Solute Transporter OCTN1/Slc22a4 Affects Disease Severity and Response to Infliximab in Experimental Colitis: Role of Gut Microbiota and Immune Modulation.

BACKGROUND: Inflammatory bowel diseases are chronic disabling conditions with a complex and multifactorial etiology, still incompletely understood. OCTN1, an organic cation transporter, could have a role in modulating the inflammatory response, and some genetic polymorphisms of this molecule have been associated with increased risk of inflammatory bowel diseases. Until now, limited information exists on its potential in predicting/modulating patient's response to therapies. The aim of this study was to evaluate the role of OCTN1 in modifying gut microbiota and mucosal immunity in response to infliximab therapy in murine colitis. METHODS: A dextran sodium sulphate model of colitis was used to assess the clinical efficacy of infliximab administered intravenously in ocnt1 gene knockout mice and their C57BL/6 controls. Stool, colon, and mesenteric lymph node samples were collected to evaluate differences in gut microbiota composition, histology, and T cell populations, respectively. RESULTS: Octn1 -/- influences the microbiota profile and is associated with a worse dysbiosis in mice with colitis. Infliximab treatment attenuates colitis-associated dysbiosis, with an increase of bacterial richness and evenness in both strains. In comparison with wild type, octn1-/- mice have milder disease and a higher baseline percentage of Treg, Tmemory, Th2 and Th17 cells. CONCLUSIONS: Our data support the murine model to study OCTN1 genetic contribution to inflammatory bowel diseases. This could be the first step towards the recognition of this membrane transporter as a biomarker in inflammatory conditions and a predictor of response to therapies.

In this article, we evaluated the role of OCTN1, an organic cation transporter, in modifying gut microbiota and immune T cell populations, as well as its effects on experimental colitis and the response to infliximab treatment.

Intensive exercise ameliorates motor and cognitive symptoms in experimental Parkinson's disease restoring striatal synaptic plasticity.

Intensive physical activity improves motor functions in patients with Parkinson's disease (PD) at early stages. However, the mechanisms underlying the beneficial effects of exercise on PD-associated neuronal alterations have not been fully clarified yet. Here, we tested the hypothesis that an intensive treadmill training program rescues alterations in striatal plasticity and early motor and cognitive deficits in rats receiving an intrastriatal injection of alpha-synuclein (α-syn) preformed fibrils. Improved motor control and visuospatial learning in active animals were associated with a recovery of dendritic spine density alterations and a lasting rescue of a physiological corticostriatal long-term potentiation (LTP). Pharmacological analyses of LTP show that modulations of N-methyl-d-aspartate receptors bearing GluN2B subunits and tropomyosin receptor kinase B, the main brain-derived neurotrophic factor receptor, are involved in these beneficial effects. We demonstrate that intensive exercise training has effects on the early plastic alterations induced by α-syn aggregates and reduces the spread of toxic α-syn species to other vulnerable brain areas.

Decellularized esophageal tubular scaffold microperforated by quantum molecular resonance technology and seeded with mesenchymal stromal cells for tissue engineering esophageal regeneration.

Current surgical options for patients requiring esophageal replacement suffer from several limitations and do not assure a satisfactory quality of life. Tissue engineering techniques for the creation of customized "self-developing" esophageal substitutes, which are obtained by seeding autologous cells on artificial or natural scaffolds, allow simplifying surgical procedures and achieving good clinical outcomes. In this context, an appealing approach is based on the exploitation of decellularized tissues as biological matrices to be colonized by the appropriate cell types to regenerate the desired organs. With specific regard to the esophagus, the presence of a thick connective texture in the decellularized scaffold hampers an adequate penetration and spatial distribution of cells. In the present work, the Quantum Molecular Resonance® (QMR) technology was used to create a regular microchannel structure inside the connective tissue of full-thickness decellularized tubular porcine esophagi to facilitate a diffuse and uniform spreading of seeded mesenchymal stromal cells within the scaffold. Esophageal samples were thoroughly characterized before and after decellularization and microperforation in terms of residual DNA content, matrix composition, structure and biomechanical features. The scaffold was seeded with mesenchymal stromal cells under dynamic conditions, to assess the ability to be repopulated before its implantation in a large animal model. At the end of the procedure, they resemble the original esophagus, preserving the characteristic multilayer composition and maintaining biomechanical properties adequate for surgery. After the sacrifice we had histological and immunohistochemical evidence of the full-thickness regeneration of the esophageal wall, resembling the native organ. These results suggest the QMR microperforated decellularized esophageal scaffold as a promising device for esophagus regeneration in patients needing esophageal substitution.

