Testicular function in males with infantile nephropathic cystinosis.

STUDY QUESTION: Do males with the rare lysosomal storage disease infantile nephropathic cystinosis (INC) have a chance of biological fatherhood? SUMMARY ANSWER: Cryostorage of semen could be an option for approximately 20% of young males with INC, with surgical sperm retrieval from the centre of the testes providing additional opportunities for fatherhood. WHAT IS KNOWN ALREADY: Biallelic mutations in the cystinosin (CTNS) gene in INC cause dysfunction in cystine transport across lysosomal membranes and cystine accumulation throughout the body. Spontaneous paternity in cystinosis has not been described, despite the availability of cysteamine treatment. Azoospermia has been diagnosed in small case series of males with INC. ART using ICSI requires few spermatozoa, either from semen or extracted surgically from the testes of azoospermic men. However, there is limited evidence to suggest this could be successful in INC. STUDY DESIGN, SIZE, DURATION: In this prospective cohort study performed between 2018 and 2019, we performed a cross-sectional investigation of 18 male patients with INC to delineate endocrine and spermatogenic testicular function. PARTICIPANTS/MATERIALS, SETTING, METHODS: Serum hormone levels, semen samples (according to World Health Organization 2010 standards), and testicular ultrasound images were analysed in 18 male patients aged 15.4-40.5 years. Surgical sperm extraction was performed in two, and their testicular biopsies were investigated by light and electron microscopy. Past adherence to cysteamine treatment was assessed from medical record information, using a composite scoring system. MAIN RESULTS AND THE ROLE OF CHANCE: Adherence to cysteamine treatment was high in most patients. Testicular volumes and testosterone levels were in the normal ranges, with the exception of two and three older patients, respectively. Serum LH levels were above the normal range in all subjects aged ≥20 years. FSH levels were elevated in all but four males: three with spermatozoa in semen and one adolescent. Inhibin B levels were shown to be lower in older men. Testicular ultrasound revealed signs of obstruction in 67% of patients. Reduced fructose and zinc seminal markers were found in 33%, including two patients with azoospermia who underwent successful surgical sperm retrieval. Histology identified fully preserved spermatogenesis in the centre of their testes, but also tubular atrophy and lysosomal overload in Sertoli and Leydig cells of the testicular periphery. LIMITATIONS, REASONS FOR CAUTION: Limitations of this study are the small number of assessed patients and the heterogeneity of their dysfunction in cystine transport across lysosomal membranes. WIDER IMPLICATIONS OF THE FINDINGS: This study suggests that testicular degeneration in cystinosis results from the lysosomal overload of Sertoli and Leydig cells of the testicular periphery, and that this can possibly be delayed, but not prevented, by good adherence to cysteamine treatment. Endocrine testicular function in INC may remain compensated until the fourth decade of life; however, azoospermia may occur during adolescence. Cryostorage of semen could be an option for approximately 20% of young males with INC, with surgical sperm retrieval providing additional opportunities for biological fatherhood. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the Cystinosis Foundation Germany. The authors have no competing interests to declare. TRIAL REGISTRATION NUMBER: n/a.

A history of undescended testes in young men with Klinefelter syndrome does not reduce the chances for successful microsurgical testicular sperm extraction.

Klinefelter syndrome (KS) and undescended testes (UDT) are known etiologies for non-obstructive azoospermia (NOA), and coexistence of both etiologies is not uncommon. Patients with both KS and a history of UDT are therefore considered to have extremely reduced chances for paternity. We aimed to analyze the impact of previous surgically corrected unilateral or bilateral UDT on sperm retrieval rates (SRRs) by microsurgical testicular sperm extraction (mTESE) in azoospermic men with KS. Age, testicular volumes, and hypothalamo-pituitary-gonadal axis function were investigated in relation to SRRs in 29 non-mosaic KS patients (47,XXY) with a history of UDT (group 1) who underwent mTESE between 2008 and 2016 in our center and compared to the data of age- and serum testosterone-matched non-mosaic KS controls with eutopic testes at birth (group 2), and to those of 51 men with NOA and a normal male karyotype (46,XY), but previous UDT (group 3). SRRs in KS patients with surgically corrected UDT during childhood were comparable to SRRs of KS patients with eutopic testes at birth: 31% (35% in unilateral and 22% in bilateral UDT) vs. 38% (p = 0.581). SRRs and Leydig cell function in group 1 were negatively correlated with age. Significantly higher SRRs (66%) were found in euploid azoospermic men with surgically corrected UDT (p < 0.001). A history of UDT does not preclude chances for future fatherhood in young azoospermic males with KS. In one of three men with previous unilateral UDT and in one of 4-5 in those with previous bilateral UDT, spermatozoa can be harvested by mTESE during late adolescence or young adulthood for immediate or future use in assisted reproduction.

