Prognostic implications of pathologically determined tumour volume in glottic carcinomas treated by transoral laser microsurgery.

OBJECTIVES: The TNM classification system for squamous cell carcinoma (SCC) of the head and neck neglects to incorporate volumetric analysis of the primary tumour. Tumour volume (TV) has been implicated prognostically in laryngeal SCC treated by primary radiotherapy (RT), but data for patients treated surgically are lacking. We evaluated such for glottic SCCs resected by transoral laser microsurgery (TLM). DESIGN: Retrospective cohort study utilising TVs calculated as the product of tumour resection dimensions and time-to-event analyses using the Kaplan-Meier method. The prognostic ability of variables was estimated using log-rank statistics, univariate Cox regression and receiver-operating characteristics analysis where appropriate. SETTING: Tertiary referral head and neck cancer centre. PARTICIPANTS: Patients undergoing primary TLM for glottic SCC with curative intent (2007-2011) with at least 12 months follow-up data. MAIN OUTCOME MEASURES: Prognostic impact of TV on local control (LC), overall survival (OS) and disease-specific survival (DSS). RESULTS: Eligible patients (n = 129) had a median follow-up of 40 months (range 14-79 months). Median TV for all cases was 300 mm(3) (range 2-19800 mm(3)). Three-year LC, OS and DSS were 92%, 92% and 98%, respectively. Tumour volume was not a significant predictor of any oncological outcome measure. Otherwise, a significant influence of pT stage on DSS was observed and of age on OS. CONCLUSIONS: In contrast to laryngeal SCC treated by RT, TV does not appear to portend oncological outcome in glottic SCC managed specifically by TLM and consequently does not warrant incorporation into current prognostic models for such patients.

Injection of botulinum toxin for the treatment of post-laryngectomy pharyngoesophageal spasm-related disorders.

Introduction Pharyngoesophageal spasm (PES) can cause dysphagia, central valve leak (CVL), and dypshonia in post-laryngectomy patients. Botulinum toxin has been used effectively for the treatment of PES, but data regarding patient-reported outcomes and efficacy for CVL are limited. We evaluated the results of botox injection for PES spasm using subjective and objective measures. Methods Data were collected prospectively (February 2010 to August 2013) on 13 patients undergoing botox injection for PES as identified by video fluoroscopy. We collected digital voice recordings, air-pressure measurements (APMs) for speech, and quality of life (QoL) data before and after the procedure: University of Washington QoL questionnaire (UW-QoL), MD Anderson Swallowing Inventory (MDADI) and the Voice Handicap Index (VHI-30). Results APMs for a sustained vowel decreased by 18% after botox injection, whereas maximum phonatory times increased by 63% (mean increase, 8 to 13 seconds). Sustained vowel amplitude decreased (mean, 87db to 83db) with an associated reduction in sustained vowel frequency (117Hz to 77Hz). MDADI scores improved by 10.2% overall, and UW-QoL scores showed an improvement in score of 7.6%. Mean scores for VHI-30 deteriorated by 2% overall but, when considering only those patients experiencing dysphonia, an improvement of 9.4% was seen. There was an overall net reduction for the CVL cohort of 12 speech valves in the year after injection. Conclusions Our series confirm the safety and objective efficacy of botox injection for PES. QoL measurements were less convincing, and this disparity between subjective and objective measurements must be considered when treating such patients.

IgG isotype antibody responses to epitopes of the Mycobacterium bovis protein MPB70 in immunised and in tuberculin skin test-reactor cattle.

Serological assays may have merit in identifying animals in advanced stages of bovine tuberculosis, but most tests have had sub-optimal sensitivities and specificities. The Mycobacterium bovis protein MPB70 has been identified as a B-cell target with diagnostic potential in measurement of pre- and post-skin-test antibody responses. One observation, which has potential practical application, has been that skin testing with tuberculin boosts IgG(1) anti-MPB70 antibody responses in cattle with tuberculous lesions. However, serological cross-reactivities with bacteria, such as Nocardia asteroides, have been described for this protein. With the aim of identifying candidate reagents for improved diagnostic tests, this study investigated IgG isotype antibody responses to MPB70 at the epitope level and, because of the previous findings, focused on IgG(1) responses following skin testing. Screening of a panel of overlapping synthetic peptides using sera from cattle immunised with MPB70 and cattle infected with M. bovis showed that two regions of the protein (residues 21-70 and 101-120) contain dominant B-cell epitopes. No individual epitope appeared to be selectively recognised by one isotype of IgG antibody. Investigation of IgG(1) responses showed that recognition of the epitope within residues 51-70 was boosted strongly by tuberculin injections in skin-test positive cattle and that this memory response was generally a feature of cattle which were found to have macroscopic, tuberculous lesions.

