O recrutamento de neutrófilos para o sítio da infecção é um evento crucial para o combate aos microrganismos e sobrevivência na sepse. A migração destes polimorfonucleares é dirigida através de um gradiente quimiotático por meio do reconhecimento de quimiocinas por receptores acoplados a proteína G (GPCRs), os quais são regulados por quinases específicas (GRKs). Estudos prévios demonstraram que na sepse ocorre uma falência na migração de neutrófilos para o foco infeccioso em função da dessensibilização de receptores quimiotáticos via GRKs induzida pela ativação de receptores toll-like (TLRs), TLR2 e TLR4. Apesar de a ausência de TLR9 em células dendriticas ter sido relacionada a maior sobrevivência de camundongos sépticos, o papel do TLR9 atuando diretamente em neutrófilos não foi avaliado. Objetivando preencher esta lacuna, propôs-se avaliar o papel direto de TLR9 na falência de migração de neutrófilos na sepse. Os camundongos TLR9-/- apresentaram maior sobrevivência a sepse polimicrobiana avaliada por meio do modelo de ligadura e perfuração do ceco (CLP). A deficiência de TLR9 também acarretou em aumento na migração de neutrófilos para o foco da infecção, menor seqüestro de neutrófilos no pulmão, bem como, menor número de bactérias no lavado peritoneal e sangue. A ativação de TLR9 por oligodeoxinucleotídeo contendo o dinucleotídeo CpG não metilado (ODN CpG) nos neutrófilos reduziu a quimiotaxia destes em direção a quimiocina CXCL2 e expressão do receptor quimiotático CXCR2. Além disso, neutrófilos estimulados com ODN CpG apresentaram aumento na expressão da quinase tipo 2 relacionada a receptores acoplados a proteína G (GRK2). Dessa forma, a ativação de TLR9 em neutrófilos circulantes no sangue é prejudicial na sepse por reduzir a quimiotaxia destes para o foco da infecção ao induzir a dessensibilização de CXCR2 via GRK2. / The recruitment of neutrophils to the site of infection is a crucial event for combating the microorganisms and survival on sepsis. The neutrophil migration is directed by a chemotactic gradient through the recognition of chemokines by G protein-coupled receptors (GPCRs), which are regulated by specific kinases (GRKs). Previous studies have shown a failure of neutrophil migration into infectious focus on sepsis due to chemotactic receptor desensitization via GRKs induced by activation of toll- like receptors (TLRs), TLR2 and TLR4. Despite the absence of activation of TLR9 in dendritic cells have been related to increase survival of septic mice, the role of TLR9 acting directly on neutrophils was not evaluated. We proposed to verify the direct role of TLR9 in the failure of neutrophil migration on sepsis. The TLR9 knockout mice (TLR9-/-) showed high survival to polymicrobial sepsis using cecal ligation and puncture model (CLP). TLR9-/- mice had high neutrophil migration to the focus of infection, low neutrophil sequestration in the lung, as well as, few bacteria in the peritoneal exudates and blood. The activation of TLR9 by oligodeoxinucleotide containing unmethylated dinucleotide CpG (CpG ODN) in neutrophils also reduced chemotaxis toward CXCL2 and the expression of chemokine receptor CXCR2. In addition, neutrophils stimulated with CpG ODN showed increased expression of kinase-related G protein-coupled receptor type 2 (GRK2). Thus, the activation of TLR9 in blood circulating neutrophils is harmful on sepsis by reducing their chemotaxis into the site of the infection by inducing CXCR2 desensitization via GRK2.

