Hepatocellular carcinoma after direct-acting antivirals for hepatitis C is associated with KIR-HLA types predicting weak NK cell-mediated immunity.

BACKGROUND AND AIMS: Second-generation direct-acting antivirals (2G DAA) to cure HCV have led to dramatic clinical improvements. HCV-associated hepatocellular carcinoma (HCC), however, remains common. Impaired immune tumor surveillance may play a role in HCC development. Our cohort evaluated the effects of innate immune types and clinical variables on outcomes including HCC. METHODS: Participants underwent full HLA class I/KIR typing and long-term HCV follow-up. RESULTS: A total of 353 HCV+ participants were followed for a mean of 7 years. Cirrhosis: 25% at baseline, developed in 12% during follow-up. 158 participants received 2G DAA therapy. HCC developed without HCV therapy in 20 subjects, 24 HCC after HCV therapy, and 10 of these after 2G DAA. Two predictors of HCC among 2G DAA-treated patients: cirrhosis (OR, 10.0, p = 0.002) and HLA/KIR profiles predicting weak natural killer (NK) cell-mediated immunity (NK cell complementation groups 6, 9, 11, 12, OR of 5.1, p = 0.02). Without 2G DAA therapy: cirrhosis was the main clinical predictor of HCC (OR, 30.8, p < 0.0001), and weak NK-cell-mediated immunity did not predict HCC. CONCLUSION: Cirrhosis is the main risk state predisposing to HCC, but weak NK-cell-mediated immunity may predispose to post-2G DAA HCC more than intermediate or strong NK-cell-mediated immunity.

Surgical Site Complications in Open Pronation-Abduction Ankle Fracture-Dislocations With Medial Tension Failure Wounds.

OBJECTIVES: To examine the incidence of surgical site complications associated with pronation-abduction ankle fracture-dislocations with an open medial tension wound. DESIGN: Retrospective case series. SETTING: Accredited Level-1 trauma center. PATIENTS/PARTICIPANTS: Forty-eight open pronation-abduction ankle fracture-dislocations with medial tension failure wounds treated at our institution from 2014 to 2016. INTERVENTION: Immediate irrigation and debridement along with surgical stabilization of open ankle fracture-dislocation. MAIN OUTCOME MEASURES: The primary outcome measure was deep surgical site infection. Secondary outcome measures included other surgical site complications and adverse radiographic events. RESULTS: A total of 5 patients (10.4%) developed a deep surgical site infection requiring additional surgical debridement. One of the patients with a deep surgical site infection required a below-knee amputation as a result of sepsis. Adverse radiographic outcomes included 3 fibular nonunions (6.3%), 3 implant failures related to syndesmotic fixations (6.3%), one periimplant fracture (2.1%), and postoperative collapse of the tibial plafond in 3 patients (6.3%). CONCLUSIONS: Open pronation-abduction ankle fracture-dislocations with medial tension failure wounds remain a challenging and potentially devastating injury. Our study suggests that with appropriate surgical debridement, early stabilization, and primary wound closure, acceptable outcomes with a relatively low risk of surgical site complications can be achieved. LEVEL OF EVIDENCE: Therapeutic Level IV. See Instructions for Authors for a complete description of levels of evidence.

Association of the patterns of use of medications with mortality of COVID-19 infection: a hospital-based observational study.

OBJECTIVES: SARS-CoV-2 enters cells using the ACE2 receptor. Medications that affect ACE2 expression or function such as angiotensin receptor blockers (ARBs) and ACE inhibitors (ACE-I) and metformin have the potential to counter the dysregulation of ACE2 by the virus and protect against viral injury. Here, we describe COVID-19 survival associated with ACE-I, ARB and metformin use. DESIGN: This is a hospital-based observational study of patients with COVID-19 infection using logistic regression with correction for pre-existing conditions and propensity score weighted Cox proportional hazards models to estimate associations between medication use and mortality. SETTING: Medical record data from the US Veterans Affairs (VA) were used to identify patients with a reverse transcription PCR diagnosis of COVID-19 infection, to classify patterns of ACE inhibitors (ACE-I), ARB, beta blockers, metformin, famotidine and remdesivir use, and, to capture mortality. PARTICIPANTS: 9532 hospitalised patients with COVID-19 infection followed for 60 days were analysed. OUTCOME MEASURE: Death from any cause within 60 days of COVID-19 diagnosis was examined. RESULTS: Discontinuation of ACE-I was associated with increased risk of death (OR: 1.4; 95% CI 1.2-1.7). Initiating (OR: 0.3; 95% CI 0.2-0.5) or continuous (OR: 0.6; 95% CI 0.5-0.7) ACE-I was associated with reduced risk of death. ARB and metformin associations were similar in direction and magnitude and also statistically significant. Results were unchanged when accounting for pre-existing morbidity and propensity score adjustment. CONCLUSIONS: Recent randomised clinical trials support the safety of continuing ACE-I and ARB treatment in patients with COVID-19 where indicated. Our study extends these findings to suggest a possible COVID-19 survival benefit for continuing or initiating ACE-I, ARB and metformin medications. Randomised trials are appropriate to confirm or refute the therapeutic potential for ACE-I, ARBs and metformin.

