Ligand-Independent Activation of Aryl Hydrocarbon Receptor and Attenuation of Glutamine Levels by Natural Deep Eutectic Solvent.

Natural deep eutectic solvents (NADESs) are emerging sustainable alternatives to conventional organic solvents. Beyond their role as laboratory solvents, NADESs are increasingly explored in drug delivery and as therapeutics. Their increasing applications notwithstanding, our understanding of how they interact with biomolecules at multiple levels - metabolome, proteome, and transcriptome - within human cell remain poor. Here, we deploy integrated metabolomics, proteomics, and transcriptomics to probe how NADESs perturb the molecular landscape of human cells. In a human cell line model, we found that an archetypal NADES derived from choline and geranic acid (CAGE) significantly altered the metabolome, proteome, and transcriptome. CAGE upregulated indole-3-lactic acid and 4-hydroxyphenyllactic acid levels, resulting in ligand-independent activation of aryl hydrocarbon receptor to signal the transcription of genes with implications for inflammation, immunomodulation, cell development, and chemical detoxification. Further, treating the cell line with CAGE downregulated glutamine biosynthesis, a nutrient rapidly proliferating cancer cells require. CAGE's ability to attenuate glutamine levels is potentially relevant for cancer treatment. These findings suggest that NADESs, even when derived from natural components like choline, can indirectly modulate cell biology at multiple levels, expanding their applications beyond chemistry to biomedicine and biotechnology.

AdrA as a Potential Immunomodulatory Candidate for STING-Mediated Antiviral Therapy That Required Both Type I IFN and TNF-α Production.

Several dinucleotide cyclases, including cyclic GMP-AMP synthase, and their involvement in STING-mediated immunity have been extensively studied. In this study, we tested five bacterial diguanylate cyclases from the Gram-negative bacterium Salmonella Enteritidis, identifying AdrA as the most potent inducer of a STING-mediated IFN response. AdrA wild-type (wt) or its inactive version AdrA mutant (mut) were delivered by an adenovirus (Ad) vector. Dendritic cells obtained from wt mice and infected in vitro with Ad vector containing AdrA wt, but not mut, had increased activation markers and produced large amounts of several immunostimulatory cytokines. For dendritic cells derived from STING-deficient mice, no activation was detected. The potential antiviral activity of AdrA was addressed in hepatitis B virus (HBV)-transgenic and adenovirus-associated virus (AAV)-HBV mouse models. Viremia in serum of Ad AdrA wt-treated mice was reduced significantly compared with that in Ad AdrA mut-injected mice. The viral load in the liver at sacrifice was in line with this finding. To further elucidate the molecular mechanism(s) by which AdrA confers its antiviral function, the response in mice deficient in STING or its downstream effector molecules was analyzed. wt and IFN-αR (IFNAR)-/- animals were additionally treated with anti-TNF-α (Enbrel). Interestingly, albeit less pronounced than in wt mice, in IFNAR-/- and Enbrel-treated wt mice, a reduction of serum viremia was achieved-an observation that was lost in anti-TNF-α-treated IFNAR-/- animals. No effect of AdrA wt was seen in STING-deficient animals. Thus, although STING is indispensable for the antiviral activity of AdrA, type I IFN and TNF-α are both required and act synergistically.

Effects of supplementation of tropical fruit processing by-products on lipid profile, retinol levels and intestinal function in Wistar rats.

Fruits agro-industrial by-products may have a great variety of bioactive compounds that promote health. Thus, the effects of supplementation with acerola, cashew and guava processing by-products for 28 days on retinol level, lipid profile and on some aspects related to intestinal function in rats were investigated. The animals supplemented with different fruit by-products presented similar weight gain, faecal pH values and intestinal epithelial structures; however, they showed higher moisture and Lactobacillus spp. and Bifidobacterium spp. counts in faeces compared to the control group. Supplementation with the cashew by-product decreased the blood glucose, acerola and guava by-products reduced serum lipid levels and all fruit by-products tested increased serum and hepatic retinol. The results indicated that acerola and guava by-products possess a potential hypolipidemic effect. The three fruit by-products increase the hepatic retinol deposition and the faecal populations of beneficial bacterial groups and modulated aspects of intestinal function. The findings of this study can contribute to sustainable fruticulture and support future clinical studies with the supplementation of by-products.

