Bioactivation of Bovine Bone Matrix and Collagen Scaffold Using Argon Plasma: In Vitro Study.

PURPOSE: Bone graft materials and soft tissue allografts are widely used in clinical practice to counteract physiologic postextraction site tridimensional shrinkage. The aim of this study was to test if plasma of argon treatment could have a bioactivation effect on hard and soft tissue scaffolds in clinical usage. MATERIALS AND METHODS: Forty-eight bovine bone matrix and porcine collagen samples were subdivided into two groups (test and control) of 12 samples each. The test group was treated with argon plasma (10 W, 1 bar for 12 minutes), while the control group was left untreated. Immediate cell adhesion and a proliferation assay at 72 hours were performed in the perfusion chamber of a bioreactor. Additionally, micro-CT analysis was performed on the treated and untreated scaffolds, before and after soaking in cell culture medium (four samples). RESULTS: Osteoblasts seeded on plasma-treated bone matrix significantly increased the adhesion level compared with the untreated sample (43,144.3 ± 12,442.9 vs 21,736 ± 77,27.1; P = .0083). However, 3-day proliferation tests could not achieve significant differences between groups (105,715.5 ± 21,751.5 vs 107,108.6 ± 19,343.4; P = .998). No differences were measured on fibroblast adhesion on the collagen matrix in both conditions. Plasma of argon treatment and soaking in cell culture medium did not affect the bone matrix samples. The structure of collagen matrix samples was unaltered after plasma treatment, but became enlarged after soaking. CONCLUSION: Plasma of argon may be useful to biofunctionalize bone grafts, although benefits seemed to disappear after 3 days. No biologic response was detected on collagen matrix scaffolds. In vivo studies are needed to draw final clinical conclusions.

New bone ingrowth into ß-TCP/HA graft activated with argon plasma: a histomorphometric study on sinus lifting in rabbits.

BACKGROUND: In a previous experimental study, new bone was found growing within granules of HA/ß-TCP. In vitro and experimental studies have shown increased protein adsorption and cell adhesion graft material bioactivated with Argon plasma. The aims of the present experiment were to study new bone ingrowth into ß-TCP/HA granules used as filler material for sinus lifting and the influence on the healing of the bioactivation of the graft with argon plasma. METHODS: Sinus lifting was carried out in 20 rabbits using 60% HA and 40% ß-TCP as filler material either bio-activated with argon plasma (plasma group) or left untreated (control group). The antrostomies were closed with collagen membranes. Biopsies representing the healing after 2 and 10 weeks were collected, and ground sections were prepared for histomorphometric analyses. Various regions of the elevated space were analyzed both around (outer bone; OB) and inside (interpenetrating bone network, IBN) the graft particles. RESULTS: After 2 weeks of healing, 8.2% and 9.3% (n = 10; p = 0.635) of total new bone (OB + IBN) was found in the plasma and control groups, respectively. Small fractions of IBN were found, spreading from the periphery inward of the graft particles. After 10 weeks of healing, the total new bone was 34.0% in the plasma and 31.3% in Control groups (n = 9; p = 0.594). The respective fractions of IBN were 18.0% and 16.0%. New bone was penetrating from the peripheral regions inside the remnants of graft particles, where it was forming a network of bridges in continuity to the remnants of biomaterial through its porosities. The biomaterial decreased in proportion between 2 and 10 weeks from 52.1 to 28.3% in the plasma group, and from 52.5% to 31.9% in the control group. CONCLUSION: The bio-activation with argon plasma on a synthetic graft composed of 60% HA and 40% ß-TCP used as filler material for sinus lifting showed a tendency to improve bone formation; however, the difference with the control group was neither statistically significant nor clinically relevant.

Influence of the position of the antrostomy in sinus floor elevation on the healing of mini-implants: a randomized clinical trial.

AIM: To evaluate histologically the healing of mini-implants installed after sinus floor elevation using a lateral approach and placing the antrostomy at different level from the sinus floor. MATERIAL AND METHODS: Sinus floor elevation using a lateral approach was performed in 24 healthy volunteers. The antrostomy was randomly placed either close to the base of the sinus floor (group base) or at about 3-4 mm cranially to it (group standard). After 6 months of healing, mini-implants were installed within the grafted region, through the alveolar crest. Three months later, biopsies were collected. RESULTS: Sixteen biopsies from 16 patients were available for histological analyses. The new bone reached fractions of 40.9 ± 11.9% and 48.5 ± 20.1% at the base and standard groups, respectively (p = 0.208). Xenograft particles were found in contact with the implant surface at percentages of 12.1 ± 11.0% in the base group, and 15.9 ± 23.7% in the standard group (p = 0.674). CONCLUSIONS: Based on the present study, the choice of one or the other position of antrostomy did not influence significantly the outcome and, therefore, should be left to the preference of the surgeon.

Sinus Floor Elevation and Antrostomy Healing: A Histomorphometric Clinical Study in Humans.

