Levels Of MDC Research Articles

Identification of new bioactive molecules in platelet preparation, storage, and transfusion reactions for improved transfusion management

Platelet concentrates (PCs) intended for transfusion contain bioactive molecules that can be considered Biological Response Modifiers (BRMs), mainly originating from plasma regardless of the preparation process. During storage, NGAL and GDF-15 levels increase in single donor apheresis platelet concentrates (SDA-PC), whereas in buffy coat platelet concentrates (BC-PC), the levels of MIP1α, MCP-3, and HSAA increase, and GDF-15 levels decrease. These molecules, primarily released by leukocytes, may contribute to adverse reactions (ARs) following a PC transfusion. Notably, in SDA-PC or BC-PC transfusions that result in ARs, the levels of NGAL, HSAA, and GDF-15 are significantly elevated, while the levels of MDC and CX3CL1 are significantly reduced compared to transfusions without ARs. These biomarkers could potentially serve as predictors for PCs-induced ARs.

PBMC-Humanized Mouse Model to Assess Individual Variability in Efficacy and Toxicity of Universal CD19 Chimeric Antigen Receptor T Cell Immunotherapy

Chimeric antigen receptor T cell (CART) immunotherapy has been accumulating extraordinary breakthroughs in fighting against hematological malignancies. Particularly, universal, “off-the-shelf”, CARTs (uCARTs) are emerging as an attractive alternative to patient-derived CARTs, offering cost-effectiveness, immediate availability for treatment, and independence from patients' lymphopenic conditions. However, uCARTs share the same safety concerns with autologous CARTs while the efficacy and persistence of uCARTs vary by each product and remain uncertain. Further, T cells sourced from healthy donors for uCARTs may manifest individual-specific immune characteristics upon interaction with cancer cells and the patient's immune system. These characteristics often remain latent when assessed in vitro and complicate the prediction of uCART therapy's toxicity and efficacy on an individual basis. We previously demonstrated that the PBMC-humanized mouse model can capture efficacy and toxicity of autologous and allogeneic CD19 CARTs. We showed that the CART-induced toxicity varied by the combination of PBMC donors and CART cell donors. Here, we extend the use of PBMC-humanized mice to assess the potential toxicity, efficacy, and persistence of uCARTs. Female NSG™-DKO mice (n = 19-20 mice per donor) were irradiated and reconstituted with human PBMCs sourced from four selected donors (Donor A, B, C, and D) out of 58 available in our PBMC bank. The selection criteria were based on their pre-characterized in vivo human cytokine induction profiles, specifically IFN-gamma response levels to OKT3 treatment (Donor A: 268 pg/ml; B: 1936 pg/ml; C: 4862 pg/ml; D: 13538 pg/ml). PBMC-humanized mice were grouped into three different CD19 CART treatments; a) Allogeneic CARTs were manufactured from a healthy donor's T cells containing CD28 costimulatory domain (19-28z) and no genetic modification was made for “off-the-shelf” availability; b) uCART 1 cells (19-28z) were knocked out of TCRab via CRISPR-cas9 (19-28z); c) uCART 2 cells were knocked out of CD3 and TCRab using TALENs and magnetically depleted for remaining CD3/TCRab, and contain 4-1BB costimulatory domain. Body weight change and clinical scores were measured throughout the experiments. We measured the levels of human CD45, human CD19, and CART expansion from blood collected 8 days post-treatment using flow cytometry. We measured 54 human cytokine levels from serum collected 48 hours post-treatment using a multiplex assay. CART-induced toxicity, measured by body weight loss, dramatically varied by PBMC donors and the treatments (Figure 1). uCART 2 induced significant body weight loss in Donor B and D reaching the lowest 6 days post-treatment. Notably, all mice humanized with Donor C and treated with uCART 2 reached a humane endpoint within 12 days post-treatment. We only observed noticeable body weight loss from uCART 1 in Donor C. Allogeneic CART did not induce body weight loss across the four donors. Efficacy, measured by the % of elimination of human CD19 cells in peripheral blood compared to the PBS-treated group, also varied by PBMC donor and the treatments (Figure 2). Overall, Allogeneic CART showed the best elimination of CD19 cells. Mice humanized with Donor D responded with the poorest efficacy of the three CART treatments. CART expansion was observed only in uCART 2 treated mice that reached a humane endpoint within 6 days post-treatment. CART persistence measured 16 days post-treatment was not observed across three CARTs and this corresponds to the clinical data of uCARTs. Principal component analysis of human cytokine data revealed that uCART 2 induced the most distinct cytokine responses compared to the other CARTs, driven by IFN-gamma, IL-10, IL-5, IL-3, and GM-CSF. In Donor-A humanized mice, the same set of human cytokines (IFN-gamma, IL-10, IL-5, IL-3, GM-CSF, IL-13) was induced in response to the three CARTs, while the degree of induction varied by treatment. In contrast, Donor D-humanized mice showed vastly different responses to each treatment; uCART 2 induced significant levels of IL-6, TNF-beta, MIG/CXCL9, MDC, MCP-3, IL-13, and IL-2. In sum, we show that PBMC-humanized mice capture individual variations in the toxicity and efficacy of uCARTs. This platform can be used to preclinically assess efficacy and persistence of uCARTs as well as used to predict toxicity from the interaction between the host and healthy T cell donor of uCARTs.