An outlook on fluorescent in situ hybridization coupled to flow cytometry as a versatile technique to evaluate the effects of foods and dietary interventions on gut microbiota.

The increasing interest in the effects of the gut microbiota on host health has stimulated the investigation of the composition of this microbial community and the factors affecting these microorganisms. This review discusses the recent advances and progress applications in the use of the fluorescent in situ hybridization (FISH) coupled to flow cytometry (FC) technique (FISH-FC) in studies evaluating the gut microbiota published in the last 10 years, with particular emphasis on the effects of foods and dietary interventions. These studies have shown that FISH-FC technique is capable of detecting and quantifying several groups of bacteria found as part of the gut microbiota. FISH-FC can be considered an effective, versatile, and rapid technique to evaluate alterations in gut microbiota composition caused by different foods as assessed in studies in vitro, in vivo, and in clinical trials. Some specific probes have been most used to represent the general gut microbiota, such as those specific to Lactobacillus spp./Enterococcus spp., Bacteroidaceae/Prevotellaceae, Clostridium histolyticum, and Bifidobacterium spp. FISH-FC technique could have an important opportunity for application in studies with next-generation probiotics belonging to the gut microbiota. Optimizations of FISH-FC protocols could allow more discoveries about the gut microbiota, including the development of new probes targeting microorganisms still not explored, the analysis of individual portions of the intestine, and the proposition of novel quantitative approaches.

Autochthonous adjunct culture of Limosilactobacillus mucosae CNPC007 improved the techno-functional, physicochemical, and sensory properties of goat milk Greek-style yogurt.

We evaluated the performance of Limosilactobacillus mucosae CNPC007 as an autochthonous adjunct culture in the production of goat milk Greek-style yogurt. The techno-functional, physicochemical, and sensory characteristics of the control yogurt (containing only starter culture, CY) and the probiotic yogurt (with the probiotic strain added, PY) were assessed during 28 d of refrigerated storage. Furthermore, we determined the survival of the strain throughout the gastrointestinal tract under simulated conditions. The PY yogurt had a lower extent of proteolysis index and a higher depth of proteolysis index. These results indicate that the proteolytic enzymes of L. mucosae may have a possible action in PY. The PY formulation exhibited viscosity almost 1.5 times as high as CY over the refrigeration period, probably due to higher production of exopolysaccharides by the probiotic strain, which directly interferes with the microstructure, texture, and viscosity of the product. The PY formulation received higher scores for color, flavor, and global acceptance at 1 d of storage and higher texture scores at 28 d. The counts of L. mucosae remained high (>7 log cfu/g and >8.5 log cfu/g) throughout mouth-ileum digestion and storage, respectively, in PY. The autochthonous adjunct culture of L. mucosae CNPC007 can be used for production of a novel potentially probiotic goat yogurt without negatively affecting the general characteristics of the product quality, adding value associated with maintaining its functional potential.

High SOX9 Maintains Glioma Stem Cell Activity through a Regulatory Loop Involving STAT3 and PML.

Glioma stem cells (GSCs) are critical targets for glioma therapy. SOX9 is a transcription factor with critical roles during neurodevelopment, particularly within neural stem cells. Previous studies showed that high levels of SOX9 are associated with poor glioma patient survival. SOX9 knockdown impairs GSCs proliferation, confirming its potential as a target for glioma therapy. In this study, we characterized the function of SOX9 directly in patient-derived glioma stem cells. Notably, transcriptome analysis of GSCs with SOX9 knockdown revealed STAT3 and PML as downstream targets. Functional studies demonstrated that SOX9, STAT3, and PML form a regulatory loop that is key for GSC activity and self-renewal. Analysis of glioma clinical biopsies confirmed a positive correlation between SOX9/STAT3/PML and poor patient survival among the cases with the highest SOX9 expression levels. Importantly, direct STAT3 or PML inhibitors reduced the expression of SOX9, STAT3, and PML proteins, which significantly reduced GSCs tumorigenicity. In summary, our study reveals a novel role for SOX9 upstream of STAT3, as a GSC pathway regulator, and presents pharmacological inhibitors of the signaling cascade.

The influence of spirituality and religion on critical care nursing: An integrative review.

