Ductular Reaction Cells Display an Inflammatory Profile and Recruit Neutrophils in Alcoholic Hepatitis.

Chronic liver diseases are characterized by the expansion of ductular reaction (DR) cells and the expression of liver progenitor cell (LPC) markers. In alcoholic hepatitis (AH), the degree of DR expansion correlates with disease progression and short-term survival. However, little is known about the biological properties of DR cells, their impact on the pathogenesis of human liver disease, and their contribution to tissue repair. In this study, we have evaluated the transcriptomic profile of DR cells by laser capture microdissection in patients with AH and assessed its association with disease progression. The transcriptome analysis of cytokeratin 7-positive (KRT7+ ) DR cells uncovered intrinsic gene pathways expressed in DR and genes associated with alcoholic liver disease progression. Importantly, DR presented a proinflammatory profile with expression of neutrophil recruiting C-X-C motif chemokine ligand (CXC) and C-C motif chemokine ligand chemokines. Moreover, LPC markers correlated with liver expression and circulating levels of inflammatory mediators such as CXCL5. Histologically, DR was associated with neutrophil infiltration at the periportal area. In order to model the DR and to assess its functional role, we generated LPC organoids derived from patients with cirrhosis. Liver organoids mimicked the transcriptomic and proinflammatory profile of DR cells. Conditioned medium from organoids induced neutrophil migration and enhanced cytokine expression in neutrophils. Likewise, neutrophils promoted the proinflammatory profile and the expression of chemokines of liver organoids. Conclusion: Transcriptomic and functional analysis of KRT7+ cells indicate that DR has a proinflammatory profile and promote neutrophil recruitment. These results indicate that DR may be involved in the liver inflammatory response in AH, and suggest that therapeutic strategies targeting DR cells may be useful to mitigate the inflammatory cell recruitment in AH.

Expression of microRNA-155 in inflammatory cells modulates liver injury.

MicroRNA 155 (miR-155) is involved in immune and inflammatory diseases and is associated with liver fibrosis and steatohepatitis. However, the mechanisms involved in miR-155 regulation of liver injury are largely unknown. The role of miR-155 in acute liver injury was assessed in wild-type (WT), miR-155-/- , and miR-155-/- mice transplanted with WT bone marrow. Additionally, miR-155 expression was evaluated in liver tissue and peripheral blood mononuclear cells of patients with autoimmune hepatitis. Concanavalin A, but not acetaminophen, treatment increased the expression of miR-155 in liver tissue of WT mice. Concanavalin A induced increases in cell death, liver aminotransferases, and expression of proinflammatory cytokines (chemokine [C-X-C motif] ligands 1, 5, 9, 10, and 11; chemokine [C-C motif] ligands 2 and 20; and intercellular cell adhesion molecule 1) in miR-155-/- compared to WT mice. Importantly, these animals showed a significant decrease in cluster of differentiation 4-positive/chemokine (C-X-C motif) receptor 3-positive and forkhead box p3-positive cell recruitment but no changes in other inflammatory cell populations. Mechanistically, miR-155-deficient regulatory T cells showed increased SH2 domain-containing inositol 5-phosphatase 1 expression, a known target of miR-155. Inhibition of SH2 domain-containing inositol 5-phosphatase 1 in miR-155-/- mice restored forkhead box p3 recruitment and reduced liver cytokine expression. Transplantation of bone marrow from WT animals into miR-155-/- mice partially reversed the effect of concanavalin A on miR-155-/- mice as assessed by proinflammatory cytokines and cell death protein expression. Patients with autoimmune hepatitis showed a marked increase in miR-155 expression in the liver but reduced expression of miR-155 in peripheral blood mononuclear cells. CONCLUSION: miR-155 expression is altered in both liver tissue and circulating inflammatory cells during liver injury, thus regulating inflammatory cell recruitment and liver damage; these results suggest that maintaining miR-155 expression in inflammatory cells might be a potential strategy to modulate liver injury. (Hepatology 2018).

Pentraxin-3 modulates lipopolysaccharide-induced inflammatory response and attenuates liver injury.