MDC/CCL22 depletion in COVID-19 and post-COVID

In this article, we explore the role of macrophage-derived chemokine (MDC/CCL22) in COVID-19 immunity. The study included plasma samples of 289 patients with PCR-verified COVID-19 from specialized hospitals. The blood samples were collected at admission, approximately 7 days after the start of infection. Genetic testing of the virus was performed in nasopharyngeal swabs to determine the viral strain for each patient. We also included blood plasma of 69 convalescent patients who had recovered from COVID-19 more than a month prior to the study. Additionally, 51 healthy donors were included in the study as controls. The concentrations of MDC/CCL22 and other cytokines and chemokines were measured with multiplex analysis using Luminex MagPix Technology. The results showed that COVID-19 patients had significantly lower MDC levels in their plasma, regardless of the SARS-CoV-2 strain, compared to healthy donors. This finding suggests that MDC/CCL22 depletion may play a role in COVID-19 immunity. Furthermore, convalescent patients still showed decreased concentrations of MDC/CCL22 more than a month after infection, indicating that this depletion may persist even after recovery. We propose two mechanisms that can explain the reasons leading to MDC/CCL22 depletion. The first is binding and inactivation of this chemokine with SARS-CoV-2 peptides, making it not only undetectable for commercial kits, but also less functionally active. Another mechanism is the dysfunction of its effector cells (e.g., DCs and macrophages). Lymphopenia following COVID-19 can potentially be explained by the absence of MDC/CCL22. This may lead to a shift towards hyperactivation in the inflammatory response, potentially explaining the severity of COVID-19. This research sheds light on the importance of MDC/CCL22 in COVID-19 immunity and highlights the need for further investigation into its role in the disease. Understanding the mechanisms behind MDC/CCL22 depletion could provide new insights into the pathogenesis of COVID-19 and inform the development of potential treatments.

Cytokine profiling in patients with hepatic glycogen storage disease: Are there clues for unsolved aspects?

Hepatic Glycogen Storage Diseases (GSD) are rare genetic disorders in which the gluconeogenesis pathway is impaired. Cytokines control virtually every aspect of physiology and may help to elucidate some unsolved questions about phenotypes presented by GSD patients. This was an exploratory study in which 27 GSD patients on treatment (Ia=16, Ib=06, III=02, IXα=03) and 24 healthy age- and sex-matched subjects had plasma samples tested for a panel of 20 cytokines (G-CSF,GM-CSF, IL-1α,IL-1β, IL-4, IL-6, IL-8, IL-10, IL-13, IL-17A, GRO, IP-10/CXCL10, MCP-1/CCL2, MIP-1α/CCL3, MIP-1β/CCL4, MDC/CCL22, IFN-γ, TNF-α, TNF-β, VEGF) through a multiplex kit and analyzed in comparison to controls and among patients, regarding to clinical features as anemia, hepatic adenocarcinoma and triglyceride levels. Patients (GSD-Ia/III/IX) presented reduced levels of IL-4 (p=0.040), MIP-1α/CCL3 (p=0.003), MDC/CCL22 (p<0.001), TNF-β (p=0.045) and VEGF (p=0.043) compared to controls. When different types of GSD were compared, G-CSF was higher in GSD-Ib than -Ia (p<0.001) and than -III/IX (p=0.033) patients; IL-10 was higher in GSD-Ib than in GSD-Ia patients (p=0.019); and GSD-III/IX patients had increased levels of IP-10/CXCL10 than GSD-Ib patients (p=0.019). When GSD-I patients were gathered into the same group and compared with GSD-III/IX patients, IP10/CXCL10 and MCP-1 were higher in the latter group (p=0.005 and p=0.013, respectively). GSD-I patients with anemia presented higher levels of IL-4 and MIP-1α in comparison with patients who had not. Triglyceride level was correlated with neutrophil count and MDC levels on GSD-Ia patients without HCA. Altogether, altered levels of cytokines in GSD-I patients reflect an imbalance in immunoregulation process. This study also indicates that neutrophils and some cytokines are affected by triglyceride levels, and future studies on the theme should consider this variable.

Serum CC Chemokines as Potential Biomarkers for the Diagnosis of Major Depressive Disorder.

ObjectiveEvidence indicates a potential role of chemokines in depression-like behavior and depression-related pathophysiological processes. In the present study, we examined the serum levels of multiple chemokines, focusing on CC chemokines, in patients with major depressive disorder (MDD), with the aim to discover and identify serum chemokines-based biomarkers for MDD diagnosis.MethodsParticipants included 24 patients with MDD and 24 healthy controls. The 24-item Hamilton Depression Rating Scale (HAMD-24) was administered to assess the disease severity of patients with MDD. A total of 9 serum CC chemokines including MCP-1 (CCL-2), MIP-1α (CCL-3), MIP-1β (CCL-4), eotaxin-1 (CCL-11), MCP-4 (CCL-13), TARC (CCL-17), MIP-3α (CCL-20), MDC (CCL-22), and Eotaxin-3 (CCL-26) were measured using electrochemiluminescence immunoassays. The levels of serum CC chemokines between MDD group and control group were compared, and diagnostic values of different CC chemokines were evaluated using the receiver operating characteristic (ROC) curve method for discriminating MDD patients from healthy controls. Correlations between the levels of serum CC chemokines and depression severity (HAMD-24 scores) were evaluated using Pearson’s correlation test.ResultsPatients with MDD had higher levels of serum MIP-1α and MIP-1β and lower levels of serum MCP-1, MCP-4, TARC, MDC, and Eotaxin-3 compared to controls (all P < 0.05). Moreover, ROC curve analysis showed that the Area Under Curve (AUC) values of MIP-1α, MCP-4, TARC, and Eotaxin-3 were > 0.7 in discriminating patients with MDD from healthy controls. Furthermore, no significant relationship was found between the levels of serum CC chemokines and HAMD-24 scores in MDD group.ConclusionThese results suggested that circulating CC chemokines may hold promise in the discovery of biomarkers for diagnosing MDD.

