Synthetic Dipeptide Research Articles

Reappraisal of Pidotimod: an immunomodulatory agent with 30-year evidence.

Pidotimod (3-L-pyroglutamyl-L-thiaziolidine-4-carboxylic acid) is a synthetic dipeptide with immunomodulatory properties that is indicated for use in adults and children over 3 years of age with documented cell-mediated immunodepression during respiratory and urinary tract infections. Infections are associated with an immune response that helps fight pathogens. In this scenario, inflammatory events occur to improve the antimicrobial reaction. However, defective immunity and/or sustained inflammation may adversely affect the course of the infection. Thus, modulating immune function could be a valuable option in managing patients with infections. The multifaceted mechanism of action of Pidotimod enables it to modulate innate and adaptive immunity. Extensive evidence about Pidotimod, accumulated over the last 30 years, has provided much data on the prevention of recurrent respiratory infections in susceptible children and respiratory exacerbations in patients with chronic bronchitis. Recent studies provide interesting information on how Pidotimod affects the metabolomic profiles of patients with bronchiectasis, clinical and immunological outcomes of elderly patients with pneumonia, clinical and cellular changes in patients with allergic rhinitis and asthma, and beneficial effects on cytokines and humoral immunity in human immunodeficiency virus (HIV) patients. Preliminary experience suggests that Pidotimod can shorten the duration of COVID-19 infection and reduce clinical severity by modulating the immune response, as well as prevent vaccination-related adverse events. In conclusion, the immunomodulatory properties of Pidotimod indicate that it may be a valuable option in managing patients with respiratory infections and other immune-mediated disorders, including allergy, chronic obstructive pulmonary disease, and asthma.

Phase II trial of BXCL701 and pembrolizumab in patients with metastatic pancreatic ductal adenocarcinoma (EXPEL-PANC): Preliminary findings.

LBA4132 Background: Pancreatic ductal adenocarcinoma (PDAC) has limited therapeutic options and is thought to be a “cold” tumor that does not respond to immunotherapy, due to a tumor microenvironment (TME) consisting of a desmoplastic stroma and poor T cell infiltrate. BXCL701 is an oral synthetic dipeptide that competitively inhibits dipeptidyl peptidases DPP4, DPP8, DPP9 and fibroblast activation protein (FAP). BXCL701 exerts antitumor activity via inhibition of DPP8/9, which is associated with induction of proinflammatory cytokines, as well as inhibition of FAP, which disrupts tumor-stromal interactions. Preclinical xenograft models demonstrate synergy between BXCL701 and PD-1 blockade, reducing tumor growth and promoting an increase in intratumoral CD4+ and CD8+ T cells, macrophages and NK cells, with induction of host-protective immunity. Methods: This is a phase II trial of BXCL701 in patients with metastatic PDAC (mPDAC) following progression on 1 line of treatment for advanced disease and amenable to serial biopsies. BXCL701 is administered at 0.2 mg PO BID days 1-7 and 0.3 mg BID days 8-14 during cycle 1 (21 days) followed by 0.3 mg BID days 1-14 every 21 days in all other cycles, given with pembrolizumab 200 mg IV every 21 days (all cycles). The primary objective is to determine the 18-week progression-free survival rate (PFS18weeks). We estimate that historical 2nd-line PFS18weeks is 30% or less; using a Simon’s two-stage (minimax) design, a type I error rate of 0.05 and power of 80% if the true rate is 50%, we will need 19 patients in stage 1 and 20 in stage 2 (39 total). There is a safety lead-in phase of 6 patients. We plan to enroll 43 patients to account for a predicted 10% drop out of unevaluable patients. Correlative pharmacodynamic studies include imaging mass cytometry to examine 36 markers of the PDAC TME in tissue biopsies, as well as blood-based analyses of KRAS circulating tumor DNA, circulating markers of fibrosis, and IL-6. Enrollment began in Q3 2023 (NCT05558982). Results: Six patients have enrolled, 3 women and 3 men, median age 57.5 (range 37-80). One patient was progression-free at 18 weeks and 1 patient had stable disease (SD) at 9 weeks, not yet evaluable for the 18-week landmark. Objective response rate is 16% and disease control rate is 50% (RECIST: 1 partial response, -41%, and 2 patients with SD, -18% and 0%). Three patients had significant reductions in CA19-9 from baseline (-100%, -73%, and -97%). Median PFS and overall survival have not been reached (NR, 95% CI 1.45 months-NR and 0.92 months-NR, respectively). There have been no serious treatment-related safety events. The safety lead-in will complete after the next patient completes the 6-week safety window (1 patient was unevaluable). Conclusions: BXCL701 plus pembrolizumab is well-tolerated and shows early signs of potential clinical activity in patients with mPDAC refractory to chemotherapy. Clinical trial information: NCT05558982 .

