For more than 150 years, since the synthesis of urea by Friedrich Wöhler in 1828, molecular chemistry has developed a vast array of highly sophisticated and powerful methods for the construction of more complex molecular structures. Beyond the molecular chemistry based on the covalent bond, there lies the field of supramolecular chemistry, aims to gain control over the intermolecular bond. Supramolecular species are characterized both by the spatial arrangement of their components and by the nature of the intermolecular bonds that hold these components together. They possess well-defined structural, conformational, thermodynamic and kinetic properties. Research has been focused on utilization of such confined spaces to manipulate reaction dynamics, properties of the encapsulated guest molecules. This research presented in this thesis is a consolidated account of photophysical and photochemical reactions carried in water-soluble macrocycles, cavitands and dynamic host systems such as dendrimers and micelles. With the aid of NMR (1D and 2D) spectroscopic techniques, the host-guest complex characterization is executed.

Supramolecular systems comprised of non-covalent interactions are reversible in nature. This intrinsic reversibility of these systems is essential in achieving several functions, making it crucial to understand the dynamics of supramolecular systems. However, studies on the dynamics of supramolecular systems have always lagged behind structural and thermodynamic characterization of innumerable supramolecular systems developed. The first objective of this work was to understand the dynamics leading to a shift in the acidity constant (pKa) for 2-aminoanthracenium cation (AH+) upon binding with cucurbit[7]uril (CB[7]) host molecule. The adiabatic deprotonation of free AH+ in water was found to be inhibited in the complex with CB[7]. Different spectral characteristics for the protonated and deprotonated form of the guest molecule were used to understand the mechanism of this pKa shift associated with the binding to CB[7]. The results suggested that the pKa shift upon binding with CB[7] is a result of the slowing down of the deprotonation step in the complex, whereas the association rate constant did not change very much. The second objective of this work was to understand the role of cations on the binding dynamics of the N-phenyl-2-naphthyl amine (Ph-A-Np) binding to CB[7]. Ph-A-Np has two binding sites, which can lead to 1:1 and 2:1 host-guest complexes. The results indicate a switch in the binding mechanism for Ph-A-Np at low and high concentration regimes of sodium ions. Sodium ion was found to reduce the binding affinity of the naphthyl group to CB[7] whereas the complex formed by the phenyl group with CB[7] bound to one sodium ion was found to be stabilized. The final objective of this work was to study how structural changes to a guest molecule can affect the binding dynamics for the formation of a 2:1 “capsule” like complex with octa acid (OA). The dissociation for the OA capsule with pyrene (Py) as the encapsulated guest was shown to happen in 2.7 s previously. Two pyrene derivatives, 1-methylpyrene (MePy) and 1-pyrenemethanol (PyMeOH) were chosen as guest molecules to study the effect of these substituents on pyrene on the capsule dissociation dynamics. The results show that the residence time for the guests in the OA capsule depends on the substituents. For PyMeOH and MePy a shorter and longer residence time respectively in the capsule was observed when compared to Py. / Graduate / 2019-09-30

Supramolecular systems, which are formed by the noncovalent intermolecular interactions between molecules, are highly dynamic. The high reversibility of supramolecular systems leads to some functional features that cannot be achieved by the single chemical component. The kinetic information for the supramolecular systems can not be inferred from thermodynamic studies or structural studies. Furthermore, the information provided by the dynamic study can be employed to infer or explain the results from the thermodynamic study and the structural study. The first objective of this work was to study the dynamics and the binding mechanism of cucurbit[7]uril with a charged guest molecule (2-naphthyl-1-ethylammonium cation, NpAmH+). In