Search

Additional file 1. Strategies to train new staff and assure consistency of the intervention. List of strategies to ensure intervention consistency and train new staff.

Additional file 3. Physical Activity Unified-7 item Screener (PAU-7S). English version of Physical Activity Unified– 7 items Screener (PAU-7S) administered to children.

Estrada JL, Bautista NS, Dionisio-Sese ML. 2020. Morphological variation of two common sea grapes (Caulerpa lentillifera and Caulerpa racemosa) from selected regions in the Philippines. Biodiversitas 21: 1823-1832. Seagrapes, locally known in the Philippines as “lato” or “ar-arusip”, are economically important macroalgae belonging to the edible species of the genus Caulerpa. This study characterized and compared distinct populations of sea grapes from selected regions in the Philippines and described the influence of physicochemical parameters of seawater on their morphology. Morphometric, cluster and principal component analyses showed that morphological plasticity exists in sea grapes species (Caulerpa lentillifera and Caulerpa racemosa) found in different sites in the Philippines. These are evident in morphometric parameters namely, assimilator height, space between assimilators, ramulus diameter and number of rhizoids on stolon wherein significant differences were found. This evident morphological plasticity was analyzed in relation to physicochemical parameters of the seawater. Assimilator height of C. racemosa is significantly associated and highly influenced by water depth, salinity, temperature and dissolved oxygen whereas for C. lentillifera depth and salinity are the significant influencing factors. Highest assimilator height of sea grapes was found in Coron and Culion in Palawan due to higher salinity and temperature while in Mactan, Cebu, it is primarily attributed to lower depth and higher salinity values.

Poor solubility is the main drawback of the direct industrial exploitation of chitin, the second most abundant biopolymer after cellulose. Chemical methods are conventional to solubilize chitin from natural sources. Enzymatic hydrolysis of soluble chitinous substrates is a promising approach to obtain value-added by-products, such as N-acetylglucosamine units or low molecular weight chito-oligomers. Protein display on the bacterial membrane remains attractive to produce active enzymes anchored to a biological surface. The Lpp-OmpA system, a gene fusion of the Lpp signal sequence with the OmpA transmembrane region, represents the traditional system for targeting enzymes to the E. coli surface. EhCHT1, the amoebic chitinase, exhibits an efficient endochitinolytic activity and significant biochemical features, such as stability over a wide range of pH values. Using an extended Lpp-OmpA system as a protein carrier, we engineered E. coli to express the catalytic domain of EhCHT1 on the surface and assess the endochitinase activity as a trait. Engineered bacteria showed a consistent hydrolytic rate over a typical substrate, suggesting that the displayed enzyme has operational stability. This study supports the potential of biomembrane-associated biocatalysts as a reliable technology for the hydrolysis of soluble chitinous substrates.

Cancer incidence rates vary regionally among American Indians (AIs) and Latinos. The goal of this was to identify areas of research necessary to reduce cancer health disparities in AIs and Latinos, the two major racial/ethnic minority groups in Arizona. In an effort to better understand cancer health disparities, cancer incidence rates in AIs and Latinos in Arizona were compared to non-Hispanic Whites (NHWs). Age-adjusted incidence rates (per 100,000) were obtained from the Arizona Cancer Registry and the North American Association of Central Cancer Registries. Spearman’s rank test was used to examine correlation between county-level cancer incidence rates and socio-demographic factors. AIs and Latinos had lower incidence rates of screening for detectable cancers than NHWs. Among older men (age ≥65), however, AIs and Latinos had similar prostate cancer incidence rates to NHWs. Some of less common cancers, such as kidney, stomach, liver, and gallbladder, were more frequently diagnosed in AIs and Latinos than NHWs. AIs and Latinos were more likely to be diagnosed with advanced cancer stage, except for cervical cancer. Correlations between prostate and breast cancer incidence rates and percent urban residents as well as correlations between incidence rates of these two cancer types and population size were significantly positive. Poverty levels were inversely correlated with colorectal and lung cancer incidence rates. Our review of cancer incidence rates suggests that socio-demographic factors, such as population size (rural/urban) and poverty levels, have influenced cancer detection and incidence rates in Arizona.

Spanish-only speaking residents in the United States face barriers to receiving potentially life-saving 911 interventions such as Telephone -cardiopulmonary resuscitation (TCPR) instructions. Since 2015, 911 dispatchers have placed an increased emphasis on rapid identification of potential cardiac arrest. The purpose of this study was to describe the utilization and timing of the 911 system during suspected out-of-hospital cardiac arrest (OHCA) by Spanish-speaking callers in Metropolitan Phoenix, Arizona.The dataset consisted of suspected OHCA from 911 centers from October 10, 2010 through December 31, 2013. Review of audio TCPR process data included whether the need for CPR was recognized by telecommunicators, whether CPR instructions were provided, and the time elements from call receipt to initiation of compressions.A total of 3398 calls were made to 911 for suspected OHCA where CPR was indicated. A total of 39 (1.2%) were determined to have a Spanish language barrier. This averages to 18 calls per year with a Spanish language barrier during the study period, compared with 286 OHCAs expected per year among this population. The average time until telecommunicators recognized CPR need was 87.4s for the no language barrier group compared to 160.6s for the Spanish-language barrier group (p0.001).Time to CPR instructions started was significantly different between these groups (144.4s vs 231.3s, respectively) (p0.001), as was time to first compression, (174.4s vs. 290.9s, respectively) (p0.001).Our study suggests that Hispanic callers under-utilize the 911 system, and when they do call 911, there are significant delays in initiating CPR.

Introduction and objectives Heart failure is a common comorbidity of COPD and contributes to increased breathlessness and adverse clinical outcomes. Neural respiratory drive (NRD) is closely related to breathlessness intensity in COPD. This study aimed to investigate the impact of comorbid left heart failure (COPD-HF) on NRD in patients with COPD. We hypothesised that NRD would be higher during an inspiratory threshold loading protocol (ITL) in COPD-HF than in COPD patients without left heart failure. Methods COPD and COPD-HF patients underwent incremental ITL at 12%, 24%, 36%, 48% and 60% of maximal inspiratory mouth pressure (PImax). NRD was recorded continuously using 2nd intercostal space transcutaneous electromyography (EMGpara). EMGpara signals were converted to root mean square (RMS), normalised to peak RMS EMGpara during maximal inspiratory manoeuvres (EMGpara%max) and multiplied by respiratory rate to calculate NRD index (NRDI). NRDI in COPD and COPD-HF were compared at each load using mixed effect model repeated measurement analysis. Results 11 COPD patients without left heart failure (mean (SD) age 69(7) years, FEV1%predicted 49.3 (16.4)%, VC%predicted 99.8 (22.0)%, PIMax 55.7 (15.8)cmH2O) and 11 COPD-HF patients (mean (SD) age 72(6) years, FEV1%predicted 54.8 (13.6)%, VC%predicted 86.8 (17.4)%), PIMax 53.1 (30.9)cmH2O) were studied. mMRC dyspnoea scores were higher in COPD-HF (median (IQR) 3 (2 – 4) than in COPD (median (IQR) 2 (1 to 3), p=0.0406). 11/11 COPD patients completed all loads of the ITL protocol to 60% PImax, compared to 4/11 patients in the COPD-HF group (p=0.0039). There were significant fixed effects of diagnosis (p=0.0136), load (p Conclusions Observations of higher levels of NRDI at equivalent inspiratory threshold loads in COPD-HF suggests that heart failure further increases the mechanical load on the respiratory muscles in COPD. Contributions of potential aetiological factors, such as reduced lung and chest wall compliance, require further study.

