Structure-activity relationship in NOD2 agonistic muramyl dipeptides.

Nucleotide-binding oligomerization domain 2 (NOD2) is a receptor of the innate immune system that is capable of perceiving bacterial and viral infections. Muramyl dipeptide (MDP, N-acetyl muramyl L-alanyl-d-isoglutamine), identified as the minimal immunologically active component of bacterial cell wall peptidoglycan (PGN) is recognized by NOD2. In terms of biological activities, MDP demonstrated vaccine adjuvant activity and stimulated non-specific protection against bacterial, viral, and parasitic infections and cancer. However, MDP has certain drawbacks including pyrogenicity, rapid elimination, and lack of oral bioavailability. Several detailed structure-activity relationship (SAR) studies around MDP scaffolds are being carried out to identify better NOD2 ligands. The present review elaborates a comprehensive SAR summarizing structural aspects of MDP derivatives in relation to NOD2 agonistic activity.

NOD2 in monocytes negatively regulates macrophage development through TNFalpha.

Objective: It is believed that intestinal recruitment of monocytes from Crohn's Disease (CD) patients who carry NOD2 risk alleles may repeatedly give rise to recruitment of pathogenic macrophages. We investigated an alternative possibility that NOD2 may rather inhibit their differentiation from intravasating monocytes. Design: The monocyte fate decision was examined by using germ-free mice, mixed bone marrow chimeras and a culture system yielding macrophages and monocyte-derived dendritic cells (mo-DCs). Results: We observed a decrease in the frequency of mo-DCs in the colon of Nod2-deficient mice, despite a similar abundance of monocytes. This decrease was independent of the changes in the gut microbiota and dysbiosis caused by Nod2 deficiency. Similarly, the pool of mo-DCs was poorly reconstituted in a Nod2-deficient mixed bone marrow (BM) chimera. The use of pharmacological inhibitors revealed that activation of NOD2 during monocyte-derived cell development, dominantly inhibits mTOR-mediated macrophage differentiation in a TNFα-dependent manner. These observations were supported by the identification of a TNFα-dependent response to muramyl dipeptide (MDP) that is specifically lost when CD14-expressing blood cells bear a frameshift mutation in NOD2. Conclusion: NOD2 negatively regulates a macrophage developmental program through a feed-forward loop that could be exploited for overcoming resistance to anti-TNF therapy in CD.

A probiotic bi-functional peptidoglycan hydrolase sheds NOD2 ligands to regulate gut homeostasis in female mice.

Secreted proteins are one of the direct molecular mechanisms by which microbiota influence the host, thus constituting a promising field for drug discovery. Here, through bioinformatics-guided screening of the secretome of clinically established probiotics from Lactobacillus, we identify an uncharacterized secreted protein (named LPH here) that is shared by most of these probiotic strains (8/10) and demonstrate that it protects female mice from colitis in multiple models. Functional studies show that LPH is a bi-functional peptidoglycan hydrolase with both N-Acetyl-ß-D-muramidase and DL-endopeptidase activities that can generate muramyl dipeptide (MDP), a NOD2 ligand. Different active site mutants of LPH in combination with Nod2 knockout female mice confirm that LPH exerts anti-colitis effects through MDP-NOD2 signaling. Furthermore, we validate that LPH can also exert protective effects on inflammation-associated colorectal cancer in female mice. Our study reports a probiotic enzyme that enhances NOD2 signaling in vivo in female mice and describes a molecular mechanism that may contribute to the effects of traditional Lactobacillus probiotics.

Muramyl dipeptide CD10 monoclonal antibody immunoconjugates inhibited acute leukemia in nude mice.

