Novel endodontic sealers induce cell cytotoxicity and apoptosis in a dose-dependent behavior and favorable response in mice subcutaneous tissue.

OBJECTIVES: The objective of the present study is to evaluate the in vitro cytotoxicity and in vivo biocompatibility of two novel endodontic sealers: RealSeal XT1 and Sealapex Xpress on the subcutaneous connective tissue of mice. MATERIALS AND METHODS: The cytotoxicity was assessed by cell viability using the MTT assay (one-way ANOVA), trypan blue test (Mann-Whitney) and cell apoptosis by flow cytometer. For the subcutaneous study, polyethylene tubes filled with the sealers were implanted in 70 BALB/c mice: 6 experimental groups (n = 10/group) and 2 control groups with empty tubes (n = 5/group). At the end of experimental periods (7, 21, and 63 days), the tissue was removed and histotechnically processed. Angioblastic proliferation and edema (Fisher's exact test) were evaluated, besides thickness measurement (µm) of the reactionary granulomatous tissue and neutrophil counts (Kruskal-Wallis and Dunn's post test; Mann-Whitney) (α = 0.05). RESULTS: MTT assay, trypan blue, and analysis of apoptotic cells showed a dose-dependent direct effect: the more diluted the sealer, the less cytotoxic. Regarding the angioblastic proliferation and edema, difference between the sealers at 7 and 63 days occurred (p < 0.05). Both endodontic sealers initially promoted perimaterial tissue reaction as a foreign body granuloma and thus stimulated favorable tissue responses. CONCLUSIONS: Both sealers showed a dose-dependent effect and promoted satisfactory subcutaneous tissue response; the sealer Sealapex Xpress was less cytotoxic and more biocompatible than RealSeal XT. CLINICAL RELEVANCE: The step of root canal filling during endodontic treatment is highly important for the preservation of the periapical tissue integrity. Subcutaneous reaction to endodontic sealers enables scientific basis for clinical use.

The use of lumbar spinal stabilization techniques during the performance of abdominal strengthening exercise variations.

AIM: The lumbar spinal stabilization techniques in abdominal hollowing and bracing are known to facilitate the level of activity in the muscles transversus abdominis and obliquus internus (TA/OI). The relative timing of activation and the effect of combination with other tasks are currently unknown. The objective of this study was to determine whether the performance of abdominal hollowing and bracing could promote the voluntary preferential recruitment of TA/OI muscles when performed prior to abdominal strengthening exercise variations. METHODS: The trunk muscles TA/OI, rectus abdominis and obliquus externus were investigated using surface EMG. A pressure transducer under the lumbar spine detected spinal movement. Eighteen male subjects performed a series of 4 abdominal strengthening exercise variations. Pressure cuff and electromyographic onset times were collected and analyzed. RESULTS: TA/OI muscle site was recruited significantly earlier than the upper rectus abdominis site for the hollow with curl exercise. The TA/OI site was also ranked as first activated site in the majority of subjects during exercises where stabilization techniques of hollowing and bracing were used. CONCLUSIONS: Abdominal hollowing was an effective method for selective preferential voluntary recruitment of TA/OI site prior to the performance of the curl type abdominal strengthening exercise variation. The other variations brace with curl and hold and hollow with rotation also tend to recruit TA/OI site first, however the timing was not significantly different.

Long-term survival in a rodent model of disseminated brain tumors by combined intrathecal delivery of herpes vectors and ganciclovir treatment.

Brain tumors that have disseminated into cerebrospinal fluid (CSF) pathways are an unresolved therapeutic problem, especially in pediatric neurooncology. Here a gene therapy approach using the herpes simplex virus type 1 thymidine kinase (HSV-TK)/ganciclovir (GCV) paradigm was tested using an HSV vector in a rodent model of disseminated central nervous system tumors. 9L-gliosarcoma cells were implanted simultaneously into the brain and the CSF of syngeneic rats. Five days later, resulting intracerebral and leptomeningeal tumors were treated by intrathecal injection of a replication-conditional HSV vector. This vector was defective for the ribonucleotide reductase gene, but contained an intact HSV-tk gene. Systemic GCV treatment was started 2 days after vector application and continued for 14 days. Tumor-free, long-term survival (LTS) was achieved in 90% of the animals treated with this combined therapeutic approach, whereas only 30% LTS was found in animals that had received the vector alone and 10% LTS in untreated animals. This therapeutic response probably involves oncolytic, on-site replication of the vector, activation of GCV by a HSV-TK, and a strong immune response both to the vector and to 9L cells. Apparent vector-related mortality was observed in 20% of animals without subsequent GCV therapy, but no vector-related mortality was found when the animals were treated with GCV after vector application. Given the successful outcome of this experimental treatment and the apparent potential of GCV to control HSV-related toxicity, intrathecal application of HSV vectors combined with GCV treatment may be a promising approach for treatment of disseminated brain tumors.

