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Abstract Background Liver transplantation is the definitive treatment for many end-stage liver diseases. However, the life-long immunosuppression needed to prevent graft rejection causes clinically significant side effects. Cellular immunomodulatory therapies may allow the dose of immunosuppressive drugs to be reduced. In the current protocol, we propose to complement immunosuppressive pharmacotherapy with third-party multipotent adult progenitor cells (MAPCs), a culture-selected population of adult adherent stem cells derived from bone marrow that has been shown to display potent immunomodulatory and regenerative properties. In animal models, MAPCs reduce the need for pharmacological immunosuppression after experimental solid organ transplantation and regenerate damaged organs. Methods Patients enrolled in this phase I, single-arm, single-center safety and feasibility study (n = 3-24) will receive 2 doses of third-party MAPCs after liver transplantation, on days 1 and 3, in addition to a calcineurin-inhibitor-free "bottom-up" immunosuppressive regimen with basiliximab, mycophenolic acid, and steroids. The study objective is to evaluate the safety and clinical feasibility of MAPC administration in this patient cohort. The primary endpoint of the study is safety, assessed by standardized dose-limiting toxicity events. One secondary endpoint is the time until first biopsy-proven acute rejection, in order to collect first evidence of efficacy. Dose escalation (150, 300, 450, and 600 million MAPCs) will be done according to a 3 + 3 classical escalation design (4 groups of 3-6 patients each). Discussion If MAPCs are safe for patients undergoing liver transplantation in this study, a phase II/III trial will be conducted to assess their clinical efficacy.

Immunological diseases are typically heterogeneous in clinical presentation, severity and response to therapy. Biomarkers of immune diseases often reflect this variability, especially compared to their regulated behaviour in health. This leads to a common difficulty that frustrates biomarker discovery and interpretation - namely, unequal dispersion of immune disease biomarker expression between patient classes necessarily limits a biomarker's informative range. To solve this problem, we introduce dataset restriction, a procedure that splits datasets into classifiable and unclassifiable samples. Applied to synthetic flow cytometry data, restriction identifies biomarkers that are otherwise disregarded. In advanced melanoma, restriction finds biomarkers of immune-related adverse event risk after immunotherapy and enables us to build multivariate models that accurately predict immunotherapy-related hepatitis. Hence, dataset restriction augments discovery of immune disease biomarkers, increases predictive certainty for classifiable samples and improves multivariate models incorporating biomarkers with a limited informative range. This principle can be directly extended to any classification task., (© 2024. The Author(s).)

Background: The increasing prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) incurs substantial morbidity, mortality and healthcare costs. Detection and clinical intervention at early stages of disease improves prognosis; however, we are currently limited by a lack of reliable diagnostic tests for population screening and monitoring responses to therapy. To address this unmet need, we investigated human invariant Natural Killer T cell (iNKT) activation by fat-loaded hepatocytes, leading to the discovery that circulating soluble CD46 (sCD46) levels accurately predict hepatic steatosis., Methods: sCD46 in plasma was measured using a newly developed immuno-competition assay in two independent cohorts: Prospective living liver donors (n = 156; male = 66, female = 90) and patients with liver tumours (n = 91; male = 58, female = 33). sCD46 levels were statistically evaluated as a predictor of hepatic steatosis., Findings: Interleukin-4-secreting (IL-4 + ) iNKT cells were over-represented amongst intrahepatic lymphocytes isolated from resected human liver samples. IL-4 + iNKT cells preferentially developed in cocultures with a fat-loaded, hepatocyte-like cell line, HepaRG. This was attributed to induction of matrix metalloproteases (MMP) in fat-loaded HepaRG cells and primary human liver organoids, which led to indiscriminate cleavage of immune receptors. Loss of cell-surface CD46 resulted in unrepressed differentiation of IL-4 + iNKT cells. sCD46 levels were elevated in patients with hepatic steatosis. Discriminatory cut-off values for plasma sCD46 were found that accurately classified patients according to histological steatosis grade., Interpretation: sCD46 is a reliable clinical marker of hepatic steatosis, which can be conveniently and non-invasively measured in serum and plasma samples, raising the possibility of using sCD46 levels as a diagnostic method for detecting or grading hepatic steatosis., Funding: F.B. was supported by the Else Kröner Foundation (Award 2016_kolleg.14). G.G. was supported by the Bristol Myers Squibb Foundation for Immuno-Oncology (Award FA-19-009). N.S. was supported by a Wellcome Trust Fellowship (211113/A/18/Z). J.A.H. received funding from the European Union's Horizon 2020 research and innovation programme (Award 860003). J.M.W. received funding from the Else Kröner Foundation (Award 2015_A10)., Competing Interests: Declaration of interests University Hospital Regensburg has filed a not yet published European patent application (Registration Nr. 23 183 382.3) for sCD46 as a clinical biomarker of hepatic steatosis. J.A.H. received in-kind support from Beckman Coulter. The authors have no other conflicts of interest to declare., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Cytoreductive surgery (CRS) combined with hyperthermic intraperitoneal chemotherapy (HIPEC) is a treatment option for peritoneal carcinomatosis (PC) from colorectal cancer (CRC), which is otherwise a terminal stage of disease. Nevertheless, survival outcomes are only marginally superior to other treatments. This fact highlights the need for better strategies to control intra-abdominal disease recurrence after CRS-HIPEC, including the complementary use of immunotherapies. The aim of this study was therefore to investigate the immune phenotype of T cells in patients with PC. Fifty three patients with CRC (34 patients with PC and 19 patients without PC) were enrolled in a prospective study (clinicaltrials.gov: NCT04108936). Peripheral blood and omental fat were collected to isolate peripheral blood mononuclear cells (PBMCs) and adipose tissue mononuclear cells (ATMCs). These cells were analysed by flow cytometry using a panel focused upon T cell memory differentiation and exhaustion markers. We found a more naïve profile for CD8 + T cells in peripheral blood and intra-abdominal fat of PC patients compared to comparator group (CG) patients. Furthermore, there was an over-representation of CD4 + T cells expressing inhibitory receptors in adipose tissue of PC patients, but not in blood. Our description of intraperitoneal T cell subsets gives us a better understanding of how peritoneal carcinomatosis shapes local immune responses., (© 2023. The Author(s).)

Background: The approval of Atezolizumab / Bevacizumab therapy (Atezo/Bev) in 2020 opened up a promising new treatment option for patients with end-stage hepatocellular carcinoma (HCC). However, liver transplant (LTx) patients with HCC are still denied this therapy owing to concerns about ICI-induced organ rejection and lack of regulatory approval., Methods: A prospective observational study at a tertiary liver transplant centre monitored the compassionate, off-label use of Atezo/Bev in a single, stable LTx recipient with non-resectable HCC recurrence. Close clinical, laboratory and immunological monitoring of the patient was performed throughout a four-cycle Atezo/Bev treatment. Measured parameters were selected after a systematic review of the literature on predictive markers for clinical response and risk of graft rejection caused by ICI therapy., Results: 19 articles describing 20 unique predictive biomarkers were identified. The most promising negative prognostic factors were the baseline values and dynamic course of IL-6, alpha-fetoprotein (AFP) and the AFP/CRP ratio. The frequency of regulatory T cells (Treg) reportedly correlates with the success of ICI therapy. PD-L1 and CD28 expression level with the allograft, peripheral blood CD4 + T cell numbers and Torque Teno Virus (TTV) titre may predict risk of LTx rejection following ICI therapy. No relevant side effects or acute rejection occurred during Atezo/Bev therapy; however, treatment did not prevent tumor progression. Absence of PD-L1 expression in pre-treatment liver biopsies, as well as a progressive downregulation of CD28 expression by CD4 + T cells during therapy, correctly predicted absence of rejection. Furthermore, increased IL-6 and AFP levels after starting therapy, as well as a reduction in blood Treg frequency, correctly anticipated a lack of therapeutic response., Conclusion: Atezo/Bev therapy for unresectable HCC in stable LTx patients remains a controversial strategy because it carries a high-risk of rejection and therapeutic response rates are poorly defined. Although previously described biomarkers of rejection risk and therapeutic response agreed with clinical outcomes in the described case, these immunological parameters are difficult to reliably interpret. Clearly, there is an important unmet need for standardized assays and clinically validated cut-offs before we use these biomarkers to guide treatment decisions for our patients., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2023 Yang Zhou, Eder, Weber, Heumann, Kronenberg, Werner, Geissler, Schlitt, Hutchinson and Bitterer.)