Minimal residual disease (MRD) is one of the causes of leukemia recurrence. Previously, we developed anti-CD10 mAb conjugated to muramyl dipeptide immunoconjugate (MDP-Ab) for immune enhancement. The present study aimed to investigate anti-leukemia effect of MDP-Ab administered via different methods in leukemia ectopic graft nude mouse model. BALB/c nude mice were injected with Nalm-6 cells subcutaneously to establish leukemia xenografts in nude mice as a model. MDP-Ab or/and human lymphocytes (LYM) was injected into different sites of the nude mice. Immunohistochemistry staining of CDs in the bone marrow, liver and spleen was performed. IFN-γ was detected by ELISA. We detected the metastasis of leukemia cells to the liver, spleen and bone marrow in nude mouse leukemia model. MDP-Ab and LYM inhibited the growth of tumors, and simultaneous injection of MDP-Ab and LYM into the tumor inhibited the growth of tumors. IFN-γ levels in MDP-Ab (ca) + h-LYM (ca) group, MDP-Ab (ca) + h-LYM (ip) group, MDP-Ab (iv) + h-LYM (ip) group and PBS (ca) + h-LYM (ca) group were significantly higher than those in control group, while IFN-γ level in MDP-Ab (ca) + h-LYM (ca) group was the highest. Moreover, MDP-Ab and h-LYM promoted the expression of hCD4 and hCD8, with the highest expression in MDP-Ab (ca) + h-LYM (ca) group. In conclusion, MDP-Ab effectively promoted the production of IFN-γ, enhanced the antitumor immunity of T lymphocytes and inhibited leukemia.

Muramyl dipeptide alleviates estrogen deficiency-induced osteoporosis through canonical Wnt signaling.

Wnt signaling is a positive regulator of bone formation through the induction of osteoblast differentiation and down-regulation of osteoclast differentiation. We previously reported that muramyl dipeptide (MDP) increases bone volume by increasing osteoblast activity and attenuating osteoclast activity in receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoporotic model mice. In this study, we investigated whether MDP could alleviate post-menopausal osteoporosis through Wnt signaling regulation in an ovariectomy (OVX)-induced mouse osteoporosis model. MDP-administered OVX mice exhibited higher bone volume and bone mineral density than mice of the control group. MDP significantly increased P1NP in the serum of OVX mice, implying increased bone formation. The expression of pGSK3ß and ß-catenin in the distal femur of OVX mice was lower than that in the distal femur of sham-operated mice. Yet, the expression of pGSK3ß and ß-catenin was increased in MDP-administered OVX mice compared with OVX mice. In addition, MDP increased the expression and transcriptional activity of ß-catenin in osteoblasts. MDP inhibited the proteasomal degradation of ß-catenin via the down-regulation of its ubiquitination by GSK3ß inactivation. When osteoblasts were pretreated with Wnt signaling inhibitors, DKK1 or IWP-2, the induction of pAKT, pGSK3ß, and ß-catenin was not observed. In addition, nucleotide oligomerization domain-containing protein 2-deficient osteoblasts were not sensitive to MDP. MDP-administered OVX mice exhibited fewer tartrate-resistant acid phosphatase (TRAP)-positive cells than did OVX mice, attributed to a decrease in the RANKL/OPG ratio. In conclusion, MDP alleviates estrogen deficiency-induced osteoporosis through canonical Wnt signaling and could be an effective therapeutic for the treatment of post-menopausal bone loss. © 2023 The Pathological Society of Great Britain and Ireland.

Microbe-mediated intestinal NOD2 stimulation improves linear growth of undernourished infant mice.

The intestinal microbiota is known to influence postnatal growth. We previously found that a strain of Lactiplantibacillus plantarum (strain LpWJL) buffers the adverse effects of chronic undernutrition on the growth of juvenile germ-free mice. Here, we report that LpWJL sustains the postnatal growth of malnourished conventional animals and supports both insulin-like growth factor-1 (IGF-1) and insulin production and activity. We have identified cell walls isolated from LpWJL, as well as muramyl dipeptide and mifamurtide, as sufficient cues to stimulate animal growth despite undernutrition. Further, we found that NOD2 is necessary in intestinal epithelial cells for LpWJL-mediated IGF-1 production and for postnatal growth promotion in malnourished conventional animals. These findings indicate that, coupled with renutrition, bacteria cell walls or purified NOD2 ligands have the potential to alleviate stunting.

Blau syndrome NOD2 mutations result in loss of NOD2 cross-regulatory function.

