Objective. 312 tinnitus sufferers were studied in order to analyze: the clinical characteristics of tinnitus; the presence of tinnitus-age correlation and tinnitus-hearing loss correlation; the impact of tinnitus on subjects' life and where possible the etiological/predisposing factors of tinnitus. Results. There is a slight predominance of males. The highest percentage of tinnitus results in the decades 61–70. Of the tinnitus sufferers, 197 (63.14%) have a hearing deficit (light hearing loss in 37.18% of cases). The hearing impairment results of sensorineural type in 74.62% and limited to the high frequencies in 58.50%. The tinnitus is referred as unilateral in 59.93%, a pure tone in 66.99% and 10 dB above the hearing threshold in 37.7%. It is limited to high frequencies in 72.10% of the patients with sensorineural hearing loss (SNHL) while the 88.37% of the patients with high-frequency SNHL have a high-pitched tinnitus . Conclusion. Hearing status and age represent the principal tinnitus related factors; there is a statistically significant association between high-pitched tinnitus and high-frequency SNHL. There is no significant correlation between tinnitus severity and tinnitus loudness confirming the possibility that neural connection involved in evoking tinnitus-related negative reactions are governed by conditioned reflexes.  Tinnitus is an auditory phantom sensation, a “perception of a sound which results exclusively from activity within the nervous system without any corresponding mechanical, vibratory activity within the cochlea” with high prevalence rates in western societies –7.Epidemiological studies showed that about one third of the population experiences tinnitus at least once in their life and about 1–5% develop serious psychosocial complications; in Italy an epidemiologic study based on questions to general population upon auditory dysfunctions evidenced a tinnitus prevalence percentage in 14.5%  . It was Persistent tinnitus may rapidly become a source of serious disturbance and handicap at psychological and socio-professional levels; in fact in 1–3% of the general population, the tinnitus affect the quality of life, involving sleep disturbance, work impairment, and psychiatric distress , 12.The presence of tinnitus progressively increases with increasing age , and this is not so much correlated with senescence itself as with the frequent concomitant hearing loss. The tinnitus prevalence in fact increases to 70–85% of the hearing-impaired population .The tinnitus is much more common with a percentage >90%. There is no single pathophysiological pathway to explain the production of subjective tinnitus. All structures of the auditory system have been suggested as possible sites of generation for tinnitus, from periphery to auditory cortex , 15.In most cases, despite appropriate medical examination, the origin of tinnitus is unknown but it is well documented in literature that tinnitus and hearing deficit are often related phenomena; either sensorineural hearing loss (SNHL) as well as noise-induced hearing loss are considered the first cause of tinnitus –18. TinnAlthough tinnitus is commonly associated with hearing loss, other aetiological factors have emerged from the widest epidemiological studies of tinnitus prevalence and actually they were considered as potential causes of tinnitus and/or cofactors. As reported by Hoffman these factors include conditions such as vascular disease, diabetes, hypertension, autoimmune disorders, and degenerative neural disorders , 19, 20.Even if in the past years there was a great consensus among the studies with respect to tinnitus sufferers, the knowledge on tinnitus onset and history, clinical presentation and audiological characteristics is still incomplete. Moreover it appears clear that there is an heterogeneity within the tinnitus population, the implication being that, as stated above, there may be a variety of aetiological mechanisms causing the tinnitus. Despite this, the doctor may not be in a situation where he can provide an aetiological link between the perception of tinnitus and one or more defined underlying pathologies. Finally, the perception of tinnitus may be linked to emotional changes, which may or may not be related to an organic pathology. All these unresolved questions underline the necessity of further investigations on tinnitus population.The aim of this study was to focus on patients suffering from tinnitus who were referred to the Audiology Section of Palermo University in order to analyze both the clinical characteristics of tinnitus symptoms and, where possible, its etiological/predisposing factors; to evaluate both the tinnitus age correlation and the tinnitus hearing loss-correlation; to quantify its impact on subjects' life.The study was conducted by the Audiology Section of the Department of Bio-technology of Palermo University on 312 subjects, 176 males and 136 females, ranging from 21 to 83 years of age, who suffered from tinnitus.In this study all consecutive tinnitus patients were included except 6 subjects with coexisting psychotic disorders ; these psychotic disorders presented the only exclusion criteria of this study because they did not permited the collaboration of patients.After the personal data, all patients underwent to a careful general medical history to identify tinnitus-related pathologies and other health diseases; it followed an ENT specialist history and an otological examination. As for the analysis of data collected, the following parameters were considered: age, sex, hearing threshold, any audio vestibular-associated diseases, tinnitus laterality , tinnitus duration , tinnitus measurements and subjective disturbance caused by tinnitus.As for age, patients were divided into six age groups: from 21 to 30, 31 to 40, 41 to 50, 51 to 60, 61 to 70, and >70.The audio/vestibular system was studied through threshold audiometry (considering the frequencies 0.5–1–2–3–4–8 kHz), tympanometry with stapedius reflex, auditory brain stem responses, nystagmography and Vestibular Evoked Miogenic Potentials. If necessary, in order to identify the cause of tinnitus, other specific exams were performed: in particular laboratory studies including hematocrit, blood chemistries, thyroid studies and lipid battery; Doppler ultrasonography (USG Doppler) evaluat blood flow disturbance in vertebral and basilar artery; middle/inner ear computer tomography study (CT) and, suspecting an acoustic neuroma or neurovascular conflict, a magnetic resonance imaging/Angio CT (MRI/Angio CT). Audiometric threshold was considered as the pure tone average for the frequencies 0.5–1–2–4–8 kHz and divided into: normal hearing (<20 dB); light hearing loss (21–40 dB); moderate hearing loss (41–70 dB); severe hearing loss (71–90 dB); profound hearing loss (>90 dB).Hearing loss was also divided into three categories: conductive hearing loss; sensorineural hearing loss  and mixed loss.The audiologic measurement of tinnitus included pitch matching and loudness matching. The pitch matching was determined asking to patient to compare tinnitus characteristics with the different tones proposed in head-phones and to choose the tone with frequency more similar to that of tinnitus; on the basis of the examined frequencies (0.125–0.25–0.5–1–2–4–8 KHz) it was able to identify three categories (Low-Middle-High pitched) considering undetermined overall cases of tinnitus with a frequency outside the range. The loudness matching was determined increasing, in head-phones, the magnitude of this tone for steps of 5 dB, until the patient declared to perceive with similar magnitude the tone and his tinnitus; this increase of magnitude of tone respect to hearing threshold was considered tinnitus loudness .To evaluate the perceived severity of tinnitus and its impact on life Tinnitus Handicap Inventory (THI - Newman e coll.) was administered. This tool is a 25-item survey that is composed of three subscales: a functional subscale (12 items), an emotional subscale (8 items) and a catastrophic response subscale (5 items) which address role and physical functioning, psychological distress, desperation and loss of control, respectively. Each item has 3 potential answers with “yes” assigned 4 points, “sometimes” 2 points, and “no” 0 points. This leads to a total score ranging from 0 indicating no tinnitus handicap and 100 the worst patients' annoyance. Classically it grades five categories of tinnitus severity: slight corresponding to a score 0–16; mild (18–36); moderate (38–56); severe (58–76); catastrophic (78–100) .χ2 test, the Pearson's correlation coefficient (Pearson's r) and Anova test were calculated using the Matlab computer programme.For the statistical analysis the 312 patients took part in the study, 176 (56.41%) males and 136 (43.59%) females, aged between 21 and 83 with a mean age of 58.37. The distribution of the decades of age and sex is indicated in In 45.83% of cases, it was not possible to correlate the tinnitus with a known etiology; tinnitus was associated with chronic noise exposure in 27 cases (8.65%), acoustic trauma in 12 cases (3.85%), sudden hearing loss in 6 cases (1.92%), middle ear pathology in 35 cases (11.22%), degenerative cochlear disease as Menière and chemotherapeutic toxicity in 26 cases (8.33%). Moreover, it was diagnosed in 47 cases a coexisting Eustachian tube dysfunction and a temporomandibular Joint Syndrome in 5 cases . As for other predisposing conditions for tinnitus, cardiovascular disease  was identified in 87 patients; and metabolic disease was found in 23 cases.The hearing loss accompanies tinnitus symptoms in 63.14% of patients; the hearing loss was identified as conductive hearing loss in 10.66% of cases, as sensorineural hearing loss in 74.62% of cases and mixed type in 14.72% of cases.Of 147 patients with sensorineural hearing loss in 58.50% the hearing loss was limited to high frequencies; in the 11.56% to low frequencies; in 29.93% to a flat loss.As for audiometric threshold, it was revealed: normal hearing in 36.86% of cases; light hearing loss in 37.18% of cases, moderate hearing loss in 14.1% of cases, severe hearing loss in 9.93% of cases and profound hearing loss in 1.92% of cases . The mea Tinnitus was referred as unilateral in 187 patients (59.93%), 83 cases in the right ear and 104 cases in the left ear; bilateral in 74 patients (23.71%), and “in the head” in 51 cases (16.35%).As for tinnitus duration, it was presented in acute form in 95 patients corresponding to 30.45%, subacute in 133 cases (42.63%) and chronic in 84 patients (26.92%).In most cases, patients reported their tinnitus as a pure tone in 66.99% (209 cases); in relation to narrow-band in 27.88% (87 cases); at last, the tinnitus is not identifiable in 5.13% of cases (16 patients).The tinnitus frequency, measured by the pitch matching test, was matched to high frequencies  in 172 patients (55.13%), to middle frequencies  in 56 patients (17.95%), to low frequencies  in 68 patients (21.79%) while it was variable and not identifiable in 16 patients (5.13%).The 72.10% of the patients with sensorineural hearing loss had a high-pitched tinnitus while the 88.37% of the patients with high-frequency sensorineural hearing loss had a high-pitched tinnitus .The loudness matches for tinnitus, as estimated by audiometry, was 0–5 dB above the hearing threshold