3D Cell Migration Chip (3DCM-Chip): A New Tool toward the Modeling of 3D Cellular Complex Systems.

3D hydrogel-based cell cultures provide models for studying cell behavior and can efficiently replicate the physiologic environment. Hydrogels can be tailored to mimic mechanical and biochemical properties of specific tissues and allow to produce gel-in-gel models. In this system, microspheres encapsulating cells are embedded in an outer hydrogel matrix, where cells are able to migrate. To enhance the efficiency of such studies, a lab-on-a-chip named 3D cell migration-chip (3DCM-chip) is designed, which offers substantial advantages over traditional methods. 3DCM-chip facilitates the analysis of biochemical and physical stimuli effects on cell migration/invasion in different cell types, including stem, normal, and tumor cells. 3DCM-chip provides a smart platform for developing more complex cell co-cultures systems. Herein the impact of human fibroblasts on MDA-MB 231 breast cancer cells' invasiveness is investigated. Moreover, how the presence of different cellular lines, including mesenchymal stem cells, normal human dermal fibroblasts, and human umbilical vein endothelial cells, affects the invasive behavior of cancer cells is investigated using 3DCM-chip. Therefore, predictive tumoroid models with a more complex network of interactions between cells and microenvironment are here produced. 3DCM-chip moves closer to the creation of in vitro systems that can potentially replicate key aspects of the physiological tumor microenvironment.

Modelling the disease: H2S-sensitivity and drug-resistance of triple negative breast cancer cells can be modulated by embedding in isotropic micro-environment.

Three-dimensional (3D) cell culture systems provide more physiologically relevant information, representing more accurately the actual microenvironment where cells reside in tissues. However, the differences between the tissue culture plate (TCP) and 3D culture systems in terms of tumour cell growth, proliferation, migration, differentiation and response to the treatment have not been fully elucidated. Tumoroid microspheres containing the MDA-MB 231 breast cancer cell line were prepared using either tunable PEG-fibrinogen (PFs) or tunable PEG-silk fibroin (PSFs) hydrogels, respectively named MDAPFs and MDAPSFs. The cancer cells in the tumoroids showed changes both in globular morphology and at the protein expression level. A decrease of both Histone H3 acetylation and cyclin D1 expression in all 3D systems, compared to the 2D cell culture, was detected in parallel to changes of the matrix stiffness. The effects of a glutathionylated garlic extract (GSGa), a slow H2S-releasing donor, were investigated on both tumoroid systems. A pro-apoptotic effect of GSGa on tumour cell growth in 2D culture was observed as opposed to a pro-proliferative effect apparent in both MDAPFs and MDAPSFs. A dedicated ad hoc 3D cell migration chip was designed and optimized for studying tumour cell invasion in a gel-in-gel configuration. An anti-cell-invasion effect of the GSGa was observed in the 2D cell culture, whereas a pro-migratory effect in both MDAPFs and MDAPSFs was observed in the 3D cell migration chip assay. An increase of cyclin D1 expression after GSGa treatment was observed in agreement with an increase of the cell invasion index. Our results suggest that the "dimensionality" and the stiffness of the 3D cell culture milieu can change the response to both the gasotransmitter H2S and doxorubicin due to differences in both H2S diffusion and changes in protein expression. Moreover, we uncovered a direct relation between the cyclin D1 expression and the stiffness of the 3D cell culture milieu, suggesting the potential causal involvement of the cyclin D1 as a bio-marker for sensitivity of the tumour cells to their matrix stiffness. Therefore, our hydrogel-based tumoroids represent a valid tunable model for studying the physically induced transdifferentiation (PiT) of cancer cells and as a more reliable and predictive in vitro screening platform to investigate the effects of anti-tumour drugs.

Orchestration of Mesenchymal Stem/Stromal Cells and Inflammation During Wound Healing.

