The potential for emergence and spread of HIV drug resistance from rollout of antiretroviral (ARV) pre-exposure prophylaxis (PrEP) is an important public health concern. We investigated determinants of HIV drug resistance prevalence after PrEP implementation through mathematical modeling.A model incorporating heterogeneity in age, gender, sexual activity, HIV infection status, stage of disease, PrEP coverage/discontinuation, and HIV drug susceptibility, was designed to simulate the impact of PrEP on HIV prevention and drug resistance in a sub-Saharan epidemic.Analyses suggest that the prevalence of HIV drug resistance is influenced most by the extent and duration of inadvertent PrEP use in individuals already infected with HIV. Other key factors affecting drug resistance prevalence include the persistence time of transmitted resistance and the duration of inadvertent PrEP use in individuals who become infected on PrEP. From uncertainty analysis, the median overall prevalence of drug resistance at 10 years was predicted to be 9.2% (interquartile range 6.9%–12.2%). An optimistic scenario of 75% PrEP efficacy, 60% coverage of the susceptible population, and 5% inadvertent PrEP use predicts a rise in HIV drug resistance prevalence to only 2.5% after 10 years. By contrast, in a pessimistic scenario of 25% PrEP efficacy, 15% population coverage, and 25% inadvertent PrEP use, resistance prevalence increased to over 40%.Inadvertent PrEP use in previously-infected individuals is the major determinant of HIV drug resistance prevalence arising from PrEP. Both the rate and duration of inadvertent PrEP use are key factors. PrEP rollout programs should include routine monitoring of HIV infection status to limit the spread of drug resistance. Antiretroviral (ARV) pre-exposure prophylaxis (PrEP) is a promising HIV prevention strategy We have developed and analyzed a population model of heterosexual HIV transmission and disease progression to assess the impact of PrEP implementation For the present work, we extended our published model β, between an individual (on PrEP) of gender g, activity level k, and age i, with an (infected) individual of opposite gender g′, activity level l and age j is given by:Ψ is the number of sex acts within the partnership; γ is the probability of HIV transmission per sex-act (infectivity) based on the disease stage, Ω, and drug resistance status, Θ, of the infected partner; and ξθ is the effectiveness of PrEP. Effectiveness is defined as the probability of preventing HIV transmission per sex-act through PrEP and is given by the product of the average efficacy of PrEP, ξ  and the average level of adherence to PrEP, θ . In a partnership, where the infected partner harbors major drug-resistant variants (discussed below), the probability of transmission of resistant virus is υβ, while that of wild-type virus is (1−υ)β, and the effectiveness of PrEP against resistant virus is ιξθ. The parameters ξ, θ, υ and ι assume values between 0 and 1 , HIV drug susceptibility (drug-sensitive or drug-resistant), type of drug resistance (transmitted or acquired), and virus population dynamics of drug-resistant HIV  into 22 different HIV drug susceptibility strata , 2. Our sistance  may occuThe model's dynamical behavior was investigated using numerical methods. The key model outputs were: i) HIV incidence; ii) HIV prevalence; iii) cumulative new HIV infections; iv) proportion of cumulative new infections with transmitted resistance; v) overall prevalence of HIV drug resistance (transmitted plus acquired); vi) prevalence of transmitted resistance; and vii) prevalence of acquired resistance. PrEP was introduced  at endemic equilibrium when HIV prevalence in sexually active adults (15–49 year-olds) was approximately 20%. We made comparisons between the epidemics with and without PrEP at each simulation time-step over a 10 year interval after PrEP introduction.We performed sensitivity analyses The impact of PrEP was next determined by simulating three different scenarios: optimistic, realistic and pessimistic . For eacOur mathematical model stratifies the study population by gender, age, sexual activity level, PrEP use, HIV infection status, disease stage and HIV drug susceptibility , and itsBy contrast, the overall prevalence of drug resistance was influenced most by the duration of inadvertent PrEP use (SRRC = 0.62) and the rate of PrEP uptake (SRRC = 0.34) in previously-infected individuals. Together these two parameters explained 50.5% of the variance in overall prevalence of resistance after 10 years. Not surprisingly, the prevalence of transmitted resistance after 10 years was most influenced by the persistence time of transmitted resistance (SRRC = 0.53), explaining 28% of the variance. The rate of PrEP uptake and duration of inadvertent use in previously-infected individuals (SRRC = 0.32) explained another 10.5% and 10.2% of variance in transmitted resistance, respectively. The prevalence of acquired resistance was most sensitive to the duration of inadvertent PrEP use (SRRC = 0.74) and its rate of uptake (SRRC = 0.27) in previously-infected individuals; together these parameters explained 61.6% of the variance in the prevalence of acquired resistance after 10 years. Likewise, the rate (SRRC = 0.40) and duration (SRRC = 0.36) of inadvertent PrEP use in previously-infected individuals were most influential for the proportion of cumulative new infections with transmitted resistance, explaining 28.8% of the variance in this outcome (data not shown). Factors influencing the prevalence of drug resistance when risk compensation was assumed were similar to the above (data not shown).For each scenario, the largest decrease in infections was achieved with the non-targeted strategy and the smallest decrease with the targeted-by-activity strategy . SpecifiUnivariate sensitivity analyses of resistance prevalence confirmed that the most important factors affecting resistance prevalence were the rate and duration of use of inadvertent PrEP in previously-infected individuals. When no inadvertent PrEP use in previously infected individuals was assumed, there was a major decline in the prevalence of drug resistance , particuUsing the targeted-by-gender strategy , more infections were prevented in women compared to men. These findings were generally robust (data not shown) to single and multiple changes in the model's key structural assumptions including those related to balance in the supply and demand of sexual partnerships in the population over time Data from animal studies show that orally administered antiretrovirals can prevent infection of macaques by simian immunodeficiency virus The current model represents a significant refinement of our earlier version in terms of model structure, parameter assignment and scenario design Notwithstanding model improvements, sensitivity analyses of infections prevented confirm our earlier findings of the impact of PrEP on HIV prevention The results of our scenario analyses provide important insight into potential emergence of HIV drug resistance after PrEP implementation. The non-targeted optimistic and realistic scenarios predicted low to moderate prevalence of drug resistance (2.5% and 9.9% respectively) along with high to moderate decreases in cumulative infections . Uncertainty analysis also predicted moderate levels of overall drug resistance. With targeted optimistic and realistic scenarios, the prevalence of resistance was modestly reduced with considerable erosion (up to 70%) of infections prevented. The prevalence of drug resistance rose to over 40% in the pessimistic scenarios with minimal reduction in HIV infections. Sensitivity analyses showed that the key driver of this negative outcome was the high level of inadvertent PrEP use in the already infected population. When the pessimistic scenarios were re-simulated excluding PrEP use in previously-infected individuals, the prevalence of resistance decreased to 4.5%.There are some important limitations of our current model structure and the assumptions within it. The precise quantitative detail of our predictions will be affected by variations in the sexual activity patterns of different populations, for which data are very limited, especially on sexual mixing patterns. However, we employed a well-established template of sexual behavior The actual impact of PrEP on drug resistance will depend on the PrEP agent or agents used as well as the biological, behavioral and viral characteristics of the HIV-infected population. Although we do not model a specific PrEP agent, we used resistance-related input estimates that would be expected for a single antiretroviral drug used for PrEP such as tenofovir disoproxil fumarate We excluded from our analyses the impact of antiretroviral therapy for infected persons and various other influences on transmission . These and other refinements will be addressed in future work. Nevertheless, the important conclusion for our modeling is that the spread of HIV drug resistance could be mitigated by limiting inadvertent PrEP exposure in already infected individuals. To accomplish this, PrEP implementation programs would need to be tightly coupled with HIV testing of individuals who are candidates for PrEP and monitoring of PrEP recipients for HIV infection and drug resistance.Appendix S1Model Equations and Details.(DOC)Click here for additional data file.