general, the binding affinity of cucurbit[7]uril to the positively charged guests are very high compared with other host systems such as cyclodextrins and bile salt aggregates. In this work, the complexation of cucurbit[7]uril and NpAmH+ was studied from a kinetic point of view. Results showed that the high binding affinity of cucurbit[7]uril to NpAmH+ was due to the high association rate constant and the low dissociation rate constant for the complexation of cucurbit[7]uril and NpAmH+. Moreover, the competition between co-cations and NpAmH+ for the binding sites of cucurbituril molecules retarded the complexation process for cucurbit[7]uril binding to NpAmH+ and decreased the overall equilibrium constant for the formation of cucurbit[7]uril-NpAmH+ complex. The second objective of this work was to study the chiral recognition observed for the formation of 2:2 complexes between β-cyclodextrin and 2-naphthyl-1-ethanol (NpOH). The binding of β-cyclodextrin and NpOH leads to the formation of two 1:1 complexes and three 2:2 complexes. The binding dynamics of NpOH with β-cyclodextrin in the 1:1 complex is fast and occurs within microseconds. A much slower dynamics was observed for the formation of the 2:2 complex. Results showed that more 2:2 complex were formed for (R)-NpOH than for (S)-NpOH, which is due to the difference of the dissociation rate constant of the 2:2 complex for both NpOH enantiomers. The dissociation rate constant of the 2:2 complex for (R)-NpOH is 46.8% lower than that for (S)-NpOH while the association rate constant of the 2:2 complex are similar for both NpOH enantiomers. The third objective of this work was to study the dynamics and the binding mechanism of octa acid with pyrene. As known from the work of other researchers, the accessibility of small molecules (e.g. I- or O2) to pyrene bound to octa acid is largely limited by the octa acid capsule. In this study, a two-step successive process was observed for the complexation of octa acid and pyrene. The first step, which was related to the formation of octa acid-pyrene 1:1 complex, was sufficiently fast to be viewed as a pre-equilibrium process. The second step, which was related to the formation of octa acid-pyrene 2:1 complex, was slow on the millisecond – second time scale. The high binding affinity of octa acid to pyrene was observed, which is due to the low dissociation rate constant for the octa acid-pyrene 2:1 complex. / Graduate

Orientador: Devaney Ribeiro do Carmo / Banca: Newton Luiz Dias Filho / Banca: André Luís dos Santos / Resumo: O presente trabalho descreve a preparação, caracterização e as aplicações eletroanalíticas de silsesquioxanos e dendrímeros modificados suportados na superfície da sílica gel. A primeira etapa consistiu em ancorar o silsesquioxano ([H2N(CH2)3]8Si8O12) e o dendrímero Hexadecaamino Poli(propileno)imina (DAB-Am- 16) na superfície da 3-cloropropil sílica gel. Os materiais obtidos, SAC e SD, respectivamente, foram caracterizados pelas técnicas de Infravermelho (FTIR), RMN 29Si e 13C no estado sólido e Energia Dispersiva de Raios-X (EDX). A segunda etapa foi interagir o SAC e o SD com íons de Cu2+ e Ni2+ e posterior complexação com substâncias eletroativas, tais como: ferricianeto de potássio e nitroprussiato de sódio formando, dessa forma, os complexos binucleares: CuHCFSAC, NiHCFSAC, CuNPSAC, CuHCFSD, NiHCFSD e CuNPSD que foram caracterizados por Infravermelho (FTIR) e voltametria cíclica (VC). Os espectros na região do Infravermelho dos materiais supracitados exibiram freqüências próximas ou iguais a 2090 cm-1, atribuídas ao estiramento n(C N) característico do hexacianoferrato e nitroprussiato formados na superfície da SAC e SD. Utilizou-se estes materiais na construção do eletrodo de pasta de grafite os quais foram testados na oxidação eletrocatalítica de substâncias biologicamente importantes tais como a dipirona sódica e nitrito de sódio após um rigoroso estudo sobre o comportamento voltamétrico. O eletrodo de pasta de grafite contendo o CuHCFSAC exibiu dois pares redox com potenciais médios (Em) de: (Em)1= 0,29 V e (Em)2= 0,72 V vs Ag/AgCl (KCl =1.0 molL-1; v = 20 mV s-1) ambos atribuídos aos processos redox Cu(I)/Cu(II) e [Fe(II)(CN)6]4- / [Fe(III)(CN)6]3-, respectivamente, valores estes bem próximos para o CuHCFSD. Os eletrodos de NiHCFSAC e NiHCFSD exibiram... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: This work describes the preparation, characterization and electroanalytical properties of modified silsesquioxanes and dendrimers supported onto the silica gel surface. The first step was to anchor the silsesquioxane [(H2N(CH2)3] 8Si8O12) and dendrimer Hexadecaamino Poly(propylene)imine (DAB-Am-16) onto the 3-chloropropyl silica gel surface. The materials obtained, SAC and SD, respectively, were characterized by infrared (FTIR), 29Si NMR and 13C solid state and Energy Dispersive X-ray (EDX). The second step was interact the SAC and SD with Cu2+ and Ni2+ ions and subsequent complexation with electroactive substances, such as: ferricyanide of potassium and sodium nitroprusside forming binuclear complexes: CuHCFSAC, NiHCFSAC, CuNPSAC, CuHCFSD, NiHCFSD and CuNPSD which were characterized by Infrared (FTIR) and cyclic voltammetry (CV). The spectra in the infrared region of the above materials exhibited frequencies near or equal to 2090 cm-1 assigned to stretching n(C N) characteristic of nitroprusside and hexacyanoferrate formed onto the SD and SAC surface. We used these materials in the construction of carbon paste electrode which were tested in the electrocatalytic oxidation of biologically important substances such as sodium dipyrone and sodium nitrite after a rigorous study about voltammetric behavior. The graphite paste electrode containing CuHCFSAC exhibited two redox couples with midpoint potentials (Em) of: (Em)1 = 0.29 V and (Em)2 = 0.72 V vs. Ag/AgCl (KCl = 1.0 mol L-1, v = 20 mV s-1) both assigned to the redox processes Cu(I)/Cu(II) and [Fe(II)(CN)6]4- / [Fe(III)(CN)6]-3, respectively, values very near to the CuHCFSD. The electrodes of NiHCFSAC and NiHCFSD each exhibited two redox couples with midpoint potentials (Em): (Em)1 = 0.29 V and (Em)2 = 0.51 V vs Ag/AgCl (KCl = 1.0 mol L-1, v = 20 mV s- 1) both... (Summary complete electronic access click below) / Mestre

Made available in DSpace on 2014-06-11T19:25:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-10-01Bitstream added on 2014-06-13T20:14:08Z : No. of bitstreams: 1 gabrieljunior_s_me_ilha.pdf: 1458989 bytes, checksum: 61fbe8b93b56e59134558e478f7ab91e (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / O presente trabalho descreve a preparação, caracterização e as aplicações eletroanalíticas de silsesquioxanos e dendrímeros modificados suportados na superfície da sílica gel. A primeira etapa consistiu em ancorar o silsesquioxano ([H2N(CH2)3]8Si8O12) e o dendrímero Hexadecaamino Poli(propileno)imina (DAB-Am- 16) na superfície da 3-cloropropil sílica gel. Os materiais obtidos, SAC e SD, respectivamente, foram caracterizados pelas técnicas de Infravermelho (FTIR), RMN 29Si e 13C no estado sólido e Energia Dispersiva de Raios-X (EDX). A segunda etapa foi interagir o SAC e o SD com íons de Cu2+ e Ni2+ e posterior complexação com substâncias eletroativas, tais como: ferricianeto de potássio e nitroprussiato de sódio formando, dessa forma, os complexos binucleares: CuHCFSAC, NiHCFSAC, CuNPSAC, CuHCFSD, NiHCFSD e CuNPSD que foram caracterizados por Infravermelho (FTIR) e voltametria cíclica (VC). Os espectros na região do Infravermelho dos materiais supracitados exibiram freqüências próximas ou iguais a 2090 cm-1, atribuídas ao estiramento n(C N) característico do hexacianoferrato e nitroprussiato formados na superfície da SAC e SD. Utilizou-se estes materiais na construção do eletrodo de pasta de grafite os quais foram testados na oxidação eletrocatalítica de substâncias biologicamente importantes tais como a dipirona sódica e nitrito de sódio após um rigoroso estudo sobre o comportamento voltamétrico. O eletrodo de pasta de grafite contendo o CuHCFSAC exibiu dois pares redox com potenciais médios (Em) de: (Em)1= 0,29 V e (Em)2= 0,72 V vs Ag/AgCl (KCl =1.0 molL-1; v = 20 mV s-1) ambos atribuídos aos processos redox Cu(I)/Cu(II) e [Fe(II)(CN)6]4- / [Fe(III)(CN)6]3-, respectivamente, valores estes bem próximos para o CuHCFSD. Os eletrodos de NiHCFSAC e NiHCFSD exibiram... / This work describes the preparation, characterization and electroanalytical properties of modified silsesquioxanes and dendrimers supported onto the silica gel surface. The first step was to anchor the silsesquioxane [(H2N(CH2)3] 