In situ forming oxidised hyaluronic acid/adipic acid dihydrazide hydrogel for prevention of epidural fibrosis after laminectomy.

Post-operative epidural fibrosis is a biological response after laminectomy that may lead to clinical symptoms, such as radicular pain. An ideal material for prevention of epidural fibrosis should be able to inhibit fibroblast adhesions and reduce formation of scar tissue. An injectable hydrogel would be the material of choice for this purpose, since it could fill an irregular surgical defect completely, gelate in situ and be delivered in a minimally-invasive manner. The objective of this study was to evaluate, in vitro and in vivo, the cytocompatibility and anti-adhesive effect of an oxidised hyaluronic acid/adipic acid dihydrazide (oxi-HA/ADH) hydrogel. Different cell types present in the spine were used to test the cytocompatibility of the hydrogel. The hydrogel extraction medium had no deleterious effects on neural cells (PC-12), but reduced fibroblasts viability (NIH/3T3). Although the hydrogel did not change the release of lactate dehydrogenase from myoblasts (C2C12) and Schwann cells (RSC96), the extraction medium concentration slightly affected the mitochondrial activity of these two cell types. qPCR showed that the hydrogel down-regulated S100a and P4hb expression in NIH/3T3 cells, supporting the hypothesis that the hydrogel might inhibit fibroblast activity. The animal study showed a reduction of scar tissue formation as well as severity of adhesion between scar tissue and the dura mater in a rat laminectomy model. Superficially, the peel-off test showed significantly decreased tenacity. In conclusion, the oxi-HA/ADH hydrogel is a promising injectable and thermosensitive material for prevention of post-operative epidural fibrosis.

Gonorrhea infection increases the risk of prostate cancer in Asian population: a nationwide population-based cohort study.

This nationwide population-based retrospective cohort study evaluated the risk of developing prostate cancer among patients with gonorrhea. We identified cases of newly diagnosed gonorrhea in men between 2000 and 2010 from the Taiwan National Health Insurance Research Database. Each patient with gonorrhea was matched to four controls, based on age and index year. All subjects were followed up from the index date to December 31, 2010. The Cox proportional hazards regression model was used to assess the risk of prostate cancer. A total of 355 men were included in the study group, and 1,420 age-matched subjects without gonorrhea were included in the control group. After adjusting for age, comorbidities, urbanization level, hospital level, and monthly income, gonorrhea was significantly associated with an increased risk of prostate cancer (adjusted hazard ratio = 5.66, 95% confidence interval = 1.36-23.52). Men aged 45-70 years and those with lower monthly income were more strongly associated with prostate cancer in the study group than the control group. The higher risk for developing prostate cancer were also found in those without syphilis, without genital warts, without diabetes mellitus, without chronic obstructive pulmonary disease, without benign prostatic hypertrophy, without chronic prostatitis, and without alcoholism. The Kaplan-Meier analysis showed the risk of prostate cancer was significantly higher in the study group than in the control group. Gonorrhea may be involved in the development of prostate cancer. More intensive screening and prevention interventions for prostate cancer should be recommended in men with gonorrhea.

Evaluation of adhesion force and binding affinity of phytohemagglutinin erythroagglutinating to EGF receptor on human lung cancer cells.

PHA-E is a natural product extracted from red kidney beans, and it has been reported to induce cell apoptosis by blocking EGFR in lung cancer cells. Because EGF is the major in vivo competitor to PHA-E in clinical application, PHA-E must be proved that has better affinity to EGFR than EGF. This study would focus on how PHA-E tightly bind to EGFR and the results would compare with EGF. The adhesion force, measured by AFM, between EGFR and PHA-E was 207.14±74.42 pN that was higher than EGF (183.65±86.93 pN). The equilibrium dissociation constant of PHA-E and EGF to EGFR was 2.4 10(-9)±1.4 10(-9) and 7.3 10(-8)±2.7 10(-8), respectively, that could evaluate binding affinity. The result showed that binding affinity of PHA-E to EGFR was one order higher than EGF to EGFR. In the results of flow cytometer and confocal microscope, we found binding efficiency of EGF to EGFR was decrease as the concentration of PHA-E increased. In the analysis of Western blot, treatment of A-549 cells with PHA-E resulted in a dose-dependent decrease in EGFR phosphorylation. In conclusion, we found that PHA-E had better adhesion force and binding affinity to EGFR than that of the EGF. The interaction between PHA-E and EGFR could block EGF binding and then inhibit EGFR phosphorylation. PHA-E could be developed into a new target molecule for lung cancer treatment that could be immobilized on the drug carrier to guide therapeutic particles to the tumor site.

