<p>The diagnosis and treatment of patients with advanced tumors in the head and neck is an interesting challenge where there is a need for new approaches in diagnostics and adjuvant treatment. Differences in antigen expression between tumors and normal tissues provide a means for application of antibody-based targeting techniques. By targeting a structure that is abundant on tumor cells and limited on normal cells, radioactivity can be delivered.</p><p>The use of positron emission tomography (PET) in patients with head and neck tumors is evaluated in this thesis. PET using the tracer fluorodeoxyglucose (FDG) is found to play an important diagnostic role and often has a direct clinical impact on planned surgery or other treatment. Possible targeting structures are also investigated in this thesis, and it is concluded that the EGFR and CD44v6 stand out as possible antigens for targeting approaches of squamous cell carcinomas in the head and neck (HNSCC). A radioimmunoassay for quantification of EGFR and CD44v6 is validated and concluded to be a valuable complement to immunohistochemistry for the analysis of tumors and for the planning of radioimmunotherapy. Finally, promising results of radioimmunotherapy in tumor bearing mice with the monoclonal antibody U36 labeled with the alpha emitter astatine-211 are presented.</p><p>These results demonstrate how differences between tumors and normal tissues can be used to improve diagnostic outcomes and indicate that radioimmunotherapy can be a future adjuvant therapy or treatment of residual disease in HNSCC.</p>

The diagnosis and treatment of patients with advanced tumors in the head and neck is an interesting challenge where there is a need for new approaches in diagnostics and adjuvant treatment. Differences in antigen expression between tumors and normal tissues provide a means for application of antibody-based targeting techniques. By targeting a structure that is abundant on tumor cells and limited on normal cells, radioactivity can be delivered. The use of positron emission tomography (PET) in patients with head and neck tumors is evaluated in this thesis. PET using the tracer fluorodeoxyglucose (FDG) is found to play an important diagnostic role and often has a direct clinical impact on planned surgery or other treatment. Possible targeting structures are also investigated in this thesis, and it is concluded that the EGFR and CD44v6 stand out as possible antigens for targeting approaches of squamous cell carcinomas in the head and neck (HNSCC). A radioimmunoassay for quantification of EGFR and CD44v6 is validated and concluded to be a valuable complement to immunohistochemistry for the analysis of tumors and for the planning of radioimmunotherapy. Finally, promising results of radioimmunotherapy in tumor bearing mice with the monoclonal antibody U36 labeled with the alpha emitter astatine-211 are presented. These results demonstrate how differences between tumors and normal tissues can be used to improve diagnostic outcomes and indicate that radioimmunotherapy can be a future adjuvant therapy or treatment of residual disease in HNSCC.

Tonsillectomy is one of the most frequently performed surgical procedures in children and youths. The aim of this thesis was to study children and youths in relation to tonsil surgery with the goal of improving the care, and to describe partial tonsillectomy/tonsillotomy (TT) using radiofrequency technique (RF) (Ellman International) in comparison with the more commonly used total tonsillectomy (TE). The thesis covers studies of wo age-groups with obstructive problems, with or without recurrent tonsillitis. Randomization to surgery was done from the existing waiting list; 92 children, 5-15 years old to 49/TT and 43/TE, (I-III) and 76 youths, 16-25 years old to 32/TT and 44/TE (IV-V). The first purpose (I, IV) was to compare the two surgical techniques with respect to pain and postoperative morbidity. Pain measures were for the children the Face Pain Scale and for the youths and parents and staff a verbal-pain-rating-scale. From the first day, the TT-groups scored significantly less pain than the TE-groups. The doses of pain-killing drugs (paracetamol and diclofenac) taken were significantly less for the children and youths receiving the TT-surgery, they could stop taking pain-killers sooner, and were back to normal activity three (5-15yrs) or four (16-25yrs) days earlier compared with TE-groups. Paper II focused on the child’s behavior (Child Behavior Checklist/CBCL), experience of pain, anxiety (State-Trait-Anxiety Inventory for Children /STAIC), previous experiences of surgery/tonsillitis, and the management of pain. The children scored higher on CBCL than a normative group before surgery, but no connection was observed between CBCL rating and experience of pain reported post surgically. There was no relation between preoperative anxiety and reported pain, but the postoperative anxiety level correlated with pain. The Egroup scored higher anxiety after surgery. Previous experience of surgery or tonsillitis did not influence the postoperative pain. The nurses scored pain lower than the parents/children and under-medicated. The second purpose was to compare the long-term effects of TT and TE-surgery after one and three years (5-15yrs) and one year (16-25yrs) (III, IV). The effect on snoring was the same for both TT and TE-groups and the rate of recurrence of throat infections was low after both surgical techniques. After one year, all children (TT/TE) showed improvements on CBCL to the same degree and there was no longer a difference between total behavior and normative values. They also scored improvements in health-related quality of life (HRQL) with Glasgow-Children-Benefit-Inventory. For both TT and TE, the older group reported lower HRQL preoperatively on all dimensions of Study-Short-Form (SF-36) compared with a normal population. After one year, a large improvement was found in HRQL in both groups and there were no differences compared with a normal population. Conclusion: Preoperative obstructive problems, in combination with recurrent tonsillitis have a negative impact on HRQL. Both after TE and TT there are large improvements in HRQL, infections, obstructive, and behavior problems one to three years after surgery, indicating that both surgical methods are equally effective. With fewer postoperative complications, less pain, shorter recovery time, and lower cost, TT with RF should be considered as method of choice.