Acute-on-chronic liver injury is characterized by an important inflammatory response frequently associated with endotoxemia. In this context, acute-phase proteins such as Pentraxin-3 (PTX3) are released; however, little is known about their role in chronic liver disease. The aim of this study was to elucidate the role of PTX3 in liver injury. The role of PTX3 was evaluated in cultured human cells, liver tissue slices, and mice with acute-on-chronic liver injury. PTX3 expression was assessed in tissue and serum samples from 54 patients with alcoholic hepatitis. PTX3 expression was up-regulated in animal models of liver injury and strongly induced by lipopolysaccharide (LPS). Liver cell fractionation showed that macrophages and activated hepatic stellate cells were the main cell types expressing PTX3 in liver injury. Ex vivo and in vivo studies showed that PTX3 treatment attenuated LPS-induced liver injury, inflammation, and cell recruitment. Mechanistically, PTX3 mediated the hepatic stellate cell wound-healing response. Moreover, PTX3 modulated LPS-induced inflammation in human primary liver macrophages and peripheral monocytes by enhancing a TIR domain-containing adapter-inducing interferon-dependent response and favoring a macrophage interleukin-10-like phenotype. Additionally, hepatic and plasma PTX3 levels were increased in patients with alcoholic hepatitis, a prototypic acute-on-chronic condition; and its expression correlated with disease severity scores, endotoxemia, infections, and short-term mortality, thus suggesting that expression of PTX3 found in patients could be a counterregulatory response to injury. CONCLUSION: Experimental and human evidence suggests that, in addition to being a potential biomarker for alcoholic hepatitis, PTX3 participates in the wound-healing response and attenuates LPS-induced liver injury and inflammation; therefore, administration of PTX3 could be a promising therapeutic strategy in acute-on-chronic conditions, particularly those associated with endotoxemia. (Hepatology 2017;66:953-968).

Biomimetic photonics: jamming avoidance system in Eigenmannia.

Biomimetic photonics extract the good design of nature and mimic it with photonics. The weakly electric fish genus, Eigenmannia, has a unique neural algorithm - jamming avoidance response, to facilitate their survival in the deep dark ocean, by automatically adjusts the local transmitter carrier frequency to move away from the jamming frequency when it is within the jamming spectral range. Examining our own wireless microwave systems, the situation of inadvertent jamming is very similar as that in Eigenmannia. In this article, a biomimetic photonic approach inspired by the jamming avoidance response in a weakly electric fish genus, Eigenmannia, is naturally adopted to experimentally tackle signal jamming in wireless systems. Mimicking the system with photonics enables the proposed scheme to work for frequencies from hundreds of MHz to tens of GHz.

Integrative microRNA profiling in alcoholic hepatitis reveals a role for microRNA-182 in liver injury and inflammation.

OBJECTIVE: MicroRNAs (miRNAs) are well-known regulators of disease pathogenesis and have great potential as biomarkers and therapeutic targets. We aimed at profiling miRNAs in alcoholic hepatitis (AH) and identifying miRNAs potentially involved in liver injury. DESIGN: MiRNA profiling was performed in liver samples from patients with AH, alcohol liver disease, non-alcoholic steatohepatitis, HCV disease and normal liver tissue. Expression of miRNAs was assessed in liver and serum from patients with AH and animal models. Mimic and decoy miR-182 were used in vitro and in vivo to evaluate miR-182's biological functions. RESULTS: MiRNA expression profile in liver was highly altered in AH and distinctive from alcohol-induced cirrhotic livers. Moreover, we identified a set of 18 miRNAs predominantly expressed in AH as compared with other chronic liver conditions. Integrative miRNA-mRNA functional analysis revealed the association of AH-altered miRNAs with nuclear receptors, IGF-1 signalling and cholestasis. Interestingly, miR-182 was the most highly expressed miRNA in AH, which correlated with degree of ductular reaction, disease severity and short-term mortality. MiR-182 mimic induced an upregulation of inflammatory mediators in biliary cells. At experimental level, miR-182 was increased in biliary cells in mice fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet but not upregulated by alcohol intake or fibrosis. Inhibition of miR-182 in DDC-fed mice reduced liver damage, bile acid accumulation and inflammatory response. CONCLUSIONS: AH is characterised by a deregulated miRNA profile, including miR-182, which is associated with disease severity and liver injury. These results highlight the potential of miRNAs as therapeutic targets and biomarkers in AH.