BACKGROUND: Spiritual care could help family members and critically ill patients to cope with anxiety, stress and depression. However, health care professionals are poorly prepared and health managers are not allocating all the resources needed. AIMS AND OBJECTIVES: To critically review the empirical evidence concerning the influence of spirituality and religion (S-R) on critical care nursing. METHODS: An integrative review of the literature published in the last 10 years (2010-2019) was conducted in PubMed, Scopus, CINHAL, PsycINFO, Web of Science, Cochrane and LILACS. In addition, searches were performed in the System for Information on Grey Literature in Europe and the Grey Literature Report. Quantitative and/or qualitative studies, assessing S-R and including health care professionals caring for critically ill patients (i.e. adults or children), were included. RESULTS: Forty articles were included in the final analysis (20 qualitative, 19 quantitative and 1 with a mixed methodology). The studies embraced the following themes: S-R importance and the use of coping among critical care patients and families; spiritual needs of patients and families; health care professionals' awareness of spiritual needs; ways to address spiritual care in the intensive care unit (ICU); definition of S-R by health care professionals; perceptions and barriers of addressing spiritual needs; and influence of S-R on health care professionals' outcomes and decisions. Our results indicate that patients and their families use S-R coping strategies to alleviate stressful situations in the ICU and that respecting patients' spiritual beliefs is an essential component of critical care. Although nurses consider spiritual care to be very important, they do not feel prepared to address S-R and report lack of time as the main barrier. CONCLUSION AND IMPLICATIONS FOR PRACTICE: Critical care professionals should be aware about the needs of their patients and should be trained to handle S-R in clinical practice. Nurses are encouraged to increase their knowledge and awareness towards spiritual issues.

Dynamics of physiological responses of potentially probiotic fruit-derived Limosilactobacillus fermentum in apple and orange juices during refrigeration storage and exposure to simulated gastrointestinal conditions.

This study evaluated the dynamics of the physiological responses of potentially probiotic fruit-derived Limosilactobacillus fermentum 139 and L. fermentum 263 in apple and orange juice during 28 days of refrigeration storage (4 °C) and when submitted to simulated gastrointestinal conditions. Physiological responses were measured with multiparametric flow cytometry using propidium iodide (PI), carboxyfluorescein diacetate (cFDA) and bis-1,3-dibutylbarbutiric acid (BOX). Viable counts were enumerated with plate count. L. fermentum strains had sizes of > 30% of cell subpopulations with non-permeabilized membrane and enzymatic activities (viable cells, PI-CFDA +) in apple and orange juices during storage and viable counts of > 6 log CFU ml-1. Sizes of cell subpopulations with permeabilized membrane without enzymatic activity (dead cells, PI + cFDA-) were low (< 15%) in apple and orange juices during storage. Sizes of cell subpopulations with non-permeabilized and depolarized membrane (PI-BOX +) were decreased (14%) on day 28 of storage. The sizes of permeabilized and depolarized membrane cell (PI + BOX-) subpopulations were variable among the examined strains in juices during storage. Both strains maintained high PI-cFDA + cell subpopulation sizes (> 35%) after exposure to ileum condition and viable counts of ≥ 5 log CFU/mL. PI-BOX + cell subpopulation sizes were low (< 13%) after exposure to ileum condition. L. fermentum 139 and L. fermentum 263 are capable of maintaining a high population of physiologically active and functional cells in apple and orange juice during 28 days of refrigeration storage and when exposed to gastrointestinal conditions.

Defibrotide in hematopoietic stem cell transplantation: A multicenter survey study of the Spanish Hematopoietic Stem Cell Transplantation Group (GETH).