Terminalia chebula Retz. extract ameliorates the symptoms of atopic dermatitis by regulating anti-inflammatory factors in vivo and suppressing STAT1/3 and NF-ĸB signaling in vitro

BackgroundTerminalia chebula (TC) is a traditional medicinal plant used for treating various diseases in humans. However, pharmacological mechanisms underlying the effects of TC in atopic treatment remain unelucidated. Hypothesis/purposeWe investigated the therapeutic effects of TC extract in a mouse model of atopic dermatitis (AD) in vivo and the anti-inflammatory mechanism in vitro. Study design/methodsFor the in vivo study, AD was induced by Dermatophagoides farinae extract (Dfe) in NC/Nga mice. After 14 days of oral administration, the effects of TC concentrations of 30, 100, and 300 mg/kg were analyzed by assessing morphological changes visually; measuring serum levels of inflammatory chemokines/cytokines, IgE, histamine, MDC, TARC, RANTES, and TSLP using ELISA kits; and counting infiltrated mast cells. For in vitro analyses, we used IFNγ/TNF-α-stimulated human keratinocyte cell lines to study the mechanism of action. The production of chemokines/cytokines in the IFNγ/TNF-α-stimulated HaCaT cells was measured using ELISA and a bead array kit. The signaling pathways were analyzed by western blotting and the expression of the transcriptional factors using RT-PCR and luciferase assay. ResultsAdministration of TC significantly alleviated AD-like symptoms in vivo and decreased the ear thickness, dermatitis score, keratinization, and mast cell infiltration. It also resulted in decreased serum levels of IgE, histamine, and inflammation-related mediators MDC, TARC, RANTES, and TSLP compared with those in the Dfe treatment group. Moreover, TC downregulated the expression of the inflammatory chemokines RANTES and MDC in IFNγ/TNF-α-stimulated HaCaT cells. TC inhibited phosphorylated STAT1/3 and NK-κB subunits and nuclear translocation of NF-κB. It also suppressed the transcription of IFNγ, IL-6, IL-8 and MCP-1 in the IFNγ/TNF-α-stimulated HaCaT cells. TC and its constituents, chebulic acid, gallic acid, corlagin, chebulanin, chbulagic acid, ellagic acid, and chebulinic acid, strongly inhibited the nuclear translocation of NF-κB, STAT1, and STAT3 and decreased the expression of inflammatory cytokines at the mRNA level. ConclusionsOverall, TC extract alleviated AD-like symptoms by regulating anti-inflammatory factors in vivo and suppressing STAT1/3 and NF-κB signaling in vitro. In addition, our results show the in vivo effect of partial improvements in AD, as well as the in vitro effect on inflammatory factors by the constituents of TC. This finding provides that TC extract and its components could be potential therapeutic drugs for AD.

Inhibitory Effects of Myrrha Water Extract on Atopic Dermatitis Related Chemokines in HaCaT Cells

Due to high therapeutic potentials of natural products, the use of medicinal plants is highlighted to treat the various inflammationmediated diseases. Myrrha have been used as a traditional remedy to treat infectious and inflammatory diseases. But, its effects and mechanisms of anti atopic dermatitis (AD) have not been elucidated. The aim of this study is to evaluate anti-allergic and antiinflammatory effects of the water extract of Myrrha, in cell models and also to suggest a putative mechanism of AD actions of Myrrha. HaCaT cells were pre-treated with Myrrha for 1 h and stimulated with tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN- γ) (10 ng/mL each). After 24 h, cells were harvested To evaluate the expression of Th2 chemokines, such as C-C motif chemokine ligand 5 (CCL5, also known as RANTES), C-C chemokine ligand 17 (CCL17, also known as TARC) and C-C chemokine ligand 22 (CCL22, also known as MDC). To investigate the regulatory mechanisms of Myrrha, we also assessed the phosphorylation of signal transducer and activator of transcription 1 (STAT1) signaling pathways in HaCaT cells. Treatment of Myrrha decreased the mRNA levels of RANTES, TARC and MDC with a concentration dependent manner. In addition, Myrrha significantly reduced TNF-α and IFN-γ induced phosphorylation of STAT1. This could indicate that the Myrrha shows anti AD activity mainly through STAT1. Thus, we propose that Myrrha may be a promising anti AD skin protector, which could suggest the clinical basis for cosmetics development.

P.091 Multi-modal analysis of outcomes in pediatric mild traumatic brain injury (mTBI)

Background: mTBI is the most common type of head injury among children but diagnosing and managing symptoms are challenging due to heterogeneity. This study used multi-modal analysis to examine how acute mTBI symptoms transition to chronic deficits. Methods: Subjects included a cohort with mild TBI (n=40, 8-18 years old) and age/sex-matched controls (n=27). All participants received symptom assessment, neuropsychological evaluation, ERP assessment, neuroimaging, and serum cytokine analysis. Results were analyzed individually and in multi-modal models to identify important outcome predictors. Results: mTBI resulted in higher symptom burdens compared to controls. There were no group differences in measures of balance, ERP, FA, or MD. Female mTBI participants had lower CNSVS Neurocognition Index scores (p=0.0401) and faster reaction times (p=0.0385) than controls. Repetitive mTBI males had faster psychomotor speed than symptomatic mTBI males (p=0.0260). CTACK levels were higher in female mTBI groups (p= 0.0043), SCGF- levels were lower in male mTBI groups (p=0.0486), and MDC levels were lower in female mTBI groups (p=0.0377) compared to controls. Multi-modal models revealed key predictors from all modalities, despite most measures producing non-statistically significant results in individual analyses. Conclusions: Multi-modal analysis may afford the opportunity to delineate complex mTBI pathology and provide better identification of biomarkers than unimodal analysis.