The studies described here provide an analysis of the pathogenesis of Blau syndrome and thereby the function of NOD2 as seen through the lens of its dysfunction resulting from Blau-associated NOD2 mutations in its nucleotide-binding domain (NBD). As such, this analysis also sheds light on the role of NOD2 risk polymorphisms in the LRR domain occurring in Crohn's disease. The main finding was that Blau NOD2 mutations precipitate a loss of canonical NOD2 signaling via RIPK2 and that this loss has two consequences: first, it results in defective NOD2 ligand (MDP)-mediated NF-κB activation and second, it disrupts NOD2-mediated cross-regulation whereby NOD2 downregulates concomitant innate (TLR) responses. Strong evidence is also presented favoring the view that NOD2-mediated cross-regulation is under mechanistic control by IRF4 and that failure to up-regulate this factor because of faulty NOD2 signaling is the proximal cause of defective cross-regulation and the latter's effect on Blau syndrome inflammation. Overall, these studies highlight the role of NOD2 as a regulatory factor and thus provide additional insight into its function in inflammatory disease. Mutations in the nucleotide binding domain of the CARD15 (NOD2) gene underlie the granulomatous inflammation characterizing Blau syndrome (BS). In studies probing the mechanism of this inflammation we show here that NOD2 plasmids expressing various Blau mutations in HEK293 cells result in reduced NOD2 activation of RIPK2 and correspondingly reduced NOD2 activation of NF-κB. These in vitro studies of NOD2 signaling were accompanied by in vivo studies showing that BS-NOD2 also exhibit defects in cross-regulation of innate responses underlying inflammation. Thus, whereas over-expressed intact NOD2 suppresses TNBS-colitis, over-expressed BS-NOD2 does not; in addition, whereas administration of NOD2 ligand (muramyl dipeptide, MDP) suppresses DSS-colitis in Wild Type (WT) mice it fails to do so in homozygous or heterozygous mice bearing a NOD2 Blau mutation. Similarly, mice bearing a Blau mutation exhibit enhanced anti-collagen antibody-induced arthritis. The basis of such cross-regulatory failure was revealed in studies showing that MDP-stimulated cells bearing BS-NOD2 exhibit a reduced capacity to signal via RIPK2 as well as a reduced capacity to up-regulate IRF4, a factor shown previously to mediate NOD2 suppression of NF-κB activation. Indeed, TLR-stimulated cells bearing a Blau mutation exhibited enhanced in vitro cytokine responses that are quieted by lentivirus transduction of IRF4. In addition, enhanced anti-collagen-induced joint inflammation in mice bearing a Blau mutation was accompanied by reduced IRF4 expression in inflamed joint tissue and IRF4 expression was reduced in MDP-stimulated cells from BS patients. Thus, inflammation characterizing Blau syndrome are caused, at least in part, by faulty canonical signaling and reduce IRF4-mediated cross-regulation.

Phosphorylation of muramyl peptides by NAGK is required for NOD2 activation.

Bacterial cell wall components provide various unique molecular structures that are detected by pattern recognition receptors (PRRs) of the innate immune system as non-self. Most bacterial species form a cell wall that consists of peptidoglycan (PGN), a polymeric structure comprising alternating amino sugars that form strands cross-linked by short peptides. Muramyl dipeptide (MDP) has been well documented as a minimal immunogenic component of peptidoglycan1-3. MDP is sensed by the cytosolic nucleotide-binding oligomerization domain-containing protein 24 (NOD2). Upon engagement, it triggers pro-inflammatory gene expression, and this functionality is of critical importance in maintaining a healthy intestinal barrier function5. Here, using a forward genetic screen to identify factors required for MDP detection, we identified N-acetylglucosamine kinase (NAGK) as being essential for the immunostimulatory activity of MDP. NAGK is broadly expressed in immune cells and has previously been described to contribute to the hexosamine biosynthetic salvage pathway6. Mechanistically, NAGK functions upstream of NOD2 by directly phosphorylating the N-acetylmuramic acid moiety of MDP at the hydroxyl group of its C6 position, yielding 6-O-phospho-MDP. NAGK-phosphorylated MDP-but not unmodified MDP-constitutes an agonist for NOD2. Macrophages from mice deficient in NAGK are completely deficient in MDP sensing. These results reveal a link between amino sugar metabolism and innate immunity to bacterial cell walls.