Gonadal function in adult male patients with congenital adrenal hyperplasia.

CONTEXT: Current knowledge on gonadal function in congenital adrenal hyperplasia (CAH) is mostly limited to single-center/country studies enrolling small patient numbers. Overall data indicate that gonadal function can be compromised in men with CAH. OBJECTIVE: To determine gonadal function in men with CAH within the European 'dsd-LIFE' cohort. DESIGN: Cross-sectional clinical outcome study, including retrospective data from medical records. METHODS: Fourteen academic hospitals included 121 men with CAH aged 16-68 years. Main outcome measures were serum hormone concentrations, semen parameters and imaging data of the testes. RESULTS: At the time of assessment, 14/69 patients had a serum testosterone concentration below the reference range; 7 of those were hypogonadotropic, 6 normogonadotropic and 1 hypergonadotropic. In contrast, among the patients with normal serum testosterone (55/69), 4 were hypogonadotropic, 44 normogonadotropic and 7 hypergonadotropic. The association of decreased testosterone with reduced gonadotropin concentrations (odds ratio (OR) = 12.8 (2.9-57.3)) was weaker than the association between serum androstenedione/testosterone ratio ≥1 and reduced gonadotropin concentrations (OR = 39.3 (2.1-732.4)). Evaluation of sperm quality revealed decreased sperm concentrations (15/39), motility (13/37) and abnormal morphology (4/28). Testicular adrenal rest tumor (TART)s were present in 39/80 patients, with a higher prevalence in patients with the most severe genotype (14/18) and in patients with increased current 17-hydroxyprogesterone 20/35) or androstenedione (12/18) serum concentrations. Forty-three children were fathered by 26/113 patients. CONCLUSIONS: Men with CAH have a high risk of developing hypothalamic-pituitary-gonadal disturbances and spermatogenic abnormalities. Regular assessment of endocrine gonadal function and imaging for TART development are recommended, in addition to measures for fertility protection.

Impact of various progestins with or without transdermal testosterone on gonadotropin levels for non-invasive hormonal male contraception: a randomized clinical trial.

Although several progestins have been tested for hormonal male contraception, the effects of dosage and nature of various progestins on gonadotropin suppression combined with and without additional testosterone has not been performed in a comparative trial. The aim of this study was to evaluate the differential impact of four oral or transdermal progestins on the suppression of gonadotropins in healthy men: oral: cyproterone acetate (CPA), levonorgestrel (LNG), norethisterone acetate (NETA), and transdermal: Nestorone® (NES), all in combination with transdermal testosterone (T). Randomized clinical trial testing was performed with four progestins at two doses each. After a 2-week progestin-only treatment, transdermal T was added for further 4 weeks and was followed by a 3-week recovery period. Progestin-dose per day: CPA 10 mg/20 mg, NES 2 mg/3 mg/dose e.g. 200/300 µg/day absorbed, NETA 5 mg/10 mg, LNG 120 µg/240 µg. From an andrology outpatient clinic, 56 healthy men aged 18-50 years, with body mass index ≤33 kg × m-2 were included in the study. Serum concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were studied. Secondary outcome measure included were serum testosterone concentrations, sperm concentrations, and safety parameters. Intergroup comparisons demonstrated that CPA and LNG had the strongest effect on LH/FSH suppression. Nevertheless, every substance showed significant inhibitory effects on gonadotropin secretion, especially in combination with transdermal T. A decrease in hematocrit and insulin sensitivity as well as cholesterol subfractions and triglycerides was uniformly seen for every group. The combination of oral or transdermal progestins with a transdermal testosterone preparation is able to suppress gonadotropins. Further dose titration studies with sperm suppression as an end-point should be conducted to determine the lowest effective dose for hormonal male contraception.