Study Objectives To evaluate the contribution of long-term and short-term REM sleep homeostatic processes to REM sleep recovery and the ultradian organization of the sleep wake cycle. Methods Fifteen rats were sleep recorded under a 12:12 LD cycle. Animals were subjected during the rest phase to two protocols (2T2I or 2R2I) performed separately in non-consecutive experimental days. 2T2I consisted of 2 h of total sleep deprivation (TSD) followed immediately by 2 h of intermittent REM sleep deprivation (IRD). 2R2I consisted of 2 h of selective REM sleep deprivation (RSD) followed by 2 h of IRD. IRD was composed of four cycles of 20-min RSD intervals alternating with 10 min of sleep permission windows. Results REM sleep debt that accumulated during deprivation (9.0 and 10.8 min for RSD and TSD, respectively) was fully compensated regardless of cumulated NREM sleep or wakefulness during deprivation. Protocol 2T2I exhibited a delayed REM sleep rebound with respect to 2R2I due to a reduction of REM sleep transitions related to enhanced NREM sleep delta-EEG activity, without affecting REM sleep consolidation. Within IRD permission windows there was a transient and duration-dependent diminution of REM sleep transitions. Conclusions REM sleep recovery in the rat seems to depend on a long-term hourglass process activated by REM sleep absence. Both REM sleep transition probability and REM sleep episode consolidation depend on the long-term REM sleep hourglass. REM sleep activates a short-term REM sleep refractory period that modulates the ultradian organization of sleep states.

espanolResumen Las interacciones bioticas como depredacion, forrajeo y herbivoria han sido generalmente catalogadas como las fuerzas que determinan las variaciones espaciales y temporales de las poblaciones planctonicas, tanto de presas como de depredadores. Una especie del zooplancton, presente en aguas continentales, calificada como depredadora es el rotifero Asplanchna girodi. Sin embargo, se desconoce la composicion real de su dieta, las preferencias alimentarias y la dinamica espacial y temporal en relacion con sus presas o alimento. En esta investigacion se determinaron las preferencias troficas de A. girodi y su fluctuacion espacio-temporal, basados en items verdaderamente consumidos en el medio natural, a traves del analisis de los contenidos estomacales de individuos capturados en las aguas del embalse Riogrande II (Antioquia-Colombia), durante un ano de muestreos realizados cada 10 dias. Se observo que A. girodi presento una preferencia trofica amplia que incluyo el forrajeo de, por lo menos, cinco taxones de algas y la depredacion de ocho (8) especies de rotiferos y del cladocero Bosmina freyi. Se concluye que: 1. A. girodi es una especie omnivora, con mayor preferencia por el consumo de algas. 2. Este rotifero no fue factor de presion en la dinamica poblacional de las algas, pero posiblemente lo fue para Keratella americana. 3. No se observaron diferencias espaciales en la dieta de A. girodi, pero si existieron diferencias temporales en cuanto a la composicion de la dieta, la cual estuvo determinada por la oferta de las algas, principalmente durante florecimientos de Ceratium. 4. Este tipo de dieta se enmarca dentro de los conceptos de forrajeo optimo y oportunismo trofico, estrategias que conllevan el consumo de las especies con mayor disponibilidad (en este caso algas) y el maximo ahorro de energia, puesto que no se invierte en la busqueda de presas especificas. EnglishAbstract Biotic interactions such as predation, foraging and herbivory have been generally categorized as the forces that determine the spatial and temporal variations of planktonic populations, both preys and predators. A species of zooplankton, present in continental waters, classified as predator is the rotifer Asplanchna girodi. However, the true composition of its diet, dietary preferences and spatial and temporal dynamics in relation to its prey or food are unknown. In this research the trophic preferences of A. girodi and its spatio-temporal fluctuation were determined, based on items truly consumed in the natural environment, through analysis of stomach contents of individuals captured in the waters of Riogrande II reservoir (Antioquia-Colombia), during one year of sampling every 10 days. It was observed that A. girodi showed a broad trophic preference that included the foraging of, at least, five algae taxa and the predation of eight rotifer species and cladoceran Bosmina freyi. It is concluded that: 1. A. girodi is an omnivorous species, with a greater preference for the consumption of algae. 2. This rotifer was not a pressure factor in the population dynamics of the algae, but possibly it was for the Keratella americana. 3. No spatial differences were observed in the A. girodi diet, but there were temporal differences in the composition of the diet, which was determined by the algae supply, mainly during Ceratium blooms. 4. This type of diet is framed within the concepts of optimal foraging and trophic opportunism, strategies that involve the consumption of the most available species (in this case algae) and the maximum energy saving since it is not invested in the search for specific prey. portuguesResumo Interacoes bioticas como predacao, forrageamento e herbivoria tem sido geralmente classificadas como as forcas que determinam as variacoes espaciais e temporais de populacoes planctonicas, tanto de presas quanto de predadores. Uma especie de zooplâncton, presente em aguas continentais, classificada como predador e o rotifero Asplanchna girodi. No entanto, a composicao real de sua dieta, preferencias alimentares e dinâmicas espaciais e temporais em relacao a suas presas ou alimentos sao desconhecidas. Nesta pesquisa, as preferencias troficas de A. girodi e sua flutuacao espaco-temporal foram determinadas, com base em itens verdadeiramente consumidos no ambiente natural, atraves da analise do conteudo estomacal de individuos capturados nas aguas do reservatorio Riogrande II (Antioquia-Colombia), durante um ano de amostragem a cada 10 dias. Observou-se que A. girodi apresentou uma ampla preferencia trofica que incluiu o forrageamento de pelo menos cinco taxa de algas e a predacao de oito (8) especies de rotiferos e o cladocero Bosmina freyi. Conclui-se que: 1. A. girodi e uma especie onivora, com maior preferencia pelo consumo de algas. 2. Este rotifero nao foi um fator de pressao na dinâmica populacional das algas, mas possivelmente foi para Keratella americana. 3. Nao foram observadas diferencas espaciais na dieta de A. girodi, mas houve diferencas temporarias na composicao da dieta, que foi determinado pelo fornecimento de algas, principalmente durante a floracao de Ceratium. Este tipo de dieta enquadra-se nos conceitos de forrageamento otimo e oportunismo trofico; estrategias que envolvem o consumo de especies com maior disponibilidade (neste caso algas) e a maxima economia de energia, uma vez que nao e investido na busca por presas especificas.