Effects of Liposome-Entrapped Muramyl Tripeptide Phosphatidylethanolamine (L-MTP-PE) on the Tumor Growth and Survival of Mice Bearing Syngeneic Tumor in Combination with a Chemotherapeutic or Immunomodulatory Agent.

Antitumor activities of L-MTP-PE (Liposome entrapped myuramyl tripeptide phosphatidylethanolamine) in the combination treatment with chemo- or immune-therapeutic antitumor agents against various syngeneic tumors were tested.Against Meth A fibrosarcoma solid tumor system, L-MTP-PE showed slight but statistically significant elongation of survival days against 5-FU monotherapy in spite of its non-effect on tumor growth, when combined with 5-FU. Against liver metastasis model of M5076 carcinoma, L-MTP-PE showed a tendency of elongation of survival days by its single drug treatment, however, elongation with statistical significance was observed in the combination treatment with 5-FU in comparison with control group.These data suggest that L-MTP-PE seems to elongate the survival days of the solid tumor bearing mice and the liver metastasis model basically due to its saving effect on chemotherapeutic drug-induced immunosuppression. In the combination with an immunotherapeutic agent in mice, TNF production induced by another biological response modifier OK-432 was potentiated when primed with L-MTP-PE. L-MTP-PE also potentiate the antitumor effect of OK-432 possibly through the enhanced production of TNF-α. Combination of L-MTP-PE and OK-432 is considered to be a candidate for a new treatment model for cancer.

Simultaneous Administration of NOD-2 (MDP) and TLP-4 (LPS) Ligands to Bone Marrow Donors 24 h before Transplantation Increases the Content of Multipotent Stromal Cells (MSCs) in Bone Marrow Grafts in CBA Mice Compared to the Total Result of Their Isolated Administration.

In 3-month bone marrow transplants of CBA mice from bone marrow donors receiving single injections of TLR-4 ligand (LPS) or NOD-2 ligand (muramyl dipeptide, MDP) 24 h before transplantation, an increase in the total number of MSCs (by 2.6 and 1.9 times, respectively), as well as a slight increase in the number of nuclear cells and the mass of bone capsules (by 1.3 and 1.2 times) were observed. After combined administration of MDР and LPS to donors, the total content of MSCs in the grafts was higher by 1.6 times in comparison with the total result of their isolated administration (and by 7.2 times in comparison with the control). At the same time, the concentration of osteogenic MSCs in the grafts of all groups was almost the same and corresponded to the control level. The number of nuclear cells and the mass of bone capsules of the grafts after combined administration of LPS and MDP were close (~80%) to the sum of the results of their isolated administration. These findings suggest that activation of the stromal tissue and the success of bone marrow transplantation depend on the intensity of innate immune responses. These data can be useful for the development of optimal methods of tissue transplantation.

Structural Fine-Tuning of Desmuramylpeptide NOD2 Agonists Defines Their In Vivo Adjuvant Activity.

We report on the design, synthesis, and biological evaluation of a series of nucleotide-binding oligomerization-domain-containing protein 2 (NOD2) desmuramylpeptide agonists with improved in vitro and in vivo adjuvant properties. We identified two promising compounds: 68, a potent nanomolar in vitro NOD2 agonist, and the more lipophilic 75, which shows superior adjuvant activity in vivo. Both compounds had immunostimulatory effects on peripheral blood mononuclear cells at the protein and transcriptional levels, and augmented dendritic-cell-mediated activation of T cells, while 75 additionally enhanced the cytotoxic activity of peripheral blood mononuclear cells against malignant cells. The C18 lipophilic tail of 75 is identified as a pivotal structural element that confers in vivo adjuvant activity in conjunction with a liposomal delivery system. Accordingly, liposome-encapsulated 75 showed promising adjuvant activity in mice, surpassing that of muramyl dipeptide, while achieving a more balanced Th1/Th2 immune response, thus highlighting its potential as a vaccine adjuvant.

