout mice or from wild type mice. 5 Our data may suggest that kappa-opioid receptors are involved in some peripheral responses to the nerve stimulation, as indicated by the effect of U-50,488H, a selective kappa-opioid receptor agonist. However, the involvement of kappa-opioid receptor was also present, although less apparent, in kappa -opioid receptor knockout mice, suggesting either that this drug acts not only on kappa-opioid receptors but also on other receptor sites, such as kappa-like receptors. An alternative interpretation can be related to a sodium channel blocking action of U-50,488H, which could explain the inhibitory effects of twitch response still present but less evident in knockout strain and the lack of effect of the antagonist nor-BNI.

Objective: To evaluate the postoperative changes in circulating interleukin-2 (IL-2) concentration according to the severity of the surgical injury and other postoperative variables that could influence IL-2 production., Design: Prospective observational study., Setting: University hospital, Italy., Subjects: 43 patients about to undergo major operations (gastric and colo-rectal resection for cancer), intermediate operations (open cholecystectomy or mastectomy for cancer), and minor operations (hernia repair or breast lump); 24 healthy age and sex matched controls., Main Outcome Measures: Postoperative changes in serum concentrations of IL-2 after different degrees of surgery on the 1st, 3rd and 8th postoperative days correlated with changes in in vivo cellular mediated immunity (skin tests), duration of operation, blood transfusion or postoperative H2-blockers and antiprostaglandins., Results: There were no significant variations in IL-2 serum concentrations postoperatively on ANOVA, and when the data were normalised, there were no significant changes in the median postoperative values after minor and intermediate operations. There was a slight but not significant increase in IL-2 concentrations after major operations. Neither blood transfusion nor duration of operation correlated with postoperative changes in IL-2, while postoperative antiprostaglandins and H2-blockers seemed to provide slight but not significant protection against a reduction in IL-2 concentrations., Conclusions: Circulating IL-2 does not necessarily correlate with reported in vitro postoperative production of IL-2 and therefore seems to be of little use in monitoring immunosuppression in surgical patients.