This case study evaluated the response of objective and subjective markers of overreaching to a highly demanding conditioning training mesocycle in elite tennis players to determine 1) whether players would become functionally or non-functionally overreached, and 2) to explore how coherently overreaching markers would respond.Performance, laboratory and cardiac autonomous activity markers were evaluated in three experienced male tennis professionals competing at top 30, top 100 and top 1000 level before and after their strength and conditioning training was increased by 120, 160 and 180%, respectively, for 30 days. Every week, subjective ratings of stress and recovery were evaluated by means of a questionnaire.2max  and speed strength indices  were observed in all players. Changes of maximal heart rate , laboratory markers  and cardiac autonomous activity markers  were variable among the players.After 74, 76 and 55 h of training, increases in V̇OImproved performance provides evidence that overreaching was functional in all players. However, several overreaching markers were altered and these alterations were more pronounced in the two top 100 players. The response of overreaching indicators was not coherent. Professional tennis players compete in about 50 to 90 competitive matches per year in the Association of Tennis Professionals (ATP) tour. Accordingly, volume and intensity of conditioning need to be reduced during the season from January to mid-November, and a relatively short period of 5–6 weeks at the end of the year offers the best opportunity to substantially improve motor abilities and get into shape for the up-coming season. Players follow a demanding conditioning regime and undergo training stimuli which may differ in volume and intensity from the physiological demands imposed by the matches played throughout the season.Functional overreaching is defined as a short-term decrement in performance as a result of increased training stress. It is a usual part of the training process of elite athletes and its recovery to regular performance occurs within a few days . Non-funThis lack of widely accepted markers may be one among other reasons why the prevalence of overreaching in professional tennis is currently unknown. No study has yet attempted to monitor overreaching in high-level tennis players. Notwithstanding, sports medicine physicians have called for a rigorous assessment of overreaching indicators in tennis . HoweverIs there any evidence that the athletes monitored were non-functionally overreached?Is the response of potential overreaching markers coherent, or does this response differ in magnitude or direction between the indicators employed?Thus, the current study evaluated the impact of an intense conditioning training mesocycle on common overreaching indicators in three elite tennis players competing at different levels. The following research questions were formed:2, 7–10 years of ATP tour experience, hard-court specialists) had signed written informed consent to participate in this study which was performed in accordance with the ethical standards reported by Harris & Atkinson , indicative of cardiovascular autonomic balance, has been reported to be related to overreaching . After 1Blood samples were collected from the ante-cubital vein at rest between 8.00 and 9.00 a.m. Samples were stored at 2–8 °C and analyzed by automated clinical systems within 2.5 hours. Plasma urea was measured using an enzymatic calorimetric endpoint method . Serum total testosterone and cortisol were assessed by competitive immunoassay using direct chemiluminescence . Determinations of IGF-I were made with a solid phase, enzyme-labelled chemiluminescent immunometric assay .Athletes performed an incremental exercise test on a motor driven treadmill . Protocol, measurements and data analysis have been described in detail elsewhere . ShortlyThe ability of the neuromuscular system to overcome resistance with the highest contraction speed possible (explosive strength) and to produce the greatest impulse in the shortest time (stretch-shortening cycle strength) was assessed by countermovement jump (CMJ) and by the drop jump (DJ), respectively, following the protocol suggested by Logan et al. . Single-The German version of the EBF-52 SPORT questionnaire   was usedThe conditioning mesocycle was designed individually according to the specific aims and initial test results, and left room for weekly adjustments. The primary aim of the top 30 player was to improve endurance and speed endurance, while the top 100 player was slightly more focused on developing strength and speed strength. The top 1000 player, whose performance had been hampered by a chronic foot injury in the preceding months, aimed at regaining his previous fitness level. The conditioning training included endurance, speed and agility, as well as strength and sensorimotor training.○ Steady-state moderate aerobic training: 70–85% of AnT velocity○ Vigorous steady-state exercise and moderate interval training: 85–100% of AnT velocity2max velocity○ High-intensity intervals: Between AnT and V̇OThe intensity of endurance training was prescribed relative to the individual lactate anaerobic threshold (AnT) as determined during the initial cardiorespiratory fitness test on the treadmill.Short sprints, plyometric training, agility drills and medicine ball throws with complete recovery in between sets Cross training  and speed endurance sprints characterized by an incomplete recovery between sets (lactic)Sensorimotor training for legs, trunk and shoulders ; moderate resistance exercises for shoulder and trunk○ Strength endurance: 3–5 × 15–20 repetitions, resistance 40–55% of 1RM○ Hypertrophy: 2–4 × 10–12 repetitions, 70–80% of 1RM○ Neuromuscular adaptation: pyramid system, 3–6 × 3–8 reps 80–95% of 1RMWhole-body strength trainingFor regeneration, athletes underwent massage by a physiotherapist (1.5–4.0 h/wk), stretched (0.5–1.5 h/wk), performed low-intensity ergometer cycling (0.5–1.0 h/wk) and stayed in the Sauna 1–2 times/wk.The volume of conditioning was decreased during the last two weeks, and especially during the last week of the preparation period, while on-court tennis practice time was increased. Volume and intensity of training load were reduced on Thursdays, and Sunday was off. If possible, the three athletes practiced together as a group. All conditioning training sessions were guided, monitored and documented by an experienced conditioning coach.Overall, conditioning training volume in the preparation period was increased by 120, 140 and 180%  as compared to the training volume the players had completed on their own during the competitive season in the preceding months.Because this case study involved three subjects only, we solely focused on descriptive statistics. Data are presented either individually for each player, or as a range.Athletes completed 74, 76 and 55 h of conditioning, corresponding to 95, 96 and 93% of their scheduled training sessions. The amount and quality of endurance, strength, sensorimotor, speed and agility, as well as on-court tennis training can be depicted from After 30 days of training, body weight and percent body fat remained unchanged in the top 30  and the top 100 player , while the top 1000 player  lost 1.5 kg of body weight and reduced body fat by 1.7%. Aerobic capacity and speed strength were improved in all players. In the month following the preparation period, the top 30 player improved his ATP entry ranking by 15 positions, whereas the ranking of the top 100 (−4) and top 1000 player (−15) decreased.Resting heart rate was increased and heart rate variability was reduced after the training period in all players. Lactate and heart rate responses to maximum exercise stress as well as alterations of hormonal parameters were variable among the three players .In terms of psychophysical state, the EBF-52 scores of the general stress subcategory remained low in all players (0.4–1.7), while general recovery was constantly rated moderate . In the top 30 and the top 100 players, perceived sports-specific stress increased to levels considered to indicate over-reaching  but remained low in the top 1000 player (2.2). Sports-specific recovery showed small variation throughout the preparation period and was constantly rated between 1.8 and 4.1 .Non-functional overreaching is a transient decrement in performance capacity despite increased training stress which restores within weeks . For theThe higher performance capacity of the top 100 and especially the top 30 player allowed them to conduct higher training volumes (+25%) and higher absolute intensities than the top 1000 player. These higher workloads seem to have resulted in stronger deflections of several overreaching markers, even though performance was not compromised.2max by an extraordinary 18% within 30 days. Compared to his personal best V̇O2max recorded mid November in previous years (59 ml·kg−1·min−1), the player had entered the conditioning programme in relatively poor shape due to his foot injury (49 ml·kg−1·min−1). It is well known that after prolonged training interruption, aerobic fitness may be regained more rapidly. However, subject effort may also have been greater in the second test than in the first, even though criteria for attaining V̇O2max were fulfilled.Interestingly, the top 1000 player increased his V̇ORegarding the immediate response to maximum exercise, the two higher ranked players showed reduced maximal heart rates (HR max) despite enhanced performance. A reduced HR max has been found in highly strained or over-reached athletes , as confSports-specific perceived stress increased markedly in the first half of the preparation period in both top 100 players before displaying a plateau (top 100 player) and slightly decreasing (top 30 player), whereas it did not change substantially in the first weeks and decreased in the last weeks in the top 1000 player. In other studies, overreached athletes have shown a deterioration in mood state, psychological distress and sleep disorders before a drop in performance . However−1) in all players (−1)  .An elevated resting heart rate, observed in all three tennis professionals, has been reported in highly strained athletes  and suggLimitations in the current case study obviously comprise the small sample size and the lack of a longitudinal tracking of the markers. Also, tennis performance is conditioned by many diverse factors such as physiological, anthropometric and biomechanical characteristics, technical skills, motivation, and the occurrence of acute or chronic injuries, so that a comparison between the three different tennis players is difficult, as is the extrapolation of the presented results.Even so, the variability observed in the response of various overreaching markers seems indicative of the difficulties related to the diagnosis of overreaching states. Attempts to identify reliable, specific and sensitive parameters for nonfunctional overreaching and overtraining have not been successful so far . The chaTo conclude, enhanced performance shows that functional overreaching occurred in three professional tennis players after a prolonged intense conditioning training mesocycle. The tennis players tolerated up to 75 hours of individualized training within 30 days. Endocrine, vegetative and psychophysical markers as well as the physiological response to exercise did not show a coherent response, but the alterations were generally more pronounced in the Top 100 players. Using single markers for the identification of impending nonfunctional overreaching in sports practice may lead to false alarms. To identify athletes who are failing to cope with the stress of training, coaches and medical staff might need to monitor performance, physiological, biochemical, vegetative and psychological markers on a regular basis.


