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Background. Chronic hepatitis C virus (HCV) infection is characterized by activation of natural killer (NK) cells. Here, we asked whether HCV elimination by sofosbuvir-based direct-acting antivirals (DAAs) and the addition of ribavirin (RBV) improve NK cell function in liver transplant (LTx) recipients. Methods. We analyzed NK cell degranulation and interferon (IFN)γ-response along with STAT1 and STAT4 phosphorylation in 29 HCV-infected LTx recipients and 17 HCV-infected patients during DAA treatment. Results. Compared with uninfected LTx recipients, NK cells from HCV-infected LTx recipients were polarized toward cytotoxicity with increased CD107a-degranulation (10.1% versus 14.6%; P = 0.0263) and reduced capacity to produce IFNγ (43.0% versus 26.7%; P = 0.0002). The altered phenotype of NK cells in HCV-infected LTx recipients was accompanied by increased STAT1 (44.6% versus 87.4%; P < 0.0001) and STAT1 phosphorylation (0.7% versus 8.9%; P = 0.0005) compared with pSTAT4 IFNα-induction (29.9% versus 17.6%; P = 0.0014). Successful DAA therapy did not affect CD107a-degranulation but decreased STAT1. RBV cotreatment with DAA therapy for HCV increased CD56Bright NK cell IFNγ-responses in LTx recipients (70.9% versus 89.2%; P = 0.002), and this correlated to an increase in the inducibility of pSTAT4 (MFI 157 versus 173; P = 0.0002). Conclusions. RBV cotreatment of HCV infection improved pSTAT4-dependent IFNγ-production in NK cells. This is relevant especially for immunocompromised patients such as LTx recipients or patients with end-stage liver disease.

Background Anaplastic thyroid carcinomas are associated with rapid tumor growth, short survival time and without any promising therapy to improve the poor prognosis. In this study, expression of immunoregulative receptor CD1d and lymphocyte infiltration in different thyroid tumors as well as in healthy tissue were analyzed in order to find new targets for an immunotherapeutic approach. Methods CD1d immunohistochemistry was performed in samples of 18 anaplastic, 17 follicular, 27 papillary, and 4 medullary thyroid carcinomas as well as in 19 specimens from normal thyroid tissue and additionally in 10 samples of sarcoma, seven malignant melanoma and three spindle‐cell lung carcinoma. Furthermore, thyroid samples were stained with antibodies against CD3, CD20, CD56, CD68, and LCA in order to analyze lymphocyte infiltration. Results For the first time CD1d receptor expression on normal thyroid tissue could be demonstrated. Moreover, anaplastic thyroid carcinomas showed significantly higher expression levels compared to other thyroid samples. Most astonishingly, CD1d expression disappeared from the cellular surface and was detected rather in the cytoplasm of anaplastic thyroid carcinoma cells. In addition, histologically similar tumors to anaplastic carcinoma like sarcoma and malignant melanoma revealed distinct CD1d staining patterns. Furthermore, infiltration of T cells, B cells, and macrophages in anaplastic thyroid carcinomas was different when compared to normal thyroid tissue and all other thyroid carcinomas. Conclusions Anaplastic thyroid carcinomas show significantly higher expression of CD1d, a receptor for NKT cells, which are subject of several anticancer therapy studies. These results may offer a novel approach to explore immunotherapeutic treatment options., Anaplastic thyroid carcinomas show significantly higher expression of CD1d, a receptor for NKT cells, which are subject of several anticancer therapy studies. For anaplastic thyroid carcinomas, this may offer a new differentiation marker as well as a novel approach to explore immunotherapeutic treatment options.

Background Risks for living-liver donors are lower in case of a left liver donation, however, due to lower graft volume, the risk for small-for-size situations in the recipients increases. This study aims to prevent small-for-size situations in recipients using an auxiliary two-staged partial resection liver transplantation (LTX) of living-donated left liver lobes. Case presentation Two patients received a two-stage auxiliary LTX using living-donated left liver lobes after left lateral liver resection. The native extended right liver was removed in a second operation after sufficient hypertrophy of the left liver graft had occurred. Neither donor developed postoperative complications. In both recipients, the graft volume increased by an average of 105% (329 ml to 641 ml), from a graft-to-body-weight ratio of 0.54 to 1.08 within 11 days after LTX, so that the remnant native right liver could be removed. No recipient developed small-for-size syndrome; graft function and overall condition is good in both recipients after a follow-up time of 25 months. Conclusions Auxiliary two-staged partial resection LTX using living-donor left lobes is technically feasible and can prevent small-for-size situation. This new technique can expand the potential living-donor pool and contributes to increase donor safety.

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.

Belatacept offers superior long-term outcome relative to calcineurin inhibitor (CNI)-based immunosuppression. However, the higher frequency of early T cell-mediated rejection (TCMR) in belatacept-treated patients hampered the widespread adoption of costimulation blockade. Here, we applied gene expression analysis and whole-slide inflammatory cell quantification to assess the impact of belatacept on intragraft immune signature. We studied formalin-fixed, paraffin-embedded renal biopsies from 92 patients stratified by histopathologic diagnosis (TCMR, borderline changes, or normal) and immunosuppression regimen (belatacept, CNI). An interaction model was built to explore maintenance treatment-dependent expression level changes of immune response-related genes across diagnostic categories of normal, borderline changes, and TCMR. Ninety-one percent of genes overexpressed in TCMR showed significant correlation with whole section inflammatory load. There were 27 genes that had a positive association with belatacept treatment. These were mostly related to myeloid cells and innate immunity. Genes negatively associated with costimulation blockade (n = 14) could be linked to B-cell differentiation and proliferation. We concluded that expression levels of genes characteristic of TCMR are strongly interconnected with quantitative changes of the biopsy inflammatory load. Our results might suggest differential involvement of the innate immune system, and an altered B-cell engagement during TCMR in belatacept-treated patients relative to CNI-treated referents.

