Development of macrophages in the intestine

Macrophages (mφ) are one the most numerous leukocytes present in the healthy gut and contribute to both harmful and beneficial immune reactions. In the colon, mφ are exposed continuously to large amounts of material from the environment, including harmful agents such as invasive bacteria, viruses and parasites, as well as harmless materials such as food proteins and the commensal bacteria which inhabit the healthy intestine. As a result, mφ play an important role in helping defend the intestine against harmful invaders. However if these cells make similar reactions to harmless food proteins or commensal bacteria, it would be both wasteful and detrimental, likely leading to inflammatory diseases such as coeliac disease and Crohn’s disease. Several genes, which underlie susceptibility to Crohn’s disease are involved in controlling how macrophages respond to the microbiota, with considerable evidence indicating that this reflects a loss of the normal unresponsiveness that characterises intestinal macrophages in the healthy intestine. One of the most significant aspects of the epidemiology of Crohn’s disease is a particularly rapid increase in its incidence in childhood, suggesting that the first encounters between the microbiota and intestinal macrophages may be of critical importance in determining disease susceptibility. Given this link, it is essential that we elucidate the processes controlling macrophage seeding and development in the intestine and this was an aim of this thesis. In the adult healthy colon, two main mφ subsets can be identified: A dominant and homogenous one, made up of mature mφ, which express high levels of F4/80, MHC II, CX3CR1, are CD11bint/+, highly phagocytic and produce high amounts of IL10. The second mφ group is relatively smaller and is much more heterogeneous. These cells express intermediate levels of F4/80 and CX3CR1, are CD11b+ and can be divided into 3 subsets based on their levels of Ly6C and MHC II. These subsets represent a maturation continuum towards the mature mφ phenotype. Recent reports have suggested that resident macrophages in healthy tissues may be derived from yolk-sac and/or foetal liver precursors that seed tissues during development and subsequently self-renew locally. In contrast, it is proposed that macrophages in inflammation are generated by recruitment of blood monocytes, raising the possibility that these different origins could be exploited in therapy. However none of these studies have examined macrophages in the intestine and recent work in our laboratory has suggested that monocytes may be the precursors of macrophages in both healthy and inflamed gut of adult mice. Therefore, the aims of this thesis were to investigate the development of murine colonic mφ from birth until adulthood, examining the relative roles of the yolk sac, foetal liver and bone marrow monocytes, exploring their functions and comparing them with the well-characterised adult mφ. In addition, I also examined how mφ phenotype and functions are influenced by the microbiota using broad-spectrum antibiotics and germ free mice. Lastly, I examined the role of fractalkine and its receptor CX3CR1 in defining the development and functions of intestinal macrophages. Development of macrophages in early life The initial characterisation and comparison of colonic mφ subsets is included in Chapter 3. In this chapter, I describe a series of experiments adapting existing protocols and techniques used for examining the adult murine intestine in order to analyse the origin, phenotype and functions of murine colonic macrophages from late foetal life through to adulthood. These studies found that intestinal mφ are present before birth, with similar levels of phagocytic ability and IL10, TNFα and CD163 mRNA expression to the adult. However, the numbers and phenotype of mφ in the intestine do not reach the adult level until the 3rd week of postnatal life. This phenomenon appears to reflect the de novo recruitment of blood monocytes in a CCR2-dependent fashion at this time and throughout adult life, but not at early stages of life. In the colon of newborn mice, two macrophage populations can be observed and are clearly differentiated based on their F4/80 and CD11b expression: F4/80hi CD11bint/+ and F4/80lo CD11b+. Interestingly, unlike adult colonic F4/80hi mφ, the majority of F4/80hi neonatal cells do not express MHC II, however they gradually express this molecule as they age. In addition to acquiring MHC II expression, the two populations in the newborn colon gradually merge and from the 3rd week of life it is difficult to discriminate them reliably. My experiments show that both mφ subsets proliferate actively during the first 2 weeks of life, but this is later reduced and maintained at low levels indicating that there is no self-renewal of mature mφ. Moreover, fate-mapping analysis carried out in collaboration with Professor Frederic Geissmann, showed that yolk