Patterns of selective serotonin reuptake inhibitor use and risk of falls and fractures in community-dwelling elderly people: the Three-City cohort.

In this population-based elderly cohort, participants using selective serotonin reuptake inhibitor (SSRI) antidepressants have an increased risk of falls and fractures notably when the treatment was continued over 4 years. Among the various SSRI types, citalopram only was at significant risk for falls and fluoxetine for fractures. INTRODUCTION: Increased risk of falls and fractures has been reported in elderly users of SSRIs. However, biases were insufficiently addressed notably temporality between exposure and outcome and confounding by residual depression. Our objective was to examine the associations between SSRIs and fall or fracture incidence focusing on their chronic use and different types of SSRIs. METHODS: The population-based cohort included participants aged 65 years and above, who had not fallen before inclusion (n = 6599) or were free of recent fracture (n = 6823) and were followed up twice over 4 years. New fall and fracture events were self-reported and defined as at least two falls and one fracture, respectively, during the previous 2 years. SSRI users were compared with those taking no antidepressants. Hazard ratios (HRs) were estimated using Cox models with delayed entry and adjusted for many confounders including residual depressive symptoms. RESULTS: Incidence of falls was 19.3 % over 4 years and that of fractures 9.5 %. After multi-adjustment, SSRI intake was significantly associated with a higher risk of falls (HR, 95 % CI = 1.58, 1.23-2.03) and fractures (HR, 95 % CI = 1.61, 1.16-2.24). The risks were significantly increased by 80 % in those continuing the treatment over 4 years. Citalopram intake only was at significant risk for falls and fluoxetine for fractures. CONCLUSIONS: In this large community-dwelling elderly sample, SSRI users were at higher risk of falls and fractures. This association was not due to reverse causality or residual depressive symptoms. Different SSRI drugs may have specific adverse effects on falls and fractures.

Impact of a premature menopause on cognitive function in later life.

OBJECTIVE: To determine whether premature menopause (≤40 years) can have long-lasting effects on later-life cognition and investigate whether this association varies depending on the type of menopause and use of hormone treatment (HT). DESIGN: Population-based cohort study. SETTING: The French Three-City Study. POPULATION: Four thousand eight hundred and sixty-eight women aged at least 65 years. METHODS: Multivariable-adjusted logistic regression models were used to determine the association between age at menopause, type of menopause (surgical, natural), and the use of menopausal HT and later-life cognitive function. MAIN OUTCOME MEASURES: Performance on a cognitive test battery (at baseline and over 7 years) and clinical dementia diagnosis. RESULTS: Menopause at or before the age of 40 years, both premature bilateral ovariectomy and premature ovarian failure (non-surgical loss of ovarian function), was associated with worse verbal fluency (OR 1.56, 95%CI 1.12-1.87, P=0.004) and visual memory (OR 1.39, 95%CI 1.09-1.77, P=0.007) in later life. HT at the time of premature menopause appeared beneficial for later-life visual memory but increased the risk of poor verbal fluency. Type of menopause was not significantly associated with cognitive function. Premature menopause was associated with a 30% increased risk of decline in psychomotor speed and global cognitive function over 7 years. CONCLUSION: Both premature surgical menopause and premature ovarian failure were associated with long-term negative effects on cognitive function, which are not entirely offset by menopausal HT. In terms of surgical menopause, these results suggest that the potential long-term effects on cognitive function should form part of the risk/benefit ratio when considering ovariectomy in younger women.

The association between caffeine and cognitive decline: examining alternative causal hypotheses.

