ABSTRACT
Germ cell transplantation and testis graft represent promising biotechnologies that can be applied for the reproduction of commercial or endangered species. However, mechanisms of rejection from the host immune system might remove the transplanted donor cells/tissues and limit the surrogate production of gametes. In this work, we administered emulsion containing-immunosuppressants to verify whether they are capable to prevent immune rejection and promote survival of testis allografts in rainbow trout. In the first part of this study, we demonstrated in vitro that tacrolimus and cyclosporine were able to affect viability, inhibit leucocyte proliferation, and suppress il2 expression in vitro. In in vivo experiments, both doses of tacrolimus (0.5 and 1.5 mg/kg) and the lower dose of cyclosporine (20 mg/kg) significantly inhibited the expression of il2 in head kidney, three days post-injection. A higher dose of cyclosporine (40 mg/kg) was able to inhibit il2 expression for up to seven days post-injection. In the second part, testis allografts were conducted in fish treated weekly with emulsion containing-tacrolimus. Immunohistochemical, conventional histology, and qRT-PCR (vasa) analysis demonstrated the presence of spermatogonial cells by the fifth week, in animals treated with 0.5 mg/kg of tacrolimus similar as found in autografted group. In the group treated with the highest tacrolimus dose (1.5 mg/kg) and in the non-treated group (without immunosuppressant), no germ cells or their respective markers were detected. il2 expression in head kidney was also suppressed in grafted animals treated with tacrolimus compared to non-treated group. These results suggest that tacrolimus may be a promising immunosuppressant for testis allografts or germ cell transplantation in rainbow trout. Co-administration combining tacrolimus (at lower dose) with other immunosuppressive drugs for inhibiting other activation pathways of the immune system, as performed in human organ transplantation, could be an alternative approach to optimize the immunosuppressive effects in host organisms.
Subject(s)
Allografts/immunology , Cyclosporine/pharmacology , Immunosuppressive Agents/pharmacology , Oncorhynchus mykiss/surgery , Spermatogonia/immunology , Tacrolimus/pharmacology , Testis/transplantation , Transplantation, Homologous/veterinary , Animals , MaleABSTRACT
PROBLEM: Several mechanisms contribute to the tolerogenic state observed during pregnancy, such as the activity of the enzyme indoleamine 2, 3-dioxygenase (IDO). This initializes the catabolism of tryptophan, inducing T cells to apoptosis due to its deprivation and by the action of its catabolites in the placental microenvironment. Progesterone plays an important part on immunological tolerance mechanisms during pregnancy; however, there is no evidence it is related to IDO activity. Thus, this study aimed to investigate progesterone influence on the maternal-fetal interface of pregnant Wistar rats, by identifying IDO positive cells by immunophenotyping and flow cytometry under exogenous progesterone supplementation. METHOD OF STUDY: Placenta and embryo cells were cultured and separated into groups that received interferon γ or progesterone, supplemented or not with mifepristone. After 2 and 24 h, these were labeled with an anti-IDO and a series of antibodies specific to leucocytes and progesterone receptor and processed through flow cytometry analysis. RESULTS: Progesterone induced a significant decrease in the expression of IDO in dendritic cells and CD4+ lymphocytes. CONCLUSION: The blocking of progesterone receptor on these cells by mifepristone restored IDO expression levels and may constitute evidence of the participation of this hormone through a direct route in these cells.
Subject(s)
CD4-Positive T-Lymphocytes/drug effects , Immune Tolerance/drug effects , Indoleamine-Pyrrole 2,3,-Dioxygenase/immunology , Placenta/drug effects , Progesterone/pharmacology , Uterus/drug effects , Abortifacient Agents, Steroidal/pharmacology , Animals , Apoptosis/drug effects , Apoptosis/genetics , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , Dendritic Cells/cytology , Dendritic Cells/drug effects , Dendritic Cells/immunology , Embryo, Mammalian , Female , Gene Expression Regulation , Immunophenotyping , Indoleamine-Pyrrole 2,3,-Dioxygenase/antagonists & inhibitors , Indoleamine-Pyrrole 2,3,-Dioxygenase/genetics , Interferon-gamma/pharmacology , Maternal-Fetal Exchange/immunology , Mifepristone/pharmacology , Placenta/cytology , Placenta/immunology , Pregnancy , Primary Cell Culture , Rats , Rats, Wistar , Receptors, Progesterone/antagonists & inhibitors , Receptors, Progesterone/genetics , Receptors, Progesterone/immunology , Tryptophan/immunology , Tryptophan/metabolism , Uterus/cytology , Uterus/immunologyABSTRACT
A indoleamina 2,3-dioxigenase (IDO) é uma enzima responsável por catabolizar o aminoácido triptofano. Sua presença no ambiente uterino placentário está relacionada à tolerância imunológica ao semi-aloenxerto, pois impede a proliferação de células imunológicas maternas, seja pela falta do aminoácido, ou pela ação de alguns catabólitos oriundos da quebra do triptofano, como o ácido quinolínico, que é tóxico principalmente para os linfócitos T. Pouco se conhece sob a influência de substâncias (hormônios e citocinas) presentes na interface materno fetal e a expressão dessa enzima. Por esta razão, formulou-se a hipótese de que hormônios e interleucinas presentes na região uteroplacentária poderiam exercer algum efeito na expressão da IDO. Células oriundas da interface materno fetal de ratas Wistar foram mantidas em cultivo, onde receberam suplementação com estradiol e interferon-γ. A expressão da enzima foi avaliada pela técnica de citometria de fluxo nos períodos de 4, 24 e 48 horas e confirmação da presença proteica por imuno-histoquímica. Os resultados mostraram um aumento na expressão de IDO após a adição de estrógeno (9,03±0,81/11,25±0,25) e interferon-γ (9,03±0,81/20,43±0,60). O efeito do interferon-γ já era esperado como relatado na literatura, contudo, a elevação da expressão da IDO pela adição do estrógeno constitui nova informação sobre possíveis mecanismos envolvidos na ativação da enzima. O melhor esclarecimento desses achados poderia contribuir para uma melhor compreensão da participação dessa enzima na tolerância materno-fetal e para uma futura modulação terapêutica da mesma...