Arthroplasty surgery alters the local immune composition of the knee joint by causing increases in specific lymphocyte populations.

Abstract Objective We aimed to determine how the innate and adaptive immune cell composition and cytokine profiles of the knee joint microenvironment are altered in replaced knees compared to native knees. Methods Upon IRB approval, we obtained peripheral blood (PBMC) and synovial fluid (SF) from 14 native knees with osteoarthritis (OA) and 12 replaced knees at least 6 months after arthroplasty surgery, comprising 26 total patients. All PBMC and SF samples were analyzed by Flow Cytometry, and utilizing multidimensional reduction analysis via TriMap and clustering analysis by FlowSOM, cell clusters were phenotyped for identification of unique populations of cells. Additionally all samples were analyzed for cytokine/chemokine profiles using 65-Plex ProcartaPlex assays. Results Overall, there was no significant different in any of the innate cell populations studied, including neutrophils, monocytes (including G-MDSCs), macrophages (M1 of M2 phenotypes), and dendritic cells. B cells, CD8 effector T cells, CD4 T cells, Naive CD4 T cells, Th17 cells, and CD4 T cells with Th1-phenotypes s were all significantly increased in replaced knees compared to native knees (p &amp;lt; 0.02 in each case). The replaced knee group showed elevated levels of CXCL5, HGF, IFN-gamma, IL-6, IL-16, CXCL10, CCL2 (MCP-1), CCL8, MDC, TNF-alpha, and VEGF-A compared to the native knee group (p &amp;lt; 0.01 for all) and a decrease in CX3CL1, CXCL11, IL-3, IL-5, CXCL9, TRAIL (p &amp;lt; 0.02 in all cases). Conclusions Arthroplasty surgery results in a lymphocytic shift in immune cell populations and a generally pro-inflammatory cytokine/chemokine profile. Overall, this lymphocytic shift may have implications for development of periprosthetic infections and warrants further study.

The phytoactive constituents of Eugenia selloi B.D. Jacks (pitangatuba): Toxicity and elucidation of their anti-inflammatory mechanism(s) of action

We determined the phytochemical composition, anti-inflammatory mechanism of action, ROS/RNS scavenging capacity and systemic toxicity of a purified subfraction (S8) of Eugenia selloi. The composition of S8 was assessed by LC-ESI-QTOF-MS; the anti-inflammatory activity in RAW264.7 macrophages through NF-κB activation and biomarkers by multiplex in THP-1 cells; neutrophil migration, intravital microscopy and ICAM-1 expression in mice; NETs formation and CD11b expression; S8 scavenging capacity of ROS/RNS; toxicity in Galleria mellonella larvae model. Coumaric acid, quercetrin and vanillic acid were identified. S8 decreased NF-κB activation, IL-1β, IL-6, IL-10, MDC and MCP-1 levels, reduced neutrophil migration and ICAM-1 expression in mice; S8 did not interfere NET formation and CD11b expression, exhibited high antioxidant and showed negligible toxicity. E. selloi proved to be a promising, yet underexplored source of bioactive compounds, which can be useful employed in agribusiness and in the pharmaceutical and food industry to develop new products or human health supplies.

Expression and significance of TOLL2, TARC and MDC in placenta tissue of pregnant patients infected with syphilis

This study was developed to investigate the expression of TOLL2, TARC and MDC in placenta tissue of pregnant patients infected with syphilis and their clinical significance. For this aim, placenta samples were collected from five pregnant patients co-infected with syphilis and five undergoing full-term delivery before RT-PCR was performed to detect the mRNA expression of TLR2, TARC and MDC genes. The protein expression of TLR2, TARC and MDC genes was examined by Western Blotting. Results showed that TLR2, TARC and MDC were expressed in placental syncytiotrophoblast cells of patients with pregnancy-associated syphilis infection. TLR2 level was found significantly higher in placenta tissue of patients with pregnancy-associated syphilis infection compared with normal placenta tissue (P<0.05), so were TARC (P<0.05) and MDC genes (P<0.05). It is concluded that TOLL2, TARC and MDC levels significantly increased in the placenta tissue of pregnant patients infected with syphilis, suggesting that the three genes were involved in the molecular pathology of the patients.

​Circulating Cytokines in Myocardial Infarction Are Associated With Coronary Blood Flow.

BackgroundThe level of systemic inflammation correlates with the severity of the clinical course of acute myocardial infarction (AMI). It has been shown that circulating cytokines and endothelial dysfunction play an important role in the process of clot formation. The aim of our study was to assess the concentration of various circulating cytokines, endothelial function and blood clotting in AMI patients depending on the blood flow through the infarction-related artery (IRA).MethodsWe included 75 patients with AMI. 58 presented with ST-elevation myocardial infarction (STEMI) and 17 had non-ST-elevation myocardial infarction (non-STEMI). A flow-mediated dilation test (FMD test), thrombodynamics and rotational thromboelastometry as well as assessment of 14 serum cytokines using xMAP technology were performed.FindingsNon-STEMI-patients were characterized by higher levels of MDC, MIP-1β, TNF-α. Moreover, we observed that patients with impaired blood flow through the IRA (TIMI flow 0-1) had higher average and initial clot growth rates, earlier onset of spontaneous clots, C-reactive protein (CRP) and IL-10 compared to patients with preserved blood flow through the IRA (TIMI flow 2-3). Patients with TIMI 2-3 blood flow had higher level of IP-10. IL-10 correlated with CRP and pro-inflammatory cytokines levels, initial clot growth rate and clot lysis time in TIMI 0-1 patients. All these differences were statistically significant.InterpretationWe demonstrated that concentrations of the inflammatory cytokines correlate not only with the form of myocardial infarction (STEMI or non-STEMI), but also with the blood flow through the infarct-related artery. Inflammatory response, functional state of endothelium, and clot formation are closely linked with each other. A combination of these parameters affects the patency of the infarct-related artery.