Surgical Training for Transanal Total Mesorectal Excision in a Live Animal Model: A Preliminary Experience.

Background: In this preliminary experience, the feasibility and effectiveness of surgical training with an animal model for transanal total mesorectal excision (TaTME) were evaluated. Methods: The training was conducted in two experimental animal laboratories in Italy authorized by the Italian Ministry of Health, using female Danish Landrace pigs under the supervision of surgeons with extensive experience in TaTME, animal laboratory training and cadaver laboratory training. The procedure was divided into separate steps, and all the participants were guided step-by-step throughout the entirety of the procedure. Results: During all the editions of the animal laboratory, all the procedures were completed with no major damage to the anatomical structures or intraoperative death of the animals. Live animal tissue is very similar to human tissue, helping trainees improve their tactile feedback. The bleeding effect improved the value of the training and taught the participants how to address this complication. The lack of mesorectal tissue in pigs compared with humans was the main difference. Animal laboratories should not be considered alternatives to cadaver laboratories but as complementary training activities due to their effectiveness and lower costs. Conclusions: Surgical training in animal models for TaTME seems to be effective and could be an opportunity to improve training alongside the use of a cadaver laboratory and proctoring.

Feasibility and safety of a new endoscopic synthetic sealant nebulizing device over gastric endoscopic submucosal dissections.

BACKGROUND: Endoscopic Submucosal Dissection (ESD) is the treatment of choice of superficial neoplastic gastrointestinal lesions. Delayed bleedings and perforations are still current clinical concerns. Glubran 2 is a synthetic cyanoacrylate-derived glue nowadays already widely used as an effective tissue adhesive. ENDONEB is a novel device thought for enabling the sealant nebulization over a specific targeted surface during laparotomy, laparoscopy, and thoracotomy. The aim of this single-center preclinical animal trial is to evaluate the feasibility and safety of the same nebulization technique during ESD in the perspective that further clinical studies would demonstrate the efficacy of Glubran 2 in preventing post-ESD adverse events. METHODS: Four live Landrace pigs were enrolled. Two approximately 30-mm-wide gastric ESDs were performed in each pig (experimental ESD and control ESD). About 0.5 mL of Glubran 2 was nebulized on the experimental ESDs. Subjective perception of the feasibility of the Glubran 2 nebulization was reported. Pigs were clinically monitored at follow-up and upper GI endoscopy was performed at 24 and 48 hours, when animals were euthanized to perform a macroscopic and histological analysis of the specimens. RESULTS: No peri-procedural adverse events were reported. Glubran 2 nebulization over experimental ESDs showed to be technically easy and time-effective. Clinical and endoscopic animal monitoring was negative at follow-up. At 24 hours, the Glubran 2 film was clearly visible on the eschar of the ESDs and signs of initial hydrolysis were discernable at 48 hours. No signs of peritoneal reaction were observed at the macroscopic examination. Equal transmural inflammation was described at the histological examination of both types of ESDs. CONCLUSIONS: Safety and feasibility profiles of Glubran 2 nebulizing ENDONEB device over ESD surfaces were excellent. Further evidences and human trials are needed to investigate its effectiveness in ESDs' eschars sealing and, thus, in delayed micro-perforations and bleedings prevention and treatment.