Fate of the Uninsured Ankle Fracture: Significant Delays in Treatment Result in an Increased Risk of Surgical Site Infection.

OBJECTIVE: To examine the impact of insurance status on access to orthopaedic care and incidence of surgical site complications in patients with closed unstable ankle fractures. DESIGN: Retrospective chart review. SETTING: Certified Level-1 urban trauma center and county facility. PARTICIPANTS: Four hundred eighty-nine patients with closed unstable ankle fractures undergoing open reduction and internal fixation between 2014 and 2016. INTERVENTION: Open reduction and internal fixation of unstable ankle fracture. MAIN OUTCOME MEASURES: Time from injury to presentation, time from injury to surgery, rate of surgical site infections, and loss to follow-up. RESULTS: A total of 489 patients (70.5% uninsured vs. 29.5% insured) were enrolled. Uninsured patients were more likely to be present to an outside hospital first (P = 0.004). Time from injury to presentation at our hospital was significantly longer in uninsured patients (4.5 ± 7.6 days vs. 2.3 ± 5.5 days, P < 0.001). Time from injury to surgery was significantly longer in uninsured patient (9.4 ± 8.5 days vs. 7.3 ± 9.1 days, P < 0.001). Uninsured patients were more likely to be lost to postoperative follow-up care (P = 0.002). A logistic regression analysis demonstrated that delayed surgical timing was directly associated with an increased risk of postoperative surgical site infection (P = 0.002). CONCLUSIONS: Uninsured patients with ankle fractures requiring surgery experience significant barriers regarding access to health care. Delay of surgical management significantly increases the risk of surgical site infections in closed unstable ankle fractures. LEVEL OF EVIDENCE: Prognostic Level III. See Instructions for Authors for a complete description of levels of evidence.

Skin closure with surgical staples in ankle fractures: a safe and reliable method.

PURPOSE: The purpose of this study is to examine the rates of surgical site complications of staple closure versus suture closure following open reduction and internal fixation of closed unstable ankle fractures. METHODS: Between 2014 and 2016, a total of 545 patients with closed ankle fractures were treated at our level-1 trauma centre by means of open reduction and internal fixation. A total of 360 patients matched the inclusion criteria and were included in the final analysis of this study. This included 119 patients undergoing wound closure using sutures and 241 patients using surgical staples. The demographics, clinical data, and injury characteristics were recorded. The primary outcome measure was the adverse event of any type of surgical site complication. RESULTS: The overall rate of patients with a surgical site complication was 15.6%. There was a trend towards a higher risk of surgical site complication in patients undergoing wound closure with sutures as compared with staples (20.2% versus 13.3%); however, this difference was not statistically significant (P = 0.0897). The rate of superficial surgical site infection also trended higher in patients undergoing wound closure with sutures versus staples without demonstrating statistical significance (10.1% versus 5%, P = 0.0678). The rate of deep surgical site infection was similar in both groups. CONCLUSION: The use of metal staples remains controversial in the setting of orthopedic surgery, particularly involving the foot and ankle. The current study supports that metal staples are a safe and reliable option in the closure of traumatic ankle fractures.

SHIP1 Deficiency in Inflammatory Bowel Disease Is Associated With Severe Crohn's Disease and Peripheral T Cell Reduction.

