Biomedical subjects

H Kagami

Publications and source records attributed to H Kagami.

At least 19 recordsLinked to original sources

PubMed2007

Effective bone engineering with periosteum-derived cells.

Bone augmentation via tissue engineering has generated significant interest. We hypothesized that periosteum-derived cells could be used in place of bone marrow stromal cells (which are widely used) in bone engineering, but the differences in osteogenic potential between these 2 cell types are unclear. Here, we compared the osteogenic potential of these cells, and investigated the optimal osteoinductive conditions for periosteum-derived cells. Both cell types were induced, via bFGF and BMP-2, to differentiate into osteoblasts. Periosteal cells proliferated faster than marrow stromal cells, and osteogenic markers indicated that bone marrow stromal cells were more osteogenic than periosteal cells. However, pre-treatment with bFGF made periosteal cells more sensitive to BMP-2 and more osteogenic. Transplants of periosteal cells treated with BMP-2 after pre-treatment with bFGF formed more new bone than did marrow stromal cells. Analysis of these data suggests that combined treatment with bFGF and BMP-2 can make periosteum a highly useful source of bone regeneration.

Adolescent↗

PubMed2006

Shear stress facilitates tissue-engineered odontogenesis.

Numerous studies have demonstrated the effect of shear stress on osteoblasts, but its effect on odontogenic cells has never been reported. In this study, we focused on the effect of shear stress on facilitating tissue-engineered odontogenesis by dissociated single cells. Cells were harvested from the porcine third molar tooth at the early stage of crown formation, and the isolated heterogeneous cells were seeded on a biodegradable polyglycolic acid fiber mesh. Then, cell-polymer constructs with and without exposure to shear stress were evaluated by in vitro and in vivo studies. In in vitro studies, the expression of both epithelial and mesenchymal odontogenic-related mRNAs was significantly enhanced by shear stress for 2 h. At 12 h after exposure to shear stress, the expression of amelogenin, bone sialoprotein and vimentin protein was significantly enhanced compared with that of control. Moreover, after 7 days, alkaline phosphatase activity exhibited a significant increase without any significant effect on cell proliferation in vitro. In vivo, enamel and dentin tissues formed after 15 weeks of in vivo implantation in constructs exposure to in vitro shear stress for 12 h. Such was not the case in controls. We concluded that shear stress facilitates odontogenic cell differentiation in vitro as well as the process of tooth tissue engineering in vivo.

Alkaline Phosphatase↗

PubMed2005

Selective and efficient culturing of retinal pigment epithelial cells using a feeder layer.

BACKGROUND: The techniques to isolate and purify retinal pigment epithelial (RPE) cells from small piece of autologous tissues are extremely difficult, and it is important to develop an efficient cell culture technique for RPE cells. The purpose of this study was to investigate the effect of 3T3-J2 cells and conditioned medium from 3T3-J2 cells on the proliferation of cultured RPE cells. METHODS: RPE cells from pigmented rabbits and a human RPE-derived cell line, ARPE-19, were used. First, the effects of co-culturing RPE cells with 3T3-J2 cells on the growth of the cells were analyzed. Second, the effects of the conditioned medium from 3T3-J2 cells on the proliferation of both types of cells were investigated. And third, the effects of the conditioned medium on RPE cell culture from a surgically removed choroidal neovascular (CNV) membrane were investigated. RESULTS: The 3T3-J2 cells increased the proliferation of both rabbit RPE cells and ARPE-19 cells. The number of rabbit RPE cells cultured in a mixture of the conditioned medium from 3T3-J2 cells was significantly higher than that in the reported optimal condition, and a similar tendency was observed for ARPE-19 cells. The results from enzyme-linked immunosorbent assay showed the presence of PDGF-AB, VEGF and IGF-I in the conditioned medium. The conditioned medium also promoted selective growth of human RPE cells from CNV. DISCUSSION: The results from this study present the conditions for efficient and selective culture of primary RPE cells.

3T3 Cells↗

PubMed2005

Differentiation of transplanted mesenchymal stem cells in a large osteochondral defect in rabbit.