in 14.1% of cases; 5 dB above the hearing threshold in 28.21% of cases; 10 dB above the hearing threshold in 37.8% of cases; 15 dB above the hearing threshold in 14.1% of cases; above 15 dB in 1.28% of cases; in 4.48% it was no able to identify the tinnitus loudness .The tinnitus began suddenly in 41.02% (128 patients), increased in loudness overtime in 53.85% (168 cases), decreased in 8.33%, and was unchanged in 37.82% of participants (118 cases).Regarding tinnitus annoyance and its impact on the quality of life, the THI identified five categories of tinnitus severity: slight grade was found in 59 patients (18.91%); mild grade in 102 patients (32.69%); moderate grade in 79 patients (25.32%); severe grade in 52 patients (16.67%) and catastrophic grade in 20 patients (6.41%).P = .39), usually there is a slight predominance of males respect to females; both Johansson and Arlinger (2003), Fabijanska et al. (1999) and Palmer et al. (2002) reported a minimally higher, but not statistically significant, prevalence overall for male than for female [Tinnitus is the general term for sound perception  that cannot be attributed to an external sound source. It has been reported that the condition affects from 5% to 32% of the world population, and in 25% of these cases, it is sufficiently severe to cause them to seek medical help , 17, 18.r female , 23. Thir female . The epir female , 22, 23.Tinnitus is a multifactorial symptom, which can be induced by all types of hearing loss as well as by somatic and psychiatric disorders and pharmaceutical drugs. Axelsson (1991) described different diseases which were accompanied by tinnitus and the main causes leading to tinnitus were suggested. He was able to identify the aetiology of tinnitus in 81% of tinnitus tested cases . In 26% de facto a causal relationship between hearing loss and tinnitus but the value percentage of subjects with tinnitus associated with hearing loss is very variable ranging from 30% to 85% of cases [Revised tinnitus literature seems to assume of cases , 13. Thiof cases , 22, 27.of cases , 28.When present, the sensorineural hearing loss was found, in most cases, limited to the high frequencies (58.50%) and of light-moderate degree (51.28%). These findings were also referred by Savastano, Satar et al. and Henry et al. , 29, 30;χ2 = 66.1230, df = 6, P < .001).As for tinnitus characteristics it is accepted that there is a statistically significant association between high-pitched tinnitus and high-frequency SNHL; when subjects match their tinnitus pitch to a pure tone, most of the matches are at frequencies at which hearing is impaired –33. The More frequently, the subjective judgment of loudness matches for tinnitus was 10 dB above the hearing threshold (37.8%); this value suggests that most patients seek for specialist examination when the symptom is already disturbing , 12. r = 0.8712, P = .0544)  . The regP = .364 . It may support the actual theory that the patient's reaction to tinnitus cannot be classified as a simple function of its psychoacoustic aspects but rather as a complex interaction between acoustic phantom symptoms, somatic attention and depressive symptoms.Tinnitus is a distressing symptom, provoking an important decrease in the quality of life in 20% of tinnitus sufferers, that is strongly correlated to many factors. This paper, according to literature data, suggests that the hearing status and the old age represent the principal tinnitus-related factors; in most cases tinnitus is high-pitched and it is associated to high-frequency hearing loss with a significant statistical correlation between the two variables. The analysis of relationship between tinnitus severity (expressed as THI scores) and tinnitus loudness match clearly demonstrate lack of correlation between these variables. This finding support the possibility that neural connection involved in evoking tinnitus-related negative reactions is governed by conditioned reflexes.


Hearing impairment is the most common body system disability in veterans. In 2008, nearly 520,000 veterans had a disability for hearing loss through the Department of Veterans Affairs (VA). Changes in eligibility for hearing aid services, along with the aging population, contributed to a greater than 300% increase in the number of hearing aids dispensed from 1996 to 2006. In 2006, the VA committed to having no wait times for patient visits while providing quality clinically-appropriate care. One approach to achieving this goal is the use of group visits as an alternative to individual visits. We sought to determine: 1) if group hearing aid fitting and follow-up visits were at least as effective as individual visits, and 2) whether group visits lead to cost savings through the six month period after the hearing aid fitting. We describe the rationale, design, and characteristics of the baseline cohort of the first randomized clinical trial to study the impact of group versus individual hearing aid fitting and follow-up visits.Participants were recruited from the VA Puget Sound Health Care System Audiology Clinic. Eligible patients had no previous hearing aid use and monaural or binaural air-conduction hearing aids were ordered at the evaluation visit. Participants were randomized to receive the hearing aid fitting and the hearing aid follow-up in an individual or group visit. The primary outcomes were hearing-related function, measured with the first module of the Effectiveness of Aural Rehabilitation (Inner EAR), and hearing aid adherence. We tracked the total cost of planned and unplanned audiology visits over the 6-month interval after the hearing aid fitting.A cohort of 659 participants was randomized to receive group or individual hearing aid fitting and follow-up visits. Baseline demographic and self-reported health status and hearing-related measures were evenly distributed across the treatment arms.Outcomes after the 6-month follow-up period are needed to determine if group visits were as least as good as those for individual visits and will be reported in subsequent publication.NCT00260663 In 2008, nearly 520,000 veterans had a disability for hearing loss through the Department of Veterans Affairs (VA), making hearing impairment the most common body system disability in veterans . ChangesFurther, in 2006 the VA committed to having no wait times for patient visits while providing quality clinically-appropriate care (VHA Directive 2006-028). Implementation of Systems Redesign (SR) tools were recommended to help accomplish this goal. One of the SR principles aimed at reducing demand is the use of group visits as an alternative to individual visits. For new hearing aid users, the initial orientation and follow-up sessions may be well suited to a group format because they consist of standardized teaching elements and discussion topics relevant to all participants. While group visit are a promising approach, it is important to know that they provide at least equivalent outcomes as individual visits before recommending widespread use of group visits.Most of the research in audiology group visits has focused on group aural rehabilitation as a supplement to the standard one-on-one visit -18, and We used a non-inferiority randomized clinical trial with a factorial design to determine: 1) if group visits were at least as effective as individual visits ('non-inferior') for two types of audiology visits (hearing aid fitting and hearing aid follow-up), as measured by hearing-related function and hearing aid adherence six months after hearing aid fitting, and 2) whether group visits led to cost savings through a six month acclimatization period after the hearing aid fitting.We hypothesized that when compared to individual visits, group hearing aid fitting and follow-up visits would yield: 1) equivalent or improved hearing-related function (primary outcome) and equivalent or improved hearing aid adherence (secondary outcome), and 2) group visits would lead to cost savings from reduced audiology person-hours for initial rehabilitation, and equivalent or improved (lower) rates of unplanned visits in the six-month period after fitting.New hearing aid participants were randomly assigned to a group or individual hearing aid fitting, and to a group or individual hearing aid follow-up Figure . TherefoStudy participants were recruited from the Audiology Clinics at the Seattle and American Lake Divisions of the Puget Sound Health Care System from February 2006 through October 2007. Participants were eligible to enroll if they met the following criteria:• No previous hearing aid use• Monaural or binaural air-conduction hearing aids were fit as a result of findings at the hearing aid evaluation visit. Under the Veterans' Health Care Eligibility Reform Act of 1996 (Public Law 104-262) and the Veterans Health Administration Directive 96-069 of 1997, participants were eligible for VA hearing aids if they had a 10-100% disability rating for any medical condition, any disability rating for a hearing related condition, receiving aid and attendance or housebound benefits from the VA, former Prisoner of War, awarded a Purple Heart, receipt of benefits under Title 38, hearing loss resulting from another medical condition for which the veteran was receiving on-going VA care or that resulted from treatment of that condition, or hearing loss severe enough to interfere with the ability to participate actively in their own medical careWe excluded participants who met the following criteria:• previous hearing aid use• unable to participate in group instruction because of cognitive or physical restrictions• unable to give informed consent• unable to complete self-administered questionnaires written in simple English• unwilling or unable to return for follow-up visit or complete follow-up questionnairesAll participants being seen for a hearing aid evaluation were given a study flyer when they checked in for their hearing evaluation visit. The flyer briefly described the study and eligibility criteria. At the conclusion of the evaluation, participants who met the inclusion criteria were offered enrollment by the audiologist who conducted the evaluation. Participants who expressed interest were directed to an on-site research assistant (RA). The RA explained the study in detail in a quiet, private room and conducted the process of informed consent. Participants were offered $50  as remuneration for the time and effort of participating. Remuneration was made with one payment six months after the hearing aid fitting.Participants were randomized to group or individual hearing aid fitting and to group or individual hearing aid follow-up. Randomization assignments were prepared ahead of time in variable blocks of four or eight participants, and then placed in sealed, opaque envelopes. Thus, RAs did not know which treatment participants would receive. After randomization, participants were scheduled for the hearing aid fitting and follow-up appointments according to their randomization assignments. Participants returned for the fitting three-to-four weeks after the evaluation. The follow-up appointment was scheduled for three-to-five weeks after the fitting.After enrollment, baseline co-variates were collected and baseline assessment surveys were administered. Participants who did not have time to complete the surveys that day completed them at home and returned them in a postage-paid envelope prior to the hearing aid fitting.higher scores representing better function. The Inner EAR has an additional global question scored on a 0-10 scale. The second module (Outer EAR) covers hearing-aid related issues such as comfort, appearance, and convenience, is relevant only after hearing aid fitting and therefore was not administered at the baseline visit. Both modules have excellent psychometric reliability, validity, and responsiveness.