Novel chimeric TLR2/NOD2 agonist CL429 exhibited significant radioprotective effects in mice.

Severe ionizing radiation causes the acute lethal damage of haematopoietic system and gastrointestinal tract. Here, we found CL429, the novel chimeric TLR2/NOD2 agonist, exhibited significant radioprotective effects in mice. CL429 increased mice survival, protected mice against the lethal damage of haematopoietic system and gastrointestinal tract. CL429 was more effective than equivalent amounts of monospecific (TLR2 or NOD2) and combination (TLR2 + NOD2) of molecules in preventing radiation-induced death. The radioprotection of CL429 was mainly mediated by activating TLR2 and partially activating NOD2. CL429-induced radioprotection was largely dependent on the activation of TLR2-MyD88-NF-κB signalling pathway. In conclusion, the data suggested that the co-activation of TLR2 and NOD2 could induce significant synergistic radioprotective effects and CL429 might be a potential high-efficiency selective agent.

Production of fever mediator PGE2 in human monocytes activated with MDP adjuvant is controlled by signaling from MAPK and p300 HAT: Key role of T cell derived factor.

Fever and inflammatory responses were observed in some subjects in early clinical trials of vaccines adjuvanted with muramyl dipeptide (MDP), a NOD2 agonist. Biosynthesis of Prostaglandin E2 (PGE2) that transmits febrile signals to the brain is controlled by an inducible enzyme, Cyclooxygenase 2 (COX-2). MDP alone was not sufficient to induce expression of COX-2 and PGE2 production in vitro. Conditioned medium prepared from Peripheral Blood Mononuclear Cells (PBMCs)-derived CD3-bead purified human T cells (TCM) dramatically increased COX2 gene transcription, COX-2 protein expression, and PGE2 production in MDP-treated monocytes. We explored epigenetic changes at the COX2 promoter using Chromatin Immunoprecipitation assay (ChIP). Increase in COX2 transcription correlated with increased recruitment of RNA polymerase II (Pol II) and p300 histone acetyl transferase (HAT) to the COX2 promoter in monocytes activated with MDP and TCM. The role of p300 HAT was confirmed by using C646, an inhibitor of p300, that reduced binding of acetylated H3 and H4 histones at the COX2 promoter, COX2 transcription, and PGE2 production in monocytes. Binding of p300, Nuclear Factor Kappa B (NF-κB), and Pol II to the COX2 promoter was also sensitive to inhibitors of Mitogen-Activated Protein Kinase (MAPK) pathway and to antibodies against Macrophage-1 (Mac-1) integrin in MDP/TCM-treated monocytes. Importantly, recombinant Glycoprotein Ib alfa (GPIbα), the recently identified factor in TCM, increased binding of NF-κB, p300, and of Pol II to the COX2 promoter and COX2 transcription in MDP-treated monocytes. Our findings suggest that a second signal through Mac-1 and MAPK is triggered by a T cell derived soluble GPIbα protein leading to the assembly of the transcription machinery at the COX2 promoter and production of PGE2 in human monocytes in response to MDP/NOD2 activation.

The Brief Analysis of Peptide-combined Nanoparticle: Nanomedicine's Unique Value.

Therapeutic peptides (TPs) are biological macromolecules which can act as neurotransmitters, hormones, ion channel ligands and growth factors. Undoubtedly, TPs are crucial in modern medicine. But low bio-stability and some special adverse reactions reduce their places to the application. With the development of nanotechnology, nanoparticles (NPs) in pharmaceutical science gained much attention. They can encapsulate the TPs into their membrane or shell. Therefore, they can protect the TPs against degradation and then increase the bioavailability, which was thought to be the biggest advantage of them. Additionally, targeting was also studied to improve the effect of TPs. However, there were some drawbacks of nano TPs like low loading efficiency and difficulty to manufacture. Nowadays, lots of studies focused on improving effect of TPs by preparing nanoparticles. In this review, we presented a brief analysis of peptide-combined nanoparticles. Their advantages and disadvantages were listed in terms of mechanism. And several examples of applications were summarized.

Phenotypic Differences in Primary Murine Microglia Treated with NOD1, NOD2, and NOD1/2 Agonists.

