Background Cryptococcus is the most common cause of adult meningitis in Africa. We evaluated the activity of adjunctive sertraline, previously demonstrated to have in vitro and in vivo activity against Cryptococcus. Methods We enrolled 172 HIV-infected Ugandans with cryptococcal meningitis from August 2013 through August 2014 into an open-label dose-finding study to assess safety and microbiologic efficacy. Sertraline 100–400mg/day was added to standard therapy of amphotericin + fluconazole 800mg/day. We evaluated early fungicidal activity via Cryptococcus cerebrospinal fluid (CSF) clearance rate, sertraline pharmacokinetics, and in vitro susceptibility. Findings Participants receiving any sertraline dose averaged a CSF clearance rate of −0·37 (95%CI: −0·41, −0·33) colony forming units (CFU)/mL/day. Incidence of paradoxical immune reconstitution inflammatory syndrome (IRIS) was 5% (2/43) and relapse was 0% through 12-weeks. Sertraline reached steady state concentrations in plasma by day 7, with median steady-state concentrations of 201 ng/mL (IQR, 90–300; n=49) with 200mg/day and 399 ng/mL (IQR, 279–560; n=30) with 400mg/day. Plasma concentrations reached 83% of steady state levels by day 3. The median projected steady state brain tissue concentration at 200mg/day was 3·7 (IQR, 2·0–5·7) mcg/mL and 6·8 (IQR, 4·6–9·7) mcg/mL at 400mg/day. Minimum inhibitory concentrations were ≤2 mcg/mL for 27% (35/128), ≤4 mcg/mL for 84% (108/128), ≤6 mcg/mL for 91% (117/128), and ≤8 mcg/mL for 100% of 128 Cryptococcus isolates. Interpretation Sertraline had faster cryptococcal CSF clearance, decreased IRIS, and decreased relapse compared with historical experiences. Sertraline reaches therapeutic levels in a clinical setting. This inexpensive and off-patent oral medication is a promising adjunctive antifungal therapy. Funding National Institutes of Health, Grand Challenges Canada.

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Cell therapies, which include bioartificial liver support and hepatocyte transplantation, have emerged as potential treatments for a variety of liver diseases. Acute liver failure (ALF), acute-on-chronic liver failure, and inherited metabolic liver diseases are examples of liver diseases that have been successfully treated with cell therapies at centers around the world. Cell therapies also have the potential for wide application in other liver diseases, including non-inherited liver diseases and liver cancer, and in improving the success of liver transplantation. Here we briefly summarize current concepts of cell therapy for liver diseases.

Hereditary tyrosinemia type I (HT1) is caused by deficiency in fumarylacetoacetate hydrolase (FAH), an enzyme that catalyzes the last step of tyrosine metabolism. The most severe form of the disease presents acutely during infancy, and is characterized by severe liver involvement, most commonly resulting in death if untreated. Generation of FAH+/− pigs was previously accomplished by adeno-associated virus-mediated gene knockout in fibroblasts and somatic cell nuclear transfer. Subsequently, these animals were outbred and crossed to produce the first FAH−/− pigs. FAH-deficiency produced a lethal defect in utero that was corrected by administration of 2-(2-nitro-4-trifluoromethylbenzyol)-1,3 cyclohexanedione (NTBC) throughout pregnancy. Animals on NTBC were phenotypically normal at birth; however, animals were euthanized approximately four weeks after withdrawal of NTBC due to clinical decline and physical examination findings of severe liver injury and encephalopthy consistent with acute liver failure. Biochemical and histological analyses, characterized by diffuse and severe hepatocellular damage, confirmed the diagnosis of severe liver injury. FAH−/− pigs provide the first genetically engineered large animal model of a metabolic liver disorder. Future applications of FAH−/− pigs include discovery research as a large animal model of HT1 and spontaneous acute liver failure, and preclinical testing of efficacy of liver cell therapies, including transplantation of hepatocytes, liver stem cells, and pluripotent stem cell-derived hepatocytes.