• The Effectiveness of Auditory Rehabilitation (EAR)  measureshigher scores representing more handicap.• Hearing-related handicap was assessed with the Hearing Handicap Inventory for the Elderly (HHIE) -27 questHigher scores indicate better use of communication strategies.• The Communication Strategies scales of the Communication Profile for the Hearing Impaired (CPHI) -31 was uHigher scores indicate more co-morbitity.• Co-morbidity is a potentially confounding variable because those with greater disease burden tend to have worse health-related quality of life. We used the Seattle Index of Co-morbidity (SIC)  which isHigher scores indicate poorer health.• Health status, like co-morbidity, may be a confounding variable because of its independent effect on quality of life. We assessed general health status with the SF-12 . This sc• The Patient Health Questionnaire (PHQ)  was used• Demographic data. The VA Computerized Patient Record System (CPRS) was used to obtain age, service-connection rating, and gender.• Audiometric data obtained at the hearing aid evaluation, such as type and degree of hearing loss were also abstracted from CPRS.The first randomized intervention was the orientation portion of the hearing aid fitting appointment. All participants, regardless of randomization, received their aids during an individual 45-minute session at which time the aids were programmed for the participant's specific requirements. Hearing aid amplification characteristics were adjusted to match NAL-NL1  hearing For the group fitting, up to three participants received individual programming during one 45-minute period. They then waited while a second set of up to three participants received programming during a second 45-minute session. Then, these six participants and their guests convened for a 60-minute group orientation in a quiet conference room in or near the audiology clinic. The orientation was conducted by a hearing aid technician or audiologist. Teaching topics included: how a hearing aid works; use, care, and maintenance; acoustic feedback; battery use, replacement, and ordering; telephone use; warranty and repairs; listening and communication strategies; and realistic expectations. Functions of special features covered during the individual portion of the visit were reviewed. The format was standardized and facilitated by a PowerPoint presentation combined with hands-on practice. Participants also received a handout of the PowerPoint presentation. An example of the timing of a typical group hearing aid fitting is shown in Table This orientation was provided to the participant by the same audiologist who performed the programming, immediately following and in the same room as the programming. This session typically took 30 minutes. The same topics were covered as in the group orientation, without the aid of the PowerPoint presentation; however, a printout of the PowerPoint presentation was provided to these participants. Therefore, participants in both the individual and group randomization arms received the same information. Nonetheless, because of the individualized nature of the session, some information may have varied from participant to participant, as well as from audiologist to audiologist, depending on the participant's specific needs and requests. An example of the timing of a typical individual hearing aid fitting is shown in Table The second randomized intervention was the follow-up visit that was scheduled for three to five weeks after the fitting, depending on appointment availability.This visit was scheduled with up to five participants during a 75-minute sessions, 45 minutes of which reviewed information covered at the hearing aid orientation, and covered the effects of acquired hearing loss, coping with hearing loss, pinpointing the source of communication problems, realistic expectations, management of difficult communication situations, guidelines for hard of hearing, and guidelines for significant others. These visits were conducted by one of the staff audiologists in the same quiet conference room in or near the audiology clinic as the group orientation visits. This session was facilitated by a PowerPoint presentation, and participants were provided with a handout of the presentation. Individual adjustments, if needed, were conducted during the 30 minutes after the group portion. In this scenario, 75 minutes of audiologist time was used to see up to five participants  questionnaire measures hearing aid adherence with both dichotomous ("Do you use your hearing aids?") and continuous responses ("How many hours a day do you use your hearing aid?"). Both types of responses have good construct validity with hearing related function and hearing handicap ,38.7 representing maximum satisfaction. Construct and internal validity of the SADL are strong [• The Satisfaction with Amplification in Daily Life (SADL) ,40 is a higher scores reflecting better outcomes.• The International Outcome Inventory for Hearing Aids (IOI-HA) is a ge• Transitional measure of hearing-related quality of life (QoL). Transitional outcome measures have been shown to be more sensitive to change than serial measures taken over several points in time . This siImmediately after the fitting visit and after the follow-up visit, an RA distributed a session evaluation and knowledge retention questionnaire. The session evaluation questionnaire consisted of 11 questions pertinent to all participants regarding various mechanisms that might mediate the effectiveness of rehabilitation such as presence of family members, amount of attention and time spent with providers, session pace and information repetition, opportunities for questions, and discomfort in social situations. Four additional questions that were directed only to participants who had a group visit asked about privacy concerns, effectiveness of the PowerPoint slides, group interactions and shared insights. The knowledge retention questionnaire asked 12 questions about standard information that should have been learned in the session such as what color indicates the right and left hearing aids. The RA also collected data about the session characteristics such as how many participants and or spouses were present, length of session, wait time between programming and orientation, etc. Six months after the fitting, features of the hearing aid(s)  was obtained from CPRS chart review.Six months after the fitting visit, we mailed the final set of outcomes questionnaires to participants. These were returned by postage-paid mail. The packet consisted of the SF-12, PHQ, EAR (Inner and Outer), HHIE, CPHI, AdHeRe, SADL, IOI-HA, and transitional QoL measure. If items were left blank, an RA attempted to obtain the information via telephone.We chose the 6-month interval after the hearing aid fitting to track utilization and cost variables because pilot data indicated that 75% of all unplanned visits occurred in this timeframe. Total costs will be the sum of costs of planned (fitting and follow-up) and unplanned  audiology visits. We estimated the cost of fitting and follow-up visits based on provider time and pay grade. For unplanned visits, we estimated the cost of the visit using the cost for each CPT code associated with the visit from the VA cost accounting system. All cost estimates included fringe benefits and indirect and overhead costs.The study and all study modifications were approved by the Institutional Review Board of the University of Washington and the Research and Development Committee at VA Puget Sound Health Care System. RAs received thorough training in the process of informed consent, and their first enrollment interview received full edit reviews. Ten percent of all participant files were randomly audited for errors by the project manager. Data were entered into a Microsoft Access database using double-data entry verification.The cohort description and baseline characteristics are described in this report using straightforward descriptive statistics. Future study results will use an intent-to-treat analysis. The primary purpose of this trial was to investigate the relative effectiveness of group and individual appointments in audiology. We used a factorial design with concurrent testing of group fittings and group follow-ups to facilitate insight into the value of a group format for both visits with one trial. The primary outcome of effectiveness was hearing-related function (Inner EAR) 6 months after the hearing aid fitting.For our primary analysis, we will evaluate the overall effectiveness of group versus individual hearing aid fitting and group versus individual hearing aid follow-up for the Inner EAR using multivariate mixed model regression to adjust for factors related to hearing-related function and account for potential clustering among participants of a given group visit. Because our trial was a non-inferiority study, we will test the one-sided hypothesis that the effect of group visits does not lead to a detrimental effect beyond the clinically significant range. Secondary analyses will focus on secondary outcomes, and also on subgroup analyses for the primary outcome variable. Secondary outcomes will include hearing aid adherence, hearing handicap, communication strategies, and satisfaction with amplification. Subgroup analyses will focus on principal covariates such as use of monaural versus bilateral aids, sensorineural versus mixed hearing loss, degree of hearing loss, and the presence of specific hearing aid features such as directional microphones and telephone coils. The study was adequately powered to detect large differential treatment effects, but analyses of smaller effects will be exploratory rather than confirmatory.Economic evaluation will estimate potential cost savings from the VA's perspective for group visits compared to individual visits. We will examine the effect of the group visit intervention on the number of unplanned visits and cost of unplanned visits. Although this study was not powered to definitively determine cost-effectiveness, exploratory calculations of the unit cost to obtain a successfully treated patient  will be made. We will also conduct analyses with a secondary measure of treatment success: adherence to hearing aid use .We estimated our sample size based on a minimum clinically significant difference in Inner EAR scores of 6.0 points on a 100 point scale with standard deviation of 23.5, and a conservative loss to follow-up of 20%. We controlled for possible provider effects, site, and treatment, and accounted for most sources that induce clustering, and thus anticipate weak (<.10) within-group correlation. Enrollment of 660 participants (330/arm) would yield 90% power  to reject the one-sided null hypothesis. An important consideration was to ensure that we had adequate power to detect changes in secondary outcomes, such