Exposure to widespread drinking water chemicals, blood inflammation markers, and colorectal cancer.

Trihalomethanes (THMs) and nitrate are widespread chemicals in drinking water associated with colorectal cancer risk but mechanisms are not well understood. We explored the association between exposure to THMs and nitrate in drinking water and inflammation markers, and the link with colorectal cancer risk. A subset of 198 colorectal cancer cases and 205 controls from the multicase-control study MCC-Spain were included. Average concentration of THMs (chloroform, bromodichloromethane, dibromochloromethane, bromoform) and nitrate in tap water at the residence was estimated from age 18 until 2years before the interview ("long term") and for a recent period (3years before diagnosis). Serum levels of EGF, eotaxin, G-CSF, IL-17E, IL-1rA, IL-8, IP-10, MDC, MPO, periostin, VEGF, and C-reactive protein (CRP) were measured. We estimated the linear association between inflammation markers and exposure among controls, and the odds ratio of colorectal cancer associated with THM and nitrate exposure, and inflammation markers. A mediation analysis was conducted to identify inflammation markers in the pathway between THM/nitrate exposure and colorectal cancer. Serum concentrations of EGF, IL-8, IL-17E and eotaxin increased with recent residential levels of brominated THMs, chloroforom and/or total THM. No associations were observed for nitrate and for long-term residential THM levels. All residential exposures except chloroform were positively associated with colorectal cancer. Serum concentrations of VEGF and periostin were positively associated with colorectal cancer, while EGF was inversely associated. One protein-exposure combination (periostin-recent ingested brominated THMs) slightly mediated the association with colorectal cancer risk. Results suggest that estimated THM exposure is involved in inflammation processes. However, the study design was limited to stablish etiologically relevant associations between the protein levels and colorectal cancer risk. The lack of association between nitrate exposure and inflammation markers suggests other biological mechanisms are involved in the link with colorectal cancer.

ИММУНОЛОГИЧЕСКИЕ ПРЕДИКТОРЫ ОТТОРЖЕНИЯ ПОЧЕЧНОГО ТРАНСПЛАНТАТА В РАННЕМ ПОСЛЕОПЕРАЦИОННОМ ПЕРИОДЕ

Objective. To determine the immunological predictors of renal graft rejection in the early postoperative period. Methods. Three groups were formed out of the 197 renal graft recipients. The group PGF (n=101) was made up of patients with satisfactory primary graft function. The group PGD (n = 82) included patients with primary graft dysfunction without episodes of rejection. The group RGR (n=14) consisted of patients with primary dysfunction and renal graft rejection. On the 7&lt;sup&gt;th&lt;/sup&gt; day after transplantation the early kidney graft function was assessed on the basis ofserum creatinine levels. When the serum creatinine value was lower than 300 μmol/L the function was considered to be primary, at a creatinine concentration was equal to or higher than 300 μmol/L, as well as in the case of needfor maintenance dialysis on the first week after transplantation, the state was classified as the renal graft dysfunction. In the early postoperative period, the number of LIN-HLA-DR+ dendritic cells with the LIN-HLA-DR+CD11c+CD123- and LIN-HLA-DR+CD11c-CD123+ phenotypes in the fluid from the drainage installed to the kidney graft during surgery was determined. Predictive characteristics of the mDC and pDC levels in the drainage fluid were determined to predict renal graft rejection, and diagnostic capability of this indicator were identified. Results. It has been revealed that renal graft rejection is characterized by a significant growth of the total number of dendritic cells in the drainage fluid, mainly due to myeloid ones. Predictive characteristics were determined by the level of myeloid and plasmacytoid dendritic cells in the drainage fluid. The cut-off point of the level of myeloid dendritic cells was determined at the level of 60.32%, and for plasmacytoid dendritic cells it corresponded to 39.68%. Conclusion. With the level of myeloid dendritic cells in the drainage fluid greater or equal 60.32%, and plasmacytoid cells lower or equal 39.68%, renal graft rejection is predicted with a sensitivity of 99% and 93%, respectively, and a specificity of 89% and 91%, respectively. What this paper adds The level of dendritic cells and their subpopulations in the drainage fluid in renal graft recipients has been firstly studied. It has been established that acute renal graft rejection is associated with a high concentration of the total number of dendritic cells in the drainage fluid. More over this increase occurs mainly due to myeloid dendritic cells. The determination of the level of myeloid and plasmacytoid dendritic cells in the drainage fluid can be used as a predictor of renal graft rejection.

Effects of chloroform fraction of Fritillariae Thunbergii Bulbus on atopic symptoms in a DNCB-induced atopic dermatitis-like skin lesion model and in vitro models.