BACKGROUND: Defibrotide is approved in European Union for the treatment of severe sinusoidal obstruction syndrome (SOS) after HSCT. However, it has also been used for SOS prophylaxis, moderate SOS and in other complications such as transplant-associated thrombotic microangiopathy (TAM). The objective of this study was to evaluate current uses, effectiveness and safety of defibrotide in patients with HSCT. METHODS: This multicenter, retrospective study included patients treated with defibrotide for any indication at 28 HSCT centers of the Grupo Español de Trasplante Hematopoyetico (GETH) including the pediatric subgroup Grupo Español de Trasplante de Medula en Niños (GETMON). RESULTS: Three hundred and eighty eight patients treated with defibrotide between January 2011 and December 2018 were included. 253 patients were children, and 135 patients were adults. In total, 332 transplants were allogeneic, and the remainder were autologous. Main indications for defibrotide use were severe/very severe SOS in 173 patients, SOS prophylaxis in 135 patients, moderate SOS in 41 patients, TAM in six patients and suspected SOS in 33 patients. Overall survival (OS) at day +100 in the SOS prophylaxis group was 89% (95% CI, 87%-91%). In the group of patients with moderate and severe/very severe SOS, the OS at day +100 was 80% (95% CI, 74%-86%) and 62% (95% CI, 59%-65%), respectively (P = .0015). With a longer follow-up, median of 2 years (4 months-7 years), OS was 63% (95% CI, 59%-67%) in the SOS prophylaxis patients. OS for patients with moderate and severe/very severe SOS groups was 53% (95% CI, 47%-61%) and 26% (95% CI, 22%-30%), respectively (P = .006). 191 patients died, and SOS was the main cause of death in 23 patients (12%). CONCLUSIONS: Defibrotide has an acceptable safety profile with an improved response in severe/very severe SOS compared with historical controls, mainly in pediatric patients. Use of defibrotide for prophylaxis may improve prognosis of patients at high risk of complications due to endothelial damage such as those who receive a second transplant. SOS has an important impact on the HSCT long-term survival, as can be concluded from our study.

Influence of Upper Airway Surgery on Voice and Speech Recognition.

PURPOSE: Upper airway surgery comprises a set of techniques that modify the anatomy of the vocal tract, including tonsillectomy and septoplasty. The objective of this work is to study the changes in acoustic parameters and the effects on the identification or verification of the speaker through the speech produced after the vocal tract surgeries, comparing them with a control group. METHODS: A prospective study was performed between January 2019 and June 2019 including. The final study sample consisted of 84 patients who met the inclusion criteria. Of these, 31 underwent septoplasty, 26 tonsillectomy patients, and 27 controls. Demographic data and GRBAS evaluation were statistically evaluated. Tests were taken before surgery, 2 weeks after surgery and 3 months later. Furthermore, to establish the equal error rate, the recording of patients' voices was made with a succeeding acoustic analysis and programmed identification of the speaker through machine learning systems. RESULTS: A significant variance was observed in GRBAS, after surgery. Regarding acoustic parameters, a greater change was observed in the fundamental frequency at 2 weeks after surgery in the tonsillectomy group. Formants (F1-F3) and antiformants (AntiF1-AntiF3) changed in septoplasty group, not in tonsillectomy and control group at 3 months. When studying the impact of voice changes on the verification of the speaker through the speech, it was observed that there was a greater error in recognition in the tonsillectomy group at 2 weeks, coinciding with the results obtained in the rest of the parameters studied. CONCLUSIONS: Results suggest that upper airway surgery produces modifications in the vocal tract affecting GRBAS, acoustic parameters, including formants and antiformants, producing an effect on verification of the speaker through the speech.

Self-Efficacy, Pain Intensity, Rheumatic Disease Duration, and Hand Functional Disability on Activities of Daily Living.

BACKGROUND: Knowledge regarding the possible influence of self-efficacy, pain intensity and disease duration on hand functional disability could promote new intervention strategies for activities of daily living (ADLs) in patients with rheumatic disease (RD). These approaches could prevent the health problems and socioeconomic costs associated with these diseases. OBJECTIVE: The aims of this study were to evaluate if there are differences between the levels of perceived self-efficacy, pain intensity and disease duration among people with RD and non-RD diseases, and to analyze if hand functional disability in ADLs is related to self-efficacy, pain intensity and disease duration in a sample of patients with RD. METHODS: A multicenter, observational, cross-sectional study was conducted on a total sample of 335 participants over 50 years old (176 patients with RD and 159 individuals with non-RD). The Duruöz Hand Index, the General Perceived Self-Efficacy Scale, the Rheumatic Diseases Self-Efficacy Scale (RDS-ES), the Visual Analog Scale (VAS), and the mean time of evolution in years of the disease (disease duration) were used to analyze the possible relationships surrounding hand functional disability in ADLs. RESULTS: The comparison analysis showed significant differences between the RD/non-RD sample for the General Perceived Self-Efficacy Scale, RDS-ES, and VAS scores (p < .001). The multiple regression results showed that age, General Perceived Self-Efficacy Scale scores, RDS-ES scores, VAS scores, and disease duration (or a combination of some of them) explained the variability of hand functional disability in almost 68% of kitchen tasks, 44% of dressing tasks, 46% of hygiene and other tasks, and 47% of office tasks. DISCUSSION: Our study shows that general and domain-specific self-efficacy, pain intensity, and disease duration are predictors of the dimensions of hand functional disability in patients with RD. Early evaluation of these components with an interdisciplinary approach would help to manage hand disability properly.

