Kunihisa Taniguchi

A poly(γ-glutamic acid)–amphiphile complex as a novel nanovehicle for drug delivery system

Recently, many studies have focused on biomedical and pharmaceutical applications of self-assembled nanoparticles. In addition, several biodegradable nanoparticles have been reported to possess poor dispersion stability and poor size-controllability. However, these nanoparticles require complicated fabrication procedures using synthesis techniques. We developed an efficient method for producing nanoparticles derived from a biological origin of molecule poly(γ-glutamic acid) (γ-PGA), a cationic lipid, and doxorubicin (Dox). The complex had a size of 510 nm and was able to encapsulate over 90% of the added Dox. An in vivo assay of antitumor activity demonstrated that the complex had significant antitumor activity in sarcoma 180–bearing mice, and was effectively accumulated in solid tumors based on the EPR effect. The data suggested that this complex is a promising formulation of γ-PGA for targeted delivery to solid tumors. γ-PGA–12GP2 complexes may possess several unique advantages, including simplicity of nanoparticle preparation, high drug-carrying capacity, appropriate size to allow deeper penetration based on EPR effect into solid tumors, and lack of necessity to modify the chemical structure of the drugs. These data indicate that the γ-PGA–12GP2 complexes are potentially useful in cancer chemotherapy.

Preparation of and Tissue Response to DNA-Lipid Films

DNA-containing films have the potential to form complexes with antibiotics or cytokines by intercalation or groove-binding. This principle can be highly relevant for regenerative wound-healing around oral implants and in periodontology. In this study, we prepared DNA-lipid films and examined tissue responses to them as an indicator of their biological properties. The lipids were synthesized from the reaction of L-alanine, n-alkyl alcohol, and p-toluenesulfonic acid. We prepared the self-standing, water-insoluble DNA-lipid films by casting the DNA-lipid complex from chloroform/ethanol solution. The DNA-lipid complexes, which had 1:1 ratios of phosphate anions to cationic lipid, were found to have a double-helical B-form structure. The DNA-lipid films were almost dissolved 3 days after subcutaneous implantation in the backs of rats. There were no inflammatory reactions or inhibition of new tissue formation. We concluded that DNA-lipid films can be prepared by simple methods, and that they do not cause an unfavorable tissue response.

Solitary neurofibroma of the gingiva with prominent differentiation of Meissner bodies: a case report

BackgroundOral neurofibromas are peripheral nerve sheath tumors, similar to schwannomas. Histological variations in oral neurofibromas are relatively uncommon.Case presentationHere, we present a case of unique variation in the observed characteristics of a neurofibroma, with no relation to neurofibromatosis type-1 or von Recklinghausen disease of the skin. The neurofibroma was observed in the right mandibular gingiva of a 32-year-old Japanese woman. Histologically, it differed from conventional neurofibromas in that the tumor was composed of a mixture of fine fibrillary collagen in sheets and/or cords of neoplastic Schwann cells containing numerous clusters of Meissner bodies. Histologically, these bodies were in contact with neoplastic Schwann cells. The Meissner bodies were immunopositive for S-100 protein, neuron-specific enolase, and vimentin, but were negative for calretinin. CD34-positive spindle cells were observed around the Meissner bodies. No recurrence or signs of other tumors have been observed in the patient for 5 years after tumor resection.ConclusionTo the best of our knowledge, no formal descriptions of sporadic, solitary neurofibromas containing numerous Meissner bodies occurring in the oral cavity are available in literature. We believe that an uncommon proliferation of Meissner bodies, as seen in the present case, may result from aberrant differentiation of neoplastic Schwann cells.

