Multiple reproduction forms in the polyps of the cannonball jellyfish Stomolophus sp. 2: Probable life-cycle reversal.

The jellyfish genera Stomolophus spp. is one of the most abundant in the Pacific Ocean, yet it has not been thoroughly studied. Until recently, research has been developed and directed to its knowledge because of the economic interest in its exploitation. The genus Stomolophus in the Pacific Ocean is composed of five species (S. agaricus, S. chunii, S. collaris, S. fritillaria, and S. meleagris), and Stomolophus sp. 2 has been recently reported in the central part of the Gulf of California. Therefore, this study aimed to describe in vivo the different developmental stages of Stomolophus sp. 2 life cycle. As a result, multiple polyp reproduction forms were described, such as polyp-stolon formation, polydisc strobilation with more than 20 ephyrae formed by each strobila, and polyp formation directly from juvenile ephyra. In the degenerating phase, the polyps turned into cysts induced by stress conditions, such as changes in temperature, oxygen, and food availability. The life cycle of Stomolophus sp. 2 can be distinguished from that of S. meleagris by showing various asexual reproduction mechanisms and polydisc-like strobilation. The formation of polyps directly from the ectoderm of degenerating juvenile medusae suggests the possibility of a reversion cycle. Because of the different life cycles between S. meleagris and S. sp. 2, in addition to their morphological and genetic differences, this study proposes that Stomolophus sp. 2 should be considered a new species and suggests the name Stomolophus yaquilli, in reference to the indigenous community that lives in the species distribution area.

Effects of Yam (Dioscorea rotundata) Mucilage on the Physical, Rheological and Stability Characteristics of Ice Cream.

In the present investigation, yam mucilage was evaluated as a stabilizer and emulsifier in the formulation of vanilla flavored ice cream; physicochemical, rheological, and stability characteristics were determined. A completely randomized bifactorial design was used (yam mucilage: Carboxymethylcellulose ratio with the following levels: 100:0, 80:20, 50:50, and 20:80, and stabilizers concentration with levels of 0.4 and 0.8%). Results showed an increase in the protein content present in ice cream mixture as the amount of mucilage increases. Rheologically, it was found that ice cream has the characteristic behavior of a pseudoplastic fluid, presenting a viscoelastic structure where elastic behavior predominates. In addition, ratios with a higher content of mucilage incorporated a greater volume of air and presented the longest melting times, delaying drops falling time; in the same way mucilage gives ice cream a freezing temperature between -6.1 to -2.8 °C, indicating that the application of mucilage in food industry is possible due to its nutritional value, and it gives ice cream stability properties.

TRPA1 ankyrin repeat six interacts with a small molecule inhibitor chemotype.

TRPA1, a member of the transient receptor potential channel (TRP) family, is genetically linked to pain in humans, and small molecule inhibitors are efficacious in preclinical animal models of inflammatory pain. These findings have driven significant interest in development of selective TRPA1 inhibitors as potential analgesics. The majority of TRPA1 inhibitors characterized to date have been reported to interact with the S5 transmembrane helices forming part of the pore region of the channel. However, the development of many of these inhibitors as clinical drug candidates has been prevented by high lipophilicity, low solubility, and poor pharmacokinetic profiles. Identification of alternate compound interacting sites on TRPA1 provides the opportunity to develop structurally distinct modulators with novel structure-activity relationships and more desirable physiochemical properties. In this paper, we have identified a previously undescribed potent and selective small molecule thiadiazole structural class of TRPA1 inhibitor. Using species ortholog chimeric and mutagenesis strategies, we narrowed down the site of interaction to ankyrinR #6 within the distal N-terminal region of TRPA1. To identify the individual amino acid residues involved, we generated a computational model of the ankyrinR domain. This model was used predictively to identify three critical amino acids in human TRPA1, G238, N249, and K270, which were confirmed by mutagenesis to account for compound activity. These findings establish a small molecule interaction region on TRPA1, expanding potential avenues for developing TRPA1 inhibitor analgesics and for probing the mechanism of channel gating.

A comprehensive approach to identifying repurposed drugs to treat SCN8A epilepsy.