OBJECTIVES: To compare the histomorphometric outcomes of biopsies collected from the antrostomy and from the alveolar crest of the maxillary sinus after a sinus-lift procedure. MATERIAL AND METHODS: In 12 volunteers, sinus floor elevation was performed using collagenated corticocancellous porcine bone. Nine months after the surgery, 2 biopsies, 1 from the alveolar crest and 1 from the antrostomy, were collected for histological analysis. RESULTS: Biopsies from 11 patients were available for histological analyses (n = 11). At the alveolar crest sites, the percentage of mineralized bone was 40.1 ± 14.1%, of bone marrow was 40.1 ± 18.0%, and of the xenograft was 14.7 ± 15.2%. Small amounts of soft tissue were found. At the antrostomy sites, the percentages of mineralized bone, bone marrow, and xenograft were 26.0 ± 10.8%, 23.4 ± 17.0%, and 28.2 ± 15.7%, respectively. Soft tissue was represented by 19.7 ± 19.4%. CONCLUSION: Higher amounts of mineralized bone and bone marrow were found in the alveolar crest compared with the antrostomy.

The influence of bone-graft bio-functionalization with plasma of argon on bacterial contamination.

Plasma of argon was demonstrated to improve protein and cell adhesion on bone grafts. On the other hand, increased surface energy and hydrophilicity could potentially amplify the risks of graft surface contamination in a clinical environment. The aim of the present study was to in vitro verify if the plasma of argon treatment could alter the graft characteristics affecting its ability to remain sterile. Six graft materials produced by different company were selected for this study, and randomly split by allocation either in the test (Plasma of argon treatment for 20') or the control group (only removed from the plastic sterile vials). To replicate the surgical work flow, both test and control samples were left 2 min in the clinical environment simulated conditions. Samples were therefore transferred in a Biosafety level 2 culture room. Bacterial growth analysis was performed. Optical density at 600 nm was measured as readout of bacterial growth and, after 24 hours, colony forming unit (CFU) was evaluated. Statistical analysis was performed by using the ordinary one-way ANOVA. The optical density confirmed no significant differences within groups and the number of CFU/ml for each measured sample (test and control) failed to present significant differences. Data from the present study highlighted that surface activation using plasma of argon did not affect the degree of contamination of the bone grafts, allowing to maintain a required sterility of the surface. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 67-70, 2019.

Closed reduction of a posterior sternoclavicular joint dislocation: A case report.

Sternoclavicular joint dislocation (SCJD) is a rare injury; there are only two reported cases of SCJD that have occurred during judo practice. We present a case of an 18-year-old male athlete who fell while practicing judo and experienced upper left chest pain. He was diagnosed with posterior SCJD at another institute before being transferred to our hospital. Closed reduction was initially not possible using traditional methods. Reduction was eventually accomplished by clamping the proximal end of the clavicle using bone forceps and rotating it while pulling it upward. Many authors have reported that closed reduction is difficult if not performed within 48 h after SCJD injury. However, we were able to achieve closed reduction approximately 72 h after injury. We found that reduction might be easily accomplished by pulling the proximal end of the clavicle up and rotating it when other closed reduction methods are unsuccessful.

A phase I pharmacokinetic and pharmacodynamic study of s-3304, a novel matrix metalloproteinase inhibitor, in patients with advanced and refractory solid tumors.

PURPOSE: Matrix metalloproteinases (MMP) play a fundamental role in cancer development and progression. S-3304 is a potent, orally active, noncytotoxic inhibitor of MMPs, primarily MMP-2 and MMP-9, that prolongs survival in mice xenografts and is well tolerated in healthy volunteers. EXPERIMENTAL DESIGN: The aims of this phase I clinical trial were to determine the maximum tolerated dose, dose-limiting toxicities, pharmacokinetic profile, and intratumoral MMP inhibitory activity of single-agent S-3304 in advanced and refractory solid tumors. MMP activity was determined by film in situ zymography (FIZ). Patients had tumor biopsies before and after S-3304 administration and were also evaluated for response and survival. RESULTS: Four dose levels were explored [DL1-DL4 or 800, 1,600, 2,400, and 3200 mg twice daily (BID), respectively], and 32 patients were enrolled. Toxicities were mostly gastrointestinal. The maximum tolerated dose was not reached, but dose escalations beyond DL4 were impractical (number of capsules needed). S-3304 steady-state concentrations were reached by day 8, and day 1 mean C(max) and AUC(0-8) increases were less than dose proportional. After S-3304 administration, 17 of 18 patients experienced inhibition of MMP activity by FIZ. Strong mean inhibition of MMP activity was observed in DL1 to DL3. The negative mean inhibitory activity calculated for DL4 was due to one patient with a 397% MMP activity increase. CONCLUSION: S-3304 is safe, well tolerated, and achieves plasma concentrations above those required to inhibit MMP-2 and MMP-9. Its intratumoral MMP inhibitory activity has been shown using FIZ, which is useful as a biomarker with this and other MMP inhibitors.