BACKGROUND: Although accumulating evidence shows that mesenchymal stem cells (MSC) are a promising cell source for articular cartilage repair, the fate of transplanted MSC has not been extensively studied. METHODS: To monitor their persistence and differentiation, we labeled uninduced MSC with a fluorescent dye, PKH26, and transplanted them, in a poly-glycolic-acid scaffold, to full-thickness defects made in the weight-bearing area of rabbit femoral trochleae with hyaluronate sheets. The fate of the labeled cells was monitored for up to 8 weeks. RESULTS: Two weeks after transplantation, immature cartilage containing collagen type II had formed. By 8 weeks, this cartilage had thinned and immunolabeling for collagen type II gradually disappeared from the basal region, which became positive for collagen type I. Most chondrocytes within the regenerated cartilage were PKH26-positive and, therefore, derived from transplanted MSC, whereas osteoblasts within the regenerated bone were a mixture of donor- and host-derived cells. The thickness of the cartilage became thinner up to 8 weeks and then remained stable up to 42 weeks after surgery. DISCUSSION: These results showed that uninduced MSC were able to survive osteochondral defects and differentiated according to the environment, making a major contribution to initial cartilage formation and a partial contribution to bone regeneration.

Animals↗

PubMed2004

Transplantation of cultured salivary gland cells into an atrophic salivary gland.

Patients with dry mouth have been treated with salivary substitutes and/or medications such as pilocarpine or cevimeline hydrochloride. These treatments temporarily relieve their symptoms and induce salivation from residual tissue. However, no treatment is available for the purpose of regenerating an atrophic gland. In this study, the feasibility of a cell transplantation therapy for the atrophic submandibular glands was investigated in rats. Further, the potential of cell differentiation into a useful phenotype was assessed by immunohistochemistry together with cell tracking with the fluorescent dye PKH 26. Rat submandibular glands were excised, and the salivary gland epithelial cells were cultured for 3 weeks with 3T3 cells as a feeder layer. Ductal ligation of the submandibular gland was employed to generate an atrophic gland. One week after the operation, the ligation was removed, and the cultured cells labeled with PKH 26 were injected into the atrophic submandibular glands. As a control, the cultured cells were also injected into normal submandibular glands. Two weeks after cell transplantation, the transplanted cells were detectable in both the experimental and control groups. The cells were clustered in the connective tissue between the lobules. Four weeks after transplantation, the labeled cells were detectable in the experimental group but not in the control group. In the atrophic glands, the scattered transplanted cells were observed over a broad area of the gland but localized mainly around the acini and ductal region. Immunostaining results showed a possible involvement of the transplanted cells in ductal regeneration, while neither myoepithelial nor acinar differentiations were observed within the 4 weeks since transplantation. This study demonstrated that cell transplantation to the salivary gland is feasible, and that the transplanted cells were selectively attracted to and remained in the damaged area without affecting normal tissue.

3T3 Cells↗

PubMed2003

Small bowel bacterial overgrowth and rice malabsorption in healthy and physically disabled older adults.

AIM: To determine whether there are differences in small bowel bacterial overgrowth (SBBO) and rice digestion between healthy and disabled older adults and to estimate the influence of physical activity on these nutritional statuses. METHOD: Fifteen disabled adults who commute to a day-care centre and 11 healthy older adults participated in this study. SBBO and rice absorption were judged using a breath hydrogen test. Physical activity was estimated using a pedometer. RESULTS: The average number of steps taken per day by the disabled was 1056 +/- 243, which was statistically lower than that of the healthy, 6904 +/- 782 (P < 0.001). No SBBO-positive subject was seen in the healthy group, whereas five (33.3%) of 15 disabled older adults were SBBO-positive. After ingesting glucose solution, the triangle up H2 of disabled subjects was higher than that of the healthy subjects (7.6 +/- 2.7 versus 0.5 +/- 0.3 p.p.m., P < 0.05). Rice malabsorption was seen in one (9.1%) of 11 in the healthy and two (14.3%) of 14 in the disabled groups, which was not statistically significant. CONCLUSIONS: Disabled older people who have a physically inactive lifestyle are at risk of SBBO, probably because of a reduction in their intestinal motility. SBBO has no influence on absorption of rice, and some older adults, independent of physical condition, can not absorb rice adequately.

Aged↗

PubMed2003

Small intestinal metastasis from renal cell carcinoma exhibiting rare findings.

Small intestinal metastasis from renal cell carcinoma (RCC) has only rarely been described. We report two patients who developed small bowel metastases from RCC showing different clinicopathological characteristics. Both patients underwent hemilateral nephrectomy for RCC and developed lung metastases metachronously or simultaneously. One patient developed occlusive ileus caused by multiple polypoid tumours composed of sarcomatoid tissue in the jejunum shortly after nephrectomy. The other patient presented melaena due to bleeding from a Borrmann 2-like tumour in the jejunum six years after nephrectomy. Clinically, his disease was slow-growing. Sarcomatoid histology and Borrmann 2-like tumour in this report are rare findings in metastatic tumour of RCC in the small bowel.