8Si8O12) and dendrimer Hexadecaamino Poly(propylene)imine (DAB-Am-16) onto the 3-chloropropyl silica gel surface. The materials obtained, SAC and SD, respectively, were characterized by infrared (FTIR), 29Si NMR and 13C solid state and Energy Dispersive X-ray (EDX). The second step was interact the SAC and SD with Cu2+ and Ni2+ ions and subsequent complexation with electroactive substances, such as: ferricyanide of potassium and sodium nitroprusside forming binuclear complexes: CuHCFSAC, NiHCFSAC, CuNPSAC, CuHCFSD, NiHCFSD and CuNPSD which were characterized by Infrared (FTIR) and cyclic voltammetry (CV). The spectra in the infrared region of the above materials exhibited frequencies near or equal to 2090 cm-1 assigned to stretching n(C N) characteristic of nitroprusside and hexacyanoferrate formed onto the SD and SAC surface. We used these materials in the construction of carbon paste electrode which were tested in the electrocatalytic oxidation of biologically important substances such as sodium dipyrone and sodium nitrite after a rigorous study about voltammetric behavior. The graphite paste electrode containing CuHCFSAC exhibited two redox couples with midpoint potentials (Em) of: (Em)1 = 0.29 V and (Em)2 = 0.72 V vs. Ag/AgCl (KCl = 1.0 mol L-1, v = 20 mV s-1) both assigned to the redox processes Cu(I)/Cu(II) and [Fe(II)(CN)6]4- / [Fe(III)(CN)6]-3, respectively, values very near to the CuHCFSD. The electrodes of NiHCFSAC and NiHCFSD each exhibited two redox couples with midpoint potentials (Em): (Em)1 = 0.29 V and (Em)2 = 0.51 V vs Ag/AgCl (KCl = 1.0 mol L-1, v = 20 mV s- 1) both... (Summary complete electronic access click below)

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A bicyclo-[2.2.2]-octa-2,5-diene (BOD) is a molecular photoswitch which makes up a so-called molecular solar thermal energy storage system. A molecular photoswitch has the ability to capture, store and release solar energy as heat. These processes can be enhanced by functional groups introduction. In this thesis, the effect of functional group introductions on the BODs' solar energy storage capacity and the thermal activation energy required for energy release was investigated. Various functional groups were introduced onto the BOD, and the corresponding storage and thermal back conversion energies were computed. Correlations were identified, and further tested using a practical BOD and its related structures. Conclusively, it was revealed that a storage energy might relate to (i) an electron withdrawing functional group strength, and (ii) an electron rich atom placed next to an electron withdrawing group. A dimethyl introduced bridge was observed to reduce storage energy and seemed to have a positive effect on the thermal back conversion barrier energy. The practical BOD λmax calculation was also computed, however, the result was not so promising as expected. Very little could be said about the thermal back conversion barrier energy due to long computational time and unreliable data. / En bicyklisk-[2.2.2]-okta-2,5-dien (BOD) är en molekylär fotoväxlare som ingår i ett så-kallat molekylär sol-termisk energilagrings-system. En molekylär fotoväxlare kan fånga, lagra och släppa tillbaka solenergin i form av värme. Dessa processer kan förstärkas av funktionella gruppers introduktion. I den här kandidatuppsatsen utforskades effekten av introduktion av funktionella grupper på BODs solenergilagrings-kapacitet och den termiska aktiveringsenergin som behövs för energiutlösning.  Olika funktionella grupper introducerades på BODn, och korresponderande lagringsenergier och termiska tillbakaomvandlingsenergier beräknades. Korrelationer identifierades och testades extensivt mot en praktisk BODs och dess relaterade strukturer. Sammanfattningsvis identifierades lagringsenergins möjliga relationer till (i) en elektron-accepterande funktionell grupps stryka, och (ii) en elektron-rik atom placerade nära en elektrondragande grupp. En dimetyl introducerad bro verkade reducera lagringsenergin och positivt påverka den termiska tillbakaomvandlingsenergin. Beräkning av den praktiska BODns λmax gjordes också, men resultatet var inte det förväntade. Mycket begränsade slutsatser kunde dras om den termiska tillbakaomvandlingsenergin på grund av lång beräkningstid och opålitliga data.

