The impact of cell-cell contact, e-cadherin and EGF receptor on the cellular radiosensitivity of a431 cancer cells

Cell-cell contact is thought to be critically involved in mechanisms leading to radioresistance. Here, we assessed the influence of confluent compared to subconfluent cell culture conditions and the radiosensitizing ability of E-cadherin inhibition alone or in combination with C225-mediated EGFR inhibition in human squamous cell carcinoma cells. Our results show higher radioresistance under subconfluent growth conditions than under confluency. Delayed plating only resulted in higher radiation survival in confluently growing cells. While E-cadherin depletion significantly reduced basal clonogenic survival, increased the rate of apoptosis, and diminished cell adhesion, the cellular radiosensitivity remained unchanged under both subconfluent and confluent conditions. C225 decreased basal cell survival but failed to modify radiation survival. Additional treatment of E-cadherin knockdown cell cultures with C225 did not further reduce basal cell survival or lead to radiosensitization. Interestingly, E-cadherin silencing in 3D cell cultures did not alter radiosensitivity. Our data indicate that cell-cell contact and E-cadherin are not prominently involved in the regulation of radioresistance of human squamous cell carcinoma cells. In addition, no regulatory interaction between E-cadherin and EGFR in the radiation response was observed.     

Neurochemical consequence of steroid abuse: stanozolol-induced monoaminergic changes

An extensive literature has documented adverse effects on mental health in anabolic androgenic steroids (AAS) abusers. Depression seems a common adverse reaction in AAS abusers. Recently it has been reported that in a rat model of AAS abuse stanozolol induces behavioural and biochemical changes related to the pathophysiology of major depressive disorder. In the present study, we used the model of AAS abuse to examine possible changes in the monoaminergic system, a neurobiological substrate of depression, in different brain areas of stanozolol-treated animals. Wistar rats received repeated injections of stanozolol (5mg/kg, s.c.), or vehicle (propylene glycol, 1ml/kg) once daily for 4weeks. Twenty-four hours after last injection, changes of dopamine (DA) and relative metabolite levels, homovanilic acid (HVA) and 3,4-dihydroxy phenylacetic acid (DOPAC), serotonin (5-HT) and its metabolite levels, 5-hydroxy indolacetic acid (5-HIAA), and noradrenaline (NA) amount were investigated in prefrontal cortex (PFC), nucleus accumbens (NAC), striatum (STR) and hippocampus (HIPP). The analysis of data showed that after chronic stanozolol, DA levels were increased in the HIPP and decreased in the PFC. No significant changes were observed in the STR or in the NAC. 5-HT and 5-HIAA levels were decreased in all brain areas investigated after stanozolol exposure; however, the 5-HIAA/5-HT ratio was not altered. Taken together, our data indicate that chronic use of stanozolol significantly affects brain monoamines leading to neurochemical modifications possibly involved in depression and stress-related states.     

The Vindija Neanderthal scapular glenoid fossa: comparative shape analysis suggests evo-devo changes among Neanderthals

Although the shape of the scapular glenoid fossa (SGF) may be influenced by epigenetic and developmental factors, there appears to be strong genetic control over its overall form, such that variation within and between hominin taxa in SGF shape may contain information about their evolutionary histories. Here we present the results of a geometric morphometric study of the SGF of the Neanderthal Vi-209 from Vindjia Cave (Croatia), relative to samples of Plio-Pleistocene, later Pleistocene, and recent hominins. Variation in overall SGF shape follows a chronological trend from the plesiomorphic condition seen in Australopithecus to modern humans, with pre-modern species of the genus Homo exhibiting intermediate morphologies. Change in body size across this temporal series is not linearly directional, which argues against static allometry as an explanation. However, life history and developmental rates change directionally across the series, suggesting an ontogenetic effect on the observed changes in shape (ontogenetic allometry). Within this framework, the morphospace occupied by the Neanderthals exhibits a discontinuous distribution. The Vindija SGF and those of the later Near Eastern Neanderthals (Kebara and Shanidar) approach the modern condition and are somewhat segregated from both northwestern European (Neandertal and La Ferrassie) and early Mediterranean Neanderthals (Krapina and Tabun). Although more than one scenario may account for the pattern seen in the Neanderthals, the data is consistent with palaeogenetic evidence suggesting low levels of gene flow between Neanderthals and modern humans in the Near East after ca. 120-100 ka (thousands of years ago) (with subsequent introgression of modern human alleles into eastern and central Europe). Thus, in keeping with previous analyses that document some modern human features in the Vindija Neanderthals, the Vindija G₃ sample should not be seen as representative of ‘classic’ - that is, unadmixed, pre-contact - Neanderthal morphology.     