Ethnopharmacological relevanceFritillariae thunbergii Bulbus (FT), knowns as “Jeolpaemo (浙貝母)” in Korean traditional medicine, is a perennial plant belonging to the Liliaceae family and has been used to treat symptoms such as cough, sputum formation, and purulent pneumonia. Owing to its effects of lowering heat, removing sputum, and reducing swelling, the plant has also been used as an external prescription medicine to treat inflammation. Aim of the studyTo analyze the anti-inflammatory effects of FT-ethanol extract (FT-Et) and FT-chloroform fraction extract (FT-Cl) on 1-chloro-2,4-dinitrobenzene (DNCB)-induced atopic dermatitis (AD) in vivo and in vitro. Materials and methodsThe effect of FT-Et and FT-Cl on AD was observed using an AD-like skin lesion model induced by DNCB in vivo. HaCaT and RBL2H3 cells were used to determine the effects of FT-Et and FT-Cl in vitro. After inducing AD-like skin lesions in vivo, FT was topically applied to the skin lesion for 35 days. Epidermal thickness, dermal thickness, scratching behavior, infiltration of inflammatory cells, and expression of skin barrier proteins were measured. TARC, MDC, and IL-4 levels were analyzed using ELISA in HaCaT cells. Beta-hexosaminidase and IL-4 levels were measured in RBL2H3 cells. The expression of filaggrin (FLG), loricrin (LOR), involucrin (INV), and aquaporin-3(AQP-3) was measured by PCR. Phosphorylation of MAPKs was analyzed using Western blot technique. ResultsFT-Cl significantly reduced ear swelling, scratching behavior, SCORAD index, epidermal thickness, infiltration of inflammatory cells, and loss of skin barrier proteins. FT-Et inhibited the infiltration of mast cells and CD8+ cells and decreased the loss of skin barrier proteins. In TNF-α/IFN-γ-stimulated HaCaT cells, FT-Cl inhibited TRAC, MDC, and IL-4 expression and upregulated the expression of FLG, INV, and AQP-3, whereas FT-Et inhibited the expression of TRAC and MDC and increased the expression of FLG, INV, and AQP-3 at high concentrations. In RBL2H3, FT-Cl downregulated β-hexosaminidase and IL-4 expression. In addition, FT-Cl inhibited the phosphorylation of ERK and p-38 in HaCaT and RBL2H3 cells. ConclusionsCollectively, FT-Cl showed better effect than FT-Et in vivo and in vitro. These results suggest that a specific component present in FT-Cl acted against AD. Future research should focus on the analysis of components contained in FT-Cl and the anti-inflammatory effects of the active ingredient.

Small blood stem cells for enhancing early osseointegration formation on dental implants: a human phase I safety study

BackgroundSmall blood stem cells (SB cells), isolated from human peripheral blood, demonstrated the ability to benefit bone regeneration and osseointegration. The primary goal of our study is to examine the safety and tolerability of SB cells in dental implantation for human patients with severe bone defects.MethodsNine patients were enrolled and divided into three groups with SB cell treatment doses of 1 × 105, 1 × 106, and 1 × 107 SB cells, and then evaluated by computed tomography (CT) scans to assess bone mineral density (BMD) by Hounsfield units (HU) scoring. Testing was conducted before treatment and on weeks 4, 6, 8, and 12 post dental implantation. Blood and comprehensive chemistry panel testing were also performed.ResultsNo severe adverse effects were observed for up to 6-month trial. Grade 1 leukocytosis, anemia, and elevated liver function were observed, but related with the patient’s condition or the implant treatment itself and not the transplantation of SB cells. The levels of cytokines and chemokines were detected by a multiplex immunological assay. Elevated levels of eotaxin, FGF2, MCP-1, MDC, and IL17a were found among patients who received SB cell treatment. This observation suggested SB cells triggered cytokines and chemokines for local tissue repair. To ensure the efficacy of SB cells in dental implantation, the BMD and maximum stresses via stress analysis model were measured through CT scanning. All patients who suffered from severe bone defect showed improvement from D3 level to D1 or D2 level. The HU score acceleration can be observed by week 2 after guided bone regeneration (GBR) and prior to dental implantation.ConclusionsThis phase I study shows that treatment of SB cells for dental implantation is well tolerated with no major adverse effects. The use of SB cells for accelerating the osseointegration in high-risk dental implant patients warrants further phase II studies.Trial registrationTaiwan Clinical Trial Registry (SB-GBR001) and clinical trial registry of the United States (NCT04451486).

Day 100 Levels of Blood Immune Cells Are Prognostic Biomarkers for Allogeneic Hematopoietic Stem Cell Transplant Outcomes: Results from BMT CTN 0201