Synthesis of Peptides by Reactive Extrusion. Application to the Continuous and Solventless Preparation of Aspartame.

AbstractThe solventless synthesis of peptides from unactivated amino acids was optimized in a twin‐screw extruder (TSE). The general method enabled the preparation of dipeptides and tripeptides with no epimerization. This flow mechanochemistry approach was applied to the synthesis of aspartame, a synthetic dipeptide, prepared in a continuous mode on large scale and with high productivity.

Emergence of Dynamic Instability by Hybridizing Synthetic Self-Assembled Dipeptide Fibers with Surfactant Micelles.

Supramolecular chemistry currently faces the challenge of controlling nonequilibrium dynamics such as the dynamic instability of microtubules. In this study, we explored the emergence of dynamic instability through the hybridization of peptide-type supramolecular nanofibers with surfactant micelles. Using real-time confocal imaging, we discovered that the addition of micelles to nanofibers induced the simultaneous but asynchronous growth and shrinkage of nanofibers during which the total number of fibers decreased monotonically. This dynamic phenomenon unexpectedly persisted for 6 days and was driven not by chemical reactions but by noncovalent supramolecular interactions between peptide-type nanofibers and surfactant micelles. This study demonstrates a strategy for inducing autonomous supramolecular dynamics, which will open up possibilities for developing soft materials applicable to biomedicine and soft robotics.

A closer look at ligand specificity for cellular activation of NOD2 with synthetic muramyl dipeptide analogues.

To further understand the specificity of muramyl dipeptide (MDP) sensing by NOD2, we evaluated the compatibility of synthetic MDP analogues for cellular uptake and NAGK phosphorylation, the pre-requisite steps of intracellular NOD2 activation. Our results revealed that these two prior steps do not confer ligand stereoselectivity; yet NAGK strictly discriminates against the disaccharide NOD2 agonists for phosphorylation in vitro, despite it being indispensable for the cellular NOD2-stimulating effects of these analogues, implying potential glycosidase cleavage as a novel intermediate step for cellular activation of NOD2.

C9orf72 dipeptides trigger neuroinflammation via the NLRP3 inflammasome

AbstractBackgroundThe most common cause of both frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) is a repeat expansion in C9orf72. The expansion is translated to produce five toxic dipeptides (DPRs), which aggregate in patient brain. Neuroinflammation is observed in patients, with increased microglial activation and elevated levels of pro‐inflammatory cytokines in CSF. However, the causes of this inflammation are unknown. Here, we investigated whether DPRs activate immune cells and contribute to neuroinflammation.MethodPrimary macrophages or microglia were treated with synthetic C9orf72 dipeptides and levels of pro‐inflammatory cytokines released into the culture media were analysed by ELISA and Western blot. Cytotoxicity was measured by LDH assay. Mouse organotypic hippocampal slice cultures were also treated with DPRs and cytokine release measured.ResultOne of the DPRs, GR, caused secretion of the pro‐inflammatory cytokine, interleukin (IL)‐1β, from macrophages, microglia and in hippocampal slice cultures. GR also caused significant toxicity. IL‐1β secretion was prevented by pre‐treatment with MCC950, a drug which inhibits the NLRP3 inflammasome. The inflammasome is a protein complex which forms in the cytosol of immune cells is response to “danger” stimuli, resulting in cleavage of caspase‐1 and subsequent release of IL‐1β. We confirmed that caspase‐1 was cleaved in macrophages following GR treatment. In addition, knockout of NLRP3 prevented IL‐1β secretion in response to GR. Finally, we showed that several drugs which are clinically approved for use in other conditions rescued GR‐induced NLRP3 activation.ConclusionHere, we demonstrate that C9orf72 dipeptides could directly contribute to neuroinflammation in FTD/ALS via the NLRP3 inflammasome. Furthermore, we demonstrate that this can be inhibited pharmacologically, highlighting NLRP3 as a potential therapeutic target in FTD/ALS.

