Brucella invasion of human intestinal epithelial cells elicits a weak proinflammatory response but a significant CCL20 secretion

In spite of the frequent acquisition of Brucella infection by the oral route in humans, the interaction of the bacterium with cells of the intestinal mucosa has been poorly studied. Here, we show that different Brucella species can invade human colonic epithelial cell lines (Caco-2 and HT-29), in which only smooth species can replicate efficiently. Infection with smooth strains did not produce a significant cytotoxicity, while the rough strain RB51 was more cytotoxic. Infection of Caco-2 cells or HT-29 cells with either smooth or rough strains of Brucella did not result in an increased secretion of TNF-α, IL-1β, MCP-1, IL-10 or TGF-β as compared with uninfected controls, whereas all the infections induced the secretion of IL-8 and CCL20 by both cell types. The MCP-1 response to flagellin from Salmonella typhimurium was similar in Brucella-infected or uninfected cells, ruling out a bacterial inhibitory mechanism as a reason for the weak proinflammatory response. Infection did not modify ICAM-1 expression levels in Caco-2 cells, but increased them in HT-29 cells. These results suggest that Brucella induces only a weak proinflammatory response in gut epithelial cells, but produces a significant CCL20 secretion. The latter may be important for bacterial dissemination given the known ability of Brucella to survive in dendritic cells.Facultad de Ciencias Exacta

Successful pregnancy in a patient with short bowel syndrome after surgical rehabilitation and sGLP-2 treatment: novel report on endogenous GLP-2 levels at delivery and during breastfeeding

Pregnant patients with short bowel syndrome (SBS) and chronic intestinal failure (CIF) can successfully reach to term their pregnancies while on parenteral nutrition (PN) but with high rates of complications. The combination of rehabilitation surgery, combined with the use of novel treatment with enterohormones, especially semisynthetic glucagon-like peptide 2 (sGLP-2), has increased the chances to achieve intestinal sufficiency. Here, we report the case of a 33-year-old female with SBS/CIF (anatomy type 2), weaned off PN using sGLP-2 for 3.7 years, discontinued when she became pregnant. She was able to carry the pregnancy to term without any additional PN support. Considering that, we queried if the endogenous GLP-2 (eGLP-2) levels in this SBS patient, during the pregnancy and breastfeeding period, could be like those presented in healthy pregnant women and in non-pregnant SBS patients. Also, we inquired if there was any passage or increase in the plasmatic eGLP-2 from the fetus to the mother. Thus, we determined eGLP-2 levels in paired neonatal (cord blood) and maternal plasma samples from the SBS pregnant patient (n = 1), healthy pregnant women (controls, n = 2), and non-pregnant SBS patients (n = 12). The results indicated that the SBS pregnant patient showed higher eGLP-2 levels than non-SBS pregnant patients and healthy pregnant women along all the period studied. Furthermore, we found that the maternal sample had higher eGLP-2 levels than the neonatal sample, suggesting that fetal contribution to maternal eGLP2 levels would be minor. In conclusion, this study not only reports for the first time a case of a patient with SBS that was able to achieve intestinal adaptation after combining the use of autologous reconstructive surgery and sGLP-2, but also enlightens the possibility of carrying out an uneventful pregnancy and lactation without any nutritional support and remaining independent of sGLP-2.Fil: Gentilini, Maria Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; ArgentinaFil: Doeyo, M.. Fundación Favaloro; ArgentinaFil: Ortega, M.. Fundación Favaloro; ArgentinaFil: Illidge Perez, L.. Fundación Favaloro; ArgentinaFil: Rumbo, C.. Fundación Favaloro; ArgentinaFil: Arriola Benitez, Paula Constanza. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; ArgentinaFil: Crivelli, A.. Fundación Favaloro; ArgentinaFil: Rumbo, Martín. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Estudios Inmunológicos y Fisiopatológicos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Estudios Inmunológicos y Fisiopatológicos; ArgentinaFil: Solar, H.. Fundación Favaloro; ArgentinaFil: Gondolesi, Gabriel Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; Argentin

Brucella invasion of human intestinal epithelial cells elicits a weak proinflammatory response but a significant CCL20 secretion