Surgical site infections (SSI) in open Hepatopancreatobiliary (HPB) surgery are common complications. They worsen patients’ outcomes and prolong hospital stays. Their economic significance in the German diagnosis related groups (DRG) system is mostly unknown. To investigate their economic importance, we evaluated all cases for SSIs as well as clinical and financial parameters undergoing surgery in our centre from 2015 and 2016. Subsequently, we carried out a cost‐revenue calculation by assessing our billing data and the cost matrix of the InEK (German Institute for the Payment System in Hospitals). A total of 13.5% of the patients developed a superficial, 9% a deep incisional, and 2.4% of the patients an organ space SSI. Compared with Patients without SSI, Patients with SSI had more comorbidities, were older, and their average length of stay was extended by 19 days (P

The elusive nature of assessing immunological processes in situ in organ transplantation is one of the major impediments to improve diagnostics and treatment. Here, we present a proof-of-concept study using multiplexed in situ hybridization (ISH) (RNAscope) to detect low-abundance cytokines in formalin-fixed paraffin-embedded (FFPE) human transplant kidney biopsies in combination with immunofluorescence (IF) for cell phenotyping. We show that a multiplex IF and ISH (mIFISH) assay is feasible to identify the cellular source of cytokines and chemokines (tumor necrosis factor-α, interferon-γ, and CXCL9) in FFPE transplant kidney biopsies and that quantification of the mRNA and protein signal is also possible at single-cell resolution in the context of tissue complexity. Furthermore, the mIFISH assay allows precise quantitative assessment of tubulitis, one of the key morphological correlates of alloimmune injury. Simultaneous in situ identification and quantification of multiple cellular phenotypes and mRNA expression of proinflammatory cytokines in FFPE tissues offer a novel insight into the biology of alloimmune injury in kidney transplantation and may contribute to improved diagnostic accuracy and patient care.

Living donor liver transplantation (LDLT) from HCV-positive donors into HCV-positive recipients is not established yet (1). We report a case of combining direct antiviral agents (DAA) therapy and living liver donation from a donor with chronic-HCV-infection (genotype 3a) to a recipient with congenital chronic-HCV-infection (genotype 3a).

Background: There is an urgent need to develop and implement low cost, high-throughput standardized methods for routine molecular assessment of transplant biopsies. Given the vast archive of formalin-fixed and paraffin-embedded (FFPE) tissue blocks in transplant centers, a reliable protocol for utilizing this tissue bank for clinical validation of target molecules as predictors of graft outcome over time, would be of great value. Methods: We designed and optimized assays to quantify 19 target genes, including previously reported set of tissue common rejection module (tCRM) genes. We interrogated their performance for their clinical utility for detection of graft rejection and inflammation by analyzing gene expression microarrays analysis of 163 renal allograft biopsies, and subsequently validated in 40 independent FFPE archived kidney transplant biopsies at a single center. Results: A QPCR (Fluidigm) and a barcoded oligo-based (NanoString) gene expression platform were compared for evaluation of amplification of gene expression signal for 19 genes from degraded RNA extracted from FFPE biopsy sections by a set protocol. Increased expression of the selected 19 genes, that reflect a combination of specific cellular infiltrates (8/19 genes) and a graft inflammation score (11/19 genes which computes the tCRM score allowed for segregation of kidney transplant biopsies with stable allograft function and normal histology from those with histologically confirmed acute rejection (AR; p = 0.0022, QPCR; p = 0.0036, barcoded assay) and many cases of histological borderline inflammation (BL). Serial biopsy shaves used for gene expression were also processed for in-situ hybridization (ISH) for a subset of genes. ISH confirmed a high degree of correlation of signal amplification and tissue localization. Conclusions: Target gene expression amplification across a custom set of genes can identify AR independent of histology, and quantify inflammation from archival kidney transplant biopsy tissue, providing a new tool for clinical correlation and outcome analysis of kidney allografts, without the need for prospective kidney biopsy biobanking efforts.

The aim of this study was to examine the development of biliary epithelial damage between organ retrieval and transplantation and its clinical relevance for patients.Common bile duct samples during donor hepatectomy, after cold storage, and after reperfusion were compared to healthy controls by hematoxylin and eosin (HE) staining and immunofluorescence for tight junction protein 1 and Claudin-1. A bile duct damage score to quantify biliary epithelial injury was developed and correlated with recipient and donor data and patient outcome.Control (N=16) and donor hepatectomy bile ducts (N=10) showed regular epithelial morphology and tight junction architecture. After cold storage (N=37; p=0.0119), and even more after reperfusion (N=62; p=0.0002), epithelial damage, as quantified by the bile duct damage score, was markedly increased, and both tight junction proteins were detected with inappropriate morphology. Patients with major bile duct damage after cold storage had a significantly increased risk of biliary complications (relative risk 18.75; p0.0001) and graft loss (p=0.0004).In many cases, the common bile duct epithelium shows considerable damage after cold ischemia with further damage occurring after reperfusion. The extent of epithelial damage can be quantified by our newly developed bile duct damage score and is a prognostic parameter for biliary complications and graft loss. Possibly, in an intraoperative histological examination, this bile duct damage score may influence decision-making in transplantation surgery.