Inhibition of intracellular dipeptidyl peptidases 8 and 9 enhances parthenolide's anti-leukemic activity.

Parthenolide is selectively toxic to leukemia cells; however, it also activates cell protective responses that may limit its clinical application. Therefore, we sought to identify agents that synergistically enhance parthenolide's cytotoxicity. Using a high-throughput combination drug screen, we identified the anti-hyperglycemic, vildagliptin, which synergized with parthenolide to induce death of the leukemia stem cell line, TEX (combination index (CI)=0.36 and 0.16, at effective concentration (EC) 50 and 80, respectively; where CI <1 denotes statistical synergy). The combination of parthenolide and vildagliptin reduced the viability and clonogenic growth of cells from acute myeloid leukemia patients and had limited effects on the viability of normal human peripheral blood stem cells. The basis for synergy was independent of vildagliptin's primary action as an inhibitor of dipeptidyl peptidase (DPP) IV. Rather, using chemical and genetic approaches we demonstrated that the synergy was due to inhibition of the related enzymes DPP8 and DPP9. In summary, these results highlight DPP8 and DPP9 inhibition as a novel chemosensitizing strategy in leukemia cells. Moreover, these results suggest that the combination of vildagliptin and parthenolide could be useful for the treatment of leukemia.

The development of iron-free partially stabilized cement for use as dental root-end filling material.

AIM: To determine the effect of increasing the proportion of zinc on partially stabilized cement (PSC) produced using a one-step sol gel process. METHODOLOGY: A one-step sol-gel process of Portland cement-based PSC with Zn was synthesized by replacing iron nitrate. The crystalline phases of the PSC-Zn powder were analysed by using X-ray diffraction (XRD). The experimental groups [i.e., MTA, PSC-Fe (control), PSC with 1% Zn, PSC with 3% Zn, and PSC with 5% Zn] were immersed in simulated body fluid for 3 h, 1 and 3 days to evaluate the hydration product formation. The microstructure and surface morphology were analysed using scanning electron microscopy (SEM). Initial and final setting times of the materials were determined using an ASTM Vicat needle testing machine. To evaluate the cytotoxicity of PSC-Zn system, primary osteoblasts cell lines were used. RESULTS: The addition of increased weight percentages of Zn, resulted in a more unstable phase which favoured the formation of a monoclinic structure of C3 S with an increased hydration reaction of PSC and reduced setting time. The cytotoxicity testing of PSC with Zn revealed that the material was not toxic. CONCLUSIONS: The newly synthesized PSC-Zn material had short setting time and was biocompatible.

Intramedullary cavity as implantation site for bioartifical pancreas: preliminary in vivo study.

