Meeting the needs of adolescent post-abortion care patients in the Dominican Republic.

Counselling on contraception and contraceptive method provision are key components of post-abortion care (PAC). Some studies have suggested that adolescent PAC patients receive worse care than older women seeking these services. This study aimed to evaluate an intervention whose goal was to improve the counselling and contraceptive uptake of PAC patients, with special attention given to the needs of adolescent patients, in the four public hospitals in the Dominican Republic where PAC services were not being routinely offered. The counselling intervention effort included provider training and the development of adolescent-friendly information, education and communication (IEC) materials. Eighty-eight providers were interviewed at baseline and 6 months after the intervention was implemented. Six months after providers were trained, 140 adolescent PAC patients (< or = 19 years of age) and 134 older PAC patients (20-35 years) were interviewed about the contraceptive counselling messages and contraceptive methods they received before they were discharged from hospital. The adolescent and older PAC patients were matched on study hospital and time of arrival. Significant improvements were noted in provider knowledge and attitudes. No changes were noted in provider-reported PAC counselling behaviours, with close to 70% of providers reporting they routinely assess patients' fertility intentions, discuss contraception, assess STI/HIV risk and discuss post-abortion complications. Adolescent and older PAC patients reported receiving PAC counselling messages at similar rates. Forty per cent of adolescent PAC patients and 45% of older PAC patients who wanted to delay pregnancy were discharged with a contraceptive method. Adolescents were more likely to receive an injectable contraceptive method whereas older women were discharged with a variety of methods. The PAC counselling intervention increased provider knowledge and improved their attitudes and benefited both adolescent and older patients.

Equilibrium unfolding of the retinoid X receptor ligand binding domain and characterization of an unfolding intermediate.

The retinoid X receptor (RXR) is a ligand-activated transcription factor that plays an important role in growth and development and the maintenance of cellular homeostasis. A thermodynamic ultraviolet circular dichroism, tryptophan fluorescence and ligand binding activity with guanidine as a chemical denaturant are consistent with a two step mechanism. The dimeric LBD equilibrates with a monomeric intermediate (DeltaG(0)(H(2)O) equal to 8.3 kcal/mol) that is in equilibrium with the unfolded state (DeltaG(0)(H(2)O) equal to 2.8 kcal/mol). The intermediate was characterized by analytical ultracentrifugation, spectroscopy, and collisional fluorescence quenching, which imply that the monomeric intermediate maintains a high degree, but not all, of native secondary structure. Although intrinsic fluorescence from native and intermediate suggests little change in tryptophan environments, fluorescence intensities from fluorescein reporter groups differ significantly between the two structures. Analysis of the collisional quenching results imply that the intermediate is characterized by tryptophans with increased accessibility to small solutes and less overall compactness than the native protein.

Structural characterization of zinc-deficient human superoxide dismutase and implications for ALS.

Over 130 mutations to copper, zinc superoxide dismutase (SOD) are implicated in the selective death of motor neurons found in 25% of patients with familial amyotrophic lateral sclerosis (ALS). Despite their widespread distribution, ALS mutations appear positioned to cause structural and misfolding defects. Such defects decrease SOD's affinity for zinc, and loss of zinc from SOD is sufficient to induce apoptosis in motor neurons in vitro. To examine the importance of the zinc site in the structure and pathogenesis of human SOD, we determined the 2.0-A-resolution crystal structure of a designed zinc-deficient human SOD, in which two zinc-binding ligands have been mutated to hydrogen-bonding serine residues. This structure revealed a 9 degrees twist of the subunits, which opens the SOD dimer interface and represents the largest intersubunit rotational shift observed for a human SOD variant. Furthermore, the electrostatic loop and zinc-binding subloop were partly disordered, the catalytically important Arg143 was rotated away from the active site, and the normally rigid intramolecular Cys57-Cys146 disulfide bridge assumed two conformations. Together, these changes allow small molecules greater access to the catalytic copper, consistent with the observed increased redox activity of zinc-deficient SOD. Moreover, the dimer interface is weakened and the Cys57-Cys146 disulfide is more labile, as demonstrated by the increased aggregation of zinc-deficient SOD in the presence of a thiol reductant. However, equimolar Cu,Zn SOD rapidly forms heterodimers with zinc-deficient SOD (t1/2 approximately 15 min) and prevents aggregation. The stabilization of zinc-deficient SOD as a heterodimer with Cu,Zn SOD may contribute to the dominant inheritance of ALS mutations. These results have general implications for the importance of framework stability on normal metalloenzyme function and specific implications for the role of zinc ion in the fatal neuropathology associated with SOD mutations.

Calmodulin potentiates G beta gamma activation of phospholipase C-beta3.

Phospholipase C-beta (PLC-beta) isozymes (EC 3.1.4.11) hydrolyze the membrane phospholipid phosphatidylinositol-4,5-bisphosphate to generate intracellular second messenger signaling molecules inositol-1,4,5-trisphosphate (IP3) and diacylglycerol (DAG) in response to receptor activation and other cellular stimuli. PLCbeta1 and PLCbeta3 isozymes were previously demonstrated to bind the calcium-sensitive molecule calmodulin [McCullar JS, Larsen SA, Millimaki RA, Filtz TM. Calmodulin is a phospholipase C-{beta} interacting protein. J Biol Chem 2003;278(36):33708-13]. We have now shown through fluorescence anisotropy that calmodulin/PLCbeta3 affinities increase with increasing calcium in a physiologically relevant concentration range. The bimolecular affinity constants for calmodulin interaction with PLCbeta1 or PLCbeta3 were estimated as 260 and 200 nM, respectively, from fluorescence anisotropy data. There was no effect of calmodulin on basal or G alpha q-stimulated catalytic activity for either isozyme. However, the interaction between calmodulin and PLCbeta3 leads to potentiation of activation by the G-protein beta gamma dimer in an in vitro assay. 1321N1 cells treated with calmodulin inhibitors concurrent with and post-stimulation of muscarinic receptors significantly reduced [3H]PIP hydrolysis. Together these data are suggestive of cooperative role for calmodulin in the G-protein beta gamma dimer-stimulated activity of PLCbeta3.

