Intense and long-standing problems in burn centers in Tehran have led nurses to burnout. This phenomenon has provoked serious responses and has put the nurses, patients and the organization under pressure. The challenge for managers and nurse executives is to understand the factors which would reduce or increase the nurses' responses to burnout and develop delivery systems that promote positive adaptation and facilitate quality care. This study, as a part of more extensive research, aims to explore and describe the nurses' perceptions of the factors affecting their responses to burnout.Grounded theory was used as the method. Thirty- eight participants were recruited. Data were generated by unstructured interviews and 21 sessions of participant observations. Constant comparison was used for data analysis.Nurses' and patients' personal characteristics and social support influenced nurses' responses to burnout. Personal characteristics of the nurses and patients, especially when interacting, had a more powerful effect. They altered emotional, attitudinal, behavioral and organizational responses to burnout and determined the kind of caring behavior. Social support had a palliative effect and altered emotional responses and some aspects of attitudinal responses.The powerful effect of positive personal characteristics and its sensitivity to long standing and intense organizational pressures suggests approaches to executing stress reduction programs and refreshing the nurses' morale by giving more importance to ethical aspects of caring. Moreover, regarding palliative effect of social support and its importance for the nurses' wellbeing, nurse executives are responsible for promoting a work environment that supports nurses and motivates them. Working in a burn unit has been described as a stressful occupation . Every nWhile no health-care professional is immune to these pressures, there is evidence that suggests that areas of nursing particularly those areas we think of as critical care environments such as burn units, are often the most vulnerable to stress, and in need of much support ,7. NurseHowever, regarding emotional, attitudinal, psychosomatic, behavioral and organizational responses of these nurses to burnout, it is vital to identify the factors that involve in their perception of burnout. Some authors also referred to these factors in burn centers  and otheThe challenge for managers and nurse executives of burn centers is to understand the intervening factors and their impacts on these burn nurses' responses to burnout. As a result they can develop and promote delivery systems that support positive adaptation to stressors in burn centers, retain nurses and facilitate quality nursing care.In order to understand nurses' perceptions of factors modifying their responses to job stress and burnout, qualitative research adapted from the grounded theory method was chosen .The value of using a qualitative research method such as grounded theory is embedded in the subjective and often emotional nature of care, stress and coping. As a descriptive study, the qualitative paradigm, with its emphasis on understanding factors modifying nurses' responses to job stress and burnout from the view point of practicing nurses themselves seemed logical. Grounded theory is a theory that is derived from data, systematically gathered and analyzed through the research process  The aim In effect, grounded theory is guided by simultaneous analysis. Both analysis and data collection inform each other. The analysis process is systematic and ends when new data no longer generate new insights. This has been also described as ' category saturation' ,14.Five clinical nursing instructors participated in the pilot study. They were faculties of School of Nursing in Iran University of Medical Sciences (IUMS) and had been supervising nursing students in burn centers of Tehran for many years. Their age ranged from 40–48 years and had been working in burn centers for 7–14.5 years. The aim of pilot study was using the experiences of nursing instructors in the original study and reducing the informant and researcher bias in the interviews and participant observations .The results of the study indicated that the staff of burn units felt drained, they haven't had any motivation or desire to care and they had been working purely for their pay. The findings were strongly indicative of the symptoms of burnout. It revealed that their behavior was representative of an indication of their professional dilemma. The pilot study also indicated that social support, patients' cooperation/motivation and the nurses' unique characteristics had been modified to alter the nurses' responses to burnout. Analysis of data from the original study was conducted keeping these findings in mind.The research proposal was approved by the ethics committee of IUMS. Then permission was granted from the managers of two burn centers and their nursing administrators. Further permission and written consent was obtained from all who participated in this study.Following ethical approval, data was collected through tape- recorded, unstructured interviews. Initially data was collected in one center and analyzed. Then data gathering was initiated in the second center. There were 19 informants from the first center and 14 from the second center that participated (except 5 participants in pilot study). From this sample, 25 were nurses in different levels and positions and 8 were other members of burn team. The nurses' sample included 8 staff nurses, 8 licensed practical nurses, 2 nurses' aids, 3 head nurses, 2 supervisors, and 2 nursing administrators. Since nursing staff pointed to some issues concerning the burn team, the researcher interviewed one physician, one social worker, 2 physiotherapists, and 4 patients in the process of theoretical sampling.Criterion for recruiting nursing participants was at least one year of experience in the burn center. Patients were selected according to their desire as well as their physical and psychological stability. Selecting patients occurred by consultation with head nurses.Participants were 11 males and 22 females. The nursing staff participated were 19 females and 6 males. Twelve of the nursing staff had been working 2–3 shifts in burn centers or other hospitals as well as working in other jobs due to financial needs. Other demographic information of the nursing staff is displayed in Table Samples were recruited from all units of both centers. The first purposeful sample included 6 of nursing staff. Theoretical sampling was used after emerging the tentative theory. The basis for theoretical sampling was the questions which emerged during data analysis. At this stage the researcher interviewed nursing administrators and other members of the burn team. Theoretical sampling helped in verifying nursing staff's responses and credibility of categories and resulted in more conceptual density. No new data were emerged in the last two interviews; therefore data gathering by interviews were terminated.Interviews were conducted in a private place with mutual agreement of the interviewer and interviewees. All interviews were completed by the main researcher. The duration of interviews ranged between 30–165 minutes. All the interviews were tape recorded except one. Some notes were taken during dialogues.Unstructured interviews were conducted using a topic guide which has been drawn up by the researchers initial review of the literature related to the concepts of the subject of the study. This topic guide included the structure, process and outcome of care .The following grand tour question guided the study:" please tell me about the nursing care in your unit". Subsequent questions were based on the participants' responses and demands of the emerging theory. Interviews were terminated when data redundancy occurred.In each center after the termination of interviews, participant observations were performed in all wards at morning, evening and night. 14 sessions of observation in the first center and 7 in the second center occurred. For this purpose the researcher informed the nursing administrators of her program. By selecting all the wards in all shifts there was no need for theoretical sampling in this stage (place and time) , but theDescriptive, focused and selective observations were occurred in a non- linear fashion. Theoretical sampling occurred during focused and selective observations. Some of the questions which guided the theoretical sampling were," is there any difference between nursing cares received by different patients?", "is nursing care different in a large or small ward?"Prolonged engagement of the researcher in the field reduced the obtrusiveness. The level of participation varied from complete observation to participation in some activities. Some informal interview also occurred during the observations. Immediately after each session of observation field notes were completed systematically. Analysis of the field notes helped in determining contextual conditions and explaining variations in the nurses' responses in each context. This led to proposition of several hypotheses.Data collection, analysis and interpretation occurred simultaneously, in keeping with grounded theory methodology . After eThe process of integrating and refining the theory occurred in selective coding . In thisThe researchers accepted the perspective of Guba and Lincoln. They translated internal validity into credibility, external validity into transferability, reliability into dependability, and objectivity into confirm ability .Credibility enhanced by the researchers' describing and interpreting her experiences. For this purpose the researcher kept a field journal in which she noted the content and the process of interactions, including reactions to various events. This journal became the record of relationships and provided material for reflection. Prolonged engagement and persistent observation helped to data credibility. In this way the process of data collection and analysis took 8 months. Data triangulation and method triangulation confirmed credibility . MaximumConfirm ability requires one to show the way in which interpretations have been arrived at via the inquiry. In this study, confirm ability was established, because credibility, transferability and dependability were achieved . The sigThe findings related to factors which involved in the nurses' responses to burnout are presented in this article. Analysis and interpretation of data indicated that personal characteristics and social support have involved in the nurses' responses. These factors are presented in Table Data from interviews and participant observations indicated that special personal characteristics and personality traits have involved in the nurses' emotional, attitudinal, behavioral, and organizational responses to burnout. Personal characteristics such as conscience, religious beliefs, personal philosophy, commitment, a sense of responsibility, and altruism facilitated caring behaviors. Nurses with these characteristics were more patient and empathetic. They were more cooperative and rarely justified their faults by fatigue, workload or staff shortage. Conscience, commitment, and religious beliefs such as fear of divine requital were the most prominent traits that modified the responses to burnout. One of participants stated:" God knows. I always feel it's me there on the bed. Sometimes a patient calls