Kinase analysis in alcoholic hepatitis identifies p90RSK as a potential mediator of liver fibrogenesis.

OBJECTIVE: Alcoholic hepatitis (AH) is often associated with advanced fibrosis, which negatively impacts survival. We aimed at identifying kinases deregulated in livers from patients with AH and advanced fibrosis in order to discover novel molecular targets. DESIGN: Extensive phosphoprotein analysis by reverse phase protein microarrays was performed in AH (n=12) and normal human livers (n=7). Ribosomal S6 kinase (p90RSK) hepatic expression was assessed by qPCR, Western blot and immunohistochemistry. Kaempferol was used as a selective pharmacological inhibitor of the p90RSK pathway to assess the regulation of experimentally-induced liver fibrosis and injury, using in vivo and in vitro approaches. RESULTS: Proteomic analysis identified p90RSK as one of the most deregulated kinases in AH. Hepatic p90RSK gene and protein expression was also upregulated in livers with chronic liver disease. Immunohistochemistry studies showed increased p90RSK staining in areas of active fibrogenesis in cirrhotic livers. Therapeutic administration of kaempferol to carbon tetrachloride-treated mice resulted in decreased hepatic collagen deposition, and expression of profibrogenic and proinflammatory genes, compared to vehicle administration. In addition, kaempferol reduced the extent of hepatocellular injury and degree of apoptosis. In primary hepatic stellate cells, kaempferol and small interfering RNA decreased activation of p90RSK, which in turn regulated key profibrogenic actions. In primary hepatocytes, kaempferol attenuated proapoptotic signalling. CONCLUSIONS: p90RSK is upregulated in patients with chronic liver disease and mediates liver fibrogenesis in vivo and in vitro. These results suggest that the p90RSK pathway could be a new therapeutic approach for liver diseases characterised by advanced fibrosis.

Evaluating the Usability and Safety of Virtual Reality Application Combined with the SWalker for Functional Gait Rehabilitation.

Objective: This research evaluates from a usability point of view the combination of a developed fully immersive virtual reality (VR) solution with the SWalker robotic device. It aims to contribute to research in the exploration of immersive experiences overground with a functional gait recovery device. Materials and Methods: We evaluated the system in a pilot study with 20 healthy participants aged 85.1 (SD: 6.29). Participants used the SWalker-VR platform while testing one VR application focused on walking and the other on balance practice. Afterward, the participants answered three usability questionnaires. Results: The platform was validated in terms of safety using the Simulator Sickness Questionnaire, obtaining less than 20 points for all subscales: nausea (4.29 ± 14.47), oculomotor (0.38 ± 14.18), and disorientation (1.39 ± 14.52). For usability evaluation, the System Usability Scale provided an overall score of 70.63 ± 11.64, and the Post-Study System Usability Questionnaire (PSSUQ) rated 1.61 ± 0.54. The usability scores reported by both questionnaires were moderate and good, respectively. These results were similar in overall scores for both groups: participants with low cognitive level and participants with high cognitive level. Finally, the possible causes for the "no answered" responses on the PSSUQ were discussed. Conclusion: It is concluded that the SWalker-VR platform is reported to have adequate usability and high security by older adults. The potential interest of studying the effects of the long-term use of this platform by older adults with gait impairment is expressed. Clinical Trials reference: NCT06025981.

Cell and tissue interactions of Treponema pallidum in primary and secondary syphilitic skin lesions: an ultrastructural study of serial sections.