Wound healing is a complex process and encompasses a number of overlapping phases, during which coordinated inflammatory responses following tissue injury play dominant roles in triggering evolutionarily highly conserved principals governing tissue repair and regeneration. Among all nonimmune cells involved in the process, mesenchymal stem/stromal cells (MSCs) are most intensely investigated and have been shown to play fundamental roles in orchestrating wound healing and regeneration through interaction with the ordered inflammatory processes. Despite recent progress and encouraging results, an informed view of the scope of this evolutionarily conserved biological process requires a clear understanding of the dynamic interplay between MSCs and the immune systems in the process of wound healing. In this review, we outline current insights into the ways in which MSCs sense and modulate inflammation undergoing the process of wound healing, highlighting the central role of neutrophils, macrophages, and T cells during the interaction. We also draw attention to the specific effects of MSC-based therapy on different pathological wound healing. Finally, we discuss how ongoing scientific advances in MSCs could be efficiently translated into clinical strategies, focusing on the current limitations and gaps that remain to be overcome for achieving preferred functional tissue regeneration.

Sustainable Vegetable Oil-Based Biomaterials: Synthesis and Biomedical Applications.

One of the main criteria for ecological sustainability is that the materials produced for common use are green. This can include the use of biomaterials and materials that are environmentally friendly, biodegradable and produced at low cost. The exploration of natural resources as sustainable precursors leads to the production of biopolymers that are useful for 3D printing technology. Recently, waste vegetable oils have been found to be a good alternative source for the production of biopolymers in various applications from the engineering to the biomedicine. In this review, the processes for the synthesis of vegetable oil-based biomaterials are described in detail. Moreover, the functionalization strategies to improve the mechanical properties of these materials and the cell-material interaction for their potential use as micro-structured scaffolds in regenerative medicine are discussed.

New vegetable-waste biomaterials by Lupin albus L. as cellular scaffolds for applications in biomedicine and food.

The reprocessing of vegetal-waste represents a new research field in order to design novel biomaterials for potential biomedical applications and in food industry. Here we obtained a biomaterial from Lupinus albus L. hull (LH) that was characterized micro-structurally by scanning electron microscopy and for its antimicrobial and scaffolding properties. A good adhesion and proliferation of human mesenchymal stem cells (hMSCs) seeded on LH scaffold were observed. Thanks to its high content of cellulose and beneficial phytochemical substances, LH and its derivatives can represent an available source for fabrication of biocompatible and bioactive scaffolds. Therefore, a reprocessing protocol of LH was optimized for producing a new LH bioplastic named BPLH. This new biomaterial was characterized by chemico-physical analyses. The water uptake, degradability and antimicrobial properties of BPLH were evaluated, as well as the mechanical properties. A good adhesion and proliferation of both fibroblasts and hMSCs on BPLH were observed over 2 weeks, and immunofluorescence analysis of hMSCs after 3 weeks indicates an initial commitment toward muscle differentiation. Our work represents a new approach toward the recovery and valorization of the vegetal waste showing the remarkable properties of LH and BPLH as cellular waste-based scaffold with potential applications in cell-based food field as well as in medicine for topical patches in wound healing and bedsores treatment.

Global mapping of cancers: The Cancer Genome Atlas and beyond.

Cancer genomes have been explored from the early 2000s through massive exome sequencing efforts, leading to the publication of The Cancer Genome Atlas in 2013. Sequencing techniques have been developed alongside this project and have allowed scientists to bypass the limitation of costs for whole-genome sequencing (WGS) of single specimens by developing more accurate and extensive cancer sequencing projects, such as deep sequencing of whole genomes and transcriptomic analysis. The Pan-Cancer Analysis of Whole Genomes recently published WGS data from more than 2600 human cancers together with almost 1200 related transcriptomes. The application of WGS on a large database allowed, for the first time in history, a global analysis of features such as molecular signatures, large structural variations and noncoding regions of the genome, as well as the evaluation of RNA alterations in the absence of underlying DNA mutations. The vast amount of data generated still needs to be thoroughly deciphered, and the advent of machine-learning approaches will be the next step towards the generation of personalized approaches for cancer medicine. The present manuscript wants to give a broad perspective on some of the biological evidence derived from the largest sequencing attempts on human cancers so far, discussing advantages and limitations of this approach and its power in the era of machine learning.

Photo-Polymerization Damage Protection by Hydrogen Sulfide Donors for 3D-Cell Culture Systems Optimization.