A Microservices e-Health System for Ecological Frailty Assessment Using Wearables.

The population in developed countries is aging and this fact results in high elderly health costs, as well as a decrease in the number of active working members to support these costs. This could lead to a collapse of the current systems. One of the first insights of the decline in elderly people is frailty, which could be decelerated if it is detected at an early stage. Nowadays, health professionals measure frailty manually through questionnaires and tests of strength or gait focused on the physical dimension. Sensors are increasingly used to measure and monitor different e-health indicators while the user is performing Basic Activities of Daily Life (BADL). In this paper, we present a system based on microservices architecture, which collects sensory data while the older adults perform Instrumental ADLs (IADLs) in combination with BADLs. IADLs involve physical dimension, but also cognitive and social dimensions. With the sensory data we built a machine learning model to assess frailty status which outperforms the previous works that only used BADLs. Our model is accurate, ecological, non-intrusive, flexible and can help health professionals to automatically detect frailty.

Helicobacter pylori pathogenicity and primary antimicrobial resistance in Northern Spain.

BACKGROUND: Helicobacter pylori infection is associated with chronic gastritis, ulcers and gastric cancer. Antimicrobial resistance has increased worldwide affecting the efficacy of current treatments. Most guidelines recommend implementation of regional surveillance of primary antibiotic resistance of H pylori. Only a fraction of individuals infected with H pylori develop gastric diseases which are related to virulence factors of the bacteria. The aims of the study were to determine the primary antimicrobial resistance rates of H pylori and to know the virulence factors prevalence of strains circulating in Southern Europe. MATERIALS AND METHODS: Susceptibility testing by Etest to clarithromycin, levofloxacin, metronidazole, amoxicillin and tetracycline was performed in 102 isolates (99 naïve patients). The prevalence of virulence factors (cagA, vacA, oipA, babA and dupA) was evaluated in 102 H pylori isolates from patients with mild-disease symptoms and in 22 isolates from patients with severe-disease symptoms. RESULTS: Primary resistance rates were 12.1% to clarithromycin, 13.1% to levofloxacin, 24.2% to metronidazole and 0% to amoxicillin and tetracycline. Combined resistance to clarithromycin and levofloxacin was 3% and to clarithromycin and metronidazole 4%. Prevalence of virulence factors in the mild- and severe-disease group was 35.3% and 81.8% for cagA, 20.6% and 54.5% for cagA/vacAs1m1, 94.1% and 95.4% for babA2, 78.4% and 100% for oipA and 30.4% and 18.2% for dupA. CONCLUSIONS: Primary antimicrobial resistance rates were under 15% for clarithromycin and levofloxacin. The prevalence of H pylori carrying the virulent genotype cagA/vacAs1m1 was higher than 20% in the mild-disease and 54% in the severe-disease symptom group.

Intrathecal cell therapy with autologous stromal cells increases cerebral glucose metabolism and can offer a new approach to the treatment of Alzheimer's type dementia.

BACKGROUND AIMS: After recent observations that intrathecal administration of autologous bone marrow mesenchymal stromal cells (MSCs) increases cerebral metabolism in patients with severe traumatic brain injury (TBI), we examined this type of cell therapy in Alzheimer's type dementia. METHODS: Three patients with clinical diagnosis of Alzheimer's disease received every 3 months 100million autologous MSCs by intrathecal route, until a total dose of 300million. RESULTS: During cell therapy the patients showed arrest in neurological deterioration and two of them manifested clear improvement of previous symptoms. A global increase in cerebral glucose metabolism, measured using 18F-fluorodeoxyglucose positron emission tomography (18F-FDG-PET), was observed after every administration of cell therapy. CONCLUSIONS: Our present findings suggest that intrathecal administrations of autologous MSCs can be a new strategy for the treatment of Alzheimer's dementia.

Frequency, characteristics, and outcome of PTLD after allo-SCT: A multicenter study from the Spanish group of blood and marrow transplantation (GETH).