as decreases in dichotomous hearing aid adherence. Using a sample size of 264 analyzable participants in each arm and our baseline adherence rate of 95%, we will have 92% power to ensure that hearing aid adherence rates do not worsen more than 10% (relative percentage).Our study design was a factorial trial evaluating two interventions. This has the benefit of research economy and the ability to test for non-additive effects for the two interventions. For our research questions we chose not to make a multiple comparison adjustment since we are prepared to make separate conclusions for each of the two interventions. A multiple comparison correction is appropriate whenever a single conclusion is derived from the evaluation of multiple tests. We will make separate conclusions regarding group versus individual hearing fitting and regarding group versus individual follow-up.A cohort of 659 participants was enrolled from February 2006 through October 2007. Eleven participants requested to be dropped from the study, one died prior to the fitting, and three others were later determined to be ineligible, leaving 644 participants. Of these, 323 and 321 participants were randomized to individual and group fittings, respectively, and 324 and 320 participants randomized to individual and group follow up, respectively. This resulted in four different visit combinations . Our study was designed to detect differences in hearing-related function and cost throughout the six-month acclimatization period following the hearing aid fitting for each of these visit types. We randomized 659 eligible participants from February 2006 through September 2007 at the Seattle and American Lake Divisions of VA Puget Sound Health Care System to receive group or individual hearing aid fitting and group or individual hearing aid follow-up. At baseline, all measures for 644 active participants were evenly distributed across the visit combinations and representative of an older male hearing-impaired veteran population seeking their first pair of hearing aids.Data from randomized trials represent the highest level of evidence about the effectiveness of interventions . To our In addition, factors that might mediate the effectiveness of group sessions will also be considered ,19,48-60Our study was designed to determine whether positive or negative effects will predominate on average. If positive effects predominate, or uncertain effects turn out to be positive, we would expect to see better treatment effectiveness . Conversely, if negative effects predominate, or the bulk of the uncertain effects are negative, we will see worse treatment effectiveness. By gathering data about the potential mediators, we can examine whether these effects exist and how these effects may affect outcomes, giving us preliminary insight into the mechanisms by which group visits may or may not be effective.This is the first randomized trial to consider the effects on cost from providing group hearing aid visits. Costs from implementation of group visits can be incurred immediately and on a longer-term basis. Immediate cost savings can be realized with the reduced audiology person-hours required during group fitting and follow-up visits. However, the impact on longer-term costs over the rehabilitation period is less clear. If group treatment is effective, there may be more learning and information retention that leads to long-term savings from fewer subsequent unplanned visits and hearing aid repairs. On the other hand, if group sessions are less effective and participants do not learn how to use their aids optimally, the result may be more unplanned visits and more hearing aid repairs. It is also entirely possible that mixed effects will be observed: for example, improved effectiveness at greater cost. The desirable finding that effectiveness is either improved or equivalent (non-inferior) while saving costs was the hypothesis for this study.Based on prior studies, we expect that group hearing aid visits will yield equivalent or better outcomes compared to individual visits, but with lower costs associated with group visits as a result of fewer providers required to provide the care. Similar findings have been reported for group visits in chronic disease management. With the use of group visits, a number of large managed health care organizations have found lower costs ,61,62 anPrior to the HEARING trial, no randomized trials have compared group versus individual hearing aid visits; however, three non-randomized studies suggest that group visits may yield better participant outcomes than individual visits -21. CollA number of factors may have influenced how participants in this study responded to group visits. In clinical settings, patients often wait many years from first noticing a hearing loss until they first visit an audiologist for evaluation. However, veterans receive hearing aids free of charge so their threshold for seeking their first pair of hearing aids may be lower that that found in private clinics, possibly yielding new VA hearing aid users with less hearing loss than new private sector users. Degree of impairment may differentially affect receptiveness to group intervention. In addition, receiving hearing aids free of charge may change an individual's motivation and willingness to participate in a group format. For example, if the participant preferred an individual visit, they might be more likely to accept a group visit instead if they received the aids at no charge. In addition, only veterans who were eligible to receive VA hearing aids were enrolled in this study. This means that VA hearing aid participants have sustained some type of service-related disability, and/or have a substantial enough hearing loss to prevent them from participating in their medical care. Because of their military service experiences and/or the severity of their hearing loss, these veterans may respond differently to a group setting. Given these factors, our results may not be generalizable to a non-veteran population.In addition, it was not possible to blind participants or providers to treatment allocation, possibly biasing participants' perception of their treatment depending on preconceived ideas about group visits. The extent to which group hearing aid visits yield positive outcomes will depend on how cost and effectiveness interact. These data will be reported in a subsequent publication.AdHeRe: Adherence to Hearing Rehabilitation; CPHI: Communication Profile for the Hearing Impaired; CPRS: VA Computerized Patient Record System; DSM-IV: Diagnostic and Statistical Manual of Mental Disorders; EAR: Effectiveness of Aural Rehabilitation; G-G: group fitting and group follow-up; G-I: group fitting and individual follow-up; HEARING: Hearing Aid Effectiveness after Aural Rehabilitation-Individual versus Group; HHIE: Hearing Handicap Inventory for the Elderly; I-G: individual fitting and group follow-up; I-I: individual fitting and individual follow-up; IOI-HA: International Outcome Inventory for Hearing Aids; MCS: Mental Component Score; NAL-NL1: National Acoustic Laboratories-non-linear; PCS: Physical Component Score; PHQ: Patient Health Questionnaire; QoL: Quality of Life; RA: Research Assistant; SADL: Satisfaction with Amplification in Daily Life; SAI-WHAT: Screening for Auditory Impairment: Which Hearing Assessment Test; SIC: Seattle Index of Co-morbidity; SR: Systems Redesign; VA: Veteran's Affairs; VHA: Veterans Health Administration.The authors declare that they have no competing interests.MC participated in the study design and coordination, statistical analysis, and drafted the manuscript. PS participated in the study design and manuscript preparation. CL participated in the study design, cost analysis, and manuscript preparation. PH and DA participated in the study design. BY participated in the study design, coordination, and statistical analysis. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6963/9/233/prepub


Music is ever present in our daily lives, establishing a link between humans and the arts through the senses and pleasure. Sound technicians are the link between musicians and audiences or consumers. Recently, general concern has arisen regarding occurrences of hearing loss induced by noise from excessively amplified sound-producing activities within leisure and professional environments. Sound technicians' activities expose them to the risk of hearing loss, and consequently put at risk their quality of life, the quality of the musical product and consumers' hearing. The aim of this study was to measure the prevalence of high frequency hearing loss consistent with noise exposure among sound technicians in Brazil and compare this with a control group without occupational noise exposure.This was a cross-sectional study comparing 177 participants in two groups: 82 sound technicians and 95 controls (non-sound technicians). A questionnaire on music listening habits and associated complaints was applied, and data were gathered regarding the professionals' numbers of working hours per day and both groups' hearing complaint and presence of tinnitus. The participants' ear canals were visually inspected using an otoscope. Hearing assessments were performed  using a portable digital AD 229 E audiometer funded by FAPESP.There was no statistically significant difference between the sound technicians and controls regarding age and gender. Thus, the study sample was homogenous and would be unlikely to lead to bias in the results. A statistically significant difference in hearing loss was observed between the groups: 50% among the sound technicians and 10.5% among the controls. The difference could be addressed to high sound levels.The sound technicians presented a higher prevalence of high frequency hearing loss consistent with noise exposure than did the general population, although the possibility of residual confounding due to unmeasured factors such as socioeconomic status cannot be ruled out. Music is ever present in our daily lives, establishing a link between human beings and the arts through the senses and pleasure. However, if we use music inappropriately, that is, if we make abusive use of the duration, intensity and/or frequency of exposure to music, the noise may have an adverse impact on our hearing function.High frequency of hearing loss consistent with noise exposure , excluding cases of acute acoustic trauma, is characterized by the gradual and progressive loss of hearing acuity. NIHL occupies the second place among the most common hearing illnesses, only surpassed by presbyacusis ,2.In Brazil, noise has only been recognized as an ambient pollutant since March 1991 . EpidemiIt has been estimated that 25% of the worker population exposed to noise have some degree of NIHL. In spite of being the most frequent type of damage to workers' health, the prevalence of NIHL among Brazilian workers is still unknown .In the United States, approximately ten million workers have NIHL > 25 decibels (dB) .According to data from the World Health Organization (WHO), approximately 16% of the world's population endures NIHL caused by occupational exposure to noise .NIHL is generally sensorineural, bilateral, symmetrical and irreversible. Individuals with this illness are likely to present a prolonged history of exposure to high noise levels (> 85 dB(A) for