Aspartame, a Synthetic Dipeptide Mediated Biochemical and Histopathological Alterations in Hepato-nephric Tissues of Mice and Pharmaceutical Intervention by Sesame Oil

Aspartame, a Synthetic Dipeptide Mediated Biochemical and Histopathological Alterations in Hepato-nephric Tissues of Mice and Pharmaceutical Intervention by Sesame Oil

Consolidation of experimental fractures during intramedullary osteosynthesis in rats using Collapan-L and Thymogen

The article presents the results of treatment of experimental femur fractures of laboratory animals (outbred rats) against the background of the use of a biocomposite material based on synthetic hydroxyapatite, collagen and lincomycin hydrochloride (collapan-L) and an immunotropic drug from the group of synthetic dipeptides (thymogen).During the experiment, biochemical, radiological and histological studies were carried out. It was shown that during the first week after the fracture, there were no significant changes in the groups. While for the period from 7 to 21 days (according to X-ray data), processes of reparative osteogenesis proceeded more intensively (on the 14th day) in the 1st, 4th, and 5th groups than it did in the control, and 2nd and 3rd experimental groups. These changes are consistent with biochemical blood tests. The radiographs obtained on the 28th day had no significant differences.On the 45th day after the surgery, the fibrocartilage callus forming in the group where both drugs were used simultaneously (thymogen ‒ immediately after the surgery) proceeded more pronouncedly compared to other groups. Thus, on radiographs, the fracture line was barely noticeable, which indicates more actively occurring processes of osteogenesis. These assumptions are also consistent with the level of calcium in the blood serum, as well as with histological studies. Thus, on the 45th day after the surgery, the fibrocartilage callus consisted of 60.2 ± 2.1% of bone tissue, which was 4.9% higher than in the control group and 9.39% higher than the impact of the mechanical factor (rupture resistance).Thus, the results of the studies on rats with experimental femoral fractures made it possible to establish that the combined use of the tested drugs contributes to a more intensive process of reparative osteogenesis (in the 4th experimental group).

Phase II trial of BXCL701 and pembrolizumab in patients with metastatic pancreatic ductal adenocarcinoma (EXPEL-PANC).

TPS4194 Background: Pancreatic ductal adenocarcinoma (PDAC) has limited therapeutic options and is thought to be a “cold” tumor that does not respond to immunotherapy, due to a tumor microenvironment (TME) consisting of a desmoplastic stroma and poor T cell infiltrate. BXCL701 is an oral synthetic dipeptide that competitively inhibits dipeptidyl peptidases DPP4, DPP8, DPP9 and fibroblast activation protein (FAP). BXCL701 exerts antitumor activity via inhibition of DPP8/9, which is associated with induction of proinflammatory cytokines, as well as inhibition of FAP, which disrupts tumor-stromal interactions. Preclinical xenograft models demonstrate synergy between BXCL701 and an anti-PD-1 monoclonal antibody, reducing tumor growth and promoting an increase in intratumoral CD4+ and CD8+ T cells and NK cells. The combination of BXCL701 and pembrolizumab is already being studied in a phase II trial in patients with prostate cancer (NCT03910660) and a multi-cohort basket study (NCT04171219). Methods: This is a phase II trial of BXCL701 administered at 0.2 mg PO BID days 1-7 and 0.3 mg BID days 8-14 during cycle 1 (21 days) followed by 0.3 mg BID days 1-14 every 21 days in all subsequent cycles, given with pembrolizumab 200 mg IV every 21 days (all cycles). The primary objective is to determine the 18-week progression-free survival rate (PFS18weeks) in patients with metastatic PDAC treated in the second-line setting. We estimate that historical 2nd-line PFS18weeks is 30% or less; using a Simon’s two-stage (minimax) design, a type I error rate of 0.05 and power of 80% when the true response rate of 50%, we will need 19 patients in stage 1 and 20 in stage 2 (a total of 39). We plan to enroll 43 patients to account for a predicted 10% drop out of unevaluable patients. Eligible patients must have metastatic PDAC and disease progression or intolerance to only 1 line of therapy for metastatic disease, without exposure to prior immunotherapy. Patients must have measurable disease amenable to serial biopsies. Correlative pharmacodynamic studies include imaging mass cytometry to examine 34 markers of the PDAC TME in tissue biopsies, as well as blood-based analyses of KRAS circulating tumor DNA, circulating markers of fibrosis, and IL-6. Enrollment began in Q1 2023. Clinical trial information: NCT05558982 .