A cost-effective capillary dialysis apparatus (Toledo Capillary Box, TCB) developed for biomacromolecule crystal growth in microgravity and unit gravity environments can provide slow equilibration between the precipitant reservoir and capillary solutions, nurturing growth of neutron-diffraction-quality crystals. Under microgravity conditions, mass transfer of precipitants and biomacro-mol-ecules occurs under diffusion-controlled conditions, promoting slow growth and suppressing defect formation. The equilibration of common precipitants (polyethyl-ene glycol and salts such as ammonium sulfate) between capillary and reservoir solutions was measured for capillaries oriented horizontally or vertically with respect to the gravitational field at unit gravity. Precipitants equilibrated less rapidly in the vertical orientation when capillary solution densities were lower than those of the reservoir solutions. A plug filled with agarose gel was introduced in the TCB apparatus for salt precipitants since salts often exhibit relatively high free diffusion. Equilibration of the capillaries with reservoir solutions was significantly delayed for many of the salt precipitants tested. Analytical and semi-analytical models allow the prediction of precipitant equilibration of capillary and reservoir solutions under diffusion-controlled transport and show good agreement with experimental results., (© Umberto A. Kober et al. 2023.)

Checkpoint blockade is particularly based on PD-1/PD-L1-inhibiting antibodies. However, an efficient immunological tumor defense can be blocked not only by PD-(L)1 but also by the presence of additional immune checkpoint molecules. Here, we investigated the co-expression of several immune checkpoint proteins and the soluble forms thereof (e.g., PD-1, TIM-3, LAG-3, PD-L1, PD-L2 and others) in humanized tumor mice (HTM) simultaneously harboring cell line-derived (JIMT-1, MDA-MB-231, MCF-7) or patient-derived breast cancer and a functional human immune system. We identified tumor-infiltrating T cells with a triple-positive PD-1, LAG-3 and TIM-3 phenotype. While PD-1 expression was increased in both the CD4 and CD8 T cells, TIM-3 was found to be upregulated particularly in the cytotoxic T cells in the MDA-MB-231-based HTM model. High levels of soluble TIM-3 and galectin-9 (a TIM-3 ligand) were detected in the serum. Surprisingly, soluble PD-L2, but only low levels of sPD-L1, were found in mice harboring PD-L1-positive tumors. Analysis of a dataset containing 3039 primary breast cancer samples on the R2 Genomics Analysis Platform revealed increased TIM-3, galectin-9 and LAG-3 expression, not only in triple-negative breast cancer but also in the HER2 + and hormone receptor-positive breast cancer subtypes. These data indicate that LAG-3 and TIM-3 represent additional key molecules within the breast cancer anti-immunity landscape.

Competing Interests: The authors declare no conflicts of interest.

Brain metastasis still encompass very grim prognosis and therefore understanding the underlying mechanisms is an urgent need toward developing better therapeutic strategies. We uncover the intricate interactions between recruited innate immune cells and resident astrocytes in the brain metastatic niche that facilitate metastasis of melanoma and breast cancer. We show that granulocyte-derived lipocalin-2 (LCN2) induces inflammatory activation of astrocytes, leading to myeloid cell recruitment to the brain. LCN2 is central to inducing neuroinflammation as its genetic targeting or bone-marrow transplantation from LCN2 -/- mice was sufficient to attenuate neuroinflammation and inhibit brain metastasis. Moreover, high LCN2 levels in patient blood and brain metastases in multiple cancer types were strongly associated with disease progression and poor survival. Our findings uncover a previously unknown mechanism, establishing a central role for the reciprocal interactions between granulocytes and astrocytes in promoting brain metastasis and implicate LCN2 as a prognostic marker and potential therapeutic target., (© 2023. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: The single-arm phase II APT trial established trastuzumab and paclitaxel (TH) as the standard adjuvant regimen for small human epidermal growth factor receptor 2 (HER2+) tumors. However, paclitaxel causes alopecia and has high rates of neuropathy and hypersensitivity reactions. In patients with metastatic HER2+ breast cancer (BC), the combination of trastuzumab and vinorelbine (TV) is effective and well tolerated. There is a need for alternative non-anthracycline/taxane-based regimens for patients with HER2+ early-stage BC, especially for those with contraindications or who wish to avoid side effects of taxane-based regimens. Here we describe our institutional experience with adjuvant TV for patients with early-stage HER2+ BC., Methods: Clinicopathological characteristics, treatment details, and outcomes of patients with localized HER2+ BC treated with adjuvant TV from 2007 to 2021 at a large academic medical institution were collected. Study endpoints included invasive disease-free survival (IDFS), overall survival (OS), and safety/tolerability. IDFS and OS were measured from start date of TV treatment to date of event/last follow-up and date of death/last follow-up, respectively., Results: A total of 30 patients were treated with TV. All patients received trastuzumab at standard dosing and vinorelbine at a starting dose of 25 mg/m 2 either on days 1/8 or on days 1/8/21 (weekly) of a 21-day cycle with four planned cycles. Median age at diagnosis was 59 years (range: 36-81). 90.3% of patients had anatomic pathologic stage IA BC and 9.7% stage IIA BC. Of the 30 patients, 24 of them opted to pursue TV due to concerns related to alopecia, neuropathy, and other toxicities, and 6 switched from treatment with TH to TV due to toxicities. Eight patients experienced neutropenia with no cases of febrile neutropenia. No patients experienced alopecia or long-term neuropathy. With a median follow-up of 68 months (5.7 years), the 5-year IDFS rate was 90.9%, with one local and one distant recurrence. The 5-year OS was 100%., Conclusions: Trastuzumab in combination with vinorelbine in the adjuvant, early-stage setting for low-risk HER2+ BC demonstrated clinical efficacy and appeared to be well tolerated. TV warrants further evaluation as an alternative regimen to TH for patients with early-stage HER2+ BC., Competing Interests: Shannon MacLaughlin: No COI Erika Nakajima: No COI Yael Bar: No COI Jennifer Hutchinson: Advisory board participant for Novartis Jennifer Shin: No COI Beverly Moy: No COI Steven J. Isakoff: SJI declares the following relationships: institutional research funding from Genentech, PharmaMar, Abbvie, OncoPep, Merck, and AstraZeneca/MedImmune Aditya Bardia: AB declares the following relationships: Consultant/advisory board: Genentech/Roche, Immunomedics, Novartis, Pfizer, Merck, Radius Health, Spectrum Pharma, Taiho Pharm, Diiachi, Sanofi, Puma Biotechnology; Research Grant (self): Biothernostics; Research Grant (Institution): Genentech/Roche, Immunomedics, Novartis, Pfizer, Merck, Radius Health, Sanofi, Mersana AB is supported by Department of Defense Breast Cancer grant and National Comprehensive Cancer Network grant. Irene Kuter: No COI Laura M. Spring: LMS has served as a compensated consultant or received honoraria from Novartis, Puma Biotechnology, G1 Therapeutics, Daiichi Sankyo, AstraZeneca; institutional research support from Merck, Gilead, Lilly, and Phillips Dr. Spring is supported by the National Cancer Institute (grant number K12CA087723) and a National Comprehensive Cancer Network grant., (© The Author(s), 2023.)

