Hypnosis and progressive muscle relaxation for anxiolysis and pain control during extraction procedure in 8-12-year-old children: a randomized control trial.

INTRODUCTION: Hypnosis (H) and Progressive Muscle Relaxation (PMR) have proven to be effective in a variety of medical settings; there is a paucity of their practical application in paediatric dentistry. The study aimed to comparatively evaluate the role of H and PMR on anxiety, heart rate (HR), oxygen saturation (SPO2), blood pressure (BP), pain, and analgesic requirement during extraction in children. MATERIALS AND METHODS: Sixty children aged 8-12 years undergoing primary molar extractions were randomly allocated to three groups-H, PMR, and control (C). The anxiety (proposed Visual Facial Anxiety scale), HR, and SPO2 were measured pre/post-operatively with/without interventions (H, PMR, C) at 4 intervals. The BP and pain (Wong-Baker faces pain scale) were recorded pre- and post-operatively. Need for analgesic post-operatively was assessed. RESULTS: Statistically significant reduction in anxiety was noted post-extraction in H (0.30 ± 0.80), PMR (0.50 ± 0.69) (p < 0.001*). HR showed a statistically significant drop after H, PMR application. (p < 0.001*) No significant difference in SPO2 was noted in the three groups (p > 0.05). Pain control was well achieved using H (85%), PMR (70%); BP was well-regulated in the H, PMR compared to C group (p < 0.001*). Need for analgesics was reduced in H (45%), PMR (50%) versus C (100%). Both techniques H, PMR were comparable in all measures. CONCLUSION: Hypnosis and PMR are effective techniques for anxiolysis and pain control in paediatric dental patients.

SLC28A3 genotype and gemcitabine rate of infusion affect dFdCTP metabolite disposition in patients with solid tumours.

BACKGROUND: Gemcitabine is used for the treatment of several solid tumours and exhibits high inter-individual pharmacokinetic variability. In this study, we explore possible predictive covariates on drug and metabolite disposition. METHODS: Forty patients were enrolled. Gemcitabine and dFdU concentrations in the plasma and dFdCTP concentrations in peripheral blood mononuclear cell were measured to 72 h post infusion, and pharmacokinetic parameters were estimated by nonlinear mixed-effects modelling. Patient-specific covariates were tested in model development. RESULTS: The pharmacokinetics of gemcitabine was best described by a two-compartment model with body surface area, age and NT5C2 genotype as significant covariates. The pharmacokinetics of dFdU and dFdCTP were adequately described by three-compartment models. Creatinine clearance and cytidine deaminase genotype were significant covariates for dFdU pharmacokinetics. Rate of infusion of <25 mg m(-2) min(-1) and the presence of homozygous major allele for SLC28A3 (CC genotype) were each associated with an almost two-fold increase in the formation clearance of dFdCTP. CONCLUSION: Prolonged dFdCTP systemic exposures (≥72 h) were commonly observed. Infusion rate <25 mg m(-2) min(-1) and carriers for SLC28A3 variant were each associated with about two-fold higher dFdCTP formation clearance. The impacts of these covariates on treatment-related toxicity in more selected patient populations (that is, first-line treatment, single disease state and so on) are not yet clear.

Targeting aurora kinases: a novel approach to curb the growth & chemoresistance of androgen refractory prostate cancer.

Aurora Kinase (AK) based therapy targeting AK-A & B is effective against some cancers. We have explored its potential against previously unreported incurable, metastatic androgen depletion independent Prostate Cancer (ADIPC). We used androgen sensitive (AS) and ADI lines derived from Transgenic Adenocarcinoma of the Mouse Prostate (TRAMP) mice. The relevance of this model was unequivocally established through focussed array, quantitative PCR and western blotting studies; significantly greater alteration of genes (fold change and number) representing major cancer pathways was shown in ADI cells compared to AS lines. A marked enhancement of in vivo growth of the ADI subline showing the greatest degree of gene modulations [TRAMP C1 (TC1)-T5: TC1-T5] reflected this. In contrast to the parental AS TC1 line, TC1-T5 cells grew with 100% incidence in the prostate, as lung pseudometastases and migrated to the bone and other soft tissues. The potential involvement of AKs in this transition was indicated by the significant upregulation of AK-A/B and their downstream regulators, survivin and phosphorylated-histone H3 in TC1-T5 cells compared to TC1 cells. This led to enhanced sensitivity of TC1-T5 cells to the pan-AK inhibitor, VX680 and to significant reduction in in vivo tumour growth rates when AK-A and/or B were downregulated in TC1-T5 cells. This cell growth inhibition was markedly enhanced when both AKs were downregulated and also led to substantially greater sensitivity of these cells to docetaxel, the only chemotherapeutic with activity against ADI PC. Finally, use of VX680 with docetaxel led to impressive synergies suggesting promise for treating clinical ADI metastatic PC.

