Production Of Anti-inflammatory Cytokines Research Articles

Unveiling the therapeutic symphony of probiotics, prebiotics, and postbiotics in gut-immune harmony.

The gut microbiota and immune system interaction play a crucial role in maintaining overall health. Probiotics, prebiotics, and postbiotics have emerged as promising therapeutic approaches to positively influence this complex axis and enhance health outcomes. Probiotics, as live bacteria, promote the growth of immune cells, shape immune responses, and maintain gut barrier integrity. They modify the gut microbiota by fostering beneficial bacteria while suppressing harmful ones. Additionally, probiotics interact with the immune system, increasing immune cell activity and anti-inflammatory cytokine production. Prebiotics, as indigestible fibers, selectively nourish beneficial microorganisms in the gut, enhancing gut microbial diversity and activity. This, in turn, improves gut health and boosts immune responses while controlling inflammation through its immunomodulatory properties. Postbiotics, produced during probiotic fermentation, such as short-chain fatty acids and antimicrobial peptides, positively impact gut health and modulate immune responses. Ensuring quality control and standardization will be essential for successful clinical implementation of these interventions. Overall, understanding and harnessing the gut microbiota-immune system interplay offer promising avenues for improving digestive and immunological health.

Erdr1 Drives Macrophage Programming via Dynamic Interplay with YAP1 and Mid1.

Erythroid differentiation regulator 1 (Erdr1) is a stress-induced, widely expressed, highly conserved secreted factor found in both humans and mice. Erdr1 is linked with the Hippo-YAP1 signaling. Initially identified as an inducer of hemoglobin synthesis, Erdr1 emerged as a multifunctional protein, especially in immune cells. Although Erdr1 has been implicated in regulating T cells and NK cell function, its role in macrophage remains unclear. This study explored the function and mechanism of Erdr1 in macrophage inflammatory response. The data demonstrated that Erdr1 could promote anti-inflammatory cytokine production, a function that also has been reported by previous research. However, I found Erdr1 also could play a proinflammatory role. The function of Erdr1 in macrophages depends on its dose and cell density. I observed that Erdr1 expression was inhibited in M1 macrophages but was upregulated in M2 macrophages compared with unpolarized macrophages. I hypothesized that Erdr1 balances the inflammatory response by binding with distinct adaptors dependent on varying concentrations. Mechanistically, I demonstrated YAP1 and Mid1 as the two adaptor proteins of Erdr1. The Erdr1-YAP1 interaction promotes anti-inflammatory cytokine production when Erdr1 levels are elevated, whereas the Erdr1-Mid1 interaction induces proinflammatory cytokine production when Erdr1 levels are decreased. This study highlights the effects of Erdr1 on regulating cytokine production from polarized macrophages potentially by regulating YAP1 in the nonclassical Hippo pathway.

P1238 Exploring the Role of Klebsiella variicola and Klebsiella pneumoniae in Pediatric Ulcerative Colitis Pathogenesis

Abstract Background Recent studies indicated that Klebsiella (K) pneumoniae (isolated from the stool of IBD patients) and K. variicola (isolated from the mesenteric tissue of Crohn disease patients) have the potential to induce inflammation in epithelial and preadipocyte cells, exacerbating colitis in murine models. We isolated these strains from pediatric ulcerative colitis (UC) patients' appendix (relevant to UC pathogenesis given the protective effects of appendectomy) and other non-inflamed colon sections. We hypothesized that these isolates are invasive and induce inflammation in UC. These strains have not been previously studied in UC. Methods K. variicola and K. pneumoniae were isolated from two pediatric UC patients' appendices and other non-inflamed colon sections and identified using 16S DNA Sanger sequencing. Virulence of the two strains was determined by infecting Caco2 cell lines and performing adhesion and invasion assays, quantifying biofilm formation, and assessing barrier functions using transepithelial electrical resistance (TEER) measurement. Importantly, we monitored the production of pro-inflammatory (e.g., IL-8, TNF-α) and anti-inflammatory (IL-10) cytokines using reverse transcriptase quantitative polymerase chain reaction (RT-qPCR). Results K. variicola and K. pneumoniae exhibited high levels of biofilm formation compared to adherent-invasive Escherichia coli (AIEC), which are commonly isolated from IBD patients. Furthermore, the Klebsiella strains adhered to epithelial cells within 2-3 hours post infection. TEER experiments showed compromised barrier integrity after 6 hours and overnight infection. Except for K. pneumoniae isolated from the ascending colon, the tested strains exhibited a significant increase in IL-8 expression. Similarly, K. variicola from the peri-appendicular region demonstrated elevated TNF-α gene expression. Conclusion K. variicola and K. pneumoniae isolated from UC patients have the potential to form biofilms, disrupt barrier integrity, and trigger inflammatory responses. These findings unravel a potential role for pathogenic Klebsiella strains in driving UC pathogenesis. Importantly, these data shed light on the role of appendix-associated bacteria in the development of UC. Future work includes using comparative genomics to map the virulence determinants of these bacteria.

