Background: Liver fibrosis induced by non-alcoholic fatty liver disease causes peri-interventional complications in morbidly obese patients. We determined the performance of transient elastography (TE), acoustic radiation force impulse (ARFI) imaging, and enhanced liver fibrosis (ELF) score for fibrosis detection in bariatric patients. Patients and Methods: 41 patients (median BMI 47 kg/m2) underwent 14-day low-energy diets to improve conditions prior to bariatric surgery (day 0). TE (M and XL probe), ARFI, and ELF score were performed on days -15 and -1 and compared with intraoperative liver biopsies (NAS staging). Results: Valid TE and ARFI results at day -15 and -1 were obtained in 49%/88%and 51%/90%of cases, respectively. High skin-to-liver-capsule distances correlated with invalid TE measurements. Fibrosis of liver biopsies was staged as F1 and F3 in n = 40 and n = 1 individuals. However, variations (median/range at d-15/-1) of TE (4.6/2.6–75 and 6.7/2.9–21.3 kPa) and ARFI (2.1/0.7–3.7 and 2.0/0.7–3.8 m/s) were high and associated with overestimation of fibrosis. The ELF score correctly classified 87.5%of patients. Conclusion: In bariatric patients, performance of TE and ARFI was poor and did not improve after weight loss. The ELF score correctly classified the majority of cases and should be further evaluated.

published_or_final_version / abstract / toc / Medicine / Master / Doctor of Medicine

When a child is diagnosed with a chronic life threatening illness there is a significant impact on the entire family. Siblings are at risk for psychological adaptation problems because of their unique relationship with the ill child and the effect of that illness on family functioning. Few studies have been reported which examine the impact of chronic life-threatening illnesses in children on healthy siblings.The purpose of this study was to investigate the predisposing risk and protective factors that affect the psychological adaptation of healthy siblings of a child with Cystic Fibrosis (CF). A descriptive study was conducted using a convenience sample from two CF centers. The relationship between several variables was explored using non- parametric correlations.In this sample, significant negative correlations were found between parental stress and their reported financial well being and emotional or behavioral problems in the well siblings prior to the diagnosis of CF was made in the sick sibling and perceived parental support.For adolescent siblings, The Behavioral Symptoms Index (BSI) was correlated to reported stress in their parents and negatively correlated to the parental perceived support. The Emotional Symptoms Index (ESI) was correlated to the BSI. A significant negative correlation was also noted between age and Internalizing Behaviors. These relationships were not significant for the child siblings in this sample.There were no significant relationships among gender, maternal education, and caretaker for the well sibling when the child with CF was hospitalized and their Internalizing and Externalizing Behaviors. The health care teams in these clinics seldom discussed CF with the well sibling. However, for the children in this study, there was a significant negative correlation with this discussion and their Externalizing Behaviors.Findings from this study suggest that the family environment, especially parental stress and perceived social support, may affect the adaptation of the well sibling. Adolescent siblings were more at risk for this environmental influence than their younger counterparts.

Recent studies support the association between metabolic syndrome (MetS), a cluster of cardiovascular risk factors, and diastolic dysfunction. Disproportionate collagen accumulation, particularly cross-linking of collagen, plays a key role in translating interstitial fibrosis into mechanical chamber stiffness and diastolic dysfunction. Characteristic changes in the expression and activity of myocardial lysyl oxidase (LOX), a matrix modifying enzyme that catalyzes cross-linked collagen, are unclear in MetS. We established a diet-induced MetS model to study diastolic dysfunction by treating male C57BL/6 mice a high-fat high-simple carbohydrate (HFHSC) diet for 6 months. Despite blunted gene expression of LOX isoforms, MetS mice demonstrated significant increase in the ratio of protein expression of mature to proenzyme LOX, enhanced LOX activity, and increased cardiac cross-linked collagen compared with controls. This fibrotic response coincided with marked increase in left ventricular end-diastolic pressure and stiffness and impaired diastolic filling pattern. Our data demonstrate that diet-induced MetS alters the remodeling enzyme LOX, thereby increasing the amount of crosslinking and inducing diastolic dysfunction.Furthermore we examined the role of T-lymphocytes in myocardial LOX regulation in diet-induced fibrotic hearts. Female SCID mice which are devoid of functional T-lymphocytes and C57BL/6 mice were treated with HFHSC diet for 12 months. Similar to male C67BL/6, female HFHSC-fed C57BL/6 mice demonstrated significant increase in maturation and catalytic activity of myocardial LOX, cross-linking, ventricular stiffness and diastolic dysfunction. Whereas induction of LOX protein was minimal in SCID mice compared with wild-type counterparts. Correspondingly fibrillar cross-linked collagen formation and diastolic dysfunction were less prominent in SCID mice. Our results suggest a potential role of T-lymphocytes in induction of myocardial stiffness and diastolic dysfunction through modulation of LOX-dependent collagen maturation.Moreover we studied the role of leptin, an adipokine over-produced in MetS with fibrotic effects in non-cardiac tissues, as a key mediator of profibrogenic responses in the heart by administrating leptin to C57BL/6 and leptin-deficient ob/ob mice. With exogenous leptin administration ob/ob mice displayed passive diastolic filling dysfunction that coincided with increase in myocardial collagen compared with ob/ob controls. Our findings suggest profibrotic effects of leptin in the heart, primarily through predominance of collagen synthesis over degradation.