Testicular function during puberty and young adulthood in patients with Klinefelter's syndrome with and without spermatozoa in seminal fluid.

Patients with Klinefelter's syndrome experience progressive testicular degeneration resulting in impaired endocrine function and azoospermia. What proportion of adolescents develop testosterone deficiency during puberty and how many have spermatozoa in their semen is unclear to date. We aimed to investigate testicular function during puberty and young adulthood in patients with Klinefelter's syndrome and to assess testosterone effects in target tissues. The clinical data of 281 patients with non-mosaic Klinefelter's syndrome aged 10-25 years without previous testosterone replacement were reviewed. In late pubertal adolescents, semen analyses were evaluated, and testicular volumes, hormone and haemoglobin (Hb) levels, the number of CAG repeats and final height data were compared to those of 233 age-matched controls with pubertal gynaecomastia. Spontaneous pubertal virilisation to Tanner stages IV-V occurred. Serum T levels ≥10 nmol/L were reached in 62% of patients with Klinefelter's syndrome and in 85% of controls at ages 15-25 (TKFS : 12.2 ± 5.4 vs. TC : 16.6 ± 7.2 nmol/L). LHKFS levels were elevated >10 U/L in 84%, and normal in all controls (LHKFS : 18.6 ± 12.2 vs. LHC : 3.5 ± 1.6 U/L). In nine of 130 (7%) adolescents with Klinefelter's syndrome, spermatozoa (oligozoospermia) were found in semen; all had T levels >7 nmol/L and eight of nine had LH levels ≤18 U/L, while their hormone levels, number of CAG repeats and testicular volumes were not different from those of adolescents with azoospermia. Controls had normal sperm concentrations in 73% (46/63). Semen volumesKFS were normal in 55% vs. 78% in controls; HbKFS was normal in 89% (HbC : 97%). Mean final heightKFS was 185 ± 8 cm vs. 181 ± 7 cm in controls. Hypergonadotropic hypogonadism develops during early puberty in adolescents with Klinefelter's syndrome and remains compensated in over 60% during ages 15-25, with sufficient testosterone secretion for spontaneous accomplishment of pubertal development. Spermatozoa in semen are rare and associated with T levels >7 nmol/L. Parameters reflecting androgen deficiency in target tissues may help to optimise timing of testosterone substitution, which should preferably not be initiated before fertility status has been clarified.

Causes of hypogonadotropic hypogonadism predict response to gonadotropin substitution in adults.

Germ cell and Sertoli cell proliferation and maturation in human testes occur in three main waves, during the late fetal and early neonatal period and at early puberty. They are triggered by periods of increased activity of the hypothalamic-pituitary-gonadal (HPG) axis. In hypogonadotropic hypogonadism (HH), these processes are variably disturbed. The objective of this study was to explore whether success of gonadotropin replacement in HH men is predictable by the origin of HH, indicating time of onset and severity of GnRH/gonadotropin deficiency. The data of 51 adult HH patients who had undergone one cycle of hCG/FSH treatment were reviewed. Five groups were established, according to the underlying HH origin. Therapeutic success by final bi-testicular volumes (BTVs) final sperm concentrations (SC) and conception rates were compared and related to baseline parameters, indicative of the degree of HPG-axis disruption. Overall, BTVs rose from 13 ± 15 to 27 ± 15 mL, spermatogenesis was induced in 98%, with mean SCs of 15 ± 30 mill/mL, spontaneous pregnancies in 37% and additional 18% via intracytoplasmic sperm injection. Kallmann syndrome patients had the poorest responses (BTV: 16.9 ± 10 mL; SC: 3.5 ± 5.6 mill/mL), followed by patients with congenital/infancy-acquired multiple pituitary hormone deficiencies (MPHD) and patients with HH+absent puberty (BTV: 21 ± 14/24 ± 9 mL; SC: 5.5 ± 6.5/ 14.5 ± 23.8 mill/mL). HH men with pubertal arrest and with post-pubertally acquired MPHD had the best results (BTV: 36 ± 14/38 ± 16 mL; SC: 25.4 ± 34.2/29.9 ± 50.5 mill/mL). Earlier conception after 20.3 ± 11.5 months (vs. 43.1 ± 43.8; p = 0.047) of gonadotropin treatment with higher pregnancy rates (62% vs. 42%) was achieved in the two post-pubertally acquired HH subgroups, compared to the three pre-pubertally acquired. Therapeutic success was higher in patients without previously undescended testes, with higher baseline BTVs (pre- vs. post-pubertal HH: 5 ± 4 mL vs. 26 ± 16 mL; p < 0.0001) and higher baseline inhibinB levels (pre- vs. post-pubertal HH: 16.6 vs. 144.5 pg/mL; p = 0.0004). The cause of HH is a valuable predictor of outcome of gonadotropin replacement in adults.