Space structures are inevitable while covering large spans. Space structures are skeletal structures, which are lighter for the same stiffness when compared with RCC roofs. Till now, space structures, like any other metal structures have been designed assuming the joints as rigid, although there have been several publications about semi rigid joints. Of course, the publications mostly deal with 2D structures and there are very few reports on 3D structures. Space structures, by their nature fall into the latter category. The joints in a space structure are popularly called as "nodes". Generally, nodes, which ensure concentricity of member axes, are either solid or hollow. These are either cast or forged. There are other proprietary types, which do not come under the above classification, and have not been considered in this thesis. Hollow nodes are obviously more economical than solid nodes, but also more flexible. While it is prudent to prefer hollow nodes, it is equally necessary to assess their flexibility, because of its influence on the behaviour of the structure. The hollow spherical node is very popular because of its simplicity and adaptability to various forms of space frames. Double layer grids, which are the most popular forms for roofing applications, are being increasingly implemented. While the hollow spherical node is well suited for double layer grids, an evolutionary development has been what is called as the hollow octahedral node (this node is simply referred to as the 'Octa ' node in this thesis). Chapter 1 introduces space frames and double layer grids in particular, with the advantages of using double-layer grids. Jointing systems available around the world are briefed and the node connector used in the present study is introduced with a brief write-up on its advantages and disadvantages. This chapter also explores the available literature and, the scope and objectives of the thesis are mentioned. Chapter 2 introduces 3D finite element models of the hollow spherical and octahedral nodes. The stiffness matrixes of these nodes have been derived by conducting analyses on the computer for six sizes each of the Octa and spherical nodes. Using the stiffness matrix of the node, a new method of incorporating this into the regular analysis of a space truss has been developed. The new method proposed yields realistic values for the forces in the members and takes into account the elastic deflections in the node under the action of member forces. Implementation of the proposed method has been carried out by writing a custom program using state-of-the-art object oriented programming techniques. A sample problem has been analyzed using this program to demonstrate the effect of including joint flexibility. The effect of flexibility of nodes on the effective length of compression members in double-layer grids has been evaluated. The effect of compliance on the dynamic characteristics of a space frame has also been evaluated for the sample space frame with flexible joints. The analysis program has been modified to evaluate the natural frequencies of the system using rigid or flexible nodes. The study of the Octanode and spherical node under the action of uniaxial compression and tension dominates the contents of Chapter 3. The two types of nodes have been analyzed using commercially available finite element software considering material nonlinearity. The stress patterns from the analyses have been examined thoroughly. Two consistent methods for fixing the load at yield in both uniaxial compression and tension have been proposed using the load-displacement curve. Yield loads for all the nodes have been evaluated using both the methods and the results agree well between the two methods. Three material yield values have been selected for each of the node size for evaluating the yield values viz. 240,320 and 415 MPa. The members of a double layer grid are connected to the nodes by bolts and holes are drilled in the nodes for this purpose. The bolthole patterns differ between two popular types of double-layer grids. Both these bolthole patterns have been modeled separately in the above exercise and the results for these two have been shown to be approximately the same. The effect of varying diameters of the boltholes on the response of the nodes has been examined. Relationships between the yield load, diameter, thickness and material yield have been developed using the method of least squares. The differences in the behaviour of the nodes under uniaxial compression and tension have been discussed. Ramberg Osgood type of relationships have been worked out for all the load-displacement curves obtained from the analyses. The simulation of non-linear behaviour of nodes with cracks with plastic crack closing forces have been carried out with useful insights into the behaviour of the two types of nodes in uniaxial compression and tension. Chapter 4 is devoted largely for studying the two types of nodes under the influence of biaxial load combinations. The combinations studied are dual compression, dual tension and compression-tension. In all cases equal loads