Intraarterial delivery of adenovirus vectors and liposome-DNA complexes to experimental brain neoplasms.

This study investigated the intraarterial delivery of genetically engineered replication-deficient adenovirus vectors (AVs) and cationic liposome-plasmid DNA complexes (lipoDNA) to experimental brain tumors. Adenovirus or lipoDNA was injected into the internal carotid artery (ICA) of F344 rats harboring intracerebral 9L gliosarcomas, using bradykinin (BK) to selectively permeabilize the blood-tumor barrier (BTB). Brain and internal organs of the animals were collected 48 hr after vector injection and stained for expression of the marker gene product, beta-galactosidase (beta-Gal). Intracarotid delivery of AV to 9L rat gliosarcoma without BTB disruption resulted in transgene expression in 3-10% of tumor cells distributed throughout the tumor. Virus-mediated expression of beta-gal gene products in this tumor model was particularly high in small foci (< or = 0.5 mm), which had invaded the normal brain tissue surrounding the main tumor mass. In these foci more than 50% of tumor cells were transduced. BK infusion increased the amount of transgene-expressing cells in larger tumor foci to 15-30%. In the brain parenchyma only a few endothelial cells expressed beta-gal owing to AV-mediated gene transfer. Intracarotid delivery of lipoDNA bearing a cytoplasmic expression cassette rendered more than 30% of the tumor cells positive for the marker gene without BTB disruption. The pattern of distribution was in general homogeneous throughout the tumor. BK infusion was able to increase further the number of transduced tumor cells to more than 50%. Although lipoDNA-mediated gene transfer showed increased efficacy as compared with AV-mediated gene transfer, it had less specificity since a larger number of endothelial and glial cells also expressed the transgene. AV and lipoDNA injections, in the absence and presence of BK, also resulted in transduction of peripheral organs. AV showed its known predilection for liver and lung. In the case of lipoDNA, parenchymal organs such as liver, lung, testes, lymphatic nodes, and especially spleen, were transduced. These findings indicate that intracarotid application of AV and lipoDNA vectors can effectively transduce tumor cells in the brain, and that BTB modulation by BK infusion can further increase the number of transgene-expressing tumor cells.

Therapeutic efficiency and safety of a second-generation replication-conditional HSV1 vector for brain tumor gene therapy.

A second-generation replication-conditional herpes simplex virus type 1 (HSV) vector defective for both ribonucleotide reductase (RR) and the neurovirulence factor gamma34.5 was generated and tested for therapeutic safety and efficiency in two different experimental brain tumor models. In culture, cytotoxic activity of this double mutant HSV vector, MGH-1, for 9L gliosarcoma cells was similar to that of the HSV mutant, R3616, which is defective only for gamma34.5, but was significantly weaker than that of the HSV mutant hrR3, which is defective only for RR. The diminished tumoricidal effect of the gamma34.5 mutants could be accounted for by their reduced ability to replicate in 9L cells. The MGH-1 vector did not achieve significant prolongation of survival in vivo in the syngeneic 9L rat gliosarcoma model for either single brain tumor focus or multiple intracerebral and leptomeningeal tumors, when the vector was applied intratumorally or intrathecally, respectively, and with or without subsequent ganciclovir (GCV) treatment. In identical 9L brain tumor models with single and multiple foci, application of hrR3 with or without GCV was previously shown to result in marked long-term survival. Contrary to the findings with intrathecal injection of hrR3, no vector-related mortality was observed in any animals treated with MGH-1. Thus, in these rat brain tumor models, the double mutant, replication-conditional HSV vector MGH-1 showed a higher therapeutic safety than the RR-minus vector, hrR3, but had clearly decreased therapeutic efficiency compared to hrR3. The development of new HSV vectors for brain tumor gene therapy will require a balance between maximizing therapeutic efficacy and minimizing toxicity to the brain. Standardized application in brain tumor models as presented here will help to screen new HSV vectors for these requirements.