sac-derived precursors contribute only minimally to the pool of colonic mφ, even at early life stages. Conversely, additional fate mapping studies suggested that most intestinal macrophages are derived from Flt3+ progenitors. Taken together, the results in this chapter demonstrate that blood monocytes are vital in replenishing the intestinal macrophage pool in the steady state, setting them apart from other tissue macrophages, which derive from primitive progenitors. Investigating the effect of the microbiota on intestinal macrophage subsets In Chapter 4, I assessed the effects of the commensal microbiota on intestinal mφ, using two different approaches: First, I assessed the function and gene expression of colonic macrophages following administration of broad-spectrum antibiotics. My results showed that this did not alter the numbers, phenotype, intracellular cytokine production or mRNA expression by macrophages. Several reasons may account for this, including dose/nature of antibiotics, length of administration or lifespan of macrophages. To overcome these issues, I compared the phenotype of colonic mφ in germ free (GF) and conventionally (CNV) reared mice of different ages in collaboration with Dr David Artis. Absolute absence of microbiota in GF mice severely impacted Ly6Chi monocyte recruitment to the colon, suggesting that constant recruitment of monocytes to the gut is at least in part due to the microbial burden. The biggest differences between GF and CNV mice were evident at 3 weeks of age, when GF mice had a much lower number and frequency of monocyte-derived cells than their CNV counterparts. By 12 weeks of age, Ly6Chi mφ populations from GF mice were partially restored, although the expression of MHC II by F4/80hi mφ remained reduced. Additionally, I FACS-purified F4/80hi cells from GF and CNV adults and sent RNA for microarray analysis, the results of which we are waiting to receive. This data will provide further information regarding how GF intestinal mφ differ from those found in conventional animals. Role of the CX3CL1-CX3CR1 axis in mφ development and function As mature colonic mφ express high levels of the chemokine receptor CX3CR1 (fractalkine), finally, in Chapter 5 I went on to investigate the role of CX3CL1-CX3CR1 axis in colonic lamina propria. In addition to the high expression of CX3CR1 by colonic mφ, its ligand, CX3CL1 has been reported to be expressed at high levels by the intestinal epithelium. Furthermore, as there is strong evidence that the CX3CL1-CX3CR1 axis may be involved in inflammation in several tissues, we hypothesised this axis might play a role in mφ function in the gut. To this end, I examined mφ phenotype, activation status and survival following in vitro co-culture of WT or CX3CR1-deficient bone marrow-derived mφ with an epithelial cell line modified to express either the soluble or membrane-bound forms of CX3CL1. I also examined the development of chemically induced colitis in CX3CR1-deficient mice. Finally, since it has been reported by the lab of Oliver Pabst, that the lack of CX3CR1 results in reduced IL10 production by intestinal mφ, I compared the ability of WT and CX3CR1-deficient mice to prime T cells after being fed with ovalbumin together with an adjuvant. The results from this chapter failed to show any definitive role of the CX3CL1-CX3CR1 axis in mφ function in either the steady state or in the setting of inflammation. My in vitro studies did not show any significant difference between WT and CX3CR1 deficient intestinal mφ in terms of survival, or co-stimulatory molecule expression, nor did bone marrow mφ (BMM) from CX3CR1 KO mice show differences in co-stimulatory molecules and pro-inflammatory cytokine production with or without stimulation by LPS. Moreover, the responses of wild type BMM were not altered by exposure to exogenous CX3CL1 either in soluble form, or when expressed as a transmembrane form by epithelial cells. The in vivo assessment of CX3CR1 during inflammation, Ly6Chi CX3CR1int cells increased after 4 days on DSS, however, the lack of CX3CR1 failed to confer protection from colitis in a consistent manner, suggesting that there may be more factors responsible for colonic inflammation apart from the CX3CL1-CX3CR1 axis. Taken together, the results of this thesis highlight that important cellular changes take place during the development of mφ in the intestine. In addition, the presence or absence of microbiota plays a crucial role in this development with acquisition of MHC II depending at least in part on the presence of microbes. Microarray data obtained from purified F4/80hi mφ populations of GF and CNV mice may reveal interesting differences and suggest how mφ phenotype and function may be regulated by the microbiota. Finally, I have shown that the CX3CL1-CX3CR1 axis plays a redundant role in the regulation of intestinal mφ phenotype and function with mφ from CX3CR1-deficient animals appearing to function normally in both health and disease