O recrutamento de neutrófilos para o sítio da infecção é um evento crucial para o combate aos microrganismos e sobrevivência na sepse. A migração destes polimorfonucleares é dirigida através de um gradiente quimiotático por meio do reconhecimento de quimiocinas por receptores acoplados a proteína G (GPCRs), os quais são regulados por quinases específicas (GRKs). Estudos prévios demonstraram que na sepse ocorre uma falência na migração de neutrófilos para o foco infeccioso em função da dessensibilização de receptores quimiotáticos via GRKs induzida pela ativação de receptores toll-like (TLRs), TLR2 e TLR4. Apesar de a ausência de TLR9 em células dendriticas ter sido relacionada a maior sobrevivência de camundongos sépticos, o papel do TLR9 atuando diretamente em neutrófilos não foi avaliado. Objetivando preencher esta lacuna, propôs-se avaliar o papel direto de TLR9 na falência de migração de neutrófilos na sepse. Os camundongos TLR9-/- apresentaram maior sobrevivência a sepse polimicrobiana avaliada por meio do modelo de ligadura e perfuração do ceco (CLP). A deficiência de TLR9 também acarretou em aumento na migração de neutrófilos para o foco da infecção, menor seqüestro de neutrófilos no pulmão, bem como, menor número de bactérias no lavado peritoneal e sangue. A ativação de TLR9 por oligodeoxinucleotídeo contendo o dinucleotídeo CpG não metilado (ODN CpG) nos neutrófilos reduziu a quimiotaxia destes em direção a quimiocina CXCL2 e expressão do receptor quimiotático CXCR2. Além disso, neutrófilos estimulados com ODN CpG apresentaram aumento na expressão da quinase tipo 2 relacionada a receptores acoplados a proteína G (GRK2). Dessa forma, a ativação de TLR9 em neutrófilos circulantes no sangue é prejudicial na sepse por reduzir a quimiotaxia destes para o foco da infecção ao induzir a dessensibilização de CXCR2 via GRK2. / The recruitment of neutrophils to the site of infection is a crucial event for combating the microorganisms and survival on sepsis. The neutrophil migration is directed by a chemotactic gradient through the recognition of chemokines by G protein-coupled receptors (GPCRs), which are regulated by specific kinases (GRKs). Previous studies have shown a failure of neutrophil migration into infectious focus on sepsis due to chemotactic receptor desensitization via GRKs induced by activation of toll- like receptors (TLRs), TLR2 and TLR4. Despite the absence of activation of TLR9 in dendritic cells have been related to increase survival of septic mice, the role of TLR9 acting directly on neutrophils was not evaluated. We proposed to verify the direct role of TLR9 in the failure of neutrophil migration on sepsis. The TLR9 knockout mice (TLR9-/-) showed high survival to polymicrobial sepsis using cecal ligation and puncture model (CLP). TLR9-/- mice had high neutrophil migration to the focus of infection, low neutrophil sequestration in the lung, as well as, few bacteria in the peritoneal exudates and blood. The activation of TLR9 by oligodeoxinucleotide containing unmethylated dinucleotide CpG (CpG ODN) in neutrophils also reduced chemotaxis toward CXCL2 and the expression of chemokine receptor CXCR2. In addition, neutrophils stimulated with CpG ODN showed increased expression of kinase-related G protein-coupled receptor type 2 (GRK2). Thus, the activation of TLR9 in blood circulating neutrophils is harmful on sepsis by reducing their chemotaxis into the site of the infection by inducing CXCR2 desensitization via GRK2.

L'encéphalopathie associée au sepsis (EAS) est définie comme un dysfonctionnement cérébral diffus induit par une réponse systémique à une infection. Chez les patients septiques, l'imagerie par résonance magnétique (IRM) a indiqué à la fois des anomalies de la substance grise (SG) et blanche (SB) associées à des troubles cognitifs graves, y compris le delirium. Pour améliorer notre compréhension des changements hémodynamiques, métaboliques et structuraux associés au sepsis, différentes séquences d'IRM ont été réalisées chez des rats ayant subi une injection ip de solution saline ou de lipopolysaccharide bactérien (LPS) 2,5h plus tôt ou une ligature et ponction caecale 24h plus tôt. Après ip LPS, l'IRM de contraste de phase a été réalisée pour étudier le flux des artères cérébrales antérieures et moyennes et le marquage des spins artériels (ASL) pour étudier la perfusion des structures cérébrales de la SB et SG. Des séquences d'imagerie par diffusion pondérée (DWI) ont été utilisées pour évaluer les changements structurels. Après la chirurgie CLP, ASL a été utilisé pour étudier les changements de la microcirculation. L'imagerie pondérée en T2, l'imagerie du tenseur de diffusion (DTI) et les statistiques spatiales basées sur les faisceaux (TBSS) ont été réalisées pour caractériser les événements structurels dans différentes structures cérébrales. Après imagerie, les animaux ont été sacrifiés et leur cerveau a été traité pour l'histologie afin de détecter l'enzyme synthétisant les prostaglandines vasoactives cyclooxygénase-2 (COX-2) et le canal hydrique astrocytaire aquaporin-4 (AQP4) dont l'expression peut être régulée à la hausse, évaluer la présence d'immunoglobulines périvasculaires (Ig) indiquant une rupture de la barrière hémato-encéphalique (BHE) et étudier la morphologie des glies puisque la microglie et l’astroglie changent de morphologie lors des conditions inflammatoires. L'IRM n'a indiqué aucun changement hémodynamique dans la substance grise après