and I ignore, but I tell myself, what I expected if I were on this bed? I fear god and say to myself, his authority is great and whatever I do, I will see the reflection of my doings". Some of the participants pointed to interest and love in caring of burn victims. One of participants stated:" these patients are different from other ones. I have worked more than 18 years in general hospitals, I haven't worked more than 7 years in burn centers, but I think that was a blessing in disguise, I am glad because clinical work for a burn patient means love, means everything, believe me. I don't care the managers' behaviors, workload, nursing shortage and other deficiencies, because I love burn victims. Believe me". Many of the nursing staff, distanced from patients, they had immoral beliefs and demonstrated humiliation and reproach in their behaviors. They related these attitudes to fatigue, micro and macro conditions in burn centers, and loss of motivation; but participant observation indicated that, this isn't the case for all the nurses. Nurses, who had been known as good nurses, were very calm and intimate with their patients and focused on the patients' needs. The researcher wrote in one of her field notes:" she is very calm and speaks with compassion. She makes jokes and patients are relaxed with her. She follows the principles and procedures more strictly than others". Nurses in all levels were under pressure of workload, low pay, staff shortage, environmental conditions of burn units and other structural inhibitors, but as the excerpts of interviews indicated, appraisal of these inhibitors was different in the presence of specific personal characteristics.Data indicated that in some instances, when there were a number of inhibitors and they were long stay, even positive personal characteristics couldn't work. This was often the case in infectious dressing rooms and busy wards. The worst kinds of treating patients were seen in these places. It seemed that the inhibitory factors which are persistent and too frequent interact with personal characteristics and finally overcome the positive characteristics. One of participants stated:" patients expect to receive care, expect a friendly encounter which does not happen. To tell the truth, some days I am excessively distressed and tired that I don't have the patience to answer the patient' questions and concerns. I emotionally can not do what I really want to do on a daily basis for the patients. The reason is persistent day and night problems occurring with this job". This process is displayed in Table Data strongly indicated that nurses' appraisal of the patients' characteristics have influenced some aspects of their attitudinal and behavioral responses. When the appraisal was positive, the relationships improved, and when it was negative, relationships deteriorated. Positive appraisal occurred often when the patients were cooperative and motivated for recovery and in cases where they had an advantage of high socio- economic class, cultural and educational levels, or whenever they stimulated the nurses' senses of compassion and pettiness. Negative appraisal occurred often when the patients were from lower socio- cultural levels, addicts or there was a possibility of having acquired immune deficiency syndrome (AIDS) or hepatitis.The first group was treated kindly and more respectfully. The use of humiliating words and reproach towards them diminished and as a result the aggressive behavior and physical withdrawal less occurred. One of participants stated:" ...I am more supportive and compassionate towards children, those who are very alone, who have no one to love and care for them, those who have committed self-inflicted burns, a woman whose husband caused her to burn herself and doesn't have any one to support her. In many occasions I have even paid them to buy juice from outside the hospital. I feel that these patients are needy".The second group was treated very unethical. They encountered a humiliating, reproaching, and aggressive behavior. One of participants stated:" ...he fights when I am dressing him, he pulls his hand and leg, he isn't cooperative, and he has no class. They drain all my energy to the point that I don't want to talk to them. I think they are mentally retarded. They keep still when I shout at them just the way that the children act. I tell them I'll pull your ear, and I'll beat you up. This is the behavior that has worked with them ".Moreover patients with extensive burns, whose survival was an improbable event, not only were badly treated, but also were sometimes ignored and received poor care. In other words, they received only those treatments which had been ordered by physicians to prevent being reprimanded by supervisors. One of the participants justified herself and stated:" if you want the truth, a patient with 90% burns can not benefit from tetracycline ointment, but it's there in his order, I prefer to spend my time with a patient who has a better chance of surviving. Right or wrong I don't apply the ointment, because I can spend that time for a patient who will survive". We can conclude that burnout has made nurses to modify their caring behaviors to fit the different type of patients they care for.As described later, nurse's and patient's characteristics modified the nurse's responses to burnout and altered caring behaviors. More analysis and interpreting of data indicated that interaction between these two variables resulted in a more powerful combination that alters responses to burnout and identifies the kind of caring behavior. This process is displayed in table Table One of participants stated: "I take care of some patients with love and conscience and take care of the other patients only with conscience and some nurses doesn't have love at all, I take care of silent, calm and lonely patients better, I perform routine care for the other". Another participant also stated:" it seems I give positive energy to the patients to whom I am more interested and take care of them with more love. I've seen that they respond better to therapeutic measures". Therefore interaction between nurses and patients' characteristics has a very powerful effect on the nurses' responses to burnout and determines their nature of caring behavior.Data from interviews and participant observations strongly suggested that social support influences the nurses' responses to burnout.Supportive behavior of head nurses, nursing administrators and coworkers modified the nurses' responses. Among these, head nurse's the support was the most effective factor. Nurses believed that they do not have any motivation or desire to perform well when they are not supported well. One of participants stated:" we have a very close loving relationship with the head nurses who are supportive. In that situation I do many things for her. Her supportive attitude and caring/positive attitude helps me a lot. Do you understand?" another participant believed that he couldn't endure if the nursing administrator weren't supportive. He stated:" I have seen that she is doing a good job. I have been so stressed at times that I have thought of quitting. The nursing administrator has changed my mind during those stressful moments by being caring, loving and supportive and I have decided to stay in spite of low pay and benefits." Support from peers also modified the nurses' responses. They could tolerate more with the support of their coworkers. One of participants stated:" by god, I like every single one of them. It seems like we live together seven hours a day (major part of our day). We are so familiar with each other's character and behavior patterns. You may not believe this, these relationships has been very helpful and caused us to have a very strong unit. We care for each other in times of weakness, illness or pressure".It is worthy of mention that the effect of social support on the nurses' responses to burnout was not as powerful as the nurses' and patients' characteristics. Social support influenced the emotional and in some cases the attitudinal responses, but it didn't have enough power to alter the organizational and behavioral responses, therefore it didn't change caring behaviors significantly. Moreover, there were not any data indicative of the modifying effect of the factors proposed in this article on the psycho- somatic responses to burnout.Findings of this study indicated that nurses' and patients' characteristics and social support modified the nurses' responses to burnout. These factors altered the nurses' perceptions of the inhibitory factors; in other words they could have a more positive appraisal and this in turn modified their responses.Lazarus and Folkman 1988) proposed that the initiative of behavioral manifestations is a transactional appraisal which is important for the person's wellbeing and betrays the confrontation as noxious, useful, threatening or requiring struggle 88 propos. In thisIn this study, when inhibitory factors were persistent and frequent, even positive personal characteristics defeated. Selye (1976) concluded that the influence of stressors on a person depends on the number of stressors that must be confronted in a time, the duration of confrontation and existence of previous experience with the same stressors . This haPatients with different characteristics treated differently. Nurses changed their caring behavior with different patients. It seemed that they didn't have enough emotional and physical energy and motivation for caring for all patients; but some patients stimulated their emotions and gave them the needed positive energy for caring. In other words, patients' characteristics could both reduce and intensify the nurses' responses to burnout. When these characteristics appraised as positive, caring behaviors improved, and when it was negative, the worst kind of behaviors occurred. Maslach (1982) in congruence with this conclusion believed that there is little evidence that caring is a uniform state , and BenPatients with extensive burns received the worst kind or care. Nurses stated that caring for these patients is futile. Data implied that caring for these patients have been incongruous with the nurses' values. Meltzer and Huckabay (2004) in their study of the relationship between critical care nurses' perceptions of futile care and its effect on burnout, concluded that feeling of emotional exhaustion in these nurses was highly influenced by the frequency with which nurses were involved in life- sustaining interventions that conflicted with the nurses' values and standards in term of what the nurses thought are ethically appropriate and could result in improvement in a patient's condition and outcome .It was the interaction between nurses' and patients' characteristics that identified the caring behavior. Some authors have found according to their clinical practice that nurses have the ability to adjust their approach and their style of interaction with different patients. These authors proposed that they not only alter the nature of dialogue and the tone of voice to meet each patient's needs, but also adjust their affective response. They pointed that delineation of these behaviors would be a significant contribution, yet to date these styles of care have not been explored . In thisSocial support from head nurses, nursing administrators and peers made nurses to endure and tolerate in the face of problems. Lazarus (1976) proposed that