Hospitalidad y muerte en la Ruta Jacobea navarra. Evidencias arqueológicas e históricas

Una ruta tan transitada a lo largo de más de un milenio como es la ruta jacobea, ha hecho que la hospitalidad y la muerte estén íntimamente ligadas a ella. Además de numerosa documentación referente a este hecho, existen una serie de evidencias arqueológicas en Navarra que conviene tener en cuenta, pues son diversos los enterramientos conocidos y excavados a lo largo del Camino y en otros lugares, que muestran la presencia de peregrinos o bien miembros de alguna de las cofradías santiaguistas de la región, aspecto que se manifiesta, entre otros elementos, por haber sido enterrados con una vieira, símbolo inequívoco de haber realizado la pe- regrinación al sepulcro del Apóstol. Las diversas evidencias arqueológicas se han localizado junto a la ermita de San Salvador de Ibañeta, en Roncesvalles, en Pamplona (entorno de la Catedral, plaza del Castillo, casa del Condestable), en Santa María de Eunate, en el Santo Sepulcro de Estella, en el Santo Sepulcro de Torres del Río, en San Saturnino de Artajona, en el despoblado de Arlás en Peralta y en Santa María de Ujué.Such a well-trodden route as the Camino de Santiago in the last millennium is inevitably bound up with hospitality and death. As well as voluminous documentation referring to these aspects, there is archeological evidence in Navarra that one has to bear in mind, because of the burials that are known and have been excavated along the Camino and in other places, which reveal the presence of pilgrims or members of one of the guilds associated with St James in the region, shown by the fact that a shell, a symbol of having done the pilgrimage to Santiago, is included among other elements of burial. Diverse archeological evidence has been located beside the church of St Salvador of Ibañeta in Roncesvalles, also in Pamplona (by the Cathedral, the Plaza del Castillo and the Casa del Condestable), in Santa Maria de Eunate and in the Holy Sepulchre of Estella, in the Holy Sepulchre of Torres del Río, in San Saturnino in Artajona, and in the abandoned areas of Arlás in Peralta and in Santa María de Ujué

Coronary artery high take-off in rodents and the possible involvement of Smad2

The coronary arteries (CAs) supply the mammalian heart with oxygenated blood. They arise from the right and left aortic valve sinuses at the aortic root. In humans, the occurrence of a CA arising ectopically from the tubular aorta, a condition called high take-off (HTO), is rare (A (Smad2C>A) allele is associated with HTO in this species. In order to test whether HTO occurs in association with Smad2C>A in other rodents, we examined the anatomical origin of the CAs, by means of stereomicroscopy and a corrosion cast technique, in 3,388 specimens belonging to 17 rodent species. In addition, Smad2 DNA sequence from M. musculus was compared by Blastn analyses with that from six of the species examined in which this sequence is known (Mus spretus, Rattus norvegicus, Apodemus sylvaticus, Myodes glareolus, Mesocricetus auratus, Microtus agrestis). HTO occurred in nine out of 17 species studied. The incidence of HTO ranged from 0.4% to 6.5% (low) in three species and from 15.9% to 25% (high) in six species. The Smad2 sequence showed similarities higher than 75% for the whole gene, and higher than 71% for the intron sequence that includes the rs29725537:C>A Single Nucleotide Polymorphism. The Smad2C>A allele was identified only in M. spretus and A. sylvaticus, with high incidences of HTO, whereas R. norvegicus, M. glareolus, M. auratus and M. agrestis showed low or null incidences. We conclude that HTO is a common trait in rodents, which does not lead to cardiac pathology probably due to the intramyocardial condition of their CAs, as opposed to the human subepicardial CAs. The Smad2C>A allele may be involved in the development of HTO and probably other phenotypes in different rodent species.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech. Funding: CGL2017-85090-P, FPU15/03209, UMAJI75 and FEDER

