International Intraductal Papillary Mucinous Neoplasms Registry: Long-Term Results Based on the New Guidelines.

OBJECTIVE: The aim of this study was to analyze the outcomes of a long-term intraductal papillary mucinous neoplasm (IPMN) registry and evaluate new guidelines. METHODS: A prospectively maintained IPMN registry involving 6 centers in Europe and the United States was used to collect the data. Patients with more than 1-year follow-up and no malignancy diagnosed within the first 3 months of surveillance were included. RESULTS: From 1999 to 2014, 620 patients were included. The median follow-up time was 3 years. Thirty-seven (6%) patients developed malignancy with a median time from IPMN diagnosis to malignancy of 10.3 months. The 1-, 5-, and 10-year actuarial rates of disease-free survival were 97%, 93%, and 92% respectively. Four hundred thirty-one patients met criteria for low-risk branch duct IPMN consisting of cyst size less than 3 cm, with no solid component or main duct dilation. Eight malignancies were diagnosed in this subgroup, all of them within the first 5 years. From this subcohort, 112 patients had a follow-up time of more than 5 years, and no malignancy was diagnosed. CONCLUSIONS: In IPMN lesions with low-risk features at baseline, the risk of progression to malignancy after the first 5 years of follow-up was minimal. Furthermore, the main cyst characteristics remained unchanged during their surveillance.

Nitric oxide has a regulatory effect in the germination of conidia of Colletotrichum coccodes.

Nitric oxide (NO) was first detected in mammals and has since been found in plants and in micro-organisms such as bacteria. NO is an important signalling molecule involved in a number of critical signal transduction pathways. To date, NO has not been directly detected in fungi, and little research on NO and fungi has been completed. Here, the role of NO in the germination of Colletotrichum coccodes conidia was investigated. Conidia were germinated on microscope slides, treated with chemicals to block NO, to add NO, and/or to detect NO, and assessed for their stage of development over 24 h. NO was detected in germinating conidia at all stages of development. Exogenous NO delayed germination, while treatment with NO inhibitors accelerated germination, suggesting NO may have a regulatory effect in germination. The differential effect of the various inhibitors suggests the fungal isoform of nitric oxide synthase (NOS) may be biochemically similar to mammalian constitutive NOS.

The role of the jasmonate response in plant susceptibility to diverse pathogens with a range of lifestyles.

Plants defend themselves against attack from insects and pathogens with various resistance strategies. The jasmonate and salicylate signaling pathways are two induced responses that protect plants against these attackers. Knowledge of the range of organisms that are affected by each response is important for understanding how plants coordinate their defenses against multiple attackers and the generality of effect of different resistance mechanisms. The jasmonate response is known to protect plants against a wide range of insect herbivores; in this study, we examined the role of the jasmonate response in susceptibility to eight pathogens with diverse lifestyles in the laboratory and field. Recent biochemical models suggest that the lifestyle of the pathogen (necrotroph versus biotroph) should predict whether the jasmonate response will be involved in resistance. We tested this by examining the susceptibility of wild-type (cv Castlemart with no known genes for resistance to the pathogens used) and jasmonate-deficient mutant tomato (Lycopersicon esculentum) plants (def1) and by employing rescue treatments of the mutant. Plant susceptibility to five of the eight pathogens we examined was reduced by the jasmonate response, including two bacteria (Pseudomonas syringae and Xanthomonas campestris), two fungi (Verticillium dahliae and Fusarium oxysporum f. sp. lycopersici), and an oomycete (Phytophthora infestans). Susceptibility to three fungi was unaffected (Cladosporium fulvum, Oidium neolycopersici, and Septoria lycopersici). Our results indicate that the jasmonate response reduces damage by a wide range of pathogens from different lifestyles, a result that contrasts with the emerging picture of diseases on Arabidopsis. Thus, the generality of jasmonate-based resistance of tomato challenges the view that ecologically distinct plant parasites are resisted via different mechanisms.

[Etiological factors of fungemia in the Hospital San Martín in La Plata]. / Agentes etiológicos de fungemia en el Hospital San Martín de La Plata.