BACKGROUND: The intramedullary cavity is a widely distributed well-vascularized microenvironment capable of sustaining grafts, and is a potential site for islet transplantation. The bone marrow offers sufficient space that may also be suitable for bioartificial pancreas (BAP) implantation. OBJECTIVE: To evaluate the feasibility of bone marrow as an implantation site for BAPs. MATERIALS AND METHODS: A calcium phosphate cement chamber satisfies the criteria for immunoisolation. Mouse insulinoma cells were suspended with agarose gel and enclosed in a calcium phosphate cement chamber to create a BAP, which was implanted in the intramuscular space in diabetic swine or the intramedullary cavity in diabetic dogs. Blood glucose and C-peptide concentrations were determined perioperatively. RESULTS: In the swine, the mean ± SD blood glucose concentration decreased from 413 ± 24 mg/dL to 285 ± 47 mg/dL, and was maintained in the range of 285 to 336 mg/dL for 15 days. It increased to 368 to 450 mg/dL after the BAPs were implanted in the intramuscular space. In the dogs, the blood glucose concentration decreased from 422 ± 32 mg/dL to 247 ± 52 mg/dL, and was maintained in the range of 247 to 347 mg/dL after the BAPs were implanted in the intramedullary cavity. The C-peptide concentration increased from 6.1 ± 2.8 pmol/L to 104.7 ± 16.4 pmol/L when the BAPs were implanted in the intramedullary cavity. CONCLUSION: This study indicates superior effectiveness of BAPs implanted in the intramedullary cavity compared with the intramuscular space. This observation may be attributed to the greater oxygen tension in the bone marrow. The BAPs in direct contact with the circulatory system receive sufficient blood flow for function and survival. This preliminary study demonstrates that the intramedullary cavity may be an implantation site for BAP transplantation.

Chitosan/gelatin hydrogel prolonged the function of insulinoma/agarose microspheres in vivo during xenogenic transplantation.

PURPOSE: A chitosan/gelatin solution with glycerol 2-phosphate disodium salt hydrate in liquid phase at room temperature becomes a hydrogel at 37 degrees C. The material can be used as an injectable cell carrier into the human body for gelation in situ. We hoped that the chitosan/gelatin hydrogel provided extra protection for insulinoma/agarose microspheres during xenogenic transplantation. MATERIALS AND METHODS: Mouse insulinoma was microencapsulated in agarose as microspheres, which were macroencapsulated in chitosan/gelatin hydrogel. Insulin secreting profiles were first demonstrated in vitro. Diabetic rats were injected subcutaneously with insulinoma/agarose microspheres or insulinoma/agarose microspheres suspended in chitosan/gelatin solution. The nonfasting blood glucose concentrations (NFBG) of diabetic rats were measured perioperatively. Rats were humanely killed 1 month postoperatively and the hydrogel was retrieved for histological examination. RESULTS: The insulinoma/agarose microspheres continually secreted insulin for 1 month when macroencapsulated in chitosan/gelatin hydrogel in vitro. The NFBG of diabetic rats injected with insulinoma/agarose microspheres decreased to euglycemic status albeit hyperglycemia was restored within 10 days. The NFBG of diabetic rats injected with chitosan/gelatin hydrogel, which contained insulinoma/agarose microspheres, was maintained at less than 200 mg/dL for 25 days. The histological section revealed immune cell infiltration and accumulation within the hydrogel and around the iusulinoma/agarose microspheres that may have contributed to the slowly increasing NFBG after day 25. CONCLUSION: This study showed that chitosan/gelatin hydrogel can be used as a cell carrier for an injectable bioartificial pancreas; the hydrogel prolonged the function of cells encapsulated in agarose microspheres during xenogenic transplantation.

Preparation and surface characterization of HMDI-activated 316L stainless steel for coronary artery stents.

Poor compatibility between blood and metallic coronary artery stents is one reason for arterial restenosis. Immobilization of anticoagulant agents on the stent's surface is feasible for improving compatibility. We examined possible surface-coupling agents for anticoagulant agent immobilization. Hexamethylene diisocyanate (HMDI) and 3-aminopropyl-triethoxysilane (APTS) were examined as surface-coupling agents to activate 316L stainless steel (e.g., stent material). The activated surface was characterized using Fourier transformation infrared spectroscopy (FTIR), atomic force microscope (AFM), surface plasmon resonance (SPR), and trinitrobenzene sulfonic acid (TNBS) assay. In FTIR analysis, HMDI and APTS were both covalently linked to 316L stainless steel. In AFM analysis, it was found that the HMDI-activated surface was smoother than the APTS-activated one. In SPR test, the shift of SPR angle for the APTS-activated surface was much higher than that for the HMDI-activated surface after being challenged with acidic solution. TNBS assay was used to determine the amount of immobilized primary amine groups. The HMDI-activated surface was found to consist of about 1.32 micromol/cm(2) amine group, whereas the APTS-activated surface consisted of only 0.89 micromol/cm(2) amine group. We conclude that the HMDI-activated surface has more desirable surface characteristics than the APTS-activated surface has, such as chemical stability and the amount of active amine groups.