Immune checkpoint inhibitors have revolutionized treatment of advanced melanoma, but commonly cause serious immune-mediated complications. The clinical ambition of reserving more aggressive therapies for patients least likely to experience immune-related adverse events (irAE) has driven an extensive search for predictive biomarkers. Here, we externally validate the performance of 59 previously reported markers of irAE risk in a new cohort of 110 patients receiving Nivolumab (anti-PD1) and Ipilimumab (anti-CTLA-4) therapy. Alone or combined, the discriminatory value of these routine clinical parameters and flow cytometry biomarkers was poor. Unsupervised clustering of flow cytometry data returned four T cell subsets with higher discriminatory capacity for colitis than previously reported populations, but they cannot be considered as reliable classifiers. Although mechanisms predisposing some patients to particular irAEs have been described, we are presently unable to capture adequate information from pre-therapy flow cytometry and clinical data to reliably predict risk of irAE in most cases., Competing Interests: MK is a Beckman Coulter Life Sciences associate. SH has received consulting fees and speaker’s honoraria from BMS and Merck Sharp & Dohme (MSD). The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Glehr, Riquelme, Yang Zhou, Cordero, Schilling, Kapinsky, Schlitt, Geissler, Burkhardt, Schmidt, Haferkamp, Hutchinson and Kronenberg.)

Purpose: To explore the immunogenicity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines in patients with breast cancer based on type of anticancer treatment., Methods: Patients with breast cancer had anti-spike antibody concentrations measured ⩾14 days after receiving a full SARS-CoV-2 vaccination series. The primary endpoint was IgA/G/M anti-spike antibody concentration. Multiple regression analysis was used to analyze log 10 -transformed antibody titer concentrations., Results: Between 29 April and 20 July 2021, 233 patients with breast cancer were enrolled, of whom 212 were eligible for the current analysis. Patients who received mRNA-1273 (Moderna) had the highest antibody concentrations [geometric mean concentration (GMC) in log 10 : 3.0 U/mL], compared to patients who received BNT162b2 (Pfizer) (GMC: 2.6 U/mL) (multiple regression adjusted p  = 0.013) and Ad26.COV2.S (Johnson & Johnson/Janssen) (GMC: 2.6 U/mL) ( p  = 0.071). Patients receiving cytotoxic therapy had a significantly lower antibody titer GMC (2.5 U/mL) compared to patients on no therapy or endocrine therapy alone (3.0 U/mL) ( p  = 0.005). Patients on targeted therapies (GMC: 2.7 U/mL) also had a numerically lower GMC compared to patients not receiving therapy/on endocrine therapy alone, although this result was not significant ( p  = 0.364). Among patients who received an additional dose of vaccine ( n  = 31), 28 demonstrated an increased antibody response that ranged from 0.2 to >4.4 U/ mL., Conclusion: Most patients with breast cancer generate detectable anti-spike antibodies following SARS-CoV-2 vaccination, though systemic treatments and vaccine type impact level of response. Further studies are needed to better understand the clinical implications of different antibody levels, the effectiveness of additional SARS-CoV-2 vaccine doses, and the risk of breakthrough infections among patients with breast cancer., Competing Interests: Competing interests: Elyssa Denault: No COI Erika Nakajima: No COI Vivek Naranbhai: No COI Jennifer Hutchinson: Advisory board participant for Novartis Lindsey Mortensen: No COI Elizabeth Neihoff: No COI Caroline Barabell: No COI Amy Comander: No COI Dejan Juric: Consulting: Novartis, Genentech, Inc., EMD Serono, Eisai, Ipsen, Syros, Vibliome Therapeutics, Relay Therapeutics, MapKure, Petra Pharma, Silverback Therapeutics, PIC Therapeutics; Research Funding (To the institution): Novartis, Genentech, Inc., Eisai, EMD Serono, Pfizer, Syros, Takeda, Amgen, InventisBio, Dizal Pharma, Celgene, Infinity Pharmaceuticals. Irene Kuter: No COI Theresa Mulvey: No COI Jeffrey Peppercorn: Employment (spouse): GlaxoSmithKline, Consulting (self) Abbott Labs Aron S Rosenstock: No COI Jennifer Shin: No COI Neelima Vidula: Research funding to the institution (MGH): Daehwa, Pfizer, Merck, Novartis, and Radius, Advisory board participation: AbbVie, OncoSec Seth A Wander: Consulting/Advisory board: Foundation Medicine, Veracyte, Eli Lilly, Hologic, Biovica; institutional research funding from Genentech. Beverly Moy: No COI Leif W. Ellisen: No COI Steven J. Isakoff: Institutional research funding from Genentech, PharmaMar, Abbvie, OncoPep, Merck, and AstraZeneca/MedImmune A. John Iafrate: Invitae (royalties); Consulting (Paige.ai, Kinnate, Oncoclinicas Brasil, Repare); Funding from Peter and Ann Lambertus Family Foundation Justin F. Gainor: Served as a compensated consultant or received honoraria from Bristol-Myers Squibb, Takeda, Loxo/Lilly, Blueprint, Oncorus, Regeneron, Gilead, Moderna, AstraZeneca, EMD Serono, Pfizer, Novartis, iTeos, Karyopharm, Silverback Therapeutics, Merck, and GlydeBio; research support from Novartis; institutional research support from Bristol-Myers Squibb, Tesaro, Moderna, Blueprint, Jounce, Array Biopharma, Merck, Adaptimmune, Novartis, and Alexo; and has an immediate family member who is an employee with equity at Ironwood Pharmaceuticals. Aditya Bardia: Consultant/advisory board: Genentech/Roche, Immunomedics, Novartis, Pfizer, Merck, Radius Health, Spectrum Pharma, Taiho Pharm, Diiachi, Sanofi, Puma Biotechnology; Research Grant (self): Biothernostics; Research Grant (Institution): Genentech/Roche, Immunomedics, Novartis, Pfizer, Merck, Radius Health, Sanofi, Mersana. Dr. Bardia is supported by Department of Defense Breast Cancer grant and National Comprehensive Cancer Network grant. Laura M. Spring: Compensated consultant or received honoraria from Novartis, Puma Biotechnology; institutional research support from Merck, Gilead, Lilly Dr. Spring is supported by the National Cancer Institute [grant number K12CA087723] and a National Comprehensive Cancer Network grant., (© The Author(s), 2022.)