Introduction Breathlessness is a cardinal feature of COPD and is frequently distressing and disabling. Thorough assessment of breathlessness is necessary for its effective management, but this is often not achieved in clinical practice. Although no single tool for subjectively measuring breathlessness and disease impact is universally accepted, the questionnaire based modified (m)MRC Dyspnoea Scale and COPD Assessment Test (CAT) scores are widely used in clinical practice and research. Lung function tests and other objective physiological biomarkers in routine clinical practice correlate poorly with patient-reported breathlessness. Neural respiratory drive (NRD), as quantified by transoesophageal diaphragm electromyography (EMGdi), is closely related to breathlessness during cardiopulmonary exercise testing in COPD. Surface parasternal intercostal muscle EMG activity (sEMGpara) can be used as a non-invasive alternative to EMGdi. This study aimed to investigate the relationship between mMRC dyspnoea, and CAT scores, and sEMGpara in COPD patients at rest and when breathing against an inspiratory threshold load. Methods sEMGpara was recorded in 17 COPD patients (mean±SD FEV151.09%±15.1% predicted,14 male, age 70.9±6.6 years), breathing against an inspiratory threshold load at 60% of maximum mouth inspiratory pressure (60% PImax). mMRC dyspnoea and CAT scores were recorded pre-protocol at rest. Mean peak root mean square (RMS) sEMGpara per breath was calculated, and relationships with mMRC dyspnoea and CAT scores were determined by correlation analysis. Results Significant positive correlations were observed between mMRC dyspnoea and sEMGpara at 60%PImax (Spearman r=0.624, p=0.007) and between CAT scores and sEMGpara at 60%PImax (Pearson r=0.497, p=0.042) (figure 1). There were no significant correlations between mMRC dyspnoea score and sEMGpara at rest (Spearman r=−0.048, p=0.853) or FEV1% predicted (r=−0.143, p=0.584), or between CAT score and sEMGpara at rest (Pearson r=0.107, p=0.682) or FEV1% predicted (r=−0.081, p=0.756). Conclusion sEMGpara recorded at 60% PImax, but not at rest, correlated with patient-reported breathlessness and disease impact in COPD. This suggests that sEMGpara recorded at 60% PImax could provide a clinically-useful objective physiological correlate of breathlessness-related quality of life in COPD.

Introduction Breathlessness is a dominant, distressing, symptom of chronic obstructive pulmonary disease (COPD) which has a significant impact on quality of life. Neural respiratory drive (NRD), quantified as diaphragm electromyogram (EMGdi) activity, is closely related to breathlessness intensity in COPD. The surface mechanomyogram (sMMG), which can be recorded using skin-surface triaxial accelerometers, measures the lateral vibration of muscle fibres and is considered to be the mechanical counterpart of the surface EMG (sEMG). The principal aim of this study was to investigate the relationship between the sMMG of the diaphragm (sMMGdi) and breathlessness in COPD. We hypothesised that there would be strong positive relationships between sMMGdi and modified Borg (mBorg) breathlessness intensity during an inspiratory threshold loading protocol. Methods Nine COPD patients (7 male, median (IQR) age 70 (64–78) years, BMI 25.6 (24.8–30.1) kg/m2, FEV144.7 (37.1–63.9)%predicted), underwent an incremental inspiratory loading protocol at 12%, 24%, 36%, 48% and 60% of maximum mouth inspiratory pressure (PImax). Left and right sEMGdi and sMMGdi activity were recorded using 2 pairs of sEMG electrodes, and one pair of triaxial accelerometers, positioned over the 7th/8th intercostal spaces in the anterior axillary lines bilaterally. sEMG and sMMG signals were analysed using fixed sample entropy (fSampEn) and the relationships with mBorg breathlessness intensity during each inspiratory load were determined using correlation analysis. Results Moderate to strong correlations were observed between mBorg breathlessness intensity and the sEMG and sMMG signals, expressed as fSampEn, recorded at all sites (table 1). Conclusion The relationships observed between mBorg breathlessness intensity and sMMGdi, and sEMGdi, expressed as fSampEn, demonstrates the potential value of sMMGdi as a complementary tool to sEMGdi for the investigation of breathlessness in COPD. sMMGdi could provide a novel non-invasive physiological correlate of breathlessness intensity in COPD and other chronic respiratory diseases.

The future of solid organ transplantation is challenged by an increasing shortage of available allografts. Xenotransplantation of genetically modified porcine organs offers an answer to this problem. Strategies of genetic modification have ‘humanized’ the porcine model towards clinical relevance. Most notably, these approaches have aimed at either antigen reduction or human transgene expression. The object of this study was to evaluate the relative effects of both antigen reduction and direct complement regulation on the human-anti-porcine complement dependent cytotoxicity response. Genetically modified animals were created through CRISPR/Cas9-directed mutation and human transgene delivery. Pigs doubly deficient in GGTA1 and CMAH genes were compared to pigs of the same background that expressed a human complement regulatory protein (hCRP). A third animal was made deficient in GGTA1, CMAH and B4GalNT2 gene expression. Cells from these animals were subjected to measures of human antibody binding and antibody-mediated complement-dependent cytotoxicity by flow cytometry. Human IgG and IgM antibody binding was unchanged between the double knockout and the transgenic hCRP double knockout pig. IgG and IgM binding was reduced by 49.1 and 43.2 % respectively by silencing the B4GalNT2 gene. Compared to the double knockout, human anti-porcine cytotoxicity was reduced by 8 % with the addition of a hCRP (p = .032); It was reduced by 21 % with silencing the B4GalNT2 gene (p = .012). Conclusions: Silencing the GGTA1, CMAH and B4GalNT2 genes in pigs achieved a significant antigen reduction. Changing the porcine carbohydrate profile effectively mediates human antibody-mediated complement dependent cytoxicity.

BACKGROUND Disrupting the porcine GGTA1 and CMAH genes [double knockout (DKO)] that produce the gal-α(1,3)-gal and N-glycolylneuraminic acid xenoantigens reduces human antibody binding to porcine peripheral blood mononuclear cells. It is important to examine rejection pathways at an organ-specific level. The object of this study is to evaluate the human preformed antibody reactivity against DKO renal microvascular endothelial cells (RMEC) in vitro. METHODS Characteristics of DKO RMEC were analyzed using flow cytometry. Human IgG/M binding to primary RMEC, immortalized RMEC (iRMEC), and iRMEC-deficient in B4GALNT2 genes were examined using flow cytometric crossmatch assay. RESULTS Porcine RMEC expressed gal-α(1,3)-gal, N-glycolylneuraminic acid, and Dolichos biflorus agglutinin glycans recognized by human preexisting antibodies in humans. Antigenicity of DKO RMEC was lower than GGTA1 KO RMEC. The disruption of B4GALNT2 gene in DKO iRMEC further reduced human IgG/IgM binding. CONCLUSIONS Silencing the porcine GGTA1, CMAH, and B4GALNT2 genes is an effective strategy to reduce human preformed antibody binding to RMEC. Porcine RMEC will be a useful reagent for the further study of xenoimmunology.

A profound thrombocytopenia limits hepatic xenotransplantation in the pig-to-primate model. Porcine livers also have shown the ability to phagocytose human platelets in the absence of immune-mediated injury. Recently, inactivation of the porcine ASGR1 gene has been shown to decrease this phenomenon. Inactivating GGTA1 and CMAH genes has reduced the antibody-mediated barrier to xenotransplantation; herein, we describe the effect that these modifications have on xenogeneic consumption of human platelets in the absence of immune-mediated graft injury.Wild type (WT), ASGR1, GGTA1, and GGTA1CMAH knockout pigs were compared for their xenogeneic hepatic consumption of human platelets. An in vitro assay was established to measure the association of human platelets with liver sinusoidal endothelial cells (LSECs) by immunohistochemistry. Perfusion models were used to measure human platelet uptake in livers from WT, ASGR1, GGTA1, and GGTA1 CMAH pigs.GGTA1, CMAH LSECs exhibited reduced levels of human platelet binding in vitro when compared with GGTA1 and WT LSECs. In a continuous perfusion model, GGTA1 CMAH livers consumed fewer human platelets than GGTA1 and WT livers. GGTA1 CMAH livers also consumed fewer human platelets than ASGR1 livers in a single-pass model.Silencing the porcine carbohydrate genes necessary to avoid antibody-mediated rejection in a pig-to-human model also reduces the xenogeneic consumption of human platelets by the porcine liver. The combination of these genetic modifications may be an effective strategy to limit the thrombocytopenia associated with pig-to-human hepatic xenotransplantation.