are applied along two orthogonal; directions in the horizontal plane. Stress patterns have been examined for each type of load combination and yield values for each case have been obtained using one of the methods proposed in chapter 3. These have been compared with the corresponding uniaxial values in both compression and tension. Some useful inferences have been possible by studying the behaviour of the nodes under the various biaxial load combinations. In each case, relationships between the biaxial yield load, uniaxial yield load, diameter of node, thickness of node and material yield of node have been obtained using the method of least squares. The nodes have been analyzed under some selected Multi-axial loading and combinations of load which cause yield based on the second method proposed in Chapter 3 have been obtained and tabulated. However, a proper and thorough study of the nodes under multi-axial loading proved to be beyond the scope of this thesis. Chapter 5 contains the contributions made towards developing new methods and algorithms for obtaining the several results of chapters 2, 3 and 4, using object oriented programming (OOP) techniques. The contributions have been in Object Pascal, the underlying language of Delphi, a popular RAD tool developed by Borland/Inprise of USA. Several new modules have been developed to reliably handle the large amounts of data generated by the hundreds of analyses detailed in chapters 2,3 and 4. The ease with which new methods were possible to be incorporated into existing software using OOP has been demonstrated, with source code examples. Comparisons with other types of tools available and die advantages of using OOP have also been demonstrated using the experience during the preparation of this thesis. A strong case for OOP as an indispensable tool for the researcher has been made. Chapter 6: Several important conclusions and suggestions for future work have been made. Appendix 1 contains a brief note on the Method of Least Squares. Appendix 2 contains a small write-up on Delphi and OOP. Concepts of OOP have been briefly described and comparisons between three popular OOP languages have been attempted. A brief description of the features in Delphi's Object Pascal has also been provided. Appendix 3 contains the listing of Unit Arrays, which is a general purpose unit developed to make handling of large arrays easy. Several matrix calculations have been implemented which make the unit extremely useful for programmers. Appendix 4 contains the full listing of program FormK, which has been developed for chapter 2 to derive the fall stiffness matrix of a space frame node. The program picks up results from several analyses, forms a few columns of the stiffness matrix and then fills up the rest using the cyclic symmetry present in the space frame node. This program is given in full, with the intention that other researchers may find it useful to use it as-is or use after making small alterations to suit their circumstances. OOP is known for fast, reliable and easy ways of implementing modifications to existing code. Appendix 5 provides the full listing of the Object Pascal program for extracting Eigenvalues of a space truss with rigid joints or flexible joints. The incorporation of flexibility of the joints proposed in chapter 2 has been implemented. Descriptions of the program's implementations have been provided in chapter 5. Bibliography contains the alphabetical list of references.

Los fotorreceptores de los mamíferos, las células externas de la retina que detectan la luz, carecen de capacidad de autorregeneración tras una lesión. En los estadios avanzados de las IRD, los fotorreceptores se pierden pero la estructura interna de la retina se conserva durante largos periodos de tiempo, aunque con una importante remodelación sináptica y gliosis. Para estas condiciones, las terapias regenerativas dirigidas a reemplazar los fotorreceptores y establecer sinapsis funcionales con las neuronas internas de la retina viables restantes podrían permitir la recuperación de la visión en pacientes que de otro modo serían ciegos. En otras palabras, la terapia con células madre está dirigida al tratamiento de las degeneraciones de la retina en aquellas situaciones en las que se han perdido los fotorreceptores y, por lo tanto, no es posible restaurar la funcionalidad a través de enfoques más comunes como la terapia de reemplazo de genes. Se han desarrollado y caracterizado células madre embrionarias humanas (hESC) o células madre pluripotentes inducidas (iPSC) derivadas de células precursoras de fotorreceptores (PRPC) contenidas en organoides de retina (RO) para terapias experimentales regenerativas. Aunque la terapia con células madre es un campo que ha mostrado resultados prometedores en animales de laboratorio, no hay informes que evalúen la seguridad y eficacia de su uso en perros. Con un inyector subretiniano que fue modificado para acomodar el gran tamaño de los agregados celulares, inyectamos con éxito las células en el espacio subretiniano (SRS) canino utilizando un procedimiento quirúrgico sencillo (inyección manual