[Feminizing genitoplasty for prepubertal children and teenagers female]. / Génitoplastie féminisante dans les anomalies du développement sexuel de l'enfant pré-pubère et de l'adolescente au Cameroun. À propos de 9 cas.

OBJECTIVE: The aim of this study was to report our experience after 10 years of practice of feminizing genitoplasty in prepubertal and adolescent patients with disorders of sex development (DSD) assigned females as females in a developing country. METHODOLOGY: This was a cross-sectional, descriptive and retrospective study over a period of 9 years. All pre-pubertal (8-12 years) and adolescent patients female sex assigned with DSD who had willfully consented to the surgery with their guardians and underwent feminizing genital surgery were enrolled in the study. Data collection included: age at presentation, precise diagnosis, surgical procedures, complications, cosmetic result and duration of follow-up. Each patient had a precise diagnosis and the surgery was planned after discussion with the multidisciplinary team. Cosmetic results were assessed based on: appearance of the clitoris and separation of the vaginal and urethral openings. RESULTS: Nine patients raised as females with a median age of 8 years (IR: 10.75) were recorded. Surgery was performed at a median age of 11 years (IR: 9.5). In this series, 6 had a 46, XY karyotype with varying diagnoses: partial androgen insensitivity syndrome (n=2); 5-alphareductase insufficiency (n=2); 17-ketoreductase insufficiency (n=2); gonadal dysgenesis with a mutation in the NR5A1 gene (n=2), 2 had ovostesticular DSD, (karyotypes 46, XX), and 1 had mixed gonadal dysgenesis (karyotype 45, X/46, XY). Partial or total gonad(s) removal in accordance with assigned gender was the most common associated procedure. It was bilateral in 7 cases and unilateral in 2 cases. Follow-up ranged from 3 months to 4.5 years (median: 26 months, IR:18.25). One patient had acute urinary retention in the early follow-up. No other complication such as incision bleeding was recorded. The cosmetic appearance of the external genitalia was satisfactory in all patients. CONCLUSION: Feminizing genital surgery in Cameroon remains a major challenge and should seldom be realized without a precise diagnosis. Late age at presentation is peculiar to our setting; however, it gives room for the patients' participation and input to decisions that will have a life-long personal impact on their lives in terms of psychosocial development and fertility. LEVEL OF EVIDENCE: 3.

Are biomarkers associated with sensitivity to physical activity?

BACKGROUND: Activity-based treatments play an integral role in managing musculoskeletal conditions including low back pain. However, while therapeutic exercise has been shown to reduce pain in such conditions, certain individuals experience a paradoxical pain increase in response to exercise. The physiological processes underlying this sensitivity to physical activity (SPA) are not fully understood, however stress and inflammation have been shown to contribute to SPA. The present cross-sectional study investigated whether physiological indicators of stress (cortisol) and inflammation (IL-6) help explain SPA. METHODS: Twenty-seven patients with chronic low back pain and 21 healthy controls completed a 1-h exercise session of standardized physical tasks. SPA was calculated from the difference between post- and pre-exercise pain levels. Participant's saliva was collected at several timepoints for cortisol and IL-6 levels quantification. Their waking cortisol response was calculated to reflect their cortisol regulation. Reactivity of IL-6 and cortisol was calculated to reflect changes in these measures during exercise. RESULTS: IL-6 reactivity was significantly and positively correlated with SPA among participants with low back pain. In contrast, neither cortisol waking response nor cortisol reactivity was significantly correlated within the low back pain group. No significant differences in IL-6 reactivity, cortisol reactivity or cortisol waking response were observed. CONCLUSION: These findings are the first to link SPA to an objective biomarker among people with low back pain. These findings help describe the physiological mechanisms of SPA and can support new clinical research that targets the inflammatory response of patients with chronic low-back pain and elevated SPA. SIGNIFICANCE: This study reveals a correlation between SPA and an objective salivary biomarker of IL-6 in people with low back pain, improving our understanding of this clinically relevant subjective experience.

The effect of smoking cessation counselling in pregnant women: a meta-analysis of randomised controlled trials.