Background The Galα(1,3)Gal epitope (α-GAL), created by α-1,3-glycosyltransferase-1 (GGTA1), is a major xenoantigen causing hyperacute rejection in pig-to-primate and pig-to-human xenotransplantation. In response, GGTA1 gene-deleted pigs have been generated. However, it is unclear whether there is a residual small amount of α-Gal epitope expressed in GGTA1−/− pigs. Isoglobotrihexosylceramide synthase (iGb3s), another member of the glycosyltransferase family, catalyzes the synthesis of isoglobo-series glycosphingolipids with an α-GAL-terminal disaccharide (iGb3), creating the possibility that iGb3s may be a source of α-GAL epitopes in GGTA1−/− animals. The objective of this study was to examine the impact of silencing the iGb3s gene (A3GalT2) on pig-to-primate and pig-to-human immune cross-reactivity by creating and comparing GGTA1−/− pigs to GGTA1−/−- and A3GalT2−/−-double-knockout pigs. Methods We used the CRISPR/Cas 9 system to target the GGTA1 and A3GalT2 genes in pigs. Both GGTA1 and A3GalT2 genes are functionally inactive in humans and baboons. CRISPR-treated cells used directly for somatic cell nuclear transfer produced single- and double-gene-knockout piglets in a single pregnancy. Once grown to maturity, the glycosphingolipid profile (including iGb3) was assayed in renal tissue by normal-phase liquid chromatography. In addition, peripheral blood mononuclear cells (PBMCs) were subjected to (i) comparative cross-match cytotoxicity analysis against human and baboon serum and (ii) IB4 staining for α-GAL/iGb3. Results Silencing of the iGb3s gene significantly modulated the renal glycosphingolipid profile and iGb3 was not detected. Moreover, the human and baboon serum PBMC cytotoxicity and α-GAL/iGb3 staining were unchanged by iGb3s silencing. Conclusions Our data suggest that iGb3s is not a contributor to antibody-mediated rejection in pig-to-primate or pig-to-human xenotransplantation. Although iGb3s gene silencing significantly changed the renal glycosphingolipid profile, the effect on Galα3Gal levels, antibody binding, and cytotoxic profiles of baboon and human sera on porcine PBMCs was neutral.

Reactions of trans-[Rh(CO)(Cl){P{(CH2)14}3P2(Rh–P2)}] (1) with ZnPh2, MeLi, or NaBH4 give the chloride-substitution products trans-[Rh(CO)(Ph){P{(CH2)14}3P2(Rh–P2)}] (3, 89 %), trans-[Rh(CO)(Me){P{(CH2)14}3P2(Rh–P2)}] (94 %, 92 % purity), and trans-[Rh(CO)(H2BH2){P{(CH2)14}3P2(Rh–P2)}] (5, 99 %). The crystal structures of 3, 5, and the corresponding (fortuitously obtained) bromide complex are determined, and the ability of the X–Rh–CO moieties to rotate is interpreted in terms of their effective radii and the void space within the dibridgehead diphosphine cage. Reaction of 1 and excess PMe3 gives the diphosphine substitution product trans-[Rh(CO)(Cl)(PMe3)2] (89 %) and the dibridgehead diphosphine P[(CH2)14]3P (58 %). Complex 5 serves as a catalyst precursor for the hydroformylation of 1-octene (neat or THF, 60 °C, 75 psig 1:1 CO/H2). The locus of catalysis with respect to the diphosphine cage is discussed.

Neoadjuvant therapy (NAT) is a useful therapeutic option. However, some patients respond poorly to it and can even show tumor progression. It is important to define factors that can predict response to NAT. This is a retrospective cohort study to define histopathological factors predicting response to NAT in gastric tubular carcinoma. This study has enrolled 80 patients receiving chemotherapy for locally advanced gastric carcinoma. 44.5% of the patients were men; mean age was 64.49 years. Only 5.7% of the patients showed a complete response to therapy, 10% had grade 1, 21.4% grade 2, and 62.9% grade 3 regression. On follow-up, 43.8% of the patients showed recurrence of disease (57.1% distant metastasis) and 33.8% eventually died of it. We found a statistically significant association between response and prognosis. We found a statistically significant association between regression and perineural, vascular, and lymph vessel invasion. Logistic regression model showed that only lymph vessel invasion had independent influence. Lymph vessel invasion not only indicated lack of response to therapy, but also higher incidence of lymph node involvement in the gastrectomy specimen. Our study indicates that the presence of vascular or perineural invasion in the endoscopic biopsies and high histopathological grade predict poor response to therapy. This seems peculiar, for undifferentiated tumors are supposed to have better response to therapy. Our study indicates that undifferentiated tumors respond worse to therapy. Furthermore, studies are necessary to define lack of response, to help avoid neoadjuvant therapy in unfavorable cases.

Antipig antibodies are a barrier to clinical xenotransplantation. We evaluated antibody binding of waitlisted renal transplant patients to 3 glycan knockout (KO) pig cells and class I swine leukocyte antigens (SLA).Peripheral blood mononuclear cells from SLA identical wild type (WT), α1, 3-galactosyltransferase (GGTA1) KO, GGTA1/ cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) KO, and GGTA1/ CMAH /b1,4 N-acetylgalactosaminyl transferase (B4GalNT2) KO pigs were screened for human antibody binding using flow cytometric crossmatch (FCXM). Sera from 820 patients were screened on GGTA1/CMAH/B4GalNT2 KO cells and a subset with elevated binding was evaluated further. FCXM was performed on SLA intact cells and GGTA1/SLA class I KO cells after depletion with WT pig RBCs to remove cell surface reactive antibodies, but leave SLA antibodies. Lastly, human and pig reactive antibodies were eluted and tested for cross-species binding and reactivity to single-antigen HLA beads.Sequential glycan KO modifications significantly reduce antibody binding of waitlisted patients. Sera exhibiting elevated binding without reduction after depletion with WT RBCs demonstrate reduced binding to SLA class I KO cells. Human IgG, eluted from human and pig peripheral blood mononuclear cells, interacted across species and bound single-antigen HLA beads in common epitope-restricted patterns.Many waitlisted patients have minimal xenoreactive antibody binding to the triple KO pig, but some HLA antibodies in sensitized patients cross-react with class I SLA. SLA class I is a target for genome editing in xenotransplantation.