An immunohistochemical and ultrastructural study of vasomotor nerves in the microvasculature of human dental pulp

Immunohistochemical methods for S-100 protein and neurone-specific enolase showed two types of nerve endings around the pulp microvasculature: the free endings, which comprise the major neural component and are distributed in all types of microvessel such as arterioles, venules and capillaries; and the varicose endings. The varicose ending was a less frequent, minor component observed only in the arterioles. Both immunohistochemical and ultrastructural observations confirmed that the varicose endings were the terminal axons of efferent vasomotor nerves. Further extensive ultrastructural examinations on the vasomotor nerves added the following new findings to our previous reports. Vasomotor nerves sometimes ramified into more than two terminal axons around arterioles, and most of these ramified axons ended in the adventitia-media junction of the arteriolar wall; however, nerve endings occasionally penetrated into the media. These findings suggest an intimate structural association between vasomotor nerves and arterioles in regulating the arteriolar microcirculation in human dental pulp by stimulating smooth-muscle cells not only of the outermost but also of the inner layers. Furthermore, the deep penetration of terminal axons into the arteriolar wall seems to provide effective regulation of pulpal blood flow under physiological and pathological conditions.

Ultrastructure of the neuromuscular junction of vasomotor nerves in the microvasculature of human dental pulp

Vasomotor nerves in human dental pulp were more closely related to arterioles than to venules. Most were composed of unmyelinated fibres, which were mainly adrenergic. They appeared close to arterioles that were surrounded by a few layers of contractile smooth-muscle cells. The smaller arterioles with a diameter of 10-15 microns received a more intimate innervation by vasomotor nerves than did the larger. These vessels occasionally showed much narrower neuromuscular junctions than previously reported. Most of these nerve fibres were identified as adrenergic by the presence of chromaffin-positive synaptic vesicles detected by ultrastructural enzyme histochemistry. Their function appeared to be to regulate the blood flow and/or the blood pressure by stimulating smooth-muscle cells, resulting in contraction and a change in the calibre of the vessels. Capillaries and venules, which have a higher permeability, received weaker innervation by the vasomotor nerves than did arterioles. The intimate relation between vasomotor nerves and arterioles is related to the function of dental pulp in normal and pathological conditions.

Tumor-induced lymphangiogenesis in cervical lymph nodes in oral melanoma-bearing mice

BackgroundMetastasis via the lymphatic system is promoted by lymphangiogenesis. Alterations of the lymphatic channels during the progression of metastasis to regional lymph nodes (LNs) remain unexplored. To examine whether tumor-induced LN lymphangiogenesis controls metastasis to regional LNs, we investigated cervical LN metastasis in a mouse model of oral melanoma.MethodsInjection of B16F10 melanoma cells into mouse tongues replicated spontaneous cervical LN metastasis. We performed histological, immunofluorescent, and histomorphometric analyses of tumor-reactive lymphadenopathy and lymphangiogenesis in tumor-associated LNs. We investigated the expression of vascular endothelial growth factor (VEGF)-C and its receptor, VEGF receptor-3 (VEGFR-3), in tumor cells and tissues, and LNs by reverse transcription polymerase chain reaction and immunofluorescence.ResultsTumor-associated LNs comprised sentinel LNs (SLNs) before and after tumor cell invasion (tumor-bearing SLNs), and LNs adjacent or contralateral to tumor-bearing SLNs. Extensive lymphangiogenesis appeared in SLNs before evidence of metastasis. After metastasis was established in SLNs, both LNs adjacent and contralateral to tumor-bearing SLNs demonstrated lymphangiogenesis. Interaction between VEGF-C-positive melanoma cells and VEGFR-3-positive lymphatic vessels was evident in tumor-associated LNs.ConclusionsLN lymphangiogenesis contributes a progression of tumor metastasis from SLNs to other regional LNs.

Effects of bFGF on suppression of collagen type I accumulation and scar tissue formation during wound healing after mucoperiosteal denudation of rat palate

Objective. The purpose of this study was to evaluate the effect of basic fibroblast growth factor (bFGF) on collagen changes after mucoperiosteal denudation of rat palate. Material and methods. A total of 36 male Wistar rats were divided into control, scar, sham, and bFGF groups. In the scar, sham, and bFGF groups, lateral palatal mucoperiosteum was excised to form scar tissue on the palate. In the bFGF group, bFGF solution was injected into the operated area 1 week postoperatively. At 6 weeks postoperatively, the distribution of collagen type I and the 3-dimensional structure of collagen fibers were investigated under immunofluorescent and scanning electron microscopy. Result. In the bFGF group, weakly immunostained submucosa was clearly distinguishable from the strongly immunostained cervical periodontal ligament and gingiva. Collagen fibers running from submucosal tissue into the surface of underlying palatal bone comprised loosely arranged collagen fibrils. Lumen structures in collagen fibers resembled those in the control group. Conclusion. Administration of bFGF for suppression of collagen type I generation could suppress scar tissue formation and reduce connective strength with adjacent teeth and palatal bone.