eight hours a day), sufficient to have caused a loss in the levels and configurations observed in audiological tests. Hearing loss consistent with noise exposure is likely to have gradually developed over a period of between 5 and 20 years of general exposure in all locations ,8.Recently, there has been rising concern about the prevalence of NIHL caused by excessively amplified sound activities in leisure settings. Among these activities, the exposure of musicians and sound technicians  who are exposed to intense and long lasting music have been highlighted.With regard to sound technicians who begin their careers early in life and consequently are frequently exposed to high-intensity sounds, it needs to be asked how the hearing levels of these individuals are holding up, when music is the medium through which they perform their professional activities.Sound technicians are the agents who transport to consumers one of Brazil's best products: Brazilian popular music. Dealing with and watching over what this process entails is of great social relevance to the country's health, culture and economy.Moreover, decreases in hearing function worsen individuals' quality of life, communication abilities, educational achievements and enjoyment of leisure activities, and have enduring economic consequences .Thus, the present study had the aim of gauging the prevalence of high prevalence hearing loss consistent with noise exposure among sound technicians and comparing this with a control group without occupational noise exposure.This study was approved by the Ethics Committee of the Federal University of São Paulo. It was a cross-sectional comparative study between two groups: sound technicians and individuals without occupational noise exposure (controls). The sample consisted of 177 participants: 82 people working with sound systems and a sample of 95 members of the general population (control group). The inclusion criterion was that participants needed to be adults > 19 years old.The sound technician group included professionals involved in sound mixing, audio equipment operation, video material editing, video equipment operation and microphone operation, working in closed or open spaces, with at least five years of exposure to sound-producing activities ,8.The control group consisted of individuals without a history of exposure to continuous noise, i.e. individuals who were rarely exposed to music and whose work activities did not involve music. Workers exposed to high levels of industrial noise, such as factory workers, bus drivers and the like, were also excluded from this group.The following were exclusion criteria for both the sound technician and control groups: use of individual sound amplification devices (hearing aids); diagnosis of congenital hearing deficiency (genetic or acquired); reduced external acoustic ear canal; and presence of excess earwax or otorrhea.Using the criteria outlined above, we excluded 19 subjects from the sound technician group because 11 of them had been exposed to sound-producing activities for less than five years and eight subjects were less than 19 years old. In the control group, we excluded seven subjects: six because they did not fulfill the inclusion criterion regarding age and one because of the presence of excess earwax.The control group was selected by invitation. The participants in this group were health professionals, students and employees at the Federal University of São Paulo, Brazil, who were unaware of the possible presence of hearing loss.The evaluations and questionnaires were applied in the Brazilian Cochrane Center and in the studios of the television channels Record TV and Cultura TV, in São Paulo, Brazil. Portable acoustic booths were used when conducting the audiometric examinations. Rooms were deemed to be silent room when the noise level was between 55 and 60 dB, in accordance with Administrative Technical Standard no. 3214 of the Ministry of Labor and the Workplace Safety Office .The sound pressure levels in the examination locations were measured using devices that conformed to the standard ANSI S1.4-1983. These made it possible to check whether the environment within which the booth had been set up was appropriate for conducting audiometric assessments.The sound pressure levels in the places where the audiometric assessments were carried out were measured using the TES 1350A sound level meter. In the Record TV studio, the device measured a minimum of 57 dB(A). In the Cultura TV studio and in the Brazilian Cochrane Center, the estimated minima were measured as 41.6 dB(A) and 57.8 dB(A), respectively. All three locations where acoustic measurements were made were within the above mentioned standard set by the Ministry of Labor and the Workplace Safety Office .Individuals who presented a diagnosis of hearing abnormalities during the examination (at any of the hearing frequencies assessed), or a diagnosis of obstruction, were referred to the outpatient clinic of the Speech-Language-Hearing Service of Hospital São Paulo, the Federal University of São Paulo, Brazil, for detailed evaluation and further procedures as required.We investigated the participants' hearing levels at the following frequencies: 250 Hz, 500 Hz, 1000 Hz, 2000 Hz, 3000 Hz, 4000 Hz, 6000 Hz and 8000 Hz. We also performed speech audiometry .The subjects were considered to present high frequency hearing loss consistent with noise exposure if their audiograms at the frequencies of 3000 and/or 4000 and/or 6000 Hz presented hearing thresholds that were at least 25 dB higher than the thresholds for the other tested frequencies , when this compromised hearing was observed in both the air test and the bone test, in one or both ears [The sound technicians were asked to take a 14-hour hearing rest prior to the test, in accordance with Administrative Technical Standard no. 19 of the Ministry of Labor and the Workplace Safety Office, dated April 22,1998 .A basic tinnitus assessment was performed, including an audiological evaluation and a self-reported assessment. We also defined tinnitus as a ringing or other type of noise generated within the head .The audiological evaluation was performed using the portable AD229 E digital audiometer, calibrated according to the standard ANSI S3.6 (1989) ,13,14.PE) and the control group (PPNE), pairs of ratios were compared by means of the equation proposed by Pocock (1983) [To determine the sample size to be used for comparing hearing loss prevalences between the sound technician group (Pk (1983) :α Z+ βZ, such that Zα and Zβ were respectively the confidence scores linked to the confidence intervals (CI) (1-α) × 100% and (1-β) × 100%.Where Θ = PE = 0.39 and PPNE = 0.9. In the present study, it was intended to carry out the comparison at a significance level of α = 5% with an elasticity trait of 1-β of 95% and a two-tailed hypothesis test. Therefore, the equation obtained was Θ = α Z+ β Z= 1.65 + 2.94, thereby recommending the use of at least 50 participants in each group.The prevalences of hearing loss among a group of percussionists and in the general population were estimated by Cunningham (2006) , respectIn the inferential portion of the study, we used Student's t test for homogenous independent samples, whenever the comparison variable (outcome variable) was continuous, presented normal or near-normal distribution  and presented equal variances . To compare ratios, we used the chi-square test or Fisher's exact test, in accordance with the conditions established by Cochran (1954) . The tabThe minimum sound pressure levels for each of the sound technicians' workplace environments were measured in the studios of the television channel Cultura TV, in order to exemplify how much noise the workers were exposed to. In the audio operators' studio, 101 dB(A) was detected. In the sound mixing work area, videotape operators' work area and microphone operators' work area, the measurements were respectively 91.1 dB(A), 90.7 dB(A), and 94.2 dB(A). In the videotape editing and sound monitors' areas, the measurements were 89.1 dB(A) and 86.6 dB(A), respectively.Table Table Table Workers exposed to noise levels of 85 dB have a permissible maximum daily average exposure of only eight hours and workers exposed to noise levels of 103 dB(A) have a permissible maximum daily average exposure of only 7.5 minutes . HoweverWe considered that individuals whose audiograms presented thresholds of less than or equal to 25 dB at all the frequencies examined presented normal hearing, in accordance with the standards of the Workplace SafetyOffice.According to Santos & Morata 1999) , factors999 , facThere was a statistically significant difference (p = 0.009) with regard to the presence of hearing complaints between the individuals in the sound technician and control groups Table . Thus, tThere were no statistically significant differences in music listening habits and ototoxicity exposure between the groups studied (p > 0.05) Table . HoweverTinnitus is a symptom that may be found in workers suffering from high frequency hearing loss consistent with noise exposure and, according to Ferreira Júnior 1998) , such wo998 , sucOne example of this situation of noise exposure over a long period of time is provided by the British guitar player and singer Eric Clapton (age 62 years), who developed tinnitus after spending years of his career playing in front of sound amplifiers at high volumes. He has said that, if he could go back in time, he would ask to have lower amplifier volume during his concerts. He has reported that he constantly hears "whistles" in his ears . AnotherThe main outcome of the present study was the presence of hearing loss. With regard to this, there was a statistically significant difference (p < 0.001) between the sound technicians and controls  Table , thus reThe effect of music on sound technicians' hearing was found to be greater in the present study than in other studies such as Cunningham (2006) . This waOther study limitations might include residual confounding due to differences in socioeconomic status between the two study groups. This could partly explain the presence of much higher hearing loss levels among the sound technicians.There is no effective treatment for permanent hearing loss resulting from excessive exposure to noise levels. However, this condition may generally be prevented through combined measures in programs against hearing loss . Even though the efficacy of these measures  for noise protection is determined by tests performed in laboratories, their effectiveness in preventing hearing loss due to exposure to cumulative noise depends mainly upon how regularly the workers use this protection. Studies have been demonstrating that if workers do not use hearing protectors for 100% of their working hours, the effectiveness of such devices will diminish rapidly. For example, using hearing protectors for only 90% of the time will diminish their effectiveness to less than one third of their efficacy .85.4% of the participants in the sound technician group reported not using hearing protectors during their long working hours Table . This seA systematic review has compared the effectiveness of intervention programs versus no intervention regarding the use of hearing protectors among workers exposed to noise levels above 85 dB(A). It was concluded that there was very little evidence regarding the efficacy of long-term school programs for increasing the use of hearing protectors among school students. Furthermore, the available evidence did not show that provision of customized/standardized information to motivate the workers to use hearing protectors was more effective than general information ,41.Programs for preventing hearing loss in industrial organizations have been widely defended. Occupational safety and health legislation obliges employees to have preventive measures in most countries . In EuroStudies in the USA have indicated that the use of hearing protectors is increasing, but there is still much to be improved in this respect . Many faImplementation of hearing conservation programs within this population is of fundamental importance not only for the well being of these individuals, but also for the enrichment of Brazilian popular music, given that it is one of the best products of our country. The perception of hearing is related to the hearing threshold and maintenance of good perception through hearing protection measures will have the consequence of developing better professionals who can work to provide even greater value for Brazilian musicians.For this to be definitively recognized and introduced within Brazilian culture and internationally, it will continue to be highly important, as a public health measure, to develop new strategies towards increasing the use of hearing protectors.