Pancreatic metastases from renal cell carcinoma. Postoperative outcome after surgical treatment in a Spanish multicenter study (PANMEKID)

Background: Renal Cell Carcinoma (RCC) occasionally spreads to the pancreas. The purpose of our study is to evaluate the short and long-term results of a multicenter series in order to determine the effect of surgical treatment on the prognosis of these patients. Methods: Multicenter retrospective study of patients undergoing surgery for RCC pancreatic metastases, from January 2010 to May 2020. Variables related to the primary tumor, demographics, clinical characteristics of metastasis, location in the pancreas, type of pancreatic resection performed and data on short and long-term evolution after pancreatic resection were collected. Results: The study included 116 patients. The mean time between nephrectomy and pancreatic metastases' resection was 87.35 months (ICR: 1.51-332.55). Distal pancreatectomy was the most performed technique employed (50 %). Postoperative morbidity was observed in 60.9 % of cases (Clavien-Dindo greater than IIIa in 14 %). The median follow-up time was 43 months (13-78). Overall survival (OS) rates at 1, 3, and 5 years were 96 %, 88 %, and 83 %, respectively. The disease-free survival (DFS) rate at 1, 3, and 5 years was 73 %, 49 %, and 35 %, respectively. Significant prognostic factors of relapse were a disease free interval of less than 10 years (2.05 [1.13-3.72], p 0.02) and a history of previous extrapancreatic metastasis (2.44 [1.22-4.86], p 0.01). Conclusions: Pancreatic resection if metastatic RCC is found in the pancreas is warranted to achieve higher overall survival and disease-free survival, even if extrapancreatic metastases were previously removed. The existence of intrapancreatic multifocal compromise does not always warrant the performance of a total pancreatectomy in order to improve survival. (C) 2021 The Authors. Published by Elsevier Ltd

Repeated pancreatic resection for pancreatic metastases from renal cell Carcinoma: A Spanish multicenter study (PANMEKID)

Background and objectives: Recurrent isolated pancreatic metastasis from Renal Cell Carcinoma (RCC) after pancreatic resection is rare. The purpose of our study is to describe a series of cases of relapse of pancreatic metastasis from renal cancer in the pancreatic remnant and its surgical treatment with a repeated pancreatic resection, and to analyse the results of both overall and disease -free survival. Methods: Multicenter retrospective study of patients undergoing pancreatic resection for RCC pancreatic metastases, from January 2010 to May 2020. Patients were grouped into two groups depending on whether they received a single pancreatic resection (SPS) or iterative pancreatic resection. Data on short and long-term outcome after pancreatic resection were collected. Results: The study included 131 pancreatic resections performed in 116 patients. Thus, iterative pancreatic surgery (IPS) was performed in 15 patients. The mean length of time between the first pancreatic surgery and the second was 48.9 months (95 % CI: 22.2-56.9). There were no differences in the rate of postoperative complications. The DFS rates at 1, 3 and 5 years were 86 %, 78 % and 78 % vs 75 %, 50 % and 37 % in the IPS and SPS group respectively (p = 0.179). OS rates at 1, 3, 5 and 7 years were 100 %, 100 %, 100 % and 75 % in the IPS group vs 95 %, 85 %, 80 % and 68 % in the SPS group (p = 0.895). Conclusion: Repeated pancreatic resection in case of relapse of pancreatic metastasis of RCC in the pancreatic remnant is justified, since it achieves OS results similar to those obtained after the first resection

Mortality and pulmonary complications in patients undergoing surgery with perioperative SARS-CoV-2 infection: an international cohort study