Background: BMT CTN 0201 was a randomized trial of bone marrow (BM) vs. G-CSF mobilized blood (G-PB) grafts in 550 pediatric and adult patients with leukemia or MDS undergoing allogeneic hematopoietic stem cell transplantation from unrelated donors. Overall survival (OS) was equivalent between both arms, with more chronic graft-versus-host disease (GvHD) among recipients of G-PB grafts and more graft failure among recipients of BM grafts (Anasetti NEJM 2012; 367:1487-1496). We have previously reported donor plasmacytoid dendritic cell (pDC) content in BM grafts are linked with improved survival and less death from acute GvHD (Waller JCO 2014 32:2365-2372). Here, we compare the kinetics of immune cell reconstitution between BM and G-PB recipients to test the hypothesis that graft source leads to differences in blood levels of donor-derived immune cells, and that higher levels of dendritic cells are associated with lower GvHD-related death rates.Methods: Demographics among randomized patients were similar with respect to diagnosis, conditioning regimens, GvHD prophylaxis, and CMV serostatus. Flow cytometry measured immune cells in blood samples at 1, 3, 6, 12, and 24 months post-transplant in recipients of BM (N=161) or G-PB (N=147) grafts from unrelated donors. Univariate and multivariable analyses evaluated associations between donor immune cells and outcomes.Results: Reconstitution kinetics of immune cells at day +100 were associated with graft source, a history of GvHD, and subsequent GvHD-related death.G-PB recipients had higher 1 and 3-month levels of blood T cells and B cells than BM recipients, as well as higher numbers of γ/δ, CD4+ T-reg, CD4+ T cells at later time points (Table 1). In contrast, recovery kinetics for pDC, mDC,and immature CD56+CD16- NK cellswere similar between each graft arm (Table 1). B cell reconstitution kinetics were slower in BM recipients, especially in BM recipients who developed acute GvHD prior to day +100 as opposed to those who did not (in Figure 1, p < .001). No significant association existed between aGvHD history and lymphocyte reconstitution in G-PB recipients.Blood levels of immune cells at day +100 were associated both with a history of acute GvHD as well as subsequent 2-year OS, and GvHD-related death. All patients who developed grades 2-4 acute GvHD by day +100 had worse 2-year OS than non-GvHD patients (66% vs. 53%, p=.01). However, patients with a history of GvHD and >median numbers of blood mDC, CD4+, or CD8+ T cells at day +100 had better 2-year OS (95% CI): [mDC, 63 (51-74)% vs. 43 (32-55)% p=.02; CD4+ T cells, 64 (53-75)% vs. 42 (31-53)%, p=0.007; CD8+ T cells, 65 (54-76)% vs. 42 (31-53)%, p= 0.004] than GvHD-patients with <median numbers of these cells. Furthermore, significantly fewer GvHD-related deaths occurred in patients with >median numbers of mDC compared to in those with <median numbers of mDC (23% vs 40%, respectively, p<.05).In contrast, G-PB recipients with >median numbers of blood central memory CD8+ (p=.01), CD8+ (p=.004), and γ/δ T cells (p=.002) by day +100 had significantly higher 2-year OS compared to those with <median numbers of these cells. BM recipients with >median numbers of mDC had higher 2-year OS compared to those with <median numbers (p=.06).The content of blood mDC was prognostic for post-transplant outcomes for the entire cohort of patients and the subset of BM recipients. Significantly improved 2-year OS (95% CI) was observed in patients with >median numbers of mDC by day +100 post-transplant (68 (61-76)%) compared to those with <median numbers of mDC (51 (43-59)%; p=.008; Figure 2). Compared to the latter, the former also had significantly less (p=.04) treatment-related mortality at 2-years post-transplant (12 (8-18)% vs. 23 (17-31)%) and no difference in relapse (30 (23-38)%) vs. 29 (22-37)%).Conclusions: Graft source is associated with differences in reconstitution kinetics of T and B cells. Blood levels of immune cells at day +100 are cellular bio-markers for both GvHD-related death and OS in transplant recipients, and are correlated with a history of acute GvHD. Specific cell subsets prognostic for OS vary by graft source. Higher levels of blood mDC at day +100 in BM recipients are associated with improved OS, while higher numbers of T-cells predict better OS in GPB recipients. Compared to BM recipients, G-PB recipients have faster T and B cell reconstitution as well as greater resiliency of lymphogenesis following acute GvHD. [Display omitted] DisclosuresLee:Mallinckrodt: Honoraria; Amgen: Other: One-time advisory board member; Bristol-Myers-Squibb: Other: One-time advisory board member; Kadmon: Other: One-time advisory board member. Waller:Helocyte: Consultancy; Novartis Pharmaceuticals Corporation: Consultancy, Honoraria, Research Funding; Coulter Foundation: Research Funding; Katz Foundation: Research Funding; Chimerix: Equity Ownership; AMGEN: Consultancy; Cambium Medical Technologies: Equity Ownership, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Celldex: Consultancy; Cerus: Equity Ownership; National Institutes of Health: Research Funding; PRA: Consultancy.

Prognostic Utility of Serum TARC, MDC, IL10, and Soluble CD163 Levels with Positron Emission Tomography (PET) Response-Adapted Therapy for Advanced-Stage Hodgkin Lymphoma (HL): a SWOG S0816 US Intergroup Correlative Study

Prognostic Utility of Serum TARC, MDC, IL10, and Soluble CD163 Levels with Positron Emission Tomography (PET) Response-Adapted Therapy for Advanced-Stage Hodgkin Lymphoma (HL): a SWOG S0816 US Intergroup Correlative Study

Initial Treatment with Lenalidomide Plus Rituximab for Mantle Cell Lymphoma: 5-Year Follow-up and Correlative Analysis from a Multi-Center Phase II Study