Background Pig erythrocytes are potentially useful to solve the worldwide shortage of human blood for transfusion. Domestic pig erythrocytes, however, express antigens that are bound by human preformed antibodies. Advances in genetic engineering have made it possible to rapidly knock out the genes of multiple xenoantigens, namely galactose α1,3 galactose (aGal) and N-glycolylneuraminic acid (Neu5Gc). We have recently targeted the GGTA1 and CMAH genes with zinc finger endonucleases resulting in double knockout pigs that no longer express aGal or Neu5Gc and attract significantly fewer human antibodies. In this study, we characterized erythrocytes from domestic and genetically modified pigs, baboons, chimpanzees, and humans for binding of human and baboon natural antibody, and complement-mediated lysis. Methods Distribution of anti-Neu5Gc IgG and IgM in pooled human AB serum was analyzed by ELISA. Erythrocytes from domestic pigs (Dom), aGal knockout pigs (GGTA1 KO), aGal and Neu5Gc double knockout pigs (GGTA1/CMAH KO), baboons, chimpanzees, and humans were analyzed by flow cytometry for aGal and Neu5Gc expression. In vitro comparative analysis of erythrocytes was conducted with pooled human AB serum and baboon serum. Total antibody binding was accessed by hemagglutination; complement-dependent lysis was measured by hemolytic assay; IgG or IgM binding to erythrocytes was characterized by flow cytometry. Results The pooled human AB serum contained 0.38 μg/ml anti-Neu5Gc IgG and 0.085 μg/ml anti-Neu5Gc IgM. Both Gal and Neu5Gc were not detectable on GGTA1/CMAH KO erythrocytes. Hemagglutination of GGTA1/CMAH KO erythrocytes with human serum was 3.5-fold lower compared with GGTA1 KO erythrocytes, but 1.6-fold greater when agglutinated with baboon serum. Hemolysis of GGTA1/CMAH KO erythrocytes by human serum (25%) was reduced 9-fold compared with GGTA1 KO erythrocytes, but increased 1.64-fold by baboon serum. Human IgG binding was reduced 27-fold on GGTA1/CMAH KO erythrocytes compared with GGTA1 KO erythrocytes, but markedly increased 3-fold by baboon serum IgG. Human IgM binding was decreased 227-fold on GGTA1/CMAH KO erythrocytes compared with GGTA1 KO erythrocytes, but enhanced 5-fold by baboon serum IgM. Conclusions Removal of aGal and Neu5Gc antigens from pig erythrocytes significantly reduced human preformed antibody-mediated cytotoxicity but may have complicated future in vivo analysis by enhancing reactivity from baboons. The creation of the GGTA1/CMAH KO pig has provided the xenotransplantation researcher with organs and cells that attract fewer human antibodies than baboon and our closest primate relative, chimpanzee. These finding suggest that while GGTA1/CMAH KO erythrocytes may be useful for human transfusions, in vivo testing in the baboon may not provide a direct transplantation to the clinic.

Background The process of selecting transgenic cells has been one of the bottlenecks in the generation of transgenic animals by somatic cell nuclear transfer (SCNT). In particular, selection for the Gal double-knockout (Gal-DKO) genotype has been time consuming and inefficient. The objective of this work was to generate a highly efficient system to select Gal-DKO cells to be used in SCNT without affecting the efficiency in production of Gal-null pigs. Materials and methods Fetal liver–derived cells deficient in Gal-expression were initially selected by fluorescence-activated cell sorting (FACS) using IB4 conjugated to a fluorescent dye. Cells recovered by FACS were cultured and expanded, followed by a second round of selection using streptavidin magnetic beads and IB4 lectin biotin. Results Recovery efficiency of target cells was 0.04% for the first selection using FACS and 0.3% for the second round by magnetic beads. Full reprogramming was obtained on selected Gal-DKO cells after FACS and magnetic beads selection, when used for SCNT to produce the Gal-null piglets. Cells obtained from magnetic beads developed 48 colonies; the Gal-null genotype was found in 44 of them (91.7%). Three of these colonies were used to generate piglets by SCNT. From three recipients receiving embryos, two became pregnant and produced 17 piglets, all of them DKO. Conclusions Sequential selection of Gal-DKO cells by FACS/magnetic beads is a highly efficient system to generate null cells. Selected cells were successfully used to generate healthy double-knockout piglets by SCNT.

Background Clinical xenotransplantation is not possible because humans possess antibodies that recognize antigens on the surface of pig cells. Galα-1,3-Gal (Gal) and N-glycolylneuraminic acid (Neu5Gc) are two known xenoantigens. Methods We report the homozygous disruption of the α1, 3-galactosyltransferase (GGTA1) and the cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) genes in liver-derived female pig cells using zinc-finger nucleases (ZFNs). Somatic cell nuclear transfer (SCNT) was used to produce healthy cloned piglets from the genetically modified liver cells. Antibody-binding and antibody-mediated complement-dependent cytotoxicity assays were used to examine the immunoreactivity of pig cells deficient in Neu5Gc and Gal. Results This approach enabled rapid production of a pig strain deficient in multiple genes without extensive breeding protocols. Immune recognition studies showed that pigs lacking both CMAH and GGTA1 gene activities reduce the humoral barrier to xenotransplantation, further than pigs lacking only GGTA1. Conclusions This technology will accelerate the development of pigs for xenotransplantation research.

In this work, a characterization and modeling of packet loss of a Voice over Internet Protocol (VoIP) communication is developed. The distributions of the number of consecutive received and lost packets (namely gap and burst) are modeled from the transition probabilities of two-state and four-state model. Measurements show that both models describe adequately the burst distribution, but the decay of gap distribution for non-homogeneous losses is better fit by the four-state model. The respective probabilities of transition between states for each model were estimated with a proposed algorithm from a set of monitored VoIP calls in order to obtain representative minimum, maximum and average values for both models., {"references":["T. Tao, J. Lu, K. Gong, J. Gu. A four-states Markov model for burst\nerror analysis in satellite communications. International Conference on\nCommunication Technology .Vol. 1, pp. 930-934, August 2000.","Wai-Ki Ching, Michael K. Ng. Markov Chains: Models, Algorithms and\nApplications. Springer, 2006.","G. George Yin, Qing Zhang. Discrete-Time Markov Models, Two-\nTime-Scale Methods and Applications, pp. 24-28, Springer, 2005.","Hyungkeun. Lee, Hyukjoon. Lee. A packet loss recovery scheme based\non the gap statistics. C. Kim (Ed.): ICOIN 2005, LNCS 3391, pp. 627-\n634, 2005. Springer-Verlag Berlin Heidelberg, 2005.","J. R. Yee, E. J. Weldon. Evaluation of the performance of error\ncorrecting codes on a Gilbert channel. IEEE transactions on\nCommunications, Vol. 43 No. 8, 1995.","H. Toral, D. Torres, C. Hernandez, L. Estrada. Self-similarity, packet\nloss, jitter, and packet size: empirical relationships for VoIP. 18th\nInternational Conference on Electronics, Communications and\nComputers. CONIELECOMP, March 2008.","L. Estrada, D. Torres, H.Toral, \"Analytical Investigation of the\nPerformance of Packet-level FEC Techniques in VoIP\nCommunications\", WSEAS Transactions on Communications, issue 9,\nVol. 8, 2009."]}

Background The rapidly improving tools of genetic engineering may make it possible to overcome the humoral immune barrier that prevents xenotransplantation. We hypothesize that levels of human antibody binding to donor tissues from swine must approximate the antibody binding occurring in allotransplantation. It is uncertain if this is an attainable goal. Here we perform an initial analysis of this issue by comparing human antibody binding to red blood cells (RBC) isolated from knockout swine and to allogeneic or autologous human RBC. Methods Human sera were incubated with RBC isolated from various genetically engineered swine or from humans. The level of IgG and IgM binding to these cells were compared using either flow cytometry or a novel mass spectrometric assay. Results Mass spectroscopic quantitation of human antibody binding demonstrated that as few as 3 gene inactivations can reduce the levels human antibody binding to swine RBC that is as low as autologous human RBC. Flow cytometry showed that RBC from 2-gene knockout swine exhibited less human antibody binding than human blood group O allogeneic RBC in 22% of tested sera. Deletion of a third gene from pigs resulted in 30% of human samples having less IgG and IgM RBC xenoreactivity than alloreactivity. Conclusions Xenoantigenicity of swine RBC can be eliminated via gene disruption. These results suggest that the gene knockout approach may be able reduce antigenicity in other pig tissues to levels that enable the xenotransplantation humoral barrier to be overcome.