In our previous study, we observed a severe reduction in the Src homology 2-containing-inositol-phosphatase-1 (SHIP1) protein in a subpopulation of subjects from a small adult Crohn's Disease (CD) cohort. This pilot study had been undertaken since we had previously demonstrated that engineered deficiency of SHIP1 in mice results in a spontaneous and severe CD-like ileitis. Here, we extend our analysis of SHIP1 expression in peripheral blood mononuclear cells in a second much larger adult Inflammatory Bowel Disease (IBD) cohort, comprised of both CD and Ulcerative Colitis patients, to assess contribution of SHIP1 to the pathogenesis of human IBD. SHIP1 protein and mRNA levels were evaluated from blood samples obtained from IBD subjects seen at UCSF/SFVA, and compared to healthy control samples. Western blot analyses revealed that ~15% of the IBD subjects are severely SHIP1-deficient, with less than 10% of normal SHIP1 protein present in PBMC. Further analyses by flow cytometry and sequencing were performed on secondary samples obtained from the same subjects. Pan-hematolymphoid SHIP1 deficiency was a stable phenotype and was not due to coding changes in the INPP5D gene. A very strong association between SHIP1 deficiency and the presence of a novel SHIP1:ATG16L1 fusion transcript was seen. Similar to SHIP1-deficient mice, SHIP1-deficient subjects had reduced numbers of circulating CD4+ T cell numbers. Finally, SHIP1-deficient subjects with CD had a history of severe disease requiring multiple surgeries. These studies reveal that the SHIP1 protein is crucial for normal T cell homeostasis in both humans and mice, and that it is also a potential therapeutic and/or diagnostic target in human IBD.

Identification of MHC Class Ib Ligands for Stimulatory and Inhibitory Ly49 Receptors and Induction of Potent NK Cell Alloresponses in Rats.

Early studies indicate that rats may have a repertoire of MHC class Ib-reactive Ly49 stimulatory receptors capable of mounting memory-like NK cell alloresponses. In this article, we provide molecular and functional evidence for this assumption. Pairs of Ly49 receptors with sequence similarities in the lectin-like domains, but with opposing signaling functions, showed specificity for ligands with class Ia-like structural features encoded from the first telomeric MHC class Ib gene cluster, RT1-CE, which is syntenic with the H2-D/H2-L/H2-Q cluster in mice. The activating Ly49s4 receptor and its inhibitory counterparts, Ly49i4 and Ly49i3, reacted with all allelic variants of RT1-U, whereas Ly49s5 and Ly49i5 were specific for RT1-Eu NK cell cytolytic responses were predictably activated and inhibited, and potent in vivo NK alloresponses were induced by repeated MHC class Ib alloimmunizations. Additional Ly49-class Ib interactions, including RT1-Cl with the Ly49s4/Ly49i4/Ly49i3 group of receptors, were characterized using overexpressed receptor/ligand pairs, in vitro functional assays, and limited mutational analyses. Obvious, as well as subtle, Ly49-class Ib interactions led to ligand-induced receptor calibration and NK subset expansions in vivo. Together, these studies suggest that in vivo NK alloresponses are controlled by pleomorphic Ly49-class Ib interactions, some of which may not be easily detectable in vitro.

Memory T Cell Proliferation before Hepatitis C Virus Therapy Predicts Antiviral Immune Responses and Treatment Success.

The contribution of the host immune system to the efficacy of new anti-hepatitis C virus (HCV) drugs is unclear. We undertook a longitudinal prospective study of 33 individuals with chronic HCV treated with combination pegylated IFN-α, ribavirin, and telaprevir/boceprevir. We characterized innate and adaptive immune cells to determine whether kinetics of the host response could predict sustained virologic response (SVR). We show that characteristics of the host immune system present before treatment were correlated with successful therapy. Augmentation of adaptive immune responses during therapy was more impressive among those achieving SVR. Most importantly, active memory T cell proliferation before therapy predicted SVR and was associated with the magnitude of the HCV-specific responses at week 12 after treatment start. After therapy initiation, the most important correlate of success was minimal monocyte activation, as predicted by previous in vitro work. In addition, subjects achieving SVR had increasing expression of the transcription factor T-bet, a driver of Th1 differentiation and cytotoxic effector cell maturation. These results show that host immune features present before treatment initiation predict SVR and eventual development of a higher frequency of functional virus-specific cells in blood. Such host characteristics may also be required for successful vaccine-mediated protection.