Abstract BACKGROUND: We previously reported early results of a multicenter phase 2 study of lenalidomide plus rituximab as initial treatment for MCL (NEJM 2015:373:1835), which was highly effective (ORR 92%, CR 64%) and well tolerated. Here we present outcomes with 5-year follow-up, as well as exploratory analysis of immunologic biomarkers and minimal residual disease measurement. METHODS: The study includes both induction and maintenance. Lenalidomide was administered at 20 mg daily on days 1-21 of a 28-day cycle for 12 cycles during induction, followed by dose reduction to 15 mg during maintenance. Standard dose rituximab was administered weekly x 4 during cycle 1, then once every other cycle. Treatment was continuous until progression, with option to stop therapy after 3 years. Plasma samples were obtained at baseline, C2D1 and C4D1 to measure cytokines (IFN γ , IL10, IL12p70, IL12, IL1 β , IL2, IL4, IL6, IL8, TNF α) and chemokines (MCP1, MCP4, Eotaxin, IP10, MDC, Eotaxin3, TARC, MIP1 α , MIP1 β, IL8), using MesoScale Discovery V-plex assays. The cytokine/chemokine levels over time, and their associations with responses and treatment-emergent rash were analyzed with Wilcoxon signed-rank and rank-sum tests. MRD was assessed using ClonoSEQTM (Adaptive Biotechnologies, Seattle WA) on subjects with available paired pre-treatment tissue and post-induction PB samples. RESULTS: 38 subjects requiring therapy were enrolled at 4 centers from 7/2011 to 4/2014. The median age was 65 and MIPI scores evenly distributed between low-, intermediate-, and high-risk. Median follow-up was 58 months with range of 36-70 months. Two patients were inevaluable - one withdrew consent, other was intolerant of tumor flare. Twenty-two (61%) of the evaluable 36 patients remain in remission, including 13 (36%) beyond 5 years. Nineteen patients remain on treatment and 3 opted to stop therapy after 3 years. Eleven patients had progression - 3 with primary refractory disease and 8 following initial responses (4 CRs with PFS of 18, 38, 39 and 49 months, 4 PRs with PFS at 14, 25, 28 and 43 months). Six evaluable patients died, 3 from progression, 3 from unrelated comorbidities. Median PFS and DOR have not been reached. The 3-yr and 4-yr PFS rates were estimated at 80.3% (95% CI = 63.0%, 90.1%) and 69.7% (95% CI = 50.6%, 82.6%), with 3-yr and 4-yr OS rates at 91.9% (95% CI = 76.9%, 97.3%) and 82.6% (95% CI = 65.3%, 91.9%). TEAEs during maintenance included asymptomatic grade 3-4 cytopenias (42% neutropenia, 5% thrombocytopenia, 3% anemia), and grade 1-2 infections (45% URI, 16% UTI, 13% sinusitis and 8% cellulitis), all managed in outpatient settings. Five patients required brief hospitalization for IV antibiotics: 1 (3%) developed neutropenic fever in setting of cholecystitis, 3 (8%) had pneumonia and 1 (3%) had recurrent UTI. Secondary malignancies included squamous-cell carcinoma in 2 subjects, basal-cell carcinoma in 1 subject, and melanoma in situ in 2 subjects. Merkel-cell carcinoma developed in an 86-year-old subject after 18 months of therapy, and pancreatic cancer was diagnosed in a 68-year-old patient after 12 months of therapy. Compared to baseline, cytokine levels at C2D1 and C4D1 showed Th1/Th2 modulation, with significant increase in IFNγ (p=0.004 and p=0.007) and IL4 (p=0.004 and p=0.017), as well as significant decrease in IL2 (p=0.046 and p=0.032), IL10 (p=0.004 and p=0.007) and TNFα (p CONCLUSIONS: Lenalidomide and rituximab as initial treatment for MCL can achieve high rate of complete responses and MRD negativity with durable remissions beyond 4 years. The nature of toxicity was not significantly affected by continuous treatment, particularly in the context of close follow-up. Further evaluation of this active regimen in larger, randomized frontline trials is warranted (ClinicalTrials.gov - NCT01472562). Disclosures Ruan: Cell Medica: Research Funding; Pharmacyclics: Research Funding; Seattle Genetics: Consultancy, Research Funding; Celgene: Consultancy, Research Funding. Martin: Celgene: Consultancy; Janssen: Consultancy, Honoraria, Other: travel expenses; Teva: Research Funding; Novartis: Consultancy; Genentech: Consultancy; Gilead: Consultancy, Other: travel expenses. Cerchietti: Celgene: Research Funding; Lymphoma Research Foundation: Research Funding; Leukemia and Lymphoma Society: Research Funding; Weill Cornell Medicine - New York Presbyterian Hospital: Employment. Schuster: Celgene: Consultancy, Research Funding; Seattle Genetics: Consultancy; Novartis: Consultancy, Research Funding; Bristol-Myers Squibb: Consultancy, Research Funding; Genentech: Consultancy, Research Funding; Nordic Nanovector: Consultancy; Gilead: Consultancy, Research Funding; Janssen: Consultancy, Honoraria; Merck: Research Funding. Svoboda: Seattle Genetics: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Pharmacyclics: Research Funding; Kite: Consultancy; Merck: Research Funding; Celgene: Research Funding. Furman: Verastem: Consultancy; Genentech: Consultancy; Janssen: Consultancy, Honoraria; TG Therapeutics: Consultancy; Abbvie: Consultancy, Honoraria; Gilead: Consultancy; Pharmacyclics: Consultancy, Honoraria; Sunesis: Consultancy. Leonard: Roche: Consultancy; Celgene: Consultancy, Research Funding.

Environment-dependent alterations of immune mediators in urban and rural South African children with atopic dermatitis.

In order to improve targeted therapeutic approaches for children with atopic dermatitis (AD), novel insights into the molecular mechanisms and environmental exposures that differentially contribute to disease phenotypes are required. We wished to identify AD immunological endotypes in South African children from rural and urban environments. We measured immunological, socio-economic and environmental factors in healthy children (n = 74) and children with AD (n = 78), in rural and urban settings from the same ethno-linguistic AmaXhosa background in South Africa. Circulating eosinophils, monocytes, TARC, MCP-4, IL-16 and allergen-specific IgE levels were elevated, while IL-17A and IL-23 levels were reduced, in children with AD regardless of their location. Independent of AD, children living in a rural environment had the highest levels of TNFα, TNFβ, IL-1α, IL-6, IL-8, IL-21, MCP-1, MIP-1α, MIP-1β, MDC, sICAM1, sVCAM1, VEGFA, VEGFD and Tie2, suggesting a generalized microinflammation or a pattern of trained immunity without any specific TH polarization. In contrast, IL-15, IL-22, Flt1, PIGF and βFGF were highest in urban children. Rural healthy children had the lowest levels of food allergen-specific IgG4. Early life nutritional factors, medications, animal exposures, indoor environment, sunlight exposure, household size, household income and parental education levels were associated with differences in circulating cytokine levels. This study highlights the immunological impact of environmental exposures and socio-economic status in the manifestation of immune endotypes in children with AD living in urban and rural areas, which are important in selecting appropriately matched immunological therapies for treatment of AD.