Photons detected in the active nerve by photographic technique.

The nervous system is one of the most complex expressions of biological evolution. Its high performance mostly relies on the basic principle of the action potential, a sequential activation of local ionic currents along the neural fiber. The implications of this essentially electrical phenomenon subsequently emerged in a more comprehensive electromagnetic perspective of neurotransmission. Several studies focused on the possible role of photons in neural communication and provided evidence of the transfer of photons through myelinated axons. A hypothesis is that myelin sheath would behave as an optical waveguide, although the source of photons is controversial. In a previous work, we proposed a model describing how photons would arise at the node of Ranvier. In this study we experimentally detected photons in the node of Ranvier by Ag+ photoreduction measurement technique, during electrically induced nerve activity. Our results suggest that in association to the action potential a photonic radiation takes place in the node.

Federation of European Laboratory Animal Science Associations recommendations of best practices for the health management of ruminants and pigs used for scientific and educational purposes.

Most ruminants and pigs used for scientific and educational aims are bred not for these purposes but in a farm environment. Given the wide range of diseases that these species might have, ensuring that the animals' health status is appropriate can be complex and challenging. The Federation of European Laboratory Animal Science Associations has previously published recommendations for the health monitoring of experimental colonies of pigs (1998) and, respectively, calves, sheep and goats (2000). Unfortunately, the uptake of those recommendations was poor and insufficiently reported in scientific publications. These new recommendations for best practice focus on the main species of ruminants (cattle, sheep and goats) and pigs. They provide general and specific information helpful for designing a health management programme for the suppliers and for the user establishments, as well as guidance on animal procurement. Critical thinking based on the fields of use of the animals is promoted, aiming to help in taking informed decisions rather than establishing an exhaustive exclusion list for pathogens. Implementing the best health and welfare management practices should be done under the guidance of a competent attending veterinarian, with expertise and sufficient authority to take the appropriate action, doubled by excellent communication skills. It is strongly recommended that the user establishment's veterinarian works in close collaboration with the supplier's veterinarian.

Duodenal-jejunal bypass improves nonalcoholic fatty liver disease independently of weight loss in rodents with diet-induced obesity.

Nonalcoholic fatty liver disease (NAFLD) is the most common cause of liver-related mortality. NAFLD is associated with obesity, hepatic fat accumulation, and insulin resistance, all of which contribute to its pathophysiology. Weight-loss is the main therapy for NAFLD, and metabolic surgery is the most effective treatment for morbid obesity and its metabolic comorbidities. Although has been reported that Roux-en-Y gastric bypass can reverse NAFLD, it is unclear whether such effects result from reduced weight, from a lower calorie-intake, or from the direct influence of surgery on mechanisms contributing to NAFLD. We aimed to investigate whether gastrointestinal (GI) bypass surgery could induce direct effects on hepatic fat accumulation and insulin resistance, independently of weight reduction. Twenty Wistar rats on a high-fat diet underwent duodenal-jejunal-bypass (DJB) or sham operation and were pair fed (PF) for 15 wk after surgery to obtain a matched weight. Outcome measures include ectopic fat deposition, expression of genes and proteins involved in fat metabolism, insulin-signaling, and gluconeogenesis in liver and muscle. Despite no differences in body weight and calorie intake, DJB showed lower ectopic fat accumulation, improved peripheral and hepatic insulin sensitivity, and enhanced lipid droplet degradation. In both tissues, DJB increased insulin signaling, whereas hepatic key enzymes involved in gluconeogenesis and de novo lipogenesis were decreased. These findings suggest that DJB can reverse, independently of weight loss, ectopic fat deposition and insulin resistance, two features of NAFLD that share a mutual pathway, in which perilipin-2 (PLIN2) seems to be the main player, supporting further investigation into strategies that target the gut to treat metabolic liver diseases.NEW & NOTEWORTHY Our findings suggest that duodenal-jejunal bypass can reverse, independently of weight loss, ectopic fat deposition and insulin resistance, two features of nonalcoholic fatty liver disease that share a mutual pathway, in which perilipin-2 seems to be the main player. Our study supports further investigation into the role of proximal small intestine exclusion in the pathophysiology of nonalcoholic fatty liver disease to uncover less invasive treatments that mimic the effects of metabolic surgery and aims to prevent and treat metabolic liver disease.