Antineoplastic Agents↗

PubMed2001

Gastric cancer with Virchow's and multiple lung metastases showing a remarkable response to preoperative chemotherapy: report of a case.

We report herein a rare case with advanced gastric cancer combined with group 4 lymph node and lung metastases that responded remarkably to neoadjuvant chemotherapy. A 65-year-old man was found to have a well-differentiated type 3 gastric cancer that invaded the duodenum locally and was accompanied with Virchow's, para-aortic lymph nodes, and multiple lung metastases based on physical, endoscopic, and radiological examinations. In addition, his carbohydrate antigen (CA) 19-9 was elevated to 3965U/ml, and CA72-4 to 46U/ml. Prior to surgery, he was treated with 5-fluorouracil (5-FU; 500mg/body per day) and low-dose cisplatinum (CDDP; 10mg/body per day) as neoadjuvant chemotherapy for 6 weeks. As a result, a partial response was obtained in all lesions, and CA19-9 and CA72-4 decreased to 463U/ml and 9.4U/ml, respectively. Four weeks after the completion of neoadjuvant chemotherapy, a distal gastrectomy was performed, and a histopathological examination of the resected specimen showed a grade 2 response to chemotherapy. Immunohistochemically, the thymidylate synthase expression level was very low in the tumor tissues, which might account for the good response to the combination chemotherapy with 5-FU and CDDP observed in the present case.

Adenocarcinoma↗

PubMed2001

Restriction of proliferation of primordial germ cells by the irradiation of Japanese quail embryos with soft X-rays.

Primordial germ cells (PGCs) are the progenitor cells for the gametes. Avian PGCs are located in the central region of the area pellucida at the blastoderm stage. Shortly after further incubation, they migrate to the extra-embryonic germinal crescent, and then as soon as the blood vessels form, they enter the circulation and finally settle in the gonadal primordium. We have developed a simple method using soft X-ray irradiation (18 kV power, 20 cm distance) to reduce the number of PGCs in Japanese quail embryos, which should be useful in preparing recipient embryos for PGC-transfer studies. When embryos were exposed to the soft X-rays for 40 s before incubation, the concentration of circulating PGCs was less than one-fifth that in controls after 2 days of incubation. Embryos at day 6 of incubation contained approximately half the number of PGCs compared to controls when they were exposed before or at day 2 of incubation. Irradiation for 40 s is recommended taking into consideration the restriction of proliferation of PGCs, and viability and hatchability.

Animals↗

PubMed2001

Suicide gene therapy for human oral squamous cell carcinoma cell lines with adeno-associated virus vector.

The purpose of this study was to test the possibility of gene transfer as a new therapy for oral cancer. Adeno-associated virus (AAV) has already been used in the fields of cystic fibrosis and Parkinson's disease as a potential vector for gene therapy because of its wide host range, high transduction efficiency, and lack of cytopathogenicity. Four human oral squamous cell carcinoma cell lines were transduced with an AAV vector containing the beta-galactosidase gene (AAVlacZ) in vitro. Gene transduction efficiency was from 20 to 50% at a multiplicity of infection (MOI; for the purposes of this study the number of vector genomes per target cell) of 1x10(3), and nearly 100% of each cell line were transduced at an MOI of 1x10(4). Next, four cell lines were transduced with an AAV vector containing the herpes simplex virus thymidine kinase (HSVtk) gene, which sensitizes transduced cells to ganciclovir (GCV). Subsequent administration of GCV resulted in nearly 100% tumor cell killing at an MOI of 1x10(4) and from 70 to 80% tumor cell killing at an MOI of 1x10(3). These results suggest that AAV-mediated gene transfer of HSVtk and administration of GCV has potential as a new therapy for oral squamous cell carcinoma.

Antiviral Agents↗

PubMed2001

Cloning of porcine IGF1 receptor cDNA and detection of sequence polymorphisms using RT-PCR.

A cDNA encoding the porcine type 1 insulin-like growth factor receptor (IGF1R) was amplified by reverse transcriptase-polymerase chain reaction (RT-PCR). The sequence of a 4.2-kb product was determined and had an open reading frame, encoding 1367 amino acids with 98.1 and 95.2% sequence similarity to the human and rat IGF1R, respectively. In the comparison of RT-PCR derived IGF1R sequences from 12 unrelated pigs, 12 silent sequence variants were found.