CD147/EMMPRIN and CD44 are potential therapeutic targets for metastatic prostate cancer.

Prostate cancer (CaP) is a major health problem in males in Western countries. Current therapeutic approaches are limited and many patients die of secondary disease (metastases). There is no cure for metastatic castration-resistant prostate cancer (CRPC). Targeting tumor-associated antigens is fast emerging as an area of promise to treat late stage and recurrent CaP. Extracellular matrix metalloproteinase inducer, EMMPRIN (CD147) is a multifunctional glycoprotein that can modify the tumor microenvironment by activating proteinases, inducing angiogenic factors in tumor and stromal cells, and regulating growth and survival of anchorage-independent tumor cells (micrometastases) and multidrug resistance (MDR). CD44 is a multifunctional protein involved in cell adhesion, migration and drug resistance, and is a primary receptor for hyaluronan (HA), a major component of the extracellular matrix (ECM) with a critical role in cell signaling and cell-ECM interactions in cancer. Our recent studies indicate both CD147 and CD44 are involved in cancer drug resistance and play very important roles in CaP metastasis. Thus, CD147 and CD44 may be ideal therapeutic targets to control metastatic and CRPC disease. This review will discuss their putative roles in CaP metastasis and MDR, and give an overview of literature regarding their expression on human CaP tissues. Additional focus will be on the potential of therapeutic strategies targeting CD147 and CD44 to prevent CaP metastasis and overcome drug resistance.

Molecular and traditional chemotherapy: a united front against prostate cancer.

Castrate resistant prostate cancer (CRPC) is essentially incurable. Recently though, chemotherapy demonstrated a survival benefit ( approximately 2months) in the treatment of CRPC. While this was a landmark finding, suboptimal efficacy and systemic toxicities at the therapeutic doses warranted further development. Smart combination therapies, acting through multiple mechanisms to target the heterogeneous cell populations of PC and with potential for reduction in individual dosing, need to be developed. In that, targeted molecular chemotherapy has generated significant interest with the potential for localized treatment to generate systemic efficacy. This can be further enhanced through the use of oncolytic conditionally replicative adenoviruses (CRAds) to deliver molecular chemotherapy. The prospects of chemotherapy and molecular-chemotherapy as single and as components of combination therapies are discussed.

Palliative care services for those with chronic lung disease.

Excellent palliative care is available for patients with advanced lung cancer. Whether the same services are available for those with nonmalignant respiratory disease is less clear. A questionnaire was sent to 210 named respiratory physicians, each representing a major hospital in England, Wales, and Northern Ireland. A total of 107 replies were received; the response rate was 51.0%. Respondents cared for patients with chronic obstructive pulmonary disease, asbestosis, and diffuse parenchymal lung disease but only a third had responsibility for cystic fibrosis. Physicians were supported by a mean of 3.4 respiratory nurse specialists per department and 73.8% had a specialist lung cancer nurse. In only 16 cases (20.3%) did that nurse extend care to those with nonmalignant disease. Only a minority reported easy access to hospice in-patient care or day care. About 21.5% of the respondents had formal policies in place for care of patients with chronic respiratory disease nearing the end of life, but 87.9% of respondents had no formal process for initiating end of life discussions with those with terminal respiratory illness. Patients with advanced nonmalignant respiratory disease have less universal access to specialist palliative care services than do those with malignant lung disease, and in the majority of hospitals there is no formalized approach to end of life care issues with patients with chronic lung disease.

Reflecting the scope and work of palliative care teams today: an action research project to modernise a national minimum data set.

The Minimum Data Set (MDS) for UK specialist palliative care services was developed in 1995 to provide annual data on palliative care services. Data collected is used for local and national purposes including service management, monitoring and audit, the commissioning of services and the development of national policy. The emergence of Payment by Results and HealthCare Resource Groups, which will have an impact on the funding processes, together with identified limitations of the current MDS resulted in a project to revise the MDS. An action research approach was used for the project and had distinctive phases including modifying the MDS, a pilot phase and an expert panel consultation. Modifications to all the sections of the MDS and changes to terminology were made. The action research approach enabled revisions made based upon a national consensus and met the changing provision of specialist palliative care services for the UK.

Comparative evaluation of shear bond strength of conventional composite resin and nanocomposite resin to sandblasted primary anterior stainless steel crown.