Background: The impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on postoperative recovery needs to be understood to inform clinical decision making during and after the COVID-19 pandemic. This study reports 30-day mortality and pulmonary complication rates in patients with perioperative SARS-CoV-2 infection. Methods: This international, multicentre, cohort study at 235 hospitals in 24 countries included all patients undergoing surgery who had SARS-CoV-2 infection confirmed within 7 days before or 30 days after surgery. The primary outcome measure was 30-day postoperative mortality and was assessed in all enrolled patients. The main secondary outcome measure was pulmonary complications, defined as pneumonia, acute respiratory distress syndrome, or unexpected postoperative ventilation. Findings: This analysis includes 1128 patients who had surgery between Jan 1 and March 31, 2020, of whom 835 (74·0%) had emergency surgery and 280 (24·8%) had elective surgery. SARS-CoV-2 infection was confirmed preoperatively in 294 (26·1%) patients. 30-day mortality was 23·8% (268 of 1128). Pulmonary complications occurred in 577 (51·2%) of 1128 patients; 30-day mortality in these patients was 38·0% (219 of 577), accounting for 81·7% (219 of 268) of all deaths. In adjusted analyses, 30-day mortality was associated with male sex (odds ratio 1·75 [95% CI 1·28–2·40], p\textless0·0001), age 70 years or older versus younger than 70 years (2·30 [1·65–3·22], p\textless0·0001), American Society of Anesthesiologists grades 3–5 versus grades 1–2 (2·35 [1·57–3·53], p\textless0·0001), malignant versus benign or obstetric diagnosis (1·55 [1·01–2·39], p=0·046), emergency versus elective surgery (1·67 [1·06–2·63], p=0·026), and major versus minor surgery (1·52 [1·01–2·31], p=0·047). Interpretation: Postoperative pulmonary complications occur in half of patients with perioperative SARS-CoV-2 infection and are associated with high mortality. Thresholds for surgery during the COVID-19 pandemic should be higher than during normal practice, particularly in men aged 70 years and older. Consideration should be given for postponing non-urgent procedures and promoting non-operative treatment to delay or avoid the need for surgery. Funding: National Institute for Health Research (NIHR), Association of Coloproctology of Great Britain and Ireland, Bowel and Cancer Research, Bowel Disease Research Foundation, Association of Upper Gastrointestinal Surgeons, British Association of Surgical Oncology, British Gynaecological Cancer Society, European Society of Coloproctology, NIHR Academy, Sarcoma UK, Vascular Society for Great Britain and Ireland, and Yorkshire Cancer Research

Salmonella enterica serovar Typhimurium travels to mesenteric lymph nodes both with host cells, and autonomously

Salmonella infection is a globally important cause of gastroenteritis and systemic disease, and is a useful tool to study immune responses in the intestine. Although mechanisms leading to immune responses against Salmonella have been extensively studied, questions remain about how bacteria travel from the intestinal mucosa to the mesenteric lymph nodes (MLN), a key site for antigen presentation. Here, we used a mouse model of infection with Salmonella enterica serovar Typhimurium (STM) to identify changes in intestinal immune cells induced during early infection. We then used fluorescently-labelled STM to identify interactions with immune cells, from the site of infection, through migration in lymph, to the MLN. We show that viable STM can be carried in the lymph by any subset of migrating dendritic cells, but not by macrophages. Moreover, approximately half of the STM in lymph are not associated with cells at all, and travel autonomously. Within the MLN, STM associates with dendritic cells and B cells, but predominantly with MLN-resident macrophages. In conclusion, we describe the routes used by STM to spread systemically in the period immediately after infection. This deeper understanding of the infection process could open new avenues for controlling it