In 1891, Von Recklinghausen first established the association between the development of osteoporosis in the presence of overt hyperthyroidism. Subsequent reports have demonstrated that BMD loss is common in frank hyperthyroidism, and, to a lesser extent, in subclinical presentations. With the introduction of antithyroid medication in the 1940s to control biochemical hyperthyroidism, the accompanying bone disease became less clinically apparent as hyperthyroidism was more successfully treated medically. Consequently, the impact of the above normal thyroid hormones in the pathogenesis of osteoporosis may be presently underrecognized due to the widespread effective treatments. This review aims to present the current knowledge of the consequences of hyperthyroidism on bone metabolism. The vast number of recent papers touching on this topic highlights the recognized impact of this common medical condition on bone health. Our focus in this review was to search for answers to the following questions. What is the mechanisms of action of thyroid hormones on bone metabolism? What are the clinical consequences of hyperthyroidism on BMD and fracture risk? What differences are there between men and women with thyroid disease and how does menopause change the clinical outcomes? Lastly, we report how different treatments for hyperthyroidism benefit thyroid hormone-induced osteoporosis. T score) at any site. Characteristics of osteoporosis include increased risk of fragility fractures, a deterioration of bone architecture, and in addition to low bone mass. The distinction of secondary osteoporosis is applied to low BMD as a result of factors beyond postmenopausal and senile osteoporosis. The myriad of secondary causes includes glucocorticoid therapy, connective tissue disorders, malabsorption syndromes, diabetes mellitus, and hyperthyroidism [The World Health Organization (WHO) defines osteoporosis as bone mineral density (BMD) 2.5 or more standard deviations below that of a young adult  along with a small amount of T3 , which is significantly more biologically active than T4. Additionally, T3 is peripherally created by the enzymatic conversion of T4 into T3 by the deiodinase enzymes D1 and D2. A third deiodinase enzyme, D3, converts T3 and T4 into inactive hormone metabolites reverse T4 (rT2) and rT3 [α and TRβ, exist [α1, TRα2, and TRβ1 mediate T3 action though their wide distribution in tissues. There is extensive expression of thyroid hormone receptors within bone, indicating that thyroid hormones can influence this tissue type. Expression of TRα and TRβ has been identified in chondrocytes, osteoblasts, and osteoblastoma and osteoclastoma cell lines [α1 expression in bone is about 10-fold higher than TRβ1 [α1 as the primary mediator of T3 action on bone [α1 receptors in a mutant mouse model (TRα (0/0), display developmentally delayed ossification, growth retardation, and impaired bone mineralization despite being euthyroid, highlighting the requirement for thyroid hormone in normal bone metabolism. In contrast, TRβ mutant mice TRβ (−/−) exhibit normal skeletal development in young mice. There was a persistent elevation of T3 and T4 levels in these TRβ (−/−) mice related to disruption of normal negative hormonal feedback at the hypothalamus and pituitary through TRβ. The excessive stimulation of TRα1 in these adult mice from the elevated T3 and T4 levels resulted in accelerated bone remodeling leading to adult osteoporosis [T3 action is mediated by its nuclear T3 hormone receptors (TRs). Two receptor isoforms, TRβ, exist , 5 and tll lines , 6. TRα1han TRβ1 , and stu on bone –10. The oporosis –11. ClinClassically, the deleterious effects on BMD in hyperthyroidism have been attributed to high levels of the circulating thyroid hormones T4 and T3. However, more recent publications have also implicated low TSH levels alone in the development of reduced skeletal integrity. This is clinically relevant, as subclinical hyperthyroidism is defined by normal T3 and T4 levels with an isolated suppression of TSH and has been correlated with changes to human bone density over time –14. The n = 15) and age and sex-matched controls to reconstruct bone remodeling curves, it was determined that there was a 9.6% loss of mineralized bone with each cycle in the hyperthyroid group [The continuous process of bone remodeling through balanced resorption and deposition is essential to maintain the integrity and strength of the human skeleton . During id group . Conversid group .The accelerated bone remodeling cycle in hyperthyroid states results in increased bone resorption and a subsequent increased release of calcium into systemic circulation , 27. HigIn addition to the release of calcium from bone into serum, levels of bone turnover markers are also elevated. Serum concentrations of alkaline phosphate, osteocalcin, osteoprotegerin, and FGF-23 correlate directly with the severity of biochemical hyperthyroidism –37. InteIn contrast, the urinary bone turnover marker, collagen-derived pyridinium cross-links, rapidly normalizes with correction of thyroid hormone serum levels. In the presence of elevated free T3 (fT3), collagen-derived pyridinium cross-links are elevated in urine; however, they rapidly return to normal shortly after initiation of therapy , 39.Does the severity and duration of thyrotoxicosis correlate with the rate, extent and location of bone loss?Osteoporosis is a uniform feature of untreated and sustained thyrotoxicosis. Numerous reports have described a consistent decrease in BMD and increase in fracture risk in untreated overt hyperthyroidism . This efThe severity of hyperthyroidism appears to influence the degree of bone loss and increase the probability of osteoporosis when looking at the relative degree of TSH suppression, rather than looking at free T4 and free T3 levels. The aforementioned cross-sectional study by Svare et al.  reportedn = 686) [n = 6722), which estimated a 31% higher fracture risk  in patients with TSH <0.10 mU/L compared to TSH 1.0–1.49 mU/L [Thyrotoxicosis is an established risk factor for fracture later in life –51. A prn = 686) . A TSH ln = 686) . The fran = 686) . Support.49 mU/L .The choice of initial antithyroid therapy used to correct the hyperthyroidism may be clinically important, as highlighted by studies looking at subsequent fracture risk. In a historical follow-up study of 617 patients with toxic goiter, patients treated with radioactive iodine alone, rather than being cotreated with antithyroid medications, had an increased fracture risk at the spine and forearm in comparison to age- and gender-matched control . This inIs the bone loss or osteoporosis due to hyperthyroidism reversible by normalizing thyroid function?The initiation of antithyroid therapy and successful achievement of an euthyroid state can reverse the extent of osteoporosis induced by overt hyperthyroidism , 54–58. At least three reports suggest after several years of euthyroidism, thyrotoxicosis-induced osteoporosis can be restored to normal , 56, 68.n = 27, mean age = 43.7). Comparable with previous investigations, the increase in BMD at the lumbar spine, femoral neck, and distal radius was significantly higher after 12 months of combination treatment [Studies carried out with rodents have established that bisphosphonate therapy ameliorates the excessive bone loss caused by hyperthyroidism due to excessive T4 hormone supplementation . Researcreatment .Z-scores (age-matched controls) and overall BMD values in vitamin D deficient patients (<25 nmol/L) compared to hyperthyroid patients with sufficient vitamin D levels (>25 nmol/L) [Therapy of hyperthyroidism-induced osteoporosis includes adequate Vitamin D supplementation and bisphosphonate therapy, in addition to antithyroid treatment. The continuous bone resorption and high serum calcium levels seen in sustained hyperthyroidism result in decreased levels of active Vitamin D, suggesting a therapeutic role for oral Vitamin D supplementation. A 2010 study examined the relationship between Vitamin D levels and BMD in patients with newly diagnosed hyperthyroidism . The rep nmol/L) , supportAs a result of its potent inhibition of osteoclast activity, calcitonin administration has also been investigated as a potential adjunct to therapy of hyperthyroidism-induced osteoporosis when used in combination with antithyroid medications. No additional benefit was seen in BMD or bone resorption markers at any dose of intranasal calcitonin although the BMD was significantly increased in all patients treated with antithyroid therapy and intranasal calcitonin at varying doses for 9 months . The conWhat impact does menopause or gender have on clinical outcomes of bone turnover in subclinical hyperthyroidism?Subclinical hyperthyroidism is a biochemical definition, with suppressed or undetectable TSH levels and normal concentrations of T3 and T4. Subclinical hyperthyroidism incidence increases with age, especially in women, and is present in an estimated 1.5% of women over the age of 60 . Serum TEvaluation of the association between the presence of subclinical hyperthyroidism and the risk of development of osteoporosis has produced conflicting results from many small studies; therefore, BMD and fracture risk in this population are better examined by a subdivision dependent on estrogen levels found in pre- and postmenopausal women.n = 37) measuring BMD in pre- and postmenopausal women with endogenous subclinical hyperthyroidism. It was observed that the BMD was not significantly affected in premenopausal patients, in direct contrast to postmenopausal women who had significantly decreased BMD at the femoral neck and radius, sites of predominantly cortical bone.Subclinical hyperthyroidism has consistently been correlated with an increased risk of reduced BMD , 76–78. Multiple studies of premenopausal women with subclinical hyperthyroidism, either in comparison to normal controls or postmenopausal subjects, have consistently found normal or near-normal BMD levels. It is unclear if such studies have been able to remove the bias of age, as postmenopausal women may just have had longer durations of low TSH , 78–84. Nearly half of the cases of osteoporosis in men are found to have an underlying secondary cause. The effects on BMD in male patients with endogenous subclinical hyperthyroidism have not been widely studied or reported. A recent retrospective analysis of hip fracture risk in patients with subclinical thyroid dysfunction revealed a hazard ratio (HR) of 4.91 in men with endogenous subclinical hyperthyroidism in comparison to a HR of 2.42 in postmenopausal females of the same category . The rean = 2004) examined fracture risk patients with subclinical hyperthyroidism, defined as a TSH below the reference range (≤0.4 mU/L) and a normal total T4 and T3, to age- and gender-matched controls [Subclinical hyperthyroidism has also been implicated with an increased incidence of fracture at any site. A retrospective study  [There is an increased risk of bone fracture in individuals who are overreplaced with exogenous thyroid hormone. Fracture risk is related to the degree of TSH suppression (TSH < 0.1 mU/L versus TSH 0.1–0.5 mU/L) and patient factors, such as age . For wom17, 684) . No incr17, 684) . Fractur17, 684) . TherefoCan bone loss due to exogenous thyroid hormone overreplacement be reversed?Appropriate thyroxine supplementation in euthyroid individuals who maintain TSH levels within the reference range has not been implicated in BMD reduction . It has A prospective study by Kung and Yeung  evaluateEffects of exogenous subclinical hyperthyroidism on bone are minimal in premenopausal women presumably due to the positive effects of estrogen on skeletal health. A cross-sectional study by Schneider et al.  evaluateBisphosphonates are indicated for the treatment of osteoporosis induced by overt hyperthyroidism. In a randomized trial, Rosen et al.  evaluateSimilar to the conclusions discussed previously in overt hyperthyroidism, the use of intranasal calcitonin has not been found to benefit BMD in patients treated with exogenous subclinical hyperthyroidism. Kung and Yeung  found noRegulation of skeletal development and maintenance of skeletal integrity is regulated in part by a normal balance of thyroid hormones. Rodent studies have implicated both thyroid hormones and pituitary TSH in regulation of bone remodeling, although it remains unclear if T3, TSH, or the combination of both are responsible for the increased turnover and subsequent osteoporosis observed in the hyperthyroid state. Overt hyperthyroidism leads to osteoporosis and increased fracture risk in patients regardless of their age or gender. Prevention of fractures attributed to low BMD in hyperthyroid patients begins with antithyroid therapy, as reversibility of hyperthyroid-induced osteoporosis is evident a few years after achievement of an euthyroid state. Supplementation with Vitamin D and bisphosphonates may further increase BMD in these patients. The fine control of bone integrity by thyroid hormones is exhibited by the detrimental consequences on BMD and increased fracture risk by subclinical hyperthyroidism in postmenopausal women and men, and in postmenopausal women treated with suppressive doses of thyroid hormone. Postmenopausal women and men with endogenous subclinical hyperthyroidism should be considered for bone density measurement and subsequent antithyroid therapy. Postmenopausal women on thyroid supplementation in suppressive doses should have bone density testing, with appropriate titration of suppressive therapy, and supplementation with calcium and bisphosphonates.