the resources reducing the potential harm could be found in the environment and essentially in others who indicated one can rely on them . Social This study as a part of more extensive research (PhD dissertation) identified the most important factors that intervened in the nurses' responses to burnout in burn centers of Tehran. Nurses had responded to burnout. These responses included emotional, attitudinal, psychosomatic, behavioral and organizational. This part of study indicated that the nurses and patients characteristics and interaction between these two factors had a very powerful effect on the responses and determined the kind of caring behavior. Moreover social support from managers (e.g. head nurses and nursing administrators) and peers modified some of the nurses' responses to burnout.The influence of positive personal characteristics, especially conscience, religious beliefs, philosophy, commitment, a sense of responsibility and altruism on the nurses' responses to burnout, the finding that, long lasting and persistent problems in the work setting can deteriorate even the personal characteristics, and regarding the numerous problems in burn centers of Tehran, there is an urgent need for helping the nurses. We suggest that due to the intense staff shortage in these centers, the managers try to keep these nurses. The only way they can do this is by using stress reduction programs. Data strongly indicated that these nurses need to rest periodically to preserve energy and to refresh their morale. Moreover, giving importance to moral and ethical aspects of care by managers could be helpful and motivating. Changing burn patients' inherent characteristics and their other characteristics such as poverty and socio- cultural level is not a possible alternative. Promoting nurses' morale is possible and must be done promptly if we want our burn survivors receive at least an ethical effective care.Social support made the problems tolerable. Therefore we recommend nurse executives in burn centers of Tehran promote a work environment that help to decrease the perception of pressures and increase perceptions of social support. The best solution is of course eliminating the micro and macro conditions which overshadow the nurses' responses and caring behaviors.This grounded theory created several hypotheses in this stage. Nurses with specific traits were more resistant to burnout and were more caring. It is suggestive of conducting a quantitative research to test the relationship between personality traits, burnout and caring behaviors. Differences in caring behavior for different patients were related to the influence of burnout on the nurses' personality traits and appraisal of patients' characteristics. It was a very new finding that needs to be investigated in more detail.The dramatic effect of social support on the nurse' perceptions of pressures in burn centers of Tehran is suggesting of identifying the relationship between social support and the level of burnout the nurses experience in these burn centers.However the nature of a qualitative research like this, limits it's generalizability; therefore we suggest conducting more qualitative research in other critical care units to support these findings.Application of the findings of this study and conducting the suggested studies would help the managers of burn centers to enhance an environment conductive to morale, promote natural and ethical care, refresh the nurses' positive emotions and facilitate a supportive environment for their nurses.The author(s) declare that they have no competing interests.FR initiated and designed the research, collected and analyzed the data and wrote the paper. FO was the main supervisor, helped in analysis, and revised and edited the drafts. MN was co- supervisor and revised the drafts.The pre-publication history for this paper can be accessed here:


Neutrophil infiltration is one of the critical cellular components of an inflammatory response during peritonitis. The adhesion molecules, P-selectin and intercellular adhesion molecule (ICAM)-1, mediate neutrophil-endothelial cell interactions and the subsequent neutrophil transendothelial migration during the inflammatory response. Despite very strong preclinical data, recent clinical trials failed to show a protective effect of anti-adhesion therapy, suggesting that the length of injury might be a critical factor in neutrophil infiltration. Therefore, the objective of this study was to determine the role of P-selectin and ICAM-1 in neutrophil infiltration into the peritoneal cavity during early and late phases of peritonitis.in situ immunohistochemical staining. Comparisons between groups were made by applying ANOVA and student t-test analysis.Peritonitis was induced in both male wild-type and P-selectin/ICAM-1 double deficient (P/I null) mice by cecal ligation-puncture (CLP). Peripheral blood and peritoneal lavage were collected at 6 and 24 hours after CLP. The total leukocyte and neutrophil contents were determined, and neutrophils were identified with the aid of CLP induced a severe inflammatory response associated with a significant leukopenia in both wild-type and P/I null mice. Additionally, CLP caused a significant neutrophil infiltration into the peritoneal cavity that was detected in both groups of mice. However, neutrophil infiltration in the P/I null mice at 6 hours of CLP was significantly lower than the corresponding wild-type mice, which reached a similar magnitude at 24 hours of CLP. In contrast, in peritonitis induced by intraperitoneal inoculation of 2% glycogen, no significant difference in neutrophil infiltration was observed between the P/I null and wild-type mice at 6 hours of peritonitis.The data suggest that alternative adhesion pathway(s) independent of P-selectin and ICAM-1 can participate in neutrophil migration during peritonitis and that the mode of stimuli and duration of the injury modulate the neutrophil infiltration. Sepsis is a common cause of morbidity and mortality following surgery or trauma, and is characterized by activation of a systemic inflammatory response, severe hypotension, and major damage to multiple organs . Althougendothelium (P-selectin and E-selectin) [The selectins mediate neutrophil rolling while the β-2 integrins are important for firm adhesion and transendothelial migration . The selelectin) . ICAM-1 electin) . ICAM-1 electin) .In vivostudies have shown that administration of small molecule ligands, and/or neutralizing antibodies to the selectins, ICAM-1 or β2-integrins can protect tissues from injury following endotoxin exposure, bacterial infection, or ischemia [et al. has shown that soluble P-selectin attenuated skeletal muscle reperfusion injury by inhibition of the classical complement pathway [et al. have demonstrated a significant reduction in neutrophil infiltration into the peritoneal cavity at 4 hours of streptoccocal peritonitis in ICAM-1 mutant mice [et al. has shown that P-selectin and ICAM-1 significantly contributed to liver and lung injury at 4 hours of systemic endotoxemia in ICAM-1 and P-selectin/ICAM-1 double mutant mice [et al. have shown that P-selectin and ICAM-1 double mutant mice exhibited complete loss of neutrophil migration into the peritoneum during S. pneumoniae-induced peritonitis [et al. have shown no differences in survival between wild type and ICAM-1-deficient mice following an intra-peritoneal injection with E. coli, S. auerus, or P. aeruginosa [ischemia -10. Howe pathway . Geneticant mice . Anotherant mice . Furtheritonitis . In contruginosa . More imruginosa . In an aruginosa . Additioruginosa . This moAll chemicals were purchased from Sigma Chemical , unless otherwise noted.Guide for the Care and Use of Laboratory Animals . Experimental protocols were reviewed and approved by the Michigan State University Animal Use and Care Committee.Only adult male mice  were used in this study. All animals received humane care in compliance with the tm1BaySelptm1Bay, backcrossed to C57BL/6, were used in this study. Breeding pairs of double-knockout mice were purchased directly from Jackson Laboratory  and bred under the guidance of University Laboratory Animal Resources at Michigan State University. The wild-type (WT) mice were male C57BL/6, which were acclimated to the animal laboratory environment for one week before the start of experimentation. Before and after surgery, all the animals had unlimited access to food and water.Gene-targeted double mutant mice deficient in P-selectin and ICAM-1 (P/I double mutant), C57BL/6-Icam1et al. [Polymicrobial sepsis was induced by CLP as previously described by Chaudry, et al. . SurgicaAdult male mice  were lightly anesthetized by inhalation anesthesia with methoxyflurane. A volume of 2% sterile glycogen solution, equal to 10% of body weight, was injected intraperitoneally (i.p.). Six hours after glycogen inoculation, mice were euthanized and peritoneal lavage and blood samples were collected. The collected samples were identified with ID numbers to assure a blind fashion performance of the tests and data analysis. Total leukocyte and differential counts were performed as described below.Blood samples were obtained from the right ventricle via a left anterior thoracotomy at the time of sacrifice, using a sterile heparinized syringe containing 50 μl of heparin (100 USP Units/ml). Blood smears were prepared and stained with Wright-stain  for differential cell counts. The total number of peripheral blood leukocytes was determined by lysing the red blood cells using 3% acetic acid solution with the aid of a Neubauer hemocytometer. The remaining blood was centrifuged, and plasma was collected and stored at -70°C. A portion of the liver was fixed in buffered 10% formalin and embedded in paraffin, and a second portion was snap frozen in liquid nitrogen and stored at -70°C until used for immunohistochemistry staining.The peritoneal fluids were collected using repetitive (2 times) instillation and withdrawal of 2 and 5 ml respectively of sterile saline solution using a syringe with a 22 G needle. The peritoneal lavage sample was placed on ice, immediately processed for centrifugation at 4°C, and the supernatant and the cell pellet were collected separately. The cell pellet was used for total and differential cell counts. Differential cell counts were determined on cytospin preparations of peritoneal lavage stained with Wright-stain (LeukoStat). The total number of peritoneal leukocytes was determined as described above for the peripheral blood leukocyte count. Additionally, the presence of neutrophils in peritoneal lavage was confirmed by immunohistochemical staining of cytospin preparation using a primary antibody (IgG2a) specific to mouse neutrophil as described below. Further, the neutrophil content was quantified by measuring the myeloperoxidase (MPO) level of peritoneal lavage as described below.The MPO contents of the peritoneal lavage supernatant and the cell pellet were quantified as previously published by our laboratory . Briefly2 clone, PharmMingen, San Diego, CA) and a biotin-conjugated secondary antibody. Tissue sections were counter stained with hematoxylin  and mounted with DAKO Mounting Media . The samples were examined using a Nikon light microscope interfaced with a spot 24-Bit Digital Color Camera.To confirm the identity of neutrophils in peritoneal lavage, immunohistochemical staining was performed using acetone-fixed cytospin cell preps. The primary antibody  