Burlak C, Wang ZY, Chihara RK, Lutz AJ, Wang Y, Estrada JL, Tector AJ. Identification of human preformed antibody targets in GTKO pigs. Xenotransplantation 2012; 19: 92–101. © 2012 John Wiley & Sons A/S. Abstract: Background: Human preformed antibodies continue to recognize porcine xenografts, despite the advent of α-galactosyltransferase knockout (GTKO) pigs. This study examined the potential reactivity of human preformed IgG and IgM antibodies toward antigens in the GTKO pig liver. Methods: Human serum was analyzed for the concentration of IgG, IgM, anti-αgal antibody, anti-non-αgal antibody and cytotoxicity toward domestic and GTKO fibroblasts and liver sinusoidal endothelial cells (LSEC). We detected preformed antibodies in human serum directed toward GTKO pig liver cells and tissue samples using advanced proteomic techniques. The targets of preformed antibodies were identified by MALDI TOF TOF mass spectrometry and validated by confocal microscopy, immunoblot, and immunoprecipitation. Results: Human serum used in this study contained 2.06 μg/ml IgG and 0.013 μg/ml IgM directed toward GTKO fibroblasts. Human IgG and IgM bound to GTKO LSEC in a dose-dependent manner and were cytotoxic. We detected 357 protein spots recognized by human IgG and 233 by human IgM. Two hundred and nineteen proteins were common to both human IgG and IgM. Mass spectrometry identified numerous immunoreactive proteins, of which 19 were membrane proteins on liver cells. The most significant to this study were α-enolase, CFTR, and E-cadherin, which were abundant in GTKO pig tissues and expressed on the surface of GTKO LSEC. Human IgG captured α-enolase, CFTR, and E-cadherin by immunoprecipitation validating the proteomic identification. Conclusion: These experiments indicate that several membrane antigens in GTKO pigs could be recognized directly by human IgG or IgM. Further studies on the contribution of these antigens to antibody-mediated xenograft rejection are necessary.

Protein levels and specifically casein are important in the industrial processes of milk, especially in clotting and clot formation. Characterization of genotype frequencies is important to determine the genetic value of the animals. This experiment was carried out to determine the genotype frequencies and to establish the correlation between haplotypes and production phenotypes. Milk and blood samples were taken from 54 first parity Holstein heifers from farms in the Sabana de Bogotá (2000-3000 meters above sea level;12-18°C). Milk samples were analyzed for protein, casein, fat and lactose percent, whereas the blood was used to genotype the animals for the k-casein gene (AA, AB and BB). Also, data was taken from the Dairy Herd Improvement Program of the Holstein Association of Colombia for the 305 day 2X milk production. The data were used to determine the allelic and genotypic frequency of the k-casein gene and the correlation between the haplotypes and protein content, casein percent and 305 day 2X milk production projection. The genotype frequency was 57.41% for haplotype AA, 37.04% for haplotype AB and 5.55% for haplotype BB. The allelic frequency was 75.93% for allele A and 24.07% for allele B. There was a high significant correlation between the haplotypes and the protein percent in the milk (R>.975) being the BB haplotype superior to the AB and AA haplotypes. On the other hand, there was no significant correlation between the haplotypes and the casein percent nor the 305 day 2x milk production projection, even though the BB haplotype always showed higher valúes than the others.

We describe the present status of an ongoing project to develop a Tandem-ElectroStatic-Quadrupole (TESQ) accelerator facility for Accelerator-Based (AB)-BNCT. The project final goal is a machine capable of delivering 30 mA of 2.4 MeV protons to be used in conjunction with a neutron production target based on the (7)Li(p,n)(7)Be reaction. The machine currently being constructed is a folded TESQ with a high-voltage terminal at 0.6 MV. We report here on the progress achieved in a number of different areas.

BACKGROUND Hepatic failure has been treated successfully with clinical extracorporeal perfusions of porcine livers. However, dog-to-pig and pig-to-baboon liver xenotransplant models have resulted in severe bleeding secondary to liver xenograft-induced thrombocytopenia. Kupffer cells (KC) are abundant phagocytic cells in the liver. KC express the CD11b/CD18 receptor, which has been implicated in chilled platelet binding and phagocytosis through interaction with platelet surface proteins and carbohydrates. We sought to identify the role of KC CD18 in liver xenograft-induced thrombocytopenia. METHODS Primary pig KC were characterized by flow cytometry, immunoblots, and quantitative polymerase chain reaction. Pig KC were used in inhibition assays with fluorescently labeled human platelets. The CD18 receptor was targeted for siRNA knockdown. RESULTS Domestic and α1,3-galactosyltransferase double knockout porcine KC cultures were approximately 92% positive for CD18 as detected by quantitative polymerase chain reaction and flow cytometry. Use of CD18 blocking antibodies resulted in reduction of human platelet binding and phagocytosis. Additionally, asialofetuin, not fetuin, inhibited platelet phagocytosis suggesting the involvement of an oligosaccharide-binding site. Furthermore, reduced CD18 expression by siRNA resulted in decreased human platelet binding. CONCLUSIONS Our data suggest that primary pig KC bind and phagocytose human platelets with involvement of CD18. Further understanding and modification of CD18 expression in pigs may result in a liver xenograft with reduced thrombocytopenic effects, which could be used as a bridge to allogeneic liver transplantation.

Few studies have tested a broad range of theory-based determinants of injection-related risk and protective behaviors. The current study hypothesize relationships among attitudinal beliefs, normative influences, perceived self-efficacy, intentions, and performing these behaviors consistent with the theory of planned behavior and related models. The conceptual model was tested in a cross-sectional sample of 895 drug injectors recruited from street settings in the southwestern United States and who completed a face-to-face interview. Structural equation modeling results showed intention and self-efficacy most consistently predicted needle sharing and bleaching considering all other model variables. Discussion focuses on the value of using these theoretical concepts to examine needle sharing and other behaviors among injection drug users and their relevance for developing HIV-reduction interventions.

SUMMARY Purpose: Epilepsy is highly prevalent in developing countries like Honduras, with few studies evaluating this finding. This population-based study evaluated the impact of an 8-year public health and educational intervention program in reducing symptomatic epilepsies in rural Salama ´, Honduras. Methods: We used the capture and recapture method including review of charts, previous databases, key informants from the community, and a second house-to-house survey for epilepsy. Epilepsy incidence and prevalence day after the interventions was May 5, 2005. Residents with active epilepsy with onset after May 1997 were offered neurologic evaluation, electroencephalography, and brain tomography. New data over 8 years were compared to preintervention data from the initial baseline 1997 study utilizing prevalence ratios and confidence intervals. Other calculations utilized chi square or Fisher’s exact tests. Key Findings: Thirty-three of 36 patients with onset of active epilepsy after 1997 accepted evaluations to determine etiology. Symptomatic etiology was found in 58.3%. Neurocysticercosis (NCC) was again the most frequent cause (13.9%), followed by perinatal insults (11.1%). Epilepsy secondary to NCC was significantly reduced from 36.9% in 1997 (p = 0.02). The incidence (35.7/100,000) and prevalence (11.8/1,000) of active epilepsy were not significantly reduced when compared to the incidence (92.7/ 100,000) and prevalence (15.4/1,000) of active epilepsy in 1997. Significance: Our cohort appears to indicate that health and educational community interventions can reduce preventable epilepsy from NCC in a hyperendemic population in a low-resource, developing country. Plans are underway for the Honduran Government to institute this rural model countrywide.