BACKGROUND: Pregnant smokers are often prescribed counselling as part of multicomponent cessation interventions. However, the isolated effect of counselling in this population remains unclear, and individual randomised controlled trials (RCTs) are inconclusive. OBJECTIVE: To conduct a meta-analysis of RCTs examining counselling in pregnant smokers. SEARCH STRATEGY: We searched the CDC Tobacco Information and Prevention, Cochrane Library, EMBASE, Medline and PsycINFO databases for RCTs evaluating smoking cessation counselling. SELECTION CRITERIA: We included RCTs conducted in pregnant women in which the effect of counselling could be isolated and those that reported biochemically validated abstinence at 6 or 12 months after the target quit date. DATA COLLECTION AND ANALYSIS: Overall estimates were derived using random effects meta-analysis models. MAIN RESULTS: Our search identified eight RCTs (n = 3290 women), all of which examined abstinence at 6 months. The proportion of women that remained abstinent at the end of follow up was modest, ranging from 4 to 24% among those randomised to counselling and from 2 to 21% among control women. The absolute difference in abstinence reached a maximum of only 4%. Summary estimates are inconclusive because of wide confidence intervals, albeit with little evidence to suggest that counselling is efficacious at promoting abstinence (odds ratio 1.08, 95% confidence interval 0.84-1.40). There was no evidence to suggest that efficacy differed by counselling type. CONCLUSIONS: Available data from RCTs examining the isolated effect of smoking cessation counselling in pregnant women are limited but sufficient to rule out large treatment effects. Future RCTs should examine pharmacological therapies in this population.

[Outcomes of cumulative transfers of fresh and frozen embryos in in vitro fertilization in women infected by hepatitis B virus]. / Issues des transferts cumulés d'embryons frais et congelés en fécondation in vitro chez les femmes infectées par le virus de l'hépatite B.

OBJECTIVE: To assess the impact of hepatitis B virus (HBV) infection in women on in vitro fertilization (IVF) outcomes. METHODS: An observational monocentric case-control cohort study conducted between 2012 and 2019 compared the outcomes of the first cycle of IVF between 64 woman infected with HBV and 128 seronegative controls. Frozen embryos transfers made within 18 months of the puncture were included. The exclusion criteria were severe infections, viral co-infection in women, any viral infection in their spouse, or lack of fresh embryo transfer. The matching was performed according to age, primary infertility or secondary, conventional or intracytoplasmic injection IVF technique and date of attempt. The main analysis focused on cumulative live births rates (LBR). RESULTS: The clinical and ovarian stimulation characteristics were comparable except for a longer period of infertility in the HBV group. The LBR in the HBV group, when compared to controls, was not different after transfer of fresh (14.06 vs. 25.00% P=0.08) or frozen embryos (4.17 vs. 18.92% P=0.08), but significantly decreased in cumulative analysis (15.63 vs. 35.94% P=0.003). HBV infection was negatively associated with LBR in multivariate analysis OR=0.38 (95% CI 0.14-0.92) P<0.05. The implantation rate was lower in the HBV group versus controls, in fresh (14.89 vs. 27.72% P=0.02) and frozen (3.03 vs. 21.65% P=0.01) embryo transfers. CONCLUSION: This study suggests a negative impact of HBV infection in women on the cumulative LBR after IVF.

Distinguishing risk factors for the onset of cravings, withdrawal symptoms and tolerance in novice adolescent smokers.

AIM: While many studies report determinants of adolescent cigarette smoking, few identify risk factors for nicotine dependence (ND). This study distinguished between risk factors for three hallmarks of ND including cravings, withdrawal symptoms and tolerance. METHODS: A total of 319 novice smokers were followed every 3 months from first puff on a cigarette until the end of secondary school. Outcomes included time to first report of cravings, withdrawal symptoms and tolerance. RESULTS: Female sex, inhalation, smoking a whole cigarette, weekly smoking, daily smoking and alcohol use each independently increased the incidence of the onset of cravings. Inhalation, weekly smoking, daily smoking and alcohol use predicted the onset of withdrawal symptoms. Withdrawal symptoms, smoking a whole cigarette, monthly smoking, daily smoking and friends and siblings smoking increased the incidence of the onset of tolerance. None of parental education, impulsivity, novelty seeking, self-esteem, depression, stress, parental smoking, physical activity, or participation in sports teams was associated with the outcomes. CONCLUSION: The hallmarks of early ND are related to intensity and frequency of cigarette use. Avoidance of daily smoking may be particularly important in preventing the onset of ND symptoms and sustained smoking.

Conversion of a CHO cell culture process from perfusion to fed-batch technology without altering product quality.

