Enhanced presence of serotonin in nasal cavity after autologous stimulation.

OBJECTIVE: Serotonin, which is a vasoactive amine, is an important neurotransmitter and is involved in many behavioral and psychological phenomena, such as pain, appetite, mood, and sleep. The primary purpose of our study was to investigate the effect of high-pressure administration of sterile physiological saline isotonic solution (HpPSIS) into nasal cavity and to determine the expression of the serotonin. PATIENTS AND METHODS: The study was made in two branches, the previous with 14 volunteers, the subsequent study with 40 patients with mild anxiety disorder. The middle third of the inferior turbinate epithelial cells on the right nostril was scraped using a sterile curette and indicated as (pre), then, a spray of sterilized isotonic solution at high pressure on the left nostril was delivered, and 5 minutes later a similar stimulation was delivered on the same nostril. The stimulation was made with a specific spray dispenser. The middle third of the inferior turbinate epithelial cells on the left nostril was scraped using a sterile curette and indicated as (post). Then, based on the first part of our study, we started the second part and gave a treatment on forty new patients with anxiety disorder. RESULTS: The results of these studies highlight the possibility of endogenous enhancement of serotonin by stimulation of mast cells. In the first part of the study, Serotonin significantly increased in protein extracts after treatment (64.35±5.33 vs. 10.97±2.17; unpaired two tailed t-test, t=9.8, df=24, p≤0.0001; F=6.035; DFn=12; DFd=12). In the second part of the study, in patients treated with HpPSIS, we observed improvement of mood, after one, two and three months, with a statistically significant reduction of DASS-21, while no reduction was observed in control patients, treated with normal pressure commercial spray. CONCLUSIONS: This pilot study showed that the topical treatment of HpPHIS increases serotonin levels in nasal cavity. The observation reported in this study opens the way to a new valid strategy to enhance the level of endogenous serotonin. We observed a significant improvement of ASI on patients during HpPHIS therapy.

Cationic liposomes as potential carriers for ocular administration of peptides with anti-herpetic activity.

In the present study the preparation, characterization and activity of cationic liposomes containing the secretory form of herpes simplex virus type 1 (HSV-1) glycoprotein B (gB1s) or two related polylysine rich peptides, namely DTK1 and DTK2, were described. The immunotherapeutic potential of these HSV antigens containing liposomes was examined with a rabbit ocular model of HSV-1 infection. Our study indicates that the liposomes (i) are able to encapsulate quantitatively gB1s and around 30% the DTK peptides, (ii) are characterized by dimensions compatible with ocular applications and (iii) can release the peptide comparably to the free solution. In addition, neutralization studies demonstrated that an anti-DTK specific polyclonal antiserum can inhibit HSV-1 infection, indicating that such peptides could be a good immunogen/antigen in an anti-HSV vaccine formulation. Although the vaccination protocol did not induce protection against the eye disease, a significative protection against a lethal ocular challenge was detectable together with the absence of reactivation episodes from latency on the survived animals. In this respect, the use of cationic liposomes coupled to gB1s and DTK peptides, as a local ocular vaccine, could represent an interesting approach in order to obtain a possible efficacy in protecting animals against a subsequent HSV-1 ocular challenge.

Biological variation responses in fMLP-OMe analogs, introducing bulky protecting groups on the side-chain of hydrophilic residues at position 2.

for-Met-Ser(Bzl)-Phe-OMe, for-Met-Cys(Bzl)-Phe-OMe, for-Met-Tyr(Bzl)-Phe-OMe and for-Met-Lys(Z)-Phe-OMe were synthesized to investigate the importance of a bulky protecting group on the side-chain of a hydrophilic residue at position 2 on the biological activities of human neutrophils. Our results indicate that these compounds do not trigger a good chemotactic response, which, in any case, is not improved with respect to that induced by the analogs with the unprotected residues. Instead, both superoxide anion production and, particularly, lysozyme release are more efficient.

Biological activity in human neutrophils of di-tripeptides, analogues of the chemotactic fMLP.

for-Met-Ser(for-Met-Leu-Phe)-Phe-OMe 1 and for-Met-Lys(for-Met-Leu-Phe)-Phe-OMe 2 were synthesized in order to investigate biological activities on human neutrophils of crosslinked di-tripeptides. Our results seem to highlight that the tested di-tripeptides (i) do not step up chemotaxis, (ii) can elicit superoxide anion production which is dependent on the nature of the residue at position 2, chosen in the tripeptide that is crosslinked to the fMLP-OMe.

A hydrophilic residue at position 2 can improve specific biological responses in fMLP-OMe analogs.

The peptides for-Met-Ser-Phe-OMe 1, for-Met-Cys-Phe-OMe 2, for-Met-Lys-Phe-OMe 3, and for-Met-Tyr-Phe-OMe 4 were synthesized in order to investigate the importance of a hydrophilic side-chain on the residue at position 2 on biological activities of human neutrophils. Our results seem to highlight that this type of substitution does not facilitate good chemotaxis, although it elicits both superoxide anion production and particularly lysozyme release, in some cases even more potent than the parent fMLP-OMe, if the hydrophilicity is associated with steric hindrance.