Jornadas Nacionales de Robótica y Bioingeniería 2023: Libro de actas

Las Jornadas de Robótica y Bioingeniería de 2023 tienen lugar en la Escuela Técnica Superior de Ingeniería Industrial de la Universidad Politécnica de IVIadrid, entre los días 14 y 16 de junio de 2023. En este evento propiciado por el Comité Español de Automática (CEA) tiene lugar la celebración conjunta de las XII Jornadas Nacionales de Robótica y el XIV Simposio CEA de Bioingeniería. Las Jornadas Nacionales de Robótica es un evento promovido por el Grupo Temático de Robótica (GTRob) de CEA para dar visibilidad y mostrar las actividades desarrolladas en el ámbito de la investigación y transferencia tecnológica en robótica. Asimismo, el propósito de Simposio de Bioingeniería, que cumple ahora su decimocuarta dicción, es el de proporcionar un espacio de encuentro entre investigadores, desabolladores, personal clínico, alumnos, industriales, profesionales en general e incluso usuarios que realicen su actividad en el ámbito de la bioingeniería. Estos eventos se han celebrado de forma conjunta en la anualidad 2023. Esto ha permitido aunar y congregar un elevado número de participantes tanto de la temática robótica como de bioingeniería (investigadores, profesores, desabolladores y profesionales en general), que ha posibilitado establecer puntos de encuentro, sinergias y colaboraciones entre ambos. El programa de las jornadas aúna comunicaciones científicas de los últimos resultados de investigación obtenidos, por los grupos a nivel español más representativos dentro de la temática de robótica y bioingeniería, así como mesas redondas y conferencias en las que se debatirán los temas de mayor interés en la actualidad. En relación con las comunicaciones científicas presentadas al evento, se ha recibido un total de 46 ponencias, lo que sin duda alguna refleja el alto interés de la comunidad científica en las Jornadas de Robótica y Bioingeniería. Estos trabajos serán expuestos y presentados a lo largo de un total de 10 sesiones, distribuidas durante los diferentes días de las Jornadas. Las temáticas de los trabajos cubren los principales retos científicos relacionados con la robótica y la bioingeniería: robótica aérea, submarina, terrestre, percepción del entorno, manipulación, robótica social, robótica médica, teleoperación, procesamiento de señales biológicos, neurorehabilitación etc. Confiamos, y estamos seguros de ello, que el desarrollo de las jornadas sea completamente productivo no solo para los participantes en las Jornadas que podrán establecer nuevos lazos y relaciones fructíferas entre los diferentes grupos, sino también aquellos investigadores que no hayan podido asistir. Este documento que integra y recoge todas las comunicaciones científicas permitirá un análisis más detallado de cada una de las mismas

Revolution: Museo de las estrellas un paseo por la fama : Hollywood

Convocatoria proyectos de innovación de Extremadura 2020/2021Se describe un proyecto llevado a cabo entre 13 centros educativos extremeños que consistió en desarrollar cinco unidades de trabajo gamificadas, cinco historias detectivescas con misterios por resolver, donde se ponían a prueba las habilidades de lógica, la capacidad de observación, de concentración y de atención de los alumnos. Los objetivos principales de la propuesta fueron: promover la puesta en práctica de proyectos intercentros; impulsar pedagogías activas; desarrollar la competencia digital a través del uso de las pedagogías emergentes lo que ha permitido llevar a cabo una enseñanza presencial, híbrida y virtual y atender a la diversidadExtremaduraES

Global attitudes in the management of acute appendicitis during COVID-19 pandemic: ACIE Appy Study

Background: Surgical strategies are being adapted to face the COVID-19 pandemic. Recommendations on the management of acute appendicitis have been based on expert opinion, but very little evidence is available. This study addressed that dearth with a snapshot of worldwide approaches to appendicitis. Methods: The Association of Italian Surgeons in Europe designed an online survey to assess the current attitude of surgeons globally regarding the management of patients with acute appendicitis during the pandemic. Questions were divided into baseline information, hospital organization and screening, personal protective equipment, management and surgical approach, and patient presentation before versus during the pandemic. Results: Of 744 answers, 709 (from 66 countries) were complete and were included in the analysis. Most hospitals were treating both patients with and those without COVID. There was variation in screening indications and modality used, with chest X-ray plus molecular testing (PCR) being the commonest (19\ub78 per cent). Conservative management of complicated and uncomplicated appendicitis was used by 6\ub76 and 2\ub74 per cent respectively before, but 23\ub77 and 5\ub73 per cent, during the pandemic (both P < 0\ub7001). One-third changed their approach from laparoscopic to open surgery owing to the popular (but evidence-lacking) advice from expert groups during the initial phase of the pandemic. No agreement on how to filter surgical smoke plume during laparoscopy was identified. There was an overall reduction in the number of patients admitted with appendicitis and one-third felt that patients who did present had more severe appendicitis than they usually observe. Conclusion: Conservative management of mild appendicitis has been possible during the pandemic. The fact that some surgeons switched to open appendicectomy may reflect the poor guidelines that emanated in the early phase of SARS-CoV-2