Animals↗

PubMed2000

Acceleration of rat salivary gland tissue repair by basic fibroblast growth factor.

A model of atrophic rat submandibular gland was used to examine the ability of basic fibroblast growth factor (bFGF) to accelerate tissue repair. The gland duct was separated carefully from associated blood vessels and nerve, and ligated with a 8-0 suture under a surgical microscope. Two weeks after ligation, the glandular tissue showed severe atrophy and weight loss (to 26% of that in a sham-operated group). Thereafter, the ligature was removed and various amounts of bFGF, isoproterenol or saline were instilled retrogradely through the duct. Both isoproterenol and bFGF increased cell proliferation significantly. bFGF accelerated the proliferation of various cell types, including both acinar and ductal. The proliferative effects of bFGF peaked at a dose of 1 ng/gland. When bFGF (1 ng/gland) was administered to the atrophic gland, its weight increased to 125% of the glands in saline-treated control animals after 2 weeks. The effects of bFGF were also examined in normal submandibular glands: bFGF stimulated cell proliferation, but the effective concentration was at least 50 times higher than that required in the atrophic gland. The results from immunohistochemical tests against anti-FGF receptor-type 1 antibody demonstrated increased immunoreactivity in the damaged gland, which might be involved in the difference in the response to bFGF between damaged and normal glands. Overall, the results indicate that bFGF can accelerate tissue repair in salivary gland.

Adrenergic beta-Agonists↗

PubMed2000

Effects of basic fibroblast growth factor on cultured rat and human submandibular salivary gland cells.

Basic fibroblast growth factor (bFGF) is a strong mitogen for most mesoderm- and ectoderm-derived cells. Although bFGF exists in rat and human salivary glands, its physiological role in those glands is unknown. In this study, the effects of bFGF were investigated in monolayer culture of normal rat and human submandibular gland cells. Epithelial cells from rat and human submandibular glands were cultivated with the aid of 3T3 cells as a feeder layer. The effects of different concentrations of bFGF on the second passage of these cultured cells were examined. In both the rat and human cells, the percentage of bromodeoxyuridine (BrdU)-positive cells gradually increased up to 50 ng/ml, and then increased sharply at 100 ng/ml. However, at concentrations higher than 100 ng/ml, the percentages of BrdU-positive cells reached a plateau. In both rat and human cells, total cell numbers at 100 ng/ml bFGF were significantly higher than those of the control group from culture day 4. On the other hand, the morphology of the cultured cells showed no difference either with or without bFGF. These results indicate that a major effect of bFGF on salivary gland epithelial cells is to act as a mitogenic stimulus.

3T3 Cells↗

PubMed2000

Construction and function of a recombinant adenovirus encoding a human aquaporin 1-green fluorescent protein fusion product.

Transfer of the human aquaporin 1 (hAQP1) gene provides a novel way to potentially correct the severe salivary hypofunction associated with therapeutic radiation for head and neck cancer. To facilitate the study of individual cells transduced with this gene, we have designed a fusion product of the hAQP1 and jellyfish green fluorescent protein (GFP) cDNAs. An expression plasmid, pACCMVhAQP1GFP, and a recombinant adenovirus, AdhAQP1GFP, encoding this fusion product were constructed. Both the recombinant plasmid and virus directed the expression of the encoded, 55-kDa fusion protein (hAQP1GFP), which was detected in the plasma membranes of several epithelial cell lines (293, SMIE, and A5). hAQP1GFP was functionally active and facilitated fluid movement across a polarized salivary epithelial cell monolayer (approximately 5-fold noninfected controls) in response to an osmotic gradient. In response to a hypotonic challenge, individual epithelial cells expressing the fusion protein exhibited significantly more capacitance (used herein as an indicator of cell swelling) than control cells. Conversely, in response to a hypertonic challenge, individual infected cells shrunk more rapidly (approximately 2- to 3-fold) and to a greater extent than control cells. We conclude that AdhAQP1GFP is a useful experimental tool to identify and study individual cells expressing a water channel transgene.

Adenoviridae↗

PubMed2000

Medial (intra-cisternal) acoustic neuromas.

