The synthesis and the in vitro cytotoxicity studies of bisnaphthalimidopropyl polyamine derivatives against colon cancer cells and parasite Leishmania infantum

Bisnaphthalimidopropyl derivatives (BNIPSpd, BNIPDaoct, BNIPDanon, BNIPDadec, BNIPDpta and BNIPDeta) were synthesised in yields ranging from 50% to 70% and their cytotoxicity against colon cancer cells (Caco-2) and the parasite Leishmania infantum determined using the MTT assay. Cytotoxicity within Caco-2 cells was manifested with IC(50) values between 0.3 and 22 microM. Compounds with the central longer alkyl chains exhibited the highest cytotoxicity. Against L. infantum, IC(50) values were encompassed within a narrower concentration range of 0.47-1.54 microM. In the parasites, the presence of nitrogen in the central chain and the length of the central alkyl chains did not especially enhance cytotoxicity. This may be due to the way these compounds are transported in the cells.     

Dosimetric assessment of the exposure of radiotherapy patients due to cone-beam CT procedures

Cone-beam computed tomography (CBCT) is widely used for pre-treatment verification and patient setup in image-guided radiation therapy (IGRT). CBCT imaging is employed daily and several times per patient, resulting in potentially high cumulative imaging doses to healthy tissues that surround exposed target organs. Computed tomography dose index (CTDI) is the parameter used by CBCT equipment as indication of the radiation output to patients. This study aimed to increase the knowledge on the relation between CBCT organ doses and weighted CTDI (CTDI_W) for a thorax scanning protocol. A CBCT system was modelled using the Monte Carlo (MC) radiation transport program MCNPX2.7.0. Simulation results were validated against half-value layer (HVL), axial beam profile, patient skin dose (PSD) and CTDI measurements. For organ dose calculations, a male voxel phantom (“Golem”) was implemented with the CBCT scanner computational model. After a successful MC model validation with measurements, a systematic comparison was performed between organ doses (and their distribution) and CTDI dosimetry concepts [CTDI_W and cumulative dose quantities f₁₀₀(150) and \({\text{CTD}}{{\text{I}}_\infty }\)]. The results obtained show that CBCT organ doses vary between 1.2 ± 0.1 mGy and 3.3 ± 0.2 mGy for organs located within the primary beam. It was also verified that CTDI_W allows prediction of absorbed doses to tissues at distances of about 5 cm from the isocentre of the CBCT system, whereas f₁₀₀(150) allows prediction of organ doses at distances of about 10 cm from the isocentre, independently from its location. This study demonstrates that these dosimetric concepts are suitable methods that easily allow a good approximation of the additional CBCT imaging doses during a typical lung cancer IGRT treatment.

     

Detection of human neutrophil elastase (HNE) on wound dressings as marker of inflammation

Applied Microbiology and Biotechnology - Chronic wound fluids have elevated concentration of human neutrophil elastase (HNE) which can be used as inflammation/infection marker. Our goal is to...     

Seasonal variation in plasma lipids and lipases in young healthy humans

Although intermediate metabolism is known to follow circadian rhythms, little information is available on the variation in lipase activities (lipoprotein and hepatic lipase, LPL and HL, respectively) and lipids throughout the year.

In a cross-sectional study, we collected and analysed blood from 245 healthy students (110 men and 135 women) between 18 and 25 years old from the University of Barcelona throughout the annual campaign (March, May, October and December) of the blood bank. All subjects gave their written informed consent to participate. All blood samples were taken after breakfast at 8:00 and 11:00 am.

Plasma glucose, total plasma protein, triacylglycerides (TAG), free fatty acids (FFA), free cholesterol and esterified cholesterol (FC and TC, respectively), cholesterol in low-density lipoproteins (cLDL), cholesterol in high-density lipoproteins (cHDL), phospholipids (PL) and lipase activities (LPL and HL) were determined. Cosinor analysis was used to evaluate the presence (significance of fit cosine curve to data and variance explained by rhythm) and characteristics of possible 12-month rhythms (acrophase, MESOR and amplitude).

Statistically significant seasonal rhythms were detected for all the variables studied except proteins, with most of them peaking in the winter season. The lowest value for cLDL and the HL occurs in summer, while for cHDL and the LPL it is in winter.

These findings demonstrate for the first time that in physiological conditions, plasma LPL and HL activities and lipids follow seasonal rhythms. The metabolic significance of this pattern is discussed.     

