Identifying the degree of major histocompatibility complex matching in genetically unrelated dogs with the use of microsatellite markers.

BACKGROUND: The dog has served as an important experimental model for biomedical research such as transplantation and developing immunosuppressive agents. Although major histocompatibility complex (MHC) in dogs is a dominant factor of graft rejection, it has not been well investigated in dogs compared with human. For that reason, imprecise cross-matching or time-consuming sequence-based typing methods have generally been used to choose specific donor and recipient pairs. Investigation of matching distribution of MHC in dogs with the use of simple and accurate methods would be beneficial for biomedical researchers. The aim of this study was to identify the diversity of dog leukocyte antigen (DLA) types in genetically unrelated dogs by means of microsatellite markers. METHODS: Thirty-three Beagle and Shih-Tzu dogs, which were negative in cross-matching, were chosen. The genomic DNA was isolated from peripheral blood leukocytes, and highly polymorphic short tandem repeats located in MHC class I and II were amplified with the use of specific primers. RESULTS: Among all of the dogs, MHC matching groups, including class I full match-class II full match (M-M), class I full match-class II haplo match (M-H), class I haplo match-class II full match (H-M), class I haplo match-class II haplo match (H-H) groups, were ∼1.55%, 0.39%, 1.94%, and 6.59%, respectively. MHC class I nonmatch-class II nonmatch (U-U) groups were 58.14% of the total dogs. CONCLUSIONS: Because differences of histocompatibility between donor and recipient leads to various allograft rejections, knowledge of the distribution of MHC matching in unrelated dogs would be helpful in designing studies and to get more accurate results from experiments using dog transplantation models.

Hepatic differentiation of porcine embryonic stem cells for translational research of hepatocyte transplantation.

Porcine embryonic stem cells (ES) are considered attractive preclinical research tools for human liver diseases. Although several studies previously reported generation of porcine ES, none of these studies has described hepatic differentiation from porcine ES. The aim of this study was to generate hepatocytes from porcine ES and analyze their characteristics. We optimized conditions for definitive endoderm induction and developed a 4-step hepatic differentiation protocol. A brief serum-free condition with activin A efficiently induced definitive endoderm differentiation from porcine ES. The porcine ES-derived hepatocyte-like cells highly expressed hepatic markers including albumin and α-fetoprotein, and displayed liver characteristics such as glycogen storage, lipid production, and low-density lipoprotein uptake. For the first time, we describe a highly efficient protocol for hepatic differentiation from porcine ES. Our findings provide valuable information for translational liver research using porcine models, including hepatic regeneration and transplant studies, drug screening, and toxicology.

Fabrication of a biodegradable xenoantigen-free rat liver scaffold for potential drug screening applications.

BACKGROUND: The increasing market in biological pharmaceuticals raises the demand for human test systems. Although 2-dimensional (2D) models are mostly used for these purposes, these models not mimic responses of 3-dimensional (3D) native tissue. METHODS: After generation of a rat liver scaffold using 0.1% sodium dodecyl sulfate, we characterized the histology, blood vessel integrity, and residual DNA as well as retained amounts of collagen and glycosaminoglycan (GAG). Then, we examined the susceptibility of extracellular matrix (ECM) to enzymatic remodeling. Finally, a mixed lymphocyte reaction (MLR) was performed to evaluate the in vitro immunogenicity of the ECM against human peripheral blood mononuclear cells (PBMCs). RESULTS: Histologic examination of decellularized liver revealed the removal of nuclear and cytoplasmic materials with preservation of architecture. The vascular network was intact after decellularization. Biochemical analysis of ECM components revealed that only a negligible amount of DNA was retained compared with the native liver with preservation of large amounts of GAG and collagen. Scaffolds were degraded in response to collagenase treatment. MLR demonstrated that decellularized matrices did not exert any xenostimulatory response against human PBMCs. CONCLUSION: Our findings suggested that naturally derived rat liver scaffolds show natural biocompatibility besides the ability to preserve the intact 3D structure and components. Because of these characteristics, the whole decellularized rat liver can retain many aspects of native tissue structure and function upon recellularization enabling it to be used for drug screening.