Background: Steatotic livers are more prone to rejection, but are often transplanted owing to the shortage of available organs. Type II NKT (T2NKT) cells are liver-resident lymphocytes that react to lipids presented by CD1d. The role of T2NKT cells in rejection of fatty liver transplants is unclear, partly because of a lack of T2NKT cell markers and their very low frequency in blood. Here, we quantify human T2NKT cells in blood and liver tissue by flow cytometry and provide a strategy for their enrichment and expansion., Methods: Human T2NKT cells were identified as CD3 + CD56 + CD161 + TCR-γᵹ - TCRVα7.2 - and TCRVα24 - cells. T2NKT cells were enriched from blood by sequential positive selection using CD56 and CD3 microbeads. These were subsequently FACS-sorted to purity then expanded in vitro for 3 weeks using anti-CD3/CD28 beads and TGF-β1., Results: The frequency of human T2NKT cells in blood was very low (0.8 ± 0.4% of CD3 + T cells) but they were a more abundant population in liver (6.3 ± 0.9%). Enriched T2NKT cells expressed the transcription factor PLZF. A novel subset of FoxP3 + T2NKT cells was discovered in blood and liver tissue. T2NKT cells were expanded in culture by 15- to 28-fold over 3 weeks, during which time they maintained expression of all identifying markers, including PLZF and FoxP3., Conclusions: Our work defines new strategies for identifying and isolating T2NKT cells from human blood and liver tissue. We showed that this rare population can be expanded in vitro in order to obtain experimentally amenable cell numbers. Further, we identified a novel T2NKT cell subset that stably expresses FoxP3, which might play a role in regulating innate-like lymphocyte responses in steatotic liver transplants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yang Zhou, Werner, Glehr, Geissler, Hutchinson and Kronenberg.)

The introduction of clinical antibodies against programmed death-1 (PD-1) and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) has revolutionized cancer treatment. Immune checkpoint blockade has enormous therapeutic potential and is widely prescribed for treating various cancers. However, immune-related adverse events in checkpoint blockade-treated patients are common and limit its clinical application. Despite efforts to understand the etiology of immune-related adverse events, the underlying cellular reactions remain elusive. Recently, our group identified a subset of patients with metastatic melanoma that are predisposed to hepatitis after combined PD-1 and CTLA-4 blockade. These patients are characterized by pre-treatment expansion of effector memory CD4 + T cells (T EM cells) in blood. We attributed this expansion to chronic or recurrent subclinical immune responses against cytomegalovirus (CMV) infection. Accordingly, baseline expansion of T EM cells is a reliable biomarker of hepatitis risk that identifies a subgroup of patients who might benefit from prophylactic CMV treatment with valganciclovir., (© 2022 The Authors. Journal der Deutschen Dermatologischen Gesellschaft published by John Wiley & Sons Ltd on behalf of Deutsche Dermatologische Gesellschaft.)

"Humanized" mice have been widely used for the characterization of human cancer progression and as a powerful preclinical model. Standardization of multicolor phenotyping could help to identify immune cell patterns involved in checkpoint-related complications. Therefore, we applied established protocols for immune cell profiling to our humanized Patient-Derived Xenograft (hPDX) model. hPDX are characterized by the co-existence of a human immune system and a patient-derived tumor transplant. These mice possess a human-like immune system after CD34 + stem cell transplantation while the reconstitution level of the immune system was not related to the quantity of transplanted CD34 + cells. Contamination ≤ 1.2% by CD3 + cells in the hematopoietic stem cell (HSC) transplant did not trigger abnormal T cell maturation. Different B and T cell differentiation stages were identified, as well as regulatory T cells (Tregs) and exhausted T cells that expressed TIGIT, PD-1, or KLRG1. Overall, the application of standardized protocols for the characterization of immune cells using flow cytometry will contribute to a better understanding of immune-oncologic processes.

Treatment of advanced melanoma with combined immune checkpoint inhibitor (ICI) therapy is complicated in up to 50% of cases by immune-related adverse events (irAE) that commonly include hepatitis, colitis and skin reactions. We previously reported that pre-therapy expansion of cytomegalovirus (CMV)-reactive CD4 + effector memory T cells (T EM ) predicts ICI-related hepatitis in a subset of patients with Stage IV melanoma given αPD-1 and αCTLA-4. Here, we develop and validate a 10-color flow cytometry panel for reliably quantifying CD4 + T EM cells and other biomarkers of irAE risk in peripheral blood samples. Compared to previous methods, our new panel performs equally well in measuring CD4 + T EM cells (agreement = 98%) and is superior in resolving CD4 + CD197 + CD45RA - central memory T cells (T CM ) from CD4 + CD197 + CD45RA + naive T cells (T naive ). It also enables us to precisely quantify CD14 + monocytes (CV = 6.6%). Our new "monocyte and T cell" (MoT) assay predicts immune-related hepatitis with a positive predictive value (PPV) of 83% and negative predictive value (NPV) of 80%. Our essential improvements open the possibility of sharing our predictive methods with other clinical centers. Furthermore, condensing measurements of monocyte and memory T cell subsets into a single assay simplifies our workflows and facilitates computational analyses., Competing Interests: MK is a Beckman Coulter Life Sciences associate. SH has received consulting fees and speaker’s honoraria from BMS and Merck Sharp & Dohme (MSD). NA is employed by MVZ for Laboratory Medicine Raubling. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Schilling, Glehr, Kapinsky, Ahrens, Riquelme, Cordero, Bitterer, Schlitt, Geissler, Haferkamp, Hutchinson and Kronenberg.)

Background: Plateletpheresis using a leukocyte reduction system (LRS) traps donor WBCs in the LRS chamber, which may lead to lymphopenia, especially in frequent plateletpheresis donors. It seems plausible that this might cause adverse effects. However, current knowledge about potential confounders and donor health impacts is incomplete., Donors and Methods: Recent platelet donors and donations collected at University Hospital Regensburg from 2016 to 2019 using the Terumo BCT Trima Accel LRS system were retrospectively analyzed and compared with historical platelet donors and donations collected mainly with Fresenius Kabi Amicus non-LRS system from 2010 to 2013. Additionally, recent donors were prospectively surveyed using a health-related topics questionnaire., Results: Analysis of 819 recent donors with 11,254 blood counts and 1464 questionnaires and 1011 historical donors with 12,848 blood counts revealed that increased annual platelet donation frequencies were associated with decreased lymphocyte counts in both groups. Median lymphocyte counts in recent donors with no versus ≥24 previous annual donations declined from 2.0 to 1.2 × 10 3 /μL (p < 2.2 × 10 -16 ), and those in historical donors with no versus ≥24 previous annual donations decreased from 2.0 to 1.5 × 10 3 /μL (p = 6 × 10 -4 ), respectively. The questionnaire results showed that donation frequency and lymphopenia were not associated with upper respiratory tract infection (URTI) incidence or duration, but platelet donors who concomitantly donated granulocytes had significantly shorter URTI durations than those who did not (p = .008)., Conclusion: This study confirmed that plateletpheresis-associated lymphopenia occurs in LRS and to a lesser degree in non-LRS platelet donors, but revealed no evidence of a negative impact on donor health., (© 2021 The Authors. Transfusion published by Wiley Periodicals LLC on behalf of AABB.)

Objectives: This validation study investigated a flow cytometric apoptosis assay according to good manufacturing practice (GMP)., Background: Extracorporeal photopheresis (ECP) is a treatment for various immunological diseases and cutaneous T-cell lymphomas. It is based on the induction of apoptosis by 8-methoxypsoralene and ultraviolet A light. The quantification of apoptosis is therefore essential for ECP improvements. However, despite numerous publications on apoptosis, validated technical details are lacking., Methods and Materials: Mononuclear cells were collected by apheresis and treated by ECP or camptothecin. Samples taken before and after ECP were cultured for 24, 48 and 72 h and analysed for apoptosis and viability of T cells and monocytes by flow cytometry with Annexin V and 7-AAD staining. Accuracy of the assay, intra- and inter-assay precision and the pre-analytical and analytical stability of the analytes were the investigated parameters., Results: Our data indicate that the median intra- and inter-assay precision coefficient of variation for T cells was 3.86% and 4.80%, respectively. Pre-analytical stability of T cells and monocytes was ensured during short-term storage for up to 2 h on ice. After staining, analytical stability was limited to 30 min, likely because of ongoing apoptosis and loss of monocytes due to plastic adhesion., Conclusion: The results of this validation study show that the assay is GMP-compliant and that its reliability, accuracy and precision are acceptable. While pre-analytical stability of the cells was compatible with on-site procedures, our analytical stability data indicate that this assay is not suited for batch mode analysis of ECP products., (© 2021 The Authors. Transfusion Medicine published by John Wiley & Sons Ltd on behalf of British Blood Transfusion Society.)

