A PTHrP Gradient Drives Mandibular Condylar Chondrogenesis via Runx2.

The mandibular condylar cartilage (MCC) is an essential component of the temporomandibular joint, which orchestrates the vertical growth of the mandibular ramus through endochondral ossification with distinctive modes of cell differentiation. Parathyroid hormone-related protein (PTHrP) is a master regulator of chondrogenesis; in the long bone epiphyseal growth plate, PTHrP expressed by resting zone chondrocytes promotes chondrocyte proliferation in the adjacent layer. However, how PTHrP regulates chondrogenesis in the MCC remains largely unclear. In this study, we used a Pthrp-mCherry knock-in reporter strain to map the localization of PTHrP+ cells in the MCC and define the function of PTHrP in the growing mandibular condyle. In the postnatal MCC of PthrpmCherry/+ mice, PTHrP-mCherry was specifically expressed by cells in the superficial layer immediately adjacent to RUNX2-expressing cells in the polymorphic layer. PTHrP ligands diffused across the polymorphic and chondrocyte layers where its cognate receptor PTH1R was abundantly expressed. We further analyzed the mandibular condyle of PthrpmCherry/mCherry mice lacking functional PTHrP protein (PTHrP-KO). At embryonic day (E) 18.5, the condylar process and MCC were significantly truncated in the PTHrP-KO mandible, which was associated with a significant reduction in cell proliferation across the polymorphic layer and a loss of SOX9+ cells in the chondrocyte layers. The PTHrP-KO MCC showed a transient increase in the number of Col10a1+ hypertrophic chondrocytes at E15.5, followed by a significant loss of these cells at E18.5, indicating that superficial layer-derived PTHrP prevents premature chondrocyte exhaustion in the MCC. The expression of Runx2, but not Sp7, was significantly reduced in the polymorphic layer of the PTHrP-KO MCC. Therefore, PTHrP released from cells in the superficial layer directly acts on cells in the polymorphic layer to promote proliferation of chondrocyte precursor cells and prevent their premature differentiation by maintaining Runx2 expression, revealing a unique PTHrP gradient-directed mechanism that regulates MCC chondrogenesis.

Periostin of human periodontal ligament fibroblasts promotes migration of human mesenchymal stem cell through the αvß3 integrin/FAK/PI3K/Akt pathway.

BACKGROUND AND OBJECTIVE: The periodontal ligament (PDL) is characterized by rapid turnover, high remodeling capacity and high inherent regenerative potential compared with other connective tissues. Periostin, which is highly expressed in the fibroblasts in the PDL, has been widely discussed in relation to collagen fibrillogenesis in the PDL. Recently, several reports have indicated periostin in cell migration. The aim of this study was to examine whether human PDL fibroblasts (hPDLFs) with high levels of periostin expression promote the migration of human bone marrow mesenchymal stem cells (hMSCs). MATERIAL AND METHODS: The migration of hMSCs was examined by transwell chamber migration assay under different conditions: medium alone, hPDLFs, human dermal fibroblasts, recombinant periostin, integrin αvß3 blocking antibody (anti-CD51/61 antibody) and inhibitors of FAK (PF431396) and PI3K (LY294002). Phosphorylation of FAK and Akt in hMSCs under stimulation of periostin was examined by western blotting. RESULTS: The migration assay revealed that the number of migrated hMSCs by hPDLFs was significantly larger than those by dermal fibroblasts, periostin small interfering RNA hPDLFs and medium alone. Furthermore, recombinant periostin also strongly induced hMSC migration. The addition of anti-CD51/61 antibody, PF431396 and LY294002 caused a significant reduction in the number of migrated hMSCs respectively. The anti-CD51/61 antibody inhibited both FAK and Akt phosphorylations under periostin stimulation. PF431396 inhibited both FAK and Akt phosphorylations. LY294002 inhibited only Akt phosphorylation, and FAK phosphorylation was not influenced under periostin stimulation. CONCLUSION: Periostin expression in hPDLFs promotes the migration of hMSCs through the αvß3 integrin/FAK/PI3K/Akt pathway in vitro.

The effect of oral management on the severity of oral mucositis during hematopoietic SCT.