l'administration de ip LPS, alors qu'une perfusion cérébrale accrue a été montrée au niveau du corps calleux comme indiqué par l'ASL. DTI a indiqué une augmentation de la diffusion des molécules d’eau parallèlement aux fibres du corps calleux. Ces changements étaient accompagnés d'une dégradation de BHE dans la SB ainsi que la substance grise corticale et striatale adjacente tel est indiqué par la présence périvasculaire d'IgG, sans aucun changement majeur de COX-2 vasculaire ou de morphologie des glies du coprs calleux. Le dysfonctionnement du SNC induit par le sepsis a résulté en une augmentation du contraste pondéré en T2 dans le cortex, le striatum et la base du cerveau, une diminution de la perfusion sanguine dans le cortex et une augmentation de la diffusion hydrique du corps calleux et du striatum ventral. Ces changements ont été associés dans la SB à des modifications de la morphologie des glies et dans la substance grise à une expression constitutive de COX-2 et AQP4 plus faible dans le cortex cérébral. La comparaison entre CLP ayant subit ou non une IRM sous anesthésie à l'isoflurane a montré une réponse inflammatoire réduite tel est indiqué par l'expression de COX- 2, une activation réduite des glies ainsi qu’une lésion réduite de la BHE dans le CLP subissant une IRM sous anesthésie. Collectivement, nos résultats suggèrent que des changements hémodynamiques peuvent survenir en l'absence de flux altéré dans les artères irriguant le cerveau antérieur. Ensuite, l'altération de la structure de la SB est une étape précoce de la pathogenèse de l’EAS qui peut résulter soit de la dégradation de la BHE, soit de l'activation des glies. Cette étude sous-tend l'effet délétère d'une seule exposition à l'anesthésie à l'isoflurane qui peut être atténuée par une seconde exposition chez les rats ayant subi une laparotomie ainsi que les effets de l'inflammation systémique induite par le CLP sur les glies pouvant être atténués par imagerie sous anesthésie à l'isoflurane. / Sepsis-associated encephalopathy (SAE) refers to central nervous system dysfunction during the systemic inflammatory response to infection. In septic patients with encephalopathy MRI has indicated both gray and white matter abnormalities that were associated with worse cognitive outcome including delirium. To improve our understanding of sepsis-associated hemodynamic, metabolic, and structural changes, different MRI sequences were performed in rats that either underwent an i.p injection of saline or bacterial lipopolysaccharide (LPS) 2.5h earlier or cecal ligation and puncture (CLP) 24h earlier. After ip LPS, phase contrast MRI was performed to study anterior and middle cerebral arteries flow and Arterial Spin Labeling (ASL) to study perfusion of white and grey matter brain structures. Diffusion Weighted Imaging (DWI) sequences was used to assess structural changes. After CLP surgery, ASL was used to study microcirculation changes. T2-Weighted Imaging, Diffusion Tensor Imaging (DTI) and tract-based spatial statistics (TBSS) were performed to characterize structural events in different brain structures. After imaging, animals were sacrificed and their brains processed for histology to detect the vasoactive prostaglandin-synthesizing enzyme cyclooxygenase-2 (COX-2) and the astrocytic aquaporin-4 water channel (AQP4) the expression of which can be upregulated during inflammation, to assess the presence of perivascular immunoglobulins (Ig) indicating blood-brain barrier (BBB) leakage and to study glia cell morphology as both microglia and astrocytes are known to change their morphology in inflammatory conditions. Magnetic resonance rat brain imaging indicated no hemodynamic changes in the grey matter after ip LPS administration while an increased CBF was shown in corpus callosum white matter as indicated by ASL. DTI indicated increased water diffusion parallel to fibers of the corpus callosum white matter. These changes were accompanied by BBB breakdown in the white matter and adjacent cortical and striatal grey matter as indicated by the perivascular presence of IgG, but no major changes in vascular COX-2 or white matter glia cell morphology. CLP induced sepsis-associated CNS dysfunction resulted in higher T2-weighted contrast intensities in the cortex, striatum and base of the brain, decreased blood perfusion distribution to the cortex and increased water diffusion in the corpus callosum and ventral striatum compared to sham surgery. These changes were associated in the white matter with modifications in glia cells morphology and in the grey matter with lower expression of constitutive COX-2 expression and AQP4 in the cerebral cortex. The comparison between CLP that underwent or not MRI under isoflurane anesthesia indicated reduced inflammatory response as indicated by COX-2 expression, reduced glia activation and reduced BBB damage in CLP that underwent MRI under isoflurane anesthesia. Collectively, our results suggest that hemodynamic changes may occur in the absence of altered flow in forebrain irrigating arteries. Then, altered white matter structure is an early step in SAE pathogenesis that may result either from BBB breakdown or glial cells activation. This study underlies the deleterious effects of a single exposure to isoflurane anesthesia that may be mitigated by a second exposure in sham-operated rats and the effects of CLP-induced systemic inflammation on glial cells that can be attenuated by imaging under isoflurane anesthesia.