Finally, hydrogen peroxide treatment of implants resulted in an almost stoichiometric TiO2, rich in hydroxyl groups on the surface. Machined commercially pure titanium implants served as controls. Scanning Auger Electron Spectroscopy, Scanning Electron Microscopy, and Atomic Force Microscopy revealed no significant differences in oxide thickness or surface roughness parameters, but differences in the surface chemical composition and apparent topography were observed. After surface preparation, the implants were inserted in cortical bone of rabbits and evaluated after 1, 3, and 6 weeks. Light microscopic evaluation of the tissue response showed that all implants were in contact with bone and had a large proportion of newly formed bone within the threads after 6 weeks. There were no morphological differences between the four groups. Our study shows that a high degree of bone contact and bone formation can be achieved with titanium implants of different surface composition and topography.In a series of experimental studies, the bone formation around systematically modified titanium implants is analyzed. In the present study, three different surface modifications were prepared and evaluated. Glow-discharge cleaning and oxidizing resulted in a highly stoichiometric TiO The objective and rationale for the approach are presented in an earlier report . In shorin vivo –3. WhereIn previous studies, we examined the response of bone around threaded titanium implants with different surface modifications . The formation of bone was evaluated morphometrically after 1, 3, 6, 7, 12, and 52 weeks. At the earlier time periods, the formation of bone was less around especially the very smooth electropolished implants. The results suggested that the type of surface modification performed mainly influenced the early phase of bone regeneration around the implants. In the present study, we have examined the biological response of cortical bone around titanium implants modified with respect to both surface structure and chemical composition by glow-discharge plasma techniques and by hydrogen peroxide treatment, 1, 3, and 6 weeks after implantation.At first, the implants were electropolished since morphology and microstructure of the surface oxides depend strongly on the microstructure of the underlying metal over which the oxide grows. Machined and mechanically polished surfaces consist of a plastically deformed, amorphous layer which may extend several microns into the bulk of the material. Electropolishing or other etching treatments remove this amorphous surface layer, resulting in surfaces that have a polycrystalline termination in which the grain structure of the material is visible. The thinnest thermal oxides on titanium have a very homogenous and essentially featureless morphology.in situ characterization of the resulting surface properties. Glow-discharge techniques are widely used for cleaning, sterilizing, and modification of biomaterial surfaces , 4–9. Thex vivo and with a physical vapor deposition (PVD) technique on Ti-alloy that compared with silicon, TiN surfaces do not adsorb as much albumin/fibrinogen as silicon surfaces do. However, the protein layer on TiN (fibril meshwork) entrapped mainly red cells and no leukocytes compared with silicon adsorbing platelets retaining red blood cells and a large amount of leukocytes. N-ion implantation is another surface treatment technique used with the purpose of increasing the protection against ion release (corrosion) and eventually increases biocompatibility [There are different methods to grow surface oxides on metals in a controlled way, electrochemically or by glow discharge plasmas. Other techniques used are, for example, physical vapor deposition (PVD) techniques and thermal oxidation. The anodic oxidation and discharge (DC) plasma oxidation techniques enhance the diffusion rate of oxygen and titanium atoms and make it possible to grow thick oxides on metals . With gltibility , 19. 2O2, is a strong oxidizer and is also used in cleaning solutions [In vitro studies of the interaction between metallic titanium and more concentrated H2O2 have shown that titanium peroxide (TiO22-) and titanium superoxide (TiO2-) were formed [The third surface modification method used is based on previous studies –23 on tholutions . In vitre formed –23.In the present study, we have used glow-discharge plasma procedures, developed in our laboratory  for prepReported results show that neither of these, relatively dramatic, surface modifications lead to significant differences in early bone healing around the titanium implants in this model. In the present study, we have examined the biological response in cortical bone 1, 3, and 6 weeks after implantation of titanium implants modified by changing the surface characteristics .∅3.75 mm, length 4.0 mm) were manufactured by machining from a commercially pure (99.7%) titanium rod. They were divided into four different groups. The preparation procedures are briefly summarized in Ninety-six threaded implants  were ultrasonically cleaned in successive baths of trichloroethylene, acetone, and ethanol of analysis grade purity for ~15 minutes in each.in situ in pure oxygen  for 5 minutes in order to form a pure surface oxide consisting of titanium dioxide (TiO2). The other group was subjected to glow-discharge plasma (2.0 kV) in pure nitrogen gas  for 5 minutes to form a thin titanium nitride film.Two groups (24 + 24 samples) of machined implants were modified using a glow-discharge procedure, carried out in different gases and resulting in the formation of two different surface compositions, titanium oxide and titanium nitride layers, respectively. The glow discharge method has been described in detail elsewhere, and it will only be briefly described here . The imp2O2  solution for 40 hours at 8°C, in order to form a sterile, peroxidized surface [γ-irradiated at 28.9 kGy for 25 h at 30°C. Bacteriological and sterile tests were proved negative for all four sample groups.The fourth group (24 samples) was, after the ultrasonic cleaning procedure, incubated in a 10 mM H surface –23. AfteμA, e-beam diameter of 180 μm, and energy resolution of 0.6%. Relative concentrations (in atomic percent) of the detected elements were calculated from their peak-to-peak values in differentiated spectra after correction with the elemental sensitivity factors [2O5, was 5.2 nm/min, which corresponds to approximately 2.6 nm/min for TiO2.The surface elemental composition of two samples of each preparation type was analyzed with Scanning Auger Electron Spectroscopy (AES), . Survey spectra (30–1730 eV) in two or five points located in the thread portion of each implant and depth profiles in one or two points at each sample were recorded. All spectra were taken at 5 keV primary electron energy with an e-beam current of 1.5  factors . This prα source and a monochromated Al-α source, respectively, of a PHI 5500-system .Since the Auger electron signals from Ti and N overlap, X-ray photoelectron spectroscopy (XPS) was also used for a more accurate determination of the chemical composition of the nitride sample. Both survey spectra (0–1100 eV binding energy) and high-resolution spectra of the Ti 2p, N 1s and O 1s peaks were recorded, using the standard Mg-Scanning Electron Microscopy  was used to obtain an overall picture of the surface topography of the samples. SEM micrographs were taken at several randomly chosen areas on the implant surfaces. 3N4 tips were used for imaging in the contact mode. One sample of each preparation type was analyzed at ten randomly chosen areas, (1 × 1 μm2 and 256 × 256 pixels) on the flat part at the bottom of the implant. This location on the implant was shown in a previous study to give essentially the same result as analysis on the threaded part [A quantitative characterization of the nanoscale surface topography and roughness was carried out by Atomic Force Microscopy . Standard SiRrms) of each imaged area was quantitatively evaluated using the computer software of the AFM instrument, and mean values were calculated for each type of surface. In addition, the AFM images were also used to calculate the surface area enlargement (Adiff). This parameter represents the enlargement in surface area (in percent of the projected area) caused by surface roughness in the range from a few nm (resolution of the images) up to 1 μm (size of the imaged area). The surface area enlargement was estimated from the sum of the area of all triangles formed by three adjacent pixels divided by the projected image area [Ra-value) was obtained by an optical profilometer for three-dimensional measurements, TopScan3D  [The surface roughness (age area . AdditioGermany) .Twenty-four adult New Zealand white female rabbits, weighing 3-4 kg, were used. The experiments were approved by the Local Ethics Committee. The animals were allowed to run free in a specially designed room with food and water ad libitum. The procedures for surgery and implant insertion are described in detail in previous reports , 3. In sen bloc, further immersed in glutaraldehyde overnight and then postfixed in osmium tetroxide for two hours. After dehydration, the undecalcified specimens were embedded in plastic resin, L R White .The animals were sacrificed with an overdose of barbiturates intravenously and fixed by perfusion with 2.5% glutaraldehyde in 0.05 M sodium cacodylate buffer, pH 7.2. The implants and surrounding tissue were removed μm thickness were prepared [Ground sections of 10–15 prepared  and examThe Fisher exact test for paired samples was used.2O2 incubated samples, respectively, were nearly stoichiometric titanium dioxide. The depth profiles also showed a similar oxide thickness (2-3 nm) for these three groups. The relative concentrations (in atomic %) of the detected elements present on the sample surfaces, as measured by AES, are presented in Glow-discharge plasma treatment in pure nitrogen resulted in 3 nm thick stoichiometric titanium nitride films, as judged from the depth profiles for the TiLMM + NKVV and TiLMV peaks, respectively –33. The γ-irradiation rather than wet chemical procedures and autoclaving.Further, when comparing the preparations made in our earlier studies –3, it ca2O2 incubated samples. The quantitative AFM results are given in μm) which are oriented in the cutting direction. The surface roughness parameters, Rrms, as measured by AFM and optical profilometry were 26 nm and 0.6 μm, respectively, and in agreement with previous work. The seemingly large difference of these results stems from the difference in measured topographical features between the two methods. AFM gives information on a nanometer scale , while the optical technique gives information on a micrometer scale .Figures 2, which presumably leads to the observed differences in topography. Quantitatively, the two plasma treated surfaces differ somewhat: the oxide sample has a lower roughness, while for the nitride it is similar to the control sample. At the submicron level, the plasma oxidized samples had a lower Rrms value than the other implant surfaces. The Ra-values were within the range of 0.4–0.7 μm. The nitrided sample, however, has a larger surface enlargement than the oxidized surface  were filled with bone. There were no qualitative differences between the different implant types. The implant surface towards the marrow cavity was covered by a layer of bone which was in continuity with the cortex. This newly formed bone had commonly a woven character . The parThe sequence of bone formation around the implants essentially followed the same pattern as that previously described for machined, electropolished, and anodized titanium implants in the cortical bone of rabbits , 37 as w2 decrease the diffusion of Ti ions from the bulk metal, which may be of potential interest when implanted in vivo [The biocompatibility and kinetics of the bone formation process, may be either enhanced or suppressed by the surface properties. Excessive release of metal ions from the material may be one such potentially suppressing effect. From a biological point of view, strategies may therefore be to optimize the surface properties in order to reduce the negative effects , 39. Bot in vivo , 41. 