Mixed Bacillus subtilis and Lactiplantibacillus plantarum-fermented feed improves gut microbiota and immunity of Bamei piglet.

Antibiotics are widely used in the breeding production of Bamei pigs, affecting the quality and safety of pork and causing enormous harm to human health, the environment, and public health. The use of probiotic fermented feed to replace antibiotic feed is one of the solutions, which has the potential to improve the intestinal microbiota, promote animal growth, and enhance immunity. The purpose of this study was to evaluate the effect of fermented feed with Lactiplantibacillus (L.) plantarum QP28-1a or Bacillus (B.) subtilis QB8a on feed, growth performance, gut microbiota, and immunity of weaned piglets. A total of 60 freshly weaned piglets from the Tibetan Plateau were randomly divided into five groups and fed basal feed, L. plantarum fermented feed, B. subtilis fermented feed, mixed fermented feed, and antibiotic fermented feed for 60 days, respectively. The results showed fermented feed supplemented with L. plantarum QP28-1a or B. subtilis QB8a significantly lowered the pH of the feed (P < 0.05), produced lactic acid and acetic acid, inhibited the growth of harmful bacteria in the feed, and reduced the feed conversion rate in the group fed mixed fermented feed (P < 0.05). The fermented feed increased the α-diversity and prominently altered the β-diversity of the intestinal microbiota, increasing the relative abundance of beneficial bacteria such as Lactobacillus and Turicibacter and decreasing the relative abundance of conditional pathogens such as Streptococcus and Clostridium, improving the intestinal microbiota of the Bamei piglets. Notably, the mixed fermented feed improved the immunity of Bamei piglets by modulating the production of pro-inflammatory cytokines, anti-inflammatory cytokines, and inflammatory-related signaling pathways. Spearman's correlation analysis revealed that the increased expression of immune-related cytokines may be associated with a significant enrichment of Lactobacillus, Prevotellaceae, Erysipelotrichaceae, and Ruminococcaceae in the gut. In conclusion, the probiotic fermented feed maintained an acidic environment conducive to suppressing pathogens, reduced the feed conversion ratio, optimized the intestinal microbiota, improved immunity, and alleviated intestinal inflammation that may be caused by weaning, demonstrating the excellent application prospects of L. plantarum QP28-1a and B. subtilis QB8a fermented feed in the feeding of Bamei piglets.

Зміни рівнів про- і протизапальних цитокінів у жінок із хронічним ендометритом та можливість медикаментозної корекції цих рівнів на преконцепційному етапі

The incidence of chronic endometritis is more than twice higher the prevalence of other clinical forms of intrauterine pathology, leading to infertility, unsuccessful attempts of in vitro fertilization and spontaneous abortion. There is a lack of data on the alteration of pro- and anti-inflammatory cytokine production in chronic endometritis and their role in the subsequent development of miscarriage. Purpose - to investigate the levels of pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α) and interferon gamma (INF-γ), as well as the anti-inflammatory cytokine interleukin 10 (IL-10) in cervical mucus of women with chronic endometritis and to assess the impact of the developed comprehensive preconception treatment on normalizing cytokine disorders in these women. Materials and methods. The levels of pro-inflammatory cytokines TNF-α and INF-γ, as well as the anti-inflammatory cytokine IL-10, were determined using the enzyme-linked immunosorbent assay method in the cervical mucus of 426 women with chronic endometritis who were planning pregnancy. Among them, 168 patients (the Group I) received preconception comprehensive treatment, which included broad-spectrum antibacterial agents (azithromycin), hormonal estrogen-progestin therapy (Femoston 2/10) and L-arginine. The remaining 258 women (the Group II) did not receive treatment for chronic endometritis at the preconception stage. The control group consisted of 30 healthy individuals. Data analysis was performed using mathematical statistical methods, calculating the mean sample values (M), variance (σ) and standard errors of the mean (m). The Student’s t-test was applied, and probabilities were determined using the statistical program «STATISTICA» (StatSoft Inc., USA). Results. In the cervical mucus of women with chronic endometritis, an increase in the levels of pro-inflammatory cytokines INF-γ by 2.8 times (p&lt;0.001) and TNF-α by 2 times (p&lt;0.001) was detected, along with a 2-fold decrease in the concentration of the anti-inflammatory cytokine IL-10 (p&lt;0.001). The shift in the cytokine balance towards pro-inflammatory cytokines is associated with a high frequency of miscarriage in these women. After the treatment of chronic endometritis, women had a 2.6 times lower level of INF-γ (p&lt;0.05), a 2 times lower level of TNF-α (p&lt;0.01), and a 1.7 times higher level of IL-10 (p&lt;0.05) compared to untreated patients. Conclusions. Women with chronic endometritis experience a cytokine imbalance, characterized by an excessive increase in pro-inflammatory cytokines and a decrease in anti-inflammatory cytokines. Women with chronic endometritis who are planning pregnancy require comprehensive preconception treatment, which includes the administration of azithromycin, femoston 2/10 and L-arginine in addition to traditional preconception folate prophylaxis. This treatment, by promoting improved conditions for implantation, leads to a 15% increase in the pregnancy rate and a 1.9-fold reduction in the frequency of spontaneous abortion. The research was carried out in accordance with the principles of the Helsinki Declaration. The study protocol was approved by the Local Ethics Committee of the participating institution. The informed consent of the patient was obtained for conducting the studies. No conflict of interests was declared by the authors.