The purpose of the study was studying the influence of different NOD agonists on the morphological phenotype of primary murine microglia and to examine their influence on characteristic cytokines. Primary CD11b-positive cells were isolated from the brain of neonatal mice. The microglial phenotype of the cells was examined by ionized calcium-binding adapter molecule (Iba)1 staining. After14 days in culture, these cells were stimulated by iE-DAP, L18-MDP, or M-TriDAP as NOD1, NOD2, and NOD1/2 agonists, respectively. The cellular morphology was recorded and compared to the phenotype of cells cultured in medium alone or after LPS stimulation. The cells developed a specific phenotype only after treatment with the NOD2 agonist L18-MDP. These cells were characterized by straight extensions carrying tiny spikes and had a high ramification index. This was in sharp contrast to all other treatments, which always resulted in an amoeboid phenotype typically shown by activated microglia in vivo and by cultured microglia in vitro. The staining intensity of IL-6 and TNF-α did not reveal any clear difference independent of the NOD agonist treatment. In contrast, an increased staining intensity was observed for IL-10 after L18-MDP treatment. The NOD2 agonist L18-MDP induced a morphologically distinct phenotype characterized by microspike-decorated dendritiform extensions and a high degree of ramification in primary murine microglia. Increased ramification index and elevated staining intensity of anti-inflammatory IL-10 as hallmarks suggest that a M2-like phenotype of microglia was induced.

T cell-derived soluble glycoprotein GPIbα mediates PGE2 production in human monocytes activated with the vaccine adjuvant MDP.

Vaccine adjuvants containing analogs of microbial products activate pattern recognition receptors (PRRs) on antigen-presenting cells, including monocytes and macrophages, which can cause prostaglandin E2 (PGE2) release and consequently undesired inflammatory responses and fever in vaccine recipients. Here, we studied the mechanism of PGE2 production by human monocytes activated with muramyl dipeptide (MDP) adjuvant, which activates cytosolic nucleotide-binding oligomerization domain 2 (NOD2). In rabbits, administration of MDP elicited an early increase in PGE2 followed by fever. In human monocytes, MDP alone did not induce PGE2 production. However, high amounts of PGE2 and the proinflammatory cytokines IL-1ß and IL-6 were secreted by monocytes activated with MDP in the presence of conditioned medium obtained from CD3 bead-isolated T cells (Tc CM) but not from those isolated without CD3 beads. Mass spectrometry and immunoblotting revealed that the costimulatory factor in Tc CM was glycoprotein Ib α (GPIbα). Antibody-mediated blockade of GPIbα or of its receptor, Mac-1 integrin, inhibited the secretion of PGE2, IL-1ß, and IL-6 in MDP + Tc CM-activated monocytes, whereas recombinant GPIbα protein increased PGE2 production by MDP-treated monocytes. In vivo, COX2 mRNA abundance was reduced in the liver and spleen of Mac-1 KO mice after administration of MDP compared with that of treated wild-type mice. Our findings suggest that the production of PGE2 and proinflammatory cytokines by MDP-activated monocytes is mediated by cooperation between two signaling pathways: one delivered by MDP through NOD2 and a second through activation of Mac-1 by T cell-derived GPIbα.

NOD2 agonist murabutide alleviates radiation-induced injury through DNA damage response pathway mediated by ATR.

Injury-induced by ionizing radiation (IR) severely reduces the quality of life of victims. The development of radiation protectors is regarded as one of the most resultful strategies to alleviate damages caused by IR exposure. In the present study, we investigated the radioprotective effects of the agonist of nucleotide-binding-oligomerization-domain-containing proteins 2 called murabutide (MBD) and clarified the potential mechanisms. Our results showed that the pretreatment with MBD effectively protected cultured cells and mice against IR-induced toxicity and the pretreatment with MBD in vitro and in vitro also inhibited apoptosis caused by IR exposure. The downregulation of γ-H2AX and the upregulation of ATR signaling pathways by MBD treatment indicated that the radioprotective effects of MBD were due to the stimulation of DNA damage response (DDR) pathway to repair DNA double-strand breaks caused by IR exposure. As the radioprotective effects of MBD were diminished by the ATR selective inhibitor rather than the ATM inhibitor, ATR pathway was confirmed to be a more crucial checkpoint pathway in mediating the stimulation of DDR pathway by MBD. Taken together, our data provide a novel and effective protector to relieve the injury induced by IR exposure.