To determine the distribution of etiologic agents of fungemia in San Martin Hospital, La Plata, we retrospectively studied 81 consecutive episodes of fungemia, diagnosed in 46 adults and 35 preterm newborn (PNB) hospitalized from November 1998 to August 2001. The diagnosis was achieved by blood culture obtained by venipuncture and by catheter aspiration and was processed using BactAlert and lysis-centrifugation technique. Isolated yeasts were identified employing API 32C system and additional tests. Candida parapsilosis (28.4%), C. albicans (25.9%) and C. tropicalis (25.9%) were predominant as etiological agents (80%). Other species of Candida (C. pelliculosa, C. kefyr and C. guillermondii), Malassezia pachydermatis, Cryptococcus neoformans and Histoplasma capsulatum were recovered in low percentage (each one < or = 7%). C. parapsilosis was predominant as causative agent among PNB male (47.4%), C. albicans among adult women (41.7%) and C. tropicalis among adult men (32.3%). The species of Candida (C. parapsilosis, C. tropicalis and C. albicans) were predominant as etiologic agents of fungemia, with a different distribution in the episodes which occurred in adults and PNB patients, and also according to gender in both groups.

Agentes etiológicos de fungemia en el Hospital San Martín de La Plata / Distribution of the etiological agents of fungemia in the San Martin Hospital of La Plata

Para determinar la distribución de los agentes causales de fungemia en el Hospital Interzonal General de Agudos San Martín, de La Plata, Argentina, se estudió retrospectivamente la etiología de 81 episodios consecutivos ocurridos en 46 adultos y 35 recién naciddos pre-término (RNPT) internados entre noviembre de 1998 y agosto de 2001. El diagnóstico se hizo a partir de cultivos de sangre obtenida por punción venosa y/o a través de catéter, procesados con el equipo BactAlert y la técnica de lisis-centrifugación. La identificación de los aislamientos fue realizada con el equipo API 32C para levaduras y pruebas adicionales. Candida parapsilosis (28,4 por ciento), C.albicans (25,9 por ciento) y C.tropoicalis (25,9 por ciento) predominaron como agentes causales (en conjunto 80 por ciento). Otras especies de Candida (C.pelliculosa, C.kefyr y C.guillermondii), Malassezia pachydermatis, Cryptococcus neoformans e Histoplasma capsulatum se recuperaron en menor porcentaje (individualmente <_7 por ciento). C.parapsilosis predominó como agente causal de fungemia en RNPT varones (47,4 por ciento), C albicans en mujeres adultas (41,7 por ciento) y C.tropicalis en varones adultos (32,3 por ciento). Las especies de Candida (C.parapsilosis, C.tropicalis y C.albicans) predominaron como agentes causales de fungemia en nuestro hospital, con una distribución diferente en los episodios ocurridos en los pacientes adultos y RNPT y entre los varones y mujeres de ambos grupos.

Agentes etiológicos de fungemia en el Hospital San Martín de La Plata / Distribution of the etiological agents of fungemia in the San Martin Hospital of La Plata

Para determinar la distribución de los agentes causales de fungemia en el Hospital Interzonal General de Agudos San Martín, de La Plata, Argentina, se estudió retrospectivamente la etiología de 81 episodios consecutivos ocurridos en 46 adultos y 35 recién naciddos pre-término (RNPT) internados entre noviembre de 1998 y agosto de 2001. El diagnóstico se hizo a partir de cultivos de sangre obtenida por punción venosa y/o a través de catéter, procesados con el equipo BactAlert y la técnica de lisis-centrifugación. La identificación de los aislamientos fue realizada con el equipo API 32C para levaduras y pruebas adicionales. Candida parapsilosis (28,4 por ciento), C.albicans (25,9 por ciento) y C.tropoicalis (25,9 por ciento) predominaron como agentes causales (en conjunto 80 por ciento). Otras especies de Candida (C.pelliculosa, C.kefyr y C.guillermondii), Malassezia pachydermatis, Cryptococcus neoformans e Histoplasma capsulatum se recuperaron en menor porcentaje (individualmente <_7 por ciento). C.parapsilosis predominó como agente causal de fungemia en RNPT varones (47,4 por ciento), C albicans en mujeres adultas (41,7 por ciento) y C.tropicalis en varones adultos (32,3 por ciento). Las especies de Candida (C.parapsilosis, C.tropicalis y C.albicans) predominaron como agentes causales de fungemia en nuestro hospital, con una distribución diferente en los episodios ocurridos en los pacientes adultos y RNPT y entre los varones y mujeres de ambos grupos. (AU)