specific to mouse neutrophil , the biotin-conjugated secondary antibody , and a Vectastain avidin-biotin complex reagent and 3,3'-diaminobenzidine chromogen kits  were used as previously described in detail . The expt-test by way of StatView version 5.0.1 software© for Windows. Comparisons between multiple groups and various time points were analyzed using ANOVA with subsequent Fisher's PLSD test. P ≤ 0.05 was considered significant.All data were expressed as means ± standard error of the mean. Comparisons between two groups were performed using an unpaired The Jackson Laboratory, where the P/I null breeding pairs were purchased, had initially tested the double knockout of the P/I null mice. In addition, the ICAM-1/P-selectin deficiency was confirmed in our laboratory in randomly selected litter mice tissue samples using RT-PCR and immunohistochemical staining of the liver tissue, as previously published by our laboratory (18). The ICAM-1 expression was determined in all the animals used in this study. Figure Clinical signs of sepsis were manifested as quietness, lack of response to stimulus, absence of socializing behavior, ruffled hair coat and lack of appetite. All mice subjected to CLP displayed these signs that became significant at 24 hours of CLP, and no striking differences were observed between the wild-type and P/I null mice.To establish the blood cell parameters under normal physiological conditions in wild-type and P/I null mice, peripheral blood samples were collected from randomly selected mice. In this article these mice are denoted as "Control", which were not subjected to the sham or CLP operation. The leukocyte and differential counts of peripheral blood were measured. As shown in Table i.p. inoculation of 2% glycogen. Further, the inflammatory response to the sham operation, which elicited a trauma-induced peritonitis, caused leukopenia in both wild-type and P/I null mice. As shown in Table The induction of CLP-induced peritonitis resulted in a significant leukopenia in wild-type as well as P/I null mice as compared to their control counterparts Table . It is i6 vs. P/I = 3.6 ± 1.2 × 106). However, it is interesting to note that monocytes/macrophages were the predominant cell type present in the peritoneal cavities of the wild-type mice, whereas in the P/I null mice lymphocytes were the predominant cell type (Table p = 0.08).Similar to peripheral blood, the cell parameters of peritoneal fluid under normal physiological conditions in the wild-type and P/I null mice were determined in randomly selected mice. The mice were not subjected to the sham or CLP operation. As Table Sepsis induced by CLP  caused a significant leukocyte infiltration into the peritoneal cavities of both wild-type and P/I null mice when compared to their corresponding sham group Figure . The perPeritonitis was also examined in response to 2% glycogen. A significant leukocyte infiltration into the peritoneal cavity occurred in both wild-type and P/I groups at 6 hours after 2% glycogen injection Figure . SimilarTo further quantify the degree of neutrophil infiltration into the peritoneal cavity, the MPO levels of both the peritoneal lavage cell pellets and supernatants were measured. Although the MPO levels of the cell pellets indicated the presence of a significant number of neutrophils in the peritoneal cavity after CLP, the MPO values did not always correlate with the absolute number of infiltrated neutrophils. The Wright's staining of cytospin preparations of the peritoneal cells demonstrated highly activated phagocytes with vacuolized cytoplasm often containing numerous intracellular bacteria and in most cases with loss of cellular integrity Figure . Thus, iPrevious studies have indicated that tissue MPO activity can be affected by other factors . and that MPO values may not indicate the true presence of neutrophils. Additionally, recent studies have identified MPO as the cellular component of macrophages  (21), as well as histiocytes (22). Therefore, an immunohistochemical staining technique was used to confirm the presence of neutrophils in the peritoneal cavity. The cytospin preparations of the peritoneal lavages were immunostained with a specific antibody to mouse neutrophil. Figure Neutrophil infiltration is coordinated by the interplay of the adhesion molecules and the chemoattractants, which plays an important role in inflammatory tissue injury. Recent clinical trials of anti-adhesion therapy did not demonstrate a protective effect in trauma-induce shock despite very strong pre-clinical data (15). Further studies to clear this disparity suggested that the model systems applied and the length of injury are important factors that might modulate the underlying mechanism of neutrophil activation and migration (15). Therefore, this study was undertaken to examine the role of P-selectin and ICAM-1 molecules in the wild-type and P/I null mice subjected to a short and longer periods of peritonitis induce by CLP. The CLP model, chosen to mimic the normal course of sepsis in humans and animals, induced the classic signs of sepsis and a significant systemic inflammatory response. This response was associated with increased leukocyte infiltration into the peritoneal cavities of both the wild-type and P/I null mice. The leukocyte influx into the peritoneal cavity consisted predominantly of neutrophils i.e., 62–80%), which significantly increased in both wild-type and P/I null mice at 6 and 24 hours of CLP. At the early phase of CLP , the total number of neutrophils infiltrated into the peritoneal cavities of P/I null mice were significantly lower than those of the corresponding wild-type mice, which reached to a comparable level at 24 hours of CLP. In contrast to CLP-induced peritonitis, peritonitis induced by 2% glycogen exhibited no significant differences in the number of neutrophils infiltrated into the peritoneal cavities between the wild-type and those of the P/I null mice at 6 hours  or chemically-induced peritonitis (i.e 2% glycogen) and during the late phase of CLP . The data suggests that there might be a functional role for these adhesion molecules during the initial stages of the inflammatory response, and as the inflammatory process progresses, deficient adhesion mechanisms are bypassed. This notion has also been proposed in leukocyte recruitment in an adhesion .2-integrins  expressed on neutrophils has shown to be a critical event mediating stable neutrophil adhesion and migration across the vascular endothelial barrier [per se, is not a critical factor that results in dysfunctional β2-integrin-mediated migration. Other adhesion molecule(s), ligand(s), and/or yet unknown counter-receptor(s) could mediate neutrophil infiltration. For example, ICAM-2, a ligand for β2-integrins, could be a potential candidate [Interaction of ICAM-1 expressed on the surface of vascular endothelial cells with the β barrier -28. Alth barrier . Furtherandidate . Other sandidate ,31. Thisandidate . Neutropandidate ,34. Addiandidate . Neutropandidate . The imp2-integrin-mediated cell adhesion and motility of neutrophils has been described [2 integrins, such as those produced during coagulation as well as complement activation and tissue factor, which could facilitate neutrophil adhesion and infiltration into the peritoneal cavity [It is possible that neutrophil infiltration utilizes other secondary or tertiary adhesion pathways, and/or is facilitated by proteins that mediate the function(s) of the adhesion molecules. For example, a novel glycosylphosphatidyl inositol-anchored protein (GPI-80) that may regulate βescribed . Additiol cavity ,40. Morel cavity . It has l cavity .i.p. inoculation of Streptococcus pneumonia [et al. study, the inflammatory response and neutrophil infiltration into the peritoneal cavity was studied at 4 hours of the induction of peritonitis, while, in our study neutrophil infiltration was evaluated at 6 and 24 hours of peritonitis. In support of this reasoning is the study reported by Mizgerd et al. who have shown a compromised neutrophil migration into the peritoneal cavity of E-/P-selectin and ICAM-1 mutant mice in response to the injection of Streptococcus pneumoniae at 4 hours after injection, with no impairment present at 24 hours of bacteria injection [et al. demonstrated that ICAM-1 is not necessary for neutrophil migration during glycogen-induced peritonitis, as shown otherwise by other authors [Streptococcus pneumonia organisms. In our study, the data of peritoneal supernatant MPO levels have demonstrated a significant level of MPO released into the peritoneal cavity in mice subjected to CLP (Figure in situ immunohistochemical staining of the peritoneal cells clearly confirms the presence of neutrophils in the cytospin preparations (Figure Contrary to several other studies that have demonstrated the functional importance of P-selectin in models of myocardial infarction and inflammatory lung and liver injury, the data presented in this study indicated that P-selectin is not essential for neutrophil migration into the peritoneal cavity -45. In sneumonia . The difnjection . Additio authors . FurtherP Figure . In Bulls Figure . The datStreptococcus- or LPS-induced peritonitis, the present study employed polymicrobial septic peritonitis as well as chemical and trauma-induced peritonitis. Published studies have shown quantitative and qualitative differences in the inflammatory response induced by gram-positive, gram-negative, polymicrobial sepsis and purified endotoxin [Another important difference between the results of the study presented in this article and those of previous studies may relate to the employment of essentially different models of sepsis and inflammation. While previous studies employed hemorrhagic shock-, ndotoxin -52. Thesndotoxin ,54.The data presented in this study have shown a clear, time-dependent neutrophil migration and infiltration into peritoneal cavity in responses to peritonitis, which is independent of P-selectin and ICAM-1 adhesion molecules. The negative nature of the data presented here and the failure of the anti-adhesion clinical trials, are of great importance. These findings demand innovative alternative approaches to neutrophil transmigration in inflammatory response and suggest that targets other than these adhesion molecules need to be identified. A better understanding of the mechanisms leading to neutrophil migration is critical for the development of new therapeutic strategies for treating inflammatory disease without compromising the host's immune response mechanisms.None declared.CR participated in the technical procedures for CLP, collection of blood and tissue samples, and MPO assay. KH and HA participated in preparation of tissue sample for immunohistochemistry and special staining. KH also assisted in preparation of the manuscript. EC conceived the study, and participated in its design, coordination, and preparation of the manuscript. All authors read and approved the final manuscript.CLP, cecal ligation-puncture; ICAM-1, Intercellular adhesion molecule-1; mAbs, monoclonal antibodies; MPO, Myeloperoxidase; P/I null mice, P-selectin/ICAM-1-deficient miceThe pre-publication history for this paper can be accessed here:


From 1963 to 1971, 117 babies with open spina bifida were treated non-selectively from birth. In 2002 we reviewed all the survivors by postal questionnaire and telephone call. The aims were to find out how many were living independently in the community or were in open employment or drove a car. In addition to these achievements we recorded health, medication and admissions to hospital and asked how much daily help they needed.Ascertainment was 100%. There had been 63 deaths, mainly of the most severely affected. The mean age of the 54 survivors was 35 years. The outcome in terms of disability ranged from apparent normality to total dependency. It reflected both the neurological deficit, which had been recorded in infancy in terms of sensory level, and events in the CSF shunt history. Overall about 2 in 5 of the survivors lived independently in the community, 2 in 5 drove a car, 1 in 5 was in competitive employment and 1 in 5 could walk 50 metres.Although those who survived to age 35 years tended to be less disabled, 2 in 5 continued to need daily care. Neurosurgical intervention in babies with open spina bifida had dramatic results in terms of survival. However, the disability and the complications of the survivors were often severe -5. Many In 1963 the Regional Neurosurgical Unit at Addenbrooke's Hospital, Cambridge, England offered treatment to all cases of open spina bifida, without any attempt at selection. Between 1963 and 1971, after a detailed neurological examination, 117 babies  had their open spinal defects closed within 48 hours of birth. A ventriculo-atrial cerebrospinal fluid (CSF) shunt was inserted for hydrocephalus when required.In 2002 all survivors were surveyed by confidential questionnaire and telephone interview. They were asked about health, disability and achievements in terms of living independently, driving a car and working in open employment. Causes of death for those who had died were obtained from medical records and from the Office of National Statistics. The study was approved by the Cambridge Local Research Ethics Committee.2.When first surveyed at the mean age 4 years, the cohort had been classified into four groups according to sensory level to pin prick recorded in infancy [Ascertainment was 100%. Twenty (37%) of the 54 survivors responded to the questionnaire and all survivors or a carer or relative were interviewed by telephone.Figure Of the original 117 babies there were 54 survivors (46%) age range 31–38 years. Of these 24 were male and 29 female and there was one who had undergone a gender change from male to female. The disability of the survivors ranged from blindness with paraplegia and double incontinence to apparent normality. Table Only 16 (30%) remained community walkers defined as being able to walk ≥50 metres with or without aids. Ten of the 16 could walk at least a kilometre. Table Of the 54 survivors, eight (15%) never had a shunt, seven of whom had little or no disability and a sensory level below L3. The remaining 46 had had a ventriculo-atrial shunt inserted. In 16 the shunt had never been revised. The other 30 had had a total of 104 revisions: for shunt insufficiency (65), infection (15), detachment (14), extrusion or leaking wound of back (5), unknown (5). In 9 patients revisions were done only before the age of 2 , and in 21 between the ages of 2 and 35 . Elective revisions were not performed; and shunts were inserted or revised only in response to definite clinical need. Of those who had revisions, 75% had had symptoms of raised intracranial pressure.Nearly half of the survivors had been in hospital during the previous 5 years. The main reasons were urological (7 patients), neurosurgical (3), and sepsis (7). Pressure sores were responsible for four of the admissions for sepsis, and 12 patients were currently being treated at home for pressure sores. Only 11 patients (20%) were fully continent of bladder and bowel without the use of catheters or appliances. Two patients needed nocturnal respiratory support, two were totally blind following shunt dysfunction and four others had severe visual defects. Endocrine conditions were common: two patients had diabetes mellitus, one had adrenal hyperplasia, one had primary azoospermia and six had had precocious puberty. Twenty-four patients were on long-term therapy: antihypertensives (12), anticonvulsants (10), antibacterials (10) and antidepressants (4). Eight patients needed regular analgesics for musculo-skeletal pain, mainly backache.Seven women and two men had become parents. One man had minimal disability and no detectable sensory loss; the other had undergone percutaneous epididymal sperm aspiration followed by intracytoplasmic sperm injection. None of the 13 children had visible spina bifida.Twenty-two individuals (41%) lived independently in the community, 11 of them used wheelchairs. A further 12 (22%) were personally independent but had supervision and help when required. The remaining 20 37%) needed help daily for dressing, shaving, toilet or nursing care (mainly pressure sores). Ten of these still lived with a parent now aged 52–77, two women were in the care of their partners, five were in residential establishments and three lived in the community with help from social services (Table 7% neededTwenty-nine (54%) of the survivors had passed the driving test, but 9 had discontinued driving. Eleven others were unfit to drive on account of poor sight (3), epilepsy (3) or severe cognitive or perceptual defects (5).Nine men and four women were in open employment. All had an IQ ≥80 and five used wheelchairs. Three did clerical work, three were teachers, two were unskilled manual workers and the remainder were a business executive, accountant, engineer, van driver and builder. Three were studying in addition to working full time. Three men and two women were in sheltered employment.Twenty-seven survivors 50%) had one or more achievements in terms of living completely independently in the community (22), driving a car (20) or working in open employment 13). Achievements were related to sensory level in infancy and to shunt history  are unlikely to be seriously disabled and could be achievers in adulthood. Babies who cry during a routine heel prick have a sensory level of S1 or below and are likely to remain community walkers in adulthood. Those with sensation to pin prick in the saddle area  are likely to have bladder and bowel control.Events in the history of the CSF shunt also had a profound effect on outcome and achievement. Revisions of shunt were associated with poor achievement particularly when the revisions were needed after the age of 2. The cranial sutures have usually fused by the age of 2 after which the skull is less expansile rendering the brain more susceptible to pressure . Table 5The main strength of the study is the community basis, which provides social as well as clinical data enabling the realities of adulthood to be seen against the optimistic forecasts of the early years . Less thAlthough outcome in childhood of early operated spina bifida has been widely reported -5,11, thThese data may help health professionals counselling parents of a child with spina bifida. They show a range of possible outcomes in adulthood when parents may no longer be able or willing to look after their child -16. Two The authors declare that they have no competing interests.


Mood disorders are a major public health problem in the United States as well as globally. Less information exists however, about the health burden resulting from subsyndromal levels of depressive symptomatology, such as feeling sad, blue or depressed, among the general U.S. population.As part of an optional Quality of Life survey module added to the U.S. Behavioral Risk Factor Surveillance System, between 1995–2000 a total of 166,564 BRFSS respondents answered the question, "During the past 30 days, for about how many days have you felt sad, blue, or depressed?" Means and 95% confidence intervals for sad, blue, depressed days (SBDD) and other health-related quality of life (HRQOL) measures were calculated using SUDAAN to account for the BRFSS's complex sample survey design.Respondents reported a mean of 3.0 (95% CI = 2.9–3.1) SBDD in the previous 30 days. Women  reported a higher number of SBDD than did men . Young adults aged 18–24 years reported the highest number of SBDD, whereas older adults aged 60–84 reported the fewest number. The gap in mean SBDD between men and women decreased with increasing age. SBDD was associated with an increased prevalence of behaviors risky to health, extremes of body mass index, less access to health care, and worse self-rated health status. Mean SBDD increased with progressively higher levels of physically unhealthy days, mentally unhealthy days, unhealthy days, activity limitation days, anxiety days, pain days, and sleepless days.Use of this measure of sad, blue or depressed days along with other valid mental health measures and community indicators can help to assess the burden of mental distress among the U.S. population, identify subgroups with unmet mental health needs, inform the development of targeted interventions, and monitor changes in population levels of mental distress over time. Mood disorders are a major public health problem in the United States as well as globally, imposing a substantial burden of disability, impaired quality of life, and death if they remain untreated -3. NatioMental health disorders due to depression, anxiety and substance use are not only burdensome by themselves, but they can complicate existing physical disorders and also increase risk for other physical comorbidity ,12,13. FFor at least one year since 1995, more than one-third of state health departments have assessed the number of recent days that adults "felt sad, blue or depressed" using the Behavioral Risk Factor Surveillance System (BRFSS). This measure has good construct validity when compared with other BRFSS health-related quality-of-life (HRQOL) domains related to mental health , and hasUsing a large multi-state sample, this study is the first to focus on the prevalence of self-reported "sad, blue or depressed" days (SBDD) overall and in sociodemographic subgroups in the United States. It also examines the construct validity of the measure.The BRFSS, which is designed to monitor behavioral health risks in the United States, is an annual random-digit-dialed telephone survey of the non-institutionalized civilian population aged 18 years or older conducted in all states and the District of Columbia . As part2), normal weight (18.5< 25.0 kg/m2), overweight (25.0 to < 30.0 kg/m2), or obese (≥ 30.0 kg/m2) [Respondents answered standard BRFSS questions about age, race/ethnicity, education, employment, income, marital status, health status, physical health, mental health, activity limitation, access to care, and the presence of certain health conditions such as hypertension and diabetes. Respondents also answered questions about how often they engaged in behaviors risky to health, such as smoking and binge drinking, and in health promoting behaviors, such as using a seat belt and exercising. Each respondent's body mass index (BMI), (weight in kilograms divided by the square of height in meters), was classified according to the National Institutes of Health criteria as either underweight (<18.5 kg/m0 kg/m2) .Individual responses were weighted to reflect the age and sex distribution of each state's population during each survey year. To account for the BRFSS's complex sample survey design, means (M) and 95% confidence intervals (CI) for SBDD and other HRQOL measures were calculated using SUDAAN . Because mean SBDD varied by five-year age groups, the analyses