African-Americans remain underrepresented in clinical research despite experiencing a higher burden of disease compared to all other ethnic groups in the United States. The purpose of this article is to describe the study design and discuss strategies used to recruit and retain African-American smokers in a pharmacokinetic study.The parent study was designed to evaluate the differences in the steady-state concentrations of bupropion and its three principal metabolites between African-American menthol and non-menthol cigarette smokers. Study participation consisted of four visits at a General Clinical Research Center (GCRC) over six weeks. After meeting telephone eligibility requirements, phone-eligible participants underwent additional screening during the first two GCRC visits. The last two visits (pharmacokinetic study phase) required repeated blood draws using an intravenous catheter over the course of 12 hours.Five hundred and fifteen African-American smokers completed telephone screening; 187 were phone-eligible and 92 were scheduled for the first GCRC visit. Of the 81 who attended the first visit, 48 individuals were enrolled in the pharmacokinetic study, and a total of 40 individuals completed the study (83% retention rate).Although recruitment of African-American smokers into a non-treatment, pharmacokinetic study poses challenges, retention is feasible. The results provide valuable information for investigators embarking on non-treatment laboratory-based studies among minority populations. Although African-Americans experience a disproportionately high burden of disease ,2, they Tobacco-related morbidity and mortality is higher among African-Americans compared to other racial/ethnic groups in the United States . HoweverThe study protocol was reviewed and approved by the University of Kansas Medical Center Human Subjects Committee. This study was conducted within the General Clinical Research Center (GCRC) at the University of Kansas Medical Center.®, Glaxo SmithKline) by evaluating the effects of menthol in mentholated cigarettes on the pharmacokinetic (PK) profiles of bupropion and its three principal metabolites: hydroxybupropion, threohydrobupropion, and erythrohydrobupropion. The study aimed to recruit and enroll 20 African-American smokers of mentholated cigarettes (menthol smokers) matched 1:1 with 20 African-American smokers of non-mentholated cigarettes (non-menthol smokers) by gender, number of cigarettes per day smoked (CPD), and body mass index (BMI). Pharmacokinetic (PK) parameters of bupropion and its three principal metabolites were assessed at steady state under smoking and non-smoking conditions. During the smoking condition which lasted 10-15 days, subjects smoked their usual brand of cigarettes. This period was followed by a non-smoking condition lasting another 10-15 days. The PK parameters were then assessed between 1) menthol and non-menthol smokers, and 2) smoking and non-smoking conditions.Figure Participants were recruited using clinic-based and community-based strategies. Clinic-based strategies included informing medical center staff about the study through flyers, broadcast emails, departmental meetings, and employee newsletters. Posters and flyers were distributed at a community health center that serves a predominantly African-American patient population. Invitation letters were sent to former research participants who gave written consent to be contacted in the future. Community strategies included paid advertisements in neighborhood newspapers, the major city paper, and on two radio stations with large African-American audiences. Research staff provided flyers to African-American business owners and religious organizations. Those interested in the study were asked to call the study office and speak with a study coordinator who assessed their eligibility for the study.2. Additionally, each participant must have smoked either mentholated or non-mentholated cigarettes exclusively for the past year. Consistent with contraindications for bupropion use, exclusion criteria included predisposition to seizures, a diagnosis of bulimia or anorexia nervosa in the past year, an unstable medical or psychiatric illness, alcohol dependency within the last year, or a myocardial infarction in the last month. In addition, individuals who planned to move from the metropolitan area in the next month, or who had used other forms of tobacco in the past 30 days, or had used bupropion in the past 30 days were excluded from participation, as were those currently using any prescription or other medications contraindicated or with known interaction with bupropion, those reporting illicit drug use and women who were pregnant, breastfeeding, or contemplating pregnancy in the next month.Eligible individuals identified themselves as African-American or Black, aged 18 years or older. They smoked at least 10 cigarettes per day and had a BMI between 18 and 45 kg/mEligibility screening was multi-staged: telephone eligibility screening was followed by medical screening, and then, lastly, by screening for adverse events with medication and adherence to medication.Study staff specifically informed callers that this was a "non-treatment study to find out how the body breaks down a medication called Zyban". Research staff verified individuals' willingness to adhere to study instructions. Interested individuals were screened on the phone according to the eligibility criteria described above. Those eligible were informed that medical screening would include an additional medical history, a physical examination and laboratory tests to rule out contraindications to bupropion use. They were instructed to fast overnight prior to the medical screening visit. A written copy of these instructions was mailed to all individuals who had passed the telephone screening.Phone-eligible individuals came to the General Clinical Research Center (GCRC) after an overnight fast. The study staff discussed the details of the study with participants during the informed consent process. Individuals signed a written informed consent form. They provided a 20 ml blood sample for a complete blood count, a complete metabolic profile, and cotinine analysis. Women also gave a urine sample for a pregnancy test. The GCRC nurse took a medical history and examined the participants. Participants completed a baseline smoking history questionnaire and also completed a smoking topography using a CreSSmicro smoking topography device . The study staff and the study physician reviewed the results of the laboratory tests, medical history and physical examination findings to determine eligibility. Eligible participants were given a sterile container for collecting urine beginning 24 hours prior to the next visit to verify their menthol status. They were also given a special cigarette carrying container (Smartpak) to use during the study.Medically eligible participants were given a 7-10 day supply of placebo in a container with a Medication Event Monitoring System (MEMS) cap, an adherence verification device . On their second visit, participants were asked about adverse events using the Common Terminology Criteria for Adverse Events, version 3.0 . Unused study medication was collected and counted, and data were downloaded from the MEMS cap. Individuals who did not tolerate the placebo medication or used less than 75% of the prescribed dose were excluded from continued participation. Such individuals were excluded with the rationale that they were unlikely to take the active medication as prescribed during the entire study. Participants were enrolled in the pharmacokinetics phase of the study after they passed the medical adherence eligibility screening (Visit 2), and were scheduled for Visit 3. The participants continued to use the MEMS cap for the entire study to monitor bupropion use. The study staff instructed participants to bring their MEMS cap and all medications to each GCRC visit.ad lib for the first 10-15 days (smoking condition) and to quit smoking for the remaining 10-15 days of the study (non-smoking condition). Blood samples were drawn for pharmacokinetics (PK) analysis on two occasions, 10-15 days after the commencement of bupropion while participants were still smoking (PK 1), and again at days 20-25 (PK 2) when they were required not to smoke (non-smoking condition). The blood samples at Visit 3 (PK 1) provided PK parameters when participants were exposed to both bupropion and menthol in cigarettes. Samples at Visit 4 (PK 2) provided PK parameters when participants were exposed only to bupropion but not to menthol. At each PK visit, approximately 10 ml of blood specimens for PK were taken through an intravenous line inserted into the participant's arm prior to ingestion of 150 mg bupropion-SR and at 1, 2, 2.5, 3, 3.5, 4, 5, 6, 8, and 12 hours after ingestion of the first dose of 150 mg bupropion-SR. Use of the second dose of bupropion for the day was delayed until after all blood draws were completed.After a seven-day placebo run-in period and an initial three-day dosing period of 150 mg/day, participants were given 300 mg/day (150 mg 2×/day) sustained-release bupropion for 20-25 days. Participants were asked to smoke their usual brand of cigarettes Once participants were scheduled for visits, reminder letters were mailed one week before each visit, and reminder telephone calls were placed two days and one day before each visit. The study staff provided information about the procedures that participants would undergo in the upcoming visit and answered questions. Participants who missed an appointment were contacted by telephone to reschedule the appointment, provided they were still within the study visit window. The research team met weekly to review recruitment and retention progress and to provide feedback to the study staff.Monetary compensation was discussed with participants during the informed consent process. Participants were given $50 Visa gift cards for each screening visit and $150 Visa gift cards for each pharmacokinetic study visit. Fifty dollar Visa gift cards were given to the participants for the use of Smartpak and MEMS devices. Additional $50 Visa gift cards were given to participants who were able to abstain from smoking during the non-smoking phase (verified with Nic check).This is a descriptive summary and, as such, we summarized categorical variables by frequencies and percentages and continuous variables by means and standard deviations using SAS statistical package, version 9.1.Recruitment was conducted from February 2006 to February 2007, during which time a total of 515 African-American smokers were screened via telephone. Interested individuals learned about the study from newspapers (58.7%), word of mouth (14.6%), and flyers (8.5%) See Figure In total, 92 smokers were scheduled for Visit 1, and 81 kept their appointments. Figure Table This study demonstrated that African-American smokers can be successfully recruited to participate in an intensive GCRC-based non-treatment study. Given the intensity of participation required for the study, including 