Inhibin B, AMH, but not INSL3, IGF1 or DHEAS support differentiation between constitutional delay of growth and puberty and hypogonadotropic hypogonadism.

In pre-pubertal boys ≥ 14 years, the differentiation between constitutional delay of growth and puberty (CDGP) and hypogonadotropic hypogonadism (HH) is challenging, as current diagnostic tools have limitations in sensitivity and specificity. The aim of this study was to assess the usefulness of markers of gonadal activity, growth axis activation and adrenarche in differentiation between pre-pubertal CDGP and HH. This retrospective study was carried out between 2006 and 2015 in an academic out-patient referral centre. The clinical data of 94 boys, aged 13.9-23.2 years and referred for "pubertal delay" were reviewed. Definite diagnoses were established on initial work-up and clinical follow-up: 24 boys were diagnosed with HH, 22 boys with CDGP, pre-pubertal (PP CDGP) at referral and 28 boys with CDGP, early pubertal at referral (EP CDGP), the latter serving as control group. Twenty patients were excluded from evaluation because of previous sex steroid treatment or associated chronic disease. Inhibin B and AMH were measured in all (n = 74); INSL3, IGF1, IGFBP3 and DHEAS in a subset of patients (n = 45) in serum of first presentation. Inhibin B and AMH were higher in boys with PP CDGP than in boys with HH: inhibin B: 87.6 ± 42.5 vs. 19.8 ± 13.9 pg/mL; p < 0.001; AMH: 44.9 ± 27.1 vs. 15.4 ± 8.3 ng/mL; p < 0.001. Receiver operating characteristics (ROC) for the diagnosis of PPCDGP vs. HH (inhibin B ≥ 28.5 pg/mL): sensitivity: 95%, specificity: 75%; AUC: 0.955. In combination with an AMH cut-off ≥20 ng/mL the specificity increased to 83%. INSL3, IGF1, IGFBP3 and DHEAS levels were not different. In boys with EP CDGP, inhibin B and IGF1 levels were highest (138.7 ± 59.9 pg/mL/289.7 ± 117 ng/mL), whereas AMH levels were lowest (11.7 ± 9.1 ng/mL). Sertoli cell markers are helpful for establishing a prognosis, whether a boy with pubertal delay will enter puberty spontaneously, whereas Leydig cell, growth and adrenal markers are not.

Age and markers of Leydig cell function, but not of Sertoli cell function predict the success of sperm retrieval in adolescents and adults with Klinefelter's syndrome.