To evaluate and compare the shear bond strength of conventional composite resin and nanocomposite resin to sandblasted primary anterior stainless steel crown. The study samples consisted of 30 primary anterior stainless steel crowns (Unitek TM, size R4), embedded in resin blocks with crown, in test groups of 15 samples each. Mounting of the crown was done using resin block with one crown each. Sandblasting was done and the bonding agent Prime and Bond NT (Dentsply) was applied on the labial surface of the primary anterior sandblasted crown. The composite resin and nanocomposite resin were placed into the well of Teflon jig and bonded to Stainless Steel Crowns. The cured samples were placed in distilled water and stored in incubator at 37 degrees C for 48 hours. Shear bond strength was measured using universal testing machine (Hounsefield U.K. Model, with a capacity of 50 KN). Independent sample 't' test revealed a nonsignificant (P < 0.385) difference between mean shear bond strength values of conventional and nanocomposite group. The bond strength values revealed that nanocomposite had slightly higher mean shear bond strength (21.04 +/- 0.56) compared to conventional composite (20.78 +/- 0.60). It was found that conventional composite resin and nanocomposite resin had statistically similar mean shear bond strength, with nanocomposite having little more strength compared to conventional composite.

Consistent cytotoxic-T-lymphocyte targeting of immunodominant regions in human immunodeficiency virus across multiple ethnicities.

Although there is increasing evidence that virus-specific cytotoxic-T-lymphocyte (CTL) responses play an important role in the control of human immunodeficiency virus (HIV) replication in vivo, only scarce CTL data are available for the ethnic populations currently most affected by the epidemic. In this study, we examined the CD8(+)-T-cell responses in African-American, Caucasian, Hispanic, and Caribbean populations in which clade B virus dominates and analyzed the potential factors influencing immune recognition. Total HIV-specific CD8(+)-T-cell responses were determined by enzyme-linked immunospot assays in 150 HIV-infected individuals by using a clade B consensus sequence peptide set spanning all HIV proteins. A total of 88% of the 410 tested peptides were recognized, and Nef- and Gag-specific responses dominated the total response for each ethnicity in terms of both breadth and magnitude. Three dominantly targeted regions within these proteins that were recognized by >90% of individuals in each ethnicity were identified. Overall, the total breadth and magnitude of CD8(+)-T-cell responses correlated with individuals' CD4 counts but not with viral loads. The frequency of recognition for each peptide was highly correlated with the relative conservation of the peptide sequence, the presence of predicted immunoproteasomal cleavage sites within the C-terminal half of the peptide, and a reduced frequency of amino acids that impair binding of optimal epitopes to the restricting class I molecules. The present study thus identifies factors that contribute to the immunogenicity of these highly targeted and relatively conserved sequences in HIV that may represent promising vaccine candidates for ethnically heterogeneous populations.

Vpr is preferentially targeted by CTL during HIV-1 infection.

The HIV-1 accessory proteins Vpr, Vpu, and Vif are essential for viral replication, and their cytoplasmic production suggests that they should be processed for recognition by CTLs. However, the extent to which these proteins are targeted in natural infection, as well as precise CTL epitopes within them, remains to be defined. In this study, CTL responses against HIV-1 Vpr, Vpu, and Vif were analyzed in 60 HIV-1-infected individuals and 10 HIV-1-negative controls using overlapping peptides spanning the entire proteins. Peptide-specific IFN-gamma production was measured by ELISPOT assay and flow-based intracellular cytokine quantification. HLA class I restriction and cytotoxic activity were confirmed after isolation of peptide-specific CD8(+) T cell lines. CD8(+) T cell responses against Vpr, Vpu, and Vif were found in 45%, 2%, and 33% of HIV-1-infected individuals, respectively. Multiple CTL epitopes were identified in functionally important regions of HIV-1 Vpr and Vif. Moreover, in infected individuals in whom the breadth of HIV-1-specific responses was assessed comprehensively, Vpr and p17 were the most preferentially targeted proteins per unit length by CD8(+) T cells. These data indicate that despite the small size of these proteins Vif and Vpr are frequently targeted by CTL in natural HIV-1 infection and contribute importantly to the total HIV-1-specific CD8(+) T cell responses. These findings will be important in evaluating the specificity and breadth of immune responses during acute and chronic infection, and in the design and testing of candidate HIV vaccines.

Enhanced activity in parathyroid hormone-(1-14) and -(1-11): novel peptides for probing ligand-receptor interactions.