Mycobacterial antigen-specific antibody responses in bovine tuberculosis: an ELISA with potential to confirm disease status.

Serological assays may help to identify animals in advanced stages of bovine tuberculosis, but most of the tests available have suboptimal sensitivities and specificities. This study was designed to determine whether the antibody responses to defined antigens (rMPB70, rMPB64 and rMPB59) of Mycobacterium bovis at the immunoglobulin subclass level could be used to develop improved serological tests. In experimentally infected cattle it was found that the predominant serum antibody response was to rMPB70, and that an IgG1 response to this antigen was boosted strongly by skin testing. Studies in naturally infected cattle suggested that this memory IgG1 anti-rMPB70 response may be able to differentiate between skin test-reactor animals with and without lesions by comparing the ratio of the antibody response before and after skin testing. The study has provided a clearer understanding of the kinetics of antibody responses to defined mycobacterial antigens at the subclass level in bovine tuberculosis and has made it possible to develop a novel ELISA system which may be useful in disease diagnosis.

Assessment of defined antigens for the diagnosis of bovine tuberculosis in skin test-reactor cattle.

The continued use of purified protein derivative (PPD) tuberculin is considered to be the main factor which limits the specificity of diagnostic tests for bovine tuberculosis (TB). This study evaluated a whole blood interferon-gamma (IFN-gamma) assay and compared the diagnostic potential of PPD with two tuberculosis-specific antigens, ESAT-6 and MPB70. To provide estimates of sensitivity and specificity, responses were measured in 180 skin test-reacting cattle, of which 131 were confirmed as tuberculous, and in 128 cattle from TB-free herds. For the skin test reactors, there was a positive correlation between the IFN-gamma responses to PPD from Mycobacterium bovis (PPDB) and PPD from Mycobacterium avium (PPDA), indicating cross-reactivity between these complex antigens which are the basis of the skin test. In comparisons of the ESAT-6 IFN-gamma test with a PPD IFN-gamma test (using PPDB compared with PPDA), there was a decrease in sensitivity (76.3 per cent vs 89.3 per cent), but a clear increase in specificity (99.2 per cent vs 92.2 per cent). The provision of high specificity, even with lower sensitivity, offers major benefits for testing in areas with a low incidence of TB.

Pre-operative overnight pulse oximetry to predict high dependency unit intervention in children undergoing adenotonsillectomy for obstructive sleep apnoea.

Background: Post-operative high dependency unit beds are often requested for patients undergoing adenotonsillectomy for obstructive sleep apnoea. This study evaluated the utilisation of high dependency unit beds for such cases at our institution, a paediatric tertiary referral centre. Method: A retrospective case note review of patients admitted to the high dependency unit following adenotonsillar surgery for obstructive sleep apnoea, over a two-year period, was performed. Results: Sixty-six cases were identified. Thirty-nine patients underwent pre-operative overnight pulse oximetry; of these, 30 patients had desaturations noted. Seventeen patients had significant post-operative desaturations. These were predicted in all 11 patients who had undergone pre-operative pulse oximetry. The remaining six had not undergone pre-operative pulse oximetry. Nineteen patients required high dependency unit care; eight had experienced post-operative desaturations. Conclusion: High dependency unit care may be required following adenotonsillectomy for obstructive sleep apnoea. In this study, pre-operative overnight pulse oximetry had 100 per cent sensitivity in predicting post-operative desaturations, and may therefore aid the appropriate utilisation of high dependency unit beds for patients undergoing adenotonsillectomy for obstructive sleep apnoea.

Natural history of flexed knee gait in diplegic cerebral palsy evaluated by gait analysis in children who have not had surgery.

Eighteen children with diplegic cerebral palsy and no history of orthopaedic surgery had two gait analyses a mean of 6.3 years apart to analyse the effects of time on their gait. The mean age of the children at first analysis was 7.7 years (range 4.4-13.3 years). The data was analysed as a whole group (18 children) and as two sub-groups of nine children: those with a shorter follow-up (mean 5.0 years) and those with a longer follow-up (mean 7.5 years) between analyses. The following significant bilateral changes were seen in the whole group and longer follow-up sub-group: deterioration in the range of knee flexion, mid-stance knee flexion, peak knee extension in stance and hamstring length and an improvement in mean and maximum hip rotation. Temporal data showed no significant changes once normalised. There were no bilateral significant changes in data from children evaluated at a mean of 5 years follow-up. GMFCS scores generally improved over time despite the significant increase in flexed knee gait. There was no significant change in gait deviation index in any group over time. There was an increase in body mass index in 16 children but there was no correlation between this and the degree of mid-stance knee flexion. These findings may have implications for longer term follow-up of children with cerebral palsy into adulthood.