Antibody-based targeting techniques play an increasingly important role in cancer research. By targeting a structure that is abundant in tumour cells, but rare in healthy tissues, an antibody can mediate the delivery of radioactivity specifically to tumour cells in the body. This idea is particularly appealing for head and neck squamous cell carcinoma (HNSCC), as the advanced stages have a large fraction of spread disease that is difficult to treat with procedures available today. In this thesis, we have investigated possible radioimmunotargeting structures for HNSCC, and found that CD44v6 is a suitable target for antibody-based radiotherapy and diagnostics in this patient group. We have identified radiohalogens as attractive nuclides for such use, and have investigated the possibility of radiohalogenating the anti CD44v6 chimeric monoclonal antibody (cMAb) U36. Several feasible labelling methods were identified, using both direct and indirect labelling. The cMAb U36 was then successfully labelled with 211At and 131I, and preclinically evaluated for therapeutic use. Results proved the astatinated conjugate to be most efficient in this context, demonstrating a specific and dose-dependent cytotoxicity. The cMAb U36 was then evaluated for diagnostic use in thyroid anaplastic carcinoma, using 124I as the diagnostic nuclide. Results in tumour-bearing mice were promising, with all of the tumours identified in micro-PET studies. These results demonstrate how antibody-based radionuclide targeting can provide more sensitive and specific methods for identifying and treating head and neck cancer, and hopefully help improve long-term survival rates for this patient group in the future.

The distribution of cell types and synapses is well characterized in the rodent olfactory bulb (OB), and from that plausible models of odor processing have been constructed. Individual olfactory sensory neurons (OSNs) express only 1 of ~1000 odorant receptors (ORs) and send their axons to specific synaptic targets in the OB glomerular neuropil. Each glomerulus is innervated exclusively by OSN axons expressing the same OR. The distribution of these glomeruli is conserved across animals, as is the numerical relationship between number of expressed ORs and number of glomeruli in the OB. Our objective is to extend such results to the level of the human OB to determine how its cellular and synaptic organization, and more specifically how the number and distribution of its glomeruli, compare to what has been elucidated in mice. As there are ~2,000 glomeruli for ~1,000 ORs in mice, we predicted ~700 glomeruli in humans based on the ~350 intact OR genes identified in the human through genomic studies. Using immunohistochemistry, the organization of cells and synapses in human OBs was evaluated and quantified. While the laminar structure of the OB is broadly conserved between species, in the human OB the laminar organization as well as additional structural features suggest a less rigorously organized OB than in rodents, perhaps suggesting that odor processing in the human OB may be less efficient than in mice. Of particular note, the total number of glomeruli in the human OB differs significantly from predicted and demonstrates a high degree of variability amongst specimens, thus far ranging from approximately 3000 - 9000/OB. These results indicate that the principles of OR-homotypic axon convergence developed from mouse studies may not be readily applicable to the human, and that central processing of odor signals in the human may differ from those characterized in the mouse.