Post-transplant lymphoproliferative disorder (PTLD) is an infrequent complication of allogeneic stem cell transplant (allo-SCT). AIMS: To estimate the frequency and management of PTLD in Spain and to identify prognostic factors influencing outcomes. METHODS: Multicenter, retrospective analysis of allo-SCT performed in 14 transplant units over a 15-year period. RESULTS: 102 PTLD were diagnosed among 12 641 allo-SCT, leading to an estimated frequency of 0.8%. PTLD was diagnosed at a median of 106 days after SCT. Eighty-seven cases (85%) were diagnosed between 2007 and 2013. At diagnosis, 22% and 17% of the patients had gastrointestinal tract and CNS involvement. Eighty-seven (85%) received rituximab treatment, alone or in combination with immunosuppression reduction, with an ORR of 50.6%. With a median follow-up for survivors of 58 months, the 2-year overall survival (OS) was 33% and the PTLD-related mortality 45%. Age ≥ 40 years, malignant underlying disease, non-response to rituximab, and severe thrombocytopenia or lymphocytopenia at PTLD diagnosis were associated with worse overall survival. CONCLUSIONS: Only a small proportion of allografted patients were diagnosed a PTLD. Its clinical course was highly aggressive, and prognosis poor, especially in those failing rituximab. The prognostic impact found of the platelet, and lymphocyte count at diagnosis requires further confirmation.

Validation of a multivariable prediction model for post-engraftment invasive fungal disease in 465 adult allogeneic hematopoietic stem cell transplant recipients.

BACKGROUND: Recently, we reported a simple prognostic score for post-engraftment invasive fungal disease (IFD) obtained in 404 adult allogeneic hematopoietic stem cell transplant (alloSCT) (training cohort). OBJECTIVES: We aim to validate this score in an external cohort assessing the 1-year cumulative incidence (CI) of post-engraftment IFD. Additionally, we analyse the type of IFD and incidence of IFD according to type of prophylaxis. PATIENTS/METHODS: We included 465 consecutive adult recipients surviving >40 days who engrafted and were discharged without prior IFD (median age 45 years, range, 14-69). RESULTS: Patients classified as low-risk, 139; intermediate-risk, 162; and high-risk, 164 (35% vs 27% in the training cohort, P = 0.03). The CI of probable/proven IFD in the validation cohort was 8% vs 11% in the training cohort (P = 0.006). The only voriconazole prophylaxis used in the training cohort was 100 mg/12 h, 65% vs 27% in the validation cohort, but 38% received 200 mg/12 h. Thus, the validation cohort showed a lower CI of IFD (P = 0.009). The post-engraftment IFD score was validated, showing a CI of IFD for low-, intermediate- and high-risk of 3%, 6% and 14%, respectively (P < 0.001). CONCLUSION: To our knowledge, this is the first prognostic index to predict the occurrence of post-engraftment IFD after alloSCT that has been validated in an external cohort.

Effect of Septoplasty On Voice Performance: Nasalance and Acoustic Study.

BACKGROUND: Septoplasty is a surgical technique for the correction of the nasal septum that may alter the vocal tract. The aim of our study is to assess whether this technique modifies nasalance and acoustic parameters, and their clinical implications in voice perception. METHODOLOGY: A prospective study was performed between January 2017 and June 2017 including 2 groups of patients: those undergoing septoplasty, and a control group. Subjective nasality questionnaire, objective nasalance with nasometer, and GRBAS (Grade, Roughness, Breathiness, Asthenia, Strain) assessment were statistically analysed. In addition, a recording of patients' voices was made with a subsequent acoustic analysis. Samples were taken: pre-surgically, 2 weeks after surgery and after 3 months. RESULTS: After septoplasty, a significant difference was observed in GRBAS, nasality questionnaire and nasometer nasalance, when compared with the control group. As for the acoustic analysis, no differences were observed in most parameters (F0, Jitter, Shimmer, HNR, NHR, Formants F1-F3), except for the antiF3 antiformant, which showed significant changes in all the vowels studied. CONCLUSIONS: Septoplasty can produce changes in the vocal tract, with an increase in initial nasalance but with subsequent normalization. Besides, minor changes were found in the acoustic analysis but with no clinical relevance.

Transient Expression of Transgenic IL-12 in Mouse Liver Triggers Unremitting Inflammation Mimicking Human Autoimmune Hepatitis.