Treatment of advanced melanoma with combined PD-1/CTLA-4 blockade commonly causes serious immune-mediated complications. Here, we identify a subset of patients predisposed to immune checkpoint blockade-related hepatitis who are distinguished by chronic expansion of effector memory CD4 + T cells (T EM cells). Pre-therapy CD4 + T EM cell expansion occurs primarily during autumn or winter in patients with metastatic disease and high cytomegalovirus (CMV)-specific serum antibody titres. These clinical features implicate metastasis-dependent, compartmentalised CMV reactivation as the cause of CD4 + T EM expansion. Pre-therapy CD4 + T EM expansion predicts hepatitis in CMV-seropositive patients, opening possibilities for avoidance or prevention. 3 of 4 patients with pre-treatment CD4 + T EM expansion who received αPD-1 monotherapy instead of αPD-1/αCTLA-4 therapy remained hepatitis-free. 4 of 4 patients with baseline CD4 + T EM expansion given prophylactic valganciclovir and αPD-1/αCTLA-4 therapy remained hepatitis-free. Our findings exemplify how pathogen exposure can shape clinical reactions after cancer therapy and how this insight leads to therapeutic innovations.

Background: Incisional surgical site infections (iSSI) in hepatopancreatobiliary (HPB) surgery usually lead to prolonged hospital stays, consume valuable resources, and impact on patients' outcome. Prophylactic closed incision negative pressure wound therapy (ciNPWT) to decrease wound complications has become available. Owing to an increasing number of studies, evidence for superiority in many indication areas has accumulated; however, in general surgery, there are a few data and those have shown contradictory results., Methods: In this monocentric, prospective, randomized, controlled, two-armed study, the influence of ciNPWT on incisional surgical site infection rates after HPB operations will be investigated. A total of 222 patients will be randomized 1:1 to an interventional group (7-day treatment with ciNPWT) or a control group (treated with gauze dressing). The primary parameter to evaluate efficacy is the rate of incisional SSIs within 30 days after surgery. Additionally, several clinically relevant secondary outcomes will be assessed., Discussion: A reduction in the rate of incisional SSIs would not only lead to a significant cost reduction and shorter postoperative length of stay, but may also improve postoperative quality of life for patients. While earlier publications have shown advantages for ciNPWT, recent studies did not confirm a positive effect regarding iSSI rate. Even if iSSI rate is not reduced, findings obtained from the secondary endpoints may be of clinical relevance, such as reduction of wound complication rates., Trial Registration: This trial has been registered in the German Clinical Trials Register, DRKS 00015136 . Registered on 19 February 2019 and has been approved by the local ethics committee of the University of Regensburg: 18-1225-101.

Background: Although landmark trials in the metastatic (CLEOPATRA) and neo-adjuvant (NeoSphere; TRYPHAENA) settings identified all-grade diarrhea as a pertuzumab-associated adverse event, it was not classified as dose-limiting. In actual practice, diarrhea is often a reason for treatment modifications., Objectives: To compare the risk of pertuzumab-associated diarrhea in actual practice to the risks in randomized controlled trials., Methods: We conducted a retrospective cohort study of HER2/neu-positive breast cancer patients who received a pertuzumab-containing regimen between January 2012 and August 2015. We calculated the risk of diarrhea with 95% confidence limits (CLs), and then used two-sample t-tests to compare the risk between trials and actual practice., Results: A total of 27 patients in the study cohort received a pertuzumab-containing treatment regimen for HER2/neu-positive breast cancer. The overall risk of all-grade and severe diarrhea in actual practice was 70% (95% CLs 55-90%) and 37% (95% CLs 20-66%), respectively. No severe diarrhea was observed in the metastatic setting, and the risk of all-grade diarrhea (44%, 95% CLs 21-92%) was similar to the CLEOPATRA study (67%). The risk of all-grade diarrhea in the neo-adjuvant setting was 83% (95% CLs 68-100%), compared to 46% in the NeoSphere trial (p = 0.03). The risk of severe diarrhea (Grade 3-4) in the neo-adjuvant setting was 47% (95% CLs 27-80%) versus 6% in the NeoSphere (p < 0.0001) and 12% in the TRYPHAENA (p < 0.01) trials., Conclusions: The risk of all-grade and severe diarrhea associated with neoadjuvant pertuzumab use for HER2/neu-positive breast cancer was greater in actual practice than in trials.

Background: Extracorporeal photopheresis (ECP) is an immunosuppressive treatment that involves leukocyte apheresis, psoralen and UV light treatment, and subsequent reinfusion. Patients treated with ECP are usually immunosuppressed. Bacterial contamination therefore poses a much unwanted risk, but incidence data are lacking., Patients and Methods: We screened all 1922 consecutive ECP procedures scheduled within a roughly 3-year period for eligibility. Those with missing data on ECP method (inline or offline) or type of venous access (peripheral or central) were excluded. ECPs with complete aerobic and anaerobic microbial testing of baseline patient blood samples (n = 1637) and of ECP cell concentrates (n = 1814) were included in the analysis., Results: A test for microbial contamination was positive for 1.82% of the cell concentrates, with central venous access was the most significant risk factor for the contamination (odds ratio = 19). Patient blood samples were positive in 3.85% of cases, but no patients became septic. Staphylococcus spp. were most abundant, and products with bacterial contamination did not cause side effects after reinfusion. There were no significant differences in contamination rates between inline and offline ECP., Conclusion: These findings stress the importance of sterile procedures and the benefits of using peripheral over central venous access for reducing the risk of bacterial contamination in ECP., (© 2020 The Authors. Transfusion published by Wiley Periodicals, Inc. on behalf of AABB.)

