Nicolas, Clara T.

Abstract Introduction Identification of hospitalized patients with suddenly unfavorable clinical course remains challenging. Models using objective data elements from the electronic health record may miss important sources of information available to nurses. Methods We recorded nurses’ perception of patient potential for deterioration in 2 medical and 2 surgical adult hospital units using a 5-point score at the start of the shift (the Worry Factor [WF]), and any time a change or an increase was noted by the nurse. Cases were evaluated by three reviewers. Intensive care unit (ICU) transfers were also tracked. Results 31 159 patient-shifts were recorded for 3185 unique patients during 3551 hospitalizations, with 169 total outcome events. Out of 492 potential deterioration events identified, 380 (77%) were confirmed by reviewers as true deterioration events. Likelihood ratios for ICU transfer were 17.8 (15.2–20.9) in the 24 hours following a WF > 2, and 40.4 (27.1–60.1) following a WF > 3. Accuracy rates were significantly higher in nurses with over a year of experience (68% vs 79%, P = 0.04). The area under the receiver operator characteristic curve (AUROC) was 0.92 for the prediction of ICU transfer within 24 hours. Discussion This is a higher accuracy than most published early warning scores. Conclusion Nurses’ pattern recognition and sense of worry can provide important information for the detection of acute physiological deterioration and should be included in the electronic medical record.

ABSTRACTObjectivesThe congenital lung malformation volume ratio (CVR) is a prenatal ultrasound measurement that parameterizes congenital lung malformation (CLM) size. The aims of this study were to use serial measurements to create estimated growth curves of fetal CVR for asymptomatic and symptomatic neonates with CLM and to investigate whether a discriminant prognostic model based on these measurements could predict accurately which fetuses with CLM will require invasive respiratory support at delivery and should therefore be delivered at a tertiary‐care facility.MethodsThis was a retrospective study of fetuses diagnosed prenatally with CLM at three tertiary‐care children's hospitals between 2009 and 2016. Those with two or more sonographic measurements of CVR were included. Serial fetal CVR measurements were used to create estimated growth curves for neonates with and those without respiratory symptoms at delivery, defined as requiring invasive respiratory support for the first 24 h after delivery. A discriminant model based on serial CVR measurements was used to calculate the dynamic probability of the need for invasive respiratory support. The performance of this model overall and in preterm and term neonates was compared with those using maximum CVR thresholds of 1.0 and 1.6.ResultsOf the 147 neonates meeting the inclusion criteria, 16 (10.9%) required postnatal invasive respiratory support. The estimated CVR growth curve models showed different growth trajectories for asymptomatic and symptomatic neonates, with significantly higher CVR in symptomatic neonates, and values peaking late in the second trimester at around 25 weeks' gestation in asymptomatic neonates. All prognostic methods had high accuracy for the prediction of the need for invasive respiratory support in term neonates, but the discriminant model had the best performance overall (area under the receiver–operating characteristics curve (AUC) = 0.88) and in the preterm population (AUC = 0.85).ConclusionsThe estimated CVR growth curves showed different growth patterns in asymptomatic and symptomatic neonates with CLM. The dynamic discriminant model performed well overall and particularly in neonates that were carried to term. Development of an externally validated clinical tool based on this analysis could be useful in determining the site of delivery for fetuses with CLM. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Ex vivo CRISPR/Cas9‐mediated gene editing in hepatocytes using homology‐directed repair (HDR) is a potential alternative curative therapy to organ transplantation for metabolic liver disease. However, a major limitation of this approach in quiescent adult primary hepatocytes is that nonhomologous end‐joining is the predominant DNA repair pathway for double‐strand breaks (DSBs). This study explored the hypothesis that ex vivo hepatocyte culture could reprogram hepatocytes to favor HDR after CRISPR/Cas9‐mediated DNA DSBs. Quantitative PCR (qPCR), RNA sequencing, and flow cytometry demonstrated that within 24 hours, primary mouse hepatocytes in ex vivo monolayer culture decreased metabolic functions and increased expression of genes related to mitosis progression and HDR. Despite the down‐regulation of hepatocyte function genes, hepatocytes cultured for up to 72 hours could robustly engraft in vivo. To assess functionality long‐term, primary hepatocytes from a mouse model of hereditary tyrosinemia type 1 bearing a single‐point mutation were transduced ex vivo with two adeno‐associated viral vectors to deliver the Cas9 nuclease, target guide RNAs, and a 1.2‐kb homology template. Adeno‐associated viral Cas9 induced robust cutting at the target locus, and, after delivery of the repair template, precise correction of the point mutation occurred by HDR. Edited hepatocytes were transplanted into recipient fumarylacetoacetate hydrolase knockout mice, resulting in engraftment, robust proliferation, and prevention of liver failure. Weight gain and biochemical assessment revealed normalization of metabolic function. Conclusion: The results of this study demonstrate the potential therapeutic effect of ex vivo hepatocyte‐directed gene editing after reprogramming to cure metabolic disease in a preclinical model of hereditary tyrosinemia type 1.