Gene therapy for brain tumors.

Gene therapy has opened new doors for treatment of neoplastic diseases. This new approach seems very attractive, especially for glioblastomas, since treatment of these brain tumors has failed using conventional therapy regimens. Many different modes of gene therapy for brain tumors have been tested in culture and in vivo. Many of these approaches are based on previously established anti-neoplastic principles, like prodrug activating enzymes, inhibition of tumor neovascularization, and enhancement of the normally weak anti-tumor immune response. Delivery of genes to tumor cells has been mediated by a number of viral and synthetic vectors. The most widely used paradigm is based on the activation of ganciclovir to a cytotoxic compound by a viral enzyme, thymidine kinase, which is expressed by tumor cells, after the gene has been introduced by a retroviral vector. This paradigm has proven to be a potent therapy with minimal side effects in several rodent brain tumor models, and has proceeded to phase 1 clinical trials. In this review, current gene therapy strategies and vector systems for treatment of brain tumors will be described and discussed in light of further developments needed to make this new treatment modality clinically efficacious.

Selective delivery of herpes virus vectors to experimental brain tumors using RMP-7.

RMP-7, a bradykinin analog, has been shown to selectively open the blood-tumor barrier for the delivery of chemotherapeutic drugs to brain tumors. In contrast to bradykinin, RMP-7 has no hypotensive effects and has been approved for human use. This study was initiated to determine whether RMP-7 would open the blood-tumor barrier to virus vectors encoding tumor-killing genes in an experimental model. The herpes virus vector used, hrR3, which encodes virus thymidine kinase gene and the lacZ reporter gene, is defective in a gene encoding ribonucleotide reductase, replicates selectively in dividing tumor cells and not in postmitotic neural cells. It was determined that an optimum dose of RMP-7 (1.5-3.0 microg/kg over 10-15 minutes) enhanced viral delivery to brain tumors in rats bearing intracranial 9 L gliosarcomas when infused through the carotid artery immediately prior to virus vector application. Maximum expression of the lacZ reporter gene occurred at 3 days after intracarotid infusion. By 8 days, transgene expression was largely confined to tumor foci away from the main tumor mass. Viral delivery was essentially specific to tumor cells, with little transgene expression elsewhere in the brain. Minimal uptake and pathology was noted in the kidney, spleen, and liver. These findings indicate that intracarotid delivery of RMP-7 can augment the selective delivery of virus vectors to brain tumors in an experimental rat model, with the potential for application to human brain tumors.

Effectiveness of trabeculectomy in glaucoma.

A follow-up study of 90 eyes in 60 patients subjected to trabeculectomy between 1967 and 1972 showed that intraocular pressure was controlled at the onset in 84% of the eyes, and eventually controlled in over 97%. Only 11% of the eyes required further medication and 5.5% further surgery. Subconjunctival drainage was established by means of a bleb in 91%. Trabeculectomy produced a high significant fall in intraocular pressure (P less than .001) and a parallel rise in aqueous outflow facility. The absolute fall in intraocular pressure was constant whether the preoperative pressure was high or low and whether or not the postoperative pressure remained above 20 mm Hg. This method was virtually free of major operative and postoperative complications when used appropriately; and it can be modified during the operation to deal with peripheral anterior synechiae. The anterior chamber remained formed or, if lost, was speedily re-formed. The anterior chamber rarely flattened postoperatively.

Is 2% hydroxypropylmethylcellulose a safe solution for intraoperative clinical applications?

We examined 2% hydroxypropylmethylcellulose prepared for clinical intraocular surgical procedures in various laboratories. Each sample contained a variety of particulate debris of botanical origin, although in varying amounts. The identified material was also seen in a sample of the raw material from which all the clinical material had been prepared. Our conclusion is that filtration methods, which are the physical methods used to purify the product, are at worst ineffective and at best inadequate. Until proper laboratory and clinical studies confirm an acceptable level of adverse reactions, we recommend that this material not be used clinically.