[ES] El principal objetivo de esta tesis fue evaluar el valor nutritivo de la pulpa granulada de remolacha (PR) producida en diferentes zonas azucareras de España. Inicialmente se caracterizó la composición química de PR procedentes del norte y sur de España, cosechadas en invierno y verano, respectivamente. Se compararon la composición química y el contenido de ácidos grasos y aminoácidos de 12 muestras de PR, agrupadas según su procedencia. La composición de la materia seca (MS) de la PR, se caracteriza por un bajo contenido en proteína cruda (8.7%), dentro del 62% de la fibra neutro detergente (FND), pero rica en FND (39.4%) y fibra soluble en detergente neutro (45.7%), y bajo en grasa bruta (1.2%) y almidón (0.9%). El análisis de componentes principales (CP) de los principales principios inmediatos indicó que ya los dos primeros CP explican el 62% de la variabilidad en la composición de la PR. Los valores de los autovectores de los CP indicaron un importante peso del contenido en almidón y FND en el primer CP y del contenido en cenizas en el segundo CP. Posteriormente se determinó la digestibilidad de un pienso control con sustitución de 20% de PR de cinco orígenes distintos, y su posible efecto en el valor nutritivo de estas PR para conejos de engorde. Se utilizaron 90 conejos de 42 días de edad con 1.49 ± 0.208 kg de peso vivo, alojados en jaulas metabólicas individuales y con alimentación ad libitum. La digestibilidad de los componentes de los piensos se determinó por el método convencional in vivo de recolección fecal total. La digestibilidad de cada una de las pulpas de remolacha se determinó mediante el método de sustitución. No se apreció efecto de la sustitución de componentes del pienso control por el 20% de PR de diferentes procedencias en el consumo voluntario de los conejos. La inclusión de PR en las dietas aumentó la digestibilidad de materia seca (MS), materia orgánica, energía bruta y de todas las fracciones fibrosas, y disminuyó la digestibilidad de la proteína bruta. Respecto al efecto de la procedencia, se observaron diferencias significativas (P<0.05) en la digestibilidad de todas las fracciones fibrosas, siendo la PR recolectada en verano la de mayor digestibilidad. El origen de la PR también afectó su valor nutritivo, con valores de energía digestible entre 11.8 y 14.5 MJ/kg MS y de proteína digestible entre 38 y 66 g/kg MS. Finalmente, para evaluar el efecto del nivel de inclusión de pulpa de remolacha, con o sin melaza, sobre el rendimiento del crecimiento, se utilizaron un total de 470 conejos de 28 días de edad, con 612.2 ± 103.4 gr. de peso vivo. Los animales fueron asignados aleatoriamente a cinco tratamientos dietéticos: Control, sin pulpa de remolacha; PR20 y PR40, con 20 y 40% de pulpa de remolacha sin melaza, respectivamente; y PRM20 y PRM40, con 20 y 40% de pulpa de remolacha con melaza, respectivamente. El consumo diario de alimento (CDA) y la ganancia media diaria (GMD) se controlaron a los 28, 49 y 59 días de edad. También se determinaron las características de la canal y del tracto digestivo a los 59 días de edad. La mortalidad y la morbilidad fueron controladas diariamente. La mortalidad durante el período de crecimiento fue mayor en los grupos PRM que en los PR (+9.2%; P<0,05). A mayor inclusión de PR, con o sin melaza, menor CDA y GMD de los animales, así como el rendimiento a la canal, la proporción de hígado y el porcentaje de grasa disecable de sus canales. Sin embargo, la mejor eficiencia alimenticia durante los últimos 10 días se obtuvo con el grupo PRM40. Respecto a los parámetros digestivos, cuanto mayor fue la inclusión de pulpa de remolacha, con o sin melaza, mayor el peso del tracto gastrointestinal vacío y ciego. De hecho, una mayor inclusión de pulpa de remolacha disminuyó el pH y la materia seca, y disminuyó el contenido total de ácidos grasos volátiles del ciego, más rico en ácido acético, pero más pobre en propiónico, isobutirico, isovalérico y valérico. / [CA] L'objectiu principal d'aquesta tesi era avaluar el valor