en bolo sin vitrectomía previa). Pudimos monitorizar la supervivencia y las características de las células injertadas a lo largo del tiempo utilizando un enfoque de imagen multimodal que incluía la detección mediante fotografía del fondo de ojo y/o cSLO de los genes reporteros fluorescentes (tdTomato o GFP) expresados por las PRPC. Esto nos permitió superar un reto importante encontrado en otros estudios, donde las células donantes no fluorescentes fueron seguidas sólo por OCT o detectadas por histología después de la terminación. La visualización de las PRPC fluorescentes en el animal vivo nos permitió diferenciarlas de las células del huésped. En consonancia con lo descrito anteriormente, observamos dos patrones temporales de pérdida de células del donante: una reducción temprana del número de células injertadas en la primera semana del trasplante que no dependía del estado de la inmunosupresión (IS), y un rechazo retardado del injerto, observado en aquellos perros que no estaban inmunosuprimidos. De hecho, hasta donde sabemos, no hay estudios que evalúen el tiempo de pérdida de fotorreceptores tras el trasplante, con y sin IS, controlando simultáneamente la transferencia de material citoplasmático entre las células del donante y del huésped. Aquí observamos signos compatibles con el rechazo del trasplante en animales que no recibieron IS sistémico, así como en un único perro cuyo tratamiento con IS se interrumpió. El grado de inflamación clínica variaba entre los animales, pero la vasculitis retiniana, el vítreo turbio y la inflamación de la retina eran comunes en todos los perros con rechazo de las células del donante. Estos signos se detectaron por primera vez entre 1-2 y 12 semanas después del trasplante, lo que respalda la necesidad de un seguimiento frecuente de las retinas tratadas en los meses siguientes al trasplante para detectar posibles signos tempranos de rechazo y ofrecer la oportunidad de ajustar el régimen inmunosupresor. Dado que el rechazo del trasplante en el SRS también puede producirse sin inflamación clínica manifiesta, se justifica la identificación de nuevos biomarcadores que puedan detectar la inflamación subclínica temprana para modular la respuesta inmunitaria y prolongar la supervivencia del injerto. Aunque se desconocen todos los factores que promueven la supervivencia de las células del donante, descubrimos que el IS sistémico desempeñaba un papel fundamental en la supervivencia de las hESC-PRPCs administradas por vía subretiniana, como se había informado anteriormente. En el presente estudio, algunas PRPC desarrollaron estructuras similares a pedículos, expresaron la proteína presináptica sinaptofisina y establecieron contactos con las células bipolares del anfitrión. Estos resultados alentadores preparan ahora el terreno para la evaluación funcional de estas xenosinapsis. La retinosis pigmentaria es un grupo de enfermedades genéticas que provocan una pérdida progresiva de la visión, siendo una de las principales causas de ceguera en los países desarrollados. Está bien documentado que en pacientes con retinosis pigmentaria, existe una disfunción vascular asociada que conduce al adelgazamiento de los vasos; sin embargo, las implicaciones de esta disfunción vascular en la degeneración de los fotorreceptores no se comprenden completamente. Los modelos caninos de degeneraciones retinianas hereditarias han sido de gran relevancia en el desarrollo traslacional de terapias de reemplazo génico para múltiples formas de retinosis pigmentaria, Amaurosis congénita de Leber, y enfermedad de Best. De manera similar a lo que ocurre en pacientes con retinosis pigmentaria, los perros afectados con las mismas mutaciones también experimentan remodelación vascular, sin embargo, la cinética de esta remodelación vascular en enfermedades retinianas hereditarias caninas no se ha estudiado y no se han establecido los parámetros normales en el perro. La angiografía por tomografía de coherencia óptica (OCTA) es un método novedoso de obtención de imágenes sin necesidad del uso de contraste intravenoso que permite la visualización detallada de la circulación retiniana, lo que permite el estudio de los distintos plexos vasculares por separado. Esta importante extracción de imágenes de los diferentes plexos, junto con la alta resolución de estos angiogramas, permite una cuantificación más precisa de la densidad vascular y otros parámetros, teniendo muchas aplicaciones en sujetos sanos y en pacientes con diferentes enfermedades oculares y sistémicas. Con el uso de modernas técnicas de imagen, este trabajo ha confirmado la presencia de cuatro plexos retinianos distintos. Aunque muchos estudios han informado de los datos cuantitativos de las imágenes OCTA en retinas humanas, no se han descrito parámetros vasculares caninos. Este estudio proporciona datos normativos para el SVP+ICP, DCP y WR, estableciendo con éxito un rango de referencia que puede ser consultado y comparado en futuros estudios. En los ojos humanos, el número de plexos retinianos y sus densidades disminuyen hacia la periferia, y esto es similar a lo que Engerman et al. describieron