Large-scale production of a thermostable <Emphasis Type="Italic">Rhodothermus marinus</Emphasis> cellulase by heterologous secretion from <Emphasis Type="Italic">Streptomyces lividans</Emphasis>

Background

The gene encoding a thermostable cellulase of family 12 was previously isolated from a Rhodothermus marinus through functional screening. CelA is a protein of 260 aminoacyl residues with a 28-residue amino-terminal signal peptide. Mature CelA was poorly synthesized in some Escherichia coli strains and not at all in others. Here we present an alternative approach for its heterologous production as a secreted polypeptide in Streptomyces.

Results

CelA was successfully over-expressed as a secreted polypeptide in Streptomyces lividans TK24. To this end, CelA was fused C-terminally to the secretory signal peptide of the subtilisin inhibitor protein (Sianidis et al. in J Biotechnol. 121: 498–507, 2006) from Streptomyces venezuelae and a new cloning strategy developed. Optimal growth media and conditions that stall biomass production promote excessive CelA secretion. Under optimal growth conditions in nutrient broth medium, significant amounts of mature CelA (50–90 mg/L or 100–120 mg/g of dry cell weight) are secreted in the spent growth media after 7 days. A protocol to rapidly purify CelA to homogeneity from culture supernatants was developed and specific anti-sera raised against it. Biophysical, biochemical and immmuno-detection analyses indicate that the enzyme is intact, stable and fully functional. CelA is the most thermostable heterologous polypeptide shown to be secreted from S. lividans.

Conclusion

This study further validates and extends the use of the S. lividans platform for production of heterologous enzymes of industrial importance and extends it to active thermostable enzymes. This study contributes to developing a platform for poly-omics analysis of protein secretion in S. lividans.

     

Crosstalk between glial and glioblastoma cells triggers the “go-or-grow” phenotype of tumor cells

Background

Glioblastoma (GBM), the most malignant primary brain tumor, leads to poor and unpredictable clinical outcomes. Recent studies showed the tumor microenvironment has a critical role in regulating tumor growth by establishing a complex network of interactions with tumor cells. In this context, we investigated how GBM cells modulate resident glial cells, particularly their paracrine activity, and how this modulation can influence back on the malignant phenotype of GBM cells.

Methods

Conditioned media (CM) of primary mouse glial cultures unexposed (unprimed) or exposed (primed) to the secretome of GL261 GBM cells were analyzed by proteomic analysis. Additionally, these CM were used in GBM cells to evaluate their impact in glioma cell viability, migration capacity and activation of tumor-related intracellular pathways.

Results

The proteomic analysis revealed that the pre-exposure of glial cells to CM from GBM cells led to the upregulation of several proteins related to inflammatory response, cell adhesion and extracellular structure organization within the secretome of primed glial cells. At the functional levels, CM derived from unprimed glial cells favored an increase in GBM cell migration capacity, while CM from primed glial cells promoted cells viability. These effects on GBM cells were accompanied by activation of particular intracellular cancer-related pathways, mainly the MAPK/ERK pathway, which is a known regulator of cell proliferation.

Conclusions

Together, our results suggest that glial cells can impact on the pathophysiology of GBM tumors, and that the secretome of GBM cells is able to modulate the secretome of neighboring glial cells, in a way that regulates the “go-or-grow” phenotypic switch of GBM cells.

     

GLUT1‐mediated glucose uptake plays a crucial role during Plasmodium hepatic infection

Intracellular pathogens have evolved mechanisms to ensure their survival and development inside their host cells. Here, we show that glucose is a pivotal modulator of hepatic infection by the rodent malaria parasite Plasmodium berghei and that glucose uptake via the GLUT1 transporter is specifically enhanced in P. berghei‐infected cells. We further show that ATP levels of cells containing developing parasites are decreased, which is known to enhance membrane GLUT1 activity. In addition, GLUT1 molecules are translocated to the membrane of the hepatic cell, increasing glucose uptake at later stages of infection. Chemical inhibition of GLUT1 activity leads to a decrease in glucose uptake and the consequent impairment of hepatic infection, both in vitro and in vivo. Our results reveal that changes in GLUT1 conformation and cellular localization seem to be part of an adaptive host response to maintain adequate cellular nutrition and energy levels, ensuring host cell survival and supporting P. berghei hepatic development.