Carbopol hydrogel/sorbitan monostearate-almond oil based organogel biphasic formulations: Preparation and characterization of the bigels

Purpose: To obtain and evaluate carbopol hydrogel/sorbitan monostearate-almond oil-based organogel biphasic formulations (bigels) as a semi-solid vehicle for medicated topical applications.Methods: Bigel formulations were obtained under mild conditions at a hydrogel/organogel ratio of 80/20, 70/30, and 60/40 (w/w). Their stability, viscosity, spreadability, microarchitecture, and acute skin toxicity were evaluated.Results: Two formulations, prepared at ratios of 80/20 and 70/30, were stable based on intermediate stability testing, and had a similar viscosity and spreadability (38.0 ± 1.0 mm and 37.3 ± 0.6 mm, p > 0.05, respectively). Both of these formulations had a bimodal droplet size distribution and very similar values for the droplet mean diameter (0.33 ± 0.05 μm and 2.35 ± 0.44; and 0.34 ± 0.04 μm and 2.59 ± 0.21 μm). The formulation obtained at a ratio of 60/40 was unstable during storage. The in vivo results did not reveal any signs of skin toxicity.Conclusion: Considering their beneficial properties, the developed bigels are a potential semi-solid vehicle for topical application and exhibit a moisturizing effect.Keywords: Almond oil, Bigels, Carbopol hydrogel, Moisturizing effect, Organogel, Sorbitan monostearat

Graft-versus-host disease, but not graft-versus-leukemia immunity, is mediated by GM-CSF–licensed myeloid cells

Allogeneic hematopoietic cell transplantation (allo-HCT) not only is an effective treatment for several hematologic malignancies but can also result in potentially life-threatening graft-versus-host disease (GvHD). GvHD is caused by T cells within the allograft attacking nonmalignant host tissues; however, these same T cells mediate the therapeutic graft-versus-leukemia (GvL) response. Thus, there is an urgent need to understand how to mechanistically uncouple GvL from GvHD. Using preclinical models of full and partial MHC-mismatched HCT, we here show that the granulocyte-macrophage colony-stimulating factor (GM-CSF) produced by allogeneic T cells distinguishes between the two processes. GM-CSF drives GvHD pathology by licensing donor-derived phagocytes to produce inflammatory mediators such as interleukin-1β and reactive oxygen species. In contrast, GM-CSF did not affect allogeneic T cells or their capacity to eliminate leukemic cells, retaining undiminished GvL responses. Last, tissue biopsies and peripheral blood mononuclear cells from patients with grade IV GvHD showed an elevation of GM-CSF–producing T cells, suggesting that GM-CSF neutralization has translational potential in allo-HCT

The Role of Purine Metabolites as DAMPs in Acute Graft-versus-Host Disease

Acute graft-versus-host disease (GVHD) causes high mortality in patients undergoing allogeneic hematopoietic cell transplantation (allo-HCT). An early event in the classical pathogenesis of acute GVHD is tissue damage caused by the conditioning treatment or infection that consecutively leads to translocation of bacterial products (pathogen-associated molecular patterns, PAMPs) into blood or lymphoid tissue, as well as danger-associated molecular patterns (DAMPs), mostly intracellular components which act as pro-inflammatory agents, once they are released into the extracellular space. A subtype of DAMPs are nucleotides such as adenosine triphosphate (ATP) released from dying cells that can activate the innate and adaptive immune system by binding to purinergic receptors. Binding to certain purinergic receptors leads to a pro-inflammatory microenvironment and promotes allogeneic T cell priming. After priming, T cells migrate to the acute GVHD target organs, mainly skin, liver and the gastrointestinal tract and induce cell damage which further amplifies the release of intracellular components. This review summarizes the role of different purinergic receptors in particular P2X7 and P2Y2 as well as nucleotides in the pathogenesis of GVHD

Oral Microbial Flora in Bulgarian Adolescents with Moderate Plaque-induced Gingivitis

Introduction: In children and adolescents, the most common periodontal disease is the plaque-induced gingivitis.Aim: The aim of this study was to reveal the bacterial species associated with supragingival plaque of Bulgarian adolescents diagnosed with plaque-induced gingivitis.Materials and methods: Supragingival plaque samples from 70 healthy subjects with moderate plaque-induced gingivitis (37 females and 33 males), aged 12-18 years, were obtained and examined microbiologically.Results: A total of 224 microorganisms were isolated. Gram-negative bacteria were predominant compared to Gram-positive [132 (59%) vs. 92 (41%), p<0.001]. Aerobic microorganisms were detected more often than anaerobic (151; 67.5% vs. 73; 32.5%, p<0.001). The Streptococcus mutans group and Neisseria spp. were isolated from all adolescents. The frequency of isolation of C. albicans was relatively lower – 11 (15.7%). The anaerobes showed much greater microbial diversity (12 pathogen groups were isolated). Gram-negative rods were isolated from 57 of the adolescents (isolation frequency 81.4%). F. varium, P. melaninogenica, P. intermedia and P. assacharolyticus were detected respectively in 12 (16%), 9 (12%), 8 (11%) and 7 (10%) samples. The less frequently isolated anaerobes were Gram-positive cocci, Gram-negative cocci, Bacteroides uniformis and Bifidobacterium spp. together.Conclusion: The most frequently isolated microbiota in our study is part of the normal oral bacterial flora. The presence of anaerobes such as Prevotella, Fusobacterium, Bacteroides and Porphyromonas reflects the gradual change of the flora to more complex one. The results of quantitative and qualitative evaluation of the plaque of adolescents with moderate plaque-induced gingivitis may contribute to the selection of the prevention and treatment of this disease