BACKGROUND: Numerous studies suggest that higher coffee consumption may reduce the rate of aging-related cognitive decline in women. It is thus potentially a cheap and widely available candidate for prevention programs provided its mechanism may be adequately understood. The assumed effect is that of reduced amyloid deposition, however, alternative pathways notably by reducing depression and diabetes type 2 risk have not been considered. METHODS: A population study of 1,193 elderly persons examining depressive symptomatology, caffeine consumption, fasting glucose levels, type 2 diabetes onset, serum amyloid, and factors known to affect cognitive performance was used to explore alternative causal models. RESULTS: Higher caffeine consumption was found to be associated with decreased risk of incident diabetes in men (HR = 0.64; 95% CI 0.42-0.97) and increased risk in women (HR = 1.51; 95% CI 1.08-2.11). No association was found with incident depression. While in the total sample lower ratio Aß42/Aß40 levels (OR = 1.36, 95% CI 1.05-1.77, p = 0.02) were found in high caffeine consumers, this failed to reach significance when the analyses were stratified by gender. CONCLUSIONS: We found no evidence that reduced risk of cognitive decline in women with high caffeine consumption is moderated or confounded by diabetes or depression. The evidence of an association with plasma beta amyloid could not be clearly demonstrated. Insufficient proof of causal mechanisms currently precludes the recommendation of coffee consumption as a public health measure. Further research should focus on the high estrogen content of coffee as a plausible alternative explanation.

A prospective study of diurnal cortisol and cognitive function in community-dwelling elderly people.

BACKGROUND: Elevated cortisol levels due to hypothalamic-pituitary-adrenal (HPA) axis stress response have been associated with cognitive impairment. However, the causal relationship between stress and subsequent cognitive impairment remains unclear, notably because of the small number of gender-stratified prospective studies. METHOD: Salivary cortisol secretion was evaluated in 197 non-depressed community-dwelling elderly people at three time points on the day of hospital attendance for a clinical examination and again on the following day at home, in a distinct environmental context. Cognitive performance was evaluated at baseline and at 2- and 4-year follow-up. RESULTS: Cross-sectional logistic analyses adjusted for age and education indicated that men with high morning cortisol at the hospital had higher risk of low cognitive performance in verbal fluency [odds ratio (OR) 3.0, p=0.05] and visuospatial performance (OR 5.1, p=0.03). Impairment in verbal fluency was observed in women with moderate high morning cortisol (OR 3.6, p=0.05) or moderate slow diurnal rhythm (OR 3.7, p=0.04). In longitudinal analyses, slow diurnal rhythm (flatter slope) was associated with decline over 4 years in visuospatial performance (OR 7.7, p=0.03) and visual memory (OR 4.1, p=0.03) in men, and in verbal fluency (OR 6.0, p=0.01) in women. High morning cortisol was associated with decline in visual memory in women (OR 5.1, p=0.06). CONCLUSIONS: HPA axis dysregulation seems to be associated with low cognitive performance in the elderly. Slower cortisol elimination rates could predict cognitive decline affecting principally non-verbal functioning in men and verbal functioning in women. The effects are independent of environmental context, apolipoprotein E (ApoE) genotype or psychopathology. Interventions blocking this pathway may provide new therapeutic options to prevent cognitive decline.

Modelling complex pathways between late-life depression and disability: evidence for mediating and moderating factors.

Previous research has consistently shown an association between depression and disability in the elderly but little is known about the mechanisms linking the two. Recent longitudinal population studies have shown considerable inconsistency in the criteria used to establish causality and terms such as mediation and effect modification have been frequently applied incorrectly in terms of the inferences drawn. We underline the necessity to adopt more stringent theoretical criteria for the establishment of intermediary effects in the relationship between depression and disability to better identify cross-validated potential intervention points for reducing the risk of disablement and depression.

Risk profiles for mild cognitive impairment and progression to dementia are gender specific.