NKG2D ligand expression in Crohn's disease and NKG2D-dependent stimulation of CD8+ T cell migration.

Interaction between the activating NKG2D receptor on lymphocytes and its ligands MICA, MICB, and ULBP1-6 modulate T and NK cell activity and may contribute to the pathogenesis of Crohn's disease (CD). NKG2D ligands are generally not expressed on the cell surface of normal, non-stressed cells, but expression of MICA and MICB in CD intestine has been reported. In this exploratory study, we further characterize the expression of NKG2D and its ligands, including the less well-described ULBP4-6, in CD, and test if NKG2D ligand interactions are involved in the migration of activated T cells into the affected mucosal compartments. Intestinal tissue from CD patients and healthy controls were analyzed by flow cytometry, mass cytometry, and immunohistochemistry for expression of NKG2D and ligands, and for cytokine release. Furthermore, NKG2D-dependent chemotaxis of activated CD8+ T cells across a monolayer of ligand-expressing human intestinal endothelial cells was examined. Activated lymphocytes down-regulated NKG2D expression upon accumulation in inflamed CD intestine. NKG2D expression on CD56+ T and γδ T cells from inflamed tissue seemed inversely correlated with CRP levels and cytokine release. B cells, monocytes, mucosal epithelium, and vascular endothelium expressed NKG2D ligands in inflamed CD intestine. The expression of NKG2D ligands was correlated with cytokine release, but was highly variable between patients. Stimulation of vascular intestinal endothelial cells in vitro induced expression of NKG2D ligands, including MICA/B and ULBP2/6. Blockade of NKG2D on CD8+ T cells inhibited the migration over ligand-expressing endothelial cells. Intestinal induction of NKG2D ligands and ligand-induced down-regulation of NKG2D in CD suggest that the NKG2D-ligand interaction may be involved in both the activation and recruitment of NKG2D+ lymphocytes into the inflamed CD intestine.

IL-33 promotes growth and liver metastasis of colorectal cancer in mice by remodeling the tumor microenvironment and inducing angiogenesis.

Liver metastasis is the major cause of death from colorectal cancer (CRC). Understanding its mechanisms is necessary for timely diagnosis and development of effective therapies. Interleukin-33 (IL-33) is an IL-1 cytokine family member that uniquely functions as a cytokine and nuclear factor. It is released by necrotic epithelial cells and activated innate immune cells, functioning as an alarmin or an early danger signal. Its role in invoking type 2 immune response has been established; however, it has contrasting roles in tumor development and metastasis. We identified IL-33 as a potently upregulated cytokine in a highly metastatic murine CRC cell line and examined its role in tumor growth and metastasis to the liver. IL-33 was transgenically expressed in murine and human adenocarcinoma and carcinoma cell lines and their growth and spontaneous metastasis to the liver were assessed in orthotopic models of CRC in wild-type C57Bl/6 and Il33 knockout mice. The results showed that increased expression of IL-33 in CRC cells enhanced their tumor take, growth, and liver metastasis. Tumor- rather than host-derived IL-33 induced the enhanced recruitment of CD11b+ GR1+ and CD11b+ F4/80+ myeloid cells to remodel the tumor microenvironment by increased expression of mobilizing cytokines, and tumor angiogenesis by activating endothelial cells. IL-33 expression was elevated in patient tumor tissues, induced early in adenoma development, and activated by pro-inflammatory cytokines derived from the tumor microenvironment. The data suggest that tumor-derived IL-33 modulates the tumor microenvironment to potently promote colon carcinogenesis and liver metastasis, underscoring its potential as a therapeutic target. © 2016 Wiley Periodicals, Inc.

Improved Survival Among all Interferon-α-Treated Patients in HCV-002, a Veterans Affairs Hepatitis C Cohort of 2211 Patients, Despite Increased Cirrhosis Among Nonresponders.