Orthotopic liver transplantation remains the only curative therapy for inborn errors of metabolism. Given the tremendous success for primary immunodeficiencies using ex-vivo gene therapy with lentiviral vectors, there is great interest in developing similar curative therapies for metabolic liver diseases. We have previously generated a pig model of hereditary tyrosinemia type 1 (HT1), an autosomal recessive disorder caused by deficiency of fumarylacetoacetate hydrolase (FAH). Using this model, we have demonstrated curative ex-vivo gene and cell therapy using a lentiviral vector to express FAH in autologous hepatocytes. To further evaluate the long-term clinical outcomes of this therapeutic approach, we continued to monitor one of these pigs over the course of three years. The animal continued to thrive off the protective drug NTBC, gaining weight appropriately, and maintaining sexual fecundity for the course of his life. The animal was euthanized 31 months after transplantation to perform a thorough biochemical and histological analysis. Biochemically, liver enzymes and alpha-fetoprotein levels remained normal and abhorrent metabolites specific to HT1 remained corrected. Liver histology showed no evidence of tumorigenicity and Masson’s trichrome staining revealed minimal fibrosis and no evidence of cirrhosis. FAH-immunohistochemistry revealed complete repopulation of the liver by transplanted FAH-positive cells. A complete histopathological report on other organs, including kidney, revealed no abnormalities. This study is the first to demonstrate long-term safety and efficacy of hepatocyte-directed gene therapy in a large animal model. We conclude that hepatocyte-directed ex-vivo gene therapy is a rational choice for further exploration as an alternative therapeutic approach to whole organ transplantation for metabolic liver disease, including HT1.

Acute liver failure (ALF) is a catastrophic condition that can occur after major liver resection. The aim of this study was to determine the effects of the spheroid reservoir bio‐artificial liver (SRBAL) on survival, serum chemistry, and liver regeneration in posthepatectomy ALF pigs. Wild‐type large white swine (20 kg‐30 kg) underwent intracranial pressure (ICP) probe placement followed by 85% hepatectomy. Computed tomography (CT) volumetrics were performed to measure the extent of resection, and at 48 hours following hepatectomy to assess regeneration of the remnant liver. Animals were randomized into three groups based on treatment delivered 24‐48 hours after hepatectomy: Group1—standard medical therapy (SMT, n = 6); Group2—SMT plus bio‐artificial liver treatment using no hepatocytes (0 g, n = 6); and Group3—SMT plus SRBAL treatment using 200 g of primary porcine hepatocyte spheroids (200 g, n = 6). The primary endpoint was survival to 90 hours following hepatectomy. Death equivalent was defined as unresponsive grade 4 hepatic encephalopathy or ICP greater than 20 mmHg with clinical evidence of brain herniation. All animals in both (SMT and 0 g) control groups met the death equivalent before 51 hours following hepatectomy. Five of 6 animals in the 200‐g group survived to 90 hours ( P < 0.01). The mean ammonia, ICP, and international normalized ratio values were significantly lower in the 200‐g group. CT volumetrics demonstrated increased volume regeneration at 48 hours following hepatectomy in the 200‐g group compared with the SMT ( P < 0.01) and 0‐g (P < 0.01) groups. Ki‐67 staining showed increased positive staining at 48 hours following hepatectomy ( P < 0.01). Conclusion: The SRBAL improved survival, reduced ammonia, and accelerated liver regeneration in posthepatectomy ALF. Improved survival was associated with a neuroprotective benefit of SRBAL therapy. These favorable results warrant further clinical testing of the SRBAL.