The clinical characteristics of "medial" or "intra-cisternal" acoustic neuroma (AN) treated in our institute were reviewed. Among 466 patients with ANs in our series during the last 20 years, 6 patients (1.3%) were considered to fill the criteria of medial AN definition. Compared with those with non-medial ANs, the patients with medial ANs show a tendency to have cerebellar and/or cranial nerve dysfunction (especially trigeminal and/or facial nerves) in addition to hearing loss at the time of initial presentation. On magnetic resonance imaging, medial AN is visualized as a multi-cystic mass lesion in the cerebello-pontine cistern without extension into the internal auditory canal in most cases. Although total removal of tumor was achieved in all cases, the results of preservation of facial nerve function were not satisfactory. Medial AN can be considered as a clinical, but not pathological, subtype in terms of the functional outcomes of the facial nerve and hearing.

Facial Nerve↗

PubMed2000

Salivary growth factors in health and disease.

The salivary gland is considered to be a reservoir of many growth factors in rodents. In humans, the epidermal growth factor, basic fibroblast growth factor, and insulin and insulin-like growth factor family have also been detected in this gland, but their physiological role remains unclear. In this study, we focused on bFGF, which is a well-known mitogen for various types of cells, and is present in the salivary gland as well as in saliva. The roles of bFGF in the salivary gland were investigated by three different procedures. First, the effects of bFGF on the salivary gland cells were investigated with a monolayer culture of normal submandibular gland cells. The effects of different concentrations of bFGF on the second passage of these cultured cells were examined. In both human and rat cultured submandibular gland cells, bFGF accelerated the cell proliferation at a concentration of 100 ng/mL or higher. Next, an atrophic model of the rat submandibular gland was used to examine the ability of bFGF to accelerate tissue repair. Two weeks after ductal ligation, the ligature was removed, and various amounts of bFGF, isoproterenol, or saline were administered via a retrograde duct instillation. Both isoproterenol and bFGF increased acinar and ductal cell proliferation significantly. To determine the role of bFGF in saliva, we investigated its effect on the healing process of oral mucosal defects. Four-millimeter mucosal defects were made to the depth of the periosteum in the rat palate under anesthesia. bFGF or vehicle alone was applied once only at the time of surgery as a suspension. At days 3, 5, and 7 in the bFGF group, significant increases in the degree of re-epithelialization were found in treated groups. These results indicate that its action as a mitogen stimulus is the major effect of bFGF on salivary gland cells and mucosal epithelium.

Animals↗

PubMed1999

Polarized secretion of transgene products from salivary glands in vivo.

Previously (Kagami et al. Hum. Gene Ther. 1996;7:2177-2184) we have shown that salivary glands are able to secrete a transgene-encoded protein into serum as well as saliva. This result and other published data suggest that salivary glands may be a useful target site for vectors encoding therapeutic proteins for systemic delivery. The aim of the present study was to assess in vivo if transgene-encoded secretory proteins follow distinct, polarized sorting pathways as has been shown to occur "classically" in cell biological studies in vitro. Four first-generation, E1-, type 5 recombinant adenoviruses were used to deliver different transgenes to a rat submandibular cell line in vitro or to rat submandibular glands in vivo. Subsequently, the secretory distribution of the encoded proteins was determined. Luciferase, which has no signal peptide, served as a cell-associated, negative control and was used to correct for any nonspecific secretory protein release from cells. The three remaining transgene products tested, human tissue kallikrein (hK1), human growth hormone (hGH), and human alpha1-antitrypsin (halpha1AT), were predominantly secreted (>96%) in vitro. Most importantly, in vivo, after a parasympathomimetic secretory stimulus, both hK1 and hGH were secreted primarily in an exocrine manner into saliva. Conversely, halpha1AT was predominantly secreted into the bloodstream, i.e., in an endocrine manner. The aggregate results are consistent with the recognition of signals encoded within the transgenes that result in specific patterns of polarized protein secretion from rat submandibular gland cells in vivo.

Adenoviridae↗

PubMed1999

Re-engineering the functions of a terminally differentiated epithelial cell in vivo.

Because of their easy access, and important role in oral homeostasis, mammalian salivary glands provide a unique site for addressing key issues and problems in tissue engineering. This manuscript reviews studies by us in three major directions involving re-engineering functions of salivary epithelial cells. Using adenoviral-mediated gene transfer in vivo, we show approaches to i) repair damaged, hypofunctional glands and ii) redesign secretory functions to include endocrine as well as exocrine pathways. The third series of studies show our general approach to develop an artificial salivary gland for clinical situations in which all glandular tissue has been lost.

Animals↗