Aldehyde dehydrogenase 1A1 and gelsolin identified as novel invasion-modulating factors in conditioned medium of pancreatic cancer cells

Conditioned medium (CM) from clonal sub-populations of the pancreatic cancer cell line, MiaPaCa-2 with differing invasive abilities, were examined for their effect on in vitro invasion. Conditioned medium from Clone #3 (CM#3) strongly promoted invasion, while CM from Clone #8 (CM#8) inhibited invasion in vitro. 2D DIGE followed by MALDI-TOF MS analysis of CM#3 and CM#8 identified 41 proteins which were differentially regulated; 27 proteins were down-regulated and 14 proteins up-regulated in the invasion-promoting CM#3 when compared to CM#8. Western blotting analysis confirmed the down-regulated expression of gelsolin and the up-regulation of aldehyde dehydrogenase 1A1 in CM#3. Down-regulation of aldehyde dehydrogenase 1A1 in Clone #3 CM and gelsolin levels in Clone #8 CM by siRNA transfection revealed an important involvement of these proteins in promoting and inhibiting invasion in these pancreatic cancer cell lines

RNAi knockdown of Hop (Hsp70/Hsp90 organising protein) decreases invasion via MMP-2 down regulation

We previously identified Hop as over expressed in invasive pancreatic cancer cell lines and malignant tissues of pancreatic cancer patients, suggesting an important role for Hop in the biology of invasive pancreatic cancer. Hop is a co-chaperone protein that binds to both Hsp70/Hsp90. We hypothesised that by targeting Hop, signalling pathways modulating invasion and client protein stabilisation involving Hsp90-dependent complexes may be altered. In this study, we show that Hop knockdown by small interfering (si)RNA reduces the invasion of pancreatic cancer cells, resulting in decreased expression of the downstream target gene, matrix metalloproteinases-2 (MMP-2). Hop in conditioned media co-immunoprecipitates with MMP-2, implicating a possible extracellular function for Hop. Knockdown of Hop expression also reduced expression levels of Hsp90 client proteins, HER2, Bcr-Abl, c-MET and v-Src. Furthermore, Hop is strongly expressed in high grade PanINs compared to lower PanIN grades, displaying differential localisation in invasive ductal pancreatic cancer, indicating that the localisation of Hop is an important factor in pancreatic tumours. Our data suggests that the attenuation of Hop expression inactivates key signal transduction proteins which may decrease the invasiveness of pancreatic cancer cells possibly through the modulation of Hsp90 activity. Therefore, targeting Hop in pancreatic cancer may constitute a viable strategy for targeted cancer therapy