The indoleamine 2,3-dioxygenase (IDO) is an enzyme responsible for catabolizing the tryptophan. Its presence in the placental uterine environment is related to immunological tolerance to the semi-allograft because it prevents proliferation of maternal immune cells, either by the lack of this amino acid or by the action of its catabolites, such as the quinolinic acid, which is particularly toxic for T lymphocytes. Little is known regarding the influence of hormones and cytokines on the expression of IDO in the maternal fetal interface. Therefore, the hypothesis that some hormones and interleukins present in uteroplacental region could have an effect on the expression of IDO on cultured cells was tested. Cells derived from the fetal maternal interface from Wistar rats were kept in culture and supplemented with estradiol and interferon-γ. Expression of the enzyme was assessed by flow cytometry at periods of 4, 24 and 48 hours and confirmation of the presence of protein by immunohistochemistry. The results showed an increasing of IDO expression after the addition of estrogen (9.03±0.81 to 11.25±0.25) and interferon-γ (9.03±0.81 to 20.43±0.60). The effect of interferon-γ was expected as reported in the literature, however, elevated IDO expression by estrogen represents new information on possible mechanisms involved in the enzyme activation. These findings could provide a better understanding of IDO contribution on maternal-fetal tolerance and may collaborate to future therapeutic modulation of this enzyme...
Subject(s)
Animals , Female , Pregnancy , Guinea Pigs , Estrogens , Interferon-gamma , Rats, Wistar/embryology , Flow Cytometry/veterinary , Clinical Enzyme Tests/veterinary , Immunohistochemistry/veterinary , PlacentaABSTRACT
O sistema respiratório das aves é bastante eficiente com pulmões pequenos e compactos ligados a sacos aéreos que diferem entre as espécies. Dessa forma, objetivou-se realizar a descrição anatômica dos sacos aéreos em patos em relação à sua topografia. Para tanto, foram utilizados o trato respiratório pós-cefálico de quatro patos adultos, machos e fêmeas, injetados, via sondagem traqueal, com látex corado e fixados em solução de formol a 10%. Observou-se a presença dos sacos aéreos cervical, clavicular, torácico cranial, torácico caudal e abdominal. O saco aéreo cervical distribuiu-se nos dois antímeros, na região dorsal, localizando-se dorso-lateralmente a musculatura do pescoço. O saco aéreo clavicular apresentou-se com uma distribuição irregular, formando uma porção mediana e duas laterais com inúmeras projeções entre os órgãos das regiões cervical e torácica cranial, apresentando divertículos para os ossos coracóide, úmero e esterno. Sua porção mediana formou, em alguns casos, uma projeção ao redor do coração em "forma de saia". Os sacos aéreos torácicos craniais encontraram-se caudalmente ao coração, medialmente as quatro primeiras costelas, sendo sobrepostos pelo fígado. Os sacos aéreos torácicos caudais localizaram-se em uma posição média aos sacos aéreos torácico cranial e abdominal, estendendo-se além da margem da última costela, englobando a porção cranial do saco aéreo abdominal. Os sacos aéreos abdominais encontraram-se caudo-medialmente aos sacos aéreos torácicos caudais e lateralmente ao trato digestório. Os sacos aéreos torácicos e abdominais apresentaram poucas projeções entre os órgãos, possuindo margens regulares e grande capacidade de volume.
The respiratory system of birds is very effective, comprised by small and compact lungs associated with air sacs that differ between species regarding their topography. Considering this latter aspect, this work aimed to perform an anatomical and topographical description of the air sacs in mallard (Anas platyrhynchos). Ten mallards, five males and five females, had theirpost-cephalic respiratory tract injected with colored latex and fixed by a 10% formaldehyde solution. The presence of cervical, clavicular, cranial thoracic, caudal thoracic and abdominal air sacs was observed. The cervical air sac was distributed in both antimeres situated dorso-laterally to the muscles of the neck. The clavicular air sac presented an irregular distribution, showing a middle and two lateral parts with numerous projections between the cervical and cranial thoracic organs and presenting several diverticula to the coracoids , sternum and humerus bones. Its middle portion formed, in some cases, a projection around the heart. The cranial thoracic air sacs were found caudal to the heart, medially to the first four ribs, being overplayed by the liver. The caudal thoracic air sacs were located in a middle position to the cranial thoracic and abdominal air sacs extending beyond the edge of the rib, over the cranial portion of the abdominal air sac. The abdominal air sacs were found caudal-medial to the caudal thoracic air sacs and lateral to the digestive tract. The thoracic and abdominal air sacs presented few projections between the organs, has and showed regular margins and a high volumetric capacity.