24-hour urine collection, the use of cigarette and pill monitoring devices, the number and duration of study visits, and multiple blood draws, achieving this recruitment level was considered a success. This outcome supports the feasibility of including African-Americans in laboratory-based research. Showing that African-Americans will volunteer to enroll in this type of research is important, given the persistent national problem of underrepresentation of African-Americans in clinical studies . The recMultiple factors contributed to recruitment success. Our research team has a track record of successfully engaging African-Americans in research studies, and we applied this experience to our recruitment efforts ,34,37-53The recruitment of African-American non-menthol smokers into our study was particularly challenging. Only about 20% of African-American smokers smoke non-menthol cigarettes -32,56. BThe majority of the participants who were excluded from this study after the informed consent process either did not adhere to medication regimen or did not keep the scheduled appointment. We do not know if failure to attend visit during the abstinence phase was due to the fact that some of the smokers could not stop smoking during this phase. While the exclusion criteria were carefully chosen to remove individuals who seemed unlikely to comply with our protocol, this study also provided insight into issues that warrant further investigation, such as adherence to medication among minority populations, and factors that influence minority participants to not keep the study appointments. Illicit drug use (primarily marijuana) accounted for 19.5% of those who were excluded from study after consenting to participate. Although use of illicit drugs was a clear exclusion criterion, we administered this question in person because of its sensitive nature and the need to get more accurate information along with a certificate of confidentiality on their behalf from the federal government agency for their protection.Our study demonstrates the feasibility of recruiting African-American smokers to a pharmacokinetic study, and identifies unique challenges to recruitment and retention. These challenges include intense eligibility criteria with multiple levels of eligibility, rigorous protocol, and invasive procedures. Despite these, we have demonstrated that recruiting African-Americans in non-treatment studies involving multiple blood draws and follow-up visits can be accomplished effectively. This paper provides valuable information for investigators embarking on non-treatment laboratory-based studies among minority populations.Dr. Ahluwalia is a consultant to Pfizer Inc.All authors have made substantial contributions to the intellectual content of the manuscript as follows: BF, CAB and IO participated in the acquisition of data; BF, LSC and CAB participated in drafting the manuscript; all authors participated in the concept and design of the study, interpretation of data, review and approval of the final version of the manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2288/10/6/prepub


Background and purpose Many investigations on biodegradable materials acting as an antibiotic carrier for local drug delivery are based on poly(lactide). However, the use of poly(lactide) implants in bone has been disputed because of poor bone regeneration at the site of implantation. Poly(trimethylene carbonate) (PTMC) is an enzymatically degradable polymer that does not produce acidic degradation products. We explored the suitability of PTMC as an antibiotic releasing polymer for the local treatment of osteomyelitis.Methods This study addressed 2 separate attributes of PTMC: (1) the release kinetics of gentamicin-loaded PTMC and (2) its behavior in inhibiting biofilm formation. Both of these characteristics were compared with those of commercially available gentamicin-loaded poly(methylmethacrylate) (PMMA) beads, which are commonly used in the local treatment of osteomyelitis.Results In a lipase solution that mimics the in vivo situation, PTMC discs with gentamicin incorporated were degraded by surface erosion and released 60% of the gentamicin within 14 days. This is similar to the gentamicin release from clinically used PMMA beads. Moreover, biofilm formation by Staphylococcus aureus was inhibited by approximately 80% over at least 14 days in the presence of gentamicin-loaded PTMC discs. This is similar to the effect of gentamicin-loaded PMMA beads. In the absence of the lipase, surface erosion of PTMC discs did not occur and gentamicin release and biofilm inhibition were limited.Interpretation Since gentamicin-loaded PTMC discs show antibiotic release characteristics and biofilm inhibition characteristics similar to those of gentamicin-loaded PMMA beads, PTMC appears to be a promising biodegradable carrier in the local treatment of osteomyelitis. Osteomyelitis is characterized by progressive inflammatory bone destruction . In geneSustained high local antibiotic concentrations are currently achieved by the implantation of gentamicin-loaded poly(methylmethacrylate) (PMMA) beads. Although these beads have been used to treat osteomyelitis for many years , their mCurrently, the biodegradable carrier materials most often investigated have been based on poly(lactide) (PLA) and/or poly(glycolide) (PGA) . BiodegrThe use of PLA and PGA implants has been disputed . An impoOne possible candidate material is poly(trimethylene carbonate) (PTMC), a biodegradable polymer that is relatively new in biomedical applications.In contrast to PLA/PGA, the degradation products of PTMC are not acidic . InteresThe objective of this in vitro study was to explore the suitability of PTMC as a biodegradable, antibiotic-releasing carrier, and to compare it with commercially available gentamicin-loaded PMMA beads. The study investigated 2 related issues regarding gentamicin-loaded PTMC: (1) the release kinetics and (2) its ability to inhibit biofilm formation.Polymer-grade 1,3-trimethylene carbonate (TMC) was purchased from Boehringer Ingelheim . Gentamicin sulfate was purchased from Sigma-Aldrich , and analytical-grade chloroform and diethyl ether were purchased from Merck BV . A lipase solution from Thermomyces lanuginosus  was purchased from Sigma-Aldrich and used as received. Antibiotic-loaded poly(methylmethacrylate) (PMMA) beads are commercially available under the name Septopal . Each Septopal bead (diameter: 7 mm) contains 7.5 mg of gentamicin sulfate (corresponding to 4.5 mg of gentamicin base). Staphylococcus aureus 0734  is one of the most commonly involved bacterial strains in osteomyelitis  and was High-molecular-weight PTMC (Mn = 388 × 103 g/mol) was synthesized, purified, and characterized as previously described . 2 solutPTMC precipitates with and without gentamicin were compression-molded at 50°C into films of 500 μm thickness using a laboratory press. Finally, discs with a diameter of 5 mm were punched out of the films.The surface erosion of gentamicin-loaded PTMC was visualized using SEM. A series of SEM micrographs of PTMC discs were taken after immersion in phosphate-buffered saline (PBS), pH 7.0, or in the lipase solution. After 3, 7, and 10 days, discs were taken out of the solutions for SEM evaluation. Discs were sputter-coated with gold/palladium and examined at 2.0 kV in a JEOL field emission scanning electron microscope type 6301F.Gentamicin release was determined at 37°C in PBS (in the absence of lipase) and in a lipase solution, which induces surface erosion of PTMC . 3 gentaGentamicin concentrations were measured using a procedure described by Gentamicin concentrations in lipase solutions could not be determined spectrophotometrically due to interference of the enzyme. An indirect bacterial inhibition assay, described and validated by The extent to which the released amount of antibiotics inhibits the formation of a biofilm is an index of the efficacy of the antibiotic-loaded carrier. To study biofilm formation, 2 types of bacterial growth media were prepared: TSB and a 4:1 mixture of TSB and lipase solution to allow enzymatic degradation of PTMC. 3 non-loaded PTMC discs and 3 gentamicin-loaded PTMC discs were immersed in 10 mL TSB or 10 mL TSB/lipase medium for 2 weeks at 37°C. In addition, 3 gentamicin-loaded PMMA beads were immersed in 10 mL of TSB. Medium was collected and replaced with new medium after 24, 48, 72, 144, 168, 312, and 336 h to obtain samples containing the antibiotic released during the first, second, third, seventh, and fourteenth day.S. aureus biofilms in 96-well plates  under stationary conditions. 2 μL of bacterial inoculum (approximately 1 × 106 CFU/μL) was added to wells containing 198 μL of TSB. After 24 h of incubation at 37°C, the medium was removed and the plates were washed 3 times with 220 μL of PBS. Subsequently, each well was stained with 200 μL of 1% (w/v) crystal violet in water for 30 min. After staining, the plates were gently rinsed 3 times with demineralized water. Quantitative analysis of biofilm formation was performed by adding 200 μL of ethanol/acetone (80:20) to solubilize the crystal violet. The absorbance of this crystal violet solution was measured using a FLUOstar Optima plate reader  at a wavelength of 575 nm. The absorbance (A) is proportional to the amount of crystal violet, which is directly proportional to the amount of biofilm grown in each well.These elution media were used for the formation of The maximal amount of biofilm formation was defined as the amount grown in elution media collected from non-loaded PTMC discs, and the percentage of biofilm inhibition was calculated according to the following equation:For gentamicin-loaded PMMA beads, the maximal amount of biofilm formation was defined as the amount grown in freshly inoculated TSB medium, as non-loaded PMMA beads are not commercially available. All experiments included 6 replicate wells and were performed in triplicate with separately cultured bacteria.In evaluating gentamicin release characteristics and inhibition of biofilm formation, Student's t-test for independent samples was used. A 95%  confidence interval was applied for statistical significance.The surface topographies of PTMC immersed for 3, 7, or 10 days in PBS and in lipase solution were visualized with SEM. Examples of micrographs after immersion for 7 days are shown in The cumulative release of gentamicin from PTMC discs in PBS, PTMC discs in lipase solution, and PMMA beads in PBS is shown graphically in The biofilm inhibition after 24 h in growth medium containing gentamicin released from gentamicin-loaded PTMC discs and PMMA beads during the first, second, third, seventh, and fourteenth day is graphically presented in The inhibition obtained from gentamicin-loaded PTMC discs incubated in lipase-containing growth medium was, however, better than that obtained with similar discs incubated in growth medium only (p < 0.05). After surface erosion in the lipase enzyme solution, gentamicin-loaded PTMC showed continuous, effective inhibition of biofilm formation  similar Gentamicin-loaded PMMA beads