Chemical tuning for potential antitumor fluoroquinolones

[EN] Phototoxic effects of 6,8 dihalogenated quinolones confers to this type of molecules a potential property as photochemotherapeutic agents. Two photodehalogenation processes seem to be involved in the remarkable photoinduced cellular damage. In this context, a new 6,8 dihalogenated quinolone 1 (1-methyl-6,8-difluoro-4-oxo-7-aminodimethy1-1,4-dihydroquinoline-3-carboxylic acid) was synthesized looking for improving the phototoxic properties of fluoroquinolones (FQ) and to determine the role of the photodegradation pathways in the FQ phototoxicity. With this purpose, fluorescence emissions, laser flash photolysis experiments and photodegradation studies were performed with compound 1 using 1-ethyl-6,8-difluoro-4-oxo-7-aminodimethy1-1,4-dihidroquinoline-3-carboxylic acid (2) and lomefloxacin (LFX) as reference compounds. The shortening of alkyl chain of the N(1) of the quinolone ring revealed a lifetime increase of the reactive aryl cation generated from photolysis of the three FQ and a significant reduction of the FQ photodegradation quantum yield. The fact that these differences were smaller when the same study was done using a hydrogen donor solvent (ethanol-aqueous buffer, 50/50 v/v) evidenced the highest ability of the reactive intermediate arising from 1 to produce intermolecular alkylations. These results were correlated with in vitro 3T3 NRU phototoxicity test. Thus, when PhotoIrritation-Factor (PIF) was determined for 1, 2 and LFX using cytotoxicity profiles of BALB/c 3T3 fibroblasts treated with each compound in the presence and absence of UVA light, a PIF more higher than 30 was obtained for 1 while the values for 2 and LFX were only higher than 8 and 10, respectively. Thereby, the present study illustrates an approach to modulate the photosensitizing properties of FQ with the purpose to improve the chemotherapeutic properties of antitumor quinolones. Moreover, the results obtained in this study also evidence that the key pathway responsible for the phototoxic properties associated with dihalogenated quinolones is the aryl cation generation.Financial support from Spanish government (MINECO grant CTQ2014-54729-C2-2-P and Severo Ochoa fellowship for C. A., Carlos III Institute of Health grant PI16/01877), and the Generalitat Valenciana (PROMETEO program, 2017-075). We thank M.P. Marin of IIS La Fe Microscopy Unit for confocal microscopy.Anaya-González, C.; Soldevila Serrano, S.; García-Laínez, G.; Bosca Mayans, F.; Andreu Ros, MI. (2019). Chemical tuning for potential antitumor fluoroquinolones. Free Radical Biology and Medicine. 141:150-158. https://doi.org/10.1016/j.freeradbiomed.2019.06.010S150158141Domagala, J. M., Hanna, L. D., Heifetz, C. 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D., Clements, P. J., Pruimboom-Brees, I., Aylott, M., … Krul, C. A. . (2012). The in vivo rat skin photomicronucleus assay: phototoxicity and photogenotoxicity evaluation of six fluoroquinolones. Mutagenesis, 27(6), 721-729. doi:10.1093/mutage/ges038Soldevila, S., & Bosca, F. (2012). Photoreactivity of Fluoroquinolones: Nature of Aryl Cations Generated in Water. Organic Letters, 14(15), 3940-3943. doi:10.1021/ol301694pCuquerella, M. C., Miranda, M. A., & Boscá, F. (2006). Generation of Detectable Singlet Aryl Cations by Photodehalogenation of Fluoroquinolones. The Journal of Physical Chemistry B, 110(13), 6441-6443. doi:10.1021/jp060634dFreccero, M., Fasani, E., Mella, M., Manet, I., Monti, S., & Albini, A. (2008). Modeling the Photochemistry of the Reference Phototoxic Drug Lomefloxacin by Steady-State and Time-Resolved Experiments, and DFT and Post-HF Calculations. Chemistry - A European Journal, 14(2), 653-663. doi:10.1002/chem.200701099Albini, A., & Monti, S. (2003). 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F., Nichols, J. B., Solomon, M., & Worth, D. F. (1988). 1-Substituted 7-[3-[(ethylamino)methyl]-1-pyrrolidinyl]-6,8-difluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acids. New quantitative structure activity relationships at N1 for the quinolone antibacterials. Journal of Medicinal Chemistry, 31(5), 991-1001. doi:10.1021/jm00400a017Schmidt, R., Tanielian, C., Dunsbach, R., & Wolff, C. (1994). Phenalenone, a universal reference compound for the determination of quantum yields of singlet oxygen O2(1Δg) sensitization. Journal of Photochemistry and Photobiology A: Chemistry, 79(1-2), 11-17. doi:10.1016/1010-6030(93)03746-4Garcia-Lainez, G., Martínez-Reig, A. M., Limones-Herrero, D., Consuelo Jiménez, M., Miranda, M. A., & Andreu, I. (2018). Photo(geno)toxicity changes associated with hydroxylation of the aromatic chromophores during diclofenac metabolism. 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Practice patterns and clinical outcomes in acute appendicitis differ in the elderly patient