Microsurgical testicular sperm extraction (mTESE), combined with intracytoplasmic sperm injection (ICSI) represents a chance for azoospermic men with Klinefelter's syndrome (KS) to father children. The objective of this study was to identify predictive factors for the success of mTESE from adolescents and adults with KS. The clinical data of 50 late pubertal adolescents (13-19 years) and 85 adult patients (20-61 years) with non-mosaic KS, who underwent mTESE, were analysed with respect to factors, potentially predictive of active spermatogenesis; specifically a history of cryptorchidism, age, testicular volumes, serum levels of LH, FSH, testosterone (T) and estradiol at the time of surgery. Inhibin B, AMH and INSL3 were additionally analysed in the adolescents. A younger age and a near-compensated Leydig cell function were associated with higher success of sperm retrieval via mTESE: In adolescents ≥15-19 years, spermatozoa were retrieved in 45%, compared to 31% in adults; in adolescents aged 13-14 years, spermatozoa were collected in only 10%. Adolescents with an LH ≤17.5 U/L, along with a T level ≥7.5 nmol/L had the best success rate (54%), which fell to 44% with higher LH, whereas those with low T (<7.5 nmol/L), irrespective of LH had no sperm retrieval. In adults with T levels above and LH below these thresholds, the success rate was 51%, falling to 19%, if LH was higher. When T was lower than threshold, the rate was 17%. No association between testicular volumes, serum levels of FSH, Inhibin B, AMH, estradiol and mTESE success was found. A history of cryptorchidism was associated with lower retrieval rates. A window of opportunity for an approximate 50% chance to retrieve spermatozoa via mTESE exists for young, late pubertal KS patients between age 15 and young adulthood, when Leydig cell function is at its best. In these cases, referral to a centre of expertise should be considered.

Development of specific dengue virus 2'-O- and N7-methyltransferase assays for antiviral drug screening.

Dengue virus (DENV) protein NS5 carries two mRNA cap methyltransferase (MTase) activities involved in the synthesis of a cap structure, (7Me)GpppA(2'OMe)-RNA, at the 5'-end of the viral mRNA. The methylation of the cap guanine at its N7-position (N7-MTase, (7Me)GpppA-RNA) is essential for viral replication. The development of high throughput methods to identify specific inhibitors of N7-MTase is hampered by technical limitations in the large scale synthesis of long capped RNAs. In this work, we describe an efficient method to generate such capped RNA, GpppA(2'OMe)-RNA74, by ligation of two RNA fragments. Then, we use GpppA(2'OMe)-RNA74 as a substrate to assess DENV N7-MTase activity and to develop a robust and specific activity assay. We applied the same ligation procedure to generate (7Me)GpppA-RNA74 in order to characterize the DENV 2'-O-MTase activity specifically on long capped RNA. We next compared the N7- and 2'-O-MTase inhibition effect of 18 molecules, previously proposed to affect MTase activities. These experiments allow the validation of a rapid and sensitive method easily adaptable for high-throughput inhibitor screening in anti-flaviviral drug development.

[Androgen deprivation therapy in prostate cancer. Indication and systemic consequences]. / Antiandrogene Therapie des Prostatakarzinoms: Indikation und systemische Folgen.

Androgen deprivation therapy (ADT) is effective in prolonging the progression free survival of patients with symptomatic/metastatic prostate cancer (PC). The reduction of clinical symptoms of tumour disease and the reduction of tumour growth and metastatic dissemination is accompanied by systemic consequences of testosterone deficiency. These are hot flushes, fatigue due to reduction of muscular strength and muscle mass as well as anaemia. Moreover, patients develop cognitive impairment und depressive mood. Weight gain with insulin resistance, disturbances of lipid metabolism and gynecomastia are other effects of androgen deficiency. A decrease in bone mineral density may lead to an increased susceptibility to bone fractures. There are several options to reduce these side effects of ADT, e.g. physical activity, dietary supplementation, tailored pharmacological therapy and psychotherapy. The knowledge of these adjuvant treatment options, despite their palliative character, is relevant to optimize the quality of life of these patients.

[Awareness and knowledge about risks of drinking during pregnancy in pregnant Lebanese women]. / Les conduites d'alcoolisation des femmes enceintes libanaises et leurs connaissances sur les risques et répercussions sur l'enfant à naître.