Photo-polymerized hydrogels are ideally suited for stem-cell based tissue regeneration and three dimensional (3D) bioprinting because they can be highly biocompatible, injectable, easy to use, and their mechanical and physical properties can be controlled. However, photo-polymerization involves the use of potentially toxic photo-initiators, exposure to ultraviolet light radiation, formation of free radicals that trigger the cross-linking reaction, and other events whose effects on cells are not yet fully understood. The purpose of this study was to examine the effects of hydrogen sulfide (H2S) in mitigating cellular toxicity of photo-polymerization caused to resident cells during the process of hydrogel formation. H2S, which is the latest discovered member of the gasotransmitter family of gaseous signalling molecules, has a number of established beneficial properties, including cell protection from oxidative damage both directly (by acting as a scavenger molecule) and indirectly (by inducing the expression of anti-oxidant proteins in the cell). Cells were exposed to slow release H2S treatment using pre-conditioning with glutathione-conjugated-garlic extract in order to mitigate toxicity during the photo-polymerization process of hydrogel formation. The protective effects of the H2S treatment were evaluated in both an enzymatic model and a 3D cell culture system using cell viability as a quantitative indicator. The protective effect of H2S treatment of cells is a promising approach to enhance cell survival in tissue engineering applications requiring photo-polymerized hydrogel scaffolds.

New Consensus pattern in Spike CoV-2: potential implications in coagulation process and cell-cell fusion.

Coagulopathy and syncytial formation are relevant effects of the SARS-CoV-2 infection, but the underlying molecular mechanisms triggering these processes are not fully elucidated. Here, we identified a potential consensus pattern in the Spike S glycoprotein present within the cytoplasmic domain; this consensus pattern was detected in only 79 out of 561,000 proteins (UniProt bank). Interestingly, the pattern was present in both human and bat the coronaviruses S proteins, in many proteins involved in coagulation process, cell-cell interaction, protein aggregation and regulation of cell fate, such as von Willebrand factor, coagulation factor X, fibronectin and Notch, characterized by the presence of the cysteine-rich EGF-like domain. This finding may suggest functional similarities between the matched proteins and the CoV-2 S protein, implying a new possible involvement of the S protein in the molecular mechanism that leads to the coagulopathy and cell fusion in COVID-19 disease.

Glutathione-Allylsulfur Conjugates as Mesenchymal Stem Cells Stimulating Agents for Potential Applications in Tissue Repair.

The endogenous gasotransmitter H2S plays an important role in the central nervous, respiratory and cardiovascular systems. Accordingly, slow-releasing H2S donors are powerful tools for basic studies and innovative pharmaco-therapeutic agents for cardiovascular and neurodegenerative diseases. Nonetheless, the effects of H2S-releasing agents on the growth of stem cells have not been fully investigated. H2S preconditioning can enhance mesenchymal stem cell survival after post-ischaemic myocardial implantation; therefore, stem cell therapy combined with H2S may be relevant in cell-based therapy for regenerative medicine. Here, we studied the effects of slow-releasing H2S agents on the cell growth and differentiation of cardiac Lin- Sca1+ human mesenchymal stem cells (cMSC) and on normal human dermal fibroblasts (NHDF). In particular, we investigated the effects of water-soluble GSH-garlic conjugates (GSGa) on cMSC compared to other H2S-releasing agents, such as Na2S and GYY4137. GSGa treatment of cMSC and NHDF increased their cell proliferation and migration in a concentration dependent manner with respect to the control. GSGa treatment promoted an upregulation of the expression of proteins involved in oxidative stress protection, cell-cell adhesion and commitment to differentiation. These results highlight the effects of H2S-natural donors as biochemical factors that promote MSC homing, increasing their safety profile and efficacy after transplantation, and the value of these donors in developing functional 3D-stem cell delivery systems for cardiac muscle tissue repair and regeneration.

Hydrogen Sulfide-Releasing Fibrous Membranes: Potential Patches for Stimulating Human Stem Cells Proliferation and Viability under Oxidative Stress.

The design of biomaterial platforms able to release bioactive molecules is mandatory in tissue repair and regenerative medicine. In this context, electrospinning is a user-friendly, versatile and low-cost technique, able to process different kinds of materials in micro- and nano-fibers with a large surface area-to-volume ratio for an optimal release of gaseous signaling molecules. Recently, the antioxidant and anti-inflammatory properties of the endogenous gasotramsmitter hydrogen sulfide (H2S), as well as its ability to stimulate relevant biochemical processes on the growth of mesenchymal stem cells (MSC), have been investigated. Therefore, in this work, new poly(lactic) acid fibrous membranes (PFM), doped and functionalized with H2S slow-releasing donors extracted from garlic, were synthetized. These innovative H2S-releasing mats were characterized for their morphological, thermal, mechanical, and biological properties. Their antimicrobial activity and effects on the in vitro human cardiac MSC growth, either in the presence or in the absence of oxidative stress, were here assessed. On the basis of the results here presented, these new H2S-releasing PFM could represent promising and low-cost scaffolds or patches for biomedical applications in tissue repair.