Homeopathic dilutions of Phytolacca decandra induce cytotoxicity of human breast adenocarcinoma cells

Breast cancer is a malignant tumor frequently diagnosed in the female population that representsthe main cause of cancer death in women worldwide. Recently, the use of complementary therapiessuch as homeopathy has increased to alleviate the side effects of conventional allopathic treatments.However, the cellular and molecular effects of homeopathic medicaments have not been fullyclarified. Thus, the aim of this work was to evaluate the in vitro effects of Phytolacca decandrahomeopathic preparations on human breast adenocarcinoma cells. For this, MCF-7 breastadenocarcinoma cells were cultivated in the presence or absence of Phytolacca decandra mothertincture (PdTM) or centesimal Hahnemannian dilution (CH) of Phytolacca decandra: 30 or 200 CH.Determination of apoptosis and necrosis percentages, cytokine production and morphologicalchanges were evaluated after 24, 48 and 72 hours of culture in each of the treatments. Resultsobtained in cytotoxicity analyses showed a higher percentage of death in cells treated withhomeopathic dilutions, with similar results to that presented by cells treated with Doxorubicin orPdTM. In contrast, cells treated with homeopathic dilutions of Phytolacca decandra did not showmarked differences in the production of pro and anti-inflammatory cytokines when compared tountreated. Similarly, analyses of cell morphology did not display significant differences before andafter treatment with homeopathic dilutions, when compared to untreated cells. Already, cells treatedwith PdTM presented marked cellular alterations with observation of apoptotic bodies in the threeevaluated times. Together, results show that homeopathic dilutions of Phytolacca decandra inducedcytotoxicity of human breast adenocarcinoma cells in the absence of inflammatory process.

Lonicera japonica Thunb extract ameliorates lipopolysaccharide-induced acute lung injury associated with luteolin-mediated suppression of NF-κB signaling pathway

ObjectiveLonicera japonica Thunb (LJT) is a commonly used herbal soup to treat inflammation-related diseases. However, the effect of LJT on ALI is unknown. The present study was aimed at investigating the protective effects of LJT extract (LTE) and its active ingredient luteolin (Lut) on lipopolysaccharide (LPS)-stimulated ALI and investigate its potential mechanism.Materials and methodsThe effects of LTE and Lut were explored in an ALI mouse model induced by intraperitoneal injection of lipopolysaccharide (LPS). Besides, the LPS-induced inflammation model in BEAS-2B cells was used to clarify the underlying mechanisms. The ALI pathological changes in lung tissues were tested through Haematoxylin and eosin (HE) staining. The apoptosis of cells in lung tissue and the cell model in vitro was evaluated by TUNEL assays, respectively. Meanwhile, the viability of cells in vitro was evaluated by Cell Counting Kit-8 (CCK-8) assay. The levels/concentrations of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-1β and IL-10 in BALF were detected by enzyme-linked immunosorbent assay (ELISA). Besides, through quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting, the expression of the above-mentioned inflammatory factors and key factors in the NF-κB signaling pathway was examined. The distribution of inflammatory factors in tissue was observed through immunohistochemistry (IHC) assays .ResultsIn relative to LPS-stimulated group, the in vivo study showed that LTE and different concentrations of Lut dramatically alleviated LPS-evoked lung pathological injury and lung edema based on the changes in total protein levels and lung wet/dry (W/D) ratio in the bronchoalveolar lavage fluid (BALF) from ALI mice. LTE and different concentrations of Lut also suppressed the inflammatory response, as reflected by the variations of neutrophil accumulation and the production of proinflammatory and anti-inflammatory cytokines in the lung tissues and BALF of ALI mice. The in vitro research also demonstrated that LTE and Lut visibly facilitated cell viability and restrained the apoptosis of BEAS-2B cells stimulated by LPS. Lut hindered LPS-inducible activation of NF-κB pathway in BEAS-2B cells.ConclusionThe present study proved that LTE might suppress LPS-induced acute injury and inflammation in mice and BEAS-2B cells through the Lut-caused suppression of NF-κB signal path (Figure 1).