BACKGROUND: As the era of interferon-alpha (IFN)-based therapy for hepatitis C ends, long-term treatment outcomes are now being evaluated. AIM: To more fully understand the natural history of hepatitis C infection by following a multisite cohort of patients. METHODS: Patients with chronic HCV were prospectively enrolled in 1999-2000 from 11 VA medical centers and followed through retrospective medical record review. RESULTS: A total of 2211 patients were followed for an average of 8.5 years after enrollment. Thirty-one percent of patients received HCV antiviral therapy, 15 % with standard IFN/ribavirin only, 16 % with pegylated IFN/ribavirin, and 26.7 % of treated patients achieved sustained virologic response (SVR). Cirrhosis developed in 25.8 % of patients. Treatment nonresponders had a greater than twofold increase in the hazard of cirrhosis and hepatocellular carcinoma, compared to untreated patients, whereas SVR patients were only marginally protected from cirrhosis. Nearly 6 % developed hepatocellular carcinoma, and 27.1 % died during the follow-up period. Treated patients, regardless of response, had a significant survival benefit compared to untreated patients (HR 0.58, CI 0.46-0.72). Improved survival was also associated with college education, younger age, lower levels of alcohol consumption, and longer duration of medical service follow-up-factors typically associated with treatment eligibility. CONCLUSIONS: As more hepatitis C patients are now being assessed for all-oral combination therapy, these results highlight that patient compliance and limiting harmful behaviors contribute a significant proportion of the survival benefit in treated patients and that the long-term clinical benefits of SVR may be less profound than previously reported.

Protective association of colonoscopy against proximal and distal colon cancer and patterns in interval cancer.

BACKGROUND: The protective effect of colonoscopy against proximal colorectal cancer is variable and depends on the detection and complete removal of precancerous polyps. OBJECTIVE: To estimate the efficacy of colonoscopy in a medical center with open-access screening colonoscopy since 1998. DESIGN: Nested case-control study with incidence density sampling. SETTING: University-affiliated Veterans Affairs Medical Center. PATIENTS: Colorectal cancer (CRC) cases and control subjects selected from screening age patients matched by age, gender, and date of first primary care visit. MAIN OUTCOME MEASUREMENT: Colonoscopy preceding the CRC diagnosis date. RESULTS: A total of 20.2% of CRC cases had a colonoscopy in the preceding 10 years compared with 49.0% of control subjects (adjusted odds ratio [aOR], 0.20; 95% confidence interval [CI], 0.11-0.34). Colonoscopy was strongly associated with decreased odds of both distal CRC (aOR, 0.16; 95% CI, 0.07-0.34) and proximal CRC (aOR, 0.26; 95% CI, 0.11-0.58). The fraction of cases attributed to interval cancers was 10.5%. Missed lesions predominantly localized to the cecum and rectum, and recurrent lesions clustered in the hepatic flexure. Cecal intubation rate was 93% (98% in adequately prepped patients), and the adenoma detection rate was 45.2% in the control group. LIMITATIONS: Single-center, retrospective case-control design. CONCLUSION: In an open access colonoscopy program characterized by a high cecal intubation rate and adenoma detection rate, colonoscopy was strongly associated with reduced odds of both distal and proximal CRC. Among interval cancers, missed lesions clustered in the cecum and rectum and recurrent lesions in the hepatic flexure.

Transcriptomic signatures in whole blood of patients who acquire a chronic inflammatory response syndrome (CIRS) following an exposure to the marine toxin ciguatoxin.

BACKGROUND: Ciguatoxins (CTXs) are polyether marine neurotoxins found in multiple reef-fish species and are potent activators of voltage-gated sodium channels. It is estimated that up to 500,000 people annually experience acute ciguatera poisoning from consuming toxic fish and a small percentage of these victims will develop a chronic, multisymptom, multisystem illness, which can last years, termed a Chronic Inflammatory Response Syndrome (CIRS). Symptoms of ciguatera CIRS include fatigue, cognitive deficits, neurologic deficits, pain and sensitivity to light. There are few treatment options for ciguatera CIRS since little is known about its pathophysiology. METHODS: This study characterizes the transcriptional profile in whole blood of 11 patients with ciguatera-induced CIRS and 11 normal controls run in duplicate using Agilent one color whole genome microarrays. Differential expression was determined by using a combination of moderated t-test p-value and fold change (FC). Significant genes were subjected to gene ontology, principal component analysis and SVM classification. Seven significant genes found by microarray were validated by PCR. RESULTS: Using a low stringency (p < 0.05 and FC > 1.4) and a high stringency (p < 0.01 and FC > 1.5) filter, the resulting gene sets of 185 and 55, respectively, showed clear separation of cases and controls by PCA as well as 100% classification accuracy by SVM, indicating that the gene profiles can separate patients from controls. PCR results of 7 genes showed a 95% correlation to microarray data. Several genes identified by microarray are important in wound healing (CD9, CD36, vWF and Factor XIII), adaptive immunity (HLA-DQB1, DQB2, IL18R1 and IL5RA) and innate immunity (GZMK, TOLLIP, SIGIRR and VIPR2), overlapping several areas shown to be disrupted in a mouse model of acute exposure to ciguatoxin. Another area of interest was differential expression of long, non-coding sequences, or lncRNA. CONCLUSIONS: Disruptions of innate and adaptive immune mechanisms were recorded at both the genomic and proteomic level. A disruption in the HLA-T cell receptor axis could indicate HLA haplotype sensitivity for this chronic syndrome, as noted in many autoimmune conditions. Taken together, these indicators of illness provide additional insights into pathophysiology and potential therapies.