Oral mucositis (OM) is a frequent adverse effect of allogenic or autologous hematopoietic SCT. It results from direct toxic injury to the mucosal epithelial cells by the immunosuppressive regimen. Here, we compared the incidence and severity of OM between a group of 24 patients who received proper oral management during hematopoietic SCT and a group of 24 who did not. The oral management group received pre-hematopoietic SCT instruction on oral care and an oral examination in the clean room. Differences in the incidence and severity of OM between the two groups were examined statistically. OM was observed in 14 (58.3%) patients in the oral management group and 22 (91.6%) in the control group. The median of the OM score was 1 for the oral management group (range 0 to 3) and 2 for the control group (range 0 to 3). There was a significant difference in the OM score (P<0.05) and in the incidence of OM between the two groups (P<0.01). This study shows that oral management may decrease the occurrence of OM. Our results also suggest that it is important to include an oral management provider on the hematopoietic SCT team.

Fiber system degradation, and periostin and connective tissue growth factor level reduction, in the periodontal ligament of teeth in the absence of masticatory load.

BACKGROUND AND OBJECTIVE: The periodontal ligament (PDL), which is interposed between the alveolar bone and roots, supports teeth against mechanical stress. Periostin and connective tissue growth factor (CTGF) might play essential roles in maintaining PDL fiber integrity under mechanical stress. However, this relationship has not been studied at the protein and gene levels. Therefore, the aim of this study was to assess the PDL fiber system without masticatory load to determine the structural changes in the PDL in the absence of mechanical stress. MATERIAL AND METHODS: The study included 45 Wistar male rats (12 wk of age) whose upper-right first molars were relieved from occlusion for 24 h, 72 h, 7 d or 21 d. The PDL was examined histologically, and changes in the gene and protein levels of periostin and CTGF were investigated. RESULTS: The PDL space width was reduced significantly. Histologically, an initial reduction in the fiber number and thinning of PDL fibers were observed, followed by disarrangement of the PDL fibers and their attachments to the alveolar bone; finally, the PDL fibers lost their meshwork structure. Real-time RT-PCR results revealed sharp down-regulation of the periostin and CTGF mRNA levels at 24 and 72 h, respectively, which continued throughout the experiment. Immunohistochemical analysis revealed that periostin localized to both the cellular elements and the extracellular matrix, whereas CTGF localized only to the cellular elements. Periostin and CTGF immunoreactivities became very weak without masticatory load. CONCLUSION: In the absence of mechanical stress, the PDL fiber system undergoes degradation concomitantly with a reduction in the periostin and CTGF levels in the PDL.

Consumption of medium- and long-chain triacylglycerols decreases body fat and blood triglyceride in Chinese hypertriglyceridemic subjects.

OBJECTIVES: To investigate the effects of medium- and long-chain triacylglycerol (MLCT) on blood triglyceride (TG) in Chinese hypertriglyceridemic subjects. METHODS: A double-blind controlled clinical trial was carried out, in which 112 subjects with hypertriglyceridemia were randomly divided into two dietary oil groups: (1) long-chain triacylglycerol (LCT) and (2) MLCT. All subjects were requested to ingest fixed energy and to continue their normal activity levels, and to consume LCT or MLCT oil at 25-30 g daily during the study period. Anthropometric measurements of body weight, body mass index (BMI), body fat, body fat percentage, waist and hip circumference (WC and HC), areas of subcutaneous and visceral fat by computed tomography scanning and blood biochemical markers were measured at the beginning and end of the study. RESULTS: There were 50 and 51 subjects left in LCT and MLCT groups, respectively. There were no significant differences in daily intake of energy, protein, fat and carbohydrate, as well as the daily physical activity between the two groups during the study. After 8 weeks, MLCT group showed a significant decrease in body weight, BMI, WC, HC, ratio of WC and HC, body fat, body fat percentage and subcutaneous fat when compared with the initial values. The decrease in body weight, BMI, WC, body fat and subcutaneous and visceral fat was significantly greater in MLCT group than that in the LCT group. Furthermore, the serum concentrations of TG in MLCT group were significantly lower than those in the LCT group. CONCLUSIONS: Consumption of MLCT may reduce body weight, body fat and blood TG in hypertriglyceridemic subjects under an appropriate dietary regime.

Laser capture microdissection of rat periodontal ligament for gene analysis.