State and prospect of application immunocorrection therapy in castration of piglets and 5‐month‐old boars in pig breeding farm

One of the most widespread surgical interventions is the castration of piglets and 5‐month‐old boars without breeding selection being a criterion in conditions of intensive development of the pig breeding industry. This work aims to use immunocorrection therapy, methods and system of their use, technological simplicity, and efficiency in boars castration in the pig breeding industry.The research was done with the help of the pig breeding industrial complex and Kostroma State Agricultural Academy. 87 hybrid boars at the age of 5 months (F1) were used. In total, two cycles of experiments were carried out. For the first cycle, 49 animals were divided into 7 groups (n=7) to define the effectiveness of using different methods and schemes of immunocorrection therapy in post‐castration complications and its impact on product indicators. In the second cycle of the research, performance indicators of pig‐meat production were studied on the control group (n = 9) and experimental group (n = 29) of boars by using the best immunocorrection therapy according to the first cycle of the research.Thymogen injection is a synthetic immunomodulatory dipeptide, an analog of one of the thymus hormones. Ribav (mycovit) is a natural origin alcohol‐based drug for local use immunocorrectors. Both immunocorrectors were used for the research.Research results: decrease of complications with an open surgical way of castration in the experimental group on 28,57% (control group 57,14%) especially in the group treated with thymogen and Ribav in post castration time; similar results were received considering local reactions as hyperemia and less edema; the same indicators were higher than in the control group at the closed technique of castration. It can be caused by an immune reaction to suture material ‐ Polycon; Ribav was more effective at the open technique of castration than at closed, which is explained by the accessibility of local wound treatment; the average daily gain in live weight in the groups treated with thymogen and Ribav was 0.551 ± 0.13 and 0.618 ± 0.09 versus 0.406 ± 0.04 kg in control, meat yield rate 77.0 ± 0.58 and 76.40 ± 0, 29 versus 76.85 ± 0.14 in control with 85.71% safety in the experimental groups versus 81.43% in control.Thus, Thymogen is preferable according to its manufacturability and cost indicators.In the second cycle of experiments with its use, the gain in live weight exceeded the indicators of the control group by 18.6% without significant differences in meat on the bone and its yield. At the same time, it should be noted that the safety of the livestock, as in the first series of experiments, was 7.6% higher in the experimental group (96.55 versus 88.89%).Thus, thymogen post castration injections or Ribav (mukovit) local irrigation treatment with a five‐day course at a postoperative period reduce the risk of post castration complications on 14,28 ‒ 28,57% in the condition of pig breeding intensification, reduce emersion of local reactions, increases the safety among 5‐month‐old boars by 14.28 ‐ 28.57%.As a result, it is better to use treatment schemes based on using Ribave with the closed technique of castration on small pig farms when personal care is possible.

Insight into Isomeric Diversity of Glycated Amino Acids in Maillard Reaction Mixtures.

Maillard reactions generate a wide array of amino acid- and sugar-derived intermediates; the isomeric mixtures of glycated amino acids are of particular interest. Excluding stereoisomers, regioisomers, and various anomers, most amino acids can form two monoglycated and three N,N-diglycated isomers when reacted with sugars during the Maillard reaction. Using synthetic Schiff bases and Amadori compounds as standards, we have demonstrated that diagnostic ions obtained from MS/MS fragmentations in negative ionization mode can be used effectively for the discrimination between glucose-derived Schiff bases and their corresponding Amadori compounds in both mono- and diglycated forms. The utilization of these diagnostic ions and isotopic labeling in the glycine/glucose model system revealed that milling glucose/glycine mixtures for 30 min/30 Hz at ambient temperature produced monoglycated glycine in equal proportions of Amadori and Schiff base forms, whereas diglycated glycine was a mixture of the three isomers: Schiff-Schiff, Schiff-Amadori, or Amadori-Amadori in approximately equal molar proportions. The above results were further corroborated using a synthetic histidine Amadori product, N,N-difructosyl-β-alanine, dipeptides, and ribose. Using mechanochemistry as a convenient synthetic tool in combination with MS/MS diagnostic ions, the isomeric diversity of the early stages of the Maillard reaction can be revealed.