(1) Sound technicians presented a higher prevalence of high frequency hearing loss consistent with noise exposure than did the general population, although the possibility of residual confounding by unmeasured factors such as socioeconomic status cannot be ruled out.(2) There is some urgency with regard to carrying out studies aiming towards implementing a hearing conservation program through reducing sound levels and making sound technicians aware of the importance of using hearing protectors and adopting preventive actions.The authors declare that they have no competing interests.Conception and design: RED. Coordination of the study: VMT and EKS. Data acquisition: RED. Data analysis and interpretation: RED and JFM. Drafting the manuscript: RED. Critical revision for important intellectual content: RED. Data interpretation: RED and JFM.The pre-publication history for this paper can be accessed here:


Here we describe a general technique for dissecting the primate retina to provide tissue for retinal histology, immunohistochemistry, laser capture microdissection, as well as light and electron microscopy. With the extended use of the non-human primate as a translational model, our hope is that improved understanding of the retina will provide insights into effective approaches towards attenuating or reversing the negative impact of visual disorders on the quality of life of affected individuals.The visual system in humans is considered the  gateway  to the world and plays a principal role in the plethora of sensory, perceptual and cognitive processes. It is therefore not surprising that  quality of vision  is tied to  quality of life . Despite widespread clinical and basic research surrounding the causes of visual disorders, many forms of visual impairments, such as retinitis pigmentosa and macular degeneration, lack effective treatments. Non-human primates have the closest general features of eye development to that of humans. Not only do they have a similar vascular anatomy, but amongst other mammals, primates have the unique characteristic of having a region in the temporal retina specialized for high visual acuity, the fovea Tissue should be well perfused with paraformaldehyde, glutaraldehyde, or formalin. This can be achieved through standard transcardial perfusion typically used to harvest other organs. It is recommended that shortly after sacrifice the eyes be injected with fixative just under the lens and stored in fixative.In the present study the subject was deeply sedated with ketamine hydrochloride , euthanized with an overdose of sodium pentobarbital  and perfused transcardially with 0.1 M PBS until completely exsanguinated. This is followed by a 4% paraformaldehyde solution in PBS for 5 min (~1 liter).For easier access to the eyeball it is recommended to first remove the brain. Once the brain has been removed the thin-walled orbit bone is readily apparent. Use the bone rongeurs to slowly chip away the wall of the orbit. Cut away the ocular muscles with a scalpel and remove the connective tissue from the eyeball. Carefully cut the optic nerve, this can be used in electronic microscopy studies. The eyeball should now be released from the orbital cavity.Place the eye into a Petri dish with PBS to keep the retina from drying. A dissecting microscope or table mounted magnifying light stand is useful in the dissection, but not a necessity. Remove the cornea by cutting the sclera closely to the perimeter of the cornea at the level of the ora serrata with a pair of spring scissors and remove the lens with the forceps.Use a paintbrush, forceps, and spring scissors to remove the retina from the sclera. This is done by separating the retina from the sclera and then cutting the sclera away with the scissors. One must carry out this process in small increments so as not to damage or tear the retinal tissue. The sclera is not readily separated from the remnant optic nerve so carefully cut the sclera around the optic nerve. At this point the retina has retained its curved shape and needs to be flattened for sampling. Before flattening the retina, the vitreous humour, which has the consistency of jelly, can be removed in a lump. To flatten the retina onto a slide, make several radial cuts with a scalpel blade. The residual vitreous humour can now be removed with ordinary filter paper and a paintbrush. The retinal ganglion cell layer is exposed at this point so it is imperative to be gentle when removing the vitreous humour. If the optic nerve is still attached at the optic disc, remove the optic nerve without ripping the retina using a scalpel blade and a pair of spring scissors. This is now a flat mount retina  and the fovea should be apparent as a dark patch in the temporal/ventral direction from the optic disc.2There are a number of options for sampling the retina. Here we will describe the flatmount preparation and isodentric sampling. For both procedures flatmount the retina with the optic fiber layer away from the slide. If the intention is to examine the retinal ganglion cell layer in the flatmount preparation, it is necessary to either remove or bleach the pigmented epithelium. To remove the pigmented epithelium use a paintbrush lightly to remove some of this layer, this tends to damage the photoreceptor layer. Bleaching involves soaking the retina in potassium permanganate solution (0.25%) for 1 hour, washing in distilled water, and clearing in oxalic acid (5%) for 5 minutes.If the intention is to examine cell distribution and morphology of each layer throughout the retina, then we suggest isometric sampling of the retina. The retina should be flatmounted on a slide and kept moist with PBS. For isometric sampling cut small pieces of tissue equidistant from the optic disc in the nasal, temporal, upper and lower directions . It is not necessary to bleach the pigmented epithelium in this preparation. These pieces can now be sectioned in the coronal plane with a cryostat, vibratome, or ultramicrotome depending on the research question. For cryostat sectioning, the pieces should be cryoprotected in 30% sucrose overnight and frozen in the context of a mounting medium on dry ice. Alternatively, the samples can be embedded in agar and sliced on a vibratome. The cryostat and vibratome preparations will reliably yield sections as thin as 4 m. If thinner sections are required (e.g. for electron microscope preparation), it is necessary to prepare the samples for the ultramicrotome. To do this, the sections should be post-fixed in osmium tetroxide for 1 hour under the fumehood. This is followed by dehydration in a graded ethanol series  and 100% propylene oxide . The tissue is then embedded in Epon (EMBed-812 embedding kit).In our laboratory, we routinely perform immunohistochemistry on cryosectioned retinae . In this case we are interested in the isodensity of cannabinoid receptors (CB1) in the primate retina. We also examine the effects of prenatal ethanol exposure on the primate visual system. To this end, we are interested in cell density and layer thickness in the fovea and in the peripheral retina. To accomplish this, we embed pieces of retina in Epon, slice at 700nm on an ultramicrotome, and stain with 1% toluidine blue .          Figure 1 Flatmount Retina. Radial cuts are used to flatten the retina.                    Figure 2 Immunostaining. Cryosection of peripheral retina immunostained for CB1 where the retinal ganglion cells are heavily labeled with some labeling in the inner and outer nuclear layers. The thickness of this section is 14 m and stained on the slide. RG - retinal ganglion layer; IP - inner plexiform layer; IN -  inner nuclear layer; OP -  outer plexiform layer; ON - outer nuclear layer.                    Figure 3 Fovea. Section through the fovea that was embedded in Epon and sliced on an ultramicrotrome at 700nm. The photoreceptor layer (PR) was used to align the sections.  Notice that the entire extent of the photoreceptors can be identified indicating an appropriate angle in the coronal plane.  Density measurements were taken at 300 m, 500 m, and 800 m from the center of the foveal pit using the Bioquant Imaging system. RG - retinal ganglion layer; IP - inner plexiform layer; IN -  inner nuclear layer; OP -  outer plexiform layer; ON - outer nuclear layer; scale bar = 50 m          3. Quantification of cell density in the periphery of primate retina is readily accomplished. however, in perifoveal and foveal regions, the stacking of multiple layers in the ganglion cell layer obstructs quantification. To circumvent this potential bias, the fovea and perifoveal region can be dissected from the wholemount preparation, embedded in Epon, and serially sectioned using an ultramicrotome to obtain semi-thin sections in the coronal plane2,4. There are a number of other disadvantages to the wholemount preparation, which can be overcome with alternative sampling paradigms.The preparation of the retina as a wholemount allows for the analysis of the topography and spatial distribution of either the ganglion cell layer or the endothelial cells of the retinal blood vessels5. These sections can then be removed from the slide, embedded in Epon and sliced on an ultramicrotome. With an ultramicrotome minor changes in the cutting angle can be made to ensure a standard coronal plane through the photoreceptors. Once a standard plane has been obtained, layer thickness can be measured and compared between retinal regions and subjects. Furthermore, Epon embedded tissue can be used in electron microscopy studies to reveal ultrastructural characteristics of the retina6.Taking isometric samples from the retina and sectioning in the coronal plane on either a cryostat or vibratome allows for specific examination of the different layers, which cannot be readily performed on a wholemount preparation. Sectioning in this manner also allows for the application of multiple immnohistochemistry protocols


