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.     

The use of quantitative PCR for identification and quantification of Brachyspira pilosicoli, Lawsonia intracellularis and Escherichia coli fimbrial types F4 and F18 in pig feces

This field study explored the cytokine expression in intestinal tissue and serum from 19 diarrhoeic and 9 healthy pigs in herds with a long-time history of Lawsonia intracellularis-infection. The disease, proliferative enteropathy (PE), is associated with diarrhoea and poor performance in growers and haemorrhagic diarrhoea and sudden death in finisher pigs, but the immunopathology is poorly understood. Histopathology, demonstration of L. intracellularis and porcine circovirus type 2 (PCV2) in intestinal tissue by PCR, and detection of serum antibodies to L. intracellularis, were performed. The presence of IL-1β, IL-6, IL-10, IFN-α, IFN-γ, TNF-α and TGF-β in sera was determined by immunoassays, and intestinal mRNA expression of these cytokines plus IL-12p40 was determined by qPCR. Intestinal specimens from pigs with intestinal adenomatosis (n=2), proliferative haemorrhagic enteropathy or swine dysentery (n=2), and controls (n=2) were analysed by a genome wide porcine microarray. The clinical signs of PE were not always supported by the subsequent analyses, and the presence of PCV2 may have contributed to an increased mRNA expression for IFN-γ in intestinal specimens from some pigs. The limited gene expression in the microarray analyses and the limited expression of cytokines in both sera and intestines, indicate that the immune response is poorly activated in the initial course of an infection with L. intracellularis. However, the gene encoding for insulin-like growth factor binding protein 3 (IGFBP-3) was up-regulated in two pigs with prominent mucosal proliferation.     

Reno-, hepato- and splenomegaly of common carp fingerlings (Cyprinus carpio L.) diseased in swimbladder inflammation caused by Sphaerospora renicola Dyková et Lom, 1982

The weight of internal organs (swimbladder, kidney, liver, spleen) in relation to the body weight was studied in common carp fingerlings divided into three groups on the basis of swimbladder appearance and microscopic examination of the kidney. The fish had been collected from different Hungarian fish farms at the time when swimbladder inflammation (SBI) usually occurs (in July and August). The first group comprised fish with severe signs of SBI and massive renal sphaerosporosis, the second group consisted of fish with milder swimbladder changes and/or kidney infection by a low number of Sphaerospora renicola, while the third group was constituted by infection-free common carp fry. Statistical analysis of swimbladder, kidney, liver and spleen weight in relation to the body weight revealed that in the infected groups the internal organs were substantially enlarged. This suggests that in common carp fry with SBI the swimbladder changes are accompanied by reno-, hepato- and splenomegaly.     

The surgical technique and the age of the horse both influence the outcome of mosaicplasty in a cadaver equine stifle model

Six pieces of grafts, 6.5 mm in diameter, 20 mm in length, were taken from each of 170 cadaver hindlimbs, using the cranial surface of the medial femoral trochlea for harvesting. The age of the horses varied between 4 months and 23 years. 30 limbs under the age of 12 years were selected for transplantation. Three of six grafts were transplanted into the medial femoral condyle using different combinations of tunnel depth and dilation. With ageing, a significant decline in transplantability was detected. In general, mosaicplasty cannot be recommended in horses above 11 years. Based on a previous clinical case (Bodó et al., 2000), a good surface alignment was indeed achieved with a combination of graft length drilling and dilation in most cases. However, the occasional entrapment of cartilage debris under the graft prevented perfect alignment in the present cadaver study in 27% of the grafts transplanted in this manner. Since the protrusion of grafts never exceeded 1.5 mm, we conclude that drilling 3-5 mm deeper than graft length with graft length deep dilation can avoid disadvantageous protrusion of the transplanted hyaline cartilage caps, achieving bone decompression at the same time.     

Epidemiological, clinical, echo-doppler characteristics of mitral valve endocardiosis in Cavalier King Charles in France: a retrospective study of 451 cases (1995 to 2003)

A study performed on 451 Cavalier King Charles showed that 40.6% of dogs had a left apical systolic heart murmur, whose prevalence increased with age (> 11-year-old, 100%), but was not different between males and females. Mitral valve endocardiosis represented 93.3% of the ultrasonographic abnormalities.     

The Effect of Supplementation with Fresh Browse of Ziziphus mauritiana or Combretum aculeatum on Feed Intake,Nitrogen Utilization and Growth of Grazing Range Sheep

The influence of 0, 60 or 120 min access to a grove of either Ziziphus mauritiana (Ziziphus) or Combretum aculeatum (Combretum) on forage intake and on the digestibility, growth and excretion of nutrients was determined using 40 Oudah rams grazing for 7 h/day on poor-quality dry season pasture in the Sahel. The effects of browse species and browsing duration (0, 30, 60 or 120 min) on the ruminal ammonia content were also evaluated using 8 mature fistulated rams. Ziziphus increased both total digestible organic matter and total dry matter intakes per (kg live weight)^0.75 without decreasing herbage intake, whereas 60 or 120 min access to Combretum reduced herbage intake by 6.3% and 4.2%, respectively. The digestibility of the diet decreased (p<0.05) with the duration of access to the groves. Despite the provision of more nitrogen (N) than in the control diet, ruminal NH₃-N decreased 24 h after browsing commenced. The reduced live weight gain of sheep browsing Combretum may indicate more deleterious compounds in Combretum than in Ziziphus. Ziziphus appears to have more potential than Combretum to increase sheep production in low-input crop/livestock systems.     

The epidemiology and diagnosis of bluetongue with particular reference to Corsica

Bluetongue (BT) and/or BT viruses (BTV) have been identified in the Mediterranean basin and the Balkans each year from 1998 to 2002 and in particular BTV serotype 2 in the French Island of Corsica (2000 and 2001). In response to these virus incursions, the French Veterinary Authorities carried out epidemiological studies that included virological, serological and entomological analysis, and two vaccination campaigns performed in the winter of 2000/2001 and the winter and spring of 2001 and 2002. Rapid and reliable serotype differentiation is essential at the start of an outbreak to allow an early selection of vaccine to control the spread of the virus. Thus, molecular tools, that complement conventional methods, have been developed for early detection of infection, determination of the serotype, and differentiation between natural infection and vaccination. Serological results showed that the first vaccination campaign during the winter of 2000/2001 did not provide full protection for all sheep and during the summer of 2001, 335 sheep flocks in Corsica were again infected by BTV 2 (7-fold more that in 2000). Entomological studies have demonstrated that the only proven vector of the disease, Culicoides imicola, was present in the island in 2000 and that it has successfully established itself in Corsica. The safety and immunogenicity of the commercial South African vaccine were studied. Fourteen sheep were vaccinated and then observed for clinical signs. Blood, sera, spleen and lymph nodes were collected and analyzed, and the results confirmed the safety and potency of using this vaccine to protect sheep from clinical disease. As a result, an intensive vaccination campaign was performed during winter and spring 2001/2002. No cases of BT had been observed by the end of summer 2002, indicating that the vaccination campaign has been successful in protecting sheep from infection.