In spite of the frequent acquisition of Brucella infection by the oral route in humans, the interaction of the bacterium with cells of the intestinal mucosa has been poorly studied. Here, we show that different Brucella species can invade human colonic epithelial cell lines (Caco-2 and HT-29), in which only smooth species can replicate efficiently. Infection with smooth strains did not produce a significant cytotoxicity, while the rough strain RB51 was more cytotoxic. Infection of Caco-2 cells or HT-29 cells with either smooth or rough strains of Brucella did not result in an increased secretion of TNF-α, IL-1β, MCP-1, IL-10 or TGF-β as compared with uninfected controls, whereas all the infections induced the secretion of IL-8 and CCL20 by both cell types. The MCP-1 response to flagellin from Salmonella typhimurium was similar in Brucella-infected or uninfected cells, ruling out a bacterial inhibitory mechanism as a reason for the weak proinflammatory response. Infection did not modify ICAM-1 expression levels in Caco-2 cells, but increased them in HT-29 cells. These results suggest that Brucella induces only a weak proinflammatory response in gut epithelial cells, but produces a significant CCL20 secretion. The latter may be important for bacterial dissemination given the known ability of Brucella to survive in dendritic cells.Facultad de Ciencias Exacta

Transgenic mouse model harboring the transcriptional fusion Ccl20-luciferase as a novel reporter of pro-inflammatory response

The chemokine CCL20, the unique ligand of CCR6 functions as an attractant of immune cells. Expression of CCL20 is induced by Toll-like Receptor (TLR) signaling or proinflammatory cytokine stimulation. However CCL20 is also constitutively produced at specific epithelial sites of mucosa. This expression profile is achieved by transcriptional regulation. In the present work we characterized regulatory features of mouse Ccl20 gene. Transcriptional fusions between the mouse Ccl20 promoter and the firefly luciferase (luc) encoding gene were constructed and assessed in in vitro and in vivo assays. We found that liver CCL20 expression and luciferase activity were upregulated by systemic administration of the TLR5 agonist flagellin. Using shRNA and dominant negative form specific for mouse TLR5, we showed that this expression was controlled by TLR5. To address in situ the regulation of gene activity, a transgenic mouse line harboring a functional Ccl20-luc fusion was generated. The luciferase expression was highly concordant with Ccl20 expression in different tissues. Our data indicate that the transgenic mouse model can be used to monitor activation of innate response in vivo.Laboratorio de Investigaciones del Sistema InmuneFacultad de Ciencias Exacta

Canonical and non-canonical inflammasome activation by outer membrane vesicles derived from Bordetella pertussis

Outer Membrane Vesicles (OMVs) derived from different Gram-negative bacteria have been proposed as an attractive vaccine platform because of their own immunogenic adjuvant properties. Pertussis or whooping cough is a highly contagious vaccine-preventable respiratory disease that resurged during the last decades in many countries. In response to the epidemiological situation, new boosters have been incorporated into vaccination schedules worldwide and new vaccine candidates have started to be designed. Particularly, our group designed a new pertussis vaccine candidate based on OMVs derived from Bordetella pertussis (BpOMVs). To continue with the characterization of the immune response induced by our OMV based vaccine candidate, this work aimed to investigate the ability of OMVs to activate the inflammasome pathway in macrophages. We observed that NLRP3, caspase-1/11, and gasdermin-D (GSDMD) are involved in inflammasome activation by BpOMVs. Moreover, we demonstrated that BpOMVs as well as transfected B. pertussis lipooligosaccharide (BpLOS) induce caspase-11 (Casp11) and guanylate-binding proteins (GBPs) dependent non-canonical inflammasome activation. Our results elucidate the mechanism by which BpOMVs trigger one central pathway of the innate response activation that is expected to skew the adaptive immune response elicited by BpOMVs vaccination.Fil: Elizagaray, Maia Lina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Estudios Inmunológicos y Fisiopatológicos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Estudios Inmunológicos y Fisiopatológicos; ArgentinaFil: Gomes, Marco Túlio R.. Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas; BrasilFil: Guimaraes, Erika S.. Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas; BrasilFil: Rumbo, Martín. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Estudios Inmunológicos y Fisiopatológicos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Estudios Inmunológicos y Fisiopatológicos; ArgentinaFil: Hozbor, Daniela Flavia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Biotecnología y Biología Molecular. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Biotecnología y Biología Molecular; ArgentinaFil: Oliveira, Sergio C.. Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas; BrasilFil: Moreno, Griselda Noemí. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Estudios Inmunológicos y Fisiopatológicos. Universidad Nacional de La Plata. Facultad de Ciencias Exactas. Instituto de Estudios Inmunológicos y Fisiopatológicos; Argentin