OBJECTIVE: To examine risk factors for mild cognitive impairment (MCI) and progression to dementia in a prospective community-based study of subjects aged 65 years and over. METHODS: 6892 participants who were over 65 and without dementia were recruited from a population-based cohort in three French cities. Cognitive performance, clinical diagnosis of dementia, and clinical and environmental risk factors were evaluated at baseline and 2-year and 4-year follow-ups. RESULTS: 42% of the population were classified as having MCI at baseline. After adjustment for confounding with logistic regression models, men and women classified as having MCI were more likely to have depressive symptomatology and to be taking anticholinergic drugs. Men were also more likely to have a higher body mass index, diabetes and stroke, whereas women were more likely to have poor subjective health, to be disabled, to be socially isolated, and to suffer from insomnia. The principal adjusted risk factors for men for progression from MCI to dementia in descending order were ApoE4 allele (OR = 3.2, 95% CI 1.7 to 5.7), stroke (OR = 2.8, 95% CI 1.2 to 6.9), low level of education (OR = 2.3, 95% CI 1.3 to 4.1), loss of Instrumental Activities of Daily Living (IADL) (OR = 2.2, 95% CI 1.1 to 4.5) and age (OR = 1.2, 95% CI 1.1 to 1.2). In women, progression is best predicted by IADL loss (OR = 3.5, 95% CI 2.1 to 5.9), ApoE4 allele (OR = 2.3, 95% CI 1.4 to 4.0), low level of education (OR = 2.2, 95% CI 1.3 to 3.6), subclinical depression (OR = 2.0, 95% CI 1.1 to 3.6), use of anticholinergic drugs (OR = 1.8, 95% CI 1.0 to 3.0) and age (OR = 1.1, 95% CI 1.1 to 1.2). CONCLUSIONS: Men and women have different risk profiles for both MCI and progression to dementia. Intervention programmes should focus principally on risk of stroke in men and depressive symptomatology and use of anticholinergic medication in women.

Frequent attendance and the concordance between PHQ screening and GP assessment in the detection of common mental disorders.

OBJECTIVE: Frequent Attenders (FAs) have high rates of both common mental disorders (CMD) and physical disorders, partly justifying this service use behaviour. This study examines both case and non-case concordance between CMDs as estimated by a self-report screening questionnaire and as rated by the general practitioner (GP), in FAs compared to Other Attenders (OAs). METHODS: 2275 patients of an overlapping sample of 55 GPs from 2 surveys performed 10 years apart, completed in the waiting room the Patient Health Questionnaire (PHQ) and Client Service Receipt Inventory on 6-month service use. For each patient, the GP rated mental health on a 0-4 scale, with a clear indication that scores of 2 and above referred to caseness. PHQ-CMDs included major and other depressive, anxiety, panic, and somatoform disorders, identified using the original PHQ DSM-IV criteria-based algorithms. FA was defined as the top 10% of attenders in age, sex and survey-year stratified subgroups. RESULTS: FAs had higher rates of PHQ-CMDs (42% versus 23% for OAs, p < .0001). They reported more personal and social problems, disability and had higher GP-rated physical illness. Survey-day antidepressant/anxiolytic medication prescription was higher for FAs (p < .0001), with (p = .02) but also without a CMD (p < .0001). Both GP/PHQ case and non-case concordance differed between FAs and OAs, with a non-case concordance odds ratio of 0.5 (95% CI: 0.3-0.7, p = .001) for FAs compared to OAs. CONCLUSION: Despite a greater likelihood of GPs detecting CMDs in FAs, our findings suggest a potential risk of 'over-detection' of patients not reaching CMD threshold criteria among FAs.

Chronic and remitting trajectories of depressive symptoms in the elderly. Characterisation and risk factors.

BACKGROUND: In elderly general population sub-syndromal clinically significant levels of depressive symptoms are highly prevalent and associated with high co-morbidity and increased mortality risk. However changes in depressive symptoms over time and etiologic factors have been difficult to characterise notably due to methodological shortcomings. Our objective was to differentiate trajectories of depressive symptoms over 10 years in community-dwelling elderly men and women using statistical modelling methods which take into account intra-subject correlation and individual differences as well as to examine current and life-time risk factors associated with different trajectories. METHODS: Participants aged 65 and over were administered standardised questionnaires and underwent clinical examinations at baseline and after 2, 4, 7 and 10 years. Trajectories over time of the Center for Epidemiologic Studies Depression scores were modelled in 517 men and 736 women separately with latent class mixed models which include both a linear mixed model to describe latent classes of trajectories and a multinomial logistic model to characterise the latent trajectories according to baseline covariates (socio-demographic, lifestyle, clinical, genetic characteristics and stressful life events). RESULTS: In both genders two different profiles of symptom changes were observed over the 10-year follow-up. For 9.1% of men and 25% of women a high depressive symptom trajectory was found with a trend toward worsening in men. The majority of the remaining men and women showed decreasing symptomatology over time, falling from clinically significant to very low levels of depressive symptoms. In large multivariate class membership models, mobility limitations [odds ratio (OR) = 4.5, 95% confidence interval (CI) 1.6-12.9 and OR = 4.9, 95% CI 2.3-10.7, in men and women respectively], ischemic pathologies (OR = 2.9, 95% CI 1.0-8.3 and OR = 3.1, 95% CI 1.0-9.9), and recent stressful events (OR = 4.5, 95% CI 1.1-18.5, OR = 3.2, 95% CI 1.6-6.2) were associated with a poor symptom course in both gender as well as diabetes in men (OR = 3.5, 95% CI 1.1-10.9) and childhood traumatic experiences in women (OR = 3.1, 95% CI 1.6-5.8). CONCLUSIONS: This prospective study was able to differentiate patterns of chronic and remitting depressive symptoms in elderly people with distinct symptom courses and risk factors for men and women. These findings may inform prevention programmes designed to reduce the chronic course of depressive symptomatology.