There are limited reports on the ultrastructure of syphilis skin lesions. The aim of this study has been to perform an electron microscopic investigation of the morphology and the tissue distribution of treponemes in primary and secondary cutaneous lesions. Three cases of primary syphilitic chancre and one case of secondary syphilis were included. Prominent epidermal abnormalities in the primary chancre and a perivascular inflammatory infiltrate in the secondary lesion were found by light microscopy. Ultrastructurally, spirochetes were located mainly in the blood vessel walls and dermal tissue of the chancre lesions. In the secondary syphilis case, spirochetes were more abundant between epidermal keratinocytes. Most of them adjusted to the intercellular spaces. Occasionally, the electron microscopy images were highly suggestive of an intracellular location. Both the ultrastructural and immunohistochemical examination of the primary and secondary syphilis lesions showed a paradoxical distribution of the causative microorganisms compared to the light microscopic changes. In addition, the ultrastructural findings strongly suggest that Treponema pallidum subspecies pallidum invades tissues, not only through an intercellular, but also through a transcellular pathway.

PedaleoVR: Usability study of a virtual reality application for cycling exercise in patients with lower limb disorders and elderly people.

Achieving adherence to physical exercise training is essential in elders and adults with neurological disorders. Immersive technologies are seeing wide adoption among new neurorehabilitation therapies, as they provide a highly effective motivational and stimulating component. The aim of this study is to verify whether the developed virtual reality system for pedaling exercise is accepted and could be safety, useful and motivating for these populations. A feasibility study was conducted with patients with neuromotor disorders and elderly people from Lescer Clinic and the residential group Albertia, respectively. All the participants performed a pedaling exercise session with virtual reality platform. Then, the Intrinsic Motivation Inventory, the System Usability Scale (SUS), Credibility and Expectancy Questionnaire, were assessed in the group of 20 adults (mean age = 61.1; standard deviation = 12.617, 15 males and 5 females) with lower limb disorders. While the Simulator Sickness Questionnaire, Presence Questionnaire, Game user Experience Satisfaction Scale and SUS were assessed in the group of 18 elders (mean age = 85.16; standard deviation = 5.93, 5 males and 13 females). In light of the outcomes, PedaleoVR is considered to be a credible, usable and motivational tool towards adults with neuromotor disorders to perform cycling exercise, and therefore its usage could contribute to adherence to lower limb training activities. Moreover, PedaleoVR does not generate negative effects related to cybersickness while the sensation of presence and the degree of satisfaction generated have been positively evaluated by the geriatric population. This trial has been registered at ClinicalTrials.gov under the identifier: NCT05162040, Dec 2021.

Development and Clinical Validation of a Rehabilitation Platform for Hip Fracture in Elderly Population.

Hip fracture is one of the most common traumatisms associated with falls in the elderly, severely affecting the patient's mobility and independence. In recent years, the use of robotic technology has proven to be effective in gait rehabilitation, especially for neurological disorders. However, there is a lack of research validating these devices for hip fracture in elderly patients. This paper presents the design and evaluation of a novel assistive platform for hip rehabilitation, SWalker, aimed at improving the rehabilitation of this condition. Functional validation of the SWalker platform was carried out with five healthy elderly subjects and two physiotherapists. Clinical validation was conducted with 34 patients with hip fracture. The control group ( [Formula: see text], age = 86.38±6.16 years, 75% female) followed conventional therapy, while the intervention group ( [Formula: see text], age = 86.80±6.32 years, 90% female) was rehabilitated using SWalker. The functional validation of the device reported good acceptability (System Usability Scale >85). In the clinical validation, the control group required 68.09±27.38 rehabilitation sessions compared to 22.60±16.75 in the intervention group ( [Formula: see text]). Patients in the control group needed 120.33±53.64 days to reach ambulation, while patients rehabilitated with SWalker achieved that stage in 67.11±51.07 days ( [Formula: see text]). FAC and Tinetti indexes presented a larger improvement in the intervention group when compared with the control group ( [Formula: see text] and [Formula: see text], respectively). The SWalker platform can be considered an effective tool to enhance autonomous gait and shorten rehabilitation therapy in elderly hip fracture patients. This result encourages further research on robotic rehabilitation platforms for hip fracture.

The unified protocol as an internet-based intervention for emotional disorders: Randomized controlled trial.