constitute an effective drug delivery system for local antibiotic therapy in bone infections. However, after high initial gentamicin concentrations at the site of the infection, the gentamicin concentrations drop considerably . The maiPTMC is such a surface-eroding polymer. It is an aliphatic polycarbonate, and upon degradation of the carbonate linkages by hydrolysis, carbon dioxide and 1,3-propanediol will be formed. Thus, PTMC degrades without the formation of acidic degradation products and has good compatibility with bone tissue. Subcutaneous implantation of PTMC discs in rats showed a mild-to-moderate tissue reaction around the implant . Within PTMC degrades very slowly by hydrolysis in vitro: the polymer is stable in water and in buffered solutions . HoweverInterestingly, the degradation of PTMC showed characteristics of a surface erosion process—as the loss of mass could be correlated to the decrease in thickness of the specimen, and the decrease in molecular weight was limited . This inA crucial point in the treatment of chronic osteomyelitis is the handling of the dead space and gentamicin-loaded PMMA beads can be used to treat the dead space problem. In addition to providing local delivery of antibiotics, these beads will fill the dead space. After the infection is cured, the beads can be removed and the dead space can be filled either with a bone graft or a muscle flap. A specific dead space problem is related to two-stage revision of infected joint prostheses. In these cases, PMMA beads have been used to fill the dead space, i.e. to facilitate the implantation of a new prosthesis. In the examples above, the surgeon uses PMMA beads as non-biodegradable carriers of antibiotics. Accordingly, the use of biodegradable carriers is confined to cases where the dead space problem is considered to be small.The aim of our study was to explore the suitability of biodegradable antibiotic-loaded PTMC discs in the local treatment of osteomyelitis, especially for cases where the volume of dead space is low. In buffer, PTMC released only small amounts of antibiotics during the first 2 weeks, but in a lipase solution (where PTMC degrades by surface erosion), this was accompanied by a high rate of release of antibiotics similar to that observed for non-biodegradable Septopal beads made from PMMA. In line with this, gentamicin-loaded PTMC showed less inhibition of biofilm formation for 2 weeks under non-surface eroding conditions in buffer than under surface-eroding conditions in lipase solution. Moreover, the inhibitory effect of gentamicin-loaded PTMC in the lipase enzyme solution was similar to that of clinically used Septopal beads. This indicates that under conditions that favor surface degradation of the PTMC discs, the polymer could be a suitable biodegradable carrier of antibiotics in the treatment of osteomyelitis. It should be emphasized, however, that a strict comparison of the antibiotic release behavior of Septopal beads and of our PTMC specimens is not possible due to their different geometries, masses, and degrees of gentamicin loading.In this study, macroscopic examination showed that the PTMC discs were completely resorbed within 2 weeks upon immersion in the lipase solution. As PTMC degrades by a surface erosion process, it is possible to extend the degradation time by increasing the thickness of the specimens and by reducing the surface-to-volume ratio. It has also been shown that reducing the initial molecular weight of PTMC polymer has the effect of reducing the rate of degradation . After iA next step would be to initiate animal experiments to confirm the release characteristics and antimicrobial efficacy of gentamicin-loaded PTMC discs as seen in this study. Animal experiments should also be carried out to study the long-term effects of gentamicin-loaded PTMC implants on the regeneration of bone. This is necessary to ensure that PTMC is safe during its degradation in bone, and that it can be used as a drug delivery device in orthopedic applications.In summary, gentamicin-loaded PTMC discs degrading in lipase solution showed antibiotic release kinetics and biofilm inhibition properties that are comparable to those of non-biodegradable gentamicin-loaded PMMA beads. The use of antibiotic-loaded PTMC discs does not require a second surgery for the removal of the beads after therapy; therefore, PTMC-based materials appear to be promising alternative antibiotic carriers for the local treatment of osteomyelitis.


The change in alertness was just correlated with performances on the 2-back task. Activity in the right prefrontal cortex changed depending on alertness changes on the 2- and 0-back tasks independently, which indicates that activity in this region clearly reflects the ability to overcome sleepiness; it may contribute to the function of providing sufficient activity to meet the task load demands. This study reveals characteristics of the ability to overcome sleepiness during the n-back working memory task which goes beyond the attention-control function traditionally proposed.It has been speculated that humans have an inherent ability to overcome sleepiness that counteracts homeostatic sleep pressure. However, it remains unclear which cortical substrate activities are involved in the ability to overcome sleepiness during the execution of cognitive tasks. Here we sought to confirm that this ability to overcome sleepiness in task execution improves performance on cognitive tasks, showing activation of neural substrates in the frontal cortex, by using a modified  Humans may inherently possess the ability to overcome sleepiness when faced with completing assignments accurately or within a given time frame The effects of sleep loss on various cognitive functions have been elucidated by using a simple reaction task Some behaviors for conquering sleepiness have been reported to be able to improve WM performance and the responsible neural activities. Chewing, which has potential alerting effects We hypothesized the following: the ability to overcome sleepiness might enhance WM task performance, helping to resist sleepiness in efforts to carry out a task; the ability to overcome sleepiness might depend on DLPFC activity, which may be shared with attention-control ability; and the ability to overcome sleepiness might show inter-individual differences.n-back WM task with two separate load levels To verify these hypotheses, we used a modified Fifty-five right-handed healthy volunteers participated in this study . Subjects completed a self-assessment questionnaire about their sleep habits in daily life. From the data obtained, habitual awakening time was 07:00–09:00 , habitual sleep onset was 23:00–01:00  and the average habitual sleep duration was 7.86 h (SEM: 0.11 h), as confirmed by a sleep log over a 3-day period prior to the experiment. Subjects participated in an overnight experiment in a laboratory setting, starting at 22:00 on day 1 and finishing at 10:00 on day 2 see . The expAll subjects retired to bed at 01:00 A.M. in the laboratory and were awakened immediately at 07:00 A.M. by an experimenter. The average sleep duration was 5.53 h (SEM: 0.05 h), as confirmed by using an ambulatory wrist activity recorder . Subjects had a restricted sleep duration of 2.32 h (SEM: 0.07 h) on average (29.5%). To avoid the effect of sleep inertia Subjective alertness level was evaluated immediately before and after the experiment using the Stanford Sleepiness Scale (SSS) n-back WM task with two separate load levels. For the 0-back task (low-load WM task), subjects were instructed to press a response button using their dominant hand whenever a single-digit number appeared on a 20-inch LCD computer screen. For the 2-back task (high-load WM task), they had to press a button on the right with their middle finger when the single-digit number on the screen was identical to that which had appeared last but one, otherwise to press the button on the left with their index finger. A 1500-ms baseline period preceded all the tasks. Numbers were presented in randomized order for 300 ms with a 1700-ms inter-stimulus interval. Each level of task was run in blocks of 12+n stimuli and was conducted two times; thus, 24 responses were obtained at each load level. The inter-trial interval was set at 2500 ms. From the performance data recorded, average response times (RTs) and correct response rates (%CRs) were determined.The cognitive paradigm employed in the present study consisted of a visual Changes in [oxy-Hb] and [deoxy-Hb] concentration were continuously measured with a time resolution of 0.1 s using a 22-channel fNIRS machine  at two wavelengths of near-infrared light , and [oxy-Hb] and [deoxy-Hb] levels were calculated based on the modified Beer-Lambert equation as a function of light absorbance of Hb and path length. The distance between the pair of emission and detector probes was 3.0 cm, and it was considered that the machine could measure points at a depth of 2–3 cm from the scalp , that isWe analyzed oxy-Hb data as a reflection of event-related responses in the PFC because change in oxy-Hb concentration is thought to be the most reliable index of regional cerebral blood flow. The continuous oxy-Hb data were filtered with band-pass frequencies in the range of 0.01–0.2 Hz and were standardized (z-score) in order to avoid the methodological ambiguity that changes in absolute values of Hb concentration for each recording channel were not determined, because the absolute path lengths of light through the cerebral cortex were not detectable. Changes in oxy-Hb concentration time-locked to the experimental blocks consisting of 12 trials were extracted for each experimental condition 1000 ms before the onset of the trial block (baseline) to 4400 ms after the onset , which amounted to a total duration of 5400 ms. Baseline correction of the changes in oxy-Hb concentration was performed utilizing the mean z-scores of 1000 ms prior to the onset of the experimental blocks before individual averaging. Mean z-scores of the changes in oxy-Hb concentration during the 4400 ms period after the start of the trial block were utilized for subsequent statistical analyses.To examine the difference in alertness between the pre- and post-experiment, paired-t tests were performed for SSS scores. To examine the difference in WM performance between task levels, paired-t tests were performed for RTs and %CR. To examine the relation between effects of overcoming sleepiness and WM performance, we analyzed Pearson's correlation coefficients of change in SSS level and %CR or RTs for each WM task level separately. To estimate the effects of overcoming sleepiness on the prefrontal activities corresponding to WM characteristics, we analyzed Pearson's coefficients between change in SSS level and the difference of change in oxy-Hb concentration for each channel on the 0- and 2-back tasks. To estimate the effects of overcoming sleepiness on the prefrontal activities corresponding to each WM task level separately, we analyzed Pearson's coefficients between change in SSS level and change in