Fluconazole resistant Candida auris clinical isolates have increased levels of cell wall chitin and increased susceptibility to a glucosamine-6-phosphate synthase inhibitor

In 2009 Candida auris was first isolated as fungal pathogen of human disease from ear canal of a patient in Japan. In less than a decade, this pathogen has rapidly spread around the world and has now become a major health challenge that is of particular concern because many strains are resistant to multiple class of antifungal drugs. The lack of available antifungals and rapid increase of this fungal pathogen provides an incentive for the development of new and more potent anticandidal drugs and drug combinatorial treatments. Here we have explored the growth inhibitory activity against C. auris of a synthetic dipeptide glutamine analogue, L-norvalyl-N3-(4-methoxyfumaroyl)-L-2,3- diaminopropanoic acid (Nva-FMDP), that acts as an inhibitor of glucosamine-6-phosphate (GlcN-6-P) synthase - a key enzyme in the synthesis of cell wall chitin. We observed that in contrast to FLC susceptible isolates of C. auris, FLC resistant isolates had elevated cell wall chitin and were susceptible to inhibition by Nva-FMDP. The growth kinetics of C. auris in RPMI-1640 medium revealed that the growth of FLC resistant isolates were 50–60% more inhibited by Nva-FMDP (8 μ g/ml) compared to a FLC susceptible isolate. Fluconazole resistant strains displayed increased transcription of CHS1, CHS2 and CHS3, and the chitin content of the fluconazole resistant strains was reduced following the Nva-FMDP treatment. Therefore, the higher chitin content in FLC resistant C. auris isolates may make the strain more susceptible to inhibition of the antifungal activity of the Nva-FMDP peptide conjugate.

Characterization of gluten-degrading prolyl endoprotease from Thermococcus kodakarensis.

There is increasing interest in gluten-degrading enzymes for use during food and drink processing. The industrially available enzymes usually work best at low to ambient temperatures. However, food manufacturing is often conducted at higher temperatures. Therefore, thermostable gluten-degrading enzymes are of great interest. We have identified a new thermostable gluten-degrading proline-specific prolyl endoprotease from the archaea Thermococcus kodakarensis. We then cloned and expressed it in Escherichia coli. The prolyl endoprotease was found to have a size of 70.1kDa. The synthetic dipeptide Z-Gly-Pro-p-nitroanilide was used to characterize the prolyl endoprotease and it had maximum activity at pH 7 and 77°C. The Vmax, Km and kcat values of the purified prolyl endoprotease were calculated to be 3.14mM/s, 1.10mM and 54 s-1, respectively. When the immunogenic gluten peptides PQPQLPYPQPQLPY (α-gliadin) and SQQQFPQPQQPFPQQP (γ-hordein) were used as substrates, the prolyl endoprotease was able to degrade these. Furthermore, gluten in wort was reduced when the prolyl endoprotease was used during mashing of barley malt. The discoveries open up new food processing possibilities and further the understanding of proline-specific protease diversity.

Isolation and Characterization of Novel Degradation Product of Pidotimod

A synthetic dipeptide, pidotimod (3-L-pyroglutamyl-L-thiaziolidine-4-carboxylic acid, PTD) is an immunomodulator and it possesses anti-infective activity against variety of infections. The objective of this study was to explore into and identify pidotimod’s major degradation product. Pidotimod was degraded in ICH prescribed stress conditions. pidotimod was degraded 90% in 1.0 N NaOH at 80 ºC for 6 h and the degradation product (DP2) was determined using HPLC. The HPLC method was used to separate the primary and subsidiary degradation products and carried out on a C18 column with a mobile phase of ammonium acetate buffer (pH 4.5; 10 mM) and MeOH/ACN (90:10 v/v) at a ratio of 97:03 v/v at 40 ºC, flow rate of 1.0 mL/min, and detection at 215 nm. IR, NMR and LC-MS-MS were used to identify degradation product (DP2) as 3,8-dihydroxy-tetrahydro-bisthiazolo[3,4,4a,3′,4′- d]pyrazine-5,10-dione.