Pharmacokinetics and safety assessments of high-dose and 4-week treatment with S-3304, a novel matrix metalloproteinase inhibitor, in healthy volunteers.

AIMS: To investigate the tolerability, safety and pharmacokinetics of S-3304 in healthy volunteers treated with high doses of S-3304 for 28 days. METHODS: Thirty-two healthy volunteers were recruited. Four male and four female subjects were allocated to one of four doses (800 mg, 1600 mg, 2400 mg and 3200 mg). At each dose six volunteers took active medication and two volunteers took placebo in a double-blind fashion. Volunteers took a single dose on days 1 and 28 for pharmacokinetic purposes, and took twice daily doses from day 3-27. The pharmacokinetics of S-3304 and its hydroxy metabolites were evaluated. Tolerance was based on subjective adverse events, clinical examination, vital signs, ECG and laboratory tests including haematology and biochemistry profiles using CTC grading. RESULTS: Doses up to 2400 mg twice daily were generally well tolerated. At 3200 mg twice daily, five volunteers including one randomized to placebo were withdrawn from treatment mainly due to alanine aminotransferase (ALT) elevation. C(max) of S-3304 on day 1, whose geometric mean and 95% confidence interval were 66.3 microg ml(-1) (48.8, 90.0) for 800 mg, 82.6 microg ml(-1) (69.3, 98.6) for 1600 mg, 89.5 microg ml(-1) (79.5, 100.7) for 2400 mg, and 110.5 microg ml(-1) (88.9, 137.7) for 3200 mg, respectively, was correlated with the log-transformed peak ALT (P < 0.0001 for male and P = 0.048 for female volunteers). CONCLUSIONS: In healthy volunteers the maximum tolerated dose of S-3304 was 2400 mg twice daily. ALT elevation was the most frequent dose-limiting factor and was correlated with C(max) on day 1.

Effect of a selective inhibitor of secretory phospholipase A2, S-5920/LY315920Na, on experimental acute pancreatitis in rats.

We investigated the efficacy of a potent inhibitor of secretory phospholipase A2 (sPLA2), S-5920/LY315920Na, in an experimental model of acute pancreatitis in rats. Combined intraductal injection of sodium taurocholate (5 mg/rat) and porcine pancreatic sPLA2-IB (300 microg/rat) caused severe hemorrhagic necrotizing pancreatitis resulting in high mortality, along with rapid increases of catalytic PLA2 and lipase activities in plasma and ascites and with gradual increases of plasma amylase and aspartate aminotransferase levels over 9 h after the pancreatitis. Prophylactic intravenous treatment with S-5920/LY315920Na significantly reduced mortality at 7 days, and strongly abrogated PLA2 activities in both plasma and ascites along with significant reduction of lipase activity, amylase, aspartate aminotransferase, and hemorrhage at 6 h. It also significantly reduced histological damage such as edema and parenchymal and fat necroses of the pancreatic tissue. This sPLA2 inhibitor could become an effective agent for the treatment of severe acute pancreatitis.

Human group IIA secretory phospholipase A2 induces neuronal cell death via apoptosis.

Expression of group IIA secretory phospholipase A2 (sPLA2-IIA) is documented in the cerebral cortex (CTX) after ischemia, suggesting that sPLA2-IIA is associated with neurodegeneration. However, how sPLA2-IIA is involved in the neurodegeneration remains obscure. To clarify the pathologic role of sPLA2-IIA, we examined its neurotoxicity in rats that had the middle cerebral artery occluded and in primary cultures of cortical neurons. After occlusion, sPLA2 activity was increased in the CTX. An sPLA2 inhibitor, indoxam, significantly ameliorated not only the elevated activity of the sPLA2 but also the neurodegeneration in the CTX. The neuroprotective effect of indoxam was observed even when it was administered after occlusion. In primary cultures, sPLA2-IIA caused marked neuronal cell death. Morphologic and ultrastructural characteristics of neuronal cell death by sPLA2-IIA were apoptotic, as evidenced by condensed chromatin and fragmented DNA. Before apoptosis, sPLA2-IIA liberated arachidonic acid (AA) and generated prostaglandin D2 (PGD2), an AA metabolite, from neurons. Indoxam significantly suppressed not only AA release, but also PGD2 generation. Indoxam prevented neurons from sPLA2-IIA-induced neuronal cell death. The neuroprotective effect of indoxam was observed even when it was administered after sPLA2-IIA treatment. Furthermore, a cyclooxygenase-2 inhibitor significantly prevented neurons from sPLA2-IIA-induced PGD2 generation and neuronal cell death. In conclusion, sPLA2-IIA induces neuronal cell death via apoptosis, which might be associated with AA metabolites, especially PGD2. Furthermore, sPLA2 contributes to neurodegeneration in the ischemic brain, highlighting the therapeutic potential of sPLA2-IIA inhibitors for stroke.