oxy-Hb concentration for each channel on the 0- and 2-back tasks separately.A paired-t test revealed that the mean pre-experiment SSS score  did not differ significantly from that of the mean post-experiment SSS score  . Twelve of 55 subjects showed an increased SSS level , 17 showed a decreased SSS level , and 26 showed no change in SSS level post-experiment compared to pre-experiment.A paired-t test revealed that RT was significantly shorter on the 0-back task  than on the 2-back task  , and %CR was higher on the 0-back task  than on the 2-back task  . Pearson's correlation coefficient revealed that the change in SSS level was negatively correlated with RT on the 2-back task , but not on the 0-back task  , and thePearson's correlation coefficient revealed significant positive correlations between the difference of changes in oxy-Hb concentration on the 2- and 0-back tasks, and changes in SSS level were found for the left hemispheric channels of 4 , 9 , 13 , 18  and 22  and the right hemispheric channels of 1 , 5 , 6 , 10 , 14 , 15  and 19  . PearsonThe subjects who participated in the experiment were confirmed to have similar sleep-wake habits in daily life and they experienced the same restricted sleep duration on the experimental night. However, in the post-task period, alertness was increased in some subjects but was decreased in others despite the subjects having performed the same tasks. Better RTs on the 2-back WM task were associated with the increase in alertness through performing the tasks. In contrast, however, the changes of alertness were not correlated with RTs on the 0-back task. Taken together, these findings indicate that those subjects who improved their alertness, who may have better abilities to overcome sleepiness, showed better performance on the high-load WM task, but this improved alertness did not contribute to better performance on the low-load WM task. However, as there was no significant correlation between %CRs and alertness changes on either the 2- or 0-back task, the RTs may better reflect the influence of the ability to overcome sleepiness upon performance on the current WM tasks. These results may contribute to our understanding of the individual differences in tolerance for shift work or sleep disturbance disorders Changes in oxy-Hb concentration with WM load that correlated positively to changes in alertness were observed in the bilateral DLPFC, as suggested in previous fMRI studies Recent studies have reported that maintaining attention in the evening was associated with higher activity in the evening chronotype than in the morning chronotype Finally, we recognize a limitation regarding the measurement of subjective sleepiness. Although we evaluated the relationship between task performance and subjective sleepiness, a more suitable measure for evaluating subjective sleepiness precisely would seem to be a visual analogue scale of sleepiness, rather than the SSS There remains an important question to be addressed about the relationship between the ability to overcome sleepiness and the attention-control function, which is regarded as one of the core functions of WM. Most studies have indicated a robust structure-function relationship between the left DLPFC and attention-control function


The effects of cis-dichlorodiammine platinum II (DDP) on the intestinal mucosa and the kidney were studied after single and multiple treatments with intervals of 7-45 days. After a single treatment, the jejunal epithelium underwent a transient interruption of cell proliferation followed by a hyperplastic recovery and return to control proliferative rate on Day 7. Subsequent treatments led to suboptimal recovery for all treatment intervals. In contrast, DDP induced a 6-fold increase in [3H]dT incorporation in the kidney by Day 7 which remained high until Day 21, and returned to near-control values by Day 45. After a single DDP treatment, the "recovery potential" of kidneys, measured by the proliferative response to folic-acid stress, demonstrated suboptimal proliferative reserve compartments for up to 45 days. The distinction between acute and delayed sensitivity to subsequent drug treatment was more apparent in the DDP-treated kidney than in the intestinal epithelium.


The benefits of zinc or multiple micronutrient supplementations in African children are uncertain. African children may differ from other populations of children in developing countries because of differences in the prevalence of zinc deficiency, low birth weight and preterm delivery, recurrent or chronic infections such as HIV, or the quality of complementary diets and genetic polymorphisms affecting iron metabolism.The aim of this study was to ascertain whether adding zinc or multiple micronutrients to vitamin A supplementation improves longitudinal growth or reduces prevalence of anemia in children aged 6-24 months.Randomized, controlled double-blinded trial of prophylactic micronutrient supplementation to children aged 6-24 months. Children in three cohorts - 32 HIV-infected children, 154 HIV-uninfected children born to HIV-infected mothers, and 187 uninfected children born to HIV-uninfected mothers - were separately randomly assigned to receive daily vitamin A (VA) [n = 124], vitamin A plus zinc (VAZ) [n = 123], or multiple micronutrients that included vitamin A and zinc (MM) [n = 126].Among all children there were no significant differences between intervention arms in length-for-age Z scores (LAZ) changes over 18 months. Among stunted children (LAZ below -2) [n = 62], those receiving MM had a 0.7 Z-score improvement in LAZ versus declines of 0.3 in VAZ and 0.2 in VA (P = 0.029 when comparing effects of treatment over time). In the 154 HIV-uninfected children, MM ameliorated the effect of repeated diarrhea on growth. Among those experiencing more than six episodes, those receiving MM had no decline in LAZ compared to 0.5 and 0.6 Z-score declines in children receiving VAZ and VA respectively (P = 0.06 for treatment by time interaction). After 12 months, there was 24% reduction in proportion of children with anemia (hemoglobin below 11 g/dL) in MM arm (P = 0.001), 11% in VAZ (P = 0.131) and 18% in VA (P = 0.019). Although the within arm changes were significant; the between-group differences were not significant.Daily multiple micronutrient supplementation combined with vitamin A was beneficial in improving growth among children with stunting, compared to vitamin A alone or to vitamin A plus zinc. Effects on anemia require further study.This study is registered with ClinicalTrials.gov, number .NCT00156832. An estimated 12% of South African children below five years of age are underweight, and 25% are stunted. DespiteZinc supplementation is known to improve growth in certain populations of children. A meta-analysis of zinc supplementation trials found significant beneficial effects on height in children, with the largest benefit occurring in children with low height Z-scores at baseline. NotablyAnemia in infants and children is multifactorial in origin. While iron deficiency is the most common cause, deficiencies in other nutrients that contribute to iron homeostasis - riboflavin, folic acid, vitamins B12, C and A - may also result in anemia. The higIn HIV-infected children early viremia and subsequent disease progression are superimposed on the other factors that contribute to growth faltering and anemia in uninfected children. Poor longitudinal growth becomes apparent early in infancy. Unlike weight, height fails to respond to increased feeding. It is cWe analyzed data from a randomized, controlled, double-blinded community-based trial whose primary aims were to assess the benefit of prophylactic daily zinc or multiple micronutrient supplements, both combined with vitamin A, in comparison to vitamin A alone that was used as a control group, on: 1) the primary prevention of respiratory illness and diarrhea, and 2) the effect on growth in rural South African children. SupplemThe main purpose of this analysis is to report on the second study aim - effect of supplementation on growth - and to report exploratory analyses of the effect of supplementation on anemia. More specifically, the second aim of this trial was to compare the effect on longitudinal length-for-age Z-scores of adding either zinc alone or in combination with multiple micronutrients to vitamin A supplements, and to describe how HIV, stunting and diarrheal morbidity affect the longitudinal growth response.The study was a randomized, controlled double-blinded community-based trial of prophylactic zinc supplements, or multiple micronutrient supplements including zinc, given to three cohorts of children starting at age 6 months and continuing until age 24 months. The triDetails of the study methodology and procedures, and characteristics of this community, have been previously published ,21. BrieAn allocation list was prepared using computer-generated random numbers and a block size of six. The manufacturer prepared numbered packs of tablets corresponding to the allocation list. Assignment to the three treatment arms was done separately for three cohorts of children stratified by HIV status of child and mother: HIV-infected children of HIV-infected mothers; HIV-uninfected children born to HIV-infected mothers; and HIV-uninfected children of HIV-uninfected mothers. Investigators, study staff and participants were blind to the treatment assignments.None of the children received antiretroviral therapy because of the unavailability of therapy for children in both public and private clinics during the study period. When antiretroviral therapy for children became available in clinics in the area in January 2005, all children with HIV infection in the study were referred for evaluation and initiation of therapy as appropriate.There were three supplement arms, one of which contained only 1250 IU of vitamin A. This served as the control group instead of a placebo arm because the standard of care in South Africa at the time of the study was six-monthly vitamin A supplementation to children below the age of five years. The daily dose of vitamin A used in the trial was equivalent to the 6-monthly high-dose supplementation. The other two supplements contained 10 mg of zinc as zinc gluconate in addition to 1250 IU vitamin A. The multiple micronutrient additionally contained 0.5 mg each of vitamins B1, B2 and B6; 0.9 μg vitamin B12; 35 mg vitamin C; 5 μg vitamin D; 6 mg vitamin E; 10 μg vitamin K; 0.6 mg copper as cupric gluconate; 150 μg folate; 50 μg iodine; 10 mg iron as ferrous fumarate; and 6 mg niacin. All supplements were packaged in identical blister packs of seven tablets. All trial supplement tablets had the same color, form, texture and taste. Caregivers were instructed to crush the tablets into a small volume of porridge and ensure that the entire prepared portion was consumed. Trained field workers made weekly home visits to monitor compliance and to supply study supplements. At each visit, the field workers observed preparation and administration of the tablet. Any tablets remaining from the