During the development of a new drug product, it is a common strategy to develop a first-generation process with the aim to rapidly produce material for pre-clinical and early stage clinical trials. At a later stage of the development, a second-generation process is then introduced with the aim to supply late-stage clinical trials as well as market needs. This work was aimed at comparing the performance of two different CHO cell culture processes (perfusion and fed-batch) used for the production of a therapeutically active recombinant glycoprotein at industrial pilot-scale. The first-generation process was based on the Fibra-Cel packed-bed perfusion technology. It appeared during the development of the candidate drug that high therapeutic doses were required (>100mg per dose), and that future market demand would exceed 100 kg per year. This exceeded by far the production capacity of the first-generation process, and triggered a change of technology from a packed-bed perfusion process with limited scale-up capabilities to a fed-batch process with scale-up potential to typical bioreactor sizes of 15m(3) or more. The productivity per bioreactor unit volume (in product m(-3)year(-1)) of the fed-batch process was about 70% of the level reached with the first-generation perfusion process. However, since the packed-bed perfusion system was limited in scale (0.6m(3) maximum) compared to the volumes reached in suspension cultures (15m(3)), the fed-batch was selected as second-generation process. In fact, the overall process performance (in product year(-1)) was about 18-fold higher for the fed-batch compared to the perfusion mode. Data from perfusion and fed-batch harvests samples indicated that comparable product quality (relative abundance of monomers dimers and aggregates; N-glycan sialylation level; isoforms distribution) was obtained in both processes. To further confirm this observation, purification to homogeneity of the harvest material from both processes, followed by a complementary set of studies (e.g. full physico-chemical characterization, assessment of in vitro and in vivo bioactivity, comparative pharmacokinetics and pharmacodynamics studies in relevant species, etc.) would be required. Finally, this illustrates the need to fix the production process early during the development of a new drug product in order to minimize process conversion efforts and to shorten product development time lines.

Regulation of microglial development: a novel role for thyroid hormone.

The postnatal development of rat microglia is marked by an important increase in the number of microglial cells and the growth of their ramified processes. We studied the role of thyroid hormone in microglial development. The distribution and morphology of microglial cells stained with isolectin B4 or monoclonal antibody ED1 were analyzed in cortical and subcortical forebrain regions of developing rats rendered hypothyroid by prenatal and postnatal treatment with methyl-thiouracil. Microglial processes were markedly less abundant in hypothyroid pups than in age-matched normal animals, from postnatal day 4 up to the end of the third postnatal week of life. A delay in process extension and a decrease in the density of microglial cell bodies, as shown by cell counts in the developing cingulate cortex of normal and hypothyroid animals, were responsible for these differences. Conversely, neonatal rat hyperthyroidism, induced by daily injections of 3,5,3'-triiodothyronine (T3), accelerated the extension of microglial processes and increased the density of cortical microglial cell bodies above physiological levels during the first postnatal week of life. Reverse transcription-PCR and immunological analyses indicated that cultured cortical ameboid microglial cells expressed the alpha1 and beta1 isoforms of nuclear thyroid hormone receptors. Consistent with the trophic and morphogenetic effects of thyroid hormone observed in situ, T3 favored the survival of cultured purified microglial cells and the growth of their processes. These results demonstrate that thyroid hormone promotes the growth and morphological differentiation of microglia during development.

Acute hepatitis C infection in HIV positive men who have sex with men in Paris, France, 2001-2004.

In mid-2004, three Parisian hospital wards informed the Institut de veille sanitaire of recent acute hepatitis C in HIV-infected (HIV+) men who had sex with men (MSM). These cases for whom none of the usual bloodborne routes for hepatitis C (HCV) transmission was found, reported having had unprotected sex. In October 2004, we conducted a retrospective investigation in Parisian hospital wards to explore HCV modes of transmission in recent acute hepatitis C in HIV+ MSM. Patient demographics, clinical and biological status of HIV infection, reasons for HCV testing, sexual behaviour and risk factors for HCV transmission within the 6 months before hepatitis onset were collected from medical records. An anonymous self-administered questionnaire on sexual behaviour within the six months before hepatitis onset was also offered to all cases. We identified 29 cases of acute hepatitis C in HIV+ MSM with onset from April 2001 to October 2004. HIV infection was asymptomatic for 76%. Median age at hepatitis C onset was 40 (28-54) years. In all records, were noted unprotected anal sex, fisting in 21% and a concomitant sexually transmitted infection (STI) in 41%. Median time between HIV diagnosis and HCV infection was 6.5 years (0-22). From the 11 self-administered questionnaires completed, 10 reported an STI, 8 'hard' sexual practices, 6 bleeding during sex and 5 fisting. HCV transmission probably occurred through bleeding during unprotected traumatic anal sex among HIV+ MSM and may be facilitated by STI mucosal lesions. This report stresses the continuous need to strongly advocate safer sex to MSM.