Trichormus variabilis (Cyanobacteria) Biomass: From the Nutraceutical Products to Novel EPS-Cell/Protein Carrier Systems.

A native strain of the heterocytous cyanobacterium Trichormus variabilis VRUC 168 was mass cultivated in a low-cost photobioreactor for a combined production of Polyunsaturated Fatty Acids (PUFA) and Exopolymeric Substances (EPS) from the same cyanobacterial biomass. A sequential extraction protocol was optimized leading to high yields of Released EPS (REPS) and PUFA, useful for nutraceutical products and biomaterials. REPS were extracted and characterized by chemical staining, Reversed Phase-High-Performance Liquid Chromatography (RP-HPLC), Fourier Transform Infrared Spectroscopy (FT-IR) and other spectroscopic techniques. Due to their gelation property, REPS were used to produce a photo-polymerizable hybrid hydrogel (REPS-Hy) with addition of polyethylene glycol diacrylated (PEGDa). REPS-Hy was stable over time and resistant to dehydration and spontaneous hydrolysis. The rheological and functional properties of REPS-Hy were studied. The enzyme carrier ability of REPS-Hy was assessed using the detoxification enzyme thiosulfate:cyanide sulfur transferase (TST), suggesting the possibility to use REPS-Hy as an enzymatic hydrogel system. Finally, REPS-Hy was used as a scaffold for culturing human mesenchymal stem cells (hMSCs). The cell seeding onto the REPS-Hy and the cell embedding into 3D-REPS-Hy demonstrated a scaffolding property of REPS-Hy with non-cytotoxic effect, suggesting potential applications of cyanobacteria REPS for producing enzyme- and cell-carrier systems.

An Unexpected Risk Factor for Early Structural Deterioration of Biological Aortic Valve Prostheses.

BACKGROUND: An alarming rate of early failure has been recently reported for the LivaNova (previously Sorin) Mitroflow (LivaNova, London, UK) bioprosthesis. Here, we aimed at verifying if this possible underperformance is confirmed in a large, single-center experience and identifying the risk factors associated with early deterioration. METHODS: In all, 459 Mitroflow valves have been implanted from July 2009 to December 2013 (patients' mean age 73 years; 204 women). Surviving patients have undergone yearly clinic and echocardiographic follow-up. Dysfunction was defined as moderate if the mean gradient was more than 30 mm Hg or severe if it exceeded 40 mm Hg. The population was divided on the basis of a dimensional mismatch, the model of the prosthesis (LX or DL: follow-up to 4 years), and patient's age at the time of implantation. RESULTS: Cumulative freedom from moderate valve dysfunction was 81% ± 3% at 60 months. It was lower with patient-prosthesis mismatch (71% ± 5% versus 92% ± 3%; p = 0.0065) and with the more recent DL model (at 42 months: 78% ± 6% versus 96% ± 2%; p < 0.0001). Cumulative freedom from severe dysfunction was 93% ± 2% at 5 years. Again, it was inferior among patients with a mismatch (86% ± 4% versus 100%; p = 0.0013) and for the DL model (42 months: 92.5% ± 3% versus 98.5% ± 1%; p = 0.0309). Smaller prostheses showed higher rates of early degeneration. CONCLUSIONS: The LivaNova Mitroflow valve appears to be prone to early deterioration. Smaller size prostheses should be used cautiously and avoided with patient-prosthesis mismatch. The DL model anticalcification treatment seems unable to prevent early degeneration, and possibly contributes to even earlier failure.

Injectable silk fibroin hydrogels functionalized with microspheres as adult stem cells-carrier systems.