Medicinal Herbs: Promising Immunomodulators for the Treatment of Infectious Diseases.

Humans are constantly at high risk of emerging pandemics caused by viral and bacterial infections. The emergence of new pandemics is mainly caused by evolved viruses and bacteria that are highly resistant to existing medications. The rapid evolution of infectious agents demands the urgent investigation of new therapeutic strategies to prevent and treat these infections at an early stage. One of these therapeutic strategies includes the use of medicinal herbs for their antibacterial and antiviral properties. The use of herbal medicines as remedies is very ancient and has been employed for centuries. Many studies have confirmed the antimicrobial activities of herbs against various pathogens in vitro and in vivo. The therapeutic effect of medicinal herbs is mainly attributed to the natural bioactive molecules present in these plants such as alkaloids, flavonoids, and terpenoids. Different mechanisms have been proposed for how medicinal herbs enhance the immune system and combat pathogens. Such mechanisms include the disruption of bacterial cell membranes, suppression of protein synthesis, and limitation of pathogen replication through the inhibition of nucleic acid synthesis. Medicinal herbs have been shown to treat a number of infectious diseases by modulating the immune system's components. For instance, many medicinal herbs alleviate inflammation by reducing pro-inflammatory cytokines (e.g., tumor necrosis factor-alpha (TNF-α), interleukin-1, IL-6) while promoting the production of anti-inflammatory cytokines (e.g., IL-10). Medicinal herbs also play a role in defense against viral and intracellular infections by enhancing the proliferation and functions of natural killer cells, T-helper-1 cells, and macrophages. In this review, we will explore the use of the most common herbs in preventing and treating infectious and non-infectious diseases. Using current and recently published studies, we focus on the immunomodulatory and therapeutic effects induced by medicinal herbs to enhance immune responses during diseases.

Spinal interleukin-24 contributes to neuropathic pain after peripheral nerve injury through interleukin-20 receptor2 in mice

Neuroinflammation is critically involved in nerve injury-induced neuropathic pain, characterized by local and systemic increased levels of proinflammatory cytokines. Interleukin-24 (IL-24), a key member of the IL-10 family, has been extensively studied for its therapeutic potential in various diseases, including cancer, autoimmune disorders, and bacterial infections, but whether it is involved in the regulation of neuropathic pain caused by peripheral nerve injury (PNI) has not been well established. In this study, we reported that spared nerve injury (SNI) induced a significant upregulation of IL-24 in fibroblasts, neurons, and oligodendrocyte precursor cells (OPCs, also called NG2-glia) in the affected spinal dorsal horns (SDHs), as well as dorsal root ganglions (DRGs). We also found that tumor necrosis factor α (TNF-α) induced the transcriptional expression of IL-24 in cultured fibroblasts, neurons, and NG2-glia; in addition, astrocytes, microglia, and NG2-glia treated with TNF-α exhibited a prominent increase in interleukin-20 receptor 2 (IL-20R2) expression. Furthermore, we evaluated the ability of IL-24 and IL-20R2 to attenuate pain in preclinical models of neuropathic pain. Intrathecal (i.t.) injection of IL-24 neutralizing antibody or IL-20R2 neutralizing antibody could effectively alleviate mechanical allodynia and thermal hyperalgesia after PNI. Similarly, intrathecal injection of IL-24 siRNA or IL-20R2 siRNA also alleviated mechanical allodynia after SNI. The inhibition of IL-24 reduced SNI-induced proinflammatory cytokine (IL-1β and TNF-α) production and increased anti-inflammatory cytokine (IL-10) production. Meanwhile, the inhibition of IL-20R2 also decreased IL-1β mRNA expression after SNI. Collectively, our findings revealed that IL-24/IL-20R might contribute to neuropathic pain through inflammatory response. Therefore, targeting IL-24 could be a promising strategy for treating neuropathic pain induced by PNI.

