Epithelial surface changes and spermatozoa storage in the reproductive tract of the bitch.

Spermatozoa are known to bind to the epithelial cells lining the uterine tube in various species, but information in canids is conflicting and sparse. The first aim of this study was to measure the epithelial surface outline (ESO) of different regions of the canine uterine tube in the four stages of the oestrous cycle as an indicator of a changing potential reservoir for spermatozoa. The second aim was to identify the site of sperm storage in the bitch after natural mating. Reproductive tracts were collected from bitches undergoing routine ovariohysterectomy. Histological analysis showed that, when corrected for uterine tube size, the ESO of pro-oestrus (P<0.005) and oestrus (P<0.05) tubes were larger than anoestrus, but not metoestrus, tubes. The second study examined reproductive tracts from 12 Beagle bitches at 6, 12, 24 and 48h after mating. Light and electron microscopy revealed large numbers of spermatozoa in the proximal regions of the uterus and particularly the distal utero-tubal junction (UTJ), with few present in the proximal UTJ or uterine tubes. Spermatozoa were bound by their heads to microvilli on the epithelial surface of the uterine lumen and to ciliated cells in the distal UTJ. This is the first report to measure and document differences in potential epithelial attachment sites of the uterine tubes at different stages of the oestrous cycle and to provide compelling evidence that the main spermatozoal storage site in the reproductive tract of the bitch is the distal UTJ.

The effect of cryopreservation on the capacitation status and epithelial cell attachment capability of dog spermatozoa.

Freezing and cooling of spermatozoa during cryopreservation for artificial insemination causes ultrastructural changes in the acrosome and plasma membrane which reduces longevity and fertility. Cryopreservation-induced capacitation-like changes and reduced ability of spermatozoa to bind to the cells of the reproductive tract of the bitch may contribute to the reduced fertility of cryopreserved spermatozoa. Previous studies in the dog have investigated the effects of extending and cooling spermatozoa on the plasma membrane but often only after freeze-thawing and not in conjunction with an assessment of their ability to bind to uterine tube epithelial explants. This study investigated the effect of each stage of the cryopreservation process on capacitation and attachment to the reproductive tract of the bitch. The capacitation status of spermatozoa was studied over time after cryopreservation using a chlortetracycline and Hoechst 33258 stain. The ability of spermatozoa to bind to uterine tube epithelial explants was studied using Hoechst 33342 stain. Extending, cooling and freeze-thawing promoted capacitation and decreased the spermatozoal binding ability. The effect of each stage appeared to be cumulative with the freeze-thawing stage being the most dramatic. The results suggested that the cumulative effect of each stage of the cryopreservation process results in the promotion of capacitation before spermatozoa have reached the site of fertilisation, and therefore spermatozoa have reduced ability to attach to epithelial cells. These effects are contributory factors to the reduced fertility of cryopreserved spermatozoa.

Biological actions of curcumin on articular chondrocytes.

OBJECTIVES: Curcumin (diferuloylmethane) is the principal biochemical component of the spice turmeric and has been shown to possess potent anti-catabolic, anti-inflammatory and antioxidant, properties. This article aims to provide a summary of the actions of curcumin on articular chondrocytes from the available literature with the use of a text-mining tool. We highlight both the potential benefits and drawbacks of using this chemopreventive agent for treating osteoarthritis (OA). We also explore the recent literature on the molecular mechanisms of curcumin mediated alterations in gene expression mediated via activator protein 1 (AP-1)/nuclear factor-kappa B (NF-kappaB) signalling in chondrocytes, osteoblasts and synovial fibroblasts. METHODS: A computer-aided search of the PubMed/Medline database aided by a text-mining tool to interrogate the ResNet Mammalian database 6.0. RESULTS: Recent work has shown that curcumin protects human chondrocytes from the catabolic actions of interleukin-1 beta (IL-1beta) including matrix metalloproteinase (MMP)-3 up-regulation, inhibition of collagen type II and down-regulation of beta1-integrin expression. Curcumin blocks IL-1beta-induced proteoglycan degradation, AP-1/NF-kappaB signalling, chondrocyte apoptosis and activation of caspase-3. CONCLUSIONS: The available data from published in vitro and in vivo studies suggest that curcumin may be a beneficial complementary treatment for OA in humans and companion animals. Nevertheless, before initiating extensive clinical trials, more basic research is required to improve its solubility, absorption and bioavailability and gain additional information about its safety and efficacy in different species. Once these obstacles have been overcome, curcumin and structurally related biochemicals may become safer and more suitable nutraceutical alternatives to the non-steroidal anti-inflammatory drugs that are currently used for the treatment of OA.