Agentes etiológicos de fungemia en el Hospital San Martín de La Plata. / [Etiological factors of fungemia in the Hospital San Martín in La Plata]

To determine the distribution of etiologic agents of fungemia in San Martin Hospital, La Plata, we retrospectively studied 81 consecutive episodes of fungemia, diagnosed in 46 adults and 35 preterm newborn (PNB) hospitalized from November 1998 to August 2001. The diagnosis was achieved by blood culture obtained by venipuncture and by catheter aspiration and was processed using BactAlert and lysis-centrifugation technique. Isolated yeasts were identified employing API 32C system and additional tests. Candida parapsilosis (28.4

), C. albicans (25.9

) and C. tropicalis (25.9

) were predominant as etiological agents (80

). Other species of Candida (C. pelliculosa, C. kefyr and C. guillermondii), Malassezia pachydermatis, Cryptococcus neoformans and Histoplasma capsulatum were recovered in low percentage (each one < or = 7

). C. parapsilosis was predominant as causative agent among PNB male (47.4

), C. albicans among adult women (41.7

) and C. tropicalis among adult men (32.3

). The species of Candida (C. parapsilosis, C. tropicalis and C. albicans) were predominant as etiologic agents of fungemia, with a different distribution in the episodes which occurred in adults and PNB patients, and also according to gender in both groups.

Mitochondrial alternative oxidase is not a critical component of plant viral resistance but may play a role in the hypersensitive response.

Transgenic tobacco (Nicotiana tabacum) with altered levels of mitochondrial alternative oxidase (AOX) were used to examine the potential role of this electron transport chain protein in resistance to tobacco mosaic virus. We examined the effect of AOX expression on the salicylic acid-induced resistance in susceptible plants and the resistance responses of plants harboring the N-gene. A lack of AOX did not compromise the ability of salicylic acid treatment to heighten the resistance of susceptible plants. In plants with the N-gene, a lack of AOX did not compromise the ability of the hypersensitive response to restrict the virus or the ability of the plant to develop systemic acquired resistance. Overexpression of AOX did not heighten the resistance of susceptible plants, but did result in smaller hypersensitive response lesions, suggesting a link between mitochondrial function and this programmed cell death event. We conclude that AOX is not a critical component of the previously characterized salicylhydroxamic acid-sensitive pathway important in viral resistance.

H2O2 plays different roles in determining penetration failure in three diverse plant-fungal interactions.

Fungal plant pathogens that attempt to penetrate and feed on living cells frequently trigger a localized plant defence response that results in fungal penetration failure. In the current study we demonstrate that breakdown products of the cell wall released by the localized application of hemicellulase elicit localized responses including, sequentially, extracellular H2O2 generation; accumulation of phenolic compounds; and cross-linking of proteins in the cell wall. In a detailed time-course study of three plant-fungus interactions that result in a high frequency of penetration failure, only one plant-fungus combination displayed a similar profile of responses to that induced by localized cell-wall degradation. The additional generation of extracellular O2- in one interaction, and the absence of phenolic compounds in the cell wall in another, demonstrate that plant responses to the penetration process may be influenced by activities of the penetrating fungus. Significantly, H2O2 generation was the only response detected in all three plant-fungal combinations at the correct time and place to account for penetration failure, and in all three combinations the enzymatic removal of H2O2 resulted in increased penetration success. Pharmacological studies suggest that in two of the three interactions, H2O2 generation required cytoskeletal involvement but was independent of transcription or translation, although inhibition of the latter processes increased fungal penetration. In at least one of these two interactions, the data suggest that H2O2 generation and new gene expression act within the same penetration-inhibiting pathway, possibly through the involvement of phenolic materials. However, enzymatic removal of H2O2 from the third interaction almost completely eliminated penetration failure, while interference with cytoplasmic processes had no effect, suggesting that H2O2 generation in this system did not require protoplast involvement and, alone, was necessary and sufficient to account for fungal penetration failure.