child's previous supply were counted and recorded. Use of any other micronutrient supplements during the study was documented. ChildreLength, head circumference, mid-upper arm circumference and triceps skin-fold thickness were recorded by a trained study nurse at enrolment and subsequently at scheduled visits to the study clinic. Caregivers and infants were asked to return for scheduled visits at 7, 8, 9, 12, 15, 18, 21 and 24 months of age to assess growth using anthropometry. Nurses F-test for the treatment by time interaction at a type I error level of 0.05 was used to decide whether there was an effect of treatment on longitudinal growth. Stunting was defined as a length-for-age Z-score below -2 [Length-for-age and weight-for-age Z-scores for each child at each visit were computed using the WHO Child Growth Standards. A changbelow -2 . BecauseAnemia was defined as hemoglobin below 11 g/dL. For thiA posteriori exploratory analyses were also conducted to explore the differential effect of interventions on two other relevant sets of subgroups defined by stunting and diarrhea incidence, based on prior literature describing subgroup differences and biologic plausibility. The presence of these subgroup effects was evaluated by testing for statistical interaction. For these exploratory analyses second-order statistical interactions between treatment group and subgroup over time were considered significant at p-value < 0.1.We estimated post-hoc power for our growth analyses bearing in mind that the numbers of children in the HIV-infected cohort were below the target determined by a-priori sample size calculations. In the cohorts of HIV-uninfected children we had adequate power using repeated measures on subjects to detect an effect size of 0.3 Z-scores, which is comparable to what had been observed in previous studies in the nutrition literature. In the The Ethics Review Committee of the University of KwaZulu-Natal and the Institutional Review Board of Tufts-New England Medical Center both approved the study. A Data Monitoring and Safety Board, established by the National Institutes of Health, monitored conduct of the trial. A parent or guardian for all children participating in the study provided written informed consent.Information on length Z-scores at two or more occasions was available for 317 (85%) of 373 children enrolled in the study . In the 62 children in all three cohorts who were stunted at baseline the multiple micronutrient arm showed, on average, a 0.7 Z-score improvement in length-for-age Z scores over 18 months versus declines of 0.3 in vitamin A plus zinc and 0.2 in vitamin A arms. There was a significant interaction (P = 0.029) between treatment and time in stunted children . Multiple micronutrients supplementation and addition of zinc to vitamin A ameliorated the effects of repeated diarrhea episodes on growth in the cohorts of HIV-uninfected children Figure . The proThe prevalence of anemia was 78.4% at baseline and 61.6% after 12 months respectively. In HIV-uninfected children, and in all cohorts combined, paired comparison at the two time points showed that the vitamin A and multiple micronutrient arms had significantly fewer subjects with anemia at 12 months versus baseline Table .Daily multiple micronutrient supplementation that included vitamin A and zinc, when compared to vitamin A alone or to vitamin A plus zinc, improved longitudinal growth over 18 months in children who were stunted at baseline. This is in keeping with the findings of a meta-analysis that found that multiple micronutrients, but not vitamin A or iron alone, improved longitudinal growth in children. PreviouExploratory analyses found that multiple micronutrients, when compared to vitamin A alone or with zinc, ameliorated the effect of recurrent diarrhea on stunting. Zinc has been shown to be effective in the case management of diarrhea - reducing the duration of the diarrhea episode and improving growth thereafter. To our The relatively small number of HIV-infected children in the study, and the non-ignorable pattern of missing data in this group, limits our ability to detect effects and make inferences for the cohort of HIV-infected children. All HIV-infected children with available CD4 counts who dropped out of the study either due to death or loss to follow-up had severe HIV disease based on immunologic criteria. The worse growth pattern seen in the multiple micronutrient arm is thus most likely related to the longer follow-up of stunted children in this arm rather than any impact of a particular micronutrient formulation. One may anticipate better cohort retention and very different outcomes in the presence of antiretroviral therapy. Our findings emphasize the need for evaluating how patterns of missing data and severity stratification may influence outcomes in trials involving HIV-infected children.While all three treatment arms had reductions in the proportion of children with anemia over 12 months, supplementation with multiple micronutrients tended to achieve the greatest reduction in anemia. This is supported by findings from a large multi-centre trial of multiple micronutrients that included South African; Vietnama priori hypotheses. Children with a hemoglobin level below 10 g/dL received iron treatment while continuing the study intervention. We were unable to examine whether the micronutrients prevented anemia because there was already a high prevalence of anemia at baseline. While similar numbers of children in each arm received iron treatment, fewer children in the multiple micronutrient arm remained anemic at the end of 12 months compared to the other arms. This may be due to the presence of iron in the multi-micronutrient formulation, resulting in an effectively higher total daily dose of iron than what would have been ingested through the therapeutic iron alone. The relatively smaller increment in hemoglobin in the vitamin A plus zinc arm compared to vitamin A alone suggests a possible inhibitory effect of zinc on iron absorption. The multiple micronutrient arm showed similar improvements in mean hemoglobin to the vitamin A arm despite containing zinc. It is possible that the additional micronutrients alleviated the negative effect of co-administration of iron and zinc. This is an important observation, albeit arising from exploratory analyses, that a combination of micronutrients may improve childhood morbidity while minimizing the potential negative biologic interactions between these nutrients.In this trial, anemia and hemoglobin change were not included as The effect of the micronutrient supplements on anemia and hemoglobin were not among the primary aims of the study, and thus the results of this analysis, as with any post-hoc analysis, have to be interpreted with caution. While similar numbers of children in each treatment arm received therapeutic iron the possibility of unmeasured confounding still remains. However, we think that this may improve generalizability to programmatic conditions where anemic children would receive iron therapy as standard of care. This is especially relevant to our trial, where children were enrolled for 18 months and exposure to other interventions - such as supplemental iron - is likely under non-trial conditions. The trial of zinc and multiple micronutrients in Peru by Penny et al.  also repIn summary, these findings suggest that daily multiple micronutrients added to vitamin A supplementation in infants and children may be useful in improving growth in children who are stunted or who have repeated diarrhea, and thus as a public health intervention in communities with a high prevalence of either stunting and recurrent diarrhea. The significant effect of multiple micronutrients compared to vitamin A alone that was the control group, or to vitamin A plus zinc, on longitudinal growth in this study is important for other African countries where zinc supplementation alone was not found to reverse stunting. The question of individual targeting for stunted children versus universal administration of multiple micronutrients to children in high-risk environments still needs to be resolved. Further evidence is needed to confirm the beneficial effects of micronutrients on anemia, and to determine whether similar benefits apply to HIV-infected children - especially those receiving antiretroviral therapy, which is now the standard of care. While exploratory analyses suggest an inhibitory effect of zinc on iron absorption, the co-administration of other micronutrients may negate this inhibitory effect.The authors declare that they have no competing interests.MKC designed and conducted the analyses and wrote the initial draft of the manuscript. JVdB and KAL served as project directors. MLB and AT were responsible for study design. NM supervised fieldwork. The manuscript was critically reviewed and approved by all of the authors.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2458/10/145/prepub


Sunitinib is an oral, multitargeted tyrosine kinase inhibitor that has been approved for the treatment of metastatic renal cell carcinoma. Although the majority of sunitinib-treated patients receive a clinical benefit, almost a third of the patients will not respond. Currently there is no available marker that can predict for response in these patients.We estimated the plasma levels of NT-pro-BNP  in 36 patients that were treated with sunitinib for metastatic clear-cell renal carcinoma.From the 36 patients, 9 had progressive disease and 27 obtained a clinical benefit (objective response or disease stabilization). Increases in plasma NT-pro-BNP were strongly correlated to clinical outcome. Patients with disease progression increased plasma BNP at statistically significant higher levels than patients that obtained a clinical benefit, and this was evident from the first 15 days of treatment . Median progression-free survival was 12.0 months in patients with less than 1.5 fold increases (n = 22) and 3.9 months in patients with more than 1.5 fold increases in plasma NT-pro-BNP (n = 13) .This is the first time that a potential "surrogate marker" has been reported with such a clear correlation to clinical benefit at an early time of treatment. Due to the relative small number of accessed patients, this observation needs to be further addressed on larger cohorts. More analyses, including multivariate analyses are needed before such an observation can be used in clinical practice. Sunitinib is an oral, multitargeted tyrosine kinase inhibitor of VEGFR-1, VEGFR-2, Fms-like tyrosine kinase receptor 3 (FLT3), c-KIT (stem-cell factor [SCF] receptor) and PDGFR .Cardiac toxicity is reported to be a common adverse event in sunitinib-treated patients . SchmidiWe recently reported on a cohort of patients treated with sunitinib for metastatic clear-cell renal carcinoma in our department . FollowiThe Chi-square test was used for testing significance of correlation for categorical variables. Univariate comparisons of Plasma NT-pro-BNP levels as well as changes from baseline where performed with the Wilcoxon non-parametric test. Logistic