Los elasmobranquios como modelo para la trabeculación del miocardio ventricular

La formación de las trabéculas o trabeculación en vertebrados tiene lugar durante el desarrollo embrionario de los tres tipos de mioarquitecturas ventriculares, denominados compacto, esponjoso y mixto. El tipo compacto está formado mayoritariamente por fibras musculares compactadas y el esponjoso, por trabéculas. El tipo mixto, con una capa trabeculada interna y una compacta externa, ha sido considerado la condición primitiva en los Gnatostomados. En los modelos de vertebrados estudiados, la trabeculación se inicia siguiendo dos modelos alternativos: (1) en pollo y ratón, las células endocárdicas se evaginan hacia el miocardio biestratificado, (2) en pez cebra, los miocitos del miocardio monoestratificado se invaginan hacia el endocardio. La trabeculación en un miocardio mixto no ha sido descrita aún. Hemos estudiado el desarrollo del miocardio mixto en la pintarroja (S. canicula, Elasmobranquios) utilizando histología convencional, cortes semifinos, MEB y MET. En el estadio de desarrollo 27, el ventrículo se compone de dos capas miocárdicas internamente limitadas por un endocardio, ambas separadas por gelatina cardiaca. La trabeculación comienza en el estadio 28, cuando aparecen pequeños espacios entre los cardiomiocitos, la gelatina cardiaca disminuye de grosor y el endocardio contacta focalmente con el miocardio. En el estadio 29, los espacios entre miocardiocitos aumentan de tamaño y quedan recubiertos por endocardio, dando origen a lo que será la capa trabeculada. En los siguientes estadios, la capa trabeculada aumenta en complejidad y los miocardiocitos de la capa más externa proliferan y se compactan, dando lugar a la capa compacta del miocardio mixto. Concluimos que el proceso de trabeculación temprana en condrictios coincide con el descrito en tetrápodos y proponemos que este mecanismo ha sido conservado desde la aparición del miocardio de tipo mixto hasta tetrápodos y que el proceso descrito en pez cebra es una condición derivada dentro de vertebrados.CGL2017-85090-P; FPU15/03209; PRE2018-083176; FEDER Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Origen elevado de las arterias coronarias en roedores y posible implicación del gen Smad2

El corazón de los mamíferos está irrigado por las arterias coronarias derecha e izquierda originadas en los respectivos senos de la válvula aórtica. La presencia de arterias coronarias que emergen ectópicamente de la aorta tubular (“high take-off” -HTO-), es rara en humanos (A (Smad2C>A) está asociado con el HTO en esta especie. Se ha analizado un total de 3.388 especímenes de 17 especies de roedores pertenecientes a colecciones de la UMA y del Instituto de Investigación en Recursos Cinegéticos de Ciudad Real, con el fin de comprobar si el HTO es común en roedores y si se asocia con el alelo Smad2C>A. Se ha estudiado la posición en el origen de las arterias coronarias. La secuencia de ADN de Mus musculus para el gen Smad2 se comparó, mediante análisis con Blastn, con la de seis especies en las que esta secuencia se conoce (M. spretus, R. norvegicus, A. sylvaticus, M. glareolus, M. auratus, M. agrestis). El HTO aparece en nueve de las 17 especies estudiadas, con una incidencia que se ha clasificado como baja (0,4% - 6,5%) en tres especies y alta (15,9% - 25%) en seis especies. La secuencia de Smad2 mostró similitudes, entre M. musculus y las seis especies secuenciadas, superiores al 75% para el gen completo y al 71% para la secuencia intrónica, que incluye al polimorfismo de un solo nucleótido (SNP) rs29725537:C>A. El alelo Smad2C>A fue identificado solo en especies con incidencia alta de HTO (M. spretus y A. sylvaticus), mientras que las especies con incidencia baja (R. norvegicus y M. auratus) y nula (M. glareolus y M. agrestis) mostraron el alelo silvestre. Concluimos que el HTO es un carácter común en roedores. El alelo Smad2C>A podría estar implicado en el desarrollo de HTO y probablemente en otros fenotipos en diferentes especies de roedores.CGL2017-85090-P; FPU15/03209; PRE2018-083176; FEDER Universidad de Málaga, Campus de Excelencia Internacional Andalucía Tech

Transient and intensive pharmacological immunosuppression fails to improve AAV-based liver gene transfer in non-human primates