Biological and psychological predictors of posttraumatic stress disorder onset and chronicity. A one-year prospective study.

BACKGROUND: Few studies have prospectively examined risk factors for posttraumatic stress disorder (PTSD) in the aftermath of a traumatic exposure. The aim of this study is to identify the concurrent influence of psychological and biological diatheses on PTSD onset and maintenance, taking into account socio-demographic factors and psychiatric antecedents. METHODS: A total of 123 civilians (61.8% of women) recruited in emergency units, were assessed using validated instruments during the first week and then at 1, 4, and 12 months post-trauma. Baseline assessment included evaluation of the psychological diathesis (i.e. psychiatric history and peritraumatic distress and dissociation), and the biological diathesis [i.e. cortisol, norepinephrine, epinephrine, c-reactive protein, total cholesterol, HDL cholesterol, glycosylated haemoglobin, waist-to-hip ratio (WHR), body mass index, diastolic and systolic blood pressure (SBP), and heart rate]. RESULTS: Multivariate logistic regression analyses demonstrated both psychological and biological diatheses to be independent risk factors for PTSD. Peritraumatic distress and dissociation predicted onset (1-month) and mid-term PTSD (4-months), respectively. PTSD risk was associated positively with SBP and negatively with WHR, throughout the follow-up. In addition, a higher level of 12 h-overnight urinary norepinephrine independently predicted mid-term PTSD (4-months). CONCLUSIONS: This prospective study shows that peritraumatic psychological and biological markers are independent predictors of PTSD onset with specificities according to the stage of PTSD development; the psychological diathesis, i.e. peritraumatic distress and dissociation, being a better predictor of short-term dysfunction whereas biological diathesis was also predictive of development and maintenance of PTSD.

Regulation of phosphatidylcholine biosynthesis in Plasmodium-infected erythrocytes.

Plasmodium knowlesi-infected erythrocytes efficiently incorporated choline and metabolize it into phosphatidylcholine via the de novo Kennedy pathway. No formation of either betaine or acetylcholine was detected. At physiological concentrations of external choline, isotopic equilibrium between intracellular choline and phosphocholine was reached in less than 1 h, whereas labeled phosphatidylcholine accumulated constantly, until at least 210 min. During this time, intracellular CDP-choline remained quite low compared to phosphocholine, which suggests that choline-phosphate cytidylyltransferase (EC 2.7.7.15) is the rate-limiting step of the Kennedy pathway. However, this activity was probably not saturated in situ by phosphocholine, since the external choline concentration, up to 100 microM, can regulate phosphatidylcholine biosynthesis via the level of intracellular phosphocholine. This was corroborated by the respective velocities and affinity characteristics of the three enzymatic steps involved in the Kennedy pathway. These results, together with the localization of both choline metabolites and enzyme activities, provide a precise scheme of the dynamics of de novo phosphatidylcholine biosynthesis. Concerning the alternative pathway for phosphatidylcholine biosynthesis via the methylation of phosphatidylethanolamine, we show that an increase in de novo phosphatidylcholine biosynthesis could instigate a concomitant decrease in the steps of phosphatidylethanolamine methylation, indicating that the parasite is able to modulate its phosphatidylcholine biosyntheses.