2. In the literature, there are several reports on the biological reactions at TiN surfaces, mainly prepared by PVD and CVD techniques, including the responses of bone [In the present study, the bone response to the TiN implants did not differ significantly from that to the other implanted materials, including the machined implant with a native TiO of bone –44, soft of bone , 45 bloo of bone , 46, 47, of bone , human m of bone , and ost of bone , 51. Sev2 has a net negative surface charge, thus attracting cations like calcium, [2 powder (anatase) by adsorption of lanthanum cations resulted in an increased adsorption of albumin and serum proteins in comparison with controls [Another strategy is to chemically modify the titanium and titanium oxide surfaces by incorporation of, for example, cations such as lanthanum . At physcalcium, . Chemicacontrols . Furthercontrols  and bonecontrols .These and other observations indicate that the surface charge influences the bone tissue response, possibly by influencing the types and amount of proteins that are adsorbed to the surface. The interactions between proteins, cells and implant surfaces may be influenced not only by the chemical properties of the surface but also the surface roughness which in turn may influence the wettability (hydrophilicity) which play an important role. . TherefoIn vitro studies on osteoblast-like cells [in vivo studies indicate enhanced bone response on titanium surfaces with higher surface energy [ke cells , 64 and ke cells  and titake cells . Increaske cells , as wellke cells , 65 are e energy , 67, 68 γ-irradiation and subsequent air exposure had a relatively low amount of surface contaminants. Previous studies have shown that machined and electropolished titanium implants with and without thick (180–220 nm) oxides have hydrophilic surfaces (water contact angles 15–33 degrees) with the highest contact angles observed for electropolished and machined surfaces [γ-irradiation indicates that the latter technique has a major advantage: identically prepared and cleaned, but autoclaved, machined titanium implants had 34 at % C contamination [γ-irradiated samples had only 13 at % C. The presently used methods of direct current glow discharge plasma treatment, followed by plasma oxidation or plasma nitriding, and subsequent sterilization with γ-irradiation may therefore be of interest for controlled preparation, cleaning, and sterilization of medical implants.The surfaces subjected to the present preparation techniques followed by sterilization using surfaces . In the surfaces . In the ut thick 80–220 nmsurfaces  who inve


Histological diagnosis was papillary thyroid carcinoma (PTC) in 57.1%, follicular thyroid carcinoma (FTC) in 36.4%, and Hurthle cell carcinoma in 7.8% of patients. Thus, hot thyroid nodules harbor a low but non-trivial rate of malignancy. Compared to individuals with benign hyperfunctioning thyroid nodules, those with malignant hyperfunctioning nodules are younger and more predominantly female. Also, FTC and Hurthle cell carcinoma are found more frequently in hot nodules than in general. We were unable to find any specific characteristics that could be used to distinguish between malignant and benign hot nodules.Hyperfunctioning nodules of the thyroid are thought to only rarely harbor thyroid cancer, and thus are infrequently biopsied. Here, we present the case of a patient with a hyperfunctioning thyroid nodule harboring thyroid carcinoma and, using MEDLINE literature searches, set out to determine the prevalence of and characteristics of malignant “hot” nodules as a group. Historical, biochemical and radiologic characteristics of the case subjects and their nodules were compared to those in cases of benign hyperfunctioning nodules. A literature review of surgical patients with solitary hyperfunctioning thyroid nodules managed by thyroid resection revealed an estimated 3.1% prevalence of malignancy. A separate literature search uncovered 76 cases of reported malignant hot thyroid nodules, besides the present case. Of these, 78% were female and mean age at time of diagnosis was 47 years. Mean nodule size was 4.13 ± 1.68 cm. Laboratory assessment revealed T The distinction is important, because hyperfunctioning nodules – also referred to as “autonomous,” “autonomously-functioning,” or “hot” nodules – are thought to only rarely harbor malignancy, such that fine needle aspiration (FNA) is not traditionally indicated in this circumstance [Thyroid nodules are frequently-encountered entities in clinical practice, occurring with a prevalence of 4% by palpation , 33% to umstance . Per theumstance .Here, we present the case of a woman with subclinical hyperthyroidism due to a hyperfunctioning thyroid nodule who was diagnosed with minimally-invasive follicular carcinoma after surgical resection. We also include our findings from a formal literature review on this topic, in which the historical, laboratory, and radiological features of similarly-documented cases were scrutinized to determine if there are features that differentiate hyperfunctioning thyroid carcinomas from solitary toxic adenomas. The results of a separate literature review aimed at estimating the prevalence of thyroid cancer within hot thyroid nodules are also presented. Our goal is to call attention to the fact that hyperfunctioning thyroid carcinomas are well-described in the literature , challenging the commonly-held notion that the hot thyroid nodule is very unlikely to be cancerous.A MEDLINE literature search of English-language studies published between 1950 and January 2012 with the terms, “thyroid cancer, hyperthyroidism, surgery,” “thyroid cancer, hyperfunctioning nodule, surgery,” and “thyroid cancer, hot nodule, surgery,” was performed to determine the reported prevalence of thyroid carcinoma in patients undergoing resection of solitary hyperfunctioning thyroid nodules. Another literature search, using the terms, “hyperfunctioning thyroid carcinoma,” “toxic adenoma, thyroid carcinoma,” and “hot nodule, thyroid carcinoma” was performed using MEDLINE and by reviewing the citations of relevant articles in order to collect data on reported cases of patients with a solitary thyroid nodule found to harbor thyroid carcinoma. Case series were included provided that at least some demographic and clinical details of individual subjects were described. A third MEDLINE literature search, using the terms “hot nodule,” “hyperfunctioning thyroid nodule,” and “autonomous thyroid nodules,” was performed to establish the demographic characteristics and nodule sizes of subjects with solitary hyperfunctioning thyroid nodules. In this third group, the available studies analyzed the group of hyperfunctioning nodules en masse and did not make a distinction between hyperfunctioning nodules that were benign and those that may have been malignant. Therefore, while the aggregate data from these studies most likely involved predominantly benign cases, a small number of malignant cases also may have been included. For this reason, we will subsequently refer to this group as “predominantly-benign” hyperfunctioning (or hot) nodules.Weighted averages of data were calculated by assigning a “weight” to each study , multiplying each weight by the subject mean in the corresponding study, and then taking the sum of these products.A 29-year-old teacher was referred to her local endocrinologist to evaluate a palpable left thyroid nodule. While she did not report thyrotoxic symptoms at that time, she was found to have a suppressed TSH. Thyroid scintigraphy revealed a hyperfunctioning left-sided thyroid nodule, and ultrasonography revealed it to be 2.4 cm in greatest dimension, isoechoic, and in the left lower lobe. A follow-up ultrasound one year later revealed an essentially unchanged nodule measuring 2.5 cm.The patient presented to our institution the next year, at which time she endorsed tremors, anxiety, insomnia, and oligomenorrhea. She denied local compressive symptoms, history of radiation exposure, or family history of thyroid malignancy. Her only medication was a daily multivitamin. Her weight was 106 kg with a height of 1.7 m, blood pressure of 117/73 mmHg, and heart rate of 79 beats per minute. A 1.5 cm, firm, slightly tender nodule was palpated in the left lower lobe of the thyroid; it moved with swallowing. There was no evidence of cervical lymphadenopathy or thyroid bruit. The patient had a slight, fine tremor of the hands with normal biceps deep-tendon reflexes bilaterally.123I thyroid scintigraphy revealed a left lower lobe hyperfunctioning nodule with 24-hour uptake of 27%  but normal free T4  and free T3 . Thyroid peroxidase antibody testing was negative , as were thyroglobulin antibody (< 20 IU/mL) and thyroid stimulating immunoglobulin (< 1.0) testing. Thyroid ultrasound revealed a 2.6 × 2.7 × 2.6 cm predominantly solid, isoechoic left lower lobe nodule with internal hypervascularity within a slightly enlarged but otherwise normal-appearing thyroid gland , she elected for the latter, citing the reduced risk of permanent hypothyroidism as her main deciding factor. Histologic evaluation of the surgical specimen revealed a solitary, circumscribed, and well-encapsulated tumor measuring 2.5 × 2.5 × 2.2 cm without lymphovascular invasion or extra-thyroidal extension Figure D. A focuIn order to place the current case report in context, we have attempted to establish the prevalence of malignancy within hot thyroid nodules. This involved a literature search for surgical case series of solitary hyperfunctioning nodules managed by thyroid resection. A total of 14 relevant case series were uncovered. The earliest we found was from 1967 and included 79 hot nodules that underwent surgical resection; none of those cases were found to harbor thyroid carcinoma. Among the 14 case series, carcinoma rates of intranodular carcinoma ranging from 0 – 12.5% were noted, with a weighted total of 3.1%  of malignant hot thyroid nodules, in which autonomy was highly suggested by both biochemical parameters of hyperthyroidism and scintigraphic evidence of increased