Successful muscle regeneration by a homologous microperforated scaffold seeded with autologous mesenchymal stromal cells in a porcine esophageal substitution model.

BACKGROUND: Since the esophagus has no redundancy, congenital and acquired esophageal diseases often require esophageal substitution, with complicated surgery and intestinal or gastric transposition. Peri-and-post-operative complications are frequent, with major problems related to the food transit and reflux. During the last years tissue engineering products became an interesting therapeutic alternative for esophageal replacement, since they could mimic the organ structure and potentially help to restore the native functions and physiology. The use of acellular matrices pre-seeded with cells showed promising results for esophageal replacement approaches, but cell homing and adhesion to the scaffold remain an important issue and were investigated. METHODS: A porcine esophageal substitute constituted of a decellularized scaffold seeded with autologous bone marrow-derived mesenchymal stromal cells (BM-MSCs) was developed. In order to improve cell seeding and distribution throughout the scaffolds, they were micro-perforated by Quantum Molecular Resonance (QMR) technology (Telea Electronic Engineering). RESULTS: The treatment created a microporous network and cells were able to colonize both outer and inner layers of the scaffolds. Non seeded (NSS) and BM-MSCs seeded scaffolds (SS) were implanted on the thoracic esophagus of 4 and 8 pigs respectively, substituting only the muscle layer in a mucosal sparing technique. After 3 months from surgery, we observed an esophageal substenosis in 2/4 NSS pigs and in 6/8 SS pigs and a non-practicable stricture in 1/4 NSS pigs and 2/8 SS pigs. All the animals exhibited a normal weight increase, except one case in the SS group. Actin and desmin staining of the post-implant scaffolds evidenced the regeneration of a muscular layer from one anastomosis to another in the SS group but not in the NSS one. CONCLUSIONS: A muscle esophageal substitute starting from a porcine scaffold was developed and it was fully repopulated by BM-MSCs after seeding. The substitute was able to recapitulate in shape and function the original esophageal muscle layer.

Intake of Boron, Cadmium, and Molybdenum enhances rat thyroid cell transformation.

BACKGROUND: Epidemiologic data in volcanic areas suggest that environmental factors might be involved in the increase of thyroid cancer (TC) incidence. Recent reports indicate that several heavy metals and metalloids are increased in volcanic areas. This study aims to evaluate the combined effect of three of these elements Boron (B), Cadmium (Cd), and Molybdenum (Mo) - all increased in the volcanic area of Mt. Etna, in Italy - on thyroid tumorigenesis in the rat. METHODS: Female Wistar rats prone to develop thyroid tumors by low-iodine diet and methimazole treatment received ad libitum drinking water supplemented with B, Cd, and Mo at concentrations in the range found in the urine samples of residents of the volcanic area. At 5 and 10 months animals were euthanized, and their thyroid analysed. Statistical analysis was performed with a 2-way unpaired t-test. RESULTS: No toxic effect of the three elements on the growth of the animals was observed. A significant increase of histological features of transformation was observed in thyroid follicular cells of rats treated with B, Cd, and Mo compared with those of control group. These abnormalities were associated with decreased iodine content in the thyroid. CONCLUSIONS: This study provides the evidence that slightly increased environmental concentrations of B, Cd, and Mo can accelerate the appearance of transformation marks in the thyroid gland of hypothyroid rats.

Infliximab does not increase colonic cancer risk associated to murine chronic colitis.