Photocrosslinked bovine serum albumin hydrogels with partial retention of esterase activity

Novel biohybrid hydrogels based on bovine serum albumin (BSA) were synthesized by one-pot photopolymerization of chemically modified protein in the presence of N,N'-methylenebisacrylamide (MBA) as cross-linking agent under mild conditions. Two batches of methacrylated albumins were prepared by treating the protein with different amounts of methacrylic anhydride (MAN) and the degree of substitution (DS) of primary amines was quantified via trinitrobenzesulfonic acid (TNBS) colorimetric assay. Hydrogels readily formed when a diluted buffered solution of the modified protein and MBA was exposed to LW-UV in the presence of 1-[4-(2-hydroxyethoxy)phenyl]-2-hydroxy-2-methyl-1-propan-1-one (Irgacure 2959) as the radical initiator. In contrast, no hydrogel was obtained in the absence of a polymerizable BSA, nor when the cross-linker, the radical initiator or UV light exposure was excluded from the reaction, suggesting the critical importance of the combined conditions for hydrogel formation. Hydrogels were characterized via scanning electron microscopy (SEM) and the swelling ratios were monitored at different pHs. The esterolytic activity of the novel biohybrid materials was quantitatively investigated via UV-vis spectroscopy by measuring the release of para-nitrophenol upon incubation with para-nitrophenyl acetate (p-NPA) substrate. The effect of the addition of acrylic acid co-monomer and of the monomer concentration in the catalytic activity and in the swelling behavior was also examined. Finally, the reusability of the materials following one round of catalysis was evaluated.     

Dihydroceramide desaturase and dihydrosphingolipids: debutant players in the sphingolipid arena

Sphingolipids are a wide family of lipids that share common sphingoid backbones, including (2S,3R)-2-amino-4-octadecane-1,3-diol (dihydrosphingosine) and (2S,3R,4E)-2-amino-4-octadecene-1,3-diol (sphingosine). The metabolism and biological functions of sphingolipids derived from sphingosine have been the subject of many reviews. In contrast, dihydrosphingolipids have received poor attention, mainly due to their supposed lack of biological activity. However, the reported biological effects of active site directed dihydroceramide desaturase inhibitors and the involvement of dihydrosphingolipids in the response of cells to known therapeutic agents support that dihydrosphingolipids are not inert but are in fact biologically active and underscore the importance of elucidating further the metabolic pathways and cell signaling networks involved in the biological activities of dihydrosphingolipids. Dihydroceramide desaturase is the enzyme involved in the conversion of dihydroceramide into ceramide and it is crucial in the regulation of the balance between sphingolipids and dihydrosphingolipids. Furthermore, given the enzyme requirement for O? and the NAD(P)H cofactor, the cellular redox balance and dihydroceramide desaturase activity may reciprocally influence each other. In this review both dihydroceramide desaturase and the biological functions of dihydrosphingolipids are addressed and perspectives on this field are discussed.     

Differential protein expression in tears of patients with primary open angle and pseudoexfoliative glaucoma

Primary open angle (POAG) and pseudoexfoliative glaucoma (PXG) are the most common primary and secondary forms of glaucoma, respectively. Even though the patho-physiology, aqueous humor composition, risk factors, clinical features, therapy and drug induced ocular surface changes in POAG and PXG have been widely studied, to date information concerning tear protein characterization is lacking. Tears are a source of nourishment for ocular surface tissues and a vehicle to remove local waste products, metabolized drugs and inflammatory mediators produced in several ophthalmic diseases. In glaucoma, the proteomic definition of tears may provide insights concerning patho-physiology of the disease and ocular surface modifications induced by topical therapy. Our study aimed at characterizing protein patterns in tears of patients with medically controlled POAG and PXG. A comparative tears proteomic analysis by label-free LC-MS(E) highlighted differences in the expression of several proteins in the two glaucoma sub-types and control subjects, highlighting inflammation pathways expressed in both diseases. Results were independently reconfirmed by SDS-PAGE and linear MALDI-TOF MS, validating altered levels of Lysozyme C, Lipocalin-1, Protein S100, Immunoglobulins and Prolactin Inducible Protein. Moreover, we found a differential pattern of phosphorylated Cystatin-S that distinguishes the two pathologies. The most relevant results suggest that in both pathologies there may be active inflammation pathways related to the disease and/or induced by therapy. We show, for the first time, tear protein patterns expressed under controlled intraocular pressure conditions in POAG and PXG subjects. These findings could help in the understanding of molecular machinery underlying these ophthalmologic diseases, resulting in early diagnosis and more specific therapy.     