Genetic manipulation of porcine genome to produce genetically modified pigs with high efficiency has been hampered by the unavailability of an ideal cell type. The cell type currently used for various genetic manipulations is fetal fibroblasts. These cells have very limited life span in culture, and efficiency of gene targeting is very low. In this study, we developed a simple but novel strategy to derive cell lines from adult porcine liver and adipose tissues with long life span. Small colonies with few cells became visible as early as 2 to 3 days on collagen-coated plates, and a full-grown colony took 10 to 14 days to form. These cells maintained a steady growth up to 80 population doublings with normal karyotype. Transfection of these cells with a plasmid containing a neomycin resistance gene and selected under G418 yielded clones with stable genetic modifications and extended expression of the transgene. Further, these cells were used as nuclear donors to produce somatic cell nuclear transfer (SCNT) embryos. The average fusion rates were 86.8, 80.5, and 90.4% for liver-derived cell lines (LDCs), fat-derived cell lines (FDCs), and fetal fibroblasts (FFs), respectively. We achieved a pregnancy rate of 50% with both LDCs and FDCs at day 30 and the efficiencies of generating fetuses from cloned embryos were 3.5, 2.1, and 4.0% for LDCs, FDCs and FFs, respectively.

This study was undertaken to characterize the effects of constitutive expression of the hedgehog transcriptional activator, Gli2, in porcine skin. The keratinocyte-specific human transgene, K5-hGli2 Delta N, was used to produce transgenic porcine lines via somatic cell nuclear transfer techniques. In mice, K5-hGli2 Delta N induces epithelial downgrowths resembling basal cell carcinomas. Our porcine model also developed these basal cell carcinoma-like lesions, however gross tumor development was not appreciated. In contrast to the murine model, diffuse epidermal changes as well as susceptibility to cutaneous infections were seen in the swine model. Histologic analysis of transgenic piglets revealed generalized epidermal changes including: epidermal hyperplasia (acanthosis), elongated rete ridges, parakeratotic hyperkeratosis, epidermal neutrophilic infiltration, capillary loop dilation and hypogranulosis. By 2 weeks of age, the transgenic piglets developed erythematic and edematous lesions at high contact epidermal areas and extensor surfaces of distal limb joints. Despite antibiotic treatment, these lesions progressed to a deep bacterial pyoderma and pigs died or were euthanized within weeks of birth. Non-transgenic littermates were phenotypically normal by gross and histological analysis. In summary, constitutive expression of the human hGli2 Delta N in keratinocytes, results in cutaneous changes that have not been reported in the K5-hGli2 Delta N murine model. These findings indicate a need for a multiple species animal model approach in order to better understand the role of Gli2 in mammalian skin.

OBJECTIVE Insulin resistance is greater in racial/ethnic minorities than in non-Hispanic whites (NHWs) for those with and without type 2 diabetes. Because previous research on insulin resistance in type 1 diabetes was limited to NHWs, racial/ethnic variation in an estimated measure of insulin resistance in type 1 diabetes was determined. RESEARCH DESIGN AND METHODS The sample included 79 individuals with type 1 diabetes diagnosed at age RESULTS Mean current age was 13.5 years (range 3.2–32.5) and diabetes duration was 5.7 years (0.1–19.9). eGDR was inversely associated with age. Compared with that in NHWs, age-adjusted eGDR was significantly lower among nonwhites (NHB, other/mixed, and Hispanic: Δ = −1.83, P = 0.0006). Age-adjusted eGDR was negatively associated with body fat, triglycerides, urinary albumin/creatinine, acanthosis nigricans, parental obesity, and parental insulin resistance and positively related to HDL and sex hormone–binding globulin. In multivariable analysis, lower eGDR was significantly associated with older age, nonwhite race/ethnicity, acanthosis, and lower HDL. CONCLUSIONS Minorities with type 1 diabetes are significantly more insulin resistant, as measured by eGDR, than NHWs. Exploring potential mechanisms, including disparities in care and/or physiological variation, may contribute to preventing racial/ethnic differences in insulin resistance–associated outcomes.

In this work we describe the present status of an ongoing project to develop a tandem-electrostatic-quadrupole (TESQ) accelerator facility for accelerator-based (AB) BNCT at the Atomic Energy Commission of Argentina in Buenos Aires. The project final goal is a machine capable of delivering 30 mA of 2.4 MeV protons to be used in conjunction with a neutron production target based on the (7)Li(p,n)(7)Be reaction slightly beyond its resonance at 2.25 MeV. These are the specifications needed to produce sufficiently intense and clean epithermal neutron beams, based on the (7)Li(p,n)(7)Be reaction, to perform BNCT treatment for deep-seated tumors in less than an hour. An electrostatic machine is the technologically simplest and cheapest solution for optimized AB-BNCT. The machine being designed and constructed is a folded TESQ with a high-voltage terminal at 1.2 MV intended to work in air. Such a machine is conceptually shown to be capable of transporting and accelerating a 30 mA proton beam to 2.4 MeV. The general geometric layout, its associated electrostatic fields, and the acceleration tube are simulated using a 3D finite element procedure. The design and construction of the ESQ modules is discussed and their electrostatic fields are investigated. Beam transport calculations through the accelerator are briefly mentioned. Likewise, work related to neutron production targets, strippers, beam shaping assembly and patient treatment room is briefly described.

In this study, two enucleation methods, the squeezing and the aspiration methods, were compared. The efficiency of these two methods to enucleate pig oocytes and the in vitro and in vivo viability of somatic cell nuclear transfer (SCNT) pig embryos, were evaluated. In the squeezing method, the zona pellucida was partially dissected and a small amount of cytoplasm containing metaphase II (MII) chromosomes and the first polar body (PB) were pushed out. In the aspiration method, the PB and MII chromosomes were aspirated using a beveled micropipette. After injection of fetal fibroblasts into the perivitelline space, reconstructed oocytes were fused and activated electrically, and then cultured in vitro for 6 days or transferred to surrogates. The squeezing method resulted in a higher proportion of degenerated oocytes than the aspiration method (14% vs. 5%). The squeezing method took longer to enucleate 100 oocytes (306 minutes) than the aspirating method (113 minutes). Fusion rate (72-78%) and cleavage rate (67%) were not influenced by the enucleation method but blastocyst formation was improved (P < 0.05) in oocytes enucleated by the aspiration method (5 vs. 9%). When SCNT embryos were transferred to recipients, pregnancy rates to term were similar (27%, 3/11 and 27%, 3/11) in both methods with the birth of 10 piglets/3 litters and 16 piglets/3 litters in the squeezing and the aspiration methods, respectively. Our results indicate that the aspiration method for oocyte enucleation is more efficient than the squeezing method in producing a large number of pig SCNT embryos with normal in vivo viability.

Prediction of IVF outcome on the first days of ovarian stimulation has focused clinical research for many years. The aim of this work is to predict the probability of pregnancy on the fourth day of ovarian stimulation for IVF cycle, using parameters usually determined in this stage--estradiol, antral follicle count--together with parameters determined previously: FSH on the third day of cycle and women age.One hundred and ten patients with primary infertility due to a tubal factor were recruited to participate in a prospective study. FSH was determined on the third day of spontaneous cycle. Antral follicles and estradiol were measured on the fourth day of ovarian stimulation. After oocyte pick-up, quality and quantity of oocytes and embryos and pregnancy rates were assessed.In stepwise multiple logistic regression the variables with better predictiveness over pregnancy are: antral follicles count, estradiol and woman age. The logistic regression analyses demonstrate that the capacity of the model that uses these variables to predict pregnancy is 75%, with a positive predictive value of 69% and a negative predictive value of 80%.On the fourth day of ovarian stimulation of IVF cycles, the variables with highest predictiveness are: antral follicle count, estradiol and women age. When these variables are included in a model of prediction, the capacity to predict pregnancy is 75%.