The amino-terminal portion of PTH is critical for PTH-1 receptor (P1Rc) activation. In exploring this component of the ligand receptor interaction, we recently showed that the agonist potency of the weakly active PTH-(1-14)NH(2) peptide can be enhanced by natural amino acid substitutions at several positions, including position 11 (normally leucine). Here we show that the potency of PTH-(1-14)NH(2) can be enhanced by using nonnatural amino acids that increase the length and polarizability of the position 11 side-chain. Thus, in LLC-PK(1) cells stably expressing high levels of the human P1Rc, [homoarginine([Har)(11)]PTH-(1-14)NH(2) was 30-fold more potent for cAMP production than was native PTH-(1-14)NH(2). Combining the homoarginine-11 substitution with other recently identified activity-enhancing substitutions yielded [Ala(3,12),Gln(10),Har(11),Trp(14)]PTH-(1-14)NH(2), which was 1500-fold more potent than PTH-(1-14)NH(2) (EC(50) = 0.12 +/- 0.04 and 190 +/- 20 microM, respectively) and only 63-fold less potent than PTH-(1-34) (EC(50) = 1.9 +/- 0.5 nM). The even shorter analog [Ala(3),Gln(10),Har(11)]PTH-(1-11)NH(2) was also a full cAMP agonist (EC(50) = 3.1 +/- 1.5 microM). Receptor mutations at Phe(184) and Leu(187) located near the boundary of the amino-terminal domain and transmembrane domain-1 severely impaired responsiveness to the PTH-(1-11) analog. Overall, these studies demonstrate that PTH analogs of only 11 amino acids are sufficient for activation of the PTH-1 receptor through interaction with its juxtamembrane region.

The HIV-1 regulatory proteins Tat and Rev are frequently targeted by cytotoxic T lymphocytes derived from HIV-1-infected individuals.

The HIV-1 regulatory proteins Rev and Tat are expressed early in the virus life cycle and thus may be important targets for the immune control of HIV-1-infection and for effective vaccines. However, the extent to which these proteins are targeted in natural HIV-1 infection as well as precise epitopes targeted by human cytotoxic T lymphocytes (CTL) remain to be defined. In the present study, 57 HIV-1-infected individuals were screened for responses against Tat and Rev by using overlapping peptides spanning the entire Tat and Rev proteins. CD8+ T cell responses against Tat and Rev were found in up to 19 and 37% of HIV-1-infected individuals, respectively, indicating that these regulatory proteins are important targets for HIV-1-specific CTL. Despite the small size of these proteins, multiple CTL epitopes were identified in each. These data indicate that Tat and Rev are frequently targeted by CTL in natural HIV-1 infection and may be important targets for HIV vaccines.

Copper catalyzed oxidation of Alzheimer Abeta.

Abeta derived from amyloid plaques of Alzheimer's disease-affected brain contain several oxidative posttranslational modifications. In this study we have characterized the amino acid content of human amyloid-derived Abeta and compared it with that of human synthetic Abeta subjected to metal-catalyzed oxidation. Human amyloid derived Abeta has an increased content of arginine (46%) and glutamate/glutamine residues (28%), but a decreased content of histidine residues (-32%) as compared to the expected amino acid content. Incubation of synthetic human Abeta with Cu(II), but not Fe(III), in the presence of H2O2 similarly induced a decrease in histidine residues (-79%), but also a decrease in tyrosine residues (-28%). Our results suggest that histidine and tyrosine are most vulnerable to metal mediated oxidative attack, consistent with our earlier findings that Cu coordinated via histidine residues is redox competent. Our results suggest that the loss of histidine residues in human amyloid-derived Abeta may be a result of Cu oxidation, and that unidentified post-translational mechanisms operate to modify other amino acids of Abeta in vivo.

Strategies for the synthesis of labeled peptides.

Labeled peptides synthesized by core facilities are frequently used by researchers for following trafficking of a peptide, for binding studies, to determine substrate specificity, and for receptor cross-linking studies.The membership of the Association of Biomolecular Resource Facilities was asked to participate in a study focusing on synthesis of a biotin-labeled peptide, and it was suggested that a new strategy, using Rink amide 4-methylbenzhydrylamine resin coupled with Fmoc-Lys(Dde)-OH, be used.This strategy can be used for addition of a variety of labels other than biotin and should prove useful to core facilities. Comparison of the new strategy to other strategies was performed. Biotin labeling has long been assumed to be routine and specific. Despite the assumed routine nature of synthesizing biotinylated peptides, 9 of the 34 samples submitted did not contain any of the correct product. Although synthesis using Fmoc-Lys(Dde)-OH plus biotin generally gave the highest yields, other approaches also yielded a high percentage of the correct product.Therefore, the various strategies are generally comparable. The major advantage of this new approach is that other labels such as fluorescein, dansyl groups, methyl coumarin, and potentially fluorophores and quenchers used for fluorescence resonance energy transfer (FRET) can be directly incorporated into peptides.