OBJECTIVE: To assess the awareness and knowledge of pregnant Lebanese women about the risks of drinking during pregnancy and the factors that influence their drinking patterns. MATERIALS AND METHODS: A prospective study was conducted on a sample of 107 women consulting the gynecology outpatient department of Hôtel-Dieu de France in Beirut, Lebanon, who completed the T-ACE screening test included in a 21 multiple choice questionnaire which examine knowledge and beliefs about alcohol use during pregnancy, drinking patterns and awareness of fetal alcohol exposure. RESULTS: The 107 women of our sample were all married, between 20 and 41 years old and had mostly a high educational level (86%). Most of the women (47%) were at their first pregnancy. Of the 20 women who self-reported drinking during pregnancy, 60% obtained a positive score on the T-ACE questionnaire, which indicates that more than 11% of the women engaged with potentially high risk drinking for the baby. There is not a significant difference between the different age categories or educational levels. This proportion is lower than that found in international publications. However, the rate of excessive drinking (4 drinks or more on any one occasion in females) was higher and one woman in five reported excessive drinking in the previous year. There is a high level of knowledge that alcohol use during pregnancy is harmful to the child, and the more consumption the more harmful and likely the effects, but there is confusion about the safety of small amounts of alcohol. Women (37%) think that there is a safe level of drinking during pregnancy; 29% tolerate up to one drink a month, 9% tolerate up to one drink a week and one woman thinks having one drink a day is safe. Women who actually drink during pregnancy are more likely to think that alcohol consumption to a certain level is safe. Women (31%) think that beer and/or wine are safe alcohols to a certain level during pregnancy. When asked about the source of this belief, 22% mention a gynecologist but the majority (61%) says it is a personal belief. Women (65%) in our sample are aware that alcohol use during pregnancy can lead to life-long disabilities in a child, such as delayed development (36%), birth defects/deformities (35%) and mental retardation (32%). However, up to 33% of the respondents report having no information about the effects of alcohol on the fetus and two women believe alcohol is not harmful at all. Women with lower levels of education are somewhat less knowledgeable about the risks of alcohol use during pregnancy than those with higher levels of education. There is no association between the drinking patterns of the women with their age, their professional habits and the alcohol consumption of their husbands. The women in our sample seem to be more aware of the necessity to stop smoking rather than stop drinking during pregnancy. CONCLUSION: Lebanese women are not fully aware of the recommendations and risks related to drinking during pregnancy. This is the reason why action must be taken to ensure better diffusion of these recommendations and better assessment of alcohol intake during prenatal visits.

Norovirus seasonality and the potential impact of climate change.

Seasonal variation in norovirus infection is a recognized but poorly understood phenomenon. It is likely to be based on biological, environmental and behavioural factors that regulate transmission, virulence and persistence of the virions in host populations. Understanding the seasonal dependency of norovirus infection is an important step towards understanding its epidemiology, with subsequent implementation of efficient measures of surveillance and control. Whether or not climate change could influence the seasonal patterns of norovirus infection, by impacting on its transmission, geographic distribution and prevalence, has not yet been considered. This review addresses the question.

[Predictors of prophylactic response to lithium]. / Prédicteurs de réponse prophylactique au lithium.