were directly standardized to the age distribution of adults aged 18 years or older from the 2000 U.S. Census to control for confounding by age.Respondents reported a mean of 3.0 (95% CI = 2.9–3.1) SBDD in the previous 30 days. About 43.4% of respondents reported one or more SBDD including 7.9% who reported 14 or more SBDD. Women  reported a higher number of SBDD than did men  , whereas Hispanics, Blacks, American Indians and Alaska Natives, and non-whites of another race/ethnicity reported about 4–5 SBDD; in the last three of these groups, the gaps between men and women were larger. Adults with more education reported fewer SBDD, with the gap between men and women diminishing with more education. Respondents who were unemployed or unable to work reported more SBDD than the employed. The number of SBDD decreased with increasing levels of annual household income. Widowed or separated adults reported about 6 SBDD, whereas respondents who were currently married reported the fewest number of SBDD (M = 2.5); the gap between men and women was least among those who had never married.SBDD was associated with an increased prevalence of behaviors risky to health, extremes of BMI, less access to health care, and worse self-rated health status. Respondents who reported physical inactivity, binge drinking, seldom-use or nonuse of seatbelt, or any or more cigarette smoking reported substantially higher numbers of SBDD than those who did not report engaging in these risky behaviors Table . SubjectSBDD were associated with other physical and mental HRQOL domains in expected ways. Mean SBDD increased with progressively higher levels  of physically unhealthy days, mentally unhealthy days, unhealthy days, activity limitation days, anxiety days, pain days, and sleepless days  associated with engaging in healthful behaviors . PerceivOur findings support the construct validity of the SBDD measure in this study because SBDD were associated with other physical and mental HRQOL domains in expected ways. Groups with progressively higher numbers of physically unhealthy days, activity limitation days, and pain days reported a higher number of SBDD. Moreover, these associations were more pronounced with mentally unhealthy days and anxiety days, than with physically unhealthy days, activity limitation days, and pain days. We found an exception to the linear relationship between SBDD and HRQOL measures with our measure for sleeplessness. Adults who reported 1–2 days of sleeplessness reported fewer SBDD than those who reported no days of sleeplessness. Sleep disturbance, both insomnia and hypersomnia are symptoms of depression. Those reporting no days of sleeplessness, but more SBDD, might be those with hypersomnia. Additional studies are warranted to examine this hypothesis.Besides the cross-sectional design, this study has other limitations. Only 38 states and the District of Columbia included the HRQOL supplemental module that assessed SBDD. All states and the District of Columbia, however examined mentally unhealthy days–the number of days respondents experienced poor mental health due to stress, depression or problems with emotions. Mean mentally unhealthy days in the states that assessed SBDD with the HRQOL supplemental module did not differ significantly from that in states that did not. Given the positive correlation between mentally unhealthy days and SBDD (r = 0.6), states that did not assess SBDD would most likely report similar SBDD as states that did include this measure, suggesting similar study results had all states assessed SBDD. Second, BRFSS excludes people who do not have telephones, live in institutions, and persons younger than 18 years. Third, BRFSS may under represent the severely impaired because functional capacity is required to participate in BRFSS. Including this group however, would probably only strengthen the associations we found because the variability of SBDD would increase because the severely impaired would be more likely to report more SBDD. Finally, because our findings on SBDD are based on respondents' self-reports rather than on professionally administered psychiatric evaluations, people who experience SBDD may differ from people with clinical depression.The 1999 Surgeon General's report states that mental health and mental illness "are not polar opposites but may be thought of as points on a continuum" . Use of Future research might examine in more detail the associations among SBDD, anxiety, vitality, and sleeplessness and their ability to assess mood, anxiety, and sleep disorders. It would also be useful to examine the prevalence and demographic characteristics of those who report 14 or more SBDD, and the criterion validity of this measure with other screening instruments and clinical assessments. While SBDD does not provide a strict measure of diagnosable depression as would validated screening and diagnostic assessments, SBDD and other measures such as activity limitations, alcohol or substance abuse, physical inactivity, and employment status can be useful community indicators for addressing the prevention and treatment of depressive symptoms and associated comorbidity .SBDD sad, blue or depressed daysHRQOL health-related quality of lifeBRFSS Behavioral Risk Factor Surveillance SystemCDC Centers for Disease Control and Prevention


Tracheal tumors are often misdiagnosed as asthma and are treated with inhaled steroids and bronchodilators without resolution.Here, a patient with tracheal adenoid cystic carcinoma who had been previously diagnosed with difficult asthma was reported. The possibility of the presence of localized airway obstruction was raised when the flow-volume curve suggesting fixed airway obstruction, was obtained.The presenting case report emphasizes the fact that not all wheezes are asthma. It is critical to bear in mind that if a patient does not respond to appropriate anti-asthma therapy, localized obstructions should be ruled out before establishing the diagnosis of asthma. Tracheal tumors are uncommon and often overlooked until they reach to an advanced stage. The presenting symptoms are typically prolonged cough and wheezing that can be misdiagnosed as asthma . TherefoHere, a patient with tracheal adenoid cystic carcinoma who had been previously diagnosed with difficult asthma was reported.A 29 year-old man was referred to our hospital with a 2-year history of paroxysmal attacks of dyspnea, dry cough and wheezing. He had smoked 2 packs/day cigarettes for 3 years and has been ex-smoker for 5 years. He experienced frequent sudden-onset coughing episodes followed by the development of dyspnea and wheezing a year ago. He was previously diagnosed with difficult asthma and treated with high dose inhaled corticosteroids (1600 μg budesonide) and bronchodilators. Since he was unresponsive to the therapy, he has applied to several institutions for multiple times to seek medical attention.th centimeter distal to larynx. Histopathological diagnosis was adenoid cystic carcinoma of the trachea.On admission, no stridor, wheezing and cyanosis were present and the general appearance was good. Vital signs were as follows: temperature 37°C, respiratory rate 20/min, pulse 96 beats/min, blood pressure 140/70 mmHg. The chest examination was unremarkable. The results of the routine laboratory analysis, including complete blood cell count, chemistry, arterial blood gas, urinalysis and chest x-ray were within normal ranges. On spirometric examination, flow-volume curve displayed suggestive fixed airway obstruction. Forced vital capacity (FVC) was 122 % of predicted, forced expiratory volume in one second (FEV1) was 31 % of predicted and FEV1/FVC was 21 % Figure . FOB revThe patient underwent resection surgery. At the operational site, there were severe adhesions between the mediastinal surface and the trachea. Therefore, a conservative surgery was performed. The tumor was seen on the right anterolateral wall of the trachea being expanded submucosally from the carina to the proximal end of the trachea.The patient underwent adjuvant radiation therapy after the operation. CT scan of the neck revealed resolution of the tumor Figure . Now, 3 Primary tracheal tumors are rare with the incidence of less than 0.1 % ,3. The mClinical manifestations of tracheal tumors are developed as a consequence of tumor bulk and location. Patients with tracheal tumor often have exertional shortness of breath, prolonged cough or a new onset of wheezing, which is frequently misdiagnosed as asthma. Patients are usually initially managed accordingly. However, tumor may occlude 75 % of the lumen before leading symptoms. In the literature, most of the reports highlight that there is always a remarkable delay of establishing accurate diagnosis as a result of misdiagnosis of asthma ,5,7. PeaAnother diagnostic challenge of tracheal tumor is the fact that it can occasionally be visualized by plain chest X-ray. CT scan of the chest or magnetic resonance imaging may yield more valuable data on the site and length of the tracheal lesion . The corIn the presenting report, the patient had been mistakenly diagnosed with difficult asthma because of the presence of uncontrolled asthmatic symptoms and poor lung functions despite the use of high doses corticosteroids. Nevertheless, the flow-volume curve typically displayed localized fixed obstruction of central airways. This led us to have a work-up of CT scan and FOB to rule out upper airway obstructions.Adenoid cystic carcinomas are smooth, firm, and well-circumscribed lesions. These tumors grow extremely slowly. Patients have been known to survive for 10 to 15 years with multiple lung metastases. Spread tends to occur submucosally [The optimal therapeutic approach is surgical resection and reconstruction. The surgeon should be aware of the fact that the apparent gross margin of the tumor is usually still involved with the tumor cells so that the resection should be done at least 1 cm beyond the gross tumor margin. Postoperative irradiation was recommended by most of the authors . Long-teIn this case, postoperative irradiation on curative doses has been applied for a month. Control bronchoscopic examination revealed near-complete remission.The presenting case report emphasizes the fact that not all wheezes are asthma. It is critical to bear in mind that if a patient does not respond to appropriate anti-asthma therapy, localized airway obstructions should be rule out before establishing the diagnosis of asthma.The author(s) declare that they have no competing interests.HT has seen the patient and made the diagnosis of tracheal obstruction, participated in the design of the manuscript. NK has seen the patient in the clinical, followed the patient and drafted the manuscript. SD and CK have performed the surgery and followed the patient. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:


Cognitive traits derived from neuropsychological test data are considered to be potential endophenotypes of schizophrenia. Previously, these traits have been found to form a valid basis for clustering samples of schizophrenia patients into homogeneous subgroups. We set out to identify such clusters, but apart from previous studies, we included both schizophrenia patients and family members into the cluster analysis. The aim of the study was to detect family clusters with similar cognitive test performance.Test scores from 54 randomly selected families comprising at least two siblings with schizophrenia spectrum disorders, and at least two unaffected family members were included in a complete-linkage cluster analysis with interactive data visualization.A well-performing, an impaired, and an intermediate family cluster emerged from the analysis. While the neuropsychological test scores differed significantly between the clusters, only minor differences were observed in the clinical variables.The visually aided clustering algorithm was successful in identifying family clusters comprising both schizophrenia patients and their relatives. The present classification method may serve as a basis for selecting phenotypically more homogeneous groups of families in subsequent genetic analyses. Schizophrenia is a severe mental illness which tends to run in families. Moreover, schizophrenia is a complex disorder with multiple environmental as well as genetic predisposing effects. Previous studies have shown that many neuropsychological functions are impaired in schizophrenia patients, and, to a lesser degree, also in their unaffected relatives -3. ConseRecently, cluster analysis of verbal learning and memory tests was used to divide patients with schizophrenia into subtypes. Categorization by these cognitive traits resulted in meaningful subgroups of schizophrenia . In anotIn the present study, we report a new visually aided clustering approach aimed at identifying clusters of multiply affected families with schizophrenia on the basis of performance in neuropsychological tests. In the clustering process, each family was represented by the test scores of its affected and unaffected members, and the closeness the families was defined by the maximum pairwise distance between the members of the families. To our knowledge, this is the first study in which the clustering has been applied to families instead of solely to affected subjects with schizophrenia.From a general population cohort of people born between 1940 and 1976 inclusive in Finland, a northern European country with approximately 5 million inhabitants, we identified 33,731 individuals with a diagnosis of schizophrenia, schizoaffective disorder or schizophreniform disorder. Data on the diagnosis were derived from three nation-wide computerized health care registers covering the years 1969 to 1998: the Hospital Discharge Register, the Free Medicine Register, and the Pension Register. Linking the personal identification numbers of the affected subjects to the National Population Register database allowed us to identify their family members and to construct pedigrees.Information on families with at least two members with schizophrenia, schizoaffective disorder or schizophreniform disorder, and at least two members with no diagnosis of psychiatric disorder was received from the aforementioned registers for 895 families from the whole of Finland. A blood sample for subsequent genetic analyses was drawn from 2295 subjects of 643 families. All available case note records were collected for those with a diagnosis of schizophrenia, schizoaffective disorder or schizophreniform disorder in any of the three registers. Two psychiatrists independently assessed the lifetime diagnoses for each case, according to the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) . One of n = 6), having a current substance use diagnosis (n = 6), or being mentally retarded (n = 4). Of the family members to whom no psychiatric diagnosis was assigned for their lifetime, 6 were excluded because of high age, or for a defect in vision or hearing. The final sample thus comprised 165 subjects with a psychiatric diagnosis and 94 unaffected family members from 54 families. Of the 165 subjects with a diagnosis, altogether 82 subjects had schizophrenia, while 13 subjects suffered from schizoaffective disorder, 10 from schizophreniform disorder and 12 from bipolar disorder. A nonpsychotic disorder was assigned to 48 individuals. The 94 unaffected subjects did not get any current or lifetime psychiatric diagnosis. In 51 families, at least one of the patients included in the analysis suffered from pure schizophrenia. In the remaining three families, at least one subject with schizoaffective or schizophreniform disorder was included. All families from which the subjects for the present study were drawn, represent familial schizophrenia, as in each of them there were at least one sibling with a diagnosis of pure schizophrenia, plus at least one other sibling with schizophrenia, schizoaffective disorder or schizophreniform disorder.Of those multiply affected families who already had given the blood samples, a subsample was targeted for collection of more detailed phenotypic information. This sample was selected randomly based on the data from the registers and the OPCRIT process. All subjects from the families gave a written informed consent for the study protocol comprising a diagnostic research interview and neuropsychological testing. Both patients and their family members were interviewed using the Structured Clinical Interview for DSM-IV (SCID-I for axis I disorders and SCID-II for axis II disorders) . All theA neuropsychological test battery was administered to all subjects in fixed order by well-trained examiners either after the interview during the same day, or the following day. All examiners were psychologists or advanced psychiatric nurses extensively trained and supervised with the test battery. Experienced psychologists scored all the tests.Auditory attention was assessed with the Digit Span Forward task, and verbal working memory with the Digit Span Backward task of the Wechsler Memory Scale-Revised (WMS-R) . AccordiThe Visual Span forward subtest of the WMS-R  was usedVerbal learning and memory were assessed with the California Verbal Learning Test (CVLT)  which exControlled Oral Word Association test (COWA)  was usedFour subtests of the Wechsler Adult Intelligence Test – Revised (WAIS-R)  were useM = 19 variables and N = 259 subjects, the data formed an M × N matrix x = (xik), where xik is the the value of the ith variable for the kth subject. However, there were 85 (1.7 %) missing values as not all test results were obtained for all subjects. The missing values were handled by the following procedure, which replaces an individual's missing value with an estimate obtained from a linear fit between the test with the missing value and the test that correlates with it most and that also has the individual's test result available.The variables used in cluster analysis included 17 neuropsychological test variables together with the age and the sex of the subjects. With a total of i and j by cij.1. Pairwise correlations were calculated between all test variables using only subjects with results available for both tests. We denote such correlation between the tests i for the subject k, we found the test j = j0 which had the highest value of |cij| among the tests with the value xjk available and set2. Given a missing value in the test b were found by computing linear regression of the test j0 on the test i using only subjects with results available for both tests.where the coefficients a and xk =  the normalized data for subject k and define the distance between two clusters Cr and Cs byThe families were clustered using a complete-linkage clustering algorithm. Each variable was normalized by subtracting the mean value and dividing by the standard deviation. The normalization was done to ensure that each variable contributes equally to the clustering procedure. Denote by drs = max{||xk - xl|| : xk ∈ Cr and xl ∈ Cs},drs is the maximum pairwise distance between members of the two clusters. Here ||·|| denotes the euclidean distance, . In the sense of this distance measure, two clusters are close when all subjects in both clusters are close.that is, Clustering was carried out using the following algorithm.1. Initial clusters are defined by the families.drs are merged into one larger cluster.2. The two clusters with the smallest inter-cluster distance 3. Steps 2 and 3 are repeated until a desired number of clusters remains.drs is shown by the solid red line. The inter-cluster distance between the two remaining clusters is shown by the dashed red line. Note that by using a different inter-cluster distance measure, such as the minimum pairwise distance, a different merging order would result (see the Discussion).In Figure We introduce a visualization technique that helps in identifying candidate clusters and also gives an overall picture of the main differences between the produced clusters as measured by all variables simultaneously. The method gives information about the dynamics of the clustering process and the characteristics of the candidate clusters. The upper part of Figure xi =  and the pairwise distance between two clusters Cr and Cs was defined asTo further improve the visual impression of the clusters, the neuropsychological test variables (the rows of the data image) were ordered using essentially the same procedure that was used in clustering the families. The initial clusters were now the individual variable vectors drs = max{1/|cij| : i ∈ Cr, j ∈ Cs},cij denotes the correlation between the variables i and j. Thus, at each step, the algorithm merged clusters with the highest correlating variables.where The lower part of Figure BWr,s = Br,s/(Ws + Wr), where Br,s is the mean distance between subjects from clusters r and s  and Wr is the mean distance between subjects within cluster r. The statistic BWr,s takes on a large value if the distance between the subjects from the different clusters is large compared to the distance between the subjects within the clusters themselves indicating that the two clusters are separated in the 19-dimensional space defined by the variables used. If the values of BWr,s for the proposed three clusters are significantly higher than for a random partition we take this as evidence that the clusters found indeed constitute meaningful groups also at the level of individual subjects.The clusters were obtained by treating families as single objects whose dissimilarity was measured by the pairwise test performance differences between the family members. One may therefore ask whether the clusters found still appear to be distinct groups when viewed simply as sets of individual subjects. We examined this question by dividing repeatedly the 54 families into three random clusters that had the same number of families as in the proposed three cluster solution and by computing, for each of the three pairs of the generated clusters, the ratio a priori, came from the same families. Thus, family was included as a random effect in all models with age and sex as the fixed effects. In addition, post hoc models were conducted with education years as an added fixed effect, a known confounder for cognitive functions. In all these analyses, the probability level < 0.05 indicated statistical significance. Analyses were performed using the S-Plus statistical software, version 3.4 [Further, after the cluster analysis, the proposed family clusters were examined for differences on demographic and neuropsychological measures. In addition, the patients included in the clusters were examined for the differences in clinical variables as evaluated by the OPCRIT  of the disorder. In comparing the demographic and clinical variables, the Chi-square test, or t-test, both two-tailed, were applied. The differences in the quantitative neuropsychological measures were analyzed using the linear mixed effects (LME) model, which