Study on the synthesis and characterization of peptides containing phosphorylated tyrosine.

Phosphorylation and dephosphorylation are key events in receptor-mediated and post-receptor-mediated signal transduction. Synthetic phosphopeptides have been shown to have dramatic agonist or antagonist effects in several of these signaling pathways. For its 1997 study, the Association of Biomolecular Resource Facilities (ABRF) Peptide Synthesis Research Group assessed the ability of member laboratories to synthesize phosphotyrosine peptides. Participating laboratories were requested to synthesize and submit the following crude peptide, H-Glu-Asp-Tyr-Glu-Tyr(PO3H2)-Thr-Ala-Arg-Phe-NH2, for evaluation by amino acid analysis, sequence analysis, RP-HPLC, MALDI-TOF and ESI mass spectrometry. Prior to analysis of submitted peptides from ABRF members, the Peptide Synthesis Research Group synthesized and characterized the nonphosphorylated form of the peptide, the doubly phosphorylated form and the peptides singly phosphorylated on either the first or the second tyrosine. These peptide standards were separated easily by HPLC and capillary electrophoresis and the phosphotyrosine was detected readily by Edman degradation sequence analysis. No differences were seen by amino acid analysis and the expected masses were observed by mass spectrometry. The two singly phosphorylated peptides were easily distinguished by MALDI-PSD. Analysis of the peptides submitted from member facilities revealed that all but four of the 33 samples contained the correct product as determined by HPLC and mass spectrometry. HPLC analysis indicated that 20 of the 33 submitted samples contained greater than 75% correct product, five contained less than 50% correct product and four did not contain any correct product. By ESI/MS, an additional singly charged ion at m/z 535.5 was detected in five of the 33 submitted samples; this ion was subsequently shown to represent Ac-TARF-NH2. No correlation was found to exist between coupling time and percentage correct product; however, a correlation may exist between a greater percentage of correct product and the use of non-protected phosphotyrosine.

Identification of transcripts and promoter regions of ovine adenovirus OAV287.

The ovine adenovirus isolated OAV287 represents a new group of adenoviruses that are distinct from the Mast- and Aviadenoviruses by several criteria, including genome arrangement. The OAV major late promoter and some late transcripts were previously mapped. To better define the probable coding sequences and to identify the approximate location of early promoters a partial transcription map of the genome was elucidated using a PCR-based approach. This was possible because the complete nucleotide sequence of the genome was known. The strategy permitted the identification of transcription start sites and RNA splice junctions and allowed the approximate location of promoters in the lefthand end, IVa2, E2, P32K, and E4 regions to be deduced. The data showed that lefthand end and E4 regions are controlled by three and two temporally distinct promoters, respectively. The E2 region is controlled by a single promoter, in contrast to Mastadenoviruses, where E2 expression is controlled by the E2A and E2B promoters. The p32kDa structural protein at the lefthand end and the IVa2 protein are also expressed from their own promoters. These data contribute to the first overview of transcription from a non-Mastadenovirus genome.

Erythrocyte membrane stearic to oleic acid ratio in carcinoma of the gallbladder: a preliminary study.

AIMS: The role of erythrocyte membrane stearic to oleic acid ratio (saturation index) as a marker of malignancy is still unclear, though an association has been found in colorectal carcinoma, bronchogenic carcinoma, leukaemia, lymphoma and in hepatic malignancies. This study aims to investigate the role of the saturation index in primary carcinoma of the gallbladder. METHODS: This paper describes the results of the stearic to oleic acid ratio determination in 26 subjects with either cholelithiasis or carcinoma of the gallbladder, also including a group of age- and sex-matched controls, using gas chromatography. This is the first report of the saturation index in carcinoma of the gallbladder. RESULTS: A significantly lower saturation index was observed in patients with carcinoma of the gallbladder than with cholelithiasis (t = 2.19, P = 0.043, T = 47, P < 0.05, Wilcoxon P < 0.001, F = 2192.23, P < 0.001; 95% CI 18.45-30.44) and controls (t = 2.5, P = 0.024, T = 36, P < 0.05, F = 10904.11, P < 0.001, Wilcoxon P < 0.001; 95% CI 52.42-63.39). Among the carcinoma patients a further lowering was noted in stage IV disease compared with stage III (T = 6, P < 0.05). CONCLUSIONS: These changes are probably due to a marked increase in oleic acid content at the expense of stearic acid. This lowering of the saturation index in carcinoma of the gallbladder is similar to that observed previously in the other malignancies.