nutritiu de la polpa granulada de remolatxa (PR) produïda en diferents zones de sucre d'Espanya. La composició química de PR del nord i del sud d'Espanya, recol·lectada a l'hivern i l'estiu, respectivament. Es va comparar la composició química i el contingut d'àcids grassos i aminoàcids de 12 mostres de PR, agrupades segons el seu origen. La matèria seca (MS) del PR es caracteritza per un baix contingut de proteïna crua (8,7%), dins del 62% de la fibra detergent neutra (FND), però ric en FND (39,4%) i fibra soluble en detergent neutre (45,7%) i baix en greixos bruts (1,2%) i midó (0,9%). L'anàlisi principal de components (CP) dels principals principis immediats va indicar que els dos primers CP ja expliquen el 62% de la variabilitat en la composició del PR. Els valors autovector CP van indicar un pes important del contingut de midó i FND al primer CP i del contingut de cendra al segon CP. Posteriorment, la digestibilitat d'un aliment de control es va determinar amb una substitució del 20% de PR de cinc orígens diferents i el seu possible efecte sobre el valor nutritiu d'aquests PR per engreixar conills. Es van utilitzar conills de 90 42 dies amb 1,49 ± 0,208 kg de pes corporal, allotjats en gàbies metabòliques individuals i amb alimentació ad libitum. La digestibilitat dels components dels pinsos va ser determinada pel mètode in vivo convencional de recol·lecció fecal total. La digestibilitat de cadascuna de les púlps de remolatxa es va determinar mitjançant el mètode de substitució. No hi va haver cap efecte apreciat de substituir els components de l'alimentació de control per un 20% de PR de diferents fonts sobre el consum voluntari de conills. La inclusió de PR en dietes va augmentar la digestibilitat de la matèria seca (MS), la matèria orgànica, l'energia crua i totes les fraccions fibroses i va disminuir la digestibilitat de la proteïna crua. Pel que fa a l'efecte de l'origen, es van observar diferències significatives (P < 0,05) en la digestibilitat de totes les fraccions fibroses, i el PR recollit a l'estiu és el que té la màxima digestibilitat. L'origen del PR també va afectar el seu valor nutritiu, amb valors d'energia digeribles entre 11,8 i 14,5 MJ / kg MS i proteïna digestible entre 38 i 66 g / kg MS. Finalment, sobre el rendiment del creixement, es van utilitzar un total de 470 conills de 28 dies, amb 612,2 ± 103,4 gr. de pes viu. Els animals van ser assignats aleatòriament a cinc tractaments dietètics: control, sense polpa de remolatxa; PR20 i PR40, amb polpa de remolatxa 20 i 40% sense melassa, respectivament; i PRM20 i PRM40, amb polpa de remolatxa de melassa del 20 i del 40%, respectivament. El consum diari d'aliments (CDA) i el benefici mitjà diari (GMD) es van controlar als 28, 49 i 59 dies d'edat. També es van determinar les característiques del canal i del tracte digestiu als 59 dies d'edat. La mortalitat i la morbiditat es controlaven diàriament. La mortalitat durant el període de creixement va ser superior als grups PRM que a la PR (+ 9,2%; P < 0,05). Com més gran sigui la inclusió de PR, amb o sense melassa, el CDA inferior i el GMD dels animals, així com el rendiment al canal, la proporció de fetge i el percentatge de greixos dissecables als vostres canals. Tanmateix, la millor eficiència alimentària durant els darrers deu dies es va obtenir amb el grup PRM40. Pel que fa als paràmetres digestius, com més gran sigui la inclusió de la polpa de remolatxa, amb o sense melassa, més gran sigui el pes del tracte gastrointestinal buit i cec. De fet, una major inclusió de la polpa de remolatxa va disminuir el pH i la matèria seca i va disminuir el contingut total d'àcids grassos volàtils al cec, més ric en àcid acètic, però més pobre en propionic, isobutiric, isoval I valeric, independentment de la temporada, proporciona un alt contingut de fibra, soluble i insoluble, tot i que no és una font important de proteïnes, aminoàcids o àcids grassos. / [EN] The main objective of this thesis was to evaluate the nutritional value of granulated beet pulp (PR) produced