Background: Use of cell-based medicinal products (CBMPs) represents a state-of-the-art approach for reducing general immunosuppression in organ transplantation. We tested multiple regulatory CBMPs in kidney transplant trials to establish the safety of regulatory CBMPs when combined with reduced immunosuppressive treatment., Methods: The ONE Study consisted of seven investigator-led, single-arm trials done internationally at eight hospitals in France, Germany, Italy, the UK, and the USA (60 week follow-up). Included patients were living-donor kidney transplant recipients aged 18 years and older. The reference group trial (RGT) was a standard-of-care group given basiliximab, tapered steroids, mycophenolate mofetil, and tacrolimus. Six non-randomised phase 1/2A cell therapy group (CTG) trials were pooled and analysed, in which patients received one of six CBMPs containing regulatory T cells, dendritic cells, or macrophages; patient selection and immunosuppression mirrored the RGT, except basiliximab induction was substituted with CBMPs and mycophenolate mofetil tapering was allowed. None of the trials were randomised and none of the individuals involved were masked. The primary endpoint was biopsy-confirmed acute rejection (BCAR) within 60 weeks after transplantation; adverse event coding was centralised. The RTG and CTG trials are registered with ClinicalTrials.gov, NCT01656135, NCT02252055, NCT02085629, NCT02244801, NCT02371434, NCT02129881, and NCT02091232., Findings: The seven trials took place between Dec 11, 2012, and Nov 14, 2018. Of 782 patients assessed for eligibility, 130 (17%) patients were enrolled and 104 were treated and included in the analysis. The 66 patients who were treated in the RGT were 73% male and had a median age of 47 years. The 38 patients who were treated across six CTG trials were 71% male and had a median age of 45 years. Standard-of-care immunosuppression in the recipients in the RGT resulted in a 12% BCAR rate (expected range 3·2-18·0). The overall BCAR rate for the six parallel CTG trials was 16%. 15 (40%) patients given CBMPs were successfully weaned from mycophenolate mofetil and maintained on tacrolimus monotherapy. Combined adverse event data and BCAR episodes from all six CTG trials revealed no safety concerns when compared with the RGT. Fewer episodes of infections were registered in CTG trials versus the RGT., Interpretation: Regulatory cell therapy is achievable and safe in living-donor kidney transplant recipients, and is associated with fewer infectious complications, but similar rejection rates in the first year. Therefore, immune cell therapy is a potentially useful therapeutic approach in recipients of kidney transplant to minimise the burden of general immunosuppression., Funding: The 7th EU Framework Programme., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Myeloid-derived suppressor cells (MDSC) represent a heterogeneous group of myeloid regulatory cells that were originally described in cancer. Several studies in animal models point to MDSC as important players in the induction of allograft tolerance due to their immune modulatory function. Most of the published studies have been performed in animal models, and the data addressing MDSCs in human organ transplantation are scarce. We evaluated the phenotype and function of different MDSCs subsets in 38 kidney transplant recipients (KTRs) at different time points. Our data indicate that monocytic MDSCs (Mo-MDSC) increase in KTR at 6 and 12 months posttransplantation. On the contrary, the percentages of polymorphonuclear MDSC (PMN-MDSC) and early-stage MDSC (e-MDSC) are not significantly increased. We evaluated the immunosuppressive activity of Mo-MDSC in KTR and confirmed their ability to increase regulatory T cells (Treg) in vitro . Interestingly, when we compared the ability of Mo-MDSC to suppress T cell proliferation, we observed that tacrolimus, but not rapamycin-treated KTR, was able to inhibit CD4 + T cell proliferation in vitro . This indicates that, although mTOR inhibitors are widely regarded as supportive of regulatory responses, rapamycin may impair Mo-MDSC function, and suggests that the choice of immunosuppressive therapy may determine the tolerogenic pathway and participating immune cells that promote organ transplant acceptance in KTR., (Copyright © 2020 Iglesias-Escudero, Sansegundo-Arribas, Riquelme, Merino-Fernández, Guiral-Foz, Pérez, Valero, Ruiz, Rodrigo, Lamadrid-Perojo, Hutchinson, Ochando and López-Hoyos.)

Cell therapy is a promising strategy for treating patients suffering from autoimmune or inflammatory diseases or receiving a transplant. Based on our preclinical studies, we have generated human autologous tolerogenic dendritic cells (ATDCs), which are being tested in a first-in-man clinical trial in kidney transplant recipients. Here, we report that ATDCs represent a unique subset of monocyte-derived cells based on phenotypic, transcriptomic, and metabolic analyses. ATDCs are characterized by their suppression of T cell proliferation and their expansion of Tregs through secreted factors. ATDCs produce high levels of lactate that shape T cell responses toward tolerance. Indeed, T cells take up ATDC-secreted lactate, leading to a decrease of their glycolysis. In vivo, ATDCs promote elevated levels of circulating lactate and delay graft-versus-host disease by reducing T cell proliferative capacity. The suppression of T cell immunity through lactate production by ATDCs is a novel mechanism that distinguishes ATDCs from other cell-based immunotherapies., (Copyright © 2019 Elsevier Inc. All rights reserved.)

We investigate the self-assembly of a palmitoylated (C16-chain at the N terminus) peptide fragment in comparison to the unlipidated peptide EELNRYY, a fragment of the gut hormone peptide PYY3-36. The lipopeptide C16-EELNRYY shows remarkable pH-dependent self-assembly above measured critical aggregation concentrations, forming fibrils at pH 7, but micelles at pH 10. The parent peptide does not show self-assembly behaviour. The lipopeptide forms hydrogels at sufficiently high concentration at pH 7, the dynamic mechanical properties of which were measured. We also show that the tyrosine functionality at the C terminus of EELNRYY can be used to enzymatically produce the pigment melanin. The enzyme tyrosinase oxidises tyrosine into 3,4-dihydroxyphenylalanine (DOPA), DOPA-quinone and further products, eventually forming eumelanin. This is a mechanism of photo-protection in the skin, for this reason controlling tyrosinase activity is a major target for skin care applications and EELNRYY has potential to be developed for such uses.

We investigate the impact of lipidation on the self-assembly of two peptide fragments from the gastrointestinal peptide hormone PYY 3-36 . The lipopeptides C 16 IKPEAP and C 16 IKPEAPGE contain the first 6 and 8 amino acid residues, respectively, from the PYY 3-36 peptide sequence, with a palmitoyl C 16 tail attached at the N-terminus. These lipopeptides form spherical micelles in aqueous solution, above a critical micelle concentration (cmc), which is pH-dependent. Modeling of small-angle X-ray scattering data along with molecular dynamics simulations shows the formation of micelles with a hydrophobic interior and a well-hydrated exterior. The lipopeptides have a disordered conformation over the pH and temperature ranges studied. The cmc is found to be independent of temperature, pointing to athermal micellization. In contrast to the presence of hydrated micelles in solution, β-sheet amyloid fibrils form in dried samples. Thus, the nanostructure of lipidated PYY 3-36 fragment peptides can be tuned by control of pH or concentration, for future applications.

Background: Several sensitive methods are available for red blood cell (RBC) antibody screening. Among these, gel and glass card systems have demonstrated comparably good performance in retrospective studies and are widely used in routine patient diagnostics, but their performance in prospective studies has not been sufficiently characterised., Patients and Methods: Gel card (Bio-Rad DiaMed) and glass bead-based (Ortho Clinical Diagnostics) column agglutination technologies were used to screen for antibodies prospectively (group A) and for antibody identification in stored and fresh samples known to contain RBC antibodies retrospectively (group B). Untreated reagent RBCs and either papain-treated (Bio-Rad) or ficin-treated panel C cells (Ortho) were used for antibody identification., Results: RBC-reactive antibodies were detected in 22 of 1000 group A samples, three of which tested positive only by gel card agglutination, and four only by glass bead agglutination (including one false positive each). Group B comprised 202 sera with known antibodies: 33 of these samples contained 36 antibodies detected only by gel card agglutination, whereas 9 samples contained antibodies detectable only by glass bead-based agglutination. Discrepancies mostly involved weak antibodies reactive by enzyme only. Two sera contained antibody mixtures that neither system detected completely. Of note, in antibody differentiation batches one and two, anti-Lua was reactive in 7 of 7 and 1 of 8 samples, respectively., Conclusion: Both column agglutination tests for red cell antibodies had equal sensitivity and specificity with unstored samples. In stored samples, weak and enzyme-only antibodies were more frequently detected with the gel card system., Competing Interests: This work was supported by Bio-Rad DiaMed. This does not alter the adherence to all PLOS ONE policies on sharing data and materials.