Hydrogels are good candidate materials for cell delivery scaffolds because they can mimic the physical, chemical, electrical and biological properties of most of the native tissues. In this study, composite biosynthetic hydrogels were produced by combining the bio-functionality of silk fibroin (SF) with the structural versatility of polyethylene-glycol-diacrylated (PEGDa). The formation of a photopolymerizable PEGDa-SF hydrogel (PSFHy) was optimized for 3D-cell culture. Functionalization of the 3D-PSFHy with protein microspheres (MS) was required to increase the porosity and cell-adhesive properties of the material. Cardiac mesenchymal stem cells, which were cultured within the MS-embedding PSFHy, exhibited good viability and expression of proteins that are characteristic of the initial phases of the cardiac muscle differentiation process. Further, the addition of chondroitin sulfate into the scaffolds improved the cell viability. A cell-preconditioning of the scaffold was also performed, suggesting a potential application of these sponge-like scaffolds for analysing the effects of several extracellular microenvironments, produced by different kinds of cells, on the stem cells fate. The results presented herein highlight on the possibility to use the PSFHys functionalized with MS as stem cell-carrier systems with sponge-like properties, potential ultrasound-imaging contrast agents and controlled biochemical factor delivery.

H2S-releasing nanoemulsions: a new formulation to inhibit tumor cells proliferation and improve tissue repair.

The improvement of solubility and/or dissolution rate of poorly soluble natural compounds is an ideal strategy to make them optimal candidates as new potential drugs. Accordingly, the allyl sulfur compounds and omega-3 fatty acids are natural hydrophobic compounds that exhibit two important combined properties: cardiovascular protection and antitumor activity. Here, we have synthesized and characterized a novel formulation of diallyl disulfide (DADS) and α-linolenic acid (ALA) as protein-nanoemulsions (BAD-NEs), using ultrasounds. BAD-NEs are stable over time at room temperature and show antioxidant and radical scavenging property. These NEs are also optimal H2S slow-release donors and show a significant anti-proliferative effect on different human cancer cell lines: MCF-7 breast cancer and HuT 78 T-cell lymphoma cells. BAD-NEs are able to regulate the ERK1/2 pathway, inducing apoptosis and cell cycle arrest at the G0/G1 phase. We have also investigated their effect on cell proliferation of human adult stem/progenitor cells. Interestingly, BAD-NEs are able to improve the Lin- Sca1+ human cardiac progenitor cells (hCPC) proliferation. This stem cell growth stimulation is combined with the expression and activation of proteins involved in tissue-repair, such as P-AKT, α-sma and connexin 43. Altogether, our results suggest that these antioxidant nanoemulsions might have potential application in selective cancer therapy and for promoting the muscle tissue repair.

Amino-terminal residues of ΔNp63, mutated in ectodermal dysplasia, are required for its transcriptional activity.

p63, a member of the p53 family, is a crucial transcription factor for epithelial development and skin homeostasis. Heterozygous mutations in TP63 gene have been associated with human ectodermal dysplasia disorders. Most of these TP63 mutations are missense mutations causing amino acidic substitutions at p63 DNA binding or SAM domains that reduce or abolish the transcriptional activity of mutants p63. A significant number of mutants, however, resides in part of the p63 protein that apparently do not affect DNA binding and/or transcriptional activity, such as the N-terminal domain. Here, we characterize five p63 mutations at the 5' end of TP63 gene aiming to understand the pathogenesis of the diseases and to uncover the role of ΔNp63α N-terminus residues in determining its transactivation potential.

Glutathione-garlic sulfur conjugates: slow hydrogen sulfide releasing agents for therapeutic applications.

Natural organosulfur compounds (OSCs) from Allium sativum L. display antioxidant and chemo-sensitization properties, including the in vitro inhibition of tumor cell proliferation through the induction of apoptosis. Garlic water- and oil-soluble allyl sulfur compounds show distinct properties and the capability to inhibit the proliferation of tumor cells. In the present study, we optimized a new protocol for the extraction of water-soluble compounds from garlic at low temperatures and the production of glutathionyl-OSC conjugates during the extraction. Spontaneously, Cys/GSH-mixed-disulfide conjugates are produced by in vivo metabolism of OSCs and represent active molecules able to affect cellular metabolism. Water-soluble extracts, with (GSGaWS) or without (GaWS) glutathione conjugates, were here produced and tested for their ability to release hydrogen sulfide (H2S), also in the presence of reductants and of thiosulfate:cyanide sulfurtransferase (TST) enzyme. Thus, the TST catalysis of the H2S-release from garlic OSCs and their conjugates has been investigated by molecular in vitro experiments. The antiproliferative properties of these extracts on the human T-cell lymphoma cell line, HuT 78, were observed and related to histone hyperacetylation and downregulation of GAPDH expression. Altogether, the results presented here pave the way for the production of a GSGaWS as new, slowly-releasing hydrogen sulfide extract for potential therapeutic applications.

p63 threonine phosphorylation signals the interaction with the WW domain of the E3 ligase Itch.