Background: Appendicitis is the most frequent global abdominal surgical emergency. An ageing population, who often exhibit atypical symptoms and delayed presentations, challenge conventional diagnostic and treatment paradigms. Objectives: This study aims to delineate disparities in presentation, management, and outcomes between elderly patients and younger adults suffering from acute appendicitis. Methods: This subgroup analysis forms part of ESTES SnapAppy, a time-bound multi-center prospective, observational cohort study. It includes patients aged 15 years and above who underwent laparoscopic appendectomy during a defined 90-day observational period across multiple centers. Statistical comparisons were performed using appropriate tests with significance set at p < 0.05. Results: The study cohort comprised 521 elderly patients (≥65 years) and 4,092 younger adults (18–64 years). Elderly patients presented later (mean duration of symptoms: 7.88 vs. 3.56 days; p < 0.001) and frequently required computed tomography (CT) scans for diagnosis (86.1% vs. 54.0%; p < 0.001). The incidence of complicated appendicitis was higher in the elderly (46.7% vs. 20.7%; p < 0.001). Delays in surgical intervention were notable in the elderly (85.0% operated within 24 h vs. 88.7%; p = 0.018), with longer operative times (71.1 vs. 60.3 min; p < 0.001). Postoperative complications were significantly higher in the elderly (27.9% vs. 12.9%; p < 0.001), including severe complications (6.9% vs. 2.4%; p < 0.001) and prolonged hospital stays (7.9 vs. 3.6 days; p < 0.001). Conclusions: Our findings highlight significant differences in the clinical course and outcomes of acute appendicitis in the elderly compared to younger patients, suggesting a need for age-adapted diagnostic pathways and treatment strategies to improve outcomes in this vulnerable population

Global disparities in surgeons’ workloads, academic engagement and rest periods: the on-calL shIft fOr geNEral SurgeonS (LIONESS) study

: The workload of general surgeons is multifaceted, encompassing not only surgical procedures but also a myriad of other responsibilities. From April to May 2023, we conducted a CHERRIES-compliant internet-based survey analyzing clinical practice, academic engagement, and post-on-call rest. The questionnaire featured six sections with 35 questions. Statistical analysis used Chi-square tests, ANOVA, and logistic regression (SPSS® v. 28). The survey received a total of 1.046 responses (65.4%). Over 78.0% of responders came from Europe, 65.1% came from a general surgery unit; 92.8% of European and 87.5% of North American respondents were involved in research, compared to 71.7% in Africa. Europe led in publishing research studies (6.6 ± 8.6 yearly). Teaching involvement was high in North America (100%) and Africa (91.7%). Surgeons reported an average of 6.7 ± 4.9 on-call shifts per month, with European and North American surgeons experiencing 6.5 ± 4.9 and 7.8 ± 4.1 on-calls monthly, respectively. African surgeons had the highest on-call frequency (8.7 ± 6.1). Post-on-call, only 35.1% of respondents received a day off. Europeans were most likely (40%) to have a day off, while African surgeons were least likely (6.7%). On the adjusted multivariable analysis HDI (Human Development Index) (aOR 1.993) hospital capacity &gt; 400 beds (aOR 2.423), working in a specialty surgery unit (aOR 2.087), and making the on-call in-house (aOR 5.446), significantly predicted the likelihood of having a day off after an on-call shift. Our study revealed critical insights into the disparities in workload, access to research, and professional opportunities for surgeons across different continents, underscored by the HDI

Multivariable analysis to determine if HIV-1 Tat dicysteine motif is associated with neurodevelopmental delay in HIV-infected children in Malawi