Choline kinase activity in Plasmodium-infected erythrocytes: characterization and utilization as a parasite-specific marker in malarial fractionation studies.

Choline kinase (EC 2.7.1.32) was investigated in plasmodium falciparum-infected erythrocytes. Disrupted infected erythrocytes had a choline kinase activity of 1.9 +/- 0.2 nmol phosphorylcholine/10(7) infected cells per h, whereas the activity in normal uninfected erythrocytes was less than 6 pmol/10(7) cells per h. A broad alkaline optimal pH (7.9-9.2) was observed. The Km values for choline and ATP were 79 +/- 20 microM, and 1.3 +/- 0.3 mM, respectively. ATP concentrations higher than 12 mM inhibited choline kinase. Maximal activity was registered with a Mg2+ concentration of 10 mM, whereas its replacement by Mn2+, or other divalent cations, involved a decrease in choline kinase activity of at least 75%. Inhibition by products of the reaction, such as phosphorylcholine and ADP was investigated. In plasmodium knowlesi-infected erythrocytes, choline kinase had similar properties, but with a much higher specific activity of 16.4 +/- 2.1 nmol/10(7) infected cells per h. Subcellular fractionation of P. knowlesi-infected erythrocyte suspensions revealed that choline kinase was located exclusively in the cytosol of the parasite. We show that this enzyme is a useful index of parasite cytosolic content leakage, when infected erythrocytes are fractionated by saponin lysis or nitrogen decompression.

Selective elimination of malaria infected erythrocytes by a modified phospholipase A2 in vitro.

Pig pancreatic phospholipase A2 does not act on normal erythrocytes, but the membrane penetrating capacity is enhanced by the covalent attachment of one fatty acyl chain to Lys-116 of the enzyme. Taking advantage of the impaired packing of phospholipids in the membrane of Plasmodium infected erythrocytes it was demonstrated that a lauric acid derivative of phospholipase A2 is capable of exclusively attaching the infected erythrocytes in vitro, leaving the uninfected cells undisturbed. The chemically modified phospholipase A2 appeared to cause death of the parasite in cell cultures of infected erythrocytes.

Lifelong endocrine fluctuations and related cognitive disorders.

The aim of this review is to examine the relationship between endocrine fluctuation and cognitive functioning. A plethora of in vitro and in vivo studies has demonstrated the neuroprotective role of estrogens and their impact on the neurotransmitter systems implicated in cognition. Recent hormonal replacement therapy (HRT) trials in non-demented post-menopausal women suggest a temporary positive effect (notably on verbal memory), and four recent meta-analyses converge to suggest a possible protective effect in relation to Alzheimer's disease (reducing risk by 29 to 44%). However, data from the only large randomised controlled trial published to date, the Women's Health Initiative Memory Study, did not confirm these observations and have even suggested an increase in dementia risk for women using HRT compared to controls. Several methodological differences between observation studies and controlled trials with regard to patient group, type, timing and duration of HRT, cognitive measures and analyses, are discussed to explain these discrepancies. The association between hormonal serum level and cognitive functioning remains controversial suggesting high inter-individual vulnerability in risk. Moreover, research on the impact of endocrine functioning on cognition during the female reproductive cycle suggests life-long fluctuations in vulnerability. Etiological models taking into account the interaction of clinical, reproductive, and menstrual events throughout life may provide a more valid approach in understanding the effects of steroids on the brain and in determining sub-groups at heightened risk. Cognitive disorders in the elderly are more likely be related to cumulated lifelong exposure to steroids, rather than to a specific exposure to a given steroid. Multifactorial models based on an exhaustive view of all hormonal events throughout reproductive life together with other risk factors (notably genetic risk factors related to estrogen receptor polymorphisms) should be explored to clarify the role of hormonal risk factors, or protective factors for cognitive dysfunction and dementia.

BDNF promoter methylation and genetic variation in late-life depression.