Targeting matrix metalloproteinases in inflammatory conditions.

The matrix metalloproteinases (MMPs) and their endogenous regulators, the tissue inhibitors of MMPs (TIMPs) are responsible for the physiological remodelling of the extracellular matrix (ECM) in healthy connective tissues. MMPs are also involved in the regulation of cell behaviour via the release of growth factors and cytokines from the substrates they cleave, increasing the magnitude of their effects. Excess MMP activity is associated with ECM destruction in various inflammatory conditions, such as osteoarthritis (OA), while MMP under-activity potentially impairs healing by promoting fibrosis and preventing the effective removal of scar tissue. Both direct (TIMPs, small molecule MMP inhibitor drugs, blocking antibodies and anti-sense technologies) and indirect (glucocorticoids and non-steroidal anti-inflammatory drugs, statins, anti-sense technologies and various phytochemicals) strategies for MMP inhibition have been proposed and investigated. The strategy of MMP inhibition for degenerative and neoplastic diseases has been relatively unsuccessful due to undesired sequelae, often caused by non-selectivity of the MMP inhibition method. Therapeutic strategies for MMP-related conditions ideally should regulate MMP activity in order to maintain the optimum balance between MMPs and TIMPs. By avoiding complete inhibition it may be possible to prevent the complications of MMP over- and under-activity. Furthermore, MMP sub-type specificity is critical for minimising detrimental off-target effects that have been observed with broad-spectrum MMP inhibitors. Any potential MMP inhibitor or modulator must be subjected to rigorous pharmacokinetic, toxicity and safety studies and data obtained using in vitro models must be verified in clinically relevant animal models before therapeutic use is considered.

Mesenchymal stem cells in connective tissue engineering and regenerative medicine: applications in cartilage repair and osteoarthritis therapy.

Defects of load-bearing connective tissues such as articular cartilage, often result from trauma, degenerative or age-related disease. Osteoarthritis (OA) presents a major clinical challenge to clinicians due to the limited inherent repair capacity of articular cartilage. Articular cartilage defects are increasingly common among the elderly population causing pain, reduced joint function and significant disability among affected patients. The poor capacity for self-repair of chondral defects has resulted in the development of a large variety of treatment approaches including Autologous Chondrocyte Transplantation (ACT), microfracture and mosaicplasty methods. In ACT, a cartilage biopsy is taken from the patient and articular chondrocytes are isolated. The cells are then expanded after several passages in vitro and used to fill the cartilage defect. Since its introduction, ACT has become a widely applied surgical method with good to excellent clinical outcomes. More recently, classical ACT has been combined with tissue engineering and implantable scaffolds for improved results. However, there are still major problems associated with the ACT technique which relate mainly to chondrocyte de-differentiation during the expansion phase in monolayer culture and the poor integration of the implants into the surrounding cartilage tissue. Novel approaches using mesenchymal stem cells (MSCs) as an alternative cell source to patient derived chondrocytes are currently on trial. MSCs have shown significant potential for chondrogenesis in animal models. This review article discusses the potential of MSCs in tissue engineering and regenerative medicine and highlights their potential for cartilage repair and cell-based therapies for osteoarthritis and a range of related osteoarticular disorders.

Biofilms and their relevance to veterinary medicine.

Bacteria are renowned for their ability to tolerate and adapt to a wide range of adverse environmental conditions. The primary mechanism that facilitates these adaptations is thought to be the capacity to form and maintain biofilms. Within a biofilm, bacteria become attached to a surface where they exist in complex communities which are able to interact with each other through intracellular communication and thus rapidly adapt to changing environments. The organisms within biofilms are notorious for their resistance towards the host immune response and antibacterial agents compared to their free-living planktonic counterparts. Consequently, biofilms are of significant importance to both clinical and veterinary science. However, although bacterial infections are widely reported in animals their association with biofilms is rarely discussed. The aim of this review is to look at the characteristics of biofilm infections in humans and to relate this knowledge to veterinary science in order to assess their relevance in this area.