Comparison of laterally condensed .06 and .02 tapered Gutta-Percha and sealer in vitro.

The purpose of this in vitro study was to compare the quality of the seal in canals prepared in a standardized manner and obturated with a .06 or a .02 tapered gutta-percha master cone using lateral condensation. Forty-four extracted human anterior teeth with single, straight canals were divided into two experimental groups of 20 teeth each and two control groups of 2 teeth each. The teeth were instrumented with Series 29 Profile .06 tapered rotary nickel-titanium files to a master apical file of 0.46 mm. Teeth in group 1 were obturated with a .02 tapered master gutta-percha cone and Roth 801 sealer using lateral condensation. Teeth in group 2 were obturated similarly, except a .06 tapered master gutta-percha cone was used. The depth of spreader penetration was recorded in millimeters. Positive control teeth were instrumented but not filled. Negative control teeth were instrumented, obturated, and externally sealed. The teeth were placed into a coronal leakage apparatus that contained an upper and lower reservoir of trypticase soy broth separated by the tooth. A 24-h growth of Proteus vulgaris in 0.25 ml of trypticase soy broth was placed in the coronal reservoir every 7 days for 70 days and incubated at 37 degrees C. Student's t test was used to determine whether there was a difference in spreader penetration between the groups, and a Fisher's exact test was used to determine whether there was a difference in bacterial leakage. The positive and negative controls validated the testing model. When a .02 tapered master cone was used, the spreader penetrated significantly closer to working length than when a .06 tapered master cone was used (p < 0.05). The difference between the groups in the number of samples that demonstrated complete bacterial penetration was not significant (p > 0.05).

A turnover study in the male rat of plasma-bound 59Fe, 114Inm and 109Cd with particular reference to the gonad.

After intravenous doses of the plasma-bound radionuclides 59Fe, 114Inm and 109Cd, only a minute percentage localizes in the rat testis and remains largely unchanged with time. Intratesticular injection of appropriately reduced volumes led to much higher proportionate percentage retention of 14, 65 and 11 for 59Fe, 114Inm and 109Cd, respectively. By this route, significant feedback of the elements escaping initial binding was prevented. Distinct but different testicular turnovers were now discernible. As a receptor of fluid and spermatozoa from the testicular tubules, the epididymis provides an indication of entry into and interaction of the metals with spermatogenic cells. For 59Fe no measurable changes were detected, whereas a progressive increase in epididymal 114Inm occurred, which had not reached a plateau by 70 days. 109Cd, now demonstrated within the testicular tubules by autoradiography, remained at constant organ level for upwards of 16 days but had declined by 25% by 57 days. At this point, the epididymis showed a five-fold increase in the radionuclide, declining to one-half this value by 126 days. Since 109Cd is carrier free, the data reflect a body turnover of dietary cadmium. These results, overall, are compatible with the entry of a proportion of each radionuclide into the seminiferous tubules and reaction with spermatogenic cells. Possible interpretations of the observed differences are presented.

Massive bifrontal epidermoid tumor.

There have been 27 cases of epidermoid tumor reported to arise from the frontal or ethmoid sinus area. We report the case of a virtually asymptomatic massive bifrontal epidermoid tumor arising from the ethmoid sinus in a 76-year-old male.

A behavioral approach to coaching football: improving the play execution of the offensive backfield on a youth football team.

Because of the emphasis on winning, the difficulties involved in assessing performance, and the lack of frequent and contingent reinforcement, a behavioral approach to coaching football was used. The players, all nine- or ten-year-old males, were members of an offensive backfield on a Pop Warner football team. Three frequently-run offensive plays were broken down into a series of five behaviorally defined stages, permitting construction of checklists suitable for observing the players during both game and scrimmage sessions. The intervention consisted of the presentation and explanation of the appropriate checklist, and frequent contingent reinforcement in the form of feedback and recognition for instances of desired play execution. Performance gains averaging 20% occurred for each of the three plays after, and not before, the staggered introduction of each intervention. The results suggest that behavioral specification and positive reinforcement of desired play execution is a viable approach to the coaching of football.