Microarray applications to understand the impact of exposure to environmental contaminants in wild dolphins (Tursiops truncatus).

It is increasingly common to monitor the marine environment and establish geographic trends of environmental contamination by measuring contaminant levels in animals from higher trophic levels. The health of an ecosystem is largely reflected in the health of its inhabitants. As an apex predator, the common bottlenose dolphin (Tursiops truncatus) can reflect the health of near shore marine ecosystems, and reflect coastal threats that pose risk to human health, such as legacy contaminants or marine toxins, e.g. polychlorinated biphenyls (PCBs) and brevetoxins. Major advances in the understanding of dolphin biology and the unique adaptations of these animals in response to the marine environment are being made as a result of the development of cell-lines for use in in vitro experiments, the production of monoclonal antibodies to recognize dolphin proteins, the development of dolphin DNA microarrays to measure global gene expression and the sequencing of the dolphin genome. These advances may play a central role in understanding the complex and specialized biology of the dolphin with regard to how this species responds to an array of environmental insults. This work presents the creation, characterization and application of a new molecular tool to better understand the complex and unique biology of the common bottlenose dolphin and its response to environmental stress and infection. A dolphin oligo microarray representing 24,418 unigene sequences was developed and used to analyze blood samples collected from 69 dolphins during capture-release health assessments at five geographic locations (Beaufort, NC, Sarasota Bay, FL, Saint Joseph Bay, FL, Sapelo Island, GA and Brunswick, GA). The microarray was validated and tested for its ability to: 1) distinguish male from female dolphins; 2) differentiate dolphins inhabiting different geographic locations (Atlantic coasts vs the Gulf of Mexico); and 3) study in detail dolphins resident in one site, the Georgia coast, known to be heavily contaminated by Aroclor 1268, an uncommon polychlorinated (PCB) mixture. The microarray was able to distinguish dolphins by sex, geographic location, and corroborate previously published health irregularities for the Georgia dolphins. Genes involved in xenobiotic metabolism, development/differentiation and oncogenic pathways were found to be differentially expressed in GA dolphins. The report bridges the advancements in dolphin genome sequencing to the first step towards providing a cost-effective means to screen for indicators of chemical toxin exposure as well as disease status in top level predators.

Structural brain abnormalities in patients with inflammatory illness acquired following exposure to water-damaged buildings: a volumetric MRI study using NeuroQuant®.

Executive cognitive and neurologic abnormalities are commonly seen in patients with a chronic inflammatory response syndrome (CIRS) acquired following exposure to the interior environment of water-damaged buildings (WDB), but a clear delineation of the physiologic or structural basis for these abnormalities has not been defined. Symptoms of affected patients routinely include headache, difficulty with recent memory, concentration, word finding, numbness, tingling, metallic taste and vertigo. Additionally, persistent proteomic abnormalities in inflammatory parameters that can alter permeability of the blood-brain barrier, such as C4a, TGFB1, MMP9 and VEGF, are notably present in cases of CIRS-WDB compared to controls, suggesting a consequent inflammatory injury to the central nervous system. Findings of gliotic areas in MRI scans in over 45% of CIRS-WDB cases compared to 5% of controls, as well as elevated lactate and depressed ratios of glutamate to glutamine, are regularly seen in MR spectroscopy of cases. This study used the volumetric software program NeuroQuant® (NQ) to determine specific brain structure volumes in consecutive patients (N=17) seen in a medical clinic specializing in inflammatory illness. Each of these patients presented for evaluation of an illness thought to be associated with exposure to WDB, and received an MRI that was evaluated by NQ. When compared to those of a medical control group (N=18), statistically significant differences in brain structure proportions were seen for patients in both hemispheres of two of the eleven brain regions analyzed; atrophy of the caudate nucleus and enlargement of the pallidum. In addition, the left amygdala and right forebrain were also enlarged. These volumetric abnormalities, in conjunction with concurrent abnormalities in inflammatory markers, suggest a model for structural brain injury in "mold illness" based on increased permeability of the blood-brain barrier due to chronic, systemic inflammation.