radioiodine or labeled technetium uptake. All possibly-relevant features of the 77 cases were extracted and are displayed in Table We compared age at diagnosis and female: male ratio of individuals with malignant, hyperfunctioning thyroid nodules to those with benign, toxic adenomas. Data on the latter group was compiled from a separate literature search which identified several surgical case series of predominantly-benign hot nodules, as described in the Methods section Table . SubjectThe sizes of the malignant hot nodules Table  and pred3 and T4 levels, 16 with elevations of only T3, 3 with isolated T4 excess, and 7 with subclinical hyperthyroidism [3 level was significantly higher  than mean free T4 level  [3 levels, 54 % had elevated T4 levels, and 31% were biochemically euthyroid [3 appears to be a common factor among both benign and malignant hyperfunctioning thyroid nodules. Of interest, a greater prevalence of frank biochemical hyperthyroidism can be seen in patients with malignant hyperfunctioning thyroid nodules who have larger nodules , whereas closer to half of the subjects had elevation of thyroxine (51.9%) Table . Note thyroidism . In a st5 pg/mL) . Blum evuthyroid . Thus, hs Figure B.We next turned our attention to high-risk historical features and suspicious sonographic features to determine if these were present in cases of malignant hot nodule Table . These fThe majority of malignant hot nodules were proven to be PTC (57.1%), and of these, 18.2% were the follicular variant of PTC (FVPTC). Follicular thyroid carcinoma (as seen in our patient) comprised 36.4% of cases, while Hurthle cell carcinoma was found in 7.8% of samples. By comparison, in the U.S. National Cancer Data Base (1985–1995), which includes histologic information on all thyroid nodules as a group, the prevalence of PTC was approximately 85%, FTC was 10%, and Hurthle cell carcinoma was nearly 3% . Thus, tOf note, within the 77 identified cases of malignant hot nodule, only 23 subjects received FNA of their thyroid nodule prior to resection. FNA enabled a pre-operative diagnosis of thyroid carcinoma in 43.5% of cases. However, 30.4% of subjects undergoing FNA were erroneously characterized as having benign lesions, and 17.4% of samples were nondiagnostic. Of the subjects with false negative biopsies, surgical pathology eventually revealed PTC in 3 cases (one of these was FVPTC), FTC in 1 case, and Hurthle cell carcinoma in 3 cases. Tumor size was given in only 3 instances of false negative biopsies. Two of these were of subcentimetric PTC, and it is presumed that small size was likely a chief factor leading to misdiagnosis. However, the other was a 3 cm nodule comprised wholly of Hurthle cell carcinoma.To our knowledge, the current data set is the largest and most detailed to date of patients with malignant hot thyroid nodule. In that regard, this study complements the informative and excellent 2012 review by Pazaitou-Panayiotou and colleagues examining the association of thyroid carcinoma with a broader spectrum of hyperthyroid states, including Graves’ disease and toxic multinodular goiter in addition to hyperfunctioning thyroid nodule . Of noteThere are several noteworthy discussion points and implications to the findings presented here. The first relates to the prevalence of malignancy within solitary hot nodules. As mentioned, in the available surgical case series that addressed this topic, a varied prevalence was noted, ranging from 0 – 12.5%, with a weighted average of 3.1%. Only a minority of the patients who underwent surgery in those series did so because of concerning findings from FNA, and as such, we do not believe that the data set is biased towards cases in which a post-operative diagnosis of malignancy was expected. We realize that this collection of case series includes only a small fraction of the total number of solitary hot nodules that have occurred, and also that the vast majority of malignant hot nodules likely have gone unreported. Furthermore, it is likely that malignancy goes undiagnosed in many cases of hot nodules treated with radioiodine, especially those harboring microcarcinomas. Despite these limitations, we do believe there are sufficient numbers of cases included among the collected 14 surgical case series of subjects to cite the 3.1% figure as being a fair and representative estimate of malignancy prevalence amongst solitary hyperfunctioning nodules. While this 3.1% prevalence figure is low – and in fact is lower than the estimated 5 - 15% prevalence of malignancy among all thyroid nodules  – it is Another important discussion point regards the limitations resulting from the retrospective nature of this analysis, including incomplete data and differing methodology used in many of the collected case reports and case series of malignant hot nodule. While some of the primary sources were meticulous in their case descriptions, others included less-thorough descriptions. For example, tumor size was frequently not reported, and thus it was unclear if a PTC tumor was simply an incidental microcarcinoma embedded within a larger hot nodule or a large, follicular variant of PTC comprising the entirety of the nodule. While we have included some cases in which the malignancy was a microcarcinoma and thus of uncertain clinical significance , these c123I thyroid scan, if not already done, especially if the serum TSH is in the low-normal range; if the nodule is found to be hyperfunctioning, it can then be followed [Also worth discussing are the differential prevalences of thyroid carcinoma histologic subtype found in hot nodules as compared to nodules as a group. As mentioned, there was a much higher prevalence of both FTC and Hurthle cell carcinoma in hot nodules  as compared to in all nodules . Additionally, a substantial percentage of the PTC cases found in hot nodules were the follicular variant. This may have bearing on the current algorithm for evaluation of patients with thyroid nodules as recommended in the 2009 thyroid nodule and thyroid carcinoma management guidelines [followed .As a final point of discussion, since none of the historical, biochemical or radiologic characteristics that were assessed seems to predict malignancy in the collected cases of hot nodules, one might ask if there is any utility in biopsying hot nodules. Such would be a shift from the current thyroid nodule management guidelines which views an increased nodular radiotracer uptake pattern as a reassuring characteristic from a cancer perspective . Given tThe authors declare that they have no competing interests.SM performed the literature review and drafted the manuscript. DM provided the radiology images and participated in drafting/revising the manuscript. LP provided the pathology images and participated in drafting/revising the manuscript. SW helped to provide the surgery perspective and edited the manuscript. JZ assisted with the literature review and drafting of the manuscript. All authors read and approved the final manuscript.


O. sativa accessions. The phenotyping for 14 traits was conducted in both temperate (Arkansas) and subtropical (Texas) climates and the genotyping used 154 SSRs and an indel marker. Heading, plant height and weight, and panicle length had negative correlations, while seed set and grain weight/panicle had positive correlations with harvest index across both locations. Subsequent genetic diversity and population structure analyses identified five groups in this collection, which corresponded to their geographic origins. Model comparisons revealed that different dimensions of principal components analysis (PCA) affected harvest index traits for mapping accuracy, and kinship did not help. In total, 36 markers in Arkansas and 28 markers in Texas were identified to be significantly associated with harvest index traits. Seven and two markers were consistently associated with two or more harvest index correlated traits in Arkansas and Texas, respectively. Additionally, four markers were constitutively identified at both locations, while 32 and 24 markers were identified specifically in Arkansas and Texas, respectively. Allelic analysis of four constitutive markers demonstrated that allele 253 bp of RM431 had significantly greater effect on decreasing plant height, and 390 bp of RM24011 had the greatest effect on decreasing panicle length across both locations. Many of these identified markers are located either nearby or flanking the regions where the QTLs for harvest index have been reported. Thus, the results from this association mapping study complement and enrich the information from linkage-based QTL studies and will be the basis for improving harvest index directly and indirectly in rice.Harvest index is a measure of success in partitioning assimilated photosynthate. An improvement of harvest index means an increase in the economic portion of the plant. Our objective was to identify genetic markers associated with harvest index traits using 203  Oryza sativa L.) is one of the most important staple foods In food production, optimizing grain yield, reducing production costs, and minimizing risks to the environment have been the primary objectives since the beginning of the twentieth century Crop harvest index is also highly influenced by environmental factors Classical QTL mapping reveals only a portion of the genetic control of a trait because there are only two alleles that can differ at any locus between the two parental lines. More comprehensive analyses of genetic architecture require consideration of a larger sample of the genetic variation in the species. One approach is association mapping, which maps the QTLs either among extant breeding lines with known pedigree relationships or in a diverse germplasm collection. Given pedigree and marker information, the probability for different lines in complex populations to share identity by descent QTLs can be defined, permitting estimation of the effects of each QTL O. sativaRecently, the USDA rice mini-core (URMC) subset was developed and serves as a genetically diversified panel for mining genes of interest to various users sativa URMC accessions with 155 molecular markers and phenotyped 14 traits contributable to harvest index in both temperate  and subtropical  locations. Our objectives were to identify the traits significantly correlated with harvest index per se and the markers significantly associated with component traits of harvest index. To control spurious associations, i.e., Type I error, we analyzed the genetic structure and familial relatedness in the collection. Different mapping models were tested for best fit of each trait. The chosen model was used to map markers associated with harvest index and associated traits phenotyped in two environments.We genotyped 203 O. indel with genome-wide distribution detected a total of 1993 alleles among 203 O. sativa accessions. The average number of alleles per locus was 12.86 ranging from 2 for RM338 to 57 for con673. Polymorphic Information Content (PIC) varied from 0.25 for AP5625-1 to 0.97 for con673 among the 155 markers with an average of 0.71. Nei's (1983) The set of 154 SSRs and an STRUCTURE