regression analysis was used in order to assess jointly the effects of baseline Plasma NT-pro-BNP levels and day 15 changes on disease progression. The area under the Receiver Operating Characteristic (ROC) curve is also reported. Time to event curves were calculated with the Kaplan-Meier method. Indicative cutpoint determination in PFS analysis was made using the method of Contal and O'Quigley (1999)  but therData on the patient cohort are presented on table Plasma NT-pro-BNP was increased during treatment and did not return to baseline values during the off-treatment period. This increase was not associated to changes in arterial pressure or decreased left ventricular ejection fraction, as it was assessed by echocardiogram or MUGA test. Baseline values as well as increases in plasma NT-pro-BNP were strongly correlated to clinical outcome. Patients with disease progression had lower baseline values and increased plasma BNP at statistically significant higher levels than patients that obtained a clinical benefit table , and thiMedian PFS was 6.9 months in patients (n = 15) with increases in plasma NT-pro-BNP after 15 days of treatment and 15.8 months in patients (n = 20) where plasma NT-pro-BNP remained at the baseline or lower values .We further explored the effect of change at 15 days on PFS in order to select with a statistical technique an alternative cutpoint for grouping the patients into separate groups. By testing several alternatives, the maximum value of the log-rank statistic was obtained at the 1.5 fold change. Median PFS figure  was 3.9 NT-pro-BNP levels in healthy population are depended on age and gender . In our in vitro we did not find any significant BNP production and sunitinib had no effect of on BNP at a protein level (data not shown).Plasma NT-pro-BNP levels may reflect volume overload or cardiac dysfunction in patients with heart disease. We observed a high variation in pre-treatment NT-pro-BNP levels that was not associated to disease burden or baseline cardiac function. However, Baseline levels predicted for response and sharp increases in plasma NT-pro-BNP were observed in patients that did not obtain any clinical benefit from treatment, and this was evident from the first 15 days of treatment. BNP is produced by myocardial cells, but it has been reported to be produced by many tissues including renal parenchyma as well . WhetherOn the other hand, hypoxia in large treatment-refractory tumours is a direct and sufficient stimulus for BNP induction via stabilization of HIF-1 alpha . SunitinWe report the value of plasma NT-pro-BNP measurement for predicting response to sunitinib treatment in patients with metastatic renal cell carcinoma. This is the first time that a potential "surrogate marker" has been reported with such a clear correlation to clinical benefit at an early time on treatment. Due to the relative small number of assessed patients, this observation needs to be further addressed on larger cohorts. More analyses, including multivariate analyses are needed before such an observation can be used in clinical practice.The authors declare that they have no competing interests.KTP designed the study, evaluated the patients, evaluated the results and prepared the manuscript. LFK was responsible for the treatment of the patients, collection of the data and evaluation of the results. CNP critically reviewed the manuscript and the interpretation of the data. GK performed the statistical analysis and helped with the interpretation of the data. CL performed the cardiac assessment of the patients and was involved in the preparation of the manuscript. EA and MC helped with the plasma databank and performed the immunoassay tests. CA participated in the clinical part of the study. DM supervised the project and participated in the clinical part of the study. AHK overviewed the analysis of the data and the manuscript. All authors read and approved the final manuscript.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2407/10/489/prepubTable S1. Baseline plasma NT-pro-BNP levels (pg/ml) and medianfold ratio after 15 days of sunitinib treatment according to age and gender. CB = Clinical benefit, PD = Progressive Disease.Click here for file


Hereditary angioedema (HAE) resulting from the deficiency of the C1 inhibitor (C1-INH) is a rare, life-threatening disorder. It is characterized by attacks of angioedema involving the skin and/or the mucosa of the upper airways, as well as the intestinal mucosa. In approximately 50 per cent of cases, clinical manifestations may appear during childhood. The complex management of HAE in pediatric patients is in many respects different from the management of adults. Establishing the diagnosis early, preferably before the onset of clinical symptoms, is essential in cases with a positive family history. Complement studies usually afford accurate diagnosis, whereas molecular genetics tests may prove helpful in uncertain cases. Appropriate therapy, supported by counselling, suitable modification of lifestyle, and avoidance of triggering factors  may spare the patient unnecessary surgery and may prevent mortality. Prompt control of edematous attacks, short-term prophylaxis and intermittent therapy are recommended as the primary means for the management of pediatric cases. Medicinal products currently used for the treatment of children with hereditary angioedema include antifibrinolytics, attenuated androgens, and C1-INH replacement therapy. Current guidelines favour antifibrinolytics for long-term prophylaxis because of their favorable safety profile but efficacy may be lacking. Attenuated androgens administered in the lowest effective dose are another option. C1-INH replacement therapy is also an effective and safe agent for children. Regular monitoring and follow-up of patients are necessary. The deficiency of the C1 inhibitor (C1-INH) is inherited as an autosomal dominant trait. It causes hereditary angioedema (HAE-C1-INH), which is regarded as an uncommon disorder characterized by recurrent angioedematous episodes involving the subcutis and/or the mucosa of the upper airways and the gastrointestinal tract . The diaIn 50 per cent of HAE-C1-INH patients, the manifestations of HAE-C1-INH first occur during childhood. Therefore, establishing the diagnosis early and initiating follow-up care as soon as possible are indispensable for preserving the patients quality of life. The occurrence of edematous manifestations in other members of the patient's family may assist diagnosis. This clue is present in 75 to 85 per cent of cases, whereas in the remaining 15 to 25 per cent, HAE-C1-INH results from a new gene mutation [in utero or during the neonatal period, along with those justifying the termination of pregnancy. Additionally, this diagnostic modality enables parents at genetic risk to avoid passing heritable diseases to their offspring or negative findings may encourage them to have unaffected children. Notwithstanding this, the routine use of prenatal diagnostics in HAE-C1-INH patients is impractical for several reasons. No mutation of the C1-INH gene can be detected in 8 to 10 per cent of cases [Prenatal diagnostics may recognize fetal abnormalities requiring intervention of cases . IdenticEvery patient registered with the Hungarian HAE Center undergoes testing of the C1-INH gene at the molecular genetics laboratory of the institution. The data thus obtained are collected in an international, locus-specific database . The CenType I HAE-C1-INH is characterized by reduced C4, as well as reduced antigenic and functional C1-INH levels. In Type II HAE-C1-INH C4 is reduced and antigenic C1-INH level is high or normal and the functional activity of C1-INH is reduced . ComplemAlthough HAE-C1-INH results from mutation of the C1-INH gene, molecular genetic analysis is not a prerequisite for diagnosis, as a complement study is sufficient to recognize the disease . In addiRANK [YGUM [The diagnosis of HAE-C1-INH was suggested by clinical manifestations in 21 per cent (10/49) of our pediatric patients and in five of these, a negative family history interfered with the recognition of the hereditary disorder  of other inflammatory or allergic etiologies. According to case reports published in the literature, it was misdiagnosed as edema caused by allergic asthma in a three-year-old girl  and mistIn the gastrointestinal tract, submucosal edema may be associated with colicky abdominal pain, nausea, vomiting, watery post-attack diarrhea, occasional paleness of the skin consequent to hypovolemia, prostration, dehydration, tachyarrhythmia, and fainting and it may mimic an 'acute abdominal catastrophe'. Afflicted patients are usually admitted to a surgical department for observation and are often subjected to an unnecessary operation . The mosD-dimer levels, these indices may serve as objective, but non-specific biomarkers [Recurrent abdominal complaints of unknown etiology should always raise the suspicion of HAE-C1-INH. The clinical manifestations of an edematous attack of HAE-C1-INH are accompanied by intra-abdominal abnormalities detectable by diagnostic imaging . Although US findings  are non-specific to HAE-C1-INH, abdominal US may prove a sensitive, rapid, and non-invasive differential diagnostic modality, which is particularly straightforward in pediatric patients ,30. Edemomarkers .An edematous abdominal attack occurred as the initial manifestation of HAE-C1-INH in 3 of the 49 pediatric patients. Estimating the prevalence of intestinal edema in the pediatric population is difficult, because 'belly ache' is a common symptom with a multitude of possible causes, especially in infants. One to two per cent of HAE-C1-INH attacks can occur in other localizations including the urinary bladder, the urethra, muscles and joints, kidney; pericardial or pleural effusion (known as the 'chest episode') or neurological symptoms . In our YGUM observed it in 58 per cent of cases [The characteristic prodromal sign of erythema marginatum (appearance of a map-like pattern on the skin) occurs more frequently during childhood. In a proportion of cases, this lesion can evolve as an independent phenomenon, without a subsequent attack. In our study population, this symptom occurred in 42 per cent of patients , whereasof cases . Its appof cases .ORK, we found that the earlier the onset of symptoms, the more severe will be the subsequent course of HAE-C1-INH [The time of onset, frequency, and duration of symptoms, as well as the severity of attacks all exhibit inter-individual variation and substantial differences exist even within the same family. Analyzing the time of onset of disease symptoms revealed an increase in the frequency and severity of manifestations between 3 and 6 years of age, as well as around puberty. This is probably related to the manifold physiological  changes occurring in these periods of development. In agreement with the observations of BE-C1-INH .http://www.haenet.hu.The kindergarten or school are informed of the diagnosis in writing. The patient receives a multilingual information card to carry at all