The healing of tympanic membrane (TM) perforations is a complex wound healing process including inflammation, migration of keratinocytes and tissue remodelling. Most TM perforations in human heal spontaneously, however some perforations become chronic, and the reason to why is still largely unknown. In cutaneous wound healing plasminogen (plg) has been shown to play an important role. Plg is converted into the protease plasmin regulated by two plasminogen activators (PA), urokinase type PA (uPA) and tissue-type PA (tPA). The aim of the present thesis was to evaluate the role of plg in healing of TM perforations, both in vivo and in vitro. The main objectives were to determine the healing capacity of the TM, the involvement of keratinocytes, fibrin(ogen) and inflammatory cells in the healing process. The studies were performed in plg deficient and uPA deficient mice, with littermate wild type (wt) mice as controls It was shown that myringotomies of the TMs in plg deficient mice still remained open 143 days following a perforation. The wound area was characterized by an abundant recruitment and accumulation of inflammatory cells; mainly macrophages and neutrophils, an arrested keratinocyte migration and a fibrin deposition covering the surface of the TM. The TM perforations in the wt mice all healed within 11 days. Interestingly, the myringotomies of the plg deficient mice could be closed by reconstitution with systemic injections of plg, whereas injections of PBS had no affect on the healing. To characterize mechanisms involved in the development of persistent TM perforations in plg deficient mice after a myringotomy the early inflammatory response during the first 48 hours was studied. The recruitment and accumulation of inflammatory cells in the perforated TMs was found to be similar between the plg deficient and the wt mice. Myringotomized TMs in uPA deficient mice healed similar to perforations of wt controls. Neither did the keratinocyte migration nor the occurrence of inflammatory cells differ between these genotypes. In the in vitro experiments TMs from plg deficient and wt mice, were dissected out, perforated and cultured in absence or surplus of plg. A decrease in perforation size was seen in all groups regardless of genotype or amount of plg in the medium. In conclusion, the present studies show: • Plg is essential for the healing of TM perforations in mice. • The altered healing process after a myringotomy in plg deficient mice involves a disturbed keratinocyte migration, a massive deposition of fibrin and an abundant accumulation of inflammatory cells in the wound area. • Plasminogen deficiency does not alter the early inflammatory response, following a myringotomy. • Deficiency of uPA does not influence the healing of TM perforations. • During in vitro conditions healing of TM perforations is initiated irrespectively of genotype of the explant (plg deficient or wt) or supply of plg. The increased knowledge of the involvement of plg in the healing of TM perforations may open therapeutical possibilities in the treatment of chronic TM perforations in humans.

Monogenic disorders offer a possibility for studies of genetic disturbances in hearing impairment—a knowledge which could be essential for development of future treatment options. In this thesis, the underlying genetic disturbances in neurofibromatosis 2 (NF2) and familial Meniere’s disease (FMD) were evaluated, and familial X-linked hearing impairment was described from a clinical point of view. In paper I, constitutional DNA from 116 individuals with NF2 of variable severity was studied using the array-CGH method focusing on a 7.6-Mb area surrounding the NF2 gene on chromosome 22q. Deletions were found in 20.7% of samples. In mild NF2, the deletions were small, but variable sizes of deletions were found in cases that were moderately or severely affected. Disease phenotype could not be predicted from the size of the deletions. In papers II and III, a single five-generation family with autosomal dominant FMD was described. Anticipation concerning age of onset was observed. Genome scan revealed five candidate gene regions with a LOD score of &gt; 1. Two additional families with autosomal dominant MD were analyzed for linkage to these five regions. A cumulative Zmax of 3.46 was obtained for a single 463-kb region on chromosome 12p12.3, containing only one known gene: PIK3C2G. This encodes a protein with a proposed role in hair cell regeneration in mammalian ears. No mutations were found in protein-coding sequences or exon-intron borders. In two of the three families, a shared haplotype, suggested common ancestry, was found to extend over 1.7 Mb, which could be a genomic region of importance for FMD. In paper IV, a family in which five males displayed progressive low- and mid-frequency hearing impairment from the first or second decade was described. Female carriers were affected by a high-frequency hearing impairment from the fourth decade. The family could represent a novel X-linked dominant audiophenotype.

One of the salient features of the human cochlea is the incredible dynamic range it possesses—the loudest bearable sound is 10,000,000 times greater than the softest detectable sound; this is in part due to an active process. More than twelve thousand hairlike cells known as outer hair cells are believed to expand and contract in time to amplify cochlear motions. However, the cochlea’s response is more than just the sum of its parts: the local properties of outer hair cells can have unexpected consequences for the global behaviour of the system. One such consequence is the existence of otoacoustic emissions (OAEs), sounds that (sometimes spontaneously!) propagate out of the cochlea to be detected in the ear canal. In this doctoral thesis, a classical, lumped-element model is used to study the cochlea and to simulate click-evoked and spontaneous OAEs. The original parameter values describing the microscopic structures of the cochlea are re-tuned to match several key features of the cochlear response in humans. The frequency domain model is also recast in a formulation known as state space; this permits the calculation of linear instabilities given random perturbations in the cochlea which are predicted to produce spontaneous OAEs. The averaged stability results of an ensemble of randomly perturbed models have been published in [(2008) ‘Statistics of instabilities in a state space model of the human cochlea,’ J. Acoust. Soc. Am. 124(2), 1068-1079]. These findings support one of the prevailing theories of SOAE generation. Nonlinear simulations of OAEs and the model’s response to various stimuli are performed in the time domain. Features observed in the model include the saturation of the forces generated by the OHCs, compression of amplitude growth with increasing stimulus level, harmonic and intermodulation distortion, limit cycle oscillations that travel along the cochlear membranes, and the mutual suppression of nearby linear instabilities.