A study of purified montmorillonite intercalated with 5-fluorouracil as drug carrier.

Since its introduction over 40 years ago, 5-fluorouracil (5-FU) has remained the only effective chemotherapy option available for the treatment of colorectal cancer (CRC). However, this cytotoxic anticancer drug often causes severe side effects because it does not act selectively on the tumor. It has been reported that the 5-FU showed considerable toxicity when administered by intravenous injections or via alimentary tract. Although, many materials have been developed for carrying 5-FU, there has been no clinically acceptable carrier for 5-FU till now. Montmorillonite, one of the clay minerals, consists of hydrated aluminum silicates with fine grains and large spaces between the layers. Isomorphous substitution of cations is common. In the study, we attempt to intercalate 5-FU into interlayers of montmorillonite through ion exchange. Montmorillonite was purified from crude clays of bentonite in Tai-dong, Taiwan by filtration and sedimentation. Solutions of 5-FU with different concentrations were prepared by dissolving various amounts of 5-FU into 10 ml NaOH solution. Purified montmorillonite powder was soaked in 5-FU solution for a period of time with different pH values and temperatures. In this study, we try to intercalate 5-FU into interlayers of montmorillonite to find out optimum conditions, such as soaking time, temperature, pH value, initial 5-FU concentration, etc., to prepare composites of 5-FU and montmorillonite (5-FU/mont). UV, SDT, FTIR, XRD are used to characterize the 5-FU/mont composite. From the results. 5-FU was successfully intercalated into the interlayer of montmorillonite both by free surface absorption and OH replacement. The optimum condition for 5-FU/mont preparations is 1.185 wt% of 5-FU as initial concentration under a pH value of 11.6 at a temperature of 80 degrees C and a soaking time of 2 h. The total amount of 5-FU in montmorillonite is about 87.5 mg for each gram of montmorillonite, which can be proved by thermal gravimetric analysis. The composite of 5-FU/mont is expected to achieve in situ release for colorectal cancer therapy in future applications.

In vitro effects of low-intensity ultrasound stimulation on the bone cells.

Mechanical perturbations serve as extracellular signals to a variety of cells, including bone cells. Low-intensity pulsed ultrasound produces significant multifunctional effects that are directly relevant to bone formation and resorption. Ultrasound stimulation has been shown to accelerate bone-defect healing and trabecular bone regeneration. In this study, we use an in vitro bone cell culture model to investigate the effect of low-intensity pulsed ultrasound. The rat alveolar mononuclear cell-calvaria osteoblast coculture system was used in this study. Before treatment, the bone cells were cultured for 3 days to facilitate their attachment and differentiation. Then, ultrasound exposure (frequency = 1 MHz, intensity = 0.068 W/cm(2)) or sham exposure for 20 min per day was applied until the end of the experiment. Half of the culture media were obtained on the 4th, 5th, 6th, 7th, 8th, 9th, and 10th days for the analysis of cytokines and biochemical parameters. At the end of the experiment, cells were fixed and stained for identification and quantification of the osteoblast and osteoclast cells. After low-intensity pulse ultrasound stimulation, the osteoblast cell counts were significantly increased, whereas the osteoclast cell counts were significantly decreased. The total alkaline phosphatase amount in the culture medium was increased after 7 days of ultrasound stimulation, and tumor necrosis factor-alpha in ultrasound-stimulated bone cells was significantly increased after the 7th day of culture and reached 474.77% of the control medium on the 10th day of culture. The results of this study suggest that low-intensity ultrasound treatment may have a stimulatory effect on bone-healing processes.

Phase, compositional, and morphological changes of human dentin after Nd:YAG laser treatment.