OBJECTIVE: Many previous studies of drug repurposing have relied on literature review followed by evaluation of a limited number of candidate compounds. Here, we demonstrate the feasibility of a more comprehensive approach using high-throughput screening to identify inhibitors of a gain-of-function mutation in the SCN8A gene associated with severe pediatric epilepsy. METHODS: We developed cellular models expressing wild-type or an R1872Q mutation in the Nav 1.6 sodium channel encoded by SCN8A. Voltage clamp experiments in HEK-293 cells expressing the SCN8A R1872Q mutation demonstrated a leftward shift in sodium channel activation as well as delayed inactivation; both changes are consistent with a gain-of-function mutation. We next developed a fluorescence-based, sodium flux assay and used it to assess an extensive library of approved drugs, including a panel of antiepileptic drugs, for inhibitory activity in the mutated cell line. Lead candidates were evaluated in follow-on studies to generate concentration-response curves for inhibiting sodium influx. Select compounds of clinical interest were evaluated by electrophysiology to further characterize drug effects on wild-type and mutant sodium channel functions. RESULTS: The screen identified 90 drugs that significantly inhibited sodium influx in the R1872Q cell line. Four drugs of potential clinical interest-amitriptyline, carvedilol, nilvadipine, and carbamazepine-were further investigated and demonstrated concentration-dependent inhibition of sodium channel currents. SIGNIFICANCE: A comprehensive drug repurposing screen identified potential new candidates for the treatment of epilepsy caused by the R1872Q mutation in the SCN8A gene.

Discovery of a Series of Indazole TRPA1 Antagonists.

A series of TRPA1 antagonists is described which has as its core structure an indazole moiety. The physical properties and in vitro DMPK profiles are discussed. Good in vivo exposure was obtained with several analogs, allowing efficacy to be assessed in rodent models of inflammatory pain. Two compounds showed significant activity in these models when administered either systemically or topically. Protein chimeras were constructed to indicate compounds from the series bound in the S5 region of the channel, and a computational docking model was used to propose a binding mode for example compounds.

Biophysical and Pharmacological Characterization of Nav1.9 Voltage Dependent Sodium Channels Stably Expressed in HEK-293 Cells.

The voltage dependent sodium channel Nav1.9, is expressed preferentially in peripheral sensory neurons and has been linked to human genetic pain disorders, which makes it target of interest for the development of new pain therapeutics. However, characterization of Nav1.9 pharmacology has been limited due in part to the historical difficulty of functionally expressing recombinant channels. Here we report the successful generation and characterization of human, mouse and rat Nav1.9 stably expressed in human HEK-293 cells. These cells exhibit slowly activating and inactivating inward sodium channel currents that have characteristics of native Nav1.9. Optimal functional expression was achieved by coexpression of Nav1.9 with ß1/ß2 subunits. While recombinantly expressed Nav1.9 was found to be sensitive to sodium channel inhibitors TC-N 1752 and tetracaine, potency was up to 100-fold less than reported for other Nav channel subtypes despite evidence to support an interaction with the canonical local anesthetic (LA) binding region on Domain 4 S6. Nav1.9 Domain 2 S6 pore domain contains a unique lysine residue (K799) which is predicted to be spatially near the local anesthetic interaction site. Mutation of this residue to the consensus asparagine (K799N) resulted in an increase in potency for tetracaine, but a decrease for TC-N 1752, suggesting that this residue can influence interaction of inhibitors with the Nav1.9 pore. In summary, we have shown that stable functional expression of Nav1.9 in the widely used HEK-293 cells is possible, which opens up opportunities to better understand channel properties and may potentially aid identification of novel Nav1.9 based pharmacotherapies.

Voltage sensor interaction site for selective small molecule inhibitors of voltage-gated sodium channels.

Voltage-gated sodium (Nav) channels play a fundamental role in the generation and propagation of electrical impulses in excitable cells. Here we describe two unique structurally related nanomolar potent small molecule Nav channel inhibitors that exhibit up to 1,000-fold selectivity for human Nav1.3/Nav1.1 (ICA-121431, IC50, 19 nM) or Nav1.7 (PF-04856264, IC50, 28 nM) vs. other TTX-sensitive or resistant (i.e., Nav1.5) sodium channels. Using both chimeras and single point mutations, we demonstrate that this unique class of sodium channel inhibitor interacts with the S1-S4 voltage sensor segment of homologous Domain 4. Amino acid residues in the "extracellular" facing regions of the S2 and S3 transmembrane segments of Nav1.3 and Nav1.7 seem to be major determinants of Nav subtype selectivity and to confer differences in species sensitivity to these inhibitors. The unique interaction region on the Domain 4 voltage sensor segment is distinct from the structural domains forming the channel pore, as well as previously characterized interaction sites for other small molecule inhibitors, including local anesthetics and TTX. However, this interaction region does include at least one amino acid residue [E1559 (Nav1.3)/D1586 (Nav1.7)] that is important for Site 3 α-scorpion and anemone polypeptide toxin modulators of Nav channel inactivation. The present study provides a potential framework for identifying subtype selective small molecule sodium channel inhibitors targeting interaction sites away from the pore region.

The KCNQ2/3 selective channel opener ICA-27243 binds to a novel voltage-sensor domain site.