INTRODUCTION: Since their first utilization in psychiatry as mood stabilizers in the 1940s, lithium salts have been widely studied in the medical literature. The considerable amount of data available to date, supports the use of lithium salts as first-line mood stabilizing agents, with acute antimanic and antidepressant properties and proven efficacy in the long term prevention of manic and depressive relapses. LITERATURE FINDINGS: Several predictors were reported by different authors in early articles and were confirmed later on by the medical literature. All the psychopathological, environmental, biological, neurophysiologic and genetic predictors known to date are reviewed here. PSYCHOLOGICAL PREDICTORS: Psychopathological predictors of a good response to lithium prophylaxis include: the initial good response to lithium during the first 6-12 months of treatment, considered to date to be the most reliable predictor of a favourable response to lithium; the classical pattern of elated manic episodes; a positive familial history of bipolar disorders, especially those known to be responsive to lithium; the absence of comorbid personality disorders; bipolar type I disorders; melancholic features during depressive episodes; MDI pattern in the illness course and early onset of lithium treatment. In contrast, the following have been confirmed as psychopathological predictors of poor prophylactic lithium response: mixed episodes, considered to be one of the most reliable predictors of poor response to lithium since Kraeplin's description; rapid cycling bipolar disorders; comorbid alcohol and/or drug abuse; mood disorders with incongruent psychotic features; early onset bipolar disorder before the age of 18; discontinuation of lithium treatment; high number of previous affective episodes in the illness course before lithium initiation and DMI pattern. ENVIRONMENTAL PREDICTORS: Among environmental factors, being single was found to be the only predictor of a poor response to lithium treatment in prophylaxis. BIOLOGICAL PREDICTORS: Biological predictors of a good prophylactic response to lithium include a high RBC/plasma-lithium ratio, one of the most controversial predictors of a favourable response to lithium in the literature, a higher platelet serotonin-induced calcium mobilization, and a high rate of red blood cell membrane phospholipids, especially of phosphatidylcholine, and a phospholipid implicated in lithium intracellular transport. Among neurophysiologic predictors of a favourable response to lithium, the following have been reported: brain lithium concentrations above 0.2 mEq/L when measured by 7Li-MRS; decreased cerebral intracellular pH and white matter hyper intensity at (31)P-MRS and a high intensity of loudness dependence auditory-evoked potentials (LDAEP), the latter being one of the best indicators of human cerebral serotoninergic functioning. In contrast, the following have been reported as neurophysiological predictors of a poor lithium response in prophylaxis: epileptiform anomalies with diffuse theta waves on electroencephalography, a predictor of poor response to lithium known since the descriptions of Dalen in 1965 and decreased cerebral phosphocreatine levels at (31)P-MRS, the latter being an indicator of cerebral mitochondrial dysfunction. GENETIC PREDICTORS: Genetic predictors of good response to lithium in prophylaxis include a lower-inositol-monophosphatase (IMPase-2) mRNA expression, IMPase-2 being a key enzyme of the calcium-intracellular-signalling pathway and IMPase-2 gene being studied recently as a candidate gene in bipolar disorder. A higher frequency of phospholipase C isoenzyme gamma1 (PLCG1)-5 repeat allele genes has also been associated with a good response to lithium, PLCG1 being a major enzyme of the phosphatidylinositol second messenger system. Genetic predictors of negative prophylactic lithium response include the homozygotic forms of the short allele of the serotonin transporter gene (5-HTT), the presence of the A/A subtype of tryptophan hydroxylase (TPH) gene and a high frequency of human leukocyte antigens type A3 (HLA-A3), this genotype being associated with cellular membrane anomalies implicated in alteration of lithium intracellular transport. DISCUSSION: The search for new predictors of lithium prophylactic response is currently facing several methodological problems: lack of representativity of the samples of bipolar patients enrolled in research studies, poor reliability of retrospective reconstructions of the course of the bipolar disorder before initiation of lithium treatment, absence of consensus on tools used to assess response to lithium prophylaxis in study designs, difficult access and high costs of most of the laboratory and neuroimaging techniques used in recent studies such as magnetic resonance spectroscopy and LDAEP measures, and problematic evaluation of the impact of treatment on a disorder whose natural intrinsic course is often irregular.

The VIZIER project: preparedness against pathogenic RNA viruses.

Life-threatening RNA viruses emerge regularly, and often in an unpredictable manner. Yet, the very few drugs available against known RNA viruses have sometimes required decades of research for development. Can we generate preparedness for outbreaks of the, as yet, unknown viruses? The VIZIER (VIral enZymes InvolvEd in Replication) (http://www.vizier-europe.org/) project has been set-up to develop the scientific foundations for countering this challenge to society. VIZIER studies the most conserved viral enzymes (that of the replication machinery, or replicases) that constitute attractive targets for drug-design. The aim of VIZIER is to determine as many replicase crystal structures as possible from a carefully selected list of viruses in order to comprehensively cover the diversity of the RNA virus universe, and generate critical knowledge that could be efficiently utilized to jump-start research on any emerging RNA virus. VIZIER is a multidisciplinary project involving (i) bioinformatics to define functional domains, (ii) viral genomics to increase the number of characterized viral genomes and prepare defined targets, (iii) proteomics to express, purify, and characterize targets, (iv) structural biology to solve their crystal structures, and (v) pre-lead discovery to propose active scaffolds of antiviral molecules.

Efficient transduction and long-term retroviral expression of the melanoma-associated tumor antigen tyrosinase in CD34(+) cord blood-derived dendritic cells.