BACKGROUND: Adeno-associated vectors (rAAV) have been used to attain long-term liver gene expression. In humans, the cellular immune response poses a serious obstacle for transgene persistence while neutralizing humoral immunity curtails re-administration. Porphobilinogen deaminase (PBGD) haploinsufficiency (acute intermittent porphyria) benefits from liver gene transfer in mouse models and clinical trials are about to begin. In this study, we sought to study in non-human primates the feasibility of repeated gene-transfer with intravenous administration of rAAV5 vectors under the effects of an intensive immunosuppressive regimen and to analyze its ability to circumvent T-cell immunity and thereby prolong transgene expression. METHODS: Three female Macaca fascicularis were intravenously injected with 1x1013 genome copies/kg of rAAV5 encoding the human PBGD. Mycophenolate mofetil (MMF), anti-thymocyte immunoglobulin, methylprednisolone, tacrolimus and rituximab were given in combination during 12 weeks to block T- and B-cell mediated adaptive immune responses in two macaques. Immunodeficient and immunocompetent mice were intravenously injected with 5x1012 genome copies/kg of rAAV5-encoding luciferase protein. Forty days later MMF, tacrolimus and rituximab were daily administrated to ascertain whether the immunosuppressants or their metabolites could interfere with transgene expression. RESULTS: Macaques given a rAAV5 vector encoding human PBGD developed cellular and humoral immunity against viral capsids but not towards the transgene. Anti-AAV humoral responses were attenuated during 12 weeks but intensely rebounded following cessation of the immunosuppressants. Accordingly, subsequent gene transfer with a rAAV5 vector encoding green fluorescent protein was impossible. One macaque showed enhanced PBGD expression 25 weeks after rAAV5-pbgd administration but overexpression had not been detected while the animal was under immunosuppression. As a potential explanation, MMF decreases transgene expression in mouse livers that had been successfully transduced by a rAAV5 several weeks before MMF onset. Such a silencing effect was independent of AAV complementary strand synthesis and requires an adaptive immune system. CONCLUSIONS: These results indicate that our transient and intensive pharmacological immunosuppression fails to improve AAV5-based liver gene transfer in non-human primates. The reasons include an incomplete restraint of humoral immune responses to viral capsids that interfere with repeated gene transfer in addition to an intriguing MMF-dependent drug-mediated interference with liver transgene expression

Intensive pharmacological immunosuppression allows for repetitive liver gene transfer with recombinant adenovirus in nonhuman primates

Repeated administration of gene therapies is hampered by host immunity toward vectors and transgenes. Attempts to circumvent antivector immunity include pharmacological immunosuppression or alternating different vectors and vector serotypes with the same transgene. Our studies show that B-cell depletion with anti-CD20 monoclonal antibody and concomitant T-cell inhibition with clinically available drugs permits repeated liver gene transfer to a limited number of nonhuman primates with recombinant adenovirus. Adenoviral vector–mediated transfer of the herpes simplex virus type 1 thymidine kinase (HSV1-tk) reporter gene was visualized in vivo with a semiquantitative transgene-specific positron emission tomography (PET) technique, liver immunohistochemistry, and immunoblot for the reporter transgene in needle biopsies. Neutralizing antibody and T cell–mediated responses toward the viral capsids were sequentially monitored and found to be repressed by the drug combinations tested. Repeated liver transfer of the HSV1-tk reporter gene with the same recombinant adenoviral vector was achieved in macaques undergoing a clinically feasible immunosuppressive treatment that ablated humoral and cellular immune responses. This strategy allows measurable gene retransfer to the liver as late as 15 months following the first adenoviral exposure in a macaque, which has undergone a total of four treatments with the same adenoviral vector

Renal Failure Affects the Enzymatic Activities of the Three First Steps in Hepatic Heme Biosynthesis in the Acute Intermittent Porphyria Mouse

Chronic kidney disease is a long-term complication in acute intermittent porphyria (AIP). The pathophysiological significance of hepatic overproduction of the porphyrin precursors aminolevulinate acid (ALA) and porphobilinogen (PBG) in chronic kidney disease is unclear. We have investigated the effect of repetitive acute attacks on renal function and the effect of total or five-sixth nephrectomy causing renal insufficiency on hepatic heme synthesis in the porphobilinogen deaminase (PBGD)-deficient (AIP) mouse. Phenobarbital challenge in the AIP-mice increased urinary porphyrin precursor excretion. Successive attacks throughout 14 weeks led to minor renal lesions with no impact on renal function. In the liver of wild type and AIP mice, 5/6 nephrectomy enhanced transcription of the first and rate-limiting ALA synthase. As a consequence, urinary PBG excretion increased in AIP mice. The PBG/ALA ratio increased from 1 in sham operated AIP animals to over 5 (males) and over 13 (females) in the 5/6 nephrectomized mice. Total nephrectomy caused a rapid decrease in PBGD activity without changes in enzyme protein level in the AIP mice but not in the wild type animals. In conclusion, high concentration of porphyrin precursors had little impact on renal function. However, progressive renal insufficiency aggravates porphyria attacks and increases the PBG/ALA ratio, which should be considered a warning sign for potentially life-threatening impairment in AIP patients with signs of renal failure

Hospitalidad y muerte en la Ruta Jacobea navarra. Evidencias arqueológicas e históricas