Ligands of NOD2 (Muramyl Dipeptide) and TLR4 (LPS) in 24 h after Combined In Vivo Administration Produce a Synergistic Increase in the Content of Multipotent Stromal Cells in the Bone Marrow and Peritoneal Exudate of CBA Mice.

In 24 h after combined administration of ligands of NOD2 (muramyl dipeptide) and TLR4 (LPS) receptors to CBA mice, a synergistic increase (by 10 times compared to the intact control) in cloning efficiency and content of multipotent stromal cells was observed in the bone marrow in comparison with the total effects of their individual administration (by 2.1 and 4.1 times, respectively). A similar effect was also observed in the peritoneal exudate. When ligands were administered simultaneously, the concentration of osteogenic multipotent stromal cells in the bone marrow decreased to a greater extent than in case of individual injections of the ligands, but did not drop below 7% of the control, which is apparently indicative of a decline threshold. In 3 h after simultaneous addition of the ligands in vitro to 12-day primary cultures of mouse bone marrow stromal cells, a synergistic increase in TNFα concentration was observed (32-fold increase from the level of intact control), while IL-10 concentration did not differ from the control, which is indicative of the proinflammatory nature of the process and the absence of immunosuppressive effect. These results suggest that activation of the stromal tissue depends on the intensity of innate immunity reactions.

[Muramyldipeptide - based compounds in current medicine: focus on glucosaminylmuramyl dipeptide].

The role of immune mechanisms in the pathogenesis of almost all human diseases shown in recent decades, increase in antibiotic resistance and secondary immunodeficiency, aging of the population and widespread use of immunosuppressive drugs and procedures suggest a wider use of immunomodulators in current clinical practice, but the use of most of them limits the lack of knowledge. The most promising compounds for the development as immunomodulating agents and adjuvants for a wide range of vaccines are low molecular weight fragments of peptidoglycan - muramylpeptides. The article describes the mechanisms of action of muramylpeptides, their biological effects and properties of medicines developed on their basis. Special emphasis is placed to glucosaminylmuramyl dipeptide registered in the Russian Federation under the trade name Likopid, which is currently the best - studied drug in its group. The results of Likopid studies when used as a prophylactic and therapeutic agent for infections of various localization in adults and children, for oncological diseases and complications of chemotherapy and radiation therapy, psoriasis, atopic and other diseases are presented. It is emphasized that in diseases associated with human papillomavirus and plaque psoriasis, according to current criteria of evidence - based medicine, Likopid should be classified as drug with level A efficacy (high efficiency in 80-100% of patients). High safety of Likopid in adults and children, including newborns, is noted.

Different activities of antitumor immunomodulators to induce neutrophil adherence response.

Functions of neutrophils, major participant in host defense mechanisms, are known to be regulated by various types of immunomodulators. Capacity of immunomodulators which are reported to show antitumor effect in vivo to induce neutorophil adherence response in vitro was investigated. Several bacterial immunomodulators (OK-432, Corynebacterium parvum, B.C.G.) and components of bacteria cell walls (lipopolysaccharide (LPS), lipid A, lipoteicoic acid, N-cell wall skelton (N-CWS), muramyl dipeptide (MDP)) and fungal polysaccharides (lentinan, zymosan A, etc.) were tested. Neutrophils prepared from peripheral blood of healthy men were incubated with each immunomodulator at 37°C for 60 min in 96 well plastic plates, then neutrophils adherent to substratum were stained by crystal violet and their optical density at 570 nm was measured as a parameter of neutrophil adherence. Although purified polysaccharides mainly prepared from fungi did not induce the adherent response, not only bacterial bodies and their components but also tumor necrosis factor-α (TNF-α) clearly induced it. On the base of these results, functional classification and typing of immunomodulators by different activities in neutrophil adherence was discussed.