times. Additionally, special medication for emergency use at home during acute edematous attacks is provided, along with contact information on the self-help organizations of patients.Counselling the patient and family is the initial step after diagnosis. Providing the patient and family with appropriate information is indispensable to adopting a suitable lifestyle and avoiding severe complications. At the initial visit, the child and parents are counselled in person and ongoing consultation is offered using a telephone hotline and the website of the HAE Center Helicobacter pylori is another potential provoking factor. Accordingly, it is expedient to screen patients for infection by this bacterium and when necessary, administer eradication therapy even during childhood [By triggering edematous attacks, certain factors can influence the time of onset and localization of symptoms. Provoking factors include trauma, emotional stress, surgery or diagnostic manipulation of the head and neck region, physiological changes of sexual hormones , changes of the weather, specific foodstuffs and medicinal products . The inchildhood .If despite counselling, the manifestations of HAE-C1-INH recur with increasing frequency and in more severe form, it is important to search for other accompanying disorders. In children, abdominal symptoms and/or neurological signs may suggest celiac disease. Certain medicinal products  can also induce the manifestations of HAE-C1-INH. Estrogen containing contraceptives are not recommended for adolescent girls. ACEIs and ARBs are widely administered to adult patients, but data on their pediatric use are limited. Only two case reports have been published on ACEI-induced angioedema in children ,37 and nAs disease manifestation onset usually occurs between 6 to 8 years of age, prophylaxis is extremely uncommon under the age of 6 years . During This prophylactic modality involving treatment on a single occasion or over several days is intended for the prevention of a single impending attack.OMPELS et al on the duration of prophylaxis and of C1-INH supplementation are at variance with our practice [This type of prophylaxis is recommended before surgical, diagnostic interventions contemplated in the head and neck region, including dental procedures, and other operations performed under endotracheal manipulation, as these may trigger an edematous attack -UAE primarily ,5. The fpractice .During childhood, surgical interventions are less frequent, shorter in duration and may not necessarily require general anesthesia. Operations performed under intratracheal narcosis were identified in the history of eleven patients - eight of these had experienced an edematous attack after the intervention before their HAE-C1-INH was recognized. After diagnosis, short-term prophylaxis was administered before 5 procedures  performed in 4 patients. Patients on pre-existing treatment with tranexamic acid or danazol received these agents in escalated doses before minor procedures. Untreated patients received 500 U C1-INH concentrate one hour before major operations and another 500 U was kept ready during all such interventions. Short-term prophylaxis (as above) invariably prevented edema formation; the postoperative period was uneventful and no complications ensued . Since iThis type of prophylaxis is used in the presence of prodromal symptoms as well as when potentially edema-inducing pathological, physiological or environmental effects persist for a brief  period only. Upper respiratory track infections occur frequently during childhood and it is important to treat bacterial infections early and to introduce short-term prophylaxis as soon as possible and to continue its administration over the duration of the infection . In our When prodromal symptoms  occurred, tranexamic acid (40 mg/kg/day) or danazol (100-200 mg/day) administered over 2 to 3 days reduced the severity and halved the duration of subcutaneous or gastrointestinal manifestations .The literature reveals that the recommendations on introducing long-term prophylaxis are extremely heterogeneous ,14,40-44Drugs suitable for long-term prophylaxis include antifibrinolytics , attenuated androgens  and C1-INH concentrate ,5. ExperAndrogen side effects include decreased growth rate, virilization, and behavioral disorders during childhood, whereas in adolescence, menstruation irregularities and elevation of serum transaminase levels may occur . Weight The number of longitudinal studies into the effectiveness and adverse effects of LTP is limited in both adult and pediatric patients. LTP is introduced if UAE has been detected in the history, as well as the patient has experienced severe and recurrent attacks with no identifiable triggering factor. After diagnosis, the frequency and severity of disease manifestations warranted LTP in 18.36% (9/49) of our pediatric patients - in contrast to the management of adults, where 39% are receiving LTP. TXA 20 to 40 mg/kg daily was administered in two or three divided doses (maximum dose: 3 g/day split 2 or 3 times per day). Before TXA was available, epsilon-aminocaproic acid  had been used, but it was less well tolerated by patients than TXA, which is now preferred for initial prophylaxis.When antifibrinolytics failed to achieve satisfactory improvement, caused severe adverse effects, or their use was contraindicated, treatment with attenuated androgens was introduced according to the Budapest protocol , with tith and 5th years of therapy despite the escalation of its dose and this is similar to our experience with the treatment of adult patients [Danazol was well tolerated by our pediatric patients . Treatmepatients . NotwitGOSTONI as early as in 1978 already [Long-term prophylaxis does not necessarily mean uninterrupted dosage with the drug over a lifetime - although this issue is not enlarged upon in consensus guidelines. On occasion of annually scheduled, regular check-ups, the therapeutic regimen is modified frequently: drugs are discontinued and others are introduced, as well as their doses are adjusted. Although it is not mentioned by pertinent guidelines, intermittent prophylaxis may prove effective and safe, especially in the management of pediatric patients. The technical term "intermittent prophylaxis" was used in relation to danazol treatment by A already . We usedThe management of full-blown attacks depends on their localization and severity. The range of drugs suitable for emergency use has increased over recent years. In addition to the medicinal products used earlier , icatibant (a bradykinin receptor B2 antagonist) and ecallantide  have become available in clinical practice and recombinant C1-INH is under clinical trial. No experience is available yet with the pediatric use of the latter three innovative drugs, which differ from previous treatments in dosage, mode of action, and manufacturing process. By increasing the range of therapeutic alternatives, the introduction of these agents enables medical professionals to make much more specific and individualized decisions in choosing an appropriate treatment . Owing tAlthough usually resolving spontaneously, subcutaneous attacks can be controlled by treatment with antifibrinolytics and anabolic steroids. Administering increased doses of these drugs reduces the duration of attacks and prevents their escalation. Edema localized to the extremities does not progress to a severe condition but in pediatric patients it is a common cause of absenteeism from school (and adults from work). Edematous swelling of the face, lips, neck or torso, as well as substantial edema of the extremities require special management. Facial edema may progress and involve the mucosa of the upper airways. Edema of the neck can cause complications through compression, whereas subcutaneous swelling on the chest may be accompanied by pericardial or pleural effusion ,32,57. SEdema of the upper airway mucosa is a life-threatening condition potentially leading to asphyxia, which is responsible for the 30-to 40-per-cent mortality related to lack of treatment or delayed diagnosis . CurrentEdema of the gastrointestinal mucosa requires emergency treatment to relieve acute clinical symptoms  and to correct hypovolemia from post-attack watery diarrhea. Additionally, clinical manifestations may mimic the signs of an 'acute abdominal catastrophe', which warrants considering the need for surgical intervention. In patients with known HAE-C1-INH, the dramatic effect of C1-INH concentrate administered for abdominal symptoms may be of differential diagnostic value. In our experience, treatment with this agent is followed by substantial mitigation of symptoms within half an hour and their complete resolution within 12 to 24 hours. The abnormalities detected by abdominal US  also resolve within 24 to 48 hours ,30. The th C1-INH Deficiency Workshop  a roundtable conference discussed the topic of self-injection by patients, in order to lay the foundations of future international guidelines.This involves prophylactic or emergency treatment in the patient's home, administered by the summoned health care professional or as self-medication by the patient or family member. This modality has the advantage of the fastest possible treatment without the delay incurred by transportation of the patient to a health care institution. Naturally, a prerequisite to this approach is to ensure the constant availability of emergency medication in the patient's home -62. No cARTINEZ [As regards pediatric patients, medication is best administered by a health care professional. Expert assessment of the patient's condition, observation of symptomatic improvement, and intravenous drug administration are best handled by experienced professionals. In case of UAE, emergency treatment at home should be followed by hospitalization until symptoms resolve completely. Admission to hospital is similarly necessary in a severe abdominal attack, in order to rule out a possible, acute abdominal emergency. If a life-threatening condition has occurred and no expert help is available within reasonable time, the emergency medication may be administered by parents, relatives, or older pediatric patients themselves - on condition that they have acquired the technique of intravenous injection beforehand . The sucARTINEZ . In HungAs HAE is an hereditary disorder and its gene therapy is not yet available, patients must reconcile themselves to a lifelong disease experience and prolonged doctor-patient relationships. The delivery of follow-up care for HAE-C1-INH patients and the accumulation of data on their disease and management are best implemented using a centralized approach . HAE cenThe diagnosis, management, and follow up of pediatric patients with HAE-C1-INH are different from those of adults. Familiarity with specific, childhood properties is indispensable to making an accurate diagnosis, choosing the most appropriate therapy, and shaping the pediatric patients' lifestyle to enable them to live a fuller life similar to their healthy peers.The author declares that they have no competing interests.