The periodontal ligament (PDL) is a connective tissue interposed between two hard tissues, viz., the root of a tooth and the alveolar bone, which makes it difficult to obtain directly. In the study reported here, PDL, subgingival connective tissue and pulp of rat molars were extracted directly by laser capture microdissection and the gene expression of TGF-beta1 on the microdissected PDL was examined. The maxillae of rats were dissected and rapidly immersed in isopentane cooled with liquid nitrogen. Serial frontal sections of the rat first molar area were used for immunohistochemistry and for laser capture microdissection to localize the TGF-beta1 gene. Gene expression and immunohistochemical localization of TGF-beta1 also were examined in the pulp and subgingival connective tissues. TGF-beta1 was located immunohistochemically in the fibroblasts in the PDL. A considerable amount of RNA was obtained by laser capture microdissection of these three tissues for analysis of gene expression. The reverse transcription- polymerase chain reaction demonstrated that glyceraldehyde 3-phosphate dehydrogenase was amplified in all three tissues, and that TGF-beta1 was detected in the PDL. Laser capture microdissection makes it possible to analyze the gene expression of PDL and expression of TGF-beta1 in the PDL suggests that this gene could function in maintaining PDL.

[Sequential changes in haemostatic activity during prolonged surgery and its alteration with continuous heparin infusion].

We measured molecular markers to study sequential changes in the hemostatic activity and its alteration by intraoperative continuous heparin infusion in patients, undergoing surgeries of 10 hours or longer for oral cancers. The heparin was infused continuously from the beginning of microsurgery until the end of anaesthesia to maintain an activated partial thromboplastin time between 50 to 70 seconds in the heparin group. In the control group, the concentrations of thrombin-antithrombin III complex (TAT), fragment 1 + 2 (F1 + 2) and D-dimer increased, and the soluble fibrin monomer complex (SFMC) became positive 2-6 hours after the induction of anesthesia. With continuous heparinization, the changes in measured molecular markers were clearly inhibited compared with the control group. The hemostatic activities increased progressively from the early stages of surgery, and the intraoperative continuous heparin infusion was effective in suppressing the hypercoagulable state during prolonged surgery.

Trehalose suppresses lipopolysaccharide-induced osteoclastogenesis bone marrow in mice.

We have previously shown that trehalose suppresses bone loss in ovariectomized (OVX) mice by way of inhibiting osteoclast differentiation in bone marrow. Also, trehalose inhibits the secretion of interleukin-6 in bone marrow cell cultures, resulting in a decrease in osteoclast formation. In this study, we examined the effect of trehalose on osteoclastogenesis using another model of bone resorption, namely lipopolysaccharide (LPS)-stimulated osteoclast induction. Mice were given trehalose (1g/kg) by gastric intubation for 5 consecutive days, and 24 hours later, 14 mg/kg of LPS was injected intraperitoneally. Trehalose significantly suppressed LPS-induced tumor necrosis factor (TNF)-alpha production after 90 min and decreased the number of osteoclasts in the bone marrow 48 hours after LPS injection. These results indicate that trehalose suppresses excessive osteoclastogenesis not only in OVX mice but also in a LPS-induced bone resorption mouse model and further suggest that the latter finding may be mediated at least in part through a decrease in TNF-alpha production.

Prophylactic action of oral fluconazole against fungal infection in neutropenic patients. A meta-analysis of 16 randomized, controlled trials.

BACKGROUND: Fluconazole is used widely for fungal prophylaxis. Although studies with bone marrow transplantation (BMT) recipients clearly showed the usefulness of oral fluconazole, results of the studies in neutropenic patients other than BMT recipients have been inconsistent. Therefore, the authors performed a meta-analysis to evaluate the efficacy of fluconazole prophylaxis during chemotherapy-induced neutropenia. METHODS: The authors identified reports that were not restricted to those in English and not restricted to published trials through MEDLINE, CANCERLIT, or the data base of the Pfizer company. The authors included prospective, randomized studies comparing oral fluconazole with placebo, no treatment, or oral polyenes as prophylaxis for fungal infections in neutropenic patients. Two independent authors extracted data from 16 trials with 3734 patients enrolled. The outcome measures were the development of fungal-related death, systemic and superficial fungal infections, the use of empiric intravenous amphotericin-B, and infections or colonization with fluconazole-resistant fungi. The summarized odds ratios (ORs) were calculated using the Mantel-Haenszel method and the DerSimonian-Laird method. RESULTS: Prophylactic fluconazole was not effective in reducing fungal-related death or in reducing proven, systemic fungal infections in non-BMT patients (OR, 0.91; 95% confidence interval [CI], 0.30-2.82 and OR, 0.85; 95% CI, 0.47-1.55, respectively). However, fluconazole was very effective in reducing superficial fungal infections (OR, 0.44; 95% CI, 0.24-0.80), even when it was given in lower doses (50-200 mg per day). There was no increase in proven, systemic infection of fluconazole-resistant fungi, although colonization of those fungi increased. When the results were combined in studies in which the incidence of systemic fungal infections was > 15%, fluconazole was effective in reducing such infections (OR, 0.23; 95% CI, 0.15-0.36). CONCLUSIONS: The current analyses failed to find an effect of fluconazole on both fatal fungal infection and systemic fungal infection in non-BMT patients. Further studies on severely neutropenic patients are warranted because prophylactic fluconazole seemed to be effective when the incidence of systemic fungal infection was expected to be > 15%.