Pidotimod increases inflammation in wounded zebrafish embryos

Pidotimod (PDT) is a synthetic dipeptide molecule which can improve immune responses in mice and humans, protecting hosts from infection. However, the exact mechanism of protection remains ill-defined. The effect of pidotimod has not yet been investigated in the inflammatory response of zebrafish. In this study, we used tail wound and infection models of zebrafish to study the effect of PDT on inflammation. We found that zebrafish larvae were sensitive to PDT immersion causing toxicity at doses above 50 μg/mL. The tail wound assay showed that PDT increased the recruitment of neutrophils and macrophages to the wound site and promoted the transcription of the pro-inflammatory cytokine il1b. However, we did not observe protection of uropathogenic Escherichia coli or Mycobacterium marinum infected zebrafish larvae following PDT treatment. This study provides a new platform for PDT research, which is worthy of further research to identify further effects of PDT therapy.

In vitro and in silico characterization of adiponectin-receptor agonist dipeptides

The aim of this study is to develop a dipeptide showing an adiponectin receptor 1 (AdipoR1) agonistic effect in skeletal muscle L6 myotubes. Based on the structure of the AdipoR1 agonist, AdipoRon, 15 synthetic dipeptides were targeted to promote glucose uptake in L6 myotubes. Tyr-Pro showed a significant increase in glucose uptake among the dipeptides, while other dipeptides, including Pro-Tyr, failed to exert this effect. Tyr-Pro induces glucose transporter 4 (Glut4) expression in the plasma membrane, along with adenosine monophosphate-activated protein kinase (AMPK) activation. In AdipoR1-knocked down cells, the promotion by Tyr-Pro was ameliorated, indicating that Tyr-Pro may directly interact with AdipoR1 as an agonist, followed by the activation of AMPK/Glut4 translocation in L6 myotubes. Molecular dynamics simulations revealed that a Tyr-Pro molecule was stably positioned in the two potential binding pockets (sites 1 and 2) of the seven-transmembrane receptor, AdipoR1, anchored in a virtual 1-palmitoyl-2-oleoyl-phosphatidylcholine membrane. In conclusion, we demonstrated the antidiabetic function of the Tyr-Pro dipeptide as a possible AdipoR1 agonist.

Reparative histogenesis of bone tissue in femoral fractures in rats using biocomposite material along with immunocorrection

The paper studies the effect of the RVI biocomposite material belonging to the group of osteoplastic biocomposite materials, the RV-2 immunomodulator – a synthetic dipeptide inducing an immunocorrective effect, and combinations of these drugs on the reparative histogenesis of bone tissue in femoral fractures in rats. It was found that the remodeling of the primary bone callus into the secondary one in the fracture of the studied animals was of a diverse nature. This process was the most pronounced in the group where the components were used in complex, i.e. the bone defect was filled with RVI during the surgery, as well as RV-2 was injected intramuscularly to rats at a dose of 10 mcg per 1 kg of live weight for five days, starting immediately after the surgery. Well-formed coarse-fibrous connective tissue callus was recorded in animals of this group. The connective tissue was stained more intensely which indicates a denser arrangement of fibers in the callus. Focal cartilage tissue spanning bone fragments was observed within the callus. At the periphery of the site the cartilaginous callus was subjected to endochondral ossification with replacement by coarse-fibrous trabeculae with elements of lamellar bone tissue having haversian canals in the center. The inter-girdle spaces were filled with elements of the myeloid bone marrow in the forming bone tissue. Markedly proliferated osteoblasts were visible in the cambial layer of the periosteum. The bone tissue ratio increased up to (60.21 ± 2.62)%, which significantly exceeded the same indicator in the control group and in all experimental groups. The low content of connective tissue and the high ratio of bone tissue indicated more active osteogenesis processes and reparative regeneration in comparison with other groups.

Effect of dipeptide on intestinal peptide transporter 1 gene expression: An evaluation using primary cultured chicken intestinal epithelial cells.