AIM: To explore the influence of Infliximab (IFX) on cancer progression in a murine model of colonic cancer associated to chronic colitis. METHODS: AOM/DSS model was induced in C57BL/6 mice. Mice were injected with IFX (5 mg/kg) during each DSS cycle while control mice received saline. Body weight, occult blood test and stool consistency were measured to calculate the disease activity index (DAI). Mice were sacrificed at week 10 and colons were analyzed macroscopically and microscopically for number of cancers and degree of inflammation. MTT assay was performed on CT26 to evaluate the potential IFX role on metabolic activity and proliferation. Cells were incubated with TNF-α or IFX or TNF-α plus IFX, and cell vitality was evaluated after 6, 24 and 48 h. The same setting was used after pre-incubation with TNF-α for 24 h. RESULTS: IFX significantly reduced DAI and body weight loss in mice compared with controls, preserving also colon length at sacrifice. Histological score was also reduced in treated mice. At macroscopic analysis, IFX treated mice showed a lower number of tumor lesions compared to controls. This was confirmed at microscopic analysis, although differences were not statistically significant. In vitro, IFX treated CT26 maintained similar proliferation ability at MTT test, both when exposed to IFX alone and when associated to TNF-α. CONCLUSION: IFX did not increase colonic cancer risk in AOM-DSS model of cancer on chronic colitis nor influence directly the proliferation of murine colon cancer epithelial cells.

A novel closed-chest porcine model of chronic ischemic heart failure suitable for experimental research in cardiovascular disease.

Cardiac pathologies are among the leading causes of mortality and morbidity in industrialized countries, with myocardial infarction (MI) representing one of the major conditions leading to heart failure (HF). Hitherto, the development of consistent, stable, and reproducible models of closed-chest MI in large animals, meeting the clinical realism of a patient with HF subsequent to chronic ischemic necrosis, has not been successful. We hereby report the design and ensuing application of a novel porcine experimental model of closed-chest chronic ischemia suitable for biomedical research, mimicking post-MI HF. We also emphasize the key procedural steps involved in replicating this unprecedented model, from femoral artery and vein catheterization to MI induction by permanent occlusion of the left anterior descending coronary artery through superselective deployment of platinum-nylon coils, as well as endomyocardial biopsy sampling for histologic analysis and cell harvesting. Our model could indeed represent a valuable contribution and tool for translational research, providing precious insights to understand and overcome the many hurdles concerning, and currently quenching, the preclinical steps mandatory for the clinical translation of new cardiovascular technologies for personalized HF treatments.

Effects of high-fat diet exposure during fetal life on type 2 diabetes development in the progeny.

Nutrition during fetal life is a critical factor contributing to diabetes development in adulthood. The aim of our study was to verify: 1) whether a high-fat (HF) diet in young adult mice induces alterations in beta-cell mass, proliferation, neogenesis, and apoptosis, as well as insulin sensitivity and secretion; 2) whether these alterations may be reversible after HF diet suspension; 3) the effects in a first (F1) and second generation (F2) of mice without direct exposure to a HF diet after birth. Type 2 diabetes developed in adult mice on a HF diet, in F1 mice that were HF diet-exposed during fetal or neonatal life, and in F2 mice whose mothers were HF diet-exposed during their fetal life. beta-cell mass, replication, and neogenesis were high in HF diet-exposed mice and decreased after diet suspension. beta-cell mass and replication remained high in F1 mice and decreased in F2 mice whose mothers were exposed to a HF diet. beta-cell neogenesis was present in adult mice on a HF diet and in F1 mice that were HF diet-exposed during fetal and/or neonatal life. We conclude that a HF diet during fetal life, particularly if combined with the same insult during the suckling period, can induce the type 2 diabetes phenotype, which can be directly transmitted to the progeny even in the absence of additional dietary insults.

Longitudinal study of cardiac electrical activity in anesthetized guinea pigs by contactless magnetocardiography.