in different sugar-growing areas of Spain. Initially the chemical composition of PR from the north and south of Spain, harvested in winter and summer, respectively, was characterized. The chemical composition and fatty acid and amino acid content of 12 PR samples, grouped according to their origin, were compared. The composition of the dry matter (DM) of the PR is characterized by a low crude protein content (8.7%), within 62% of neutral detergent fiber (NDF), but rich in NDF (39.4%) and fiber. soluble in neutral detergent (45.7%), and low in crude fat (1.2%) and starch (0.9%). The analysis of principal components (PC) of the main immediate principles indicated that the first two PCs already explain 62% of the variability in the composition of the PR. The values of the CP eigenvectors indicated an important weight of the starch and NDF content in the first CP and the ash content in the second CP. Subsequently, the digestibility of a control feed with 20% replacement of PR from five different origins was determined, and its possible effect on the nutritional value of these PR for fattening rabbits. Ninety 42-day-old rabbits with 1.49 ± 0.208 kg live weight were used, housed in individual metabolic cages and fed ad libitum. The digestibility of feed components was determined by the conventional in vivo method of total fecal collection. The digestibility of each of the beet pulps was determined using the substitution method. No effect was seen of replacing components of the control feed with 20% PR from different sources on the voluntary consumption of the rabbits. The inclusion of PR in the diets increased the digestibility of dry matter (DM), organic matter, gross energy and all fibrous fractions, and decreased the digestibility of crude protein. Regarding the effect of origin, significant differences (P<0.05) were observed in the digestibility of all fibrous fractions, with the PR collected in summer being the one with the highest digestibility. The origin of PR also affected its nutritional value, with digestible energy values between 11.8 and 14.5 MJ/kg DM and digestible protein between 38 and 66 g/kg DM. Finally, to evaluate the effect of the level of inclusion of beet pulp, with or without molasses, on growth performance, a total of 470 rabbits of 28 days of age were used, with 612.2 ± 103.4 gr. live weight. The animals were randomly assigned to five dietary treatments: Control, without beet pulp; PR20 and PR40, with 20 and 40% beet pulp without molasses, respectively; and PRM20 and PRM40, with 20 and 40% beet pulp with molasses, respectively. Daily feed intake (DAI) and average daily gain (ADG) were monitored at 28, 49 and 59 days of age. The characteristics of the carcass and digestive tract were also determined at 59 days of age. Mortality and morbidity were monitored daily. Mortality during the growth period was higher in the PRM than in the PR groups (+9.2%; P<0.05). The greater the inclusion of PR, with or without molasses, the lower the ADC and ADG of the animals, as well as the carcass yield, the proportion of liver and the percentage of dissectable fat in their carcasses. However, the best feed efficiency during the last 10 days was obtained with the PRM40 group. Regarding the digestive parameters, the greater the inclusion of beet pulp, with or without molasses, the greater the weight of the empty and blind gastrointestinal tract. In fact a greater inclusion of beet pulp decreased the pH and dry matter, and decreased the total content of volatile fatty acids of the caecum, richer in acetic acid, but poorer in propionic, isobutyric, isovaleric and valeric acids. Regarding the effect of the harvest season on the chemical characteristics of granulated sugar beet pulp, regardless of the season, it provides high fiber content, soluble and insoluble, although it is not an important source of protein / Arce Cabrera, ON. (2024). Utilización de pulpa granulada de remolacha en la alimentación de conejos en cebo [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/206201