Share 35 was implemented in 2013 to direct livers to the most urgent candidates by prioritizing Model for End‐Stage Liver Disease (MELD) ≥ 35 patients. We aim to evaluate this policy's impact on costs and mortality. Our study includes 834 wait‐listed patients and 338 patients who received deceased donor, solitary liver transplants at Mayo Clinic between January 2010 and December 2014. Of these patients, 101 (30%) underwent transplantation after Share 35. After Share 35, 29 (28.7%) MELD ≥ 35 patients received transplants, as opposed to 46 (19.4%) in the pre–Share 35 era (P = 0.06). No significant difference in 90‐day wait‐list mortality (P = 0.29) nor 365‐day posttransplant mortality (P = 0.68) was found between patients transplanted before or after Share 35. Mean costs were $3,049 (P = 0.30), $5226 (P = 0.18), and $10,826 (P = 0.03) lower post‐Share 35 for the 30‐, 90‐, and 365‐day pretransplant periods, and mean costs were $5010 (P = 0.41) and $5859 (P = 0.57) higher, and $9145 (P = 0.54) lower post‐Share 35 for the 30‐, 90‐, and 365‐day posttransplant periods. In conclusion, the added cost of transplanting more MELD ≥ 35 patients may be offset by pretransplant care cost reduction. Despite shifting organs to critically ill patients, Share 35 has not impacted mortality significantly. Liver Transplantation 23:11–18 2017 AASLD.

Abstract Donor organ shortage is the main limitation to liver transplantation as a treatment for end-stage liver disease and acute liver failure. Liver regenerative medicine may in the future offer an alternative form of therapy for these diseases, be it through cell transplantation, bioartificial liver (BAL) devices, or bioengineered whole organ liver transplantation. All three strategies have shown promising results in the past decade. However, before they are incorporated into widespread clinical practice, the ideal cell type for each treatment modality must be found, and an adequate amount of metabolically active, functional cells must be able to be produced. Research is ongoing in hepatocyte expansion techniques, use of xenogeneic cells, and differentiation of stem cell-derived hepatocyte-like cells (HLCs). HLCs are a few steps away from clinical application, but may be very useful in individualized drug development and toxicity testing, as well as disease modeling. Finally, safety concerns including tumorigenicity and xenozoonosis must also be addressed before cell transplantation, BAL devices, and bioengineered livers occupy their clinical niche. This review aims to highlight the most recent advances and provide an updated view of the current state of affairs in the field of liver regenerative medicine.

Cell therapy is an emerging form of treatment for several liver diseases, but is limited by the availability of donor livers. Stem cells hold promise as an alternative to the use of primary hepatocytes. We performed an exhaustive review of the literature, with a focus on the latest studies involving the use of stem cells for the treatment of liver disease. Stem cells can be harvested from a number of sources, or can be generated from somatic cells to create induced pluripotent stem cells (iPSCs). Different cell lines have been used experimentally to support liver function and treat inherited metabolic disorders, acute liver failure, cirrhosis, liver cancer, and small-for-size liver transplantations. Cell-based therapeutics may involve gene therapy, cell transplantation, bioartificial liver devices, or bioengineered organs. Research in this field is still very active. Stem cell therapy may, in the future, be used as a bridge to either liver transplantation or endogenous liver regeneration, but efficient differentiation and production protocols must be developed and safety must be demonstrated before it can be applied to clinical practice.