Both in epithelial development as well as in epithelial cancers, the p53 family member p63 plays a crucial role acting as a master transcriptional regulator. P63 steady state protein levels are regulated by the E3 ubiquitin ligase Itch, via a physical interaction between the PPxY consensus sequence (PY motif) of p63 and one of the 4 WW domains of Itch; this substrate recognition process leads to protein-ubiquitylation and p63 proteasomal degradation. The interaction of the WW domains, a highly compact protein-protein binding module, with the short proline-rich sequences is therefore a crucial regulatory event that may offer innovative potential therapeutic opportunity. Previous molecular studies on the Itch-p63 recognition have been performed in vitro using the Itch-WW2 domain and the peptide interacting fragment of p63 (pep63), which includes the PY motif. Itch-WW2-pep63 interaction is also stabilized in vitro by the conformational constriction of the S-S cyclization in the p63 peptide. The PY motif of p63, as also for other proteins, is characterized by the nearby presence of a (T/S)P motif, which is a potential recognition site of the WW domain of the IV group present in the prolyl-isomerase Pin1. In this study, we demonstrate, by in silico and spectroscopical studies using both the linear pep63 and its cyclic form, that the threonine phosphorylation of the (T/S)PPPxY motif may represent a crucial regulatory event of the Itch-mediated p63 ubiquitylation, increasing the Itch-WW domains-p63 recognition event and stabilizing in vivo the Itch-WW-p63 complex. Moreover, our studies confirm that the subsequently trans/cis proline isomerization of (T/S)P motif by the Pin1 prolyl-isomerase, could modulate the E3-ligase interaction, and that the (T/S)pPtransPPxY motif represent the best conformer for the ItchWW-(T/S)PPPxY motif recognition.

Screening for E3-ubiquitin ligase inhibitors: challenges and opportunities.

The ubiquitin proteasome system (UPS) plays a role in the regulation of most cellular pathways, and its deregulation has been implicated in a wide range of human pathologies that include cancer, neurodegenerative and immunological disorders and viral infections. Targeting the UPS by small molecular regulators thus provides an opportunity for the development of therapeutics for the treatment of several diseases. The proteasome inhibitor Bortezomib was approved for treatment of hematologic malignancies by the FDA in 2003, becoming the first drug targeting the ubiquitin proteasome system in the clinic. Development of drugs targeting specific components of the ubiquitin proteasome system, however, has lagged behind, mainly due to the complexity of the ubiquitination reaction and its outcomes. However, significant advances have been made in recent years in understanding the molecular nature of the ubiquitination system and the vast variety of cellular signals that it produces. Additionally, improvement of screening methods, both in vitro and in silico, have led to the discovery of a number of compounds targeting components of the ubiquitin proteasome system, and some of these have now entered clinical trials. Here, we discuss the current state of drug discovery targeting E3 ligases and the opportunities and challenges that it provides.

Structure of the cyclic peptide [W8S]contryphan Vn: effect of the tryptophan/serine substitution on trans-cis proline isomerization.

The structural characterization of [W8S]contryphan Vn, an analogue of Contryphan Vn with tryptophan 8 substituted with a serine residue (W8S), was performed by NMR spectroscopy, molecular dynamics simulations and fluorescence spectroscopy. Contryphan Vn, a bioactive cyclic peptide from the venom of the cone snail Conus ventricosus, contains an S-S bridge between two cysteines and a D-tryptophan. Like other Contryphans, [W8S]contryphan Vn has proline 7 isomerized trans, while the proline 4 has nearly equivalent populations of cis and trans configurations. The thermodynamic and kinetic parameters of the trans-cis isomerization of proline 4 were measured. The isomers of [W8S]contryphan Vn with proline 4 in cis and trans show structural differences. The absence of the salt bridge between the same Asp2 and Lys6, present in Contryphan Vn, may be attributed to the lack of the hydrophobic side chain of Trp8 where it likely protects the electrostatic interactions. These results may contribute to identifying, in these cyclic peptides, the structural determinants of the mechanism of proline trans-cis isomerization, this being also an important step in protein folding.