Follicular fluid concentrations of lipids and their metabolites are associated with intraovarian gonadotropin-stimulated androgen production in women undergoing in vitro fertilization.

CONTEXT: Although growing evidence points toward a role of lipotoxicity in the development of hyperandrogenesis, the main feature of polycystic ovary syndrome, few studies directly assessed this association in vivo in humans, and none targeted the ovarian milieu. OBJECTIVE: The main objective of this study was to correlate follicular fluid (FF) T levels with lipids, lipid metabolites, and inflammation markers. DESIGN: This was a cross-sectional study. SETTING: Recruitment was performed in two fertility clinics at one private and one academic center. PARTICIPANTS: Eighty women requiring in vitro fertilization were recruited during one of their scheduled visit at the fertility clinic. All women aged between 18 and 40 years with a body mass index between 18 and 40 kg/m(2) were invited to participate. INTERVENTION(S): There were no interventions. MAIN OUTCOME MEASURE(S): At the time of oocyte aspiration, FF was collected and analyzed for total T, lipids [nonesterified fatty acids (NEFAs) plus triglycerides], NEFA metabolites (acylcarnitines; markers of ineffective NEFAs ß-oxidation), and inflammatory marker composition. The hypothesis being tested was formulated before the data collection. RESULTS: FF T levels were significantly correlated with FF levels of lipids (r = 0.381, P = .001; independently of IL-6), acylcarnitines (r ≥ 0.255, all P = .008; not independently of lipids), and IL-6 (r = 0.300, P = .009, independently of lipids). Additionally, FF lipid levels were significantly and strongly correlated with acylcarnitines (r ≥ 0.594; all P < .001). CONCLUSIONS: These results suggest that ovarian androgen production is related to intraovarian exposure to lipids, independently of inflammation and mainly through ineffective NEFA ß-oxidation (as shown by higher acylcarnitine levels). Inflammation is also associated with intraovarian androgenesis, independently of lipids.

Novel muscle chloride channel (CLCN1) mutations in myotonia congenita with various modes of inheritance including incomplete dominance and penetrance.

Autosomal-dominant and -recessive myotonia congenita are caused by mutations in the skeletal muscle voltage-gated chloride channel gene (CLCN1). We searched for mutations in this gene in 20 unrelated families with myotonia congenita. We identified 11 different mutations in 10 families. Two of five new mutations (Ala313Thr and Ile556Asn) were both autosomal recessive and dominant with either reduced penetrance or incomplete dominance. Mutations in the CLCN1 gene do not therefore necessarily behave in a classic Mendelian manner.

Intrathecal synthesis of IgG subclasses in multiple sclerosis and in acquired immunodeficiency syndrome (AIDS).

Serum and cerebrospinal fluid (CSF) of 50 neurological patients (24 multiple sclerosis (MS), ten acquired immunodeficiency syndrome (AIDS) and 16 other neurological diseases (OND)) and ten controls were analyzed by enzyme-linked immunosorbent assay (ELISA) for IgG subclass quantification and for the calculation of intrathecal synthesis (ITS). Total IgG was determined by two methods: electroimmunodiffusion (EID) and ELISA. A highly significant correlation was established between both methods. The existence of ITS was proved by the IgG/albumin ratio, the IgG index, Tourtellotte's formula, and Schuller's formula. In AIDS patients all IgG subclasses showed an increase in the CSF, whereas in sera only the IgG1 was significantly increased. CSF of MS patients showed a predominant increase of IgG1 whereas no significant modification of IgG subclasses was observed in sera. In most of the AIDS patients there was an ITS of IgG1, IgG3 and IgG4, but rarely (3/10) IgG2. In contrast, a polyclonal ITS of IgG was exceptional (1/24) in MS patients. No significant correlation could be established between clinical data and IgG subclass ITS in MS. The variations of each IgG subclass in serum and in ITS were not significantly correlated. Measurement of each IgG subclass and calculation of ITS seems essential in order to analyze any subclass antibody repertory inside the central nervous system.

Bronchodilatation with a metered-dose inhaler plus an extension, using tidal breathing vs jet nebulization.