The Unified Protocol (UP) as a transdiagnostic intervention has primarily been applied in the treatment of anxiety disorders and in face-to-face-settings. The current study investigated the efficacy of a 10-week Internet-based adaptation of the UP for anxiety, depressive, and somatic symptom disorders. The trial was registered under DRKS00014820 at the German Clinical Trial Registry, DRKS. Participants (n = 129) were randomized to treatment or waitlist control. Significant treatment effects were found for symptom distress, satisfaction with life, positive/negative affect and markers of anxiety, depression, and somatic symptom burden (within-group Hedges' g = 0.32-1.38 and between-group g = 0.20-1.11). Treatment gains were maintained at 1- and 6-month-follow-up. Subgroup analyses showed comparable effects in participants with anxiety and depressive disorders. 26.6% dropped out of treatment and 35.38% did not provide post-treatment assessments. The results strengthen the application of the UP as an Internet-based treatment for alleviating symptom distress across emotional disorders. More research on the applicability for single disorders is needed and avenues to improve adherence and attrition rates should be explored.

Syndrome-Based Prescription to Optimize Psychotropics: Are CHROME Criteria a Game Changer?

A variety of medical and social factors have contributed over the last decades to the overuse of psychotropic drugs in people with dementia. One social factor is probably the frequent failure to provide adequate person-centered care, be it in the community or in institutional settings. This unfortunate reality has been reacted upon with numerous guidelines to reduce prescriptions of the most dangerous drugs (e.g., neuroleptics). Each psychotropic drug prescription can in principle be assessed around three dimensions: (a) adequate, (b) inadequate, and (c) chemical restraint. The CHemical Restraints avOidance MEthodology (CHROME) defined chemical restraint as any prescription based on organizational convenience, rather than justified with medical diagnosis. Two validation studies revealed that one of the main medical reasons of over- and miss-prescriptions was symptom-based prescription. By switching to syndrome-based prescription, a large proportion of drugs could be de-prescribed and some re-adjusted or kept. Paucity of research and weakness of data are not conclusive about the adequacy of specific drugs for the myriad of cases presented by patients with dementia and comorbid conditions. Clinical practice, however, leads us to believe that even under optimal care conditions, psychotropics might still contribute to quality of life if based on an adequate diagnosis. This article explains the rationale that underlies a syndromic approach aimed at optimizing psychotropic treatment in people with dementia whose significant suffering derives from their thought, affective, or behavioral problems. The results of previous validation studies of this new methodology will be discussed and conclusions for future results will be drawn.

CHROME Criteria and Quality of Life: A Pilot Study from Maria Wolff-Albertia.

BACKGROUND: Over- and potentially inappropriate prescribing of psychotropic medications is a major public health concern among people with dementia. OBJECTIVE: Describe the CHemical Restraints avOidance MEthodology (CHROME) criteria and evaluate its effects on psychotropic prescribing and quality of life (QoL). METHODS: Observational, prospective, two-wave study conducted in two nursing homes. A multicomponent program to eliminate chemical restraints and attain quality prescription of psychotropic medications was implemented. CHROME's diagnostic criteria comprise constellations of behavioral and psychological symptoms of dementia under six primary syndromic diagnoses. Since pharmacologic treatment is aimed at only one syndrome, polypharmacy is avoided. Psychotropic prescription, QoL, neuropsychiatric symptoms (NPS), and other clinical measurements were collected before and one year after the intervention. Results are presented for all residents (n = 171) and for completer subjects (n = 115). RESULTS: Mean age (SD) of the residents was 87.8 (5.7), 78.9% were women, and 68.5% suffered advanced dementia. Psychotropic prescriptions decreased from 1.9 (1.1) to 0.9 (1.0) (p < 0.0005). Substantive reduction in prescribing frequency was observed for antidepressants (76.9% pre-intervention, 33.8% post-intervention) and for atypical neuroleptics (38.8% pre-intervention, 15.1% post-intervention). There was improvement in patient's response to surroundings (p < 0.0005) and total NPS (p < 0.01), but small worsening occurred in social interaction (p < 0.02, completer subjects). Safety measurements remained stable. CONCLUSION: CHROME criteria appear to optimize psychotropic prescriptions, avoid chemical restraints, and allow external verification of quality prescriptions. Extensive use seems feasible, related to substantial reduction of prescriptions, and of benefit for people with dementia as de-prescriptions are not associated to increased NPS or QoL loss.