Machine learning approaches to investigate the impact of PCBs on the transcriptome of the common bottlenose dolphin (Tursiops truncatus).

As top-level predators, common bottlenose dolphins (Tursiops truncatus) are particularly sensitive to chemical and biological contaminants that accumulate and biomagnify in the marine food chain. This work investigates the potential use of microarray technology and gene expression profile analysis to screen common bottlenose dolphins for exposure to environmental contaminants through the immunological and/or endocrine perturbations associated with these agents. A dolphin microarray representing 24,418 unigene sequences was used to analyze blood samples collected from 47 dolphins during capture-release health assessments from five different US coastal locations (Beaufort, NC, Sarasota Bay, FL, Saint Joseph Bay, FL, Sapelo Island, GA and Brunswick, GA). Organohalogen contaminants including pesticides, polychlorinated biphenyl congeners (PCBs) and polybrominated diphenyl ether congeners were determined in blubber biopsy samples from the same animals. A subset of samples (n = 10, males; n = 8, females) with the highest and the lowest measured values of PCBs in their blubber was used as strata to determine the differential gene expression of the exposure extremes through machine learning classification algorithms. A set of genes associated primarily with nuclear and DNA stability, cell division and apoptosis regulation, intra- and extra-cellular traffic, and immune response activation was selected by the algorithm for identifying the two exposure extremes. In order to test the hypothesis that these gene expression patterns reflect PCB exposure, we next investigated the blood transcriptomes of the remaining dolphin samples using machine-learning approaches, including K-nn and Support Vector Machines classifiers. Using the derived gene sets, the algorithms worked very well (100% success rate) at classifying dolphins according to the contaminant load accumulated in their blubber. These results suggest that gene expression profile analysis may provide a valuable means to screen for indicators of chemical exposure.

Expanded use of aggressive therapies improves survival in early and intermediate hepatocellular carcinoma.

BACKGROUND: Despite the increasing annual incidence of hepatocellular carcinoma (HCC) in the USA, now estimated at 2.7 cases per 100 000 population, only a small proportion of patients receive treatment and 5-year survival rates range from 9% to 17%. OBJECTIVES: The present study examines the effects of multimodal treatment on survival in a mixed-stage HCC cohort, focusing on the impact of radical therapy in patients with Barcelona Clinic Liver Cancer (BCLC) stage B disease. METHODS: A retrospective review of the medical records of 254 patients considered for HCC treatment between 2003 and 2011 at a large tertiary referral centre was conducted. RESULTS: A total of 195 (76.8%) patients were treated with a median of two liver-directed interventions. Median survival time was 16 months. In proportional hazards analysis, radiofrequency ablation (RFA) and resection were associated with significantly improved 1- and 5-year survival among patients with BCLC stage 0-A disease. In patients with BCLC stage B disease, RFA conferred a survival benefit at 1 year and resection was associated with significantly improved survival at 5 years. CONCLUSIONS: As one of few studies to track the complete course of sequential HCC therapies, the findings of the present study suggest that HCC patients with intermediate-stage (BCLC stage B) disease may benefit from aggressive interventions not currently included in societal guidelines.

Monocyte activation by interferon α is associated with failure to achieve a sustained virologic response after treatment for hepatitis C virus infection.