Purpose of Review: Now that adoptive transfer of regulatory macrophages (Mregs) is clinically practicable, we ask whether this approach could be used to achieve self-sustaining peripheral regulation and what mechanisms may be involved., Recent Findings: Dehydrogenase/reductase 9 (DHRS9)-expressing Mregs are a specialized subset of monocyte-derived macrophages that are currently being investigated as a tolerogenic cell-based therapy. Human Mregs are defined by their capacity to convert naïve CD4 T cells to IL-10-secreting FoxP3 regulatory T cells (Tregs) through an activation-dependent process involving signals mediated by TGF-β, retinoic acid, indoleamine 2,3-dioxygenase activity, notch and progestagen associated endometrial protein (PAEP). Mreg-induced iTregs (miTregs) are a phenotypically distinct type of in-vitro-derived human iTreg that expresses butyrophilin-like protein 8 (BTNL8) and T cell immunoreceptor with Ig and ITIM domains (TIGIT). miTregs are nonspecifically suppressive of mitogen-stimulated bystander T cell proliferation and inhibit TNFα-induced maturation of monocyte-derived dendritic cells. Preclinical and clinical studies find that intravenous infusion of allogeneic Mregs leads to enrichment of circulating TIGIT Tregs., Summary: These results suggest a feed-forward mechanism by which Mreg treatment could promote solid organ transplant acceptance through rapid induction of direct pathway Tregs.

Human regulatory macrophages (Mreg) have shown early clinical promise as a cell-based adjunct immunosuppressive therapy in solid organ transplantation. It is hypothesised that recipient CD4 + T cell responses are actively regulated through direct allorecognition of donor-derived Mregs. Here we show that human Mregs convert allogeneic CD4 + T cells to IL-10-producing, TIGIT + FoxP3 + -induced regulatory T cells that non-specifically suppress bystander T cells and inhibit dendritic cell maturation. Differentiation of Mreg-induced Tregs relies on multiple non-redundant mechanisms that are not exclusive to interaction of Mregs and T cells, including signals mediated by indoleamine 2,3-dioxygenase, TGF-β, retinoic acid, Notch and progestagen-associated endometrial protein. Preoperative administration of donor-derived Mregs to living-donor kidney transplant recipients results in an acute increase in circulating TIGIT + Tregs. These results suggest a feed-forward mechanism by which Mreg treatment promotes allograft acceptance through rapid induction of direct-pathway Tregs.

Lipidation is a powerful strategy to improve the stability in vivo of peptide drugs. Attachment of a lipid chain to a hydrophilic peptide leads to amphiphilicity and the potential for surfactant-like self-assembly. Here, the self-assembly and conformation of three lipidated derivatives of the gastrointestinal peptide hormone PYY 3-36 is examined using a comprehensive range of spectroscopic, scattering, and electron microscopy methods and compared to those of the parent PYY 3-36 peptide. The peptides are lipidated at Ser(11), Arg(17), or Arg(23) in the peptide; the former is within the β-turn domain (based on the published solution NMR structure), and the latter two are both within the α-helical domain. We show that it is possible to access a remarkable diversity of nanostructures ranging from micelles to nanotapes and fibrillar hydrogels by control of assembly conditions (concentration, pH, and temperature). All of the lipopeptides self-assemble above a critical aggregation concentration (cac), determined through pyrene fluorescence probe measurements, and they all have predominantly α-helical secondary structure at their native pH. The pH and temperature dependence of the α-helical conformation were probed via circular dichroism spectroscopy experiments. Lipidation was found to provide enhanced stability against changes in temperature and pH. The self-assembled structures were investigated using small-angle X-ray scattering (SAXS) and cryogenic transmission electron microscopy (cryo-TEM). Distinct differences in nanostructure were observed for lipidated and unlipidated peptides, also depending on the position of lipidation. Remarkably, micelles containing lipopeptides with α-helical peptide conformation were observed. Gelation was observed at higher concentrations in certain pH intervals for the lipidated peptides, but not for unlipidated PYY 3-36 . Thus, lipidation, in addition to enhancing stability against pH and temperature variation, also provides a route to prepare PYY peptide hydrogels. These findings provide important insights into the control of PYY 3-36 conformation and aggregation by lipidation, relevant to the development of future therapeutics based on this peptide hormone, for example, in treatments for obesity.

Background: The frequency and intensity of wildfires is anticipated to increase as climate change creates longer, warmer, and drier seasons. Particulate matter (PM) from wildfire smoke has been linked to adverse respiratory and possibly cardiovascular outcomes. Children, older adults, and persons with underlying respiratory and cardiovascular conditions are thought to be particularly vulnerable. This study examines the healthcare utilization of Medi-Cal recipients during the fall 2007 San Diego wildfires, which exposed millions of persons to wildfire smoke., Methods and Findings: Respiratory and cardiovascular International Classification of Diseases (ICD)-9 codes were identified from Medi-Cal fee-for-service claims for emergency department presentations, inpatient hospitalizations, and outpatient visits. For a respiratory index and a cardiovascular index of key diagnoses and individual diagnoses, we calculated rate ratios (RRs) for the study population and different age groups for 3 consecutive 5-day exposure periods (P1 [October 22-26], P2 [October 27-31], and P3 [November 1-5]) versus pre-fire comparison periods matched on day of week (5-day periods starting 3, 4, 5, 6, 8, and 9 weeks before each exposed period). We used a bidirectional symmetric case-crossover design to examine emergency department presentations with any respiratory diagnosis and asthma specifically, with exposure based on modeled wildfire-derived fine inhalable particles that are 2.5 micrometers and smaller (PM2.5). We used conditional logistic regression to estimate odds ratios (ORs), adjusting for temperature and relative humidity, to assess same-day and moving averages. We also evaluated the United States Environmental Protection Agency (EPA)'s Air Quality Index (AQI) with this conditional logistic regression method. We identified 21,353 inpatient hospitalizations, 25,922 emergency department presentations, and 297,698 outpatient visits between August 16 and December 15, 2007. During P1, total emergency department presentations were no different than the reference periods (1,071 versus 1,062.2; RR 1.01; 95% confidence interval [CI] 0.95-1.08), those for respiratory diagnoses increased by 34% (288 versus 215.3; RR 1.34; 95% CI 1.18-1.52), and those for asthma increased by 112% (58 versus 27.3; RR 2.12; 95% CI 1.57-2.86). Some visit types continued to be elevated in later time frames, e.g., a 72% increase in outpatient visits for acute bronchitis in P2. Among children aged 0-4, emergency department presentations for respiratory diagnoses increased by 70% in P1, and very young children (0-1) experienced a 243% increase for asthma diagnoses. Associated with a 10 μg/m3 increase in PM2.5 (72-hour moving average), we found 1.08 (95% CI 1.04-1.13) times greater odds of an emergency department presentation for asthma. The AQI level "unhealthy for sensitive groups" was associated with significantly elevated odds of an emergency department presentation for respiratory conditions the day following exposure, compared to the AQI level "good" (OR 1.73; 95% CI 1.18-2.53). Study limitations include the use of patient home address to estimate exposures and demographic differences between Medi-Cal beneficiaries and the general population., Conclusions: Respiratory diagnoses, especially asthma, were elevated during the wildfires in the vulnerable population of Medi-Cal beneficiaries. Wildfire-related healthcare utilization appeared to persist beyond the initial high-exposure period. Increased adverse health events were apparent even at mildly degraded AQI levels. Significant increases in health events, especially for respiratory conditions and among young children, are expected based on projected climate scenarios of wildfire frequency in California and globally., Competing Interests: The authors have declared that no competing interests exist.