The bronchodilating response to two doses of 100 micrograms of salbutamol introduced into an extension device (Aero-chamber [AC]), each followed by four tidal breaths, was compared with the nebulization of 2.5 mg of salbutamol in a saline solution during tidal breathing in a crossover study of ten patients with reversible airway obstruction. The forced expiratory volume in one second (FEV1) and the forced vital capacity 30 min after the drug administration improved significantly with both methods (p less than .001). The improvement of FEV1 with the AC (52.1 percent) compared with the nebulization (55.7 percent) was similar (p greater than .05).

High pentamidine levels associated with hypoglycemia and azotemia in a patient with Pneumocystis carinii pneumonia.

We report on a patient who presented with a Pneumocystis carinii pneumonia. Intravenous pentamidine (4 mg/kg/day) was given for 14 days without the occurrence of adverse effects. During this treatment, the mean (+/- SD) serum pentamidine trough concentration was 94 +/- 16 ng/ml. Three days later, the patient was admitted because of fever, and pentamidine (4 mg/kg/day) was again started. Fasting hypoglycemia and azotemia then occurred; the mean serum trough pentamidine level was 190 +/- 10 ng/ml during this week of treatment. We conclude that the occurrence of hypoglycemia and azotemia during pentamidine therapy may not be idiosyncrasic, but seemed associated in our patient with high levels of serum pentamidine.

Tolerability of treatments for viral hepatitis.

Interferon-alpha is the most widely used antiviral drug in chronic hepatitis B and C. Tolerability is usually good and serious adverse effects are rare. Most of the adverse effects are mild or transient and do not necessitate drug withdrawal. More than 90% of patients who are given interferon-alpha achieve 6 months to 1 year of treatment without serious adverse effects. The serious adverse effects usually occur in predisposed patients with pre-existing organ dysfunction. Nevertheless, careful selection of patients for therapy and observation during therapy are recommended. Nucleoside analogues are promising drugs in the treatment of chronic hepatitis B through inhibition of viral DNA polymerase. Lamivudine has been licensed for use in this indication. Its tolerability is excellent even when used for periods of 1 year or more. The main concern is the relatively high incidence of viral resistance resulting in breakthrough during or relapse after therapy. In the treatment of chronic hepatitis C, ribavirin, in combination with interferon-alpha is currently the reference therapy. The main adverse effect is haemolytic anaemia, which necessitates careful monitoring and adjustment of dosage in many cases. Recently, large trials showed the better efficacy of pegylated interferons as compared with standard interferon. The combination of pegylated interferon with ribavirin is under evaluation.

Clinical usefulness of high-pressure liquid chromatographic determination of serum pentamidine in AIDS patients.

We have developed a reproducible HPLC method to determine serum pentamidine, which demonstrates good chromatographic performance, and is sensitive enough to measure therapeutic doses. Pentamidine is first extracted from serum by passage through a C-18 extraction cartridge. Potential interfering substances are then removed by washing with 100% methanol. Pentamidine is eluted from the extraction cartridge with 1-heptanesulfonic acid. The extract is chromatographed on a highly deactivated column for basic compounds in the presence of minimal concentrations of 1-heptanesulfonic acid as the pairing agent. Detection is by fluorescence. The method can determine serum pentamidine levels in the range of 15-600 ng/mL free of interference from other drugs. In monitoring pentamidine levels in AIDS patients with Pneumocystis carinii, we found that trough serum levels over 100 ng/mL were associated with toxicity (hypoglycemia or azotemia) in 100% of patients. With levels under 100 ng/mL, signs of toxicity were observed in only 29% of the patients. We conclude that dose adjustment based on serum levels reduces the incidence of toxicity and enhances pentamidine therapy.

Fibronectin in plasma and CSF: evidence for its intrathecal synthesis.

A new methodology, using electroimmunodiffusion, has been described for the determination of fibronectin (Fn) in plasma and CSF. Fn was determined in the plasma of 35 normal subjects, and in the plasma and the CSF of 86 patients: 10 controls, 17 definite MS, 15 other inflammatory processes, 11 degenerative diseases, 13 peripheral neuropathies and 20 other neurological diseases. The normal mean was 354 +/- 53 micrograms/ml in plasma, not influenced by sex or age, and in the CSF 2.3 +/- 0.85 micrograms/ml. A significant increase of plasma Fn was observed in each of the patients-group. In the CSF an increase of Fn was observed predominantly in inflammatory processes but not in MS. An intrathecal synthesis of Fn was established in 24/76 patients, most often in inflammatory (9/15) processes but rarely (3/17) in MS. The intrathecal synthesis was suspected from the CSF Fn/CSF Albumin ratio and calculated using a formula derived from the principles previously described for IgG intrathecal synthesis. It represents the principal source of Fn increase in the CSF. In all inflammatory processes (including MS) the eventual influence of corticoids, immunosuppressive or anti-inflammatory drugs has not been established. The significance of the CSF Fn level in MS is discussed with regard to the recent demonstration of its presence within MS lesions and on macrophages in plaques.