Differences Arising in Human Neutrophil Activation Passing from N-Formyl to N-Acetyl-oligopeptides.

N-formyl- and N-acetyl-peptides were synthesized and compared in order to understand which features can best elicit biological responses. The behavior of N-formyl-peptides confirms the previously found sequential obligations in the residues, while acetyl-derivatives do not seem suitable for an efficacious stimulation of human neutrophils. Copyright 2000 Academic Press.

An evaluation of fMLP pocket dimensions and features using formyltetrapeptides.

The formyltetrapeptides for-Met-Leu-Leu-Phe-OMe 1, for-Met-Leu-Aib-Phe-OMe 2, for-Met-Leu-Ac6c-Phe-OMe 3, for-Met-Leu-Pro-Phe-OMe 4, for-Met-Pro-Pro-Phe-OMe 5, for-Met-Aib-Aib-Phe-OMe 6, for-Met-Pro-Aib-Phe-OMe 7 and for-Met-Aib-Pro-Phe-OMe 8 were synthesized and biologically tested on human neutrophils in an attempt to evaluate the specific receptor pocket dimensions and features. Our results indicate that the shift in the Phe residue to the fourth position in these compounds strongly reduces chemotactic response, but is efficacious in triggering superoxide anion production and lysozyme release (order of potency 3 > 2 > 1 > 4 > 6 > 8 > 5 > 7). The potency of the two latter responses correlates well with the affinity data obtained in binding experiments.

fMLP-OMe analogs substituted at the methionine residue: an insight into the receptor properties.

In order to obtain an insight into the mode of binding at the for-Met-Leu-Phe-OH (fMLP) receptor, three fMLP-OMe analogs (1-3) were synthesized in which the Met residue was substituted by Gln 1, Asn 2, and Ser 3. We evaluated the influence of the charge variation and/or the shift of its position on neutrophil biological responses.

Synthesis, biological activity and conformational studies of geometrically restricted tripeptide analogues of the chemoattractant HCO-Met-Leu-Phe-OMe.

The formyl tripeptides containing 2-azetidinecarboxylic acid 2, 2-piperidinecarboxylic acid 3 and norvaline 4 in position 2 were synthesized and their biological activity was evaluated. The conformation of peptides was studied by CD and FT-IR techniques. While 2 and 3 do not show either chemotactic activity or superoxide production, 4 retains both activities.

Modifications of the amide bond at position 3 in fMLP analogs select neutrophil functions.

The formylpeptides formyl-L-methionyl-L-leucyl-L-N-methylphenylalanine methyl ester 1, formyl-L-methionyl-L-leucyl-L-2-oxy-3-phenylpropionic acid methyl ester 2 and formyl-L-methionyl-L-leucyl-L-2-aminoxy-3-phenylpropionic acid methyl ester 3 were synthesized to investigate the role of the amide bond at position 3 in biological activity in human neutrophiles. Our results indicate that this amide bond is required for optimal chemotactic activity, but not for superoxide anion production.

Synthesis and biological activity of a conformationally constrained chemotactic gamma-lactam formyl tetrapeptide.

The synthesis and biological activity on human neutrophils of for-Met-(gamma-lactam)-Leu-Phe-OMe [fM(gamma l)LP-OMe], an analogue of the chemotactic fMLP-OMe, are reported. The tetrapeptide evidences chemotactic activity as well as superoxide anion production and lysozyme release, although with lower efficacy when compared with the parent tripeptide.

Delta-selective opioid peptides containing a single aromatic residue in the message domain: an NMR conformational analysis.

The sequence of deltorphin I, a delta-selective opioid agonist, has been systematically modified by inserting conformationally constrained C alpha, alpha disubstituted apolar residues in the third position. As expected, substitution of Phe with Ac6c, Ac5c and Ac3c yields analogues with decreasing but sizeable affinity. Surprisingly, substitution with Aib yields an analogue with almost the same binding affinity of the parent compound but with a greatly increased selectivity. This is the first case of a potent and very selective opioid peptide containing a single aromatic residue in the message domain, that is, only Tyr1. Here we report a detailed conformational analysis of [Aib3]deltorphin I and [Ac6c3]deltorphin I in DMSO at room temperature and in a DMSO/water cryomixture at low temperature, based on NMR spectroscopy and energy calculations. The peptides are highly structured in both solvents, as indicated by the exceptional finding of a nearly zero temperature coefficient of Val5 NH resonance. NMR data cannot be explained on the basis of a single structure but it was possible to interpret all NMR data on the basis of a few structural families. The conformational averaging was analysed by means of an original computer program that yields qualitative and quantitative composition of the mixture. Comparison of the preferred solution conformation with two rigid delta-selective agonists shows that the shapes of [Aib3]deltorphin I and [Ac6c3]deltorphin I are consistent with those of rigid agonists and that the message domain of opioid peptides can be defined only in conformational terms.

The importance of the peptide bond at position 2 in HCO-Met-Leu-Phe-OMe analogues as shown by studies on human neutrophils.