CD5L is a pleiotropic player in liver fibrosis controlling damage, fibrosis and immune cell content.

BACKGROUND: Chronic hepatic inflammation leads to liver fibrosis, which may progress to cirrhosis, a condition with high morbidity. Our aim was to assess the as yet unknown role of innate immunity protein CD5L in liver fibrosis. METHODS: CD5L was measured by ELISA in plasma samples from cirrhotic (n = 63) and hepatitis (n = 39) patients, and healthy controls (n = 7), by immunohistochemistry in cirrhotic tissue (n = 12), and by quantitative RT-PCR in mouse liver cell subsets isolated by cell sorting. Recombinant CD5L (rCD5L) was administered into a murine model of CCl4-induced fibrosis, and damage, fibrosis and hepatic immune cell infiltration, including the LyC6hi (pro-fibrotic)-LyC6low (pro-resolutive) monocyte ratio were determined. Moreover, rCD5L was added into primary human hepatic stellate cells to study transforming growth factor ß (TGFß) activation responses. FINDINGS: Cirrhotic patients showed elevated plasma CD5L concentrations as compared to patients with hepatitis and healthy controls (Mann-Whitney test p < 0·0001). Moreover, plasma CD5L correlated with disease progression, FIB4 fibrosis score (r:0·25, p < 0·0001) and tissue expression (r = 0·649; p = 0·022). Accordingly, CCl4-induced damage increased CD5L levels in total liver, particularly in hepatocytes and macrophages. rCD5L administration attenuated CCl4-induced injury and fibrosis as determined by reduced serum transaminase and collagen content. Moreover, rCD5L inhibited immune cell infiltration and promoted a phenotypic shift in monocytes from LyC6hi to LyC6low. Interestingly, rCD5L also had a direct effect on primary human hepatic stellate cells promoting SMAD7 expression, thus repressing TGFß signalling. INTERPRETATION: Our study identifies CD5L as a key pleiotropic inhibitor of chronic liver injury. FUND: Fundació Marató TV3, AGAUR and the ISCIII-EDRF.

CO Oxidation at 20 °C on Au Catalysts Supported on Mesoporous Silica: Effects of Support Structural Properties and Modifiers.

In this work we report the effects of support structural properties and its modification with some metal oxides modifiers on the catalytic behavior of Au catalysts in the total CO oxidation at 20 °C. Au catalysts were supported on mesoporous silica materials (MSM) having different structural properties: Channel-like (SBA-15), cage-like (SBA-16), hexagonal (HMS), and disordered (DMS-1) structures. The effect of the modifier was evaluated by comparison of the catalytic response of the SBA-15-based catalysts modified with MgO, Fe2O3, TiO2, and CeO2. The chemical, structural, and electronic properties of the catalysts were investigated by a variety of techniques (metal content analysis by ICP-OES, N2 physisorption, XRD, UV-vis DRS, DRIFTS of adsorbed CO and OH regions, oxygen storage capacity (OSC), HR-TEM, and XPS). The activity of calcined catalysts in the CO oxidation reaction were evaluated at steady state conditions, at 20 °C, atmospheric pressure, and when using, as feed, a 1%CO/1%O2/98% gas mixture. The work clearly demonstrated that all Au catalysts supported on the mesoporous silicas modified with metal oxides were more active than the Au/SBA-15 and Au/MgO reference ones. The support structural properties and type of dopant were important factors influencing on the catalyst behavior. Concerning the support textural properties, it was found that the HMS substrate with the wormhole-structure offers better porosity and specific surface area than their silica counterparts having channel-like (SBA-15), cage-like (SBA-16), and disordered (DMS-1) mesoporous structures. Concerning the effect of modifier, the best catalytic response was achieved with the catalysts modified with MgO. After activation by calcination at 200 °C for 4 h, the Au/MgO/HMS catalyst exhibited the best catalytic performance, which was ascribed to the combined effects of the best structural properties, a large support oxygen storage capacity and homogeneous distribution of gold particles on the support (external and inner). Implications of the type of active sites (Au1+ or Au°), support structural properties and role of modifier on the catalytic activity are discussed.