software with multi-loci genotype data, a five-group model was identified to sufficiently explain genetic structure among 203 accessions. Ancestry of each of these accessions was inferred for assignment into a genetic group  belong to indica (IND), temperate japonica (TEJ), tropical japonica (TRJ), aus (AUS) and aromatic (ARO) (The resultant five groups of ic (ARO) , based oic (ARO) . TEJ accStatistical analysis using a mixed model demonstrated that the differences due to genotypes and genotype×location interactions were highly significant at the 0.001 level of probability for all of the 14 traits . The difSix traits were significantly correlated with harvest index and these correlation directions were the same across the two locations. The correlations with harvest index were negative for heading (−0.46 at Stuttgart and −0.61 at Beaumont), plant height (−0.50 and −0.50), plant weight (−0.36 and −0.30), panicle length (−0.45 and −0.32), while positive for seed set (0.52 and 0.61) and grain weight/panicle (0.32 and 0.40) . In the Dimension determination for PCA indicated that different dimensions should be included for testing associations for these traits. Further, relative performance of the association mapping models was also evaluated based on the criterion BIC . The sma−3 level of probability (the Bonferroni corrected significance level)  . Among 3eviously . AdditioAt Beaumont, we identified 28 markers significantly associated with harvest index traits . Among tAcross two locations, the associations of RM431 with plant height, Rid12 and RM471 with plant weight, and RM24011 with panicle length were consistently true. The four markers that associated with the same trait across both locations are called “constitutive QTL” markers, while others that associated with a certain trait only at one location are called “adaptive QTL” markers The allelic effects of the constitutive markers associated with their traits were estimated using the least square mean (LSMEAN) of phenotypic values and are presented in The average number of alleles per locus was 12.86 among 203 accessions in the URMC genotyped with 155 markers. The allele number per locus is the highest among the rice collections that have been reported to date O. sativa accessions in the URMC were divided into five model-based groups from ancestry analysis , the PCA showed the best fit with the smallest BIC value for harvest index traits. Interestingly, correction of the kinship model was not observed to be better than the naive model. Similarly, the models with Q+kinship or PCA+kinship did not perform better than the ones with only Q or PCA, either. Shao et al. qHID7-1” responsible for harvest index Ghd7” having major effects on grains per panicle, plant height and heading in rice qHID3-2” for control of harvest index sd1” sd1 that is involved in gibberellic acid biosynthesis decreases plant height, thus increases harvest index. The decreased height reduces lodging susceptiblity, is tolerant to heavy applications of nitrogen fertilizer, and can be planted at relatively high density, all contributing to improved grain yield that has resulted in the Green Revolution in cereal crops including rice Harvest index is an integrative trait including the net effect of all physiological processes during the crop cycle and its phenotypic expression is generally affected by genes responsible for non-target traits, such as heading qHID4-1 and qHID4-2 for harvest index qHID8-1qHID8-1hi2.1Other markers were associated with the traits correlated with harvest index, but not with harvest index directly in this study. These markers have been reported either nearby or flanking the QTLs for harvest index. RM5, which was associated with plant height in the Stuttgart study, was close to a reported QTL for harvest index on Chr 1 Quantitative traits show a range of sensitivities to environmental changes Environment-specific QTLs can be used for marker-assisted selection (MAS) at specific environments. For example, RM431 could be used to improve harvest index directly and indirectly through decreasing plant height and increasing seed set in Arkansas because it was co-associated with harvest index, plant height, and seed set. However, the constitutive marker-trait associations over multiple environments can be applied to MAS programs in a wide area. For example, results suggest that the constitutive markers Rid12 and RM471 could be used to improve harvest index indirectly through decreasing plant weight in the southern states of the USA.Comparison of allelic effects of these constitutive markers can classify the alleles within a marker locus into superior or inferior ones, which helps decide which to use for MAS in the southern states. For example, allele 253 bp of RM431 and allele 390 bp of RM24011 had the largest effect on decreasing two traits, plant height and panicle length, negatively associated with harvest index. Thus, these superior alleles can be introduced for improvement of harvest index indirectly through decreasing the negative traits at both locations. Conversely, the allele 411 bp of RM24011 had the largest effect on increasing the panicle length and thus would not be useful for improving harvest index using MAS at either location. Interestingly, the two alleles of Rid12 associated with plant weight had opposite effects at the two locations. Allelic choice for this marker should be dependent on the particular environment targeted for breeding.Results of the present study demonstrated that genome-wide association mapping in the URMC could complement and enrich the information derived from linkage-based QTL studies. After validation or fine mapping of these putative genomic regions, the information will help secure food production through either direct improvement of harvest index or indirect improvement via changes in seed set, grain weight per panicle, heading, plant height and weight, and panicle length using the MAS.sativa whereas the remaining belongs to other species in Oryza. Pure seed of these accessions were provided by the Genetic Stock Oryza Collection (GSOR) (www.ars.usda.gov/spa/dbnrrc/gsor) with cultivar name or designation, accession number, registration year, place of origin, longitude and latitude of origin, pedigree or genetic background (if available), morphological characteristics and references. The GSOR supplies seeds for research purposes to national and international users upon to request. In this study, only 203 O. sativa accessions were used for the following analysis because the wild relatives, O. glaberrima, nivara, rufipogon, glumaepatula and latifolia, contain many rare alleles. Rare alleles are one of the factors that increase the risk of Type I errors or spurious associations Of 217 accessions in the URMC, 203 belong to O. −1 of nitrogen as urea was applied. Weeds were controlled at both pre-planting and pre-flooding stages with locally recommended herbicides.Evaluations were conducted for 14 traits in two field locations, USDA-ARS Dale Bumpers National Rice Research Center near Stuttgart, Arkansas and USDA-ARS Rice Research Unit near Beaumont, Texas during the 2009 growing season. The Stuttgart test site is located at N 34°27′44″ and W 91°24′59″, representing a temperate climate with a 243 d frost free period and average temperature of 23.9 C during the growing season. The Beaumont test site is located at N 30°03′47″ and W 94°17′45″, representing a subtropical climate with a 253 d frost free period and an average temperature of 26.1 C during the growing season. The experiments at both locations utilized a randomized complete block design having three replications with nine plants spaced 0.3×0.6 m in each plot. Three seeds were sown in each of nine hills in a plot using a Hege 1000 grain drill planter on April 23 and May 6 of 2009 at Stuttgart and Beaumont, respectively. Each hill was thinned to a single plant right after the permanent flood was applied at five leaf stage. Before flooding, fertilizer at 55 kg haData collection followed procedures described by Yan et al. http://www.gramene.org/), and five SSRs  were amplified in house indel at the Rc locus, named Rid 12 and is responsible for rice pericarp color. Polymerase chain reaction (PCR) marker amplifications were performed as described in Agrama et al. http://www.gramene.org/). Markers labeled with different colored fluorescence and that amplified products with size differences of 20 bp or more were multiplexed together post PCR.Bulk tissue from five plants was collected from each accession as described by Brondani et al. PowerMarker version 3.25 PowerMarker was also used to calculate the average number of alleles, gene diversity, and polymorphism information content (PIC) values. The tree to visualize the phylogenetic distribution of accessions and ancestry groups was constructed using MEGA version 4 Genetic distance was calculated from the 155 molecular markers using Nei distance 2 = σg2/(σg2+σe2/n), where σg2 as the genotypic variance, σe2 as the environmental variance and n as the number of replications NTSYSpc software version 2.11 Each of the 14 phenotypic traits was modeled independently with the MIXED procedure in SASv.9.2, where genotype, location and interaction of location with genotype were defined as fixed effects while replication within a location (block effect) was a random effect. Broad-sense heritability was calculated using formula HSTRUCTURED(K), assisted with L(K), L′(K) and (L″K). The D(K) perceives the rate of change in log probability of the data between successive (K) values rather than just the log probability of the data. Determination of mixed ancestry (an accession unable to be clearly assigned to only one group) was based on 60% (Q) as a threshold to consider an individual with its inferred ancestry from one single group. Principal component analysis (PCA), that summarizes the major patterns of variation in a multi-locus data set, was performed with NTSYSpc software version 2.11 STRUCTURE for the following linkage disequilibrium analysis.The model-based program P values were compared to the Bonferroni threshold (1/155 = 6.45×10−3) to identify statistically significant loci. Allelic effects at marker loci were compared using the LSMEANS and pdiff option in the MIXED procedure, using Saxton's PDMIX800 SAS macro Following the procedures previously recommended Table S1A. Spearman correlation for each pair of 14 traits evaluated at Stuttgart, Arkansas in 2009. B. Spearman correlation for each pair of 14 traits evaluated at Beaumont, Texas in 2009.(DOC)Click here for additional data file.Table S2Fitness analysis of mapping model for harvest index traits using Bayesian information criterion (BIC) in both Arkansas and Texas.(DOC)Click here for additional data file.Table S3The marker loci associated with harvest index traits at Stuttgart, Arkansas and Beaumont, Texas in 2009.(DOC)Click here for additional data file.Table S4Comparison of allelic effect of four constitutive marker loci at two locations, Stuttgart, Arkansas and Beaumont, Texas. RM431 for Plant height, RM471 and Rid12 for Plant weight, and RM24011 for Panicle length.(DOC)Click here for additional data file.