Recognition of a common mycobacterial T-cell epitope in MPB59 of Mycobacterium bovis.

Bovine tuberculosis, which persists as a residual level of infection in many European countries, has implications not only for the economy of farming communities but also for human health. The aim of this study was to identify a common mycobacterial antigen which was recognized in bovine tuberculosis and to characterize the response to this antigen at the epitope level. A T-cell clone, phenotype CD4+, raised from an animal experimentally infected with Mycobacterium bovis was shown to proliferate in response to a panel of sonicates derived from different mycobacterial species indicating recognition of an antigen with broad specificity. This antigen was subsequently shown to be MPB59. Recognition of MPB59 at the epitope level was determined in experimental and field cases of bovine tuberculosis using a panel of synthetic peptides (20-mers with 10-residue overlaps) incorporating the signal sequence and mature protein. The results showed that in vitro interferon-gamma was predominantly produced in response to adjacent peptides numbers 10 and 11, suggesting that the dominant epitope was contained in the overlap, correlating to residues 101-110 (YYQSGLSIVM). This epitope was recognized by 54% of tuberculous cattle of mixed breeds, which suggests that it may be genetically permissive in terms of major histocompatibility complex presentation. Sequence analysis confirmed that there were only minor differences in the amino acid composition within this region for various mycobacterial species, which could explain the common T-cell recognition described in this study. Common recognition of this epitope indicates that it would have limited potential for use as a diagnostic reagent per se but may have potential for inclusion in a subunit vaccine.

T-cell recognition of mycobacterial proteins MPB70 and MPB64 in cattle immunized with antigen and infected with Mycobacterium bovis.

Defined antigenic reagents and knowledge of T-cell responses are required for the design of improved diagnostic tests for bovine tuberculosis. The limited species distribution of Mycobacterium bovis antigens MPB70 and MPB64 has indicated their potential for inclusion in future tests. The strategy adopted in this study was to define bovine T-cell responses to these antigens at the epitope level, using cattle immunized with recombinant forms of the antigens, and to compare these responses with cattle which had been experimentally infected with M. bovis. Panels of synthetic peptides (20-mers with 10-residue overlaps) were used and five epitopes were identified and found to be powerful stimulators of T-cell responses in both types of animal (residues 81-100 and 174-190 for MPB70, and residues 1-20, 41-60 and 181-200 for MPB64). Further investigation in larger numbers of cattle (n = 14) of mixed breeds from tuberculosis-infected herds confirmed that each peptide produced response in several of the cattle, but no single peptide was recognized by all animals. However, the limited numbers of animals in this study suggest that peptide reagents may identify as many positive animals as the intact antigenic protein and could form components of a future diagnostic test. The use of cattle immunized with the proteins of interest has proved to be an interesting model for studying the nature of bovine T-cell responses to defined mycobacterial proteins.

Immunochemical characterization of the MPB70/80 and MPB83 proteins of Mycobacterium bovis.

MPB70 and MPB80 (MPB70/80) and MPB83 are closely related antigens which are highly expressed in Mycobacterium bovis. MPB70/80 are soluble secreted antigens, while MPB83 is an exported lipoprotein associated with the bacterial surface. In the present study, these antigens had different mobilities in sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing and nonreducing conditions. These differences may be explained by the fact that MPB70 and MPB83 both have two internal cysteine residues which would create ring structures by disulfide bonding. We analyzed the structures of MPB70/80 and MPB83 by using monoclonal antibodies (MAbs) raised against bovine purified protein derivative or whole M. bovis cells. MAb 1-5C reacted specifically with MPB70 and MPB80, and MAb MBS43 reacted specifically with MPB83, while the other antibodies, including several previously described MAbs, bound all three antigens. MAbs and polyclonal antibodies reacted strongly with reduced protein and less well with nonreduced protein, indicating involvement of linear epitopes. Epitopes of MAbs Bov-1, 2-6B, 1-5C, and 1-1D were mapped by using synthetic peptides of MPB70. Sequence comparison showed the peptide with the 1-5C-reactive epitope to have three residues different from those in the homologous region of MPB83. Exchanges of A for S in position 112 or Q for E in position 116 abolished the reactivity of MAb 1-5C. Polyclonal rabbit antibodies to native purified MPB70 reacted strongly with peptides 6, 7, and 8 of the N-terminal half of mature MPB70. Cattle sera of experimentally M. bovis-infected animals recognized a broader spectrum of peptides. These findings indicate that there is diagnostic potential for these proteins and that there is also a possible role for antibodies in elucidation of the host-mycobacterium relationship involving a surface-bound and exposed lipoprotein, MPB83, and its highly homologous soluble secreted MPB70/80 counterparts.