The regulation of the brain-derived neurotrophic factor (BDNF) is important for depression pathophysiology and epigenetic regulation of the BDNF gene may be involved. This study investigated whether BDNF methylation is a marker of depression. One thousand and twenty-four participants were recruited as part of a longitudinal study of psychiatric disorders in general population elderly (age ⩾ 65). Clinical levels of depression were assessed using the Mini International Neuropsychiatric Interview for the diagnosis of major depressive disorder according to the Diagnostic and Statistical Manual of Mental Disorder IV criteria, and the Centre for Epidemiologic Studies Depression Scale (CES-D) for assessment of moderate to severe depressive symptoms. Buccal DNA methylation at the two most widely studied BDNF promoters, I and IV, was investigated using the Sequenom MassARRAY platform that allows high-throughput investigation of methylation at individual CpG sites within defined genomic regions. In multivariate linear regression analyses adjusted for a range of participant characteristics including antidepressant use, depression at baseline, as well as chronic late-life depression over the 12-year follow-up, were associated with overall higher BDNF methylation levels, with two sites showing significant associations (promoter I, Δ mean = 0.4%, P = 0.0002; promoter IV, Δ mean = 5.4%, P = 0.021). Three single-nucleotide polymorphisms (rs6265, rs7103411 and rs908867) were also found to modify the association between depression and promoter I methylation. As one of the largest epigenetic studies of depression, and the first investigating BDNF methylation in buccal tissue, our findings highlight the potential for buccal BDNF methylation to be a biomarker of depression.

Risk factors for late-onset generalized anxiety disorder: results from a 12-year prospective cohort (the ESPRIT study).

Generalized anxiety disorder (GAD) is a chronic and highly prevalent disorder associated with increased disability and mortality in the elderly. Treatment is difficult with low rate of full remission, thus highlighting the need to identify early predictors for prevention in elderly people. The aim of this study is to identify and characterize incident GAD predictors in elderly people. A total of 1711 individuals aged 65 years and above and free of GAD at baseline were randomly recruited from electoral rolls between 1999 and 2001 (the prospective ESPRIT study). The participants were examined at baseline and five times over 12 years. GAD and psychiatric comorbidity were diagnosed with a standardized psychiatric examination, the Mini-International Neuropsychiatry Interview on the basis of DSM-IV (Diagnostic and Statistical Manual of Mental Disorders, fourth edition) criteria and validated by a clinical panel. During the follow-up, 8.4% (95% confidence interval=7.1-9.7%) of the participants experienced incident GAD, 80% being first episodes; the incident rate being 10 per 1000 person-years. The principal predictors of late-onset incident GAD over 12 years derived from a multivariate Cox model were being female, recent adverse life events, having chronic physical (respiratory disorders, arrhythmia and heart failure, dyslipidemia, cognitive impairment) and mental (depression, phobia and past GAD) health disorders. Poverty, parental loss or separation and low affective support during childhood, as well as history of mental problems in parents were also significantly and independently associated with incident GAD. GAD appears as a multifactorial stress-related affective disorder resulting from both proximal and distal risk factors, some of them being potentially modifiable by health care intervention.

C-reactive protein gene variants: independent association with late-life depression and circulating protein levels.

C-reactive protein (CRP) is a heritable biomarker of systemic inflammation that is commonly elevated in depressed patients. Variants in the CRP gene that influence protein levels could thus be associated with depression but this has seldom been examined, especially in the elderly. Depression was assessed in 990 people aged at least 65 years as part of the ESPRIT study. A clinical level of depression (DEP) was defined as having a score of ⩾16 on The Center for Epidemiologic Studies Depression scale or a diagnosis of current major depression based on the Mini-International Neuropsychiatric Interview and according to Diagnostic and Statistical Manual of Mental Disorders-IV criteria. Five single-nucleotide polymorphisms spanning the CRP gene were genotyped, and circulating levels of high-sensitivity CRP were determined. Multivariable analyses adjusted for socio-demographic characteristics, smoking, ischemic pathologies, cognitive impairment and inflammation-related chronic pathologies. The minor alleles of rs1130864 and rs1417938 were associated with a decreased risk of depression in women at Bonferroni-corrected significance levels (P=0.002). CRP gene variants were associated with serum levels in a gender-specific manner, but only rs1205 was found to be nominally associated with both an increased risk of DEP and lower circulating CRP levels in women. Variants of the CRP gene thus influence circulating CRP levels and appear as independent susceptibility factors for late-life depression.