Studies were conducted using cultured immortalized wild type (non-CF) and mutant (CF) DeltaF508 cystic fibrosis transmembrane conductance regulator (CFTR) tracheal epithelial cells on the anti-inflammatory impact of agents that may alter ceramide and glutathione (GSH) metabolism. The CF cells demonstrated abnormally high levels of GSH and glutathione disulfide (GSSG), which could diminish intracellular production of ceramide, a key modulator of inflammation and apoptosis. Hence, additional cell culture studies were carried out with a known inducer of in situ ceramide synthesis, N-4(4-hydroxyphenyl) retinamide (fenretinide) on interleukin (IL)-8 release, intracellular ceramide content, and cellular proliferation in both the basal state and following the inflammatory stimuli of tumor necrosis factor (TNF) -alpha. Fenretinide treatment was associated with a dose-dependent increase in the cellular content of ceramide in both CF and non CF cells. Also, an inhibition of IL-8 release in the inflammatory condition of TNF-alpha treatment was observed following fenretinide treatment in the CF cells. As hyperbaric treatment of whey proteins was previously associated with improved survivability and higher GSH content in a Pseudomonas aeruginosa murine model of cystic fibrosis (CF), the anti-inflammatory role of low molecular weight peptides (< 1kDa) generated from enzymatic hydrolysis of native and pressurized whey protein isolates (WPI) was examined. Pressure treatment of WPI was associated with an enhanced protein digestibility and an altered peptide profile following in vitro digestion. The whey peptides were tested CF and non-CF lung epithelial cells to identify for their effects on GSH metabolism. The impact of the combined treatment of fenretinide and WPH was also tested in terms of apoptosis and cytokine release in cell culture. As opposed to non-CF cells, CF cells showed a strong downtrend in release of IL-8 following the combined fenretinide and whey peptide treatment. In addition, whey peptides protected wild type epithelial cells from the apoptotic effect of fenretinide. Our results suggest the usefulness of these agents as a pharmacological treatment in CF.

Cystic Fibrosis (CF) is caused by mutations in CFTR, a protein for chloride efflux in epithelial cells. VIP is a peptide that activates CFTR and improves membrane stability; VIP has 3 receptors VPAC1, VPAC2 and PAC1 that can cause CFTR phosphorylation. VIP-knockout (VIPKO) mice experience inflammation and reduced CFTR membrane localization comparable to CF phenotypes, that’s reversible after 3 weeks of VIP treatment (VIPKOT). In this thesis western blotting showed VPAC1 and VPAC2 expression increased in VIPKO and VIPKOT lung and duodenum tissues. The expression and maturation of CFTR was unchanged in both VIPKO and VIPKOT tissues. The results showed absence of VIP caused increased receptor expression in VIPKO mice, after VIP treatment VIPKO mice maintained increased receptor expression. VIP treatment reduces inflammation and restores existing CFTR membrane localization in VIPKO mice. VIP receptor expression may be important for future treatment of CF for CFTR localization and reducing tissue inflammation.