While somatic cell nuclear transfer (SCNT) has been successful in several species, many pregnancies are lost and anomalies are found in fetal and perinatal stages. In this study SCNT and artificial inseminations (AI) populations were compared for litter size, average birth weight, piglets alive at birth, stillborn, mummies, dead at the first week, intrauterine growth restriction (IUGR) and large for gestational age (LGA). Twenty-three SCNT litters (143 individuals) were compared to 112 AI litters (1300 individuals). Litter size average was 11.5 for AI and 6.2 for SCNT. Litter weight and average birth weight adjusted by litter size were significantly (p < 0.05) higher in AI than in SCNT litters. The SCNT population had a significant (p < 0.01) increase in the number of IUGRs per litter with LSmeans 7.2 +/- 1.4 versus 19.4 +/- 3.5 and means 8.0 +/- 10.8 versus 15.5 +/- 24.5 for AI and SCNT, respectively. Additionally, there was a trend for higher postnatal mortality and stillbirths in the SCNT population. These findings demonstrate that there are some differences between SCNT-derived and AI litters. SCNT-derived pigs are excellent models to study epigenetic factors and genes involved in IUGRs, and to develop effective means to improve fetal growth in humans and animals.

This longitudinal study with male, predominantly Latino, injection drug users (IDUs) was undertaken to test a model of determinants of unprotected sex guided by the theory of planned behavior (TPB). Structural equation modeling (SEM) was used to test each theory-based predictor and the overall model's efficacy in predicting current and escalation of unprotected sex in a community sample (n = 371). The hypothesized model fit the data well—with intention and self-efficacy directly predicting risk behavior. In the observed model, the theory-based factors were significantly related to T1 and T2 unprotected sex outcomes, though the effects on the latter were mediated by T1 unprotected sex. The findings are generally congruent with postulates of the theory of planned behavior; however, greater predictive efficacy of future behavior may be achieved by considering psychosocial and other factors related to Latino male IDUs' sexual behaviors.

Polo-like kinase 1 (PLK1) is a serine/threonine-protein kinase expressed during mitosis and overexpressed in multiple human cancers, including leukemia and also many solid tumors. PLK1 knockdown has been shown to block proliferation of leukemic cell lines and the clonogenic potential of tumor cells grown from patients with cancer. PLK1 inhibition is a promising strategy for the treatment of some tumors. We aim to analyze expression of PLK1 in metastatic colorectal carcinoma. Retrospective analysis of colorectal carcinomas with hepatic metastasis during follow-up receiving neoadjuvant chemotherapy (NAC), based on oxaliplatin. Immunohistochemistry for PLK-1 in paraffin-embedded tissue from the primary and also from the metastasis. 50 patients. 32% showed good histopathological response. 43% of the primaries were positive for PLK1, as opposed to 23.5% of the metastasis. Expression of PLK1 was significantly reduced in metastasis compared with the primaries (p = 0.05), what could be due to therapy or to a phenotypic change of the metastatic nodule. Analysis of the prognostic influence of PLK1 expression showed significant association between PLK1 expression in metastasis and lower overall survival (p = 0.000). We have also found a significant association between PLK1 expression and histopathological response (p = 0.02). All the tumors with high expression of PLK1 showed minor response (11/11). This study shows the association between survival and poor histopathological response to therapy and high expression of PLK1 in metastasis. Our results could open a new therapeutic approach through the inhibition of PLK1.

The observed intergenerational stress response to negative social and historical events is at the core of historical trauma theory, which has been applied to Native Americans, African Americans, and Pacific Islanders, among others. The historical and social experiences of the Mexican population living in the United States have many parallels that lend themselves to the application of historical trauma theory to macro-level and micro-level influences on access to health care, physical health status, and mental health status, including substance abuse among Mexican Americans. This article highlights the legacy of Spanish colonialism and Anglo-American neo-colonialism on Mexicans and Mexican Americans in the southwestern United States through a potential application of historical trauma theory.

Introduction Therapy of metastatic colorectal carcinoma has greatly evolved in recent years. Surgery is still the best curative option and can improve survival in stage IV disease. Neoadjuvant chemotherapy (NAC) has emerged as a widely used therapeutic option before surgery. Pathologists have developed several systems to grade response, mainly adapting the grading systems used for the response in primary esophageal or rectal tumors. There are many reports confirming the prognostic utility of these grading systems. However, there have been fewer references to the potential significance of the pattern of histological response. The objective of the present study is to describe the histopathological lesions found in the tumor bed after NAC and their potential significance in terms of prognosis. Material and methods We reviewed the files of patients with colorectal carcinoma that developed hepatic metastasis during follow-up and received NAC before surgical resection of metastasis. We gathered demographic, analytical and morphological data of the cases, and also reviewed the hepatic resection samples to measure the pathological response to chemotherapy according to Blazer's criteria, and to define the predominant patterns of response (mucin pools, fibrosis or necrosis). We also determined the presence of satellitosis, measured the thickness of the tumor-normal interface (TNI) as proposed by Maru et al., and searched for vascular and bile duct invasion. All these pieces of information were collected in an Excel database and analyzed with SPSS 20.0 for Windows statistical package. The outcome measures were disease-free survival and overall survival in months since the first surgery to resect metastatic disease. Results Fifty patients fulfilled the inclusion criteria for the present study. All of them had received a chemotherapeutic regimen mainly based on platinum, associated or not with targeted drugs (18% received anti-EGFR drugs and 24% anti-VEGFR drugs). Of the primaries, 66% were of sigmoid-rectal origin, and 32% of the cases showed a major histopathological response to therapy (including 3 cases with a complete response). In 76% of the tumors, the predominant histological pattern was necrosis, followed by fibrosis (57.4%). Mucin pools were the predominant feature in 23.4% of the tumors. We found satellitosis (microscopic tumor nodules separated by more than 1 mm from the principal tumor) in 53.2% of the cases. A prominent inflammatory reaction was found in 19% of the cases, and it was mainly composed of lymphocytes and hystiocytes (70% of the cases). Vessel invasion was seen in 30% of the cases, and perineural invasion was only found in 4%. We found no case of bile duct invasion by the tumor. The thickness of the TNI measured less than 2.5 mm in 60% of the present series. Statistical analysis of the series revealed that thickness of the tumor-liver interface was significantly associated with recurrence and overall survival. We found a significant association between response and thickness of the tumor-normal liver interface. In our series, the presence of satellitosis tended to predict a shorter DFS. The comparison of Kaplan–Meier curves with the log-rank test showed a significant association between overall survival and the presence of mucin pools and fibrosis in the tumor bed. The other histopathological factors did not predict differences in prognosis. These differences were independent of the use of targeted drugs. Discussion The pathological reports of hepatic metastasis from colorectal carcinoma resected after NAC usually indicate only the number, the size and the response of the tumor cells to therapy, apart from the distance to the resection margin of the specimen. Few reports have analyzed the possible prognostic significance of the different kinds of histopathological responses. The results of the present study indicate that those tumors with extensive pools of mucin show a significantly worse prognosis as compared to tumors with less mucin secretion. Fibrosis indicates a better prognosis, except when desmoplasia is present. Our study further supports the prognostic significance of the thickness of the tumor-hepatic interface. We conclude that pathology reports should specify the kind of histopathological response to therapy, besides grading it, because this might add significant prognostic information.