Misoprostol and declining abortion-related morbidity in Santo Domingo, Dominican Republic: a temporal association.

OBJECTIVE: To validate anecdotal reports that abortion-related complications decreased in the Dominican Republic after the introduction of misoprostol into the country. DESIGN: Retrospective records reviews and cross-sectional surveys, interviews and focus groups. SETTING: Family planning clinics, pharmacies, door-to-door canvassing and a tertiary care maternity hospital in Santo Domingo, Dominican Republic. POPULATION: Women of reproductive age in Santo Domingo, Dominican Republic. METHODS: Qualitative and quantitative methods were used. Individual interviews and focus groups of reproductive health professionals, non-governmental organisation leaders and women's group leaders (n= 50) were conducted to discover the role of misoprostol in the Dominican Republic. Local women (n= 157) were surveyed to determine their knowledge of misoprostol as an abortifacient and mystery client visits were made to 80 pharmacies in order to purchase misoprostol without a prescription. Sales data were obtained that documented when misoprostol was introduced to the Dominican Republic pharmacies. Hospital admissions for abortions from the prior eight years were reviewed and hospital emergency room consultation ledgers of 31,190 visits for the period 1994-2001 were reviewed for abortion complications. MAIN OUTCOME MEASURES: Frequencies of maternal morbidities and knowledge of misoprostol. RESULTS: Mystery clients purchased misoprostol without a prescription in nearly 64% of pharmacies; staff provided little additional information or counselling. Reliable sales data documented the introduction of misoprostol in 1986. Abortion complications decreased from 11.7% of abortions in 1986 to 1.7% in 2001. The majority of professionals interviewed felt that knowledge of these findings should be made public. CONCLUSIONS: The data were of too poor quality to validate the verbal reports reliably, but misoprostol appears to have been widely used over a period when abortion-related morbidity fell. It remains plausible that the use of misoprostol contributed to the reduction.

Effects of high-pressure processing at low temperature on the molecular structure and surface properties of beta-lactoglobulin.

High-pressure processing (HPP) was utilized to induce unfolding of beta-lactoglobulin (beta-LG). beta-Lactoglobulin solutions at concentrations of 0.5 mg/mL, in pH 7.5 phosphate buffer, were pressure treated at 510 MPa for 10 min at either 8 or 24 degrees C. The secondary structure, as determined by circular dichroism (CD), of beta-LG processed at 8 degrees C appeared to be unchanged, whereas beta-LG processed at 24 degrees C lost alpha-helix structure. Tertiary structures for beta-LG, as determined by near-UV CD, intrinsic protein fluorescence spectroscopy, hydrophobic fluorescent probe binding, and thiol group reactivity, were changed following processing at either temperature. The largest changes to tertiary structure were observed for the samples processed at 24 degrees C. Model solutions containing the pressure-treated beta-LG showed significant decreases in surface tension at liquid-air interfaces with values of 54.00 and 51.69 mN/m for the samples treated at 24 and 8 degrees C, respectively. In comparison, the surface tension for model solutions containing the untreated control was 60.60 mN/m. Changes in protein structure during frozen and freeze-dried storage were also monitored, and some renaturation was observed for both storage conditions. Significantly, the sample pressure-treated at 8 degrees C continued to display the lowest surface tension.

Interleukin-12 p40 secretion by cutaneous CD11c+ and F4/80+ cells is a major feature of the innate immune response in mice that develop Th1-mediated protective immunity to Schistosoma mansoni.

Radiation-attenuated (RA) schistosome larvae are potent stimulators of innate immune responses at the skin site of exposure (pinna) that are likely to be important factors in the development of Th1-mediated protective immunity. In addition to causing an influx of neutrophils, macrophages, and dendritic cells (DCs) into the dermis, RA larvae induced a cascade of chemokine and cytokine secretion following in vitro culture of pinna biopsy samples. While macrophage inflammatory protein 1alpha and interleukin-1beta (IL-1beta) were produced transiently within the first few days, the Th1-promoting cytokines IL-12 and IL-18 were secreted at high levels until at least day 14. Assay of C3H/HeJ mice confirmed that IL-12 secretion was not due to lipopolysaccharide contaminants binding Toll-like receptor 4. Significantly, IL-12 p40 secretion was sustained in pinnae from vaccinated mice but not in those from nonprotected infected mice. In contrast, IL-10 was produced from both vaccinated and infected mice. This cytokine regulates IL-12-associated dermal inflammation, since in vaccinated IL-10(-/-) mice, pinna thickness was greatly increased concurrent with elevated levels of IL-12 p40. A significant number of IL-12 p40(+) cells were detected as emigrants from in vitro-cultured pinnae, and most were within a population of rare large granular cells that were Ia(+), consistent with their being antigen-presenting cells. Labeling of IL-12(+) cells for CD11c, CD205, CD8alpha, CD11b, and F4/80 indicated that the majority were myeloid DCs, although a proportion were CD11c(-) F4/80(+), suggesting that macrophages were an additional source of IL-12 in the skin.