The formylpeptides formyl-methionyl-N-methylleucyl-phenylalanine methyl ester [for-Met-(NMe)Leu-Phe-OMe] 1, formyl-methionyl-2-aminotetralin-2-carboxyl-phenylalanine methyl ester [for-Met-Atc-Phe-OMe] 2, formyl-methionyl-1,2,3,4-tetrahydroisoquinoline-3-carboxyl-phenylalanine methyl ester [for-Met-Tic-Phe-OMe] 3 and formyl-methionyl-2-aminoxy-4-methylvaleryl-phenylalanine methyl ester [for-Met-OLeu-Phe-OMe] 4 were synthesized in order to investigate the role of the amide bond at position 2 on biological activities on human neutrophils. Only analogue 2, which keeps the NH group at position 2, was found to retain activity though sterically encumbered.

Conformational studies of chemotactic HCO-Met-Leu-Phe-OMe analogues.

In order to investigate the proper peptide backbone conformation able to elicit a biological activity, HCO-Met-Pro-Phe-OMe, HCO-Met-Ψ[COO]Leu-Phe-OMe, and HCO-Met-OLeu-Phe-OMe, analogues of the prototypical chemotactic peptide HCO-Met-Leu-Phe-OMe, were studied by CD and IR techniques. The results obtained comparing biological and conformational data evidence the critical presence of (i) the NH group at position 2, (ii) a rather flexible backbone, (iii) the chemical structure of the central residue which can affect the stability of a possible active conformer.

Structures of peptides from alpha-amino acids methylated at the alpha-carbon.

The structural preferences of peptides (and depsipeptides) from the achiral MeAib and Hib residues, and the chiral Iva, (alpha Me) Val, (alpha Me) Leu, and (alpha Me) Phe residues, as determined by conformational energy computations, x-ray diffraction analyses, and 1H-nmr and spectroscopic studies, are reviewed and compared with literature data on Aib-containing peptides. The results obtained indicate that helical structures are preferentially adopted by peptides rich in these alpha-amino acids methylated at the alpha-carbon. Intriguing experimental findings on the impact of the chirality of Iva, (alpha Me) Val, and (alpha Me) Phe residues on helix screw sense are illustrated.

The beta-bend ribbon spiral. Synthesis and conformational analysis in solution and in the crystal state of depsipeptides containing alpha-hydroxyisobutyric acid.

The synthesis and conformational analysis in solution (by FTIR absorption and 1H NMR) and in the crystal state (by X-ray diffraction) of three Hib-containing depsipeptides have been performed. In the crystal state Z-Aib-Hib-Aib-OMe is folded into a type-III beta-bend, while the conformation adopted by Z-Aib-Hib)2-Aib-OMe is a beta-bend ribbon spiral, characterized by two type-III beta-bends with Aib(1)-Hib(2) and Aib(3)-Hib(4) as corner residues, respectively. Both independent molecules in the asymmetric unit of t-Boc-L-Ala-Hib-L-Ala-OMe crystals are folded into a type-II beta-bend. For the Aib-Hib depsipeptides the conformation adopted in the crystal state is also that largely prevailing in solution, whereas for t-Boc-L-Ala-Hib-L-Ala-OMe the beta-bend conformation is significantly less populated in solution. A comparison is also made with: (i) the published crystal-state conformations of fully protected -(Aib)3-, -(Aib)5-, and -L-Ala-Aib-L-Ala- sequences and the beta-bend ribbon spiral generated by (Aib-L-Pro)n oligomers, and (ii) with the herewith described solution preferred conformation of Z-L-Ala-Aib-L-Ala-OMe. The possible use of Hib as an isosteric replacement for Aib in the design of conformationally constrained depsipeptides is briefly discussed.

Structure-activity relationship in the mutagenic effect of chiral or racemic 2-bromo-propanamides on Salmonella typhimurium.

Some 2-bromo-propanamides were prepared and tested for direct mutagenicity in Salmonella typhimurium TA 100. Results confirm the mutagenic activity of 2-bromo-N-benzyl-propanamide and indicate that it is independent of enantiomeric configuration. A variation in the chemical structure, namely, the addition of a methyl group at the benzylic carbon, causes the four resulting diastereomers to be devoid of any activity. Conversely, some racemic ring-substituted methoxy and/or hydroxy derivatives of the parent compound displayed mutagenic properties, causing an increase in the number of his+ revertants up to 524 per milligram per plate.

Selective mutagenic activity of 2-bromo-propanamides on Salmonella typhimurium. A structure-activity study.

Nineteen 2-halogeno-acetamides, -propanamides, and -iso-butyramides and four related epoxyamides were tested as mutagens for Salmonella typhimurium TA100. Mutagenic activity was observed in strictly selected 2-halogenoamides and in all epoxyamides. The effectiveness of 2-halogenoamides depends upon: the character of the carbon carrying the halogen (1 degree, 2 degrees, 3 degrees); the nature of the halogen (Br, Cl); the substitution at nitrogen. Some considerations concerning the selectivity observed are presented.