When roll-tilted, the subjective visual vertical (SVV) deviates up to 40° from earth-vertical and trial-to-trial variability increases with head roll. Imperfections in the central processing of visual information were postulated to explain these roll-angle dependent errors. For experimental conditions devoid of visual input, e.g. adjustments of body posture or of an object along vertical in darkness, significantly smaller errors were noted. Whereas the accuracy of verticality adjustments seems to depend strongly on the paradigm, we hypothesize that the precision, i.e. the inverse of trial-to-trial variability, is less influenced by the experimental setup and mainly reflects properties of the otoliths. Here we measured the subjective haptic vertical (SHV) and compared findings with previously reported SVV data. Twelve healthy right-handed human subjects  adjusted a rod with the right hand along perceived earth-vertical during static head roll-tilts .SHV adjustments showed a tendency for clockwise rod rotations to deviate counter-clockwise and for counter-clockwise rod rotations to deviate clockwise, indicating hysteresis. Clockwise rod rotations resulted in counter-clockwise shifts of perceived earth-vertical up to -11.7° and an average counter-clockwise SHV shift over all roll angles of -3.3° . Counter-clockwise rod rotations yielded peak SHV deviations in clockwise direction of 8.9° and an average clockwise SHV shift over all roll angles of 1.8° (± 11.1°). Trial-to-trial variability was minimal in upright position, increased with increasing roll (peaking around 120-140°) and decreased to intermediate values in upside-down orientation. Compared to SVV, SHV variability near upright and upside-down was non-significantly (p > 0.05) larger; both showed an m-shaped pattern of variability as a function of roll position.The reduction of adjustment errors by eliminating visual input supports the notion that deviations between perceived and actual earth-vertical in roll-tilted positions arise from central processing of visual information. The shared roll-tilt dependent modulation of trial-to-trial variability for both SVV and SHV, on the other hand, indicates that the perception of earth-verticality is dominated by the same sensory signal, i.e. the otolith signal, independent of whether the line/rod setting is under visual or tactile control. Although the position gain of OCR (= eye roll divided by head roll) is low in static conditions, being in the range of 5-25% of head-roll angle -21, orieHaptic perception results from the stimulation of mechanoreceptors in the skin, muscles, tendons and joints in the process of the manual exploration of an object in space ,32. To aStudies measuring deviations in the SHV for whole-body roll angles up to 90° showed roll overcompensation up to ~12° ,24 and hWade and Curthoys compared deviations from perceived earth-horizontal during eccentric constant-velocity rotation with chair velocities corresponding to roll-tilt angles of up to 40° during different psychophysical tasks  and measured OCR using video-oculography . They foUnlike adjustments of the SVV, which resulted in errors of ~20° when positioned along the earth-horizontal axis, self-adjustments along the perceived horizontal axis in darkness are, on average, accurate ,29. SimiThese observations from various paradigms measuring the perceived earth-vertical  in complete darkness suggest that the underlying internal estimate of gravity is obtained by multisensory integration that is critically affected by the presence or absence of visual orientation cues. Whereas clear differences in the accuracy of the SVV and the SHV  were demonstrated, similarities and differences concerning the precision of SVV and SHV adjustments under static conditions have not been addressed so far. Trial-to-trial variability of SVV increases with increasing whole-body roll ,9,37, peWith SHV being most similar in how the task is performed compared to SVV, we aimed to focus on this non-visual, haptic paradigm when studying effects of visual orientation cues on internal estimates of earth-vertical. For internal estimates of verticality in non-visual paradigms, certain qualitative predictions about the accuracy and precision of adjustments can be made. Regarding the accuracy of the internal estimate of vertical, smaller adjustment errors are predicted for the SHV, since visual orientation cues are missing. This is based on both previous experimental findings in SHV for roll angles up to 90° ,23,24 anIn this study we assessed the accuracy and precision of the SHV in various whole-body roll positions. These findings were then compared with the previously reported pattern of SVV. In brief, we found that the direction of rod rotation significantly influenced SHV adjustment errors, indicating hysteresis. Whereas errors in the SHV paradigm were clearly smaller and showed a distinct pattern compared to the SVV paradigm, trial-to-trial variability was noted to modulate similarly in both SHV and SVV. These findings underline the important contribution of the central processing of visual input to errors in estimated earth-vertical and indicate that the precise perception of earth-verticality is dominated by the same sensory signal, i.e. the otolith signal, independent of whether the setting is under visual or haptic control. The hysteresis noted in the haptic modality underlines the importance of controlling for the direction of object rotation in future studies implementing the haptic vertical.Twelve healthy human subjects : 24.8 ± 2.9 years, age range: 21-30 years) were studied. Before enrollment handedness was assessed in all subjects according to their verbal report and only subjects that reported being right-handed were included. Subsequently, the 13-item questionnaire adapted by Chapman and Chapman  confirmehead-roll orientation, although roll movements on the turntable were whole-body, i.e., included both head and trunk that were aligned. The rod to assess perceived earth-vertical was mounted on a safety bar in front of the subjects in the midline in 40 cm distance. This device consisted of a plastic tube, 29 cm long and 2.5 cm thick. By adding a Velcro strip to one end of the tube the two ends could be haptically distinguished. To achieve this haptic task, subjects actively explored the area in front of them in darkness with the right arm being unrestrained to determine the starting roll orientation of the rod. Subjects were allowed to reach the rod in a manner they felt most comfortable. Due to the fixed location of the rod 40 cm in the front of the midline, this will result in a slight forward movement of the unrestrained right arm, flexion in the elbow and an approximately horizontal position of the forearm for accurate grasping of the rod. Turntable position and rod orientation signals were digitized at 200 Hz and stored on a computer hard disk for offline processing.Subjects were seated upright on a turntable with three servo-controlled motor driven axes . A thermoplastic mask  that tightly covered the head in its neutral position  was applied and attached to the base plate behind the subject's head. Subjects were positioned so that the roll axis of the turntable intersected the center of the inter-aural line. A 4-point safety belt was placed around the shoulders and the hips. Pillows minimized movements of the trunk, the shoulders and the left arm; however, the right arm was spared to allow subjects to perform the rod adjustments without restraints. As the otolith organs, which have the largest impact on SVV , are sit2, peak velocity: ± 41°/s). Subjects were prompted acoustically to start the trial five seconds after the turntable stopped. They were asked to grasp the rod with their right hand using a wrap grip  or counter-clockwise (CCW) direction). Changes in head-roll position were made in 8 s  on different days.Trials were sorted according to head-roll orientation and the direction of rod rotation. Outliers were defined as data points differing more than 2 standard deviations (StdDev) from the mean. 3.7% of trials were identified as outliers and discarded. Average deviations relative to earth-vertical and the StdDev were calculated for each subject. CW deviations relative to earth-vertical have a positive sign. We will use the term "trial-to-trial variability" whenever intra-individual StdDevs are reported. Analysis of variance  with Tukey's correction for multiple comparisons was used for statistical analysis of the dependent variables . Independent variables were the direction of rod rotation (CW vs. CCW), the head roll orientation and (when appropriate) the experimental paradigm (SHV vs. SVV). Both main effects of and interactions between independent variables were analyzed.Average individual SHV adjustments are shown for all subjects in Figure As illustrated in Figure For comparison, SVV values as reported by Tarnutzer et al.  using a Individual trial-to-trial variability is shown in Figure When healthy human subjects were asked to indicate perceived earth-vertical in a haptic paradigm, the absence of visual orientation cues decreased the size of adjustment errors. The haptic task studied here, however, yielded a significant main effect for the direction of rotation of the rod, reflecting hysteresis, which was not the case for the SVV task used for comparison . OverallWhereas a distinct pattern of roll overcompensation  and roll undercompensation (at medium-sized roll angles) is known for the SVV ,5,6, shiAlthough the direction of arrow rotation in SVV paradigms is often not controlled for, hysteresis may emerge in certain conditions. Mezey et al. showed shifts in torsional eye position when watching at a slowly (4.8°/s) rotating visual line to obtain adjustments of SVV or subjective visual horizontal  into theTo explain the systematic errors in the SVV, Mittelstaedt postulated an imbalance in the tilt signal due to an unequal number of hair cells in the two macular organs , based uSeveral mechanisms may explain the reduced adjustment errors when eliminating vision. Either prior knowledge - as it may be used in the SVV - is not implemented in non-visual verticality estimates. This would avoid roll over- and undercompensation at the sake of centrally optimizing the precision of verticality estimates through prior knowledge. Discarding prior knowledge in non-visual verticality estimates while implementing it in vision-based paradigms, however, seems rather unlikely, since in the SHV the trial-to-trial of adjustments was increased only slightly and non-significantly at larger head-roll angles compared to the SVV. This observation speaks for a central optimization of trial-to-trial variability for the SHV as well. Alternatively a distinct bias signal integrated in non-visual verticality tasks could have masked roll over- and undercompensation known from the SVV. The brain might implement other mechanisms to reduce the noise in the otolith signal as for example sense gravity acting on the outstretching hand. This information could potentially be used to update a unified sense of the direction of gravity or changes in the perceived body limb position. Increasing gravitational torques by adding loads to the forearm, however, did not improve the accuracy of forearm alignments with the vertical . These oWhereas the accuracy of perceived earth-vertical differed considerably depending on the pointing device used, the precision of verticality adjustments in the SHV followed a similar pattern as in the SVV. As a consequence, the m-shaped pattern of roll-angle dependent variability modulations described for the SVV  was reprAs discussed above, systematic errors in the SHV adjustments were clearly smaller and distinct from the pattern know from the SVV. This raises the question to which extent central processing of the involved sensory signals becomes different when no visual input is available. In the simulations by Tarnutzer et al. it was shown that prior knowledge increases the precision of SVV adjustments at larger roll angles, whereas it was affected little near upright . AccordiPreviously, Bauermeister et al. noted slight roll overcompensation (up to 6°) of unimanual right-handed SHV adjustments for head-roll angles up to ± 90° . BortolaWhen using both hands to indicate perceived earth-horizontal, accurate adjustments were reported for roll-tilt angles up to 40° by Wade and Curthoys , whereasEliminating visual orientation cues improves the accuracy of internal estimates of the direction of gravity, whereas its precision is largely unaffected. These findings underline the important contribution of the central processing of visual input to errors in estimated earth-vertical and indicate that the precise perception of earth-verticality is dominated by the same sensory signal, i.e. the otolith signal, independent of whether the setting is under visual or haptic control. The significant direction-dependent differences in adjustment errors (hysteresis) noted in the haptic modality mandate the control for the direction of object rotation in future studies implementing the haptic vertical.The authors declare that they have no competing interests.JRS performed the experiments, participated in the design of the study and in drafting the manuscript. CJB participated in the study design and its coordination. DS helped formulating the study hypotheses and drafting the manuscript. AAT conceived of the study, performed the statistical analysis and participated in drafting the manuscript. All authors read and approved the final manuscript.