Una ruta tan transitada a lo largo de más de un milenio como es la ruta jacobea, ha hecho que la hospitalidad y la muerte estén íntimamente ligadas a ella. Además de numerosa documentación referente a este hecho, existen una serie de evidencias arqueológicas en Navarra que conviene tener en cuenta, pues son diversos los enterramientos conocidos y excavados a lo largo del Camino y en otros lugares, que muestran la presencia de peregrinos o bien miembros de alguna de las cofradías santiaguistas de la región, aspecto que se manifiesta, entre otros elementos, por haber sido enterrados con una vieira, símbolo inequívoco de haber realizado la pe- regrinación al sepulcro del Apóstol. Las diversas evidencias arqueológicas se han localizado junto a la ermita de San Salvador de Ibañeta, en Roncesvalles, en Pamplona (entorno de la Catedral, plaza del Castillo, casa del Condestable), en Santa María de Eunate, en el Santo Sepulcro de Estella, en el Santo Sepulcro de Torres del Río, en San Saturnino de Artajona, en el despoblado de Arlás en Peralta y en Santa María de Ujué.Such a well-trodden route as the Camino de Santiago in the last millennium is inevitably bound up with hospitality and death. As well as voluminous documentation referring to these aspects, there is archeological evidence in Navarra that one has to bear in mind, because of the burials that are known and have been excavated along the Camino and in other places, which reveal the presence of pilgrims or members of one of the guilds associated with St James in the region, shown by the fact that a shell, a symbol of having done the pilgrimage to Santiago, is included among other elements of burial. Diverse archeological evidence has been located beside the church of St Salvador of Ibañeta in Roncesvalles, also in Pamplona (by the Cathedral, the Plaza del Castillo and the Casa del Condestable), in Santa Maria de Eunate and in the Holy Sepulchre of Estella, in the Holy Sepulchre of Torres del Río, in San Saturnino in Artajona, and in the abandoned areas of Arlás in Peralta and in Santa María de Ujué

Innate functions of immunoglobulin M lessen liver gene transfer with helper-dependent adenovirus

The immune system poses obstacles to viral vectors, even in the first administration to preimmunized hosts. We have observed that the livers of B cell-deficient mice were more effectively transduced by a helper-dependent adenovirus serotype-5 (HDA) vector than those of WT mice. This effect was T-cell independent as shown in athymic mice. Passive transfer of the serum from adenovirus-naïve WT to Rag1KO mice resulted in a reduction in gene transfer that was traced to IgM purified from serum of adenovirus-naïve mice. To ascribe the gene transfer inhibition activity to either adenoviral antigen-specific or antigen-unspecific functions of IgM, we used a monoclonal IgM antibody of unrelated specificity. Both the polyclonal and the irrelevant monoclonal IgM inhibited gene transfer by the HDA vector to either cultured hepatocellular carcinoma cells or to the liver of mice in vivo. Adsorption of polyclonal or monoclonal IgMs to viral capsids was revealed by ELISAs on adenovirus-coated plates. These observations indicate the existence of an inborn IgM mechanism deployed against a prevalent virus to reduce early post-infection viremia. In conclusion, innate IgM binding to adenovirus serotype-5 capsids restrains gene-transfer and offers a mechanism to be targeted for optimization of vector dosage in gene therapy with HDA vectors

Renal failure affects the enzymatic activities of the three first steps in hepatic heme biosynthesis in the acute intermittent porphyria mouse

Chronic kidney disease is a long-term complication in acute intermittent porphyria (AIP). The pathophysiological significance of hepatic overproduction of the porphyrin precursors aminolevulinate acid (ALA) and porphobilinogen (PBG) in chronic kidney disease is unclear. We have investigated the effect of repetitive acute attacks on renal function and the effect of total or five-sixth nephrectomy causing renal insufficiency on hepatic heme synthesis in the porphobilinogen deaminase (PBGD)-deficient (AIP) mouse. Phenobarbital challenge in the AIP-mice increased urinary porphyrin precursor excretion. Successive attacks throughout 14 weeks led to minor renal lesions with no impact on renal function. In the liver of wild type and AIP mice, 5/6 nephrectomy enhanced transcription of the first and rate-limiting ALA synthase. As a consequence, urinary PBG excretion increased in AIP mice. The PBG/ALA ratio increased from 1 in sham operated AIP animals to over 5 (males) and over 13 (females) in the 5/6 nephrectomized mice. Total nephrectomy caused a rapid decrease in PBGD activity without changes in enzyme protein level in the AIP mice but not in the wild type animals. In conclusion, high concentration of porphyrin precursors had little impact on renal function. However, progressive renal insufficiency aggravates porphyria attacks and increases the PBG/ALA ratio, which should be considered a warning sign for potentially life-threatening impairment in AIP patients with signs of renal failure