Generation of Hepatic Stellate Cells from Human Pluripotent Stem Cells Enables In Vitro Modeling of Liver Fibrosis.

The development of complex in vitro hepatic systems and artificial liver devices has been hampered by the lack of reliable sources for relevant cell types, such as hepatic stellate cells (HSCs). Here we report efficient differentiation of human pluripotent stem cells into HSC-like cells (iPSC-HSCs). iPSC-HSCs closely resemble primary human HSCs at the transcriptional, cellular, and functional levels and possess a gene expression profile intermediate between that of quiescent and activated HSCs. Functional analyses revealed that iPSC-HSCs accumulate retinyl esters in lipid droplets and are activated in response to mediators of wound healing, similar to their in vivo counterparts. When maintained as 3D spheroids with HepaRG hepatocytes, iPSC-HSCs exhibit a quiescent phenotype but mount a fibrogenic response and secrete pro-collagen in response to known stimuli and hepatocyte toxicity. Thus, this protocol provides a robust in vitro system for studying HSC development, modeling liver fibrosis, and drug toxicity screening.

Assessment of Liver Fibrotic Insults In Vitro.

In vitro systems are required to evaluate potential liver fibrogenic effects of drugs and compounds during drug development and toxicity screening, respectively. Upon liver injury or toxicity, hepatic stellate cells are activated, thereby acquiring a myofibroblastic phenotype and participating in extracellular matrix deposition and liver fibrosis. The most widely used in vitro models to investigate liver fibrogenesis are primary cultures of hepatic stellate cells, which can be isolated from healthy human livers. Currently, there are no effective methods to maintain hepatic stellate cells in vitro in a quiescent phenotype. Therefore, when cells are plated, they spontaneously become activated in few days. Most in vitro studies in this area have been performed with monocultures of hepatic stellate cells in order to assess the direct effects of a given factor on hepatic stellate cell activation or the induction of inflammatory and fibrogenic responses. In this chapter, focus is put on basic protocols to isolate hepatic stellate cells from human tissue and to maintain them in culture as well as on common in vitro assays to evaluate their response to profibrogenic factors.

Integrative miRNA and Gene Expression Profiling Analysis of Human Quiescent Hepatic Stellate Cells.

Unveiling the regulatory pathways maintaining hepatic stellate cells (HSC) in a quiescent (q) phenotype is essential to develop new therapeutic strategies to treat fibrogenic diseases. To uncover the miRNA-mRNA regulatory interactions in qHSCs, HSCs were FACS-sorted from healthy livers and activated HSCs (aHSCs) were generated in vitro. MiRNA Taqman array analysis showed HSCs expressed a low number of miRNAs (n = 259), from which 47 were down-regulated and 212 up-regulated upon activation. Computational integration of miRNA and gene expression profiles revealed that 66% of qHSC-associated miRNAs correlated with more than 6 altered target mRNAs (17,28 ± 10,7 targets/miRNA) whereas aHSC-associated miRNAs had an average of 1,49 targeted genes. Interestingly, interaction networks generated by miRNA-targeted genes in qHSCs were associated with key HSC activation processes. Next, selected miRNAs were validated in healthy and cirrhotic human livers and miR-192 was chosen for functional analysis. Down-regulation of miR-192 in HSCs was found to be an early event during fibrosis progression in mouse models of liver injury. Moreover, mimic assays for miR-192 in HSCs revealed its role in HSC activation, proliferation and migration. Together, these results uncover the importance of miRNAs in the maintenance of the qHSC phenotype and form the basis for understanding the regulatory networks in HSCs.

New enterovirulent Escherichia coli serogroup 64474 showing antigenic and genotypic relationships to Shigella boydii 16.