previamente en perros. Nuestro trabajo no sólo confirma este hallazgo, sino que ahora proporciona datos cuantitativos para cuatro parámetros que se utilizan frecuentemente para caracterizar las redes vasculares. En nuestra evaluación, los angiogramas de OCTA tenían una mayor resolución en comparación con las imágenes de AF en la misma localización. Al igual que en el caso de los seres humanos, la angiografía por OCT en perros permitió identificar lechos capilares (ICP y DCP) que no se identificaban con la AF. Sin embargo, la AF proporcionó un mayor campo de visión y los artefactos que se encontraron en algunas de las exploraciones de OCTA (artefactos de movimiento y anormalidades de descorrelación debido al artefacto de proyección) no se observaron en las imágenes de AF. Cuando se compara con las imágenes obtenidas por IHC en montajes completos de retina, nuestro estudio confirma que la OCTA proporciona una buena visualización de la SVP y la DCP. También encontramos que había una subrepresentación de los vasos de pequeño calibre en la OCTA, especialmente los situados en capas altamente reflectantes (ICP). Cuando se compara con las imágenes adquiridas en las mismas localizaciones por microscopía confocal/IHC, nuestros resultados sugieren que la OCTA es una técnica valiosa para visualizar y cuantificar la vasculatura retiniana en perros, especialmente para el análisis de la VD en el DCP. Además, por IHC encontramos que el ICP se fusiona con el SVP pero no con el DCP como ocurre en las retinas humanas. Nuestro estudio ha confirmado la viabilidad del uso de OCTA en perros, proporcionando imágenes resueltas en profundidad de diferentes capas retinianas segmentadas que permiten la evaluación de plexos individuales. Esto allana el camino para otros análisis in vivo de la vasculatura de la retina canina en un amplio número de patologías de la retina con un fenotipo vascular. Además, evaluamos los cambios vasculares en el area centralis de perros afectados por varias formas de IRD que fueron visualizados por OCTA en diferentes etapas de la enfermedad. Identificamos que la DCP está más afectada que las redes vasculares más superficiales en una etapa temprana de la enfermedad. Además, confirmamos que existe una fuerte asociación entre el VD en el DCP y el grosor de la ONL, lo que sugiere que la evaluación de la vascularización en este plexo puede utilizarse como un marcador indirecto para la evaluación de los requisitos metabólicos de la retina externa. Por último, hemos validado mediante el análisis de los vasos en los montajes planos de la retina los hallazgos de la OCTA, y hemos descubierto que en los modelos caninos de IRD la migración de las células del RPE también desempeña un papel en las alteraciones vasculares de la fase posterior que se producen en los pacientes con RP. Encontramos que los cambios microscópicos observados en los vasos con degeneración eran diferentes en las distintas redes retinianas. En la DCP, se confirmó un estrechamiento y una pérdida progresiva de vasos, que acabó con la desaparición completa de esta red. En el SVP de los tres modelos, los vasos presentaban un mayor grosor de la pared debido a la deposición de material que rodea la pared vascular que, en las fases finales, conduce al estrechamiento y la oclusión vascular. En la fase final de la enfermedad, se observó que múltiples vasos de la SVP estaban rodeados de estructuras pigmentadas. El marcaje específico de RPE65 reveló que se trataba de células del PRE que habían migrado para rodear estos vasos internos de la retina. Aquí caracterizamos dos tipos diferentes de degeneración vascular que se producen en los plexos retinianos SVP y DCP, lo que podría aportar información para futuros estudios que evalúen específicamente la fisiopatología de esta degeneración vascular. Un animal del modelo crd2/NPHP5 fue tratado con terapia de reemplazo génica unilateralmente con una inyección subretiniana que cubría el area centralis. En este perro, la pérdida de ONL en el momento de la intervención de terapia génica era inferior al 50%. Al comparar las fotografías del fondo de ojo del ojo no tratado y el tratado, se identificó fácilmente una marcada preservación de la vascularización en el área que fue cubierta por la terapia génica. Las imágenes OCTA se procesaron con el programa AngioTool, y la evaluación cualitativa de las imágenes esqueletizadas mostró una regresión vascular en el ojo no tratado y una notable preservación de la integridad vascular en el ojo tratado. También demostramos que en estas enfermedades naturales, así como en un modelo de degeneración aguda de fotorreceptores inducido por la luz, el DCP se ve afectado antes que los otros plexos vasculares de la retina. La posterior disminución de la VD en el SVP+ICP que se produce en las últimas fases de la degeneración, es probablemente una respuesta al marcado adelgazamiento de la retina externa y a la capacidad de que el oxígeno transportado por los vasos coroideos llegue a las localizaciones internas de la retina, como se ha confirmado previamente en modelos animales felinos utilizando perfiles espaciales de oxigenación de la retina.