Apolipoprotein E and multiple sclerosis: a biochemical and genetic investigation.

Apolipoprotein E (apo E) is postulated to be a major lipid carrier protein in the brain involved in brain development and repair. Multiple sclerosis (MS) is a major demyelinating disease characterized by destruction of myelin and marked alteration of myelin cholesterol and lipid metabolism. We have determined serum and cerebrospinal fluid (CSF) apo E concentrations using an original time-resolved immunofluorometric assay and calculated intrathecal apo E concentration. Apo E concentrations were determined in 13 control subjects and 129 neurological patients: 34 definite MS patients, 25 with Guillain-Barré syndrome (GBS), 32 with amyotrophic lateral sclerosis (ALS) and 38 with other neurological diseases. Seven clinical parameters (sex, age, age at MS onset, duration of the disease, course, clinical status and disability score) were considered in MS patients. Significant (P < 0.01) decrease in CSF apo E was observed in MS, linked to a decrease in intrathecal apo E. The decreased CSF apo E concentration in MS patients occur independent of the apo E genotype. Apo E is considered as a neurotrophic factor in the brain. Any decrease in intrathecal apo E synthesis would thus contribute to progression of neurological diseases, such as MS.

STD risk assessment and risk-reduction counseling by recently trained family physicians.

PURPOSE: To survey recently trained family physicians about their practices and perceptions regarding sexual history taking, drug-use history taking, and safe-sex counseling. METHOD: The study was conducted with all 1991 graduates from the four family practice residency programs in Quebec, Canada. Data were collected in 1992 by using a mailed questionnaire. RESULTS: A total of 148 (80%) of the 186 contacted physicians responded. The physicians reported taking a sexual history less frequently than a drug-use history when seeing patients for a general medical examination (42% versus 71%) or a first pregnancy visit (75% versus 91%). When taken, the sexual history was often too superficial to detect risk behaviors. Consequently, safe-sex counseling was infrequent. Over 20% of the physicians worried about patients' discomfort, were uneasy discussing sexual matters, and did not feel properly trained in sexual history taking. Graduates from the family practice residency at the school that offered training in human sexuality performed better sexual history taking than did graduates from the residencies at the other schools. CONCLUSIONS: Although infections from sexually transmitted diseases and the human immunodeficiency virus are important causes of morbidity and mortality, family physicians are still not actively involved in their prevention. The study suggests that medical education might be deficient in this area and that more training in human sexuality should be provided for family physicians.

Two-dimensional 1H, 15N NMR investigation of uniformly 15N-labeled lac repressor headpiece.

15N uniformly labeled lac repressor and lac repressor headpiece were prepared. 15N NMR spectra of lac repressor were shown resolution inadequate for detailed study while the data showed that the 15N labeled N-terminal part of the protein is quite suitable for this type of study allowing future investigation of the specific interaction of the lac repressor headpiece with the lac operator. We report here the total assignment of proton 1H and nitrogen 15NH backbone resonances of this headpiece in the free state. Assignments of the 15N resonances of the protein were obtained in a sequential manner using heteronuclear multiple quantum coherence (HMQC), relayed HMQC nuclear Overhauser and relayed HMQC-HOHAHA spectroscopy. More than 80 per cent of residues were assigned by their 15NH(i)-N1H(i + 1) and 15NH(i)-N1H(i - 1) connectivities. Values of the 3JNH alpha splitting for 39 of the 51 residues of the headpiece were extracted from HMQC and HMQC-J. The observed 15NH(i)-C beta H cross peaks and the 3JNH alpha coupling constants values are in agreement with the three alpha-helices previously described [Zuiderweg, E.R.P., Scheek, R.M., Boelens, R., van Gunsteren, W.F. and Kaptein, R., Biochimie 67, 707 (1985)]. The 3JNH alpha coupling constants can be now used for a more confident determination of the lac repressor headpiece. From these values it is shown that the geometry of the ends of the second and third alpha-helices exhibit deviation from the canonical alpha-helix structure. On the basis of NOEs and 3JNH alpha values, the geometry of the turn of the helix-turn-helix motif is discussed.