Although techniques for repairing root fracture have been proposed, the prognosis is generally poor. If the fusion of a root fracture by laser is possible, it will offer an alternative to extraction. Our group has attempted to use lasers to fuse a low melting-point bioactive glass to fractured dentin. This report is focused on the phase, compositional, and morphological changes observed by means of X-ray diffractometer, Fourier transforming infrared spectroscopy, and scanning electron microscopy-energy dispersive X-ray spectroscopy in human dentin after exposure to Nd:YAG laser. The irradiation energies were from 150 mJ/ pulse-10 pps-4 s to 150 mJ/pulse-30 pps-4 s. After exposure to Nd:YAG laser, dentin showed four peaks on the X-ray diffractometer that corresponding to a-tricalcium phosphate (TCP) and beta-TCP at 20 = 30.78 degrees/34.21 degrees and 32.47 degrees/33.05 degrees, respectively. The peaks of a-TCP and beta-TCP gradually increased in intensity with the elevation of irradiation energy. In Fourier transforming infrared analysis, two absorption bands at 2200 cm(-1) and 2015 cm(-1) could be traced on dentin treated by Nd:YAG laser with the irradiation energies beyond 150 mJ/pulse-10 pps-4 s. The energy dispersive X-ray results showed that the calcium/phosphorus ratios of the irradiated area proportionally increased with the elevation of irradiation energy. The laser energies of 150 mJ/ pulse-30 pps-4 s and 150 mJ/pulse-20 pps-4 s could result in the a-TCP formation and collagen breakdown. However, the formation of glass-like melted substances without a-TCP at the irradiated site was induced by the energy output of 150 mJ/ pulse-10 pps-4 s. Scanning electron micrographs also revealed that the laser energy of 150 mJ/ pulse-10 pps-4 s was sufficient to prompt melting and recrystallization of dentin crystals without cracking. Therefore, we suggest that the irradiation energy of Nd:YAG laser used to fuse a low melting-point bioactive glass to dentin is 150 mJ/ pulse-10 pps-4 s.

Effect of anti-inflammatory medication on monocyte response to titanium particles.

Cytokines produced by macrophages in the periprosthetic membranes surrounding joint replacements have been implicated as causal agents in osteolysis and prosthetic loosening. The present study characterizes the response of human peripheral blood monocytes to titanium particles. Monocytes were obtained from donated blood and were cultured in the presence of different-sized titanium particles. Exposure to titanium-aluminum-vanadium particles significantly changed the release of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and interleukin-1 (IL-1), whereas there was no significant effect on the release of prostaglandin E(2) (PGE(2)). When monocytes were cultured with particles, the titanium alloy particles induced significantly more release of TNF-alpha and less IL-1 secretion. Ciprofloxacin inhibited production of TNF-alpha, IL-6, IL-1, and PGE(2) in human monocytes exposed to titanium particles. In contrast to ciprofloxacin, indomethacin was not a potent inhibitor of TNF-alpha production but potentiated IL-6 production in titanium-stimulated monocytes. Indomethacin had no effect on the production of IL-1 and was a potent inhibitor of PGE(2) production in titanium-stimulated monocytes. Pentoxifylline had an inhibitor effect on TNF-alpha production in titanium-stimulated monocytes. Pentoxifylline potentiated IL-6 and IL-1 production in monocytes exposed to titanium particles and had a biphasic effect on the PGE(2) production. The results of this study support our hypothesis that human monocytes release bone resorption mediators after in vitro exposure to TiAlV alloy particles. The results also demonstrate the differences of bone-resorbing mediators in response to different wear particle size. The pharmacologic agents (ciprofloxacin, pentoxifylline, and indomethacin) that can modulate the release of bone resorbing mediators such as PGE(2), TNF-alpha, IL-1, and IL-6 release from human monocytes. The results help to elucidate the differences in cellular response to wear particles but may not be directly transposed to the human situation.

A study of purified montmorillonite intercalated with 5-fluorouracil as drug carrier.