Effect of Mixed Lactiplantibacillus plantarum- and Bacillus subtilis-Fermented Feed on Growth, Immunity, and Intestinal Health of Weaner Pigs

Bamei pigs, an eximious local breed reared on the Tibetan Plateau of China, are facing problems such as feed shortages, weaning stress, and antibiotic abuse. This study aimed to improve the quality of feed, growth performance, intestinal microbiota, and immunity of Bamei pigs through feeding with potentially probiotic-fermented feed. Different feeds were administered to weaned Bamei piglets for 60 days, creating the following five experimental treatment groups: basal feed group; Lactiplantibacillus plantarum-fermented-feed group; Bacillus subtilis-fermented-feed group; mixed-fermented-feed group; and antibiotic-supplemented-feed group. The results showed that the pH, neutral detergent fiber, and acid detergent fiber of the potentially probiotic-fermented feeds were significantly reduced; organic acids were produced; and Coliform bacteria, Clostridium, and aerobic bacteria were effectively inhibited. Feeding with potentially probiotic-fermented feed not only reduced the feed-conversion ratio but also improved immunity by increasing the production of anti-inflammatory cytokines, as well as decreasing pro-inflammatory cytokines and typical inflammatory pathways. The 16s rDNA high-throughput-sequencing results showed that probiotic-fermented feed improved the diversity of intestinal microbiota, inhibited the growth of the opportunistic pathogens Clostridium and Streptococcus, increased the relative abundance of Lactobacillus and Prevotella, and promoted gut health, demonstrating the promising application prospects of potentially probiotic-fermented feed.

Efferocytosis is restricted by axon guidance molecule EphA4 via ERK/Stat6/MERTK signaling following brain injury

BackgroundEfferocytosis is a process that removes apoptotic cells and cellular debris. Clearance of these cells alleviates neuroinflammation, prevents the release of inflammatory molecules, and promotes the production of anti-inflammatory cytokines to help maintain tissue homeostasis. The underlying mechanisms by which this occurs in the brain after injury remain ill-defined.MethodsWe used GFP bone marrow chimeric knockout (KO) mice to demonstrate that the axon guidance molecule EphA4 receptor tyrosine kinase is involved in suppressing MERTK in the brain to restrict efferocytosis of resident microglia and peripheral-derived monocyte/macrophages.ResultsSingle-cell RNAseq identified MERTK expression, the primary receptor involved in efferocytosis, on monocytes, microglia, and a subset of astrocytes in the damaged cortex following brain injury. Loss of EphA4 on infiltrating GFP-expressing immune cells improved functional outcome concomitant with enhanced efferocytosis and overall protein expression of p-MERTK, p-ERK, and p-Stat6. The percentage of GFP+ monocyte/macrophages and resident microglia engulfing NeuN+ or TUNEL+ cells was significantly higher in KO chimeric mice. Importantly, mRNA expression of Mertk and its cognate ligand Gas6 was significantly elevated in these mice compared to the wild-type. Analysis of cell-specific expression showed that p-ERK and p-Stat6 co-localized with MERTK-expressing GFP + cells in the peri-lesional area of the cortex following brain injury. Using an in vitro efferocytosis assay, co-culturing pHrodo-labeled apoptotic Jurkat cells and bone marrow (BM)-derived macrophages, we demonstrate that efferocytosis efficiency and mRNA expression of Mertk and Gas6 was enhanced in the absence of EphA4. Selective inhibitors of ERK and Stat6 attenuated this effect, confirming that EphA4 suppresses monocyte/macrophage efferocytosis via inhibition of the ERK/Stat6 pathway.ConclusionsOur findings implicate the ERK/Stat6/MERTK axis as a novel regulator of apoptotic debris clearance in brain injury that is restricted by peripheral myeloid-derived EphA4 to prevent the resolution of inflammation.

PSI-1 Assessing the Impact of Hempseed Meal Supplementation on Inflammatory Cytokine Concentrations in Boer X Spanish Intact Male Goats