The role of checkpoint blockade after allogeneic stem cell transplantation in diseases other than Hodgkin’s Lymphoma

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative treatment option for many malignant high-risk hematological diseases. The Graft-vs.-Tumor (GvT) effect is the major hallmark of this treatment approach. However, disease relapse remains a major limitation. Boosting the GvT effect by checkpoint inhibitors (CI) is an attractive option in this desperate situation although potentially triggering Graft-vs.-Host Disease (GvHD). Early reports in patients with Hodgkin's lymphoma support the idea that CI therapy after HSCT is feasible and effective. We have retrospectively analyzed CI therapy for treatment of disease recurrence after allo-HSCT other than Hodgkin's lymphoma including 21 patients from eight German transplant centers. The median follow-up was 59 days. The overall response rate (ORR) was 43%. Patients receiving donor lymphocyte infusion (DLI) in combination with CI had superior response (ORR 80%). Severe acute GvHD grade III-IV and moderate to severe chronic GvHD were observed in 29% of all patients. Taken together, CI therapy in relapsed patients after HSCT, especially in combination with DLI, is effective but induces severe GvHD in a considerable proportion of patients. Thus, prospective trials or EBMT registry-based validation of different dosing and application schedules including immunosuppressive regimens in those patients are urgently needed

GvHD-associated, inflammasome-mediated loss of function in adoptively transferred myeloid-derived suppressor cells

Myeloid-derived suppressor cells (MDSC) are a naturally occurring immune regulatory population associated with inhibition of ongoing inflammatory responses. In vitro generation of MDSC from bone marrow have been shown to enhance survival in an acute model of lethal graft-versus-host disease (GvHD). However, donor MDSC infusion only partially ameliorates GvHD lethality. In order to improve the potential therapeutic benefit and ultimately survival outcomes we set out to investigate the fate of MDSC after transfer in the setting of acute GvHD (aGvHD). MDSC transferred to lethally irradiated recipients of allogeneic donor hematopoietic grafts are exposed to an intense inflammatory environment associated with aGvHD, which we now show directly undermines their suppressive capacity. Under conditioning regimen and GvHD inflammatory settings, MDSC rapidly lose suppressor function and their potential to inhibit GvHD lethality, which is associated with their induced conversion towards a mature inflammasome-activated state. We find even brief in vitro exposure to inflammasome-activating mediators negates the suppressive potential of cultured murine and human-derived MDSCs. Consistent with a role for the inflammasome, donor MDSC deficient in the adaptor ASC (Apoptosis-associated speck-like protein containing a CARD), that assembles inflammasome complexes, conferred improved survival of mice developing GvHD compared to wild-type donor MDSC. These data suggest the use of MDSC as a therapeutic approach for preventing GvHD and other systemic inflammatory conditions will be more effective when combined with approaches limiting in vivo MDSC inflammasome activation empowering MDSCs to maintain their suppressive potential

Immunization against poly--acetylglucosamine reduces neutrophil activation and GVHD while sparing microbial diversity

Microbial invasion into the intestinal mucosa after allogeneic hematopoietic cell transplantation (allo-HCT) triggers neutrophil activation and requires antibiotic interventions to prevent sepsis. However, antibiotics lead to a loss of microbiota diversity, which is connected to a higher incidence of acute graft-versus-host disease (aGVHD). Antimicrobial therapies that eliminate invading bacteria and reduce neutrophil-mediated damage without reducing the diversity of the microbiota are therefore highly desirable. A potential solution would be the use of antimicrobial antibodies that target invading pathogens, ultimately leading to their elimination by innate immune cells. In a mouse model of aGVHD, we investigated the potency of active and passive immunization against the conserved microbial surface polysaccharide poly--acetylglucosamine (PNAG) that is expressed on numerous pathogens. Treatment with monoclonal or polyclonal antibodies to PNAG (anti-PNAG) or vaccination against PNAG reduced aGVHD-related mortality. Anti-PNAG treatment did not change the intestinal microbial diversity as determined by 16S ribosomal DNA sequencing. Anti-PNAG treatment reduced myeloperoxidase activation and proliferation of neutrophil granulocytes (neutrophils) in the ileum of mice developing GVHD. In vitro, anti-PNAG treatment showed high antimicrobial activity. The functional role of neutrophils was confirmed by using neutrophil-deficient mice that had no survival advantage under anti-PNAG treatment. In summary, the control of invading bacteria by anti-PNAG treatment could be a novel approach to reduce the uncontrolled neutrophil activation that promotes early GVHD and opens a new avenue to interfere with aGVHD without affecting commensal intestinal microbial diversity