Muscle reorganisation through local injection of stem cells in the diaphragm of mdx mice

Background

The diaphragm is the major respiratory muscle affected by Duchenne muscular dystrophy (DMD) and is responsible for causing 80% of deaths. The use of mechanical forces that act on the body or intermittent pressure on the airways improves the quality of life of patients but does not prevent the progression of respiratory failure. Thus, diseases that require tissue repair, such as DMD, represent a group of pathologies that have great potential for cell therapy. The application of stem cells directly into the diaphragm instead of systemic application can reduce cell migration to other affected areas and increase the chances of muscle reorganisation. The mdx mouse is a suitable animal model for this research because its diaphragmatic phenotype is similar to human DMD. Therefore, the aim of this study was to assess the potential cell implantation in the diaphragm muscle after the xenotransplantation of stem cells.

Methods

A total of 9 mice, including 3 control BALB/Cmice, 3 5-month-old mdx mice without stem cell injections and 3 mdx mice injected with stem cells, were used. The animals injected with stem cells underwent laparoscopy so that stem cells from GFP-labelled rabbit olfactory epithelium could be locally injected into the diaphragm muscle. After 8 days, all animals were euthanised, and the diaphragm muscle was dissected and subjected to histological and immunohistochemical analyses.

Results

Both the fresh diaphragm tissue and immunohistochemical analyses showed immunopositive GFP labelling of some of the cells and immunonegativity of myoblast bundles. In the histological analysis, we observed a reduction in the inflammatory infiltrate as well as the presence of a few peripheral nuclei and myoblast bundles.

Conclusion

We were able to implant stem cells into the diaphragm via local injection, which promoted moderate muscle reorganisation. The presence of myoblast bundles cannot be attributed to stem cell incorporation because there was no immunopositive labelling in this structure. It is believed that the formation of the bundles may have been stimulated by cellular signalling mechanisms that have not yet been elucidated.     

Determination of the Inflammatory Potential of Bioaerosols from a Duck‐Fattening Unit by Using a Limulus Amebocyte Lysate Assay and Human Whole Blood Cytokine Response

Inhalation of bioaerosols from animal houses can induce acute inflammatory reactions in the respiratory tract. Determination of the concentration of airborne endotoxins is widely used to characterize this risk. In this study, the activity of bioaerosol samples from a duck‐fattening unit to induce interleukin‐1β (IL‐1β) in human blood and to react with Limulus Amebocyte Lysate (LAL) was investigated. The activity detected in the whole blood assay correlated well with the endotoxic activity found in the LAL assay (Spearmen's ρ = 0.902). However in all samples, the inflammation‐inducing potential was overestimated by the LAL assay. It is assumed that this overestimation could be, in part, a result of an overestimation of the inflammatory potential of endotoxins originating from Pseudomonadaceae by the LAL assay. Pseudomonadaceae were regularly isolated from the air of the duck‐fattening unit. The results presented here indicate that the whole blood assay can be used besides the LAL assay as an additional method to characterize the inflammation‐inducing potential of bioaerosols.     

Perinatal hypoxia induces subsequent retinal degeneration in the offspring of ovoviviparous fish, Xiphophorous maculates

OBJECTIVE: This experiment evaluated the perinatal hypoxic effect on the retina of offspring of the ovoviviparous fish. ANIMAL STUDIED: The ovoviviparous fish Xiphophorous maculates was used for the experiment. PROCEDURE: The mothers were kept in a hypoxic environment of 3.5% oxygen for 6 h, starting 30 h before hatching. Subsequently, the retinae of the offspring were fixed, sectioned at 6 microm and evaluated microscopically from the age of 1 to 35 days. RESULTS: Degeneration of the outer nuclear layer of the retina was noted on the 3rd day and severe retinal degeneration was observed on the 35th day. Immunocytochemistry confirmed apoptosis by TUNEL reaction. There was no difference in neovascularization, as revealed by vascular endothelial growth factor, between controls (group 1) and hypoxic fish (group 2). CONCLUSIONS: Perinatal hypoxia could have long-lasting effects on the central nervous system in some species.     

An investigation of Leishmania spp. in Didelphis spp. from urban and peri-urban areas in Bauru (São Paulo, Brazil)

Blood and bone marrow samples were taken from 112 Didelphis spp., collected between March 2005 and February 2006, from urban and peri-urban areas of Bauru, São Paulo State, Brazil, to evaluate the hypothesis that these animals might constitute a reservoir of Leishmania spp. Anti-Leishmania ssp. antibodies were screened in the serum samples using an enzyme-linked immuno-sorbent assay (ELISA) and the polymerase chain reaction (PCR). PCR was performed on fragments of DNA samples from Leishmania spp. using primers 13A and 13B, and showed a positive outcome in 91.6% of the 112 samples tested. Of the 107 samples analyzed by ELISA, 71% were positive. Evidence of epidemiological risk factors such as a circulating parasite and freely moving vectors suggests that Didelphis spp. may participate in the transmission cycle of Leishmania spp. in Bauru.     

In Vitro Assessment of the Equine Hoof Wall Strains in Flat Weight Bearing and After Heel Elevation

Horseshoeing is a common practice, but effects on the hoof wall are poorly understood. Strain gauges were used to document and compare hoof behavior in vitro during flat weight bearing and after artificial heel elevation. Ten front limbs of Thoroughbred race horses, shod with conventional flat shoes, were used. Eight strain gauges were symmetrically distributed around the toe, quarters, and heels. Each limb was mounted to a testing machine (Kratos K5002; Kratos Dynamômetros, Ltda., Cotia-SP-Brazil) and subjected to a load equivalent to 30% of the donor's body weight. Strains (μ) were acquired by means of a computerized system and the results compared using Friedman and Wilcoxon statistical tests. There was greater strain variation when the heels were elevated. Compression predominated during flat weight bearing, with a tendency to horizontal traction after heel elevation. The changes in strain caused by heel elevation were not always symmetrical. Elevation of the heels tensed the toe and the medial quarter horizontally, increased load at the posterior portion of the hoof capsule, and hindered its expansion.