In the recent decade, the most useful drug for the therapy of colon cancer is 5-Fluorouracil(5-FU). It has been reported to have considerable toxicity administered by intravenous injections or via alimentary treat. Though many materials have developed for drug carrier of 5-FU, there was no clinically acceptable carrier for 5-FU till now. Montmorillonite, one of clay minerals, consists of hydrated aluminum silicates that are fined grained and usually have a large space between the layers. Isomorphous substitution of cations is common. We try to intercalate 5-FU into interlayer of montmorillonite through ion-exchange. Montmorillonite with 5-FU intercalation is expected to achieve in-situ release for colon cancer therapy. In the study, 5-FU was dissolved in 100 ml distilled water as 5-FU solution. Purified montmorillonite powder will soak in 5-FU solution for a period of time ad different pH value and temperature. The intercalated amount of 5-FU in montmorillonite is measured by scanning differential thermal (SDT) analysis and UV analysis. The results showed that 4 g purified montmorillonite soaked in 0.6% 5-FU solution for 2 hours had an optimum condition for intercalation. The total amount of 5-FU in montmorillonite is about 9.13 wt%.

The influence of hydroxyapatite particles on osteoclast cell activities.

Aseptic loosening after total joint arthroplasty is a major problem in orthopedic surgery. Small particles from material wear have been reported as the main cause of implant failure. For this reason, investigation into possible wear particles from the materials used in the implant may lead to longevity after arthroplasty. Hydroxyapatite (HA) has been extensively investigated and reported as an excellent biomaterial with excellent biocompatibility. In this study, we used an in vitro osteoblast/osteoclast model to test the biocompatibility of various-sized HA particles. Primary osteoclasts/osteoblasts were co-cultured with different-sized HA particles (0.5-3.0 microm, 37-53 microm, 177-205 microm, and 420-841 microm) for 3 h, 1 day, 3 days, and 7 days. Cellular responses to the HA particles were evaluated by changes in cell counts and the secretion of transforming growth factor (TGF-beta1), alkaline phosphatase (ALP), tumor necrosis factor (TNF-alpha), prostaglandin (PGE2), and lactate dehydrogenase (LDH) in the supernatant of the culture media. The results showed that osteoblasts/osteoclasts co-cultured with HA particles smaller than 53 microm undergo the most significant changes. Cellular counts significantly decreased, and the changes were more obvious in the osteoblast population. There also was a significant decrease in TGF-beta1 concentration and a significant increase in PGE2 and LDH concentration, but there were no changes in the TNF-alpha or ALP titer. It can be concluded that larger HA particles may be quite compatible with bone cells while smaller-sized HA particles can both activate the osteoclasts and decrease the cell population of the osteoblasts. Justification for the additional expense incurred with the use of hydroxyapatite in primary total hip arthroplasty should be further evaluated.

Cytokine and prostaglandin E2 release from leukocytes in response to metal ions derived from different prosthetic materials: an in vitro study.

Cytokines produced by leukocytes in the periprosthetic membranes surrounding joint replacements have been implicated as causal agents in osteolysis and prosthetic loosening. In this study, we used an in vitro leukocyte culture system to monitor the response of leukocytes to various metal ions and their possible roles in the mechanism of aseptic loosening. Human peripheral leukocytes were isolated and incubated with various concentrations of Co2+, Cr3+, and Ti3+ ions. Leukocyte cell counts and the levels of the tumor necrosis factor-alpha (TNF-alpha), interleukin-1 (IL-1), interleukin-6 (IL-6) and prostaglandin E2 (PGE2) released into the media were analyzed at 1 h, 3 h, and 1, 3, and 7 day intervals. The results showed that adding different metal ions into leukocyte cultures did not affect the cell counts. Exposure of leukocytes to Co2+ ion increased the release of TNF-alpha, IL-6, and PGE2. Exposure of leukocytes to Cr3+ ion did not increase the release of TNF-alpha but increased the secretion of IL-6 and PGE2. In contrast, exposure of the leukocytes to Ti3+ ions was associated with a decrease in the release of TNF-alpha and PGE2 and a minimal change in IL-6 noted after 7 days' culture. The present study elucidated the possible mechanisms involved in periprosthetic osteolysis and the inflammatory response of human leukocytes to metal ions. We found that cobalt ion is the most potent stimulant for cytokines and prostaglandin secretion by leukocytes. This elucidation, in combination with other efforts to reduce the generation of wear debris and metal ions, may improve the longevity of orthopedic implants.