Mutations in WSC genes for putative stress receptors result in sensitivity to multiple stress conditions and impairment of Rlm1-dependent gene expression in Saccharomyces cerevisiae.

Intracellular signaling by mitogen-activated protein (MAP) kinase cascades plays an essential role in the cellular response to environmental stress. In the yeast Saccharomyces cerevisiae, the PKC1-regulated, stress-activated MAP kinase pathway, the MPK1 cascade, is activated by heat and by a decrease in osmolarity. The genes WSC1, WSC2 and WSC3 encode putative receptors that maintain cell wall integrity under conditions of heat stress. Genetic studies place the function of the WSC genes upstream of the MPK1 kinase cascade. To further define the role of the WSC family in the stress response we determined whether: (1) the wscdelta mutants are sensitive to other environmental stress conditions, in addition to heat shock; (2) expression from four transcriptional control elements, known to be activated by stress, is impaired in wscdelta mutants; and (3) Wsc4, a Wsc homolog, has functions that overlap with those of the other Wsc family members. We report here that deletion of WSC and PKC1 causes hypersensitivity to ethanol, hydrogen peroxide and DNA-damaging drugs. In wscdelta mutants expression of beta-galactosidase from the AP-1 response element (ARE), the heat shock response element (HSE) or the stress response element (STRE) is not reduced. In contrast, expression of a reporter gene placed under the control of the Rlm1 (transcription factor)-dependent response element is significantly reduced in wscdelta mutants. This suggests that the lysis defect of wscdelta mutants is at least in part caused by a defect in transcriptional regulation by Rlm1. Phenotypic analysis of the effect of deleting WSC4 in a wsc1delta mutant show that, unlike WSC2 or WSC3, deletion of WSC4 does not exacerbate the lysis defect of a wsc1delta strain. In contrast, deletion of WSC4 enhances the sensitivity of the wsc1delta mutant to heat shock, ethanol, and a DNA-damaging drug, suggesting that WSC4 plays a role in the response to environmental stress but that its function may differ from those of the other WSC family members.

6-hydroxydopamine-induced nuclear factor-kappa B activation in PC12 cells.

The involvement of nuclear Factor-kappa B (NF-kappa B) transcription factor in PC12 cell death triggered by the dopaminergic neurotoxin 6-hydroxydopamine (6-OHDA) was investigated. Results show that oxidative stress generated by 6-OHDA activates NF-kappa B. When the NF-kappa B activation was inhibited by parthenolide, PC12 cell death induced by 6-OHDA was significantly increased, thus suggesting an involvement of this transcription factor in a protective mechanism against 6-OHDA toxicity. To further assess this hypothesis, we studied the involvement of NF-kappa B in the protective effect of two anti-apoptotic genes, bcl-2 and bfl-1. Although Bcl-2 and Bfl-1 expression normally protects PC12 cells from 6-OHDA, parthenolide strongly decreased the beneficial effects afforded by transgene expression. These results suggest: (1) that the transcription factor NF-kappa B is likely associated with the protection of catecholaminergic PC12 cells and (2) that the protective effects afforded by bcl-2 and bfl-1 expression may be dependent on NF-kappa activation.

Molecular pathways involved in the neurotoxicity of 6-OHDA, dopamine and MPTP: contribution to the apoptotic theory in Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by a preferential loss of the dopaminergic neurons of the substantia nigra pars compacta. Although the etiology of PD is unknown, major biochemical processes such as oxidative stress and mitochondrial inhibition are largely described. However, despite these findings, the actual therapeutics are essentially symptomatical and are not able to block the degenerative process. Recent histological studies performed on brains from PD patients suggest that nigral cell death could be apoptotic. However, since post-mortem studies do not allow precise determination of the sequence of events leading to this apoptotic cell death, the molecular pathways involved in this process have been essentially studied on experimental models reproducing the human disease. These latter are created by using neurotoxic compounds such as 6-hydroxydopamine (6-OHDA), 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or dopamine (DA). Extensive study of these models have shown that they mimick, in vitro and in vivo, the histological and/or the biochemical characteristics of PD and thus help to define important cellular actors of cell death presumably critical for the nigral degeneration. This review reports recent data concerning the biochemical and molecular apoptotic mechanisms underlying the experimental models of PD and correlates them to the phenomena occurring in human disease.