Differentiation of genetically modified CD34(+) hematopoietic stem cells into dendritic cells (DCs) will contribute to the development of immunotherapeutic anticancer protocols. Retroviral vectors that have been used for the transduction of CD34(+) cells face the problem of gene silencing when integrated into the genome of repopulating stem cells. We reasoned that a high copy number of retroviral DNA sequences might overcome silencing of transgene expression during expansion and differentiation of progenitor cells into functional DCs. To prove this, we utilized a retroviral vector with bicistronic expression of the melanoma-associated antigen tyrosinase and the enhanced green fluorescent protein (EGFP). Human cord blood CD34(+) cells were transduced with vesicular stomatitis virus G-protein (VSV-G) pseudotyped Moloney murine leukemia virus (MoMuLV) particles using 100-150 multiplicity of infection. During expansion of transduced cells with immature phenotype, transgene expression was strongly silenced, but upon differentiation into mature DCs, residual transgene expression was retained. Intracellular processing of the provirally expressed tyrosinase was tested in a chromium release assay utilizing a cytotoxic T cell clone specific for a HLA-A*0201-restricted tyrosinase peptide. We suggest that retroviral transduction of tumor-associated antigens in hematopoietic progenitor cells and subsequent differentiation into DCs is a suitable basis for the development of potent anti-tumor vaccines.

Quantification of antigen-reactive T cells by a modified ELISPOT assay based on freshly isolated blood dendritic cells.

The enzyme-linked immunospot (ELISPOT) assay has become a widely employed method for quantification of antigen-reactive T lymphocytes. In recent years, various types of antigen-presenting cells (APCs) have been tested as stimulator cells in ELISPOT protocols to achieve a highly sensitive and rapid assay which is not impaired by a marked nonspecific cytokine release. However, the currently available APCs still have disadvantages, such as significant background reactivities, limited sensitivity, and time-consuming preparation procedures. Recently, we succeeded in defining a novel subpopulation of circulating dendritic cells (DCs) that can easily be prepared from human blood. These M-DC8+ DCs proved to be very effective in the induction of antigen-specific T cell responses. In the present study we provide evidence that M-DC8+ DCs are particularly well suited as APCs for the detection of antigen-specific CD8+ T cells after challenge with viral or tumor peptides in ELISPOT assays. In addition, protein-loaded M-DC8+ DCs proved to be quite efficient in the presentation of MHC class II-bound peptides, thus allowing the determination of frequencies of antigen-reactive CD4+ T cells. The use of M-DC8+ DCs as stimulator cells can improve the ELISPOT assay by combining high sensitivity, rapidity, and low background reactivity.

Fatal myocarditis associated with acute parvovirus B19 and human herpesvirus 6 coinfection.

We report on the case of a healthy young boy who developed a fulminant myocarditis due to acute coinfection with erythrovirus (parvovirus B19) and human herpesvirus 6 (HHV-6) in the absence of an antiviral immune response. We suggest that the HHV-6-induced immunosuppression enhanced dissemination of parvovirus B19, which led to fatal myocarditis.

Antibody response to the tumor-associated inhibitor of apoptosis protein survivin in cancer patients.

Antibody reactivity against survivin, a recently identified tumor-associated protein, was determined in sera from patients with lung (n = 51) or colorectal cancer (n = 49). The same collection of sera was tested for the presence of antibodies against p53. Eleven sera from lung cancer patients and four sera from colorectal cancer patients reacted with purified recombinant survivin in an ELISA (21.6% and 8.2%, respectively), whereas four sera from lung cancer patients and nine sera from colorectal cancer patients contained anti-p53 antibodies (7.8% and 18.4%, respectively). The increase in prevalence when anti-survivin and anti-p53 antibodies were determined in parallel was statistically significant (29.4% versus 7.8%, P = 0.005 in lung cancer population; 26.6% versus 8.2%, P = 0.015 in colorectal cancer population). The high prevalence of anti-survivin antibodies makes these antibodies an attractive novel marker for the diagnosis of lung and colorectal cancer, particularly in patients lacking anti-p53 antibodies.