Effect of hydroxyapatite particle size on myoblasts and fibroblasts.

After surgery, the bone and soft tissues around integrated biomaterials can be adversely affected by implant-related factors acting over a period of years. However, few studies have directly addressed the effects upon the adjacent soft tissue. The present study was designed to test the biological effects of various sized hydroxyapatite (HA) particles on myoblasts and fibroblasts. Both the myoblasts and fibroblasts were mixed in in vitro culture with 0.1% (1 mg ml(-1)) of various sized HA particles (0.5-3.0, 37-63, 177-250, 420-841 microm) for 1 h, 3 h, 1 day, 3 days and 7 days to test their effects on the cell culture. The results show that adding HA particles into a cell culture can decrease the cell count significantly. The transforming growth factor-beta1 (TGF-beta1) concentrations in the culture medium decreased significantly on addition of HA particles. When calculated as a ratio to the cell number, the TGF-beta1 titre increased most significantly in the groups of medium-sized particles. The prostaglandin E2 (PGE2) concentrations in the medium increased significantly. The changes in TGF-beta1 and PGE2 concentrations with the smallest particles were most significant and persisted longer. The inhibitory effects of the HA particles on the cell culture were mediated by the increased synthesis of PGE2. Caution should be exercised before considering the use of an HA product which could easily break down into a fine powder.

Effects of calcium phosphate bioceramics on skeletal muscle cells.

With advances in ceramics technology, calcium phosphate bioceramics have been applied as bone substitutes. The effects of implants on bony tissue have been investigated. The effects upon adjacent skeletal muscles have not been determined. The focus of this work is to elucidate the biological effects of various calcium phosphate bioceramics on skeletal muscles. Four different kinds of powder of calcium phosphate biomaterials including beta-tricalcium phosphate (beta-TCP), hydroxyapatite (HA), beta-dicalcium pyrophosphate (beta-DCP) and sintered beta-dicalcium pyrophosphate (SDCP), were tested by myoblast cell cultures. The results were analyzed by cell count, cell morphology and concentration of transforming growth factor beta 1 (TGF-beta 1) in culture medium. The cell population and TGF-beta 1 concentration of the control sample increased persistently as the time of culture increased. The changes in cell population and TGF-beta 1 concentration in culture medium of the beta-TCP and HA were quite low in the first 3 days of culture, then increased gradually toward the seventh day. The changes in cell population and TGF-beta 1 concentration in culture medium of the silica, beta-DCP, and SDCP were quite similar. They were lower during the first day of culture but increased and reached that of the control medium after 7 days' culture. Most cells on B-TCP and HA diminished in size with radially spread, long pseudopods. We conclude that HA and beta-TCP are thought to have an inhibitory effect on growth of the myoblasts. The HA and beta-TCP may interfere with the repair and regeneration of injured skeletal muscle after orthopedic surgery.

Prevascularized bone graft cultured in sintered porous beta-Ca2P2O7 with 5 wt% Na4P2O7.10H2O addition ceramic chamber.

Autogenous bone transfer is an important part of reconstructive plastic surgery. Presently available techniques have the disadvantages of limitation of available donor site, loss of donor tissue and the possibility of donor defect or deformity. In the present study, a vascularized bone graft was created and cultured in the groin area of the New Zealand rabbit. The cylindrical ceramic chambers, 15 mm in length, 6 mm in outer diameter and 3 mm in inner diameter, were prepared by the addition of sintered porous beta-Ca2P2O7 with 5 wt% Na4P2O7.10H2O. In the first group, the chambers impregnated with autogenous bone fragments and allogenous demineralized bone matrix with volume ratio 1:1 were cultured in the rabbit's groin area with saphenous vessels passing through. In the second group, the chambers were treated by the same procedures as the first group but without saphenous vessels passing through. In the third group, the chambers were not impregnated, and were cultured in the groin area with saphenous vessels. After 2, 4, 6, 8 and 12 wk of operation, the animals were killed with an overdose of intravenous pentobarbital. The viability of the osseous tissue in the chamber was evaluated by histological examination, microangiograms and fluorochrome incorporation for the three groups. The autogenous bone chips could survive and retain their osteogenic properties while packed into the sintered porous beta-Ca2P2O7 (with 5 wt% Na4P2O7.10H2O addition) ceramic chamber and implanted in the rabbit groin area up to 12wk. However, even at the longest time periods, considerable amounts of dead bone were present in the chambers. In addition, we observed bone resorption in the three groups up to 12 wk, which might be attributed to lack of physiological stress. There were significant differences in new bone formation and osseous cell viability among the three groups. The prevascularized vessels and autogenous bone chips were both necessary for the formation of new bone and osteogenic property in the chamber under these heterotopic circumstances. The biodegradable ceramic used in this study was gradually absorbed and dissolved in the physiological environment. However, the degradation debris of the ceramic caused no injury to the new bone formation. These findings support the concept of creating a preformed vascularized bone graft to reconstruct segmental bone defects.