Studies based on the analysis of housekeeping genes indicate that Escherichia coli and all Shigella species, except for Shigella boydii type 13, belong to a single species. This study analysed the phenotypic and genotypic characteristics of 23 E. coli strains isolated in different countries from faecal specimens taken from children with diarrhoea. Strains were identified using the VITEK system and typed with rabbit sera obtained against 186 somatic and 53 flagellar E. coli antigens and against 45 Shigella somatic antigens. Biochemical analysis of these strains showed a typical E. coli profile with a defined reaction against both E. coli O179 and S. boydii 16 somatic antisera. Agglutination assays for flagellar antigens showed a response against H2 in 7 (30 %) strains, H10 in 2 (9 %) strains, H32 in 12 (52 %) strains and H34 in 2 (9 %) strains, demonstrating 4 serotypes associated with this new somatic antigen 64474. A serum against one of these E. coli strains (64474) was prepared. Absorption assays of S. boydii 16 and E. coli 64474 antisera with E. coli O179 antigen removed the agglutination response against this O179 antigen completely, while the agglutination titres against both S. boydii 16 and E. coli 64474 remained the same. Four (17 %) E. coli strains showed antimicrobial resistance to piperacillin only, one (4 %) to piperacillin and trimethoprim/sulfamethoxazole, one (4 %) to ciprofloxacin, nitrofurantoin and piperacillin, and two (9 %) strains were resistant to ciprofloxacin, norfloxacin, ofloxacin, piperacillin and trimethoprim/sulfamethoxazole. With regards to PCR assays, one (4 %) of the strains was positive for Shigella gene ipaH, one (4 %) for ipaA, two (9 %) for ipaB, one (4 %) for ipaD, two (9 %) for sepA and three (13 %) for ospF. The rfb gene cluster in the E. coli strains was analysed by RFLP and compared with the gene cluster obtained from S. boydii 16. The rfb-RFLP patterns for all 23 E. coli strains were similar to those obtained for S. boydii 16. The results from PCR tests to detect rfb genes wzx (encoding O unit flippase) and wzy (encoding polymerase) belonging to a cluster related to the biosynthesis of the S. boydii 16-specific O antigen were positive in 21 (91 %) and 22 (96 %) of the strains, respectively. PCR assays to detect E. coli virulence genes were also performed. These assays detected enterotoxigenic E. coli genes ltA1 in 12 of the strains (52 %), st1a in 4 (17 %), cfa1 in 6 (26 %), cs1 in 1 (4 %), cs3 in 3 (13 %), cs13 in 9 (39 %) and cs14 in 5 (22 %) of the strains. Results from the PFGE analyses confirmed the wide geographical distribution of these strains suggesting that 64474 : H2, 64474 : H10, 64474 : H32 and 64474 : H34 are new serotypes of E. coli strains with a defined virulence capacity, and share a common O antigen with S. boydii 16.

Relación entre el transporte y laactividad física en universitarios,Santa Marta, Colombia, 2013: estudio de corte transversal / Association between travel mode, physical activity and sociodemographic variables in undergraduate students health, Santa Marta, Colombia, 2013: cross sectional study

Observar la asociación entre el medio de transporte, la actividad física y el programa de estudios en estudiantes de salud de la Universidad del Magdalena. Materiales y métodos: 302 estudiantes de salud fueron seleccionados aleatoriamente y se les aplicó el Cuestionario Mundial sobre Actividad Física. Se realizó análisis estadístico de asociación por X2. Resultados: se observa asociación estadísticamente significativa entre el medio transporte y el sexo (X2 = 7,2; p = 0,027), del total de 193(63,9 %) estudiantes que caminan o montan en bicicleta, el 52,3 % son mujeres y el47,6 % son hombres. Se encuentra asociación estadísticamente significativa entrelas variables medio de transporte y programa de estudios (X2 = 14,4; p = 0,024): deltotal de 193 estudiantes que caminan o montan en bicicleta por programa de estudios el 34,7 % son estudiantes de medicina y el 30 % son estudiantes de odontología.Igualmente se encuentra asociación entre el sexo y la inactividad física (X2 = 8,9; p =0,002), 247 (81,8 %) estudiantes realizan menos de 150 minutos de actividad física por semana. Conclusiones: en relación con el medio de transporte, tanto hombres como mujeres en su mayoría caminan o montan en bicicleta, siendo los estudiantes de medicina y odontología los que más tienden a transportarse de esta manera. En relación con la actividad física, cerca del 82 % de los estudiantes realizan menos de 150 minutos de actividad física por semana...