Abstract This study aimed to evaluate the effect of hempseed meal (HSM) supplementation on the concentrations of inflammatory cytokines in intact male goats. Effect on the concentration of pro-inflammatory cytokines, interleukin (IL) 2 , IL-6, tumor necrosis factor-alpha (TNF-α), and interferon-gamma (INF-γ) and anti-inflammatory cytokines,; IL-4 and IL10, were evaluated via chicken-ovalbumin challenge. Thirty-six Boer x Spanish intact male goats, 10 to 11 months of age and bodyweights of 44.1 ± 3.8 kg, were randomly assigned to one of the four experimental diets (n = 9 goats/diet): 0% (control), 10%, 20%, and 30% HSM diets and fed ad libitum for 60 days. Experimental diets were formulated as total mixed ration (TMR), and the nutrient content of each diet was formulated to be iso-nitrogenous (18% crude protein) and to meet or exceed the nutrient requirement of breeding male goats. On day 46, goats were injected subcutaneously with chicken ovalbumin (2.5 mg) prepared in saline solution and alhydrogel as an adjuvant. On 14 days of post-injection, serum samples were collected and analyzed for cytokines concentrations using an indirect enzyme-linked immunosorbent assay. Data were analyzed using one-way ANOVA and Kruskal-Wallis test using R version 4.2.1. Results show no significant difference (P &amp;gt; 0.05) among the experimental groups in terms of studied pro-inflammatory cytokines (IL-2, IL-6, TNF-α, and INF-γ) as well as anti-inflammatory cytokines (IL-4 and IL-10) concentrations (Table 1). Results indicate that the inclusion of HSM up to 30 % in the diet does not affect the production of pro-inflammatory and anti-inflammatory cytokines in intact male goats. This study provides valuable insights into the impact of HSM supplementation on inflammatory response in goats.

Antitumor and immunomodulatory activity of fucoidan from the brown alga Lessonia trabeculata (Lessoniaceae) on breast cancer spheroids

Introduction: The therapeutic benefits of the brown algae fucoidan in the treatment of breast cancer have attracted considerable interest in recent years. However, research using spheroids which provide relevant results in trials for antitumor and immunomodulatory products because they adequately simulate the tumor microenvironment, is limited. Objective: To evaluate the antitumor and immunomodulatory activity of Lessonia trabeculata fucoidan (LtF), native to the Peruvian Sea, on two types of multicellular tumor spheroids. Methods: The study was conducted from January to December 2021. Two types of spheroides were elaborated: from 4T1 tumor cells (MTS), and from 4T1 tumor cells+mouse splenocytes (MTSs). The antitumor activity of LtF was evaluated in MTS by quantifying cell viability with MTT. Immunomodulatory activity was determined in MTSs using the IC50 for two types of treatment: simple, fucoidan alone (LtF) and combined, fucoidan+doxorubicin (LtF+Dox). Pro-inflammatory (TNF-α, IL-6) and anti-inflammatory (IL-10, TGF-β) cytokine production was quantified by sandwich ELISA 72 h after treatment. Dox was used as positive control in all assays. Results: LtF exerted antitumor activity as evidenced by increased necrotic zone and cell debris formation compared to the untreated control. Antitumor activity was concentration dependent between 100 and 6 000 μg/ml. In MTSs, simple treatment increased IL-6 and decreased IL-10 and TGF-β production. The combined treatment significantly reduced TGF-β production. In both treatments and Dox, there was an increase in IL-6 compared to the untreated control. The highest production of IL-10 and TGF-β was observed in the untreated control, compatible with a highly immunosuppressive tumor microenvironment. Conclusions: LtF is a good candidate for the treatment of breast cancer and can immunomodulate the tumor microenvironment alone or in combination with Dox.

Glycolipids mimicking biosurfactants of the synthetic origin as new immunomodulating and anticandidal derivatives

The immunobiological effectivity of glycolipids mimicking biosurfactants of the synthetic origin was followed up using macrophages cell line RAW264.7. These derivatives with different number of mannose units connected glycosidically or through triazole linker, and all having octyl aglycone, were evaluated with respect to their structure - immunomodulation activity relationship. This comparative study showed that the structural variations of the selected derivatives influenced the immunobiological cell behaviour as concerned pro-inflammatory TNF-α, IL-6, IL-1α, IL-17, IL-12 and anti-inflammatory IL-10 cytokines production and enhancement of RAW264.7 cell proliferation. The derivatives with mannose units linked through triazole linkers exerted in some cases stronger immunomodulative potency than (di)mannosides. On the other hand, a presence of triazole linker is a less favourable for an effective candidacidal activity as determined by in vitro using Candida albicans biofilm. The design of new defined immunomodulating formulas of the synthetic origin as possible antifungal agents and prospective participants in drug delivery systems may be of interest.

Solid-state atomic hydrogen as a broad-spectrum RONS scavenger for accelerated diabetic wound healing.

