Treatment and survival of non-alcoholic steatohepatitis associated hepatocellular carcinoma.

BACKGROUND: The incidence of non-alcoholic steatohepatitis (NASH) is increasing worldwide and a poorly defined subset of patients develops end-stage liver disease and hepatocellular carcinoma (HCC). Differences in the biological behaviour, tumour characteristics, associated risk factors, treatment outcomes and overall survival of patients with NASH-HCC remain poorly defined. The aim of this study was to determine and analyze these differences in a large clinical cohort to guide treatment decisions. METHODS: 1119 patients with HCC treated in an 11 year period at the University Medical Centre of the Johannes Gutenberg University Mainz were retrospectively analyzed. RESULTS: Patients with NASH-HCC (n = 45) were older (67.6 vs. 65 years), had an increased frequency of the metabolic syndrome and complications with a higher incidence of obesity (31.1% vs. 14.7%), type II diabetes mellitus (66.7% vs. 37.85%), a higher rate of myocardial infarction (13.3% vs. 4.8%) and apoplectic stroke (8.9% vs. 2.1%) (all p < 0.05). Interestingly, liver function was preserved to a higher extent and MELD scores were significantly lower in NASH-HCC. Nonetheless, resection or orthotopic liver transplantation was performed only in 17.8% and 4.4% of NASH-HCC respectively. Overall survival was lower compared to HCC of other aetiologies. Independent of the underlying aetiology BMI exhibited a positive correlation with overall survival. CONCLUSION: Despite retained liver function, patients with NASH-associated HCC showed a decreased overall survival. With regards to the expected increasing prevalence of NASH, it will be necessary to improve screening and surveillance strategies to identify HCC in NASH early and improve survival.

Quantification of human papillomavirus cell-free DNA from low-volume blood plasma samples by digital PCR.

The incidence rate of human papillomavirus-driven oropharyngeal cancer (HPV-OPC) is increasing in countries with high human development index. HPV cell-free DNA (cfDNA) isolated from 3 to 4 mL blood plasma has been successfully used for therapy surveillance. A highly discussed application of HPV-cfDNA is early detection of HPV-OPC. This requires sensitive and specific cfDNA detection as cfDNA levels can be very low. To study the predictive power of pre-diagnostic HPV-cfDNA, archived samples from epidemiological cohorts with limited plasma volume are an important source. To establish a cfDNA detection workflow for low plasma volumes, we compared cfDNA purification methods [MagNA Pure 96 (MP96) and QIAamp ccfDNA/RNA] and digital PCR systems (Biorad QX200 and QIAGEN QIAcuity One). Final assay validation included 65 low-volume plasma samples from oropharyngeal cancer (OPC) patients with defined HPV status stored for 2-9 years. MP96 yielded a 28% higher cfDNA isolation efficiency in comparison to QIAamp. Both digital PCR systems showed comparable analytical sensitivity (6-17 copies for HPV16 and HPV33), but QIAcuity detected both types in the same assay. In the validation set, the assay had 80% sensitivity (n = 28/35) for HPV16 and HPV33 and a specificity of 97% (n = 29/30). In samples with ≥750 µL plasma, the sensitivity was 85% (n = 17/20), while in samples with <750 µL plasma, it was 73% (n = 11/15). Despite the expected drop in sensitivity with decreased plasma volume, the assay is sensitive and highly specific even in low-volume samples and thus suited for studies exploring HPV-cfDNA as an early HPV-OPC detection marker in low-volume archival material.IMPORTANCEHPV-OPC has a favorable prognosis compared to HPV-negative OPC. However, the majority of tumors is diagnosed after regional spread, thus making intensive treatment necessary. This can cause lasting morbidity with a large impact on quality of life. One potential method to decrease treatment-related morbidity is early detection of the cancer. HPV cfDNA has been successfully used for therapy surveillance and has also been detected in pre-diagnostic samples of HPV-OPC patients. These pre-diagnostic samples are only commonly available from biobanks, which usually only have small volumes of blood plasma available. Hence, we have developed a workflow optimized for small-volume archival samples. With this method, a high sensitivity can be achieved despite sample limitations, making it suitable to conduct further large-scale biobank studies of HPV-cfDNA as a prognostic biomarker for HPV-OPC.

Generation of Vascularized Neural Organoids by Co-culturing with Mesodermal Progenitor Cells.

Organoids are three-dimensional (3D) constructs generated in stem cell cultures and are thought to mimic tissue and organ development in situ. However, until recently, they often exclusively recapitulated the development of the organ`s parenchyma without the major components of the organ stroma. Here, we describe a protocol to incorporate stromal components, first of all blood vessels, by co-culturing with induced pluripotent stem cell-derived mesodermal progenitor cells. For complete details on the use and execution of this protocol, please refer to Wörsdörfer et al. (2019).

The Impact of Liver Cell Injury on Health-Related Quality of Life in Patients with Chronic Liver Disease.

BACKGROUND: Patients with chronic liver disease often suffer from unspecific symptoms and report severe impairment in the quality of life. The underlying mechanisms are multifactorial and include disease-specific but also liver related causes. The current analysis evaluated the association of hepatocellular apoptosis in non-viral chronic liver disease and health-related quality of life (HRQL). Furthermore we examined factors, which influence patient's physical and mental well-being. METHODS: A total of 150 patients with non-infectious chronic liver disease were included between January 2014 and June 2015. The German version of the Chronic Liver Disease Questionnaire (CLDQ-D), a liver disease specific instrument to assess HRQL, was employed. Hepatocellular apoptosis was determined by measuring Cytokeratin 18 (CK18, M30 Apoptosense ELISA). RESULTS: Female gender (5.24 vs. 5.54, p = 0.04), diabetes mellitus type II (4.75 vs. 5.46, p<0.001) and daily drug intake (5.24 vs. 6.01, p = 0.003) were associated with a significant impairment in HRQL. HRQL was not significantly different between the examined liver diseases. Levels of CK18 were the highest in patients with NASH compared to all other disease entities (p<0.001). Interestingly, CK18 exhibited significant correlations with obesity (p<0.001) and hyperlipidemia (p<0.001). In patients with cirrhosis levels of CK18 correlated with the MELD score (r = 0.18, p = 0.03) and were significantly higher compared to patients without existing cirrhosis (265.5 U/l vs. 186.9U/l, p = 0.047). Additionally, CK18 showed a significant correlation with the presence and the degree of hepatic fibrosis (p = 0.003) and inflammation (p<0.001) in liver histology. Finally, there was a small negative association between CLDQ and CK18 (r = -0.16, p = 0.048). CONCLUSION: Different parameters are influencing HRQL and CK18 levels in chronic non-viral liver disease and the amount of hepatocellular apoptosis correlates with the impairment in HRQL in chronic non-viral liver diseases. These findings support the role of liver-protective therapies for the improvement of the quality of life in chronic liver disease.