Neuromyelitis optica (NMO) is an inflammatory disease of central nervous system characterized by optic neuritis and longitudinally extensive acute transverse myelitis. NMO patients have cognitive dysfunctions but other clinical symptoms of brain origin are rare. In the present study, we aimed to investigate cognitive functions and brain volume in NMO. The study population consisted of 28 patients with NMO and 28 healthy control subjects matched for age, sex and educational level. We applied a French translation of the Brief Repeatable Battery (BRB-N) to the NMO patients. Using SIENAx for global brain volume  and VBM for focal brain volume (GM and WM), NMO patients and controls were compared. Voxel-level correlations between diminished brain concentration and cognitive performance for each tests were performed. Focal and global brain volume of NMO patients with and without cognitive impairment were also compared. Fifteen NMO patients (54%) had cognitive impairment with memory, executive function, attention and speed of information processing deficits. Global and focal brain atrophy of WM but not Grey Matter (GM) was found in the NMO patients group. The focal WM atrophy included the optic chiasm, pons, cerebellum, the corpus callosum and parts of the frontal, temporal and parietal lobes, including superior longitudinal fascicle. Visual memory, verbal memory, speed of information processing, short-term memory and executive functions were correlated to focal WM volumes. The comparison of patients with, to patients without cognitive impairment showed a clear decrease of global and focal WM, including brainstem, corticospinal tracts, corpus callosum but also superior and inferior longitudinal fascicles. Cognitive impairment in NMO patients is correlated to the decreased of global and focal WM volume of the brain. Further studies are needed to better understand the precise origin of cognitive impairment in NMO patients, particularly in the WM. Neuromyelitis optica (NMO) is a central nervous system (CNS) inflammatory disease characterized by optic neuritis (ON) and longitudinally extensive acute transverse myelitis (ATM). First described by Eugene Devic and others in the 19th century as a distinct disease, NMO or Devic’s disease was then classified as a subtype of multiple sclerosis (MS) Brain involvement in NMO patients seems to be more frequent than previously described. Brain MRI at the beginning of the disease is usually normal. However, after a disease course of several years, non-MS-like lesions were found in 50% of patients, whereas MS-like lesions were present in only 10% of cases Only a few NMO patients have undergone brain pathology analysis We recently demonstrated that NMO patients have cognitive dysfunctions, a finding subsequently confirmed by others In the present study, we aimed to investigate whether there are correlations between cognitive dysfunctions and global and/or focal GM and WM volume/concentration in NMO patients.All NMO patients and healthy subjects gave their written informed consent to participate in the study. The study was approved by the University Hospital of Strasbourg ethics committee.Twenty-eight patients with definite NMO according to the recently revised criteria Batterie Courte d'évaluation cognitive destinée aux patients souffrant de Sclérose En Plaques) We assessed NMO patients using the the BCcogSEP . Brain MRI was performed the same day as the neuropsychological assessment.(1) 3D high resolution T1-weighted (MPRAGE) images ;2, matrix 256×192, slice thickness 4.0 mm);(2) Axial turbo spin-echo (TSE) T2-weighted sequence  Axial FLAIR T2-weighted sequence .Global brain volume was estimated for each subject with high-resolution T1-weighted images according to the SIENAx method  The following sequences were acquired:Focal brain volume loss was also analyzed and compared for patients and controls with high-resolution T1-weighted images according to the method for VBM with SPM5  and Matlab 7.7 software .Briefly, this method involves an initial segmentation of T1-weighted MRI into GM and WM images in native space, followed by a normalization of the GM and WM images to GM and WM templates in stereotactic space. To improve segmentation, the normalization parameters obtained from the nonlinear normalization of the GM and WM images are re-applied to the original whole-brain MRI and the normalized whole-brain MRI is then segmented into WM and GM images in a second segmentation step. GM and WM concentration images  were then smoothed with a 12-mm full-width half-maximum (FWHM) Gaussian kernel.P value <0.05 and with a minimum cluster size of 30 voxels. MNI Space Utility , xjView software and Talairach software  were used to create reports about cluster localization in terms of Talairach Daemon anatomical region labels.We performed voxel-level correlations of GM or WM concentration between patients and controls, and also between NMO patients with and without cognitive impairment, using a two-sample t-test, without covariates. Statistical maps were thresholded with false discovery rate (FDR) correction with We considered patients to have abnormal cognitive performance if their results were inferior to the 5th percentile, using the norms of Dujardin P value ≤0.01 was considered statistically significant, a P value between 0.01 and 0.05 was considered as a trend, and a P value >0.05 was considered not significant.We looked for correlation between cognitive dysfunction and the following parameters: disease duration, EDSS, treatments, NMO antibodies, and also global brain volume, grey matter volume, and white matter volume . For this statistical analysis, we used the Pearson test for continuous variables, and the Spearman rank correlation test for ordinal data, using SPSS 18. To correct for multiple comparisons, a The nonparametric Mann-Whitney U test was used to compare total brain, GM, and WM volumes between patients and controls (SPSS 18).P value <0.001 and with a minimum cluster size of 30 voxels.We performed voxel-level correlations between brain concentration and cognitive performance for each subtest of the BCcogSEP using residual scores with SPM5. An explicit mask was used to ensure that only brain region with decrease concentration of matter were included in the analysis. Statistical maps were thresholded without correction with The main demographical, clinical and laboratory results are summarized in Fifteen NMO patients (54%) had cognitive impairment with more than four subtests of the BCcogSEP inferior to the 5th percentile. The results of the cognitive tests are summarized in P = 0.002, rho = –0.56) and EDSS score. A trend towards a correlation was found between PASAT 2s  PASAT 3s , and EDSS.Correlations were found between BCcog-SRT learning , sex (P = 0.36), educational level (P = 0.91), EDSS (P = 0.12) and duration of the disease (P = 0.14).The comparison of characteristics of NMO patients with cognitive impairment (N = 15) to NMO patients without cognitive impairment (N = 13) showed no difference in term of age . WM brain volume of NMO patients was also reduced compared to controls . No difference in GM brain volume was found between NMO patients and controls . Mean whole brain volume of NMO patients was 1582244.1 mm3±92733.8 and mean whole brain volume of controls was 1645341.6 mm3±68529.5. Mean WM brain volume of NMO patients was 762744.1 mm3 ± 59667.7, and mean WM brain volume of controls was 816182.4 mm3±66658.9. Mean GM brain volume of NMO patients was 819500.1 mm3±79067.8, and mean GM brain volume of controls was 829159.2 mm3±87229.7.Using SIENAx, global brain volume of NMO patients was reduced compared to controls . A trend towards a correlation was found between the number of subtests inferior to the 5th percentile to the BCcogSEP and normalized WM volume .Correlation was found between the number of subtests inferior to 5P = 0.024, rho = 0.40) and PASAT 2s .A trend towards a correlation was found between normalized WM volume and PASAT 3s . WM brain volume of NMO patients with cognitive impairment was also reduced compared to WM volume of NMO patients without cognitive impairment . No difference was found between GM brain volume of NMO with and without cognitive impairment .Using SIENAx, global brain volume of NMO patients with cognitive impairment (N = 15) was reduced compared to global brain volume of NMO patients without cognitive impairments (N = 13)  and regional white matter volume in the NMO group.Correlations were found between regional white matter volume and the PASAT 3s, DSST, phonemic fluency, forward and backward digit span, BCcog-SRT and 10/36 spatial memory test, as shown in BCcog-SRT was correlated to the fornix, and the WM of middle frontal gyrus, hippocampus and lingual gyrus. Backward digit span was correlated to WM of the left medial frontal gyrus, and forward digit span to the left middle temporal gyrus. Phonemic fluency was correlated to the WM of thalamus.DSST was correlated to the WM of the middle frontal gyrus and precentral gyrus. PASAT 3s was correlated to the WM of corpus callosum, frontal and parietal regions and the pons .Two main structures were correlated with 10/36 immediate recall: parietal lobe and limbic lobe, including parahippocampal structures. Many white matter structures were correlated with 10/36 delayed recall: parts of the visual system , parts of the frontal lobe, part of the limbic system including parahippocampal regions, part of the parietal lobe including post-central gyrus and inferior parietal lobule, and the vermis.We did not find any focal GM volume difference between NMO patients with cognitive impairment (n = 15) and without cognitive impairment. On the contrary, a large decrease of the WM when we compared NMO patients with cognitive impairment to NM patients without cognitive impairment was found . These dOur data show that 54% of NMO patients had cognitive impairment according to their BCcogSEP performances. We also found a decreased global and focal white matter volume in NMO patients compared to healthy subjects. Furthermore, we observed that the majority of the WM is decreased in NMO patients with cognitive impairment compared to NMO patients without cognitive impairment. We observed also correlations between cognitive tests performance and white matter volume. We did not find any global and focal GM differences between NMO patients and controls, and also between NMO patients with and without cognitive impairment.We demonstrated cognitive impairment in more than half of NMO patients in a previously published multicentre study In the previous two studies, there was no correlation between brain lesions and cognitive impairmentWe wanted also to better understand the relationship between white and grey matter volumes and cognitive functions in NMO patients. We did not find any decrease of brain GM volume in NMO patients, using SIENAx for global volume and VBM in SPM5 for focal volume. Moreover, the comparison of global and focal GM of NMO patients with and without cognitive impairment did not show any decreased volume. Similarly using MR spectroscopy of the brain we did not find any abnormality in the GM in an earlier study We found a decrease in WM brain volume in NMO patients, using SIENAx for global volume and VBM in SPM5 for focal volume. This result, found using two different methods, strengthens the discovery of such atrophy. Abnormalities of NAWM in the brain of NMO patients were previously found using diffusion tensor imaging We also observed correlations between cognitive test performances and WM volumes. First, we found an association between low performance for immediate and delayed spatial memory  and a reduced volume of the optic chiasm, the corpus callosum, limbic lobe including parahippocampal gyri and fronto-parieto-occipital regions including the superior longitudinal fascicle. Visual impairment could explain difficulties in performing a visual memory task. Thus, the correlation found with the optic chiasm is logical. Chronic disconnection of the corpus callosum by surgery is known to be responsible for moderate memory impairment, particularly topographical memory Reduced WM in the pons was correlated with poor performances on PASAT. Norepinephrine-synthesizing neurons that send diffuse projection from a part of the pons, the locus coeruleus, have a major role in attention, particularly in focused attention and the ability to redirect attention He et al., have recently demonstrated significantly correlations between corpus callosum, frontal regions and cognitive tests concerning verbal memory and speed of information processing The diminished WM volume of NMO patients with cognitive impairment compared to NMO without cognitive impairment is highly diffuse . The majWM atrophy detected with two methods and correlations between WM and cognitive tests, are important arguments in favour of brain WM involvement in NMO. NMO is an inflammatory disease of the CNS. AQP4 is known to be the main target antigen involved in this disease. AQP4 is a bidirectional water channel, found on all surfaces of astrocytes. The highest concentration of AQP4 is in the perivascular and peripial end-feet, in direct contact with the endothelium and pia mater, respectively. AQP4 immunoreactivity of the WM in the brain is highest and mesh-like in the chiasm, the optic nerves and the brainstem, particularly in the pons (at the floor of the fourth ventricle) There are some limitations to our study. Firstly, our cohort of NMO patients has only 43% of patients with NMO-IgG positivity. In North American patients, particularly in the Mayo Clinic , NMO-IgG positivity was reported to be 73% Fifty-four percent of the NMO patients in our study had cognitive impairment according to their performance on the French version of the BRB-N. We found also a decreased global and focal white matter volume in the brain of NMO patients compared to healthy subjects. Furthermore, NMO patients with cognitive impairment have an important atrophy of brain WM compared to NMO patients without cognitive impairment. We observed also correlations between cognitive tests performances and WM concentrations. Further studies are now needed to elucidate more precisely the origin of cognitive impairment in the brain of NMO patients, particularly in the WM. We are therefore planning to use diffusion tensor imaging (DTI) to analyse the correlations between cognitive impairment and DTI parameters. More pathological data are also needed to better understand the mechanisms of cognitive impairment in NMO patients.