Hydrogen therapy shows great promise as a versatile treatment method for diseases associated with the overexpression of reactive oxygen and nitrogen species (RONS). However, developing an advanced hydrogen therapy platform that integrates controllable hydrogen release, efficient RONS elimination, and biodegradability remains a giant technical challenge. In this study, we demonstrate for the first time that the tungsten bronze phase H0.53WO3 (HWO) is an exceptionally ideal hydrogen carrier, with salient features including temperature-dependent highly-reductive atomic hydrogen release and broad-spectrum RONS scavenging capability distinct from that of molecular hydrogen. Moreover, its unique pH-responsive biodegradability ensures post-therapeutic clearance at pathological sites. Treatment with HWO of diabetic wounds in an animal model indicates that the solid-state atomic H promotes vascular formation by activating M2-type macrophage polarization and anti-inflammatory cytokine production, resulting in acceleration of chronic wound healing. Our findings significantly expand the basic categories of hydrogen therapeutic materials and pave the way for investigating more physical forms of hydrogen species as efficient RONS scavengers for clinical disease treatment.

Generation of Human Regulatory Dendritic Cells from Cryopreserved Healthy Donor Cells and Hematopoietic Stem Cell Transplant Recipients.

Acute graft versus host disease (GVHD) remains a significant complication following hematopoietic stem cell transplant (HSCT), despite improved human leukocyte antigen (HLA) matching and advances in prophylactic treatment regimens. Previous studies have shown promising results for future regulatory dendritic cell (DCreg) therapies in the amelioration of GVHD. This study evaluates the effects of cryopreservation on the generation of DCreg, the generation of young and older DCreg in serum-free media, and the feasibility of generating DCreg from young and older HSCT patient monocytes. DCregs were generated in X-vivo 15 serum-free media from donor or patient monocytes. This study includes the use of monocytes from young and older healthy, donor, and HSCT patients with varying hematological diseases. Phenotypic differences in cell populations were assessed via flow cytometry while pro-inflammatory and anti-inflammatory cytokine production was evaluated in culture medium. The number of DCreg generated from cryopreserved monocytes of healthy donors was not significantly different from freshly isolated monocytes. DCreg generated from cryopreserved monocytes had comparable levels of co-stimulatory molecule expression, inhibitory molecule expression, and cytokine production as freshly isolated monocytes. Young and older healthy donor monocytes generated similar numbers of DCreg with similar cytokine production and phenotype. Although monocytes from older HSCT patients generated significantly fewer DCreg, DCreg from young and older HSCT patients had comparable phenotypes and cytokine production. Monocytes from young and older myelodysplastic syndrome (MDS) patients generated reduced numbers of DCreg compared to non-MDS-derived DCreg. We demonstrate that the cryopreservation of monocytes from HSCT patients of varying hematological diseases allows for the cost-effective generation of DCreg on an as-needed basis. Although the generation of DCreg from MDS patients requires further assessment, these data support the possibility of in vitro-generated DCreg as a therapy to reduce GVHD-associated morbidity and mortality in young and older HSCT recipients.

Analysis of regulatory T lymphocytes in patients with traumatic brain injury

In recent years, traumatic brain injury (TBI) has been one of the most urgent medical and social problems due to its prevalence, predominantly affecting young people of working age, causing high mortality, disability and economic costs for treatment and subsequent rehabilitation of patients. At present, the role of patients’ immune system in evolving neuroinflammation after traumatic brain injury has been proven. Treg cell populations represent an important factor determining the outcome of the disease due to promoting induction of immunological tolerance, being a significant component of immunoregulation. As a result of our study, we found a decrease in the relative content of Treg within the total lymphocyte pool of peripheral blood, which has the CD3+CD4+CD25bright phenotype in patients of the 3rd and 4th groups, in comparison with the data from control group. Moreover, a decreased relative content of Tregs (CD4+CD25+T cells) was revealed which is due to the degree of brain tissue damage and, as a result, their migration to suppress inflammation due to production of anti-inflammatory cytokines (TGF-â, IL-10). The Treg population is heterogeneous, thus prompting us for analysis of the Treg subpopulations profile based on the expression of CD45R0 and CD62L. When assessing subpopulations of regulatory T lymphocytes within CD45Ro and CD62L, significant changes were found only in patients with brain contusion. The changes were revealed within the pool of “naive” T regulatory lymphocytes with the CD3+CD4+CD25brightCD39+ Treg phenotype, capable of producing a wide range of cytokines specific for Th1, Th2, Th17, in patients with mild, moderate and severe TBI. Meanwhile, the level of highly active CD3+CD4+CD25brightCD73+ Tregs was significantly reduced in patients with moderate and severe TBI. These changes indicate an imbalance in immunoregulatory processes resulting from extensive damage to brain tissues.