GABAB receptor-mediated effects in synaptosomes of lethargic (lh/lh) mice.

Previously, we have shown a significant increase in number of GABAB receptor binding sites in neocortex and thalamus of lethargic (lh/lh) mice, a mutant strain exhibiting absence seizures. This study was performed to test our hypothesis that presynaptic GABAB receptors would inhibit [3H]GABA release to a greater degree in lh/lh mice compared with their nonepileptic littermates (designated +/+). Synaptosomes isolated from neocortex and thalamus of age-matched male lh/lh and +/+ mice were similar in uptake of [3H]GABA. In the neocortical preparation, baclofen dose-dependently inhibited [3H]GABA release evoked by 12 mM KCl, an effect mediated by GABAB receptors. The maximal inhibition (Imax) value was significantly greater (80%) in lh/lh than +/+ mice, whereas the IC50 (3 microM) was unchanged. In the thalamic preparation, the effect of baclofen (50 microM) was 58% less robust in lh/lh mice. Other effects mediated by GABAB receptors (inhibitions in Ca2+ uptake and cyclic AMP formation) were also significantly reduced in thalamic synaptosomes from lh/lh mice. These data suggest a greater presynaptic GABAB receptor-mediated effect in neocortex and a reduced effect in thalamic nuclei of lh/lh mice. It is possible that selective effects of presynaptic GABAB receptors or GABA release in neocortex and thalamic nuclei of lh/lh mice may contribute to mechanisms underlying absence seizures.

Sintered porous DP-bioactive glass and hydroxyapatite as bone substitute.

There is extensive experimental and surgical experience with the use of bone tissue to fill defects in the skeleton, to bridge non-union sites, and to pack defects in bone created from cyst curettage. DP-bioactive glass with a chemical composition of Na2O 8.4%, SiO2 39.6%, P2O5 12% and CaO 40% has been reported as an alternative bone substitute of high mechanical strength, good biocompatibility. and which has a tight bond with living tissue. The bonding layer between DP-bioactive glass and bone tissue was considered to be formed by dissolution of calcium and phosphate ions from the DP-bioactive glass into the surrounding body fluids. The biological hydroxyapatite was suspected to deposit directly onto the bonding layer. In order to confirm the interaction between the DP-bioactive glass and bone tissue, the developed bioactive glass was implanted into rabbit femur condyle for 2-32 weeks. The histological evaluation of DP-bioactive glass as a bone substitute was also investigated in the study. Porous hydroxyapatite bioceramic was used in the control group and the results were compared with those of DP-bioactive glass. The interface between the DP-bioactive glass and bone tissue examined with SEM-EPMA showed that the bioactive glass formed a reaction layer on the surface within 2 weeks after operation and formed a direct bond with natural bone. The elements contained in the bioactive glass apparently interdiffuse with the living bone and biological hydroxyapatite deposited onto the diffusion area, which was proved by EPMA and TEM. After implantation for over 8 weeks, the DP-bioactive glass was gradually biodegraded and absorbed by the living bone. Histological examination using the optical microscope showed that osteocytes grow into the inside of the DP-bioactive glass and the bioactive glass would be expected to be a part of bone.