We have also evaluated a Val655Ile single nucleotide polymorphism, which is associated with an increased risk of breast cancer, in a subset of the colorectal cancer patients and in healthy control subjects. Immunohistochemical studies revealed that while 81.8% of tumours expressed c-erbB-2, in the majority of cases equivalent levels of c-erb-B2 were seen in adjacent normal mucosa. Colon tumours were significantly more likely to express c-erbB-2 than rectal tumours (P=0.015). Only 52.4% of the metastases displayed staining patterns concordant with their primary tumour, indicating that determination of c-erbB-2 protein in colorectal tumours cannot predict the status of lymph node metastases. PCR–RFLP analysis of the Val655Ile single nucleotide polymorphism demonstrated that allele frequencies were identical between colorectal cancer patients and a control group of Caucasian subjects , indicating that it is not related to the risk of developing colorectal cancer in this population. Furthermore, there was no relationship between c-erbB-2 protein expression and gene polymorphism (P=0.58). In terms of prognosis, no association was seen between either c-erbB-2 protein expression or the presence of the Val allele and patient survival (P>0.05 in each case), suggesting that c-erbB-2 is not a prognostic marker in colorectal cancer.We have investigated c-British Journal of Cancer (2002) 86, 568–573. DOI: 10.1038/sj/bjc/6600127www.bjcancer.comCancer Research UK© 2002  Colorectal cancer is the second most common malignancy in the developed world .erbB-2  proto-oncogene, located at chromosome 17q21, encodes a 185 kDa glycoprotein which is a member of the type I kinase receptor family, and is involved in mediating a number of normal cellular processes including proliferation  in the transmembrane coding region of the c-erbB-2 gene . Tumours were collected between 1994 and 1998 at the Department of Pathology, University of Aberdeen, as part of the Aberdeen Colorectal Initiative database. Samples were routinely fixed in 10% neutral buffered formalin for 24 h and embedded in paraffin-wax. An experienced gastrointestinal pathologist confirmed the diagnosis of adenocarcinoma following review of all cases. Detailed clinicopathological data  is available for each sample. Cases of peri-operative death were excluded from survival analysis to avoid events that were not associated with disease. One hundred and sixty-three of the 244 eligible patients were alive at the most recent assessment (October 2000), with a median follow-up of 43 months (range 25–80 months).655Ile SNP in colorectal cancer. The genotype and allele frequencies of these colorectal cancer samples were compared with those of 257 healthy blood donor controls from Aberdeen, UK, consisting of 140 male and 117 female volunteers, median age 38 years . All studies obtained relevant approval from local ethical committees.A subset of 151 samples was used to determine the allele frequencies of the ValerbB-2 protein expression was analyzed in colorectal tumour samples using immunohistochemistry. An avidin/biotin/horseradish peroxidase development system was used as previously described  to prevent non-specific background staining. Following incubation with anti-c-erbB-2 monoclonal antibody , biotinylated rabbit anti-mouse immunoglobulin  and streptavidin/biotin/ horseradish peroxidase complex (Dako A/S Ltd), sites of bound antibody were visualized using liquid DAB Plus (Dako A/S Ltd). Slides were then counterstained with Mayer's haematoxylin and analyzed using light microscopy. Sections of an invasive breast adenocarcinoma were used as positive controls, and slides were incubated with TBS in place of primary antibody for negative controls.c-erbB-2. In addition, any normal colonic mucosa present on the section was scored using the same system, and the normal score subtracted from the tumour score. Using this modified system, only cases with a value of 2 were considered c-erbB-2 positive as previously described  and stored at 40C. Genomic DNA was also extracted from paraffin sections containing formalin-fixed normal colon tissue. Dewaxed and rehydrated material was placed in a microfuge tube, and digested with 0.5 mg ml−1 proteinase K for 4 h at 550C. Samples were then heated to 950C for 3 min to inactivate the enzyme, and spun briefly to pellet any cell debris. The resulting supernatant was used directly in PCR analysis.Genomic DNA was extracted from whole blood samples (5 ml) using a sodium perchlorate-chloroform extraction method . Following extraction, DNA was resuspended in 1 ml TE buffer , 1.5 mM MgCl2 (Perkin Elmer), 200 μM each of dATP, dTTP, dGTP and dCTP , 1 M betaine, 1.25U AmpliTaq Gold (Perkin Elmer) and 100 ng each of forward and reverse primers . Primers used in the analysis of the c-erbB-2 Val655Ile polymorphism in genomic DNA were 5′-AGA GAG CCA GCC CTC TGA CGT CCA T (forward primer HN-5), and 5′-TCC GTT TCC TGC AGC AGT CTC CGC A  at 55°C for 2 h, and visualized by electrophoresis on 2.5% agarose containing 0.5 μg ml−1 ethidium bromide. The 148 bp PCR product generated was cut by BsmAI into fragments of 122 and 26 bp if the Ile allele was present, whereas the product from the Val allele was cut to produce fragments of 90, 32 and 26 bp  was digested with 5 U erbB-2 protein expression in primary colorectal tumours and lymph node metastases was evaluated using the kappa test. Expression of c-erbB-2 protein, and the Val655Ile SNP, were assessed with respect to Dukes' stage, degree of differentiation, site of the primary tumour and patient gender using the chi-squared test, while patient age was examined using the Mann–Whitney U-test. The association between c-erbB-2 protein expression and the Val655Ile SNP was examined using the chi-squared test. The effects of c-erbB-2 on survival were tested using Kaplan–Meier survival plots and analyzed using the log rank test. Significance levels were set at P<0.05, and all statistical analyzes were carried out using SPSS for Windows 95 version 9.0 .The relationship between c-erbB-2 protein was examined in 249 colorectal tumours. One primary tumour gave inconsistent immunostaining results on repeated evaluations, and was excluded from further analysis. The distribution of immunostaining in primary colorectal tumours is detailed inTable 1erbB-2, only 8 out of 248 (3.2%) displayed strong immunoreactivity in the membrane of tumour cells  or 3+ (9 cases). Only 24 out of 204 (11.8%) samples displayed higher levels of c-erbB-2 in the tumour compared to normal tissue, and none of these samples reached a difference value of ⩾2  of the examined metastases displayed positive c-erbB-2 immunoreactivity  had higher c-erbB-2 expression in the primary tumour, while two (4.8%) had elevated c-erbB-2 levels in the lymph node lesion. Using the kappa test, which is a measure of agreement between paired samples, only a poor level of concordance was observed . There was no correlation between c-erbB-2 protein expression and patient gender, degree of differentiation, tumour stage, or patient age (P>0.05 in each case).The expression of c-erbB-2 gene was assessed in a subset of 151 colorectal cancer patients and in 257 Caucasian control subjects using PCR–RFLP analysis. The genotype and allele frequencies are shown inTable 4P=0.999; The frequency of a polymorphic Val allele at position 655 of the c-erbB-2 protein expression , nor was there a relationship between this polymorphism and c-erbB-2 protein expression, and the Val655Ile SNP, on patient survival were examined. No association was seen either between protein expression, or the presence of the Val allele, and patient outcome  the expression of c-erbB-2 protein in primary colorectal tumours does not accurately reflect the c-erbB-2 status of lymph node metastases; (3) a SNP at position 655 of the c-erbB-2 gene is not associated with risk of colorectal cancer, nor does it influence protein expression; and (4) c-erbB-2 protein status and Val655Ile SNP do not influence patient survival in colorectal cancer patients.The results of the present study indicate that: (1) although c-erbB-2 protein expression and tumour stage or grade, suggest that c-erbB-2 does not play a major role in the development of colorectal adenocarcinoma. There have been previous reports that have associated c-erbB-2 protein expression with more aggressive colorectal tumours. For example, colorectal tumours which subsequently developed liver metastases more frequently expressed c-erbB-2 protein than those which did not metastasise in an 8-year period  also make comparison between studies very challenging. In addition, the source of the primary antibody used, the immunohistochemical protocol and the size of the sample set may all lead to inconsistencies between reports. This highlights the need for a standardized system of c-erbB-2 classification as previously documented  showed concordance between primary and secondary tumours. While erbB-2 SNP and risk of breast cancer in a Chinese population  in this study is higher than both the control and case Val allele frequencies (0.11 and 0.15 respectively) seen in Chinese populations  was independently related to survival in 155 colorectal tumours. However, in agreement with our findings, erbB-2 expression in 293 colorectal tumours was not associated with prognosis, while erbB-2 staining intensity (membranous and/or cytoplasmic staining) in 96 colorectal tumours. As mentioned previously, these discrepancies are likely due to procedural disparities. The present study, which was carried out using highly reproducible automated immunostaining on a large series of colorectal tumours with a minimum of 25 months follow-up, has used recommended guidelines for assessing c-erbB-2 overexpression (To date, studies on the prognostic utility of c-655Ile SNP on the clinical outcome of colorectal cancer patients. In the group of patients evaluated in this study, there was no association between the presence of the Val allele and patient survival. In addition, we found no relationship between the c-erbB-2 SNP and protein expression, indicating that evaluation of this SNP would be of little benefit in colorectal adenocarcinoma.There have been no previous reports on the influence of the ValerbB-2 appears to be highly important in the development and progression of a number of epithelial malignancies, the measurement of its protein expression and evaluation of the Val655Ile SNP is unlikely to be of clinical benefit to patients with colorectal adenocarcinoma.In conclusion, while c-


The present study investigates the prognostic value of immunohistochemically detected cathepsin D expression in endometrial adenocarcinoma. Patients with surgically treated endometrial adenocarcinoma FIGO stages I-III and consecutive irradiation therapy were included in the study. When we performed immunohistochemistry to detect cathepsin D in 115 tissue specimens 35 cases showed a positive reaction. In the univariate analysis cathepsin D expression showed significant prognostic value for overall survival . In the multivariate analysis with established prognostic parameters  we found an independent prognostic value for cathepsin D . Immunohistochemical detection of cathepsin D could aid in predicting prognosis and planning therapy for patients with endometrial adenocarcinoma.