Modelling the correlation between EGFr expression and tumour cell radiosensitivity, and combined treatments of radiation and monoclonal antibody EGFr inhibitors

ABSTRACT: Purpose To estimate the effects of heterogeneity on tumour cell sensitivity to radiotherapy combined with radiosensitizing agents attributable to differences in expression levels of Epidermal Growth Factor Receptor (EGFr). Materials and methods Differences in radiosensitivity are not limited to cells of different cancer histotypes but also occur within the same cancer, or appear during radiotherapy if radiosensitizing drugs are combined with ionizing radiation. A modified biologically effective dose (MBED), has been introduced to account for changes in radiosensitivity parameters (alpha and alpha/beta) rather than changes in dose/fraction or total dose as normally done with standard biologically effective dose (BED). The MBED approach was applied to cases of EGFr over-expression and cases where EGFr inhibitors were combined with radiation. Representative examples in clinical practice were considered. RESULTS: Assuming membrane EGFr over-expression corresponds to reduced radiosensitivity (alphaH = 0.15 Gy-1 and alphaH/betaH = 7.5 Gy) relative to normal radiosensitivity (alpha = 0.2 Gy-1 and alpha/beta = 10 Gy), an increased dose per fraction of 2.42 Gy was obtained through the application of MBED, which is equivalent to the effect of a reference schedule with 30 fractions of 2 Gy. An equivalent hypo-fractionated regime with a dose per fraction of 2.80 Gy is obtained if 25 fractions are set. Dose fractionations modulated according to drug pharmacokinetics are estimated for combined treatments with biological drugs. Soft and strong modulated equivalent hypo-fractionations result from subtraction of 5 or 10 fractions, respectively. CONCLUSIONS: During this computational study, a new radiobiological tool has been introduced. The MBED allows the required dose per fraction to be estimated when tumour radiosensitivity is reduced because EGFr is over-expressed. If radiotherapy treatment is combined with EGFr inhibitors, MBED suggests new treatment strategies, with schedules modulated according to drug pharmacokinetics.     

Notes about the uses of plants by one of the last healers in the Basilicata Region (South Italy)

Human societies utilize mollusks for myriad material and spiritual ends. An example of their use in a religious context is found in Brazil's African-derived belief systems. Candomblé, an Afro-Brazilian religion introduced during the 18^th-19^th centuries by enslaved Yoruba, includes various magical and liturgical uses of mollusks. This work inventoried the species utilized by adherents and to analyzed their symbolic and magical context. Data were obtained from Candomblé temples in two cities in the northeast of Brazil-Caruaru, in the state of Pernambuco, and Campina Grande, in the state of Paraíba. Questionnaires administered to eleven adepts revealed that at least nineteen mollusk species are being used. Shells from Monetaria moneta, M. annulus and Erosaria caputserpentis were cited by all of the interviewees. Three uses stood out: divination (jogo de búzios); utilization as ritual objects; and employment as sacrificial offerings (Igbin or Boi-de-Oxalá). The jogo de búzios (shell toss), employed in West Africa, Brazil and Cuba, is of fundamental importance to the cult, representing the means by which the faithful enter in contact with the divinities (Orixás) and consult people's futures (Odu). The utilization of mollusks in Candomblé is strongly influenced by ancient Yoruba myths (Itãs) which, having survived enslavement and generations of captive labor, continue to guide the lives of Brazil's African Diaspora.     

Neuropsychological profile and clinical effects of arginine treatment in children with creatine transport deficiency

ABSTRACT: BACKGROUND: SLC6A8, an X-linked gene, encodes the creatine transporter (CRTR) and its mutations lead to cerebral creatine (Cr) deficiency which results in mental retardation, speech and language delay, autistic-like behaviour and epilepsy (CRTR-D, OMIM 300352). CRTR-D represents the most frequent Cr metabolism disorder but, differently from Cr synthesis defects, that are partially reversible by oral Cr supplementation, does not respond to Cr treatment even if precociously administrated. The precursors of Cr are the non-essential amino acids Glycine (Gly) and Arginine (Arg), which have their own transporters at the brain-blood barrier level and, therefore, their supplementation appears an attractive and feasible therapeutic option aimed at stimulating Cr endogenous synthesis and, in this way, at overcoming the block of Cr transport within the brain. However, until now the effects of Arg and/or Gly supplementation on Cr brain levels and behaviour have been controversial. METHODS: In this study five Italian male patients affected by CRTR-D were supplemented with oral LArg at a dosage of 300 mg/kg/day divided into 3 doses, for 24-36 months. Biochemical and plasmatic amino acids examinations and thyroid hormone dosages were periodically performed. Moreover, Proton and Phosphorus Magnetic Resonance Spectroscopy (MRS) was monitored during follow-up in concurrence with neuropsychological evaluations. RESULTS: During L-Arg treatment a clinical improvement in motor skills and to a lesser extent in communication and attention was observed. In addition, all patients had a reduction in the number and frequency of epileptic seizures. Daily living skills appeared also to be positively influenced by L-Arg treatment. Moreover, Total Cr and especially PhosphoCr, evaluated byproton and phosphorus spectroscopy, showed a mild increase, although well below the normal range. CONCLUSION: This study provides information to support the effectiveness of L-Arg supplement treatment in CTRT-D patients; in fact the syndromic pattern of cognitive and linguistic deficit presented by CRTR-D patients was partially altered by L-Arg supplementation especially at a qualitative clinical level. Oral L-Arg may represent not only a protective factor towards a further cognitive decline, but can lead to the acquisition of new skills.