Background: Basic science data suggest that acute kidney injury (AKI) induced by ischemia-reperfusion injury (IRI) is an inflammatory process involving the adaptive immune response. Little is known about the T-cell contribution in the very early phase, so we investigated if tubular cellular stress expressed by elevated cell cycle biomarkers is associated with early changes in circulating T-cell subsets, applying a bedside-to-bench approach., Methods: Our observational pilot study included 20 consecutive patients undergoing endovascular aortic repair for aortic aneurysms affecting the renal arteries, thereby requiring brief kidney hypoperfusion and reperfusion. Clinical-grade flow cytometry-based immune monitoring of peripheral immune cell populations was conducted perioperatively and linked to tubular cell stress biomarkers ([TIMP-2]•[IGFBP7]) immediately after surgery. To confirm clinical results and prove T-cell infiltration in the kidney, we simulated tubular cellular injury in an established mouse model of mild renal IRI., Results: A significant correlation between tubular cell injury and a peripheral decline of γδ T cells, but no other T-cell subpopulation, was discovered within the first 24 hours (r = 0.53; p = 0.022). Turning to a mouse model of kidney warm IRI, a similar decrease in circulating γδ T cells was found and concomitantly was associated with a 6.65-fold increase in γδ T cells (p = 0.002) in the kidney tissue without alterations in other T-cell subsets, consistent with our human data. In search of a mechanistic driver of IRI, we found that the damage-associated molecule high-mobility group box 1 protein HMGB1 was significantly elevated in the peripheral blood of clinical study subjects after tubular cell injury (p = 0.019). Correspondingly, HMGB1 RNA content was significantly elevated in the murine kidney., Conclusions: Our investigation supports a hypothesis that γδ T cells are important in the very early phase of human AKI and should be considered when designing clinical trials aimed at preventing kidney damage., Trial Registration: ClinicalTrials.gov, NCT01915446 . Registered on 5 Aug 2013.

π-Conjugated small molecules based on a [1]benzothieno[3,2-b]benzothiophene (BTBT) unit are of great research interest in the development of solution-processable semiconducting materials owing to their excellent charge-transport characteristics. However, the BTBT π-core has yet to be demonstrated in the form of electro-active one-dimensional (1D) nanowires that are self-assembled in aqueous media for potential use in bioelectronics and tissue engineering. Here we report the design, synthesis, and self-assembly of benzothienobenzothiophene (BTBT)-peptide conjugates, the BTBT-peptide (BTBT-C3-COHN-Ahx-VVAGKK-Am) and the C8-BTBT-peptide (C8-BTBT-C3-COHN-Ahx-VVAGKK-Am), as β-sheet forming amphiphilic molecules, which self-assemble into highly uniform nanofibers in water with diameters of 11-13(±1) nm and micron-size lengths. Spectroscopic characterization studies demonstrate the J-type π-π interactions among the BTBT molecules within the hydrophobic core of the self-assembled nanofibers yielding an electrical conductivity as high as 6.0 × 10-6 S cm-1. The BTBT π-core is demonstrated, for the first time, in the formation of self-assembled peptide 1D nanostructures in aqueous media for potential use in tissue engineering, bioelectronics and (opto)electronics. The conductivity achieved here is one of the highest reported to date in a non-doped state.

A salmonellosis outbreak occurred at a California prison in April and May 2016. In a cohort study of 371 inmates, persons who consumed dishes from the prison kitchen made from ground meat had a higher attack rate (15%) than those who did not (4%) (risk ratio 3.4, 95% CI 1.1-10.6). The ground meat product was composed exclusively of beef, mechanically separated chicken (MSC) and textured vegetable protein; eight of eight lots of the product collected from the prison and processing facility were contaminated with Salmonella enterica of eight serotypes and 17 distinct PFGE patterns, including multidrug-resistant S. Infantis. Either the MSC or the beef could have been the source of the particular strains of S. enterica isolated from patients or the product. The microbiological evidence is most consistent with MSC as the source of the high levels of S. enterica in the epidemiologically linked meat product. Our findings contribute to the growing body of evidence about the hazard posed by the use of products containing raw mechanically separated poultry in kitchens in institutions.

Recent introduction of all-oral direct-acting antiviral (DAA) treatment has revolutionized care of patients with chronic hepatitis C virus (HCV) infection. Regrettably, the high cost of DAA treatment is burdensome for healthcare systems and may be prohibitive for some patients who would otherwise benefit. Understanding how patient-related factors influence individual responses to DAA treatment may lead to more efficient prescribing. In this observational study, patients with chronic HCV infection were comprehensively monitored by flow cytometry to identify pretreatment immunological variables that predicted HCV RNA negativity within 4 weeks of commencing DAA treatment. Twenty-three patients [genotype 1a ( n  = 10), 1b ( n  = 9), and 3 ( n  = 4)] were treated with daclatasvir plus sofosbuvir (SOF) ( n  = 15), ledipasvir plus SOF ( n  = 4), or ritonavir-boosted paritaprevir, ombitasvir, and dasabuvir ( n  = 4). DAA treatment most prominently altered the distribution of CD8 + memory T cell subsets. Knowing only pretreatment frequencies of CD3 + and naive CD8 + T cells allowed correct classification of 83% of patients as "fast" (HCV RNA-negative by 4 weeks) or "slow" responders. In a prospective cohort, these parameters correctly classified 90% of patients. Slow responders exhibited higher frequencies of CD3 + T cells, CD8 + T EM cells, and CD5 high CD27 - CD57 + CD8 + chronically activated T cells, which is attributed to bystander hyperactivation of virus-non-specific CD8 + T cells. Taken together, non-specific, systemic CD8 + T cell activation predicted a longer time to viral clearance. This discovery allows pretreatment identification of individuals who may not require a full 12-week course of DAA therapy; in turn, this could lead to individualized prescribing and more efficient resource allocation.

Background: The human regulatory macrophage (Mreg) has emerged as a promising cell type for use as a cell-based adjunct immunosuppressive therapy in solid organ transplant recipients. In this brief report, dehydrogenase/reductase 9 (DHRS9) is identified as a robust marker of human Mregs., Methods: The cognate antigen of a mouse monoclonal antibody raised against human Mregs was identified as DHRS9 by immunoprecipitation and MALDI-MS sequencing. Expression of DHRS9 within a panel of monocyte-derived macrophages was investigated by quantitative PCR, immunoblotting and flow cytometry., Results: DHRS9 expression discriminated human Mregs from a panel of in vitro derived macrophages in other polarisation states. Likewise, DHRS9 expression distinguished Mregs from a variety of human monocyte-derived tolerogenic antigen-presenting cells in current development as cell-based immunotherapies, including Tol-DC, Rapa-DC, DC-10, and PGE2-induced myeloid-derived suppressor cells. A subpopulation of DHRS9-expressing human splenic macrophages was identified by immunohistochemistry. Expression of DHRS9 was acquired gradually during in vitro development of human Mregs from CD14 monocytes and was further enhanced by IFN-γ treatment on day 6 of culture. Stimulating Mregs with 100 ng/mL lipopolysaccharide for 24 hours did not extinguish DHRS9 expression. Dhrs9 was not an informative marker of mouse Mregs., Conclusion: DHRS9 is a specific and stable marker of human Mregs.

This article provides a transparent description of Mreg&#95;UKR cell products, including manufacture and quality-control processes, using the structure and vocabulary of the 'Minimum Information about Tolerogenic Antigen-presenting Cells' reporting guidelines. This information is intended as a resource for those in the field, as well as a stimulus to develop a new wave of immunoregulatory and tissue-reparative monocyte-derived cell therapies., (© 2017 Steunstichting ESOT.)