Hereditary tyrosinemia type I (HT1) results in hepatic failure, cirrhosis, and hepatocellular carcinoma (HCC) early in childhood and is caused by deficiency in the enzyme fumarylacetoacetate hydrolase (FAH). In a novel approach we used the chimeric adeno-associated virus DJ serotype (AAV-DJ) and homologous recombination to target and disrupt the porcine Fah gene. AAV-DJ is an artificial chimeric AAV vector containing hybrid capsid sequences from three naturally occurring serotypes (AAV2, 8 and 9). The AAV-DJ vector was used to deliver the knockout construct to fetal pig fibroblasts with an average knockout targeting frequencies of 5.4%. Targeted Fah-null heterozygote fibroblasts were used as nuclear donors for somatic cell nuclear transfer (SCNT) to porcine oocytes, and multiple viable Fah-null heterozygote pigs were generated. Fah-null heterozygotes were phenotypically normal, but had decreased Fah transcriptional and enzymatic activity compared to wild-type animals. Conclusion This study is the first to use a recombinant chimeric AAV vector to knockout a gene in porcine fibroblasts for the purpose of SCNT. In using the AAV-DJ vector we observed targeting frequencies that were higher than previously reported with other naturally occurring serotypes. We expect that the subsequent generation of FAH-null homozygote pigs will serve as a significant advancement for translational research in the areas of metabolic liver disease, cirrhosis and HCC.

Canadian university disclosures have been tracked from 1988 to 2000 using the modified accountability disclosure (MAD) index developed by Coy, Dixon, and Tower (1993) and Coy, Tower, and Dixon (1993) in their study of New Zealand universities. During the first eight years of the period under investigation, there was very little change in accountability disclosures. However, for the periods ending in 1997 through 2000, there has been a statistically significant annual improvement. This paper examines the reasons for these changes as indicated in the interviews with the presidents, or their designates, of Canadian universities. Factors include increased fund raising by the universities and pressures by the public and governments for universities to become more accountable, while a change in accounting pronouncements appears to have had little effect.

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Background Significant morbidity associated with acute liver failure (ALF) is from the systemic inflammatory response syndrome (SIRS). Toll-like receptor 4 (TLR4) has been shown to play an integral role in the modulation of SIRS. However, little is known about the mechanistic role of TLR4 in ALF. Also, no cell type has been identified as the key mediator of the TLR4 pathway in ALF. This study examines the role of TLR4 and Kupffer cells in the development of the SIRS following acetaminophen (APAP)-induced ALF. Materials and Methods Five groups of mice were established: untreated wild-type, E5564-treated (a TLR4 antagonist), gadolinium chloride (GdCl3)-treated (Kupffer cell-depleted), clodronate-treated (Kupffer cell-depleted), and TLR4-mutant. Following APAP administration, 72-hour survival, biochemical and histologic liver injury, extent of lung injury and edema, and pro-inflammatory gene expression were studied. Additionally, TLR4 expression was determined in livers of wild-type and Kupffer cell-depleted mice. Results Following APAP administration, wild-type, TLR4-mutant, E5564-treated, and Kupffer cell-depleted mice had significant liver injury. However, wild-type mice had markedly worse survival compared to the other 4 treatment groups. TLR4-mutant, E5564-treated, and Kupffer cell-depleted mice had less lung inflammation and edema than wild-type mice. Selected pro-inflammatory gene expression (il1b, il6, tnf) in TLR4-mutant, E5564-treated, and Kupffer cell-depleted mice was significantly lower compared to wild-type mice after acute liver injury. Conclusion This study demonstrates that survival in APAP-induced ALF potentially correlates with the level of pro-inflammatory gene expression. This study points to a link between TLR4 and Kupffer cells in the APAP model of ALF, and, more importantly, demonstrates benefits of TLR4 antagonism in ALF.