The mammalian KCNQ (Kv7) gene family is composed of five members (KCNQ1-5). KCNQ2, Q4 and Q5 (KCNQ2-5) channels co-express with KCNQ3 to form heterotetrameric voltage-gated K(+) (KCNQ2-5/3) channels that underlie the endogenous M-current and regulate neuronal excitability in CNS and PNS neurons. Openers of one or a mixture of these channels may be an attractive therapeutic agent for epilepsy and pain. Non-selective KCNQ2-5/3 activators have shown efficacy in pre-clinical and clinical studies. However, more selective pharmacological profiles, including greater KCNQ sub-type-selective activation, could provide efficacy with fewer side effects. One such compound, ICA-27243, sub-type selectively enhances the activation of KCNQ2/3 channels and also exhibits efficacy in pre-clinical anticonvulsant models; Roeloffs et al. (2008) [15]; Wickenden et al. (2008) [27]. The binding site of non-selective KCNQ2-5/3 openers maps to the S5-S6 pore domain and is altered by mutation of a tryptophan residue (Trp236 in KCNQ2, Trp265 in KCNQ3) conserved among KCNQ2-5 channels; Schenzer et al. (2005) [19]; Wuttke et al. (2005) [30]. Here we report that the activity of the KCNQ2/3 selective opener ICA-27243 is not affected by these Trp mutations and does not map to the S5-S6 domain. Rather, the selective activity of ICA-27243 is determined by a novel site within the S1-S4 voltage-sensor domain (VSD) of KCNQ channels. The sub-type-selective activity of ICA-27243 may arise from greater sequence diversity of KCNQ family members within the ICA-27243 binding pocket, allowing for more selective small molecule-protein interactions.

A phase I study of dose-dense topotecan given upfront to standard therapy in patients with small cell lung cancer.

OBJECTIVE: To determine the feasibility and maximum tolerated dose of dose-dense topotecan as induction chemotherapy before standard therapy (carboplatin plus etoposide alone or in combination with radiotherapy) in patients with small cell lung cancer (SCLC). PATIENTS AND METHODS: Chemotherapy-naive patients with SCLC and good performance status were eligible. Three 2-week cycles of dose-dense topotecan administered on days 1-3 with granulocyte colony-stimulating factor support were followed by four cycles of standard carboplatin plus etoposide therapy alone (extensive-stage SCLC) or with radiotherapy (limited-stage SCLC). The dose of topotecan was escalated from 1.5 mg/m2/day to 2.5 mg/m2/day in increments of 0.25 mg/m2/day within cohorts of 3-5 patients each. Dose-limiting toxicity was defined as any grade 3 or 4 toxicity resulting in a treatment reduction or a delay of >3 days. RESULTS: Twenty-two patients with SCLC (5 limited-stage, 17 extensive-stage) were enrolled. Treatment was well tolerated. The dose-limiting toxicities were thrombocytopenia and neutropenia, and the maximum tolerated dose of dose-dense topotecan induction therapy was 2.25 mg/m2/day. Overall, topotecan-related grade 3/4 haematological toxicities included neutropenia (n = 4), thrombocytopenia (n = 3) and febrile neutropenia (n = 1). No grade 4 non-haematological toxicities occurred. Grade 3 adverse events included nausea (n = 2), renal toxicity (n = 1) and anorexia (n = 1). Toxicity during the carboplatin plus etoposide +/- radiotherapy phase of therapy was consistent with that reported in previous trials. The overall response rate was 80% for limited-stage and 76% for extensive-stage SCLC. Median survival was 8 months in patients with limited-stage SCLC and 13.5 months for patients with extensive-stage SCLC. CONCLUSION: The results of this phase I study suggest that a regimen of sequential dose-dense topotecan and carboplatin plus etoposide is feasible, and the preliminary activity observed in patients with SCLC warrants further investigation at a starting dose of topotecan 2.25 mg/m2/day.

Patient privacy and conflicting legal and ethical obligations in El Salvador: reporting of unlawful abortions.

Postabortion care providers who breach patient confidentiality endanger women's health and violate ethics. A 1998 abortion ban in El Salvador likely spurred an increase in the number of women investigated, because many women were reported to legal authorities by health care providers. Having analyzed safeguards of confidentiality in laws and ethical guidelines, we obtained information from legal records on women prosecuted from 1998 to 2003 and identified factors that may lead to reporting through a survey of obstetrician-gynecologists (n=110). Although ethical and human rights standards oblige providers to respect patients' privacy, 80% of obstetrician-gynecologists mistakenly believed reporting was required. Most respondents (86%) knew that women delay seeking care because of fear of prosecution, yet a majority (56%) participated in notification of legal authorities.