Guinea pigs (GPs) are used for preclinical evaluation of electrophysiologic effects of new drugs, because their myocytes have human-like action potentials and ventricular repolarization's (VR) ion currents. This study was aimed to assess the reliability of magnetocardiographic (MCG) mapping for longitudinal studies of GP cardiac electrical activity. Eighteen anesthetized GPs were investigated with an unshielded 36-channel MCG instrumentation, at the age of 5 months (268.1 +/- 19 g). Twelve GPs survived and were restudied when 14 months old (595.6 +/- 90.5 g). RR, PR, QRS, QT(peak), QT(end), JT(peak), JT(end) and T(peak-end) intervals were measured from MCG waveforms. Magnetic field (MF) maps, equivalent current dipole (ECD) parameters and current density imaging were also analyzed. A significant prolongation of the PR (p < 0.05) and QRS (p < 0.001) intervals was found at 14 months. Gender-related differences of VR intervals were not significant. P(peak) and QRS(peak) MFs were similar in all animals, while T(peak) MF varied interindividually at 5 months and showed a rotation in some animals, at 14 months. The ECD strengths, measured at the P(peak), QRS(peak) and T(peak) were stronger (p < 0.01) at the age of 14 months than at 5 months. In contrast to findings in Wistar rats, age-related and gender-related differences of MCG VR parameters were not significant in GPs. Further work is necessary to clarify the variability of VR MF observed in healthy GPs.

Contactless magnetocardiographic mapping in anesthetized Wistar rats: evidence of age-related changes of cardiac electrical activity.

Magnetocardiography (MCG) is the recording of the magnetic field (MF) generated by cardiac electrophysiological activity. Because it is a contactless method, MCG is ideal for noninvasive cardiac mapping of small experimental animals. The aim of this study was to assess age-related changes of cardiac intervals and ventricular repolarization (VR) maps in intact rats by means of MCG mapping. Twenty-four adult Wistar rats (12 male and 12 female) were studied, under anesthesia, with the same unshielded 36-channel MCG instrumentation used for clinical recordings. Two sets of measurements were obtained from each animal: 1) at 5 mo of age (297.5 +/- 21 g body wt) and 2) at 14 mo of age (516.8 +/- 180 g body wt). RR and PR intervals, QRS segment, and QTpeak, QTend, JTpeak, JTend, and Tpeak-end were measured from MCG waveforms. MCG imaging was automatically obtained as MF maps and as inverse localization of cardiac sources with equivalent current dipole and effective magnetic dipole models. After 300 s of continuous recording were averaged, the signal-to-noise ratio was adequate for study of atrial and ventricular MF maps and for three-dimensional localization of the underlying cardiac sources. Clear-cut age-related differences in VR duration were demonstrated by significantly longer QTend, JTend, and Tpeak-end in older Wistar rats. Reproducible multisite noninvasive cardiac mapping of anesthetized rats is simpler with MCG methodology than with ECG recording. In addition, MCG mapping provides new information based on quantitative analysis of MF and equivalent sources. In this study, statistically significant age-dependent variations in VR intervals were found.

Equine ehrlichiosis in Italy.

The authors review equine granulocytic ehrlichiosis (EGE) in Italy from 1996 to 2002. In 1996, the first case of EGE has been observed in a horse affected with specific symptomatology (fever, lethargy, anorexia, limb edema, thrombocytopenia, and petechiae). In 1997, a seroepidemiological survey was performed in the province of Rome on 563 animals using IFAT. The authors describe the last case, which occurred on 2002 in a 15-year-old male, bay, half-breed, tick-infested horse. Clinical features included fever, lethargy, limb edema, icterus, leukocytopenia, and thrombocytopenia. Laboratory tests were positive for ELISA and IFAT and several morulae were seen in the cytoplasm of neutrophils in buffy coat smears. The authors think that in Italy the prevalence of EGE is underestimated because of the nonpathognomonic clinical symptoms, the diffusion of specific infected vector, and the nonroutine nature of specific laboratory tests.