Fate assessment of commercial 2D MoS2 aqueous dispersions at physicochemical and toxicological level

The physicochemical properties and the toxicological potential of commercially available MoS2 nanoparticles with different lateral size and degradation stage were studied in the present research work. To achieve this, the structure and stoichiometry of fresh and old aqueous suspensions of micro-MoS2 and nano-MoS2 was analyzed by Raman, while x-ray photoelectron spectroscopy allowed to identify more quantitatively the nature of the formed oxidized species. A, the toxicological impact of the nanomaterials under analysis was studied using adenocarcinomic human alveolar basal epithelial cells (A549 cells) and the unicellular fungus S. cerevisiae as biological models. Cell viability assays and reactive oxygen species (ROS) determinations demonstrated different toxicity levels depending on the cellular model used and in function of the degradation state of the selected commercial nanoproducts. Both MoS2 nanoparticle types induced sublethal damage on the A549 cells though the increase of intracellular ROS levels, while comparable concentrations reduced the viability of yeast cells. In addition, the old MoS2 nanoparticles suspensions exhibited a higher toxicity for both human and yeast cells than the fresh ones. Our findings demonstrate that the fate assessment of nanomaterials is a critical aspect to increase the understanding on their characteristics and on their potential impact on biological systems along their life cycle

The effect of the COVID-19 pandemic in intestinal rehabilitation and transplant patients, initial results of an international survey

Introduction: On January 30, 2020 the World Health Organization (WHO) declared the 2019-CoV outbreak in China as a global public health emergency and subsequently, a pandemic on March 11th. It was considered that intestinal failure and intestinal transplant patients might have a higher risk of severe complications from the COVID-19 disease, multidisciplinary intestinal failure teams had to adapt their clinical approaches in order to keep this vulnerable group of patients as safe as possible during the pandemic; but data was lacking. Therefore, in order to improve our knowledge, we designed a voluntary, international survey aiming to address the impact of the COVID-19 disease in intestinal failure and transplant patients worldwide. Patient and Methods: A retrospective, observational, multicenter survey was sent to all centers registered at the Intestinal Rehabilitation and Transplant Association (IRTA). The survey contained three modules: the 1st one consisted of 14 questions about the hospital\u27s activity during the COVID-19 pandemic. The 2nd one, contained 43 questions, was about intestinal failure patient management and outcome and the 3rd one (52 questions) focused on intestinal transplant patients. We used the Google Form platform. We aim to present the preliminary results of the first module. Statistical analysis was performed with the IBM SPSS Statistic version 25.0® program. Results: 13/42 (41%) centers responded; including centers from France, Netherlands, Italy, United States, UK, Sweden, Germany and Argentina. Only 2 centers reported moratorium on intestinal (IT) or multivisceral transplant (MVT), with a mean of 3 months (±4) [Table 1]. Since the pandemic started, 2 institutions reported 4 patients with intestinal rehabilitation or on TPN diagnosed with COVID-19 while 7 centers hospitals claimed to have had 9 patients post-IT/MTV affected by the disease. While 7 centers had their routine follow up and \u27protocol biopsies\u27 in the post-IT/MTV affected, none reported higher rates of rejection or complications. At the same time, 8 centers (77%) were affected by a mean of 15% decrease in referrals for new evaluations of intestinal failure or transplantation (compared to 2019) [Figure 1]. All centers adapted to utilizing telemedicine to follow up on IT/MVT patients. Conclusions: Many aspects of healthcare have been impacted by the COVID-19 pandemic. The survey showed that the number of affected patients has been lower than expected, the reduced number of centers required transient moratorium of their activity, but a secondary observation was that despite the availability of telemedicine, and probably related to the lockdown, there has been a significant reduction in the referrals for evaluation of intestinal failure and transplant patients, that may have the deleterious effect of the delay of treatment in health care system

Modelo experimental murino de isquemia-reperfusión intestinal y su impacto en órganos remotos

La falla multiorgánica es una de las causas de morbi-mortalidad en pacientes con trasplante intestinal. La lesión por isquemia-reperfusión intestinal (IRI) provoca ruptura de la barrera del intestino con impacto en órganos distantes, ya sea por sepsis, traslocación bacteriana o por activar una respuesta inflamatoria sistémica Objetivos: evaluar la respuesta histológica del tejido intestinal, pulmonar y hepático luego de un período de 35 minutos de isquemia intestinal por clampeo de la arteria mesentérica superior (AMS) seguidos de 60 minutos de reperfusión.Facultad de Ciencias Médica