Peptide transporter 1 (PepT1) is a transporter responsible for absorbing dipeptide and tripeptide in enterocytes and is upregulated by dipeptide in mammals. It has not been certain whether intestinal PepT1 expression is responsive to dipeptides in chickens because of the lack of in vitro study using the cultured enterocytes. This study established a primary culture model of chicken intestinal epithelial cells (IECs) in two‐dimensional monolayer culture using collagen gel by which the response of chicken PepT1 gene expression to dipeptide stimuli was evaluated. The cultured chicken IECs showed the epithelial‐like morphology attached in a patch‐manner and exhibited positive expression of cytokeratin and epithelial cadherin, specific marker proteins of epithelial cells. Moreover, the chicken IECs exhibited the gene expression of intestinal cell type‐specific marker, villin1, mucin 2, and chromogranin A, suggesting that the cultured IECs were composed of enterocytes as well as goblet and enteroendocrine cells. PepT1 gene expression was significantly upregulated by synthetic dipeptide, glycyl‐l‐glutamine, in the cultured IECs. From the results, we herein suggested that dipeptide is a factor upregulating PepT1 gene expression in chicken IECs.

In vitro and in vivo testing of nanofibrous membranes doped with alaptide and L-arginine for wound treatment

We have prepared a candidate biocompatible construct for skin wound healing based on electrospun polycaprolactone (PCL) nanofibrous membranes. The membrane material was loaded either with L-arginine or with alaptide, or with a mixture of both bioactive components. Alaptide is a spirocyclic synthetic dipeptide, an analogue of melanocyte-stimulating hormone release-inhibiting factor. L-arginine is an amino acid with a basic guanidine side chain. It is a direct precursor of nitric oxide, which plays a pivotal role in skin repair. The presence and the distribution of the additives were proved with high-performance liquid chromatography, Fourier-transform infrared spectroscopy and Raman spectroscopy. The influence of L-arginine and alaptide on the morphology of the membrane was characterized using scanning electron microscopy. No statistically significant correlation between fiber diameter and drug concentration was observed. The membranes were then tested in vitro for their cytotoxicity, using primary human dermal fibroblasts, in order to obtain the optimal concentrations of the additives for in vivo tests in a rat model. The membranes with the highest concentration of L-arginine (10 wt. %) proved to be cytotoxic. The membranes with alaptide in concentrations from 0.1 to 2.5 wt.%, and with the other L-arginine concentrations (1 and 5 wt.%), did not show high toxicity. In addition, there was no observed improvement in cell proliferation on the membranes. The in vivo experiments revealed that membranes with 1.5 wt.% of alaptide or with 1.5 wt.% of alaptide in combination with 5 wt.% of L-arginine markedly accelerated the healing of skin incisions, and particularly the healing of skin burns, i.e. wounds of relatively large extent. These results indicate that our newly-developed nanofibrous membranes are promising for treating wounds with large damaged areas, where a supporting material is needed.

Narrow-band phototherapy and sodium salt of the synthetic diamide gamma-D-glutamyl-D-tryptophan as a combined method of psoriasis treatment: comprehensive and comparative evaluation

The paper presents the results of a comparative study of various immunosuppressive treatment methods of medium severity vulgar psoriasis (PASI from 10 to 20). The parameters used were PASI indices, p53 apoptosis marker expression indices, and expression indices of T-regulatory cell marker Foxp3. The study involved 96 patients, from 18 to 60 years old, divided by randomization into three groups. Patients in the 1st group (n = 32) received narrow-band phototherapy according to a 4-day regimen. We used the Waldmann UV-7002K instrument and F79/120W-TL01 lamps generating radiation with a maximum emission at a wavelength of 311 nm. We started with a dose of 0.05–0.1 J/cm2, subsequently increased by 0.05–0.1 J/cm2 for each procedure, in the absence of erythema. In total, patients received 12–15 procedures, with a course dose of 4.7–7.65 J/cm2. In the 2nd group (n = 32), patients were treated with the sodium salt of the synthetic gamma-D-glutamyl-D tryptophan dipeptide. Patients of the 3rd group (n = 32) underwent combination therapy, which included both methods described above. As a result of treatment, the greatest decrease in the PASI index, as well as the level of expression of the marker of Foxp3 T-regulatory cells, was observed among patients who received the combined regimen. The data obtained indicate the high therapeutic efficacy of narrow-band phototherapy in combination with the sodium salt of the synthetic gamma-D glutamyl-D tryptophan synthetic dipeptide. It was also noted that the immunohistochemical marker p53, reflecting the cellular renewal of psoriatic papule keratinocytes, is an informative molecular-cellular indicator of the effectiveness of narrow-band phototherapy.