Source analysis of the Auditory Evoked Potential (AEP) has been used before to evaluate the maturation of the auditory system in both adult and children; in the same way, this technique could be applied to ongoing EEG recordings, in response to acoustic specific frequency stimuli, from children with cochlear implants (CI). This is done in oder to objectively assess the performance of this electronic device and the maturation of the child‟s hearing. However, these recordings are contaminated by an artifact produced by the normal operation of the CI; this artifact in particular makes the detection and analysis of AEPs much harder and generates errors in the source analysis process. The artifact can be spatially filtered using Independent Component Analysis (ICA); in this research, three different ICA algorithms were compared in order to establish the more suited algorithm to remove the CI artifact. Additionally, we show that pre-processing the EEG recording, using a temporal ICA algorithm, facilitates not only the identification of the AEP peaks but also the source analysis procedure. From results obtained in this research and limited dataset of CI vs normal recordings, it is possible to conclude that the AEPs source locations change from the inferior temporal areas in the first 2 years after implantation to the superior temporal area after three years using the CIs, close to the locations obtained in normal hearing children. It is intended that the results of this research are used as an objective technique for a general evaluation of the performance of children with CIs.

The discovery of otoacoustic emissions (OAE) has advanced our understanding of cochlear mechanics and the efferent auditory system. OAE are sounds generated within normal cochlea either spontaneously or in response to stimulation. The ability to measure OAE non-invasively, objectively and quickly makes a powerful tool to probe cochlear mechanics. Stimulation of the efferent auditory system causes changes in cochlear amplification processes and hence changes characteristics of OAE. Contralateral acoustic stimulation, commonly called OAE suppression, provides an index of the efferent auditory system (specifically, medial olivocochlear bundle) functioning. OAE is also a sensitive tool to demonstrate subtle changes in cochlear functioning caused by various pathological (e.g., noise exposure, aspirin toxicity, etc.) and non-pathological (e.g.,posture, efferent stimulation) factors. Although OAE are frequently used in both clinic and laboratory, their generation mechanism was not clearly understood until recently. It is currently accepted that distortion product otoacoustic emissions (DPOAE) are composed of two separate components, named wave- and place-fixed emissions. They not only arise from two different cochlear locations but also from two fundamentally different processes. Wave-fixed components arise from distortion sources and manifest a phase that is almost independent of frequency, where as, place-fixed components arise from reflection sources and have a phase that increases systematically with frequency. The overall aim of the work presented in this thesis was to use various OAE methods to examine cochlear function and the efferent auditory system. A related objective was to substantiate the functional relevance of the efferent auditory system in speech-in-noise perception, in order to address the clinical significance of measuring OAE suppression. Cochlear functioning was potentially manipulated by three treatments separately: one extrinsic (electromagnetic radiation exposure from mobile phone) and two intrinsic (posture and efferent activation). Potential changes in auditory function due to mobile phone exposure were evaluated in a within-subject study in a double-blind design (n=35).A comprehensive examination of the auditory system was conducted using audiometry,OAE and auditory event related potentials (ERP). The second experiment used mechanism-based DPOAE to investigate posture-induced changes in cochlear functioning (n=15). Similar DPOAE measurements were performed to evaluate the effect of contralateral acoustic stimulation on cochlear functioning (n=14). The last experiment examined the relationship between contralateral suppression of transient evoked otoacoustic emissions (TEOAE) and recognition of speech in noise (n=13). Results indicate that (i) acute exposure to mobile phone radiation does not cause any significant changes in auditory functions measured by TEOAE suppression, DPOAE or ERP (however, there were changes in auditory thresholds at 6 and 8 kHz), (ii) posture induced cochlear changes and contralateral acoustic stimulation cause significantly greater reduction in place-fixed components than wave-fixed components, and (iii) the efferent auditory system plays an anti-masking role in speech-in-noise recognition. It appears that wave- and place-fixed components are differentiallysensitive to changes in cochlear functioning. Collectively, the present results provide emerging empirical support for the need to separate the wave- and place-fixed components in DPOAE measurements. Because of inherent differences in the generation of wave- and place-fixed components, it is suggested that the separation of the components may improve the efficiency of DPOAE-based measures of cochlear dysfunction and also, of the efferent auditory system function.