Epithelial-Immune Cell Crosstalk Determines the Activation of Immune Cells In Vitro by the Human Cathelicidin LL-37 at Low Physiological Concentrations.

The only human cathelicidin, LL-37, is a host defense antimicrobial peptide with antimicrobial activities against protozoans, fungi, Gram(+) and Gram(-) bacteria, and enveloped viruses. It has been shown in experiments in vitro that LL-37 is able to induce the production of various inflammatory and anti-inflammatory cytokines and chemokines by different human cell types. However, it remains an open question whether such cytokine induction is physiologically relevant, as LL-37 exhibited its immunomodulatory properties at concentrations that are much higher (>20 μg/mL) than those observed in non-inflamed tissues (1-5 μg/mL). In the current study, we assessed the permeability of LL-37 across the Caco-2 polarized monolayer and showed that this peptide could pass through the Caco-2 monolayer with low efficiency, which predetermined its low absorption in the gut. We showed that LL-37 at low physiological concentrations (<5 μg/mL) was not able to directly activate monocytes. However, in the presence of polarized epithelial monolayers, LL-37 is able to activate monocytes through the MAPK/ERK signaling pathway and induce the production of cytokines, as assessed by a multiplex assay at the protein level. We have demonstrated that LL-37 is able to fulfill its immunomodulatory action in vivo in non-inflamed tissues at low physiological concentrations. In the present work, we revealed a key role of epithelial-immune cell crosstalk in the implementation of immunomodulatory functions of the human cathelicidin LL-37, which might shed light on its physiological action in vivo.

Cytochrome P450 26A1 Contributes to the Maintenance of Neuropathic Pain

The cytochrome P450 proteins (CYP450s) have been implicated in catalyzing numerous important biological reactions and contribute to a variety of diseases. CYP26A1, a member of the CYP450 family, carries out the oxidative metabolism of retinoic acid (RA), the active metabolite of vitamin A. Here we report that CYP26A1 was dramatically upregulated in the spinal cord after spinal nerve ligation (SNL). CYP26A1 was mainly expressed in spinal neurons and astrocytes. HPLC analysis displayed that the content of all-trans-RA (at-RA), the substrate of CYP26A1, was reduced in the spinal cord on day 7 after SNL. Inhibition of CYP26A1 by siRNA or inhibition of CYP26A1-mediated at-RA catabolism by talarozole relieved the SNL-induced mechanical allodynia during the maintenance phase of neuropathic pain. Talarozole also reduced SNL-induced glial activation and proinflammatory cytokine production but increased anti-inflammatory cytokine (IL-10) production. The RA receptors RARα, RXRβ, and RXRγ were expressed in spinal neurons and glial cells. The promoter of Il-10 has several binding sites for RA receptors, and at-RA directly increased Il-10 mRNA expression in vitro. Finally, intrathecal IL-10 attenuated SNL-induced neuropathic pain and reduced the activation of astrocytes and microglia. Collectively, the inhibition of CYP26A1-mediated at-RA catabolism alleviates SNL-induced neuropathic pain by promoting the expression of IL-10 and suppressing glial activation. CYP26A1 may be a potential therapeutic target for the treatment of neuropathic pain.

Adipose-Derived Mesenchymal Stromal Cells Co-Cultured with Primary Mixed Glia to Reduce Prion-Induced Inflammation.

Mesenchymal stromal cells (MSCs) are potent regulators of inflammation through the production of anti-inflammatory cytokines, chemokines, and growth factors. These cells show an ability to regulate neuroinflammation in the context of neurodegenerative diseases such as prion disease and other protein misfolding disorders. Prion diseases can be sporadic, acquired, or genetic; they can result from the misfolding and aggregation of the prion protein in the brain. These diseases are invariably fatal, with no available treatments. One of the earliest signs of disease is the activation of astrocytes and microglia and associated inflammation, which occurs prior to detectable prion aggregation and neuronal loss; thus, the anti-inflammatory and regulatory properties of MSCs can be harvested to treat astrogliosis in prion disease. Recently, we showed that adipose-derived MSCs (AdMSCs) co-cultured with BV2 cells or primary mixed glia reduce prion-induced inflammation through paracrine signaling. This paper describes a reliable treatment using stimulated AdMSCs to decrease prion-induced inflammation. A heterozygous population of AdMSCs can easily be isolated from murine adipose tissue and expanded in culture. Stimulating these cells with inflammatory cytokines enhances their ability to both migrate toward prion-infected brain homogenate and produce anti-inflammatory modulators in response. Together, these techniques can be used to investigate the therapeutic potential of MSCs on prion infection and can be adapted for other protein misfolding and neuroinflammatory diseases.