takes into account the dependence between the subjects, who, sion 3.4 .Three clusters of families were successfully identified from the study sample. The first cluster comprised 94 subjects from 17 families, the second cluster 50 subjects from 12 families, and the third cluster 115 and 25. Adding more neuropsychological test variables or leaving out the sex or the age of the subjects had little effect on the solution.The data image Figure  indicateA three cluster solution is supported by the homogeneity of the within-cluster test performance patterns of the proposed groups Figure . As showIn Figure BWr,s the randomly generated clustering solution almost always had a smaller value than the proposed clustering solution . This lends support to the visual impression that the three clusters are separate groups when viewed as subsets of individual subjects. Results were similar when the family structure was ignored and the random clusters were generated allowing subjects from the same family to be assigned to different clusters.In each of the three pairwise comparisons of the statistic p <0.001 in contrasts versus both other clusters). Overall, the clusters did not differ in clinical characteristics, except that the well-performing cluster showed better premorbid adaptation than the intermediate cluster (p = 0.04). The age of onset did not differ between the clusters . The impaired cluster did not include any patients with schizoaffective disorder, bipolar disorder or other affective psychotic disorders, while in the well-performing and intermediate clusters, these diagnoses were assigned to 14% and 11% of the subjects, respectively. About 36% of family members in all three clusters were unaffected.The demographic characteristics of the clusters of families are shown in Table The impaired cluster scored lowest in all measured neuropsychological variables, and the intermediate cluster showed consistently worse performance than the well-performing one Table . The difAs the clusters differed significantly from each other in education years, we conducted post hoc linear mixed effects models with family as the fixed effect, and age, sex and education years as the random effects (data not shown). This did not eliminate the significant differences in cognitive functioning between the well-performing and the impaired cluster. In contrasts between the well-performing and intermediate cluster, all other differences remained significant, except in the scores of Visual immediate recall, Digit Symbol, and Verbal fluency, which lost their significance. Between the intermediate and the impaired cluster, scores in Vocabulary and Digit Symbol were no longer significantly different after controlling for education years.We report on the application of a visually aided clustering algorithm to data based on performance in a set of neuropsychological test measures, these being potential endophenotypic traits in schizophrenia. We were able to successfully detect three separate family clusters comprising both schizophrenia patients and their family members. In the impaired cluster, the families scored significantly worse than those in the other two. The well-performing cluster received the highest scores in each cognitive test, and the intermediate cluster scored consistently between the other two. However, the clusters of families did not differ from each other in age, sex distribution, and, regarding the affected subjects, in the age of onset, or in most of the other clinical features. The well-performing cluster was significantly more educated than the two others, but controlling for education years did not change the main results.We tested the differences in the diagnostic class distributions (including those with no diagnosis), and although the differences did not reach statistical significance, we find it interesting that none of the subjects with schizoaffective disorder, bipolar disorder, or other affective psychotic disorders ended up into the impaired cluster. We consider this as supporting the validity of particularly the poor cluster, which seems to represent a subsample of core schizophrenia with the most defected cognitive functioning. This cluster included the same proportion of unaffected subjects than the other two clusters, and based on the clustering algorithm, these family members without any psychiatric diagnoses during their lifetime performed generally poorly, too.Global verbal memory, including the story recall from the WMS-R  and verbOur results suggest that molecular genetic analyses could benefit from prior appliance of our method, revealing meaningful family subgroups in a representative sample of familial schizophrenia. It would allow the resources to be targeted primarily for gene hunting projects among more homogeneous groups of families. Our new approach to combining data visualization and clustering appears to offer a valuable tool for identifying clusters in family-based data. Applying hierarchical clustering and the data image interactively helps to identify a reasonable value for the number of clusters in the cluster solution. By ordering the variables in the data image suitably, one gains useful insight into the test performance characteristics of the subjects in the clusters.As suggested in Palmer et al. , there mex post facto. Furthermore, our set of neuropsychological measures did not cover all those cognitive domains that previous studies have suggested as valid cognitive endophenotypes. However, it has been demonstrated in twin and in family studies [The present study is the first one in which cluster analysis of neuropsychological test variables has been conducted among a representative sample of familial schizophrenia comprising both affected and unaffected family members. The sample of the present study was randomly selected from a nationwide familial schizophrenia cohort. However, the results may not be generalizable to families with only one patient with the disorder. The similar patterns in neuropsychological performance in the clusters may be due to a variety of familial environmental effects, which are difficult to define  studies ,7,34, thIn the absence of a control sample, the present study could not test the possibility that the same clustering solution would emerge in normal families from the population. However, to our knowledge, such family clustering studies have not been conducted. In a study by Horan and Goldstein , a clustClearly, the choice of the inter-cluster distance measure can greatly influence the merging process and hence the cluster solutions obtained. The maximum pairwise distance between subjects adopted in our analysis assigns a small distance between clusters only if all subjects in the clusters are close to each other in their test performance. We also experimented with the minimum pairwise distance but the results were poor. An explanation for this emerges by studying the minimum distance measure along the first few principal component directions of the normalized test results (the directions of largest variance). It turns out that, along these directions, most families have a member with nearly average performance and whose test results therefore closely match those of many members of other families. The variance of the distribution of the pairwise minimum distances is small and modest changes in the test results can lead to significantly different cluster solutions. The mean of pairwise distances between two clusters would be a compromise between the maximum and the minimum distances, but it turned out to behave much like the minimum distance and was therefore not used.The new approach which combines clustering and data visualization was effective in identifying homogeneous subgroups of schizophrenia families with convergent cognitive test performance. Our results emerging from a sample of familial schizophrenia patients are in line with previous studies in which two extreme clusters have consistently emerged, characterized by a well-performing and a dysfunctional group of subjects, and at least one intermediate ,12,37. ONone declared.FH and LH made the cluster analysis and wrote the corresponding sections, ATH supervised the neuropsychological test data collection, performed the differences testing analyses and drafted the manuscript, TP was the clinical and diagnostic study leader and commented on the text, JH designed the models for differences testing, JL conceived the research project.The pre-publication history for this paper can be accessed here:Comparison of Cluster 1 (well-performing) and Cluster 2 (impaired). Theoriginal variables  are visualizedusing the parallel coordinate plot. The subjects classified aswell-performing are colored green and those classified as impaired arecolored red. Yellow indicates overlap of green and red. There is overlap inall the original variables.The above figure was produced using the Crystal Vision software. Thesoftware uses a so-called grand tour technique to systematically (and in acontinous manner) go through all possible rotations in the data space. Thetour can be visually monitored using the parallel coordinate plot.Click here for fileSmaller version of Additional file 1.Comparison of Cluster 1 (well-performing) and Cluster 2 (impaired). Theoriginal variables  are visualizedusing the parallel coordinate plot. The subjects classified aswell-performing are colored green and those classified as impaired arecolored red. Yellow indicates overlap of green and red. There is overlap inall the original variables.The above figure was produced using the Crystal Vision software. Thesoftware uses a so-called grand tour technique to systematically (and in acontinous manner) go through all possible rotations in the data space. Thetour can be visually monitored using the parallel coordinate plot.Click here for fileA parallel coordinate plot of the data after a rotation transformation inthe data space. The axes correspond now to linear combinations of theoriginal variables. The 6th axis from the top shows an interestingone-dimensional projection of the data. In this direction the two clustersare well separated.The above figure was produced using the Crystal Vision software. Thesoftware uses a so-called grand tour technique to systematically (and in acontinous manner) go through all possible rotations in the data space. Thetour can be visually monitored using the parallel coordinate plot.Click here for fileSmaller version of Additional file 3.A parallel coordinate plot of the data after a rotation transformation inthe data space. The axes correspond now to linear combinations of theoriginal variables. The 6th axis from the top shows an interestingone-dimensional projection of the data. In this direction the two clustersare well separated.The above figure was produced using the Crystal Vision software. Thesoftware uses a so-called grand tour technique to systematically (and in acontinous manner) go through all possible rotations in the data space. Thetour can be visually monitored using the parallel coordinate plot.Click here for fileReference for Crystal Vision softwareCrystalVision, copyright (c) 2000 by Crystal Data Technologies , is a Windows 95/98/NT package for Wintelcomputers. A demonstration version of CrystalVision is available at A movie which demonstrates the use of the software can be downloaded here.Smaller resolution CrystalVision.mpg(0.86 Mb). A higer resolution version ofthis movie is available at Click here for file