Several lines of evidence demonstrating that Plasmodium falciparum, a parasitic organism, has distinct enzymes for the phosphorylation of choline and ethanolamine.

In Plasmodium falciparum-infected erythrocyte homogenates, the specific activity of ethanolamine kinase (7.6 +/- 1.4 nmol phosphoethanolamine/10(7) infected cells per h) was higher than choline kinase specific activity (1.9 +/- 0.2 nmol phosphocholine/10(7) infected cells per h). The Km of choline kinase for choline was 79 +/- 20 microM, and ethanolamine was a weak competitive inhibitor of the reaction (Ki = 92 mM). Ethanolamine kinase had a Km for ethanolamine of 188 +/- 19 microM, and choline was a competitive inhibitor of ethanolamine kinase with a very high Ki of 268 mM. Hemicholinium 3 inhibited choline kinase activity, but had no effect on ethanolamine kinase activity. In contrast, D-2-amino-1-butanol selectively inhibited ethanolamine kinase activity. Furthermore, when the two enzymes were subjected to heat inactivation, 85% of the choline kinase activity was destroyed after 5 min at 50 degrees C, whereas ethanolamine kinase activity was not altered. Our results indicate that the phosphorylation of choline and ethanolamine was catalyzed by two distinct enzymes. The presence of a de novo phosphatidylethanolamine Kennedy pathway in P. falciparum contributes to the bewildering variety of phospholipid biosynthetic pathways in this parasitic organism.

Phospholipid uptake by Plasmodium knowlesi infected erythrocytes.

The uptake of phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylserine (PS) in Plasmodium knowlesi infected erythrocytes has been studied. Whereas uptake of phospholipids, in the absence of phospholipid transfer proteins, is negligible in control cells, the infected cells can incorporate considerable amounts of added phospholipids. The uptake is enhanced by the presence of lipid transfer proteins. Doubly labeled [3H]oleate, [14C]choline) PC does not undergo any appreciable remodelling following uptake, which strongly suggests that plasma PC is used as such for the biogenesis of the parasite membranes. Transport of extracellularly offered PS and PE towards the intraerythrocytic parasite and utilization of these lipids by the parasite are confirmed by the observation that these lipids are converted into respectively PE and PC. The extent and rate of these conversions depend on the way the phospholipids are introduced into the infected cells.

Characterization of the potent in vitro and in vivo antimalarial activities of ionophore compounds.

Large-scale in vitro screening of different types of ionophores previously pinpointed nine compounds that were very active and selective in vitro against Plasmodium falciparum; their in vitro and in vivo antimalarial effects were further studied. Addition of the ionophores to synchronized P. falciparum suspensions revealed that all P. falciparum stages were sensitive to the drugs. However, the schizont stages were three- to ninefold more sensitive, and 12 h was required for complete parasite clearance. Pretreatment of healthy erythrocytes with toxic doses of ionophores for 24 to 48 h showed that the activity was not due to an irreversible effect on the host erythrocyte. No preferential ionophore adsorption in infected or uninfected erythrocytes occurred. On the other hand, ionophore molecules strongly bound to serum proteins since increasing the serum concentration from 2 to 50% led to almost a 25-fold parallel increase in the ionophore 50% inhibitory concentration. Mice infected with the malaria parasites Plasmodium vinckei petteri or Plasmodium chabaudi were successfully treated with eight ionophores in a 4-day suppressive test. The 50% effective dose after intraperitoneal administration ranged from 0.4 to 4.1 mg/kg of body weight, and the therapeutic indices were about 5 for all ionophores except monensin A methyl ether, 5-bromo lasalocid A, and gramicidin D, whose therapeutic indices were 12, 18, and 344, respectively. These three compounds were found to be curative, with no recrudescence. Gramicidin D, which presented impressive antimalarial activity, requires parenteral administration, while 5-bromo lasalocid A has the major advantage of being active after oral administration. Overall, the acceptable levels of toxicity and the good in vivo therapeutic indices in the rodent model highlight the interesting potential of these ionophores for the treatment of malaria in higher animals.