Background HIV-1 Tat protein is implicated in HIV-neuropathogenesis. Tat C31S polymorphism (TatCS) has been associated with milder neuropathology in vitro and in animal models but this has not been addressed in a cohort of HIV-infected adults or children. Methods HIV viral load (VL) in plasma and cerebrospinal fluid (CSF) were determined and plasma HIV tat gene was sequenced. Neurodevelopmental assessment was performed using Bayley Scales of Infant Development III (BSID-III), with scores standardized to Malawian norms. The association between TatCS and BSID-III scores was evaluated using multivariate linear regression. Results Neurodevelopmental assessment and HIV tat genotyping were available for 33 children. Mean age was 19.4 (SD 7.1) months, mean log VL was 5.9 copies/mL (SD 0.1) in plasma and 3.9 copies/mL (SD 0.9) in CSF. The prevalence of TatCC was 27 %. Z-scores for BSID-III subtests ranged from −1.3 to −3.9. TatCC was not associated with higher BSID-III z-scores. Conclusions The hypothesis of milder neuropathology in individuals infected with HIV TatCS was not confirmed in this small cohort of Malawian children. Future studies of tat genotype and neurocognitive disorder should be performed using larger sample sizes and investigate if this finding is due to differences in HIV neuropathogenesis between children and adults

Alternatives to robenidine to control gastrointestinal disorders of weaner rabbits in the field

Gastrointestinal disorders due to Eimeria sp. and E. coli overgrowth cause high mortality in weaner rabbits and the interest in alternatives to coccidiostats is high. This study aimed to investigate the superiority of natural feed additives towards robenidine preserving gastrointestinal health in the field. Rabbits were divided into four groups, Control Group (CG) exclusively supplemented with robenidine, Sainfoin Group (SG) was supplemented with a combination of robenidine and sainfoin, and two additional groups were respectively supplemented with Herb-All COCC-X (garlic; conessi tree) (HG: Herbal Group) and by a combination of Herb-All COCC-X and Klinofeed (clinoptilolite) (MG: Mineral Group). Eimeria sp. (98,40%) and E. coli overgrowth (73.60%) could be confirmed as the main causes for losses. High mortality rates (SG: 30.00% - MG: 47.50%), also in the groups receiving robenidine (SG: 30.00%; CG: 45.00%), reinforced the importance of alternatives in the field. The natural additives of groups SG, HG and MG did not have a significant influence on the weight gains and the oocyst counts in the jejunum/ileum and caecum of slaughter rabbits at the end of the trial, compared to group CG. Significantly higher oocyst shedding in SG (p = 1.4E-03) and HG (p = 1.4E-05) during the trial may be explained by a higher surviving rate of diseased rabbits in those groups, fostered by beneficial effects of the additives, which should be investigated further

The cAMP analog 8-Cl-cAMP inhibits growth and induces differentiation and apoptosis in retinoblastoma cells

Retinoblastomas appear to be derived from a multipotential stem cell of the retina, due to alterations of the Rb I gene. These tumors arise only within a discrete time frame during childhood, prior to terminal differentiation of the retinal precursor cells. Treatment of retinoblastoma cells with certain agents can induce a partial differentiation of cell types resembling those of the mature retina, such as rod and cone photoreceptors, glia, conventional neurons and pigment epititelia. We have tested the effects of 8-CI-cAMP, a synthetic analog of cAMP which preferentially binds to and activates the RII subunit of protein kinase A on the Y-79 retinoblastoma cell line in vitro. Y-79 cells treated with 8-CI-cAMP produced short, branching processes and showed a substantial increase in staining for neuron-specific enolase, a marker for conventional neuronal differentiation. In contrast, dibutyryl-cAMP gives a strong increase in the glial marker glial acidic fibrillary protein. Y-79 cell proliferation was strongly inhibited by 8-CI-cAMP at concentrations as low as 5-25 mu M. 8-CI-cAMP significantly increased the rate of apoptosis of Y-79 cells in a dose-dependent manner. It also modulated expression of the RI regulatory subunit of intracellular cAMP-dependent protein kinase A, which is produced in abnormal quantities by Y-79 cells. A decrease in protein production was observed, with no clear effect on the RI subunit mRNA expression, suggesting that RI regulation occurs posttranscriptionall