Contribution of Efflux Pumps, Porins, and B-Lactamases to Multidrug Resistance in Clinical Isolates of Acinetobacter baumannii

Weinvestigated the mechanisms of resistance to carbapenems, aminoglycosides, glycylcyclines, tetracyclines, and quinolones in 90 multiresistant clinical strains of Acinetobacter baumannii isolated from two genetically unrelated A. baumannii clones: clone PFGEROC- 1 (53 strains producing the OXA-58B-lactamase enzyme and 18 strains with the OXA-24B-lactamase) and clone PFGE-HUI-1 (19 strains susceptible to carbapenems).Weused real-time reverse transcriptase PCR to correlate antimicrobial resistance (MICs) with expression of genes encoding chromosomalB-lactamases (AmpC and OXA-51), porins (OmpA, CarO, Omp33, Dcap-like, OprB, Omp25, OprC, OprD, and OmpW), and proteins integral to six efflux systems (AdeABC, AdeIJK, AdeFGH, CraA, AbeM, and AmvA). Overexpression of the AdeABC system (level of expression relative to that by A. baumannii ATCC 17978, 30- to 45-fold) was significantly associated with resistance to tigecycline, minocycline, and gentamicin and other biological functions. However, hyperexpression of the AdeIJK efflux pump (level of expression relative to that by A. baumannii ATCC 17978, 8- to 10-fold) was significantly associated only with resistance to tigecycline and minocycline (to which the TetB efflux system also contributed). TetB and TetA(39) efflux pumps were detected in clinical strains and were associated with resistance to tetracyclines and doxycycline. The absence of the AdeABC system and the lack of expression of other mechanisms suggest that tigecycline-resistant strains of the PFGE-HUI-1 clone may be associated with a novel resistance-nodulation-cell efflux pump (decreased MICs in the presence of the inhibitor Phe-ArgB-naphthylamide dihydrochloride) and the TetA(39) system

Small bowel transplantation in rats, a multicenter experience summarizing the pitfalls to be overcome

Small Bowel transplantation in rats is a highly complex microsurgical procedure because several technical complications may lead to recipient mortality and transplant failure. Our aim was to report the most common complications associated with orthotopic and heterotopic intestinal transplantation in rats in order to identify the “pitfalls” of the procedure and prevent them. A retrospective multicenter study was performed. All participant centers have established rodent transplant procedures and trained surgeons. Two hundred ninety-three complications from 264 unsuccessful intestinal transplants were reported, representing an overall failure rate of 15% of the procedures performed. Recipient complications were most frequent than donor (257 vs. 36 p<0.0001). Excessive surgical time (11/36); severe hemorrhage (12/36) and inappropriate infusion of the preservation solution in the intestinal graft (11/36) were the most common donor complications. Arterial anastomosis bleeding (50/257), venous anastomosis bleeding (35/257) and portal vein stenosis (26/257) were the most common intraoperative complications in the recipient. To maximize success rate, surgeons should optimize time and avoid bleeding during graft dissection in the donor surgery. After performing a bloodless vascular anastomosis an adequate post-operative management of the animal is mandatory to guarantee survival.Fil: Stringa, Pablo Luis. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; Argentina. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; ArgentinaFil: Andrés Moreno, Ane M.. La Paz University Hospital; EspañaFil: Lausada, Natalia Raquel. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; ArgentinaFil: Pastor Oliver, Cristina. Biomedical Research Center from Aragón; EspañaFil: Abate, Juan C.. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; ArgentinaFil: Vecchio, Leandro. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; ArgentinaFil: Rumbo, Martín. Comisión Nacional de Investigación Científica y Tecnológica; Chile. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; ArgentinaFil: Navarro Zorraquino, Marta. Biomedical Research Center from Aragón; EspañaFil: Hernández Oliveros, Francisco. La Paz University Hospital; EspañaFil: Gondolesi, Gabriel Eduardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Medicina Traslacional, Trasplante y Bioingeniería. Fundación Favaloro. Instituto de Medicina Traslacional, Trasplante y Bioingeniería; Argentina. Universidad Nacional de la Plata. Facultad de Ciencias Médicas. Laboratorio de Transplante de Órganos; Argentin