The extracellular matrix (ECM) acts as a critical factor during morphogenesis. Because the organization of the ECM directly influences the structure of tissues and organs, a determination of the way that ECM organization is regulated should help to clarify morphogenesis. We have analyzed the assembly of Del1, an ECM protein produced by endothelial cells in embryos, in the ECM. Del1 consists of three epidermal growth factor repeats (E1-E3) at its N-terminus and two discoidin domains (C1, C2) at its C-terminus. Experiments with various deletion mutants of Del1 have revealed that fragments containing the C-terminus of C1, which has a lectin-like structure, direct deposition in the ECM. The efficiency of deposition varies according to the presence of other domains in Del1. A fragment containing E3 and C1 has the strongest deposition activity, whereas fragments containing C2, which is highly homologous to C1, have low deposition activity. Digestion of ECM with hyaluronidase from bovine testis releases Del1 from the ECM, suggesting that glycosaminoglycans are involved in the deposition of Del1. In vivo gene transfer experiments have shown that fusion with the deposition domain of Del1 dramatically alters the distribution of exogenous proteins in mice. Thus, the extent of Del1 deposition may modify the organization of the ECM.

To examine the role of integrin αv subunit in the progression of squamous cell carcinoma (SCC), oral SCC cells were stably transfected with integrin αv cDNA. Integrin αv transfectants exhibited the enhancement of proliferation on type Ⅰ collagen, and seemed to have a high ability to invade type Ⅰ collagen gel. Overexpression of integrin αv led to rapid phosphorylation of focal adhesion kinase (FAK), mitogen‑activated protein kinase kinase 1/2 (MEK1/2) and extracellular signal‑regulated kinase 1/2 (ERK1/2) in SCC cells on type Ⅰ collagen. The downregulation of integrin β8 in integrin αv transfectants by its specific antisense oligonucleotide led to a decrease in type Ⅰ collagen‑stimulated activation of FAK and the MEK/ERK signaling pathway, and also suppressed the proliferation on type Ⅰ collagen and the invasiveness into type Ⅰ collagen gel. Moreover, the expression of integrin β8 was induced following transfection with integrin αv cDNA. These results indicated that the overexpression of integrin αv induces integrin αvβ8 heterodimer formation and the binding of integrin αvβ8 to type Ⅰ collagen might enhance the proliferation and invasion of SCC cells via the activation of the MEK/ERK signaling pathway.

Although the development of effective viral vectors put gene therapy on the road to commercialization, nonviral vectors show promise for practical use because of their relative safety and lower cost. A significant barrier to the use of nonviral vectors, however, is that they have not yet proven effective. This apparent lack of interest can be attributed to the problem of the low gene transfer efficiency associated with nonviral vectors. The efficiency of gene transfer via nonviral vectors has been reported to be 1/10th to 1/1000th that of viral vectors. Despite the fact that new gene transfer methods and nonviral vectors have been developed, no significant improvements in gene transfer efficiency have been achieved. Nevertheless, some notable progress has been made. In this review, we discuss studies that report good results using nonviral vectors in vivo in animal models, with a particular focus on studies aimed at in vivo gene therapy to treat cancer, as this disease has attracted the interest of researchers developing nonviral vectors. We describe the conditions in which nonviral vectors work more efficiently for gene therapy and discuss how the goals might differ for nonviral versus viral vector development and use.

In this paper, we found that Del1, an extracellular matrix protein secreted by embryonic endothelial cells, increases the efficiency of transfection in vitro. Conditioned medium containing Del1 increased transfection by the LacZ gene using several non-viral gene transfer systems, including lipoplex and polyplex methods. Experiments using deletion mutants and fragments of Del1 revealed that the third epidermal growth factor-like repeat (E3) of Del1 mediates the enhancement of gene transfer and, furthermore, that the motif CXDXXXFXCXC is essential. Incubation of Pro5 cells, a yolk sac-derived cell line, with as low as 16 pM recombinant E3 was sufficient to enhance transfection, and 1 nM recombinant E3 enhanced the transfection 12-fold. Inhibitors of endocytosis suppressed this activity of the recombinant E3. These results suggest that the E3 fragment of Del1 can be used as a general biological enhancer of non-viral gene transfer.

The condyle is the most common site of mandibular fracture. In the present study, an attempt was made to utilize three-dimensional computed tomography (3D-CT) images to evaluate mandibular condyle fractures and identify prognostic indicators of malocclusion after closed treatment. Accurate morphometric measurements were performed using 3D-CT images obtained before trauma, after trauma, and after healing. Morphometry revealed significant differences in loss of ramus height (LRH) and lateral movement length in patients with malocclusion, and significant LRH differences in patients with other maxillomandibular fractures after healing, or in those with dislocation-displacement. The present method of 3D-CT image analysis appears useful for evaluation of condylar fractures.