Spontaneous remission in acute type adult T-cell leukemia/lymphoma.

Adult T-cell leukemia/lymphoma (ATL) is a neoplastic disorder of T lymphocytes associated with human T lymphotropic virus type I (HTLV-I). The prognosis of ATL is generally poor. We present here a 79-year-old woman with spontaneous remission of acute type ATL. Spontaneous remission was preceded by surgical biopsy and pneumonia and lasted for two years until she died with pancreas cancer. Monoclonal integration of HTLV-I provirus DNA became undetectable after remission.

Pyogenic granuloma of the tongue early after allogeneic bone marrow transplantation for multiple myeloma.

Oral complications occur frequently after bone marrow transplantation (BMT). Some of them are caused by regimen-related toxicity of the preparative regimen, and others by infections. In addition, oral tissues are targets of graft-versus-host disease (GVHD). Oral granulomatous lesions are not a common complication after BMT, and are especially rare on the tongue. Such rare lesions reported in the literature, developed late after BMT with oral chronic GVHD. We present here a patient who developed pyogenic granuloma of the tongue early after allogeneic BMT done for multiple myeloma. Regimen-related mucositis, oral acute GVHD, the administration of cyclosporine A, and the preexisting macroglossia might be responsible for the formation of granuloma.

Prognostic significance of serum soluble interleukin-2 receptor level in non-Hodgkin's lymphoma: a single center study in Japan.

Interleukin 2 receptor is expressed not only on the surface of activated T or B lymphocytes, but also on certain lymphoid malignancies. The receptor is released from the cell membrane as soluble form (sIL-2R). Serum sIL-2R level is a sensitive and quantitative marker of circulating peripheral blood mononuclear cell activation or specific tumor cell growth including non-Hodgkin's lymphoma (NHL). However, the relevance of serum sIL-2R levels relating to clinical outcome in adult patients with NHL remains uncertain. Therefore, we investigated the serial serum sIL-2R levels in 28 untreated patients with NHL to evaluate its correlation with clinical characteristics. High serum sIL-2R level (>1000 U/ml) at diagnosis was associated with a high incidence of treatment failure (p=0.03) and poor overall survival (p=0.057). The serum sIL-2R levels decreased significantly after achieving complete remission (p=0.003). Further larger studies are required to evaluate whether serum sIL-2R level is an independent prognostic factor or not. However, adding this parameter to those already employed in the International Prognostic Index would perhaps provide a better prognostic index for adult patients with NHL.

Control of cell death by the smart polymeric vehicle.

We report a novel drug delivery system for apoptosis induction by a "smart" polymer vehicle possessing thermosensitivity and bioaffinity. The polymer chain was prepared by copolymerization of N-isopropylacrylamide and N-methacryloyloxysuccinimide. Cell-adhesive RGDS peptide was conjugated with the copolymer as a ligand model for bioaffinity. When the temperature was increased, nanoscale aggregates precipitated from a copolymer aqueous solution. Either dolichyl phosphate (dol-p), which is an apoptotic inducer, or dolichol was added to aggregates at around the precipitation temperature (31 degrees C), and the temperature was raised to 37 degrees C for incorporation. Aggregates incorporating dol-p or dolicol were added to a human promonocytic leukemia U937 cell suspension at 37 degrees C. When the temperature was lowered to 25 degrees C, cells underwent apoptosis in the presence of Ca2+. Probably, copolymer vehicles were concentrated on a cell surface through the binding of RGDS and integrin and the release of lipid inducers was caused by the disruption of vehicles in response to temperature.

Establishment of an in vivo human myeloid leukemia model in the SCID mouse.