Influence of carbon dioxide laser irradiation on the healing process of extraction sockets

Abstract Objective. To clarify the healing-promoting effects of carbon dioxide laser irradiation in high and low reactive-level laser therapies (HLLT and LLLT, respectively) on extraction sockets after tooth extraction. Material and methods. Forty-two 5-week-old male Wistar rats were divided into laser irradiation and non-irradiation (control) groups and compared. The laser-irradiation group underwent HLLT immediately after tooth extraction and then LLLT 1 day post-extraction. Tissue was excised 6 h and 3, 7, or 21 days after extraction and histopathologically investigated. The alveolar crest height was measured osteomorphometrically 21 days post-extraction, and granulation tissue in the extraction socket surface layer was immunohistologically investigated using anti-α-smooth muscle actin (anti-α-SMA) antibody 3 and 7 days post-extraction. Results. Many osteoclasts appeared and active bone resorption was noted in the irradiation group 3 days after extraction compared to the controls. On Day 7, new bone formation started around the extraction socket in the control group, but from the superficial to over the middle layer of the socket in the irradiation group. On Day 21, a concavity existed in the alveolar crest region in the controls, whereas this region was flat, with no concavity, in the irradiation group. On osteomorphometry, the alveolar crest height was significantly higher in the irradiation (0.7791 ± 0.0122) than the control (0.6516 ± 0.0181) group (P < 0.05). On immunostaining, many α-SMA-positive cells were noted in the control group, but very few in the irradiation group. Conclusion. Laser-irradiated extraction wound healing showed characteristics different from those of the normal healing process, suggesting a favorable healing-promoting effect.

Effects of Basic Fibroblast Growth Factor Administration on Vascular Changes in Wound Healing of Rat Palates

Objective: The purpose of this study was to investigate the vascular changes induced by mucoperiosteal denudation of rat palate and to elucidate the effects of basic fibroblast growth factor (bFGF) administration on the palatal vascular network in wound healing. Methods: A total of 117 male Wistar rats were used for the study on their 20th postnatal day. The animals were divided into three groups: a scar formation group, a basic fibroblast growth factor group, and a control group. The scar formation and basic fibroblast growth factor groups had lateral mucoperiosteum excised from the palate. In the basic fibroblast growth factor group, a solution of basic fibroblast growth factor was injected into the operated area 1 week after excision. At 6, 8, and 10 weeks postoperatively, palatal vascular changes were investigated by immunohistochemical staining and corrosion cast techniques. Results: Throughout the experimental period, there were significantly fewer vessels in the scar formation group than in the control and basic fibroblast growth factor groups. In the basic fibroblast growth factor group, the elongation of new vessels and capillary proliferation proceeded, and after 10 weeks a highly organized vascular network was established. The scar formation group showed few Volkmanns canals that were shrunken or closed, whereas the basic fibroblast growth factor group evidenced Volkmanns canals with arterioles or venules, as seen in the control. Conclusions: The results suggested that injection of basic fibroblast growth factor into palatal wounds improves the vascular supply to the operated mucosa and underlying bone during and after palatal wound healing, which may contribute to tissue remodeling of the palate during growth.

Ultrastructural relation between nerve terminals and dentine bridge formation after pulpotomy in human teeth

Close association between nerve terminals and preodontoblasts, odontoblasts and predentine was observed during healing after pulpotomy. The nerve terminals frequently contained large numbers of synaptic vesicles. Terminals with many vesicles tended to be fewer in the predentine than in the odontoblastic layer. The distribution of terminals was more dense at the stage before the regenerated odontoblasts became arranged regularly beneath the predentine. It is suggested that these terminals have some efferent role(s), especially during collagen synthesis at the early stage of dentinogenesis. The nerves may release their abundant synaptic vesicles, in addition to serving a sensory role for monitoring the increased sensitivity in the injured areas.