Una mirada rápida a los años 2000-2002 la mortalidad materna en Nicaragua: la tragedia ignorada de la mortalidad materna nos hace una pregunta fundamental ¿en cuánto valoramos la vida de una mujer? / A quick look a years 2000-2002 the maternal mortality in Nicaragua: does the unknown tragedy of the maternal mortality ask us a fundamental question in how much we value the life of a woman?

Presente un análisis breve de la muerte materna en Nicaragua en los últimos años. El enfoque de este estudio está dirigido a examinar cambios en el comportamiento de las muertes maternas en cuanto a su incidencia, su causa, el momento, lugar de la muerte y los factores condionantes. Este análisis se hizo en base en una revisión de las fichas de mortalidad materna recopiladas por el Departamento de Atención Integral de la Mujer y la información generada por el sistema de vigilancia de mortalidad materna del Ministerio de Salud (MINSA). La información presentada muestra una radiografía de la situación actual, que apunta hacia una investigación más profunda de las causa no-obstétricas de la muerte materna y recalca la importancia de utilizar evidencia para fundamentar las intervenciones preventivas actuales y futuras

Un diagnóstico nacional de la atención post aborto: Nicaragua 2003 / A national diagnosis of the attention post abortion: Nicaragua 2003

El presente documento tiene el objetivo de evaluar la situación de los servicios públicos de atención post aborto (APA) en Nicaragua. Con esta información se pretende motivar la definición e implementación de estrategias encaminadas a lograr servicios de atención post aborto sostenible, seguros, eficaces y centrados en la mujer. La relevancia de este estudio está fundamentalmetne en el hecho de que el ingreso de una mujer con complicaciones relacionadas con el aborto, puede representar una oportunidad única de contacto con el sistema de salud. Este contacto tiene que ser aprovechado para que a las mujeres se les ofrezca una atención digna y humana sin condicionar el origen de su aborto. Se estima que ocurren 31,911 abortos anuales en Nicaragua, lo que representa el ingreso de un promedio de 5,50 mujeres en las unidades de salud a los servicios de atención post aborto. La importancia de que las mujeres tengan acceso a estos servicios fue reconocido oficialmente por el estado de Nicaragua hace nueve años cuando firmó el Programa de Acción de la Conferencia Internacional de Población y Desarrollo (CIPD) en 1994. Este estudio se realizó en el 100 porciento (n=47) de las unidades de salud del Ministerio de Salud de Nicaragua habilitadas para prestar este tipo de atención

Un diagnóstico nacional de la atención postaborto: nicaragua 2003 / A national diagnosis of the attention postaborto: Nicaragua 2003

El presente documento tiene el objetivo de evaluar la situación de los servicios públicos de atención post aborto (APA) en Nicaragua. Este estudio fue realizado en el 100 porciento (n=47) de las unidades de salud del Ministerio de Salud. Incluye estrategias encaminadas a lograr servicios de atención aborto sostenibles, seguros eficaces y centrados en la mujer

Enhanced dendritic cell antigen presentation in RNA-based immunotherapy.

BACKGROUND: Dendritic cells pulsed with mRNA provide a unique approach to tumor immunotherapy. We hypothesized that increased mRNA transfection efficiency and dendritic cell maturation would improve antigen processing and presentation as well as T-cell costimulation, resulting in enhanced induction of antimelanoma immune responses. METHODS: Immature monocyte-derived dendritic cells were transfected with mRNA by passive pulsing, lipofection, or electroporation. Dendritic cells were either left untreated or matured using the double-stranded RNA poly(I:C). T-Cell cultures were generated by stimulation of naïve T-cells with each set of dendritic cells. Specific antigen presentation and specific effector T-cell generation were analyzed by an IFN-gamma release Elispot assay. RESULTS: Greatest intracellular green fluorescent protein was observed by flow cytometry following dendritic cell electroporation with green fluorescent protein mRNA. DC presentation of Mart-1/Melan A peptide, as measured by Elispot assay using a specific T-cell clone, was greatest following transfection with Mart-1/Melan A mRNA by electroporation. Maturation of dendritic cells further improved antigen presentation regardless of transfection technique. Specific Mart-1/Melan A effector T cells were produced after culture of naïve T cells with dendritic cells that were electroporated with Mart-1/Melan A mRNA and then matured, but not for dendritic cells that remained immature. CONCLUSIONS: Efficient mRNA transfection by electroporation as well as dendritic cell maturation results in increased levels of Mart-1/Melan A antigen presentation and enhanced production of antigen-specific effector T cells. This combination of strategies may be used to enhance immune responses to RNA-based dendritic cell vaccines.