India is a significant contributor to the world’s total burden of deafness. Out of all causes, almost 50% of the causes of decreased hearing are preventable. With the launch of the National Programme for Prevention and Control of Deafness, the need for an effective information, education and communication (IEC) campaign was felt. There is negligible information available about the status of awareness levels of the community about the various aspects of hearing loss. We carried out this research with the objective of getting to know the existing awareness related to hearing loss in the community to generate an evidence base for formulating various messages to be incorporated in IEC materials for dissemination in the community. We also asked the participants about their suggestions for the various information resources so that an IEC campaign could be designed accordingly.We carried out 10 focus group discussions among various groups of population and analyzed the discussion.A descriptive analysis of the observations regarding the awareness about deafness in the community and prevalent myths and suggested information resources is presented.We highlight the lacunae in the existing awareness of various causes of deafness and the preventive measures that could be taken to prevent hearing loss. The evidence generated was used to formulate relevant messages for the various target groups, which were then incorporated in development of the IEC materials for the dissemination in the community. The sense of hearing is a crucial faculty for the humans to communicate and thereby adapt in the society. At least 250 million people globally have hearing loss, i.e., more than 4% of the world’s population. At least two-thirds of these people, about 165 million, live in developing countries. related to deafness were analyzed. The basic problems found in the rural areas for the lack of health awareness were poverty, illiteracy, ignorance, poor hygiene, inadequate medical guidance, and age-old customs and beliefs. Only 28% of the mothers and 51% of the teachers had correct information regarding ear care. as a tool for collecting information. Ten FGDs were conducted with 104 participants as per the break-up given in the The dual moderator focus group variant was utilized, where one moderator ensures the session progresses smoothly, while another ensures that all the topics are covered.Sampling of participants: Sampling was done conveniently.We broadly identified groups of various stakeholders, approached them, and held discussions with a group of participants who were willing to be a part of the discussion.Location of the study: National Capital Region, New Delhi.The salient points and the main observations derived from the discussions with each group are presented.The participants said that hearing impairment is an important public health problem that has not been given due importance in medical education as well as by the general community. The role of loud music in clubs and excessive use of headphones for listening to music as a cause of hearing impairment was highlighted. All the participants were in favor of use of mass media like television and newspapers for creating awareness about deafness. They suggested that health workers should be trained in disseminating information about hearing impairment through community meetings.The doctors from different specializations opined that hearing loss as a presenting complaint was uncommon. According to pediatricians, mothers gave more emphasis to the problem of ear discharge rather than loss of hearing in their children. Elderly persons, according to ophthalmologists, were more concerned about loss of vision then loss of hearing. The commonly reported wrong practices in the community were putting mustard or garlic oil in the ear, indiscriminate use of ear buds, getting ears cleaned by roadside quacks, and wrongful use of eardrops.The role of early diagnosis and treatment of upper respiratory infections, precautions regarding use of drugs during pregnancy, and awareness about damaging effects of noise were considered important aspects that should be covered in awareness campaigns. Some experts favored television over other media, while others felt that radio had more penetration then TV particularly in rural areas. Awareness generation among doctors was as important as in the general community.Most of the auxiliary nurse midwives opined that they had been seeing about 5–10 persons with deafness in every village. Discharging ear was a very common problem in children, so much so that if the ear of a child was not discharging the mother would be worried that something is wrong with the ears of that child. They admitted their lack of awareness for methods of early diagnosis of decreased hearing in children. Usually they referred the patients to tertiary care hospitals, but acceptance of such advice was relatively poor in the community because of reasons like long queues in hospitals, loss of earning time for daily wage workers, and general indifference of hospital staff. The prevalent practices highlighted were putting oil in the ear or cleaning with objects like broom stick, matchstick, hair clip, etc. One startling observation of a participant was that mothers put breast milk in the infant’s ear as a treatment for deafness.panchayat members and anganwadi workers.Methods suggested for creating awareness among the people were health talks with women folk in the village, with teachers and students in schools during health camps, and in village meetings that involve neem or garlic with mustard oil or snake oil in the ear, and assistance of tantriks and faith healers. However, majority of them agreed that such treatments were not effective in the people they knew. According to them, IEC messages could be similar to those for polio, TB, and AIDS with the involvement of celebrities for added impact. Holding meetings with groups of people and one-to-one communication may also be useful methods.These groups were of the opinion that there is a lack of awareness about where to seek treatment for diseases of the ear. Each one of them had known at least one person suffering from deafness, indicating a high prevalence of the problem. Practices enumerated were putting nukkad nataks (street plays), screening of documentary films, and interactive sessions with deaf people. Writing articles in newspapers specially meant for children was another suggestion.All the teachers were of the opinion that a national program for deafness prevention similar to the Blindness Prevention Program should be started. The only preventable cause of hearing loss according to them was noise-induced hearing loss. They were unaware of causes like ear infections and trauma. Methods suggested by them for awareness generation were conducting competitions among school children on slogan writing, poster making, essay writing, etc.; conducting health exhibitions; and lectures by health experts on special days like World Disability Day. Parent-teacher meetings were suggested as a good forum for talking about deafness, for sensitization of the parents. For the community, they suggested use of They agreed that awareness is low in the community and efforts were needed to improve it. Ear infections and high-grade fever were quoted as causes leading to decreased hearing. They were not aware of preventive measures for ear diseases and hearing loss. They pointed out that people with decreased hearing gradually got excluded from social circles. Apart from the use of mass media like television and radio, other suggestions were use of hoardings and posters at public places like bus stops, markets and hospitals, and organizing film shows in parks.The participants cited ear infections, trauma, and any infection during pregnancy as causes of hearing loss. They were unaware that hitting on the ears, excessive listening to loud music through headphones, and indiscriminate use of ear buds could be harmful. As per them, it was important to provide services for diagnosis and treatment of ear diseases at dispensaries as well. They recommended dissemination of information in schools, institutions, and offices.Most of the participants were unaware of the causes of deafness in spite of having a child with hearing impairment. Only two participants were aware that infection during pregnancy might cause hearing impairment in the newborn. Most of the participants expressed their inability to recognize the symptoms of deafness in small children. The earliest they could think of detecting hearing impairment was in a 1.5 to 2-year-old child. Prenatal counseling at antenatal clinics was suggested as a means of making prospective parents aware of the causes, prevention, and early diagnosis of hearing impairment in children. Putting up of display boards and hoardings at prominent sites in hospital premises, in addition to use of audiovisual mass media and handbills, were suggestions for an effective IEC campaign for deafness.The importance of creating an IEC program for any community health problem cannot be overemphasized. IEC activities provide people the information they need to make informed choices about adopting and continuing healthy lifestyles.Communication inputs increase people’s awareness and concern over matters relating to health and family welfare and subsequently decide to a large extent the limits of possible improvements and changes in these areas. This proneem or garlic oil in the ear of a child and going to tantriks and faith healers for ear diseases were highlighted from discussion among patients attending OPD. Lack of awareness of importance of proper and timely vaccination for the prevention of hearing loss and lack of availability of good diagnostic and treatment facilities as well as rehabilitation facilities were also brought out. An evidence base was generated for the formulation and dissemination of messages such as, “not to get ears cleaned by road side quacks,” “not to hit anyone on the ears,” “avoid excessive exposure to loud sounds,” etc. The need for messages stressing the importance and effectiveness of rehabilitation of hearing loss was also brought out.In our qualitative study, the groups consisting of the ANMs were in our opinion the most well informed. However, they also expressed their inability to identify a child with suspected hearing loss. The FGDs brought out the issues like lack of awareness about preventable causes of deafness such as ear infections, congenital hearing loss, noise-induced hearing loss, drug-induced hearing loss, and hearing loss due to trauma. Various myths and prevalent practices such as putting mother’s milk, Studies in the past have stressed the importance of education and communication in the prevention of deafness at one level or the other.et al.,(In a study by King et al., it was fIn a German study, the statistics in 1981 showed the average age at initial diagnosis of children with hearing impairment to be between 3.3–3.4 years. In 1989, this average age came down to 13.4 months. This favorable result was attributed to the population being informed by special professional events for various target groups.et al.(Many studies have highlighted the importance of noise exposure as a preventable cause of deafness if effective IEC material is created and community is educated about the harmful effects of noise. Brookhouser et al. in theirIn a consensus conference on NIHL it was sAfter generating an evidence base for the IEC needs of the community, we propose following specific recommendations:The social and economic effects of deafness and its prevalence needs to be highlighted in the IEC materials.Awareness to be promoted in junior doctors and medical officers through CMEs and periodic updates.Training the ANMs for primary ear care and recognizing patients that require referral. Creating IEC materials and kits for the ANMs such as flip charts to help them in spreading information in the community for early diagnosis of hearing loss.Creating awareness in teachers through messages for the recognition of a child with hearing loss and to spread awareness in children about NIHL and preventing trauma to the ear.Importance of prenatal counseling and early screening for hearing loss in children to be stressed in messages for the antenatal clinics.Making information about treatment, diagnostic, and rehabilitation centers to be easily and widely available to the community through mass media.The need for disseminating information about various aspects of deafness was felt by all the groups. Awareness levels were low, wrong practices were prevalent in the community, and there was lack of serious concern for deafness as a health problem. Evidence was generated that messages needed to be formulated for primary ear care at home, early diagnosis of ear infections, screening of infants for suspected hearing loss, avoidance of high noise levels, antenatal counseling, and proper rehabilitation of people with decreased hearing. Evidence was also generated that it was felt that mass media would play a major role in an effective IEC campaign for deafness along with increased community participation through involvement of the grassroot primary health care workers.