BACKGROUND: Interferon α (IFN-α) and ribavirin can induce a sustained virologic response (SVR) in some but not all hepatitis C virus (HCV)-infected patients. The mechanism of effective treatment is unclear. One possibility is that IFN-α differentially improves the functional capacity of classic myeloid dendritic cells (mDCs) by altering expression of surface molecules or cytokines. Others have proposed that antigen-presenting cell activation could be paradoxically detrimental during HCV infection because of the production by monocytes of substances inhibitory or toxic to plasmacytoid dendritic cells. METHODS: We examined responses to in vitro IFN-α treatment of peripheral blood leukocyte samples from a retrospective treatment cohort of nearly 200 HCV-seropositive patients who had undergone antiviral therapy with ribavirin and pegylated IFN. We analyzed the variable responses of antigen-presenting cell subsets to drug. RESULTS: We found that patients achieving SVR were no more likely to have robust mDC activation in response to IFN-α than those who did not achieve SVR. Rather, patients achieving SVR were distinguished by restrained monocyte activation in the presence of IFN-α, a factor that was second in importance only to IL28B genotype in its association with SVR. CONCLUSIONS: These results suggest that interindividual variability in the response of monocytes to IFN-α is an important determinant of treatment success with IFN-α-based regimens.

Transcriptomic responses of juvenile Pacific whiteleg shrimp, Litopenaeus vannamei, to hypoxia and hypercapnic hypoxia.

Estuarine crustaceans are often exposed to low dissolved O2 (hypoxia) accompanied by elevated CO2 (hypercapnia), which lowers water pH. Acclimatory responses to hypoxia have been widely characterized; responses to hypercapnia in combination with hypoxia (hypercapnic hypoxia) are less well known. Here we used oligonucleotide microarrays to characterize changes in global gene expression in the hepatopancreas of Pacific whiteleg shrimp, Litopenaeus vannamei, exposed to hypoxia or hypercapnic hypoxia for 4 or 24 h, compared with time-matched animals held in air-saturated water (normoxia). Unigenes whose expressions were significantly impacted by treatment and/or time were used to build artificial neural networks (ANNs) to identify genes with the greatest sensitivity in pairwise discriminations between treatments at each time point and between times for each treatment. ANN gene sets that discriminated hypoxia or hypercapnic hypoxia from normoxia shared functions of translation, mitochondrial energetics, and cellular defense. GO terms protein modification/phosphorylation/cellular protein metabolism and RNA processing/apoptosis/cell cycling occurred at highest frequency in discriminating hypercapnic hypoxia from hypoxia at 4 and 24 h, respectively. For 75.4% of the annotated ANN genes, exposure to hypercapnic hypoxia for 24 h reduced or reversed the transcriptional response to hypoxia alone. These results suggest that high CO2/low pH may interfere with transcriptionally based acclimation to hypoxia or elicit physiological or biochemical responses that relieve internal hypoxia. Whether these data reflect resilience or sensitivity of L. vannamei in the face of expanding hypoxic zones and rising levels of atmospheric CO2 may be important to understanding the survival of this and other estuarine species.

Nonresponse to interferon-α based treatment for chronic hepatitis C infection is associated with increased hazard of cirrhosis.

BACKGROUND: The long-term consequences of unsuccessful interferon-α based hepatitis C treatment on liver disease progression and survival have not been fully explored. METHODS AND FINDINGS: We performed retrospective analyses to assess long-term clinical outcomes among treated and untreated patients with hepatitis C virus in two independent cohorts from a United States Veterans Affairs Medical Center and a University Teaching Hospital. Eligible patients underwent liver biopsy during consideration for interferon-α based treatment between 1992 and 2007. They were assessed for the probability of developing cirrhosis and of dying during follow-up using Cox proportional hazards models, stratified by pretreatment liver fibrosis stage and adjusted for known risk factors for cirrhosis and characteristics affecting treatment selection. The major predictor was a time-dependent covariate for treatment outcome among four patient groups: 1) patients with sustained virological response to treatment; 2) treatment relapsers; 3) treatment nonresponders; and 4) never treated patients. Treatment nonresponders in both cohorts had a statistically significantly increased hazard of cirrhosis compared to never treated patients, as stratified by pretreatment liver fibrosis stage and adjusted for clinical and psychosocial risk factors that disproportionately affect patients who were ineligible for treatment (Veterans Affairs HR=2.35, CI 1.18-4.69, mean follow-up 10 years, and University Hospital HR=5.90, CI 1.50-23.24, mean follow-up 7.7 years). Despite their increased risk for liver disease progression, the overall survival of nonresponders in both cohorts was not significantly different from that of never treated patients. CONCLUSION: These unexpected findings suggest that patients who receive interferon-α based therapies but fail to clear the hepatitis C virus may have an increased hazard of cirrhosis compared to untreated patients.