An enzymatic procedure for the purification of DNA restriction fragments without gel electrophoresis and ethidium bromide staining.

A rapid and simple enzymatic method for the purification of a DNA fragment from a restriction digest was developed. The method is based on the two features of exonuclease III activity: digestion of DNA from a 3'-OH at blunt or recessed ends and failure to initiate digestion at DNA ends with four-base 3' overhangs. Herein, we establish a method for purification of a DNA restriction fragment without any physical separation via gel electrophoresis. The elimination of the ethidium bromide staining and ultraviolet irradiation steps should increase the quality and the safety of the purified DNA, a matter of major concern in the perspective of human gene therapy. In addition, since the method described does not use the visualization of the restriction fragments or their difference in size it can be used to purify a DNA fragment from a pool of DNA fragments with the same size even when microquantities of material are available.

Nuclear factor-kappa B activation in permanent intraluminal focal cerebral ischemia in the rat.

Nuclear factor-kappa B (NF-kappa B) is an oxidative stress responsive transcription factor known to be activated in response to transient middle cerebral artery intraluminal occlusion. Since oxidative stress activation may largely occur during reperfusion, the aim of this study was to determine if permanent middle cerebral artery intraluminal occlusion without reperfusion induces NF-kappa B activation and the relationship of NF-kappa B activation to HSP70 expression and neuronal cell death. Our results suggest that permanent intraluminal occlusion is sufficient to induce NF-kappa B activation 7 h after the onset of occlusion. Interestingly, this activation seems to occur specifically in dying neurons of the penumbra area devoid of any HSP70 neuronal immunoreactivity. These findings are consistent with the suggested protective role of HSP70 expression and suggest that NF-kappa B activation observed in the penumbra area has a role in neuronal cell death after permanent intraluminal cerebral ischemia.

Extracellular toxicity of 6-hydroxydopamine on PC12 cells.

6-hydroxydopamine (6-OHDA) is usually thought to cross cell membrane through dopamine uptake transporters, to inhibit mitochondrial respiration and to generate intracellular reactive oxygen species. In this study, we show that the anti-oxidants catalase, glutathione and N-acetyl-cysteine are able to reverse the toxic effects of 6-OHDA. These two latter compounds considerably slow down 6-OHDA oxidation in a cell free system suggesting a direct chemical interaction with the neurotoxin. Moreover, desipramine does not protect PC12 cells and 6-OHDA is also strongly toxic towards non-catecholaminergic C6 and NIH3T3 cells. These results thus suggest that 6-OHDA toxicity on PC12 cells mainly involves an extracellular process.

A cautionary note on the use of stable transformed cells.

Gene transfection and ectopic expression is a widely used method in experimental biology. In the present report, we would like to point out that this approach may, in certain circumstances, lead to a modification of the transfected cell phenotype. Indeed, we observed that after transfection of bcl-2 gene in the neuronal PC12 cell line some of the selected clones have lost their neuronal and catecholaminergic characteristics, i.e. TH expression and ability to grow neurites in response to NGF. Thus, the resistance of some PC12-Bcl-2 clones against neurotoxic insults may not necessarily reflect the potential benefit afforded by Bcl-2 expression. We therefore encouraged authors to verify cell phenotype after stable transfection to avoid misinterpretation of their results.

Electromyographic activity of the lumbar extensor muscles: effect of angle and hand position during Roman chair exercise.