Myocardial fibrosis is a common pathological finding in patients with cardiovascular disease and is believed to be a major contributing factor in the development of end stage organ failure. Early events that promote the development of myocardial fibrosis are not well understood. Rapid cellular infiltration into the cardiac tissue is evident in fibrosis but the infiltrating populations and their functions have yet to be completely elucidated. The aim of this thesis was to characterize the phenotype and function of this cellular population in a model of hypertension mediated myocardial fibrosis. Furthermore, we intended to explore therapies that target this population and ameliorate fibrosis. We characterized a novel population of infiltrating cells as circulating fibroblast progenitor cells, termed fibrocytes. We determined that this population does not appear to specifically migrate in response to previously established chemotactic signals (CCL2 or CXCL12). We found that fibrocytes respond to fibrogenic stimuli (AngII and CTGF) by increasing the expression of collagen and CTGF, an early molecular mediator of fibrosis, while also promoting fibrocyte differentiation. Using an anti-hypertension treatment, we found that hypertension as a physiologic stimulus likely promotes cellular infiltration and corresponding fibrosis. We also established that treatment with activated protein C (aPC) conferred protection against the development of myocardial fibrosis, potentially by inhibiting fibrocyte recruitment and/or activation. Lastly, to assess fibrocyte involvement in the progression of human myocardial fibrosis we assessed fibrocytes in levels in the circulation of patients with ischemic heart disease compared to healthy controls. We found that patients with ischemic heart disease had an increase of circulating cells that have the potential to become fibrocytes compared to healthy controls and therefore likely contribute to myocardial fibrosis. From this data, we propose that fibrocytes are a key effector cell that directly promotes pathologic fibrosis within the injured myocardium. Understanding their migration and function is therefore essential to the development of future therapies targeting this cell type to inhibit their role in fibrosis.

The Apelin-APJ system is transiently up-regulated in murine models of cardiac dysfunction. We have previously shown that Apelin-deficient mice subjected to aortic constriction suffer from severe fibrosis. In turn, we hypothesized that Apelin deficiency will also exaggerate the fibrosis phenotype post experimental myocardial infarction, associated with changes in fibroblast cell activity. Apelin-deficient and wildtype mice were randomly subjected to sham operation or left coronary artery ligation. Apelin deficiency worsened cardiac functionality, enhanced fibrosis-related gene expression and morphology, and enhanced vimentin intermediate filament expression, which may be involved in increasing fibroblast proliferation. MicroRNA target prediction softwares predict that apelin and vimentin 3 ’UTRs are potential targets of microRNA-378 regulation, and were confirmed with Luciferase reporter assays and western blot. Apelin up-regulation may be a useful strategy for attenuating unfavorable fibrosis through down-regulating vimentin-mediated adverse fibroblast activity. MicroRNA-378 regulation may be partly responsible for changes in apelin and vimentin expression.

The pulmonary infection with Pseudomonas aeruginosa is considered as one of the main causes of health deterioration in cystic fibrosis patients (CF). Efficient management of P. aeruginosa in CF remains difficult mainly with the emergence of multidrug-resistant strains leading ultimately to death. There is a pressing need for new approaches to control these Pseudomonal infections. Current studies on the antimicrobial efficacy of liposomal antibiotics have shown conflicting results. We sought to assess whether the incorporation of clarithromycin into liposomes could improve its antibacterial activity against clinical isolate of Pseudomonas aeruginosa from CF patients. Different formulations of liposomal clarithromycin were prepared, characterized and their antibacterial activities against resistant strains of P. aeruginosa were investigated. These formulations reduced the biofilm formation, the virulence factors production and the bacterial motilities compared to free drug. The therapeutic importance of liposome containing macrolides in the management of experimental pseudomonal lung infection in animals is warranted.

The Apelin-APJ system is transiently up-regulated in murine models of cardiac dysfunction. We have previously shown that Apelin-deficient mice subjected to aortic constriction suffer from severe fibrosis. In turn, we hypothesized that Apelin deficiency will also exaggerate the fibrosis phenotype post experimental myocardial infarction, associated with changes in fibroblast cell activity. Apelin-deficient and wildtype mice were randomly subjected to sham operation or left coronary artery ligation. Apelin deficiency worsened cardiac functionality, enhanced fibrosis-related gene expression and morphology, and enhanced vimentin intermediate filament expression, which may be involved in increasing fibroblast proliferation. MicroRNA target prediction softwares predict that apelin and vimentin 3 ’UTRs are potential targets of microRNA-378 regulation, and were confirmed with Luciferase reporter assays and western blot. Apelin up-regulation may be a useful strategy for attenuating unfavorable fibrosis through down-regulating vimentin-mediated adverse fibroblast activity. MicroRNA-378 regulation may be partly responsible for changes in apelin and vimentin expression.