Subconjunctival use of allogeneic mesenchymal stem cells to treat chronic superficial keratitis in German shepherd dogs: Pilot study

Background: Chronic superficial keratitis (CSK) is an ocular condition in dogs characterized by corneal opacification leading to visual function impairment. Control of this chronic condition requires the use of topical immunomodulators or corticosteroids daily. Regenerative medicine has shown promising results in several fields of medicine. Aim: The aim of this study was to evaluate the clinical effect of allogeneic mesenchymal stem cells (MSCs) of adipose tissue applied via subconjunctival in dogs with CSK. Methods: A series of cases of eight dogs diagnosed with CSK were divided into two groups, four dogs each; the conventional treatment group received prednisolone 1% as topical eye drops and the experimental group (EG) received allogeneic MSCs transplantation. The dogs had not previously been treated for CSK. Systemic and ophthalmologic examinations were performed to exclude other abnormalities. An administered amount of MSC (1 × 106 cells each time) was injected via subconjunctival in the peri-limbal region at 0 and 30 days. The animals were followed for 110 days for clinical evaluation, and, at the same time, the images of the corneal abnormalities were obtained and analyzed in the ImageJ software. The statistical analysis was performed in the GrandPrism 7.0 software. Results: Initial and final images revealed that areas with neovascularization, inflammatory infiltrate, and opacity regressed in most eyes in both groups (7/8 eyes in each group) at the end of the 110 days, p = 0.0391 and p = 0.0078 respectively, but this response was minor in the EG comparing to conventional group (CG) (p = 0.026). No local or systemic side effects were observed. Conclusions: Despite the small melioration, MSCs treatment suggests clinical improvement in patients with CSK after 110 days without any local or systemic side effects. However, the improvement achieved was significantly less than the observed within CG. Further studies still are needed to evaluate the use and benefits of stem cells as an adjunct treatment for CSK

Comportamento do complexo DNA-proteina, do vigor e da motilidade de espermatozoides de touros submetidos a diferentes tempos de congelação

Orientador: Maria Luiza S. MelloDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: A incidência de patologias do complexo DNA-proteína, medida pelo método de metacromasia induzida, bem como os parâmetros vigor e motilidade, foram estudados em espermatozóides de touro provenientes de 42 partidas seminais congeladas a partir de 1973 a 1995. Os resultados indicam que esses parâmetros diferem estatisticamente para grupos de sêmen criopreservados por tempos diferentes. As anomalias no complexo DNA-proteína foram mais elevadas para as partidas de 1973 e 1974, diferindo de todas as demais, que por sua vez não diferiram entre si. Admite-se que o tipo de envasamento e o diluidor do sêmen tenham alguma participação nesses resultados, pois apenas as partidas de 1973 e 1974 foram envasadas em ampolas. Quanto aos parâmetros vigor e motilidade, houve diminuição nos seus valores, em função do aumento no tempo de criopreservação, porém não diretamente correlacionada ao tipo de envasamento. Especialmente para o parâmetro vigor foi observada uma distribuição não simétrica de valores inclusive para tempos de criopreservação mais recentes, como 1991 a 1995. Considerando-se que as técnicas de congelação e descongelação utilizadas neste período são muito semelhantes entre si, acredita-se que a existência de partidas seminais que apresentem vigor e motilidade muito abaixo da média do grupo possa ocorrer devido a condições de manutenção da congelação fora do padrão ideal, como por exemplo, utilizando-se níveis insatisfatórios de nitrogênio líquido no botijão de armazenamento de sêmen ou por manipulação excessiva ou inadequada das partidas seminais durante o período de criopreservação, causando exposição das mesmas à temperatura ambiente por tempo excessivo. Os dados levantados sugerem cautela na utilização de sêmen criopreservado por tempos muito longos, especialmente se associado a isto tiver sido utilizada a técnica de preservação e envasamento mais antiga. Cuidados adicionais de preservação deverão ser mantidosAbstract: The incidence of the DNA-protein complex pathologies, measured by induced metachromasy method, as welI as vigor and motility pattems, were studied in bulI sperm from 42 ftozen batches from 1973 to 1995. Results indicate that these parameters differ statistically for cryopreserved semen groups in accord to different time. Complex DNA-protein anomalies were higher for 1973 and 1974 batches, differing ftom alI the remaining, which in tum did not differ among themselves. It is acknowledged that storage type and semen dilutor have some participation in these results because on1y 1973 and 1974 batches were stored in ampules. Decrease was shown in the amounts of vigor and motility parameters, as a function of increase in cryopreservation time, however, not direct1y correlated to storage type. A non-symmetric distribution of amounts was observed especially for vigor parameter, inc1uding more recent cryopreservation time as 1991 to 1995. Considering that the fteezing and thawing techniques used by both groups during this period are very similar it is assumed that the existence of seminal batches showing vigor and motility much below group's average could be due to not maintaining ideal freezing conditions, for instance , using unsatisfying levels of liquid nytrogen in the semen storage tank or due to excessive or inadequate manipulation of semen batches during cryopreservation period, causing their exposure to environmental temperature for excessive time. ColIected data suggest caution when using cryopreserved semen for very long periods of time especialIy if the older preservation and storage technique has been utilized associated to it. Additional preservation measures need to be keptMestradoBiologia CelularMestre em Ciências Biológica