OBJECTIVE: To evaluate the effects of angle and hand position during variable-angle Roman chair (VARC) back extension exercise on lumbar paraspinal electromyographic (EMG) activity. DESIGN: Descriptive, repeated measures. SETTING: University-based musculoskeletal research laboratory. PARTICIPANTS: Two female and eight male volunteers recruited from a university setting. INTERVENTION AND OUTCOME MEASURES: Surface integrated EMG activity was recorded from the L3-L4 paraspinal region during 24 10-second repetitions of dynamic back extension exercise, each consisting of a unique VARC angle (six total) and subject hand position (four total). Lumbar paraspinal surface integrated EMG activity measured in millivolts per repetition was used for analysis. RESULTS: Significant lumbar paraspinal EMG activity was evident during each of the 24 repetitions (p < or = .05), with a 104% increase in activity noted between the lowest and highest. EMG activity increased progressively among hand positions and as the VARC angle became more horizontal. VARC angle affected EMG activity more than hand position, but the greatest impact on EMG activity was produced by modifying both angle and hand position. CONCLUSION: Lumbar paraspinal EMG activity can be altered during VARC back extension exercise by changing angle and hand position. Clinicians can use these data to develop progressive resistance exercise programs using the VARC apparatus.

A novel role for the mating type (MAT) locus in the maintenance of cell wall integrity in Saccharomyces cerevisiae.

The cell wall and stress response component (Wsc) protein family in the yeast Saccharomyces cerevisiae is encoded by at least three genes, WSC1, WSC2, and WSC3. The Wsc proteins are putative upstream activators of the RHO1-regulated PKC1-MAP kinase cascade, and are required for maintenance of cell wall integrity and the stress response. Deletion of WSC1 causes a cell lysis defect that is exacerbated by deleting WSC2 or WSC3. This cell lysis defect can be rescued by adding osmotic stabilizers, such as 1 M sorbitol, to the medium, and by overexpressing PKC1 or RHO1. To advance our understanding of the function of the WSC genes, we performed a genetic screen to identify other components of the pathways they regulate. Here we report our findings. MATa1 and MATalpha2 were identified as dosage-dependent suppressors of the lysis defect of a wsc delta mutant. Overexpression of MATa1 or MATalpha2 was found to suppress the heat shock sensitivity, in addition to the lysis defect, of the wsc delta mutant. Phenotypic suppression by these two genes, MATa1 and MATalpha2, is significantly stronger when they are overexpressed in cells of the opposite mating type. Deletion of MATa1 exacerbates the lysis defect of haploid and diploid wsc delta strains. Our results suggest that the MAT locus plays a role in responses similar to those regulated by WSC and provide evidence for a regulatory effect of the MAT locus outside the realm of cell type determination.

Increased bax expression is associated with cell death induced by ganciclovir in a herpes thymidine kinase gene-expressing glioma cell line.

The herpes simplex virus thymidine kinase gene (HSV-tk) was stably transfected into rat C6 glioma cells (C6tk) in order to characterize the mechanisms underlying cell toxicity induced in vitro by the guanosine analog ganciclovir (GCV). The results demonstrate the efficiency of the HSV-tk/GCV system in ablating most of the tumoral cells within 7 to 8 days of treatment with 20 mivroM GCV; however, a few cells still survive. C6tk cells arrest in the S phase of the cell cycle after 2 days of drug treatment before undergoing cell death. Microscopic analysis reveals dying cells with ultrastructural characteristics consistent with apoptosis; we cannot rule out, however, that necrotic cell death may also be occurring. The cytotoxicity induced by GCV is not associated with changes in the expression of p53 protein, suggesting that cell cycle arrest and cell death may occur through a p53-independent pathway. C6tk cells constitutively express Bcl-xL and Bax proteins; when exposed to GCV, Bcl-xL levels do not change but Bax accumulation is rapidly induced. These findings suggest that the balance between Bcl-xL and Bax proteins may be of importance in determining the sensitivity of tumoral cells to GCV.

p53 and Bax activation in 6-hydroxydopamine-induced apoptosis in PC12 cells.

p53, Bax and Bcl-xL proteins have been implicated in apoptotic neuronal cell death. We have investigated whether those proteins are involved in 6-OHDA-induced PC12 cell death. After a 24-h exposure to the neurotoxin (100 microM), morphological evidence for apoptosis was observed in PC12 cells. Up-regulation of p53 and Bax proteins was demonstrated 4 and 6 h, respectively, after 6-OHDA treatment; in contrast, no change in Bcl-xL levels was found. These findings suggest that p53 and Bax could be relevant markers of neuronal apoptosis as previously described in kainic acid- or ischemia-induced neuronal cell death and may participate to neuronal degeneration in Parkinson's disease.