We succeeded in establishing a human myelogenous leukemia model in severe combined immunodeficient (SCID) mice by transplanting 2 x 10(7) ML-2 cells intraperitoneally (i.p.) with cyclophosphamide (CTX) pretreatment. Two months after transplantation, 9 of 10 mice developed leukemia and leukemia cells were detected in the peripheral blood (PB) and bone marrow (BM). The main findings at autopsy were peritoneal and pleural effusions and large tumor masses involving the peritoneal organs. However, successful transplantation required injection of a large number of cells. We therefore established a new cell line, ML-2S, from the PB of a mouse with ML-2 leukemia. Although only 2 x 10(6) ML-2S cells were inoculated, ML-2S induced the same pattern of leukemic dissemination reminiscent of the parent ML-2 cells. Compared to ML-2, progression of ML-2S was slow, suggesting that ML-2S is suitable as a leukemia model to study treatment. Furthermore, we confirmed that ML-2S cells are of human origin using isoenzyme analysis and also that ML-2S and ML-2 cells have the same phenotypic character by cell surface marker analysis.

In vivo therapeutic effects of IFN-gamma on human myelogenous leukemia in a severe combined immunodeficiency mouse model.

BACKGROUND: Results of clinical trials of IFN-gamma on the treatment of various types of leukemia are not so promising, regardless of the antiproliferative activity against leukemic cells and expected immunomodulatory effects. In this study, we have re-evaluated the anti-leukemic effects of natural human IFN-gamma (nHuIFN-gamma) using an established human myelogenous leukemia model in SCID mice. MATERIALS AND METHODS: SCID mice transplanted with human myelogenous leukemia cell line ML-2S received subcutaneously 5 x 10(4) IU/mouse of nHuIFN-gamma at 5 times/week for 5 weeks. RESULT: nHuIFN-gamma significantly prolonged the lifespan of SCID mice in leukemic crisis. Percentages of ML-2S cells in the peripheral blood were also significantly decreased by the IFN-gamma treatment. Histopathological examination revealed that IFN-gamma treatment suppressed the replacement of pancreatic cells by tumor cells and the formation of tumor masses in the intestine. CONCLUSIONS: These results suggest that IFN-gamma is effective against human myeloid leukemia, especially extramedullary tumor mass-forming type in the peritoneal organs. Our results further suggest that studies employing SCID mice leukemia model would help in devising appropriate therapeutic strategies of IFN-gamma based on the specific characteristics of each leukemia subtype.

Blood coagulation activity during microsurgery.

The authors investigated blood coagulation activity in patients who underwent microsurgery. Hemostatic parameters were measured in 9 patients (10 operations) who were undergoing free tissue transfers. These parameters included prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinopeptide A (FPA), prothrombin fragment 1 + 2 (F1 + 2), and thrombin-antithrombin complex (TAT). The flap totally necrosed owing to vasospasm in 1 patient with osteomyelitis of the heel, and the FPA, F1 + 2, and TAT values significantly increased. Reexploration was required because of flap cyanosis in 1 patient with a hemangioma on the wrist, and the F1 + 2 and TAT values increased during the salvage procedure. These molecular markers could be important in indicating hypercoagulable state sensitivity, and they serve as a warning of possible vascular compromise to a surgeon.

Innovation in nursing education: development of computer-assisted thinking.

In order to enhance students' active thinking, faculty members at International University of Health and Welfare developed the CAT (Computer Assisted Thinking) program. The CAT program is different from CAI (Computer Assisted Instruction), which mainly asks users to choose correct answers. Instead, the CAT program asks users to type in short sentences. There are two functions in the CAT program: one is to keep the students' action log each time they use the program and the other is to serve as medical dictionary. An analysis of the action log revealed that the students demonstrated little skill in inferential thinking. Their observations were very concrete. In order to help the students to develop their abstract thinking skills, we need to review our curriculum.

[Studies on penetration of cefepime into respiratory tract using broncho-alveolar lavage and sputum].

We investigated broncho-alveolar distribution of cefepime (CFPM), a fourth generation cephem, using 38 BALF specimens from 19 serious pneumonia patients who underwent artificial respiratory system control. The mean broncho-alveolar CFPM level was 3.44 microgram/ml (5.34% of the mean peak blood level). We thus observed that the BALF level after a single dose of 1 g of CFPM exceeds the MIC90 of the drug against RTI causing bacteria such as Streptococcus pneumoniae, Klebsiella pneumoniae, Haemophillus influenzae, Branhamella catarrhalis, coagulase-negative Staphylococcus, Pseudomonas aeruginosa and Staphylococcus aureus.