The etiopathogenesis of autoimmune hepatitis (AIH) remains poorly understood. In this study, we sought to develop an animal model of human AIH to gain insight into the immunological mechanisms driving this condition. C57BL/6 mice were i.v. injected with adeno-associated viral vectors encoding murine IL-12 or luciferase under the control of a liver-specific promoter. Organ histology, response to immunosuppressive therapy, and biochemical and immunological parameters, including Ag-specific humoral and cellular response, were analyzed. Mechanistic studies were carried out using genetically modified mice and depletion of lymphocyte subpopulations. Adeno-associated virus IL-12-treated mice developed histological, biochemical, and immunological changes resembling type 1 AIH, including marked and persistent liver mononuclear cell infiltration, hepatic fibrosis, hypergammaglobulinemia, anti-nuclear and anti-smooth muscle actin Abs, and disease remission with immunosuppressive drugs. Interestingly, transgenic IL-12 was short-lived, but endogenous IL-12 expression was induced, and both IL-12 and IFN-γ remained elevated during the entire study period. IFN-γ was identified as an essential mediator of liver damage, and CD4 and CD8 T cells but not NK, NKT, or B cells were essential executors of hepatic injury. Furthermore, both MHC class I and MHC class II expression was upregulated at the hepatocellular membrane, and induction of autoreactive liver-specific T cells was detected. Remarkably, although immunoregulatory mechanisms were activated, they only partially mitigated liver damage. Thus, low and transient expression of transgenic IL-12 in hepatocytes causes loss of tolerance to hepatocellular Ags, leading to chronic hepatitis resembling human AIH type 1. This model provides a practical tool to explore AIH pathogenesis and novel therapies.

Successful Repeated Hepatic Gene Delivery in Mice and Non-human Primates Achieved by Sequential Administration of AAV5ch and AAV1.

In the gene therapy field, re-administration of adeno-associated virus (AAV) is an important topic because a decrease in therapeutic protein expression might occur over time. However, an efficient re-administration with the same AAV serotype is impossible due to serotype-specific, anti-AAV neutralizing antibodies (NABs) that are produced after initial AAV treatment. To address this issue, we explored the feasibility of using chimeric AAV serotype 5 (AAV5ch) and AAV1 for repeated liver-targeted gene delivery. To develop a relevant model, we immunized animals with a high dose of AAV5ch-human secreted embryonic alkaline phosphatase (hSEAP) that generates high levels of anti-AAV5ch NAB. Secondary liver transduction with the same dose of AAV1-human factor IX (hFIX) in the presence of high levels of anti-AAV5ch NAB proved to be successful because expression/activity of both reporter transgenes was observed. This is the first time that two different transgenes are shown to be produced by non-human primate (NHP) liver after sequential administration of clinically relevant doses of both AAV5ch and AAV1. The levels of transgene proteins achieved after delivery with AAV5ch and AAV1 illustrate the possibility of both serotypes for liver targeting. Furthermore, transgene DNA and RNA biodistribution patterns provided insight into the potential cause of decrease or loss of transgene protein expression over time in NHPs.

The performance of five fruit-derived and freeze-dried potentially probiotic Lactobacillus strains in apple, orange, and grape juices.

BACKGROUND: This study assessed the survival of the fruit-derived and freeze-dried L. plantarum 49, L. brevis 59, L. paracasei 108, L. fermentum 111 and L. pentosus 129 strains during frozen storage and when incorporated into apple, orange, and grape juice stored under refrigeration. Physicochemical parameters of juices containing the freeze-dried Lactobacillus strains and the survival of the test strains in the fruit juices during in vitro digestion were also evaluated. RESULTS: No decreases in survival rates (log N/log N0) of the freeze-dried cells were observed in up to 1 month of storage. The survival rates of the freeze-dried strains L. plantarum 49 and L. paracasei 108 were > 0.75 in up to 4 months of storage. All freeze-dried strains exhibited survival rates of >0.75 in up to 2 weeks of storage in apple juice; only L. plantarum 49 and L. paracasei 108 showed similar survival rates in orange and grape juices in up to 2 weeks of storage. The contents of the monitored organic acids or sugars during storage varied depending on the added strain and the type of fruit juice. At the end of in vitro digestion, L. brevis 59, L. paracasei 108 and L. fermentum 111 showed survival rates of >0.80 in apple juice. CONCLUSION: Apple juice was the best substrate for the survival of the tested freeze-dried Lactobacillus strains over time. L. paracasei 108 and L. plantarum 49 were the strains presenting the best performance for incorporation in potentially probiotic fruit juices. © 2018 Society of Chemical Industry.