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β-thalassemia major (βTM) patients require frequent blood transfusions, with consequences that span from allogenic reactions to iron overload. To minimize these effects, βTM patients periodically receive leucodepleted packed red blood cells (P-RBCs) stored for maximum 14 days. The aim of this study was to compare two alternative routine procedures to prepare the optimal P-RBCs product, in order to identify differences in their content that may somehow affect patients' health and quality of life (QoL). In method 1, blood was leucodepleted and then separated to obtain P-RBCs, while in method 2 blood was separated and leucodepleted after removal of plasma and buffycoat. Forty blood donors were enrolled in two independent centers; couples of phenotypically matched whole blood units were pooled, divided in two identical bags and processed in parallel following the two methods. Biochemical properties, electrolytes and metabolic composition were tested after 2, 7 and 14 days of storage. Units prepared with both methods were confirmed to have all the requirements necessary for βTM transfusion therapy. Nevertheless, RBCs count and Hb content were found to be higher in method-1, while P-RBCs obtained with method 2 contained less K + , iron and storage lesions markers. Based on these results, both methods should be tested in a clinical perspective study to determine a possible reduction of transfusion-related complications, improving the QoL of βTM patients, which often need transfusions for the entire lifespan.

Background : Platelet-Rich Plasma (PRP) induces bone regeneration; however, there is low evidence supporting its efficacy in bone healing. The lack of a standardized protocol of administration represents the main obstacle to its use in the clinical routine for bone defects' treatment. The purpose of this study was to characterize PRP and elucidate its osteogenic potential. Methods : Platelet count, fibrinogen levels, and growth factors concentration were measured in PRP obtained by four apheresis procedures. HOB-01-C1, a pre-osteocytic cell line, was used to examine the effects of different PRP dilutions (from 1% to 50%) on cell viability, growth, and differentiation. Gene expression of RUNX2, PHEX, COL1A1, and OCN was also assayed. Results : PRP showed a mean 4.6-fold increase of platelets amount compared to whole blood. Among the 36 proteins evaluated, we found the highest concentrations for PDGF isoforms, EGF, TGF-β and VEGF-D. PDGF-AA positively correlated with platelet counts. In three of the four tested units, 25% PRP induced a growth rate comparable to the positive control (10% FBS); whereas, for all the tested units, 10% PRP treatment sustained differentiation. Conclusions : This study showed that PRP from apheresis stimulates proliferation and differentiation of pre-osteocyte cells through the release of growth factors from platelets.

Allogeneic peripheral blood-derived (PBS) serum eye drops have been largely used in the treatment of dry eye disease (DED). Recently, cord blood has emerged as an effective alternative serum source (cord blood serum, CBS), containing a higher amount of growth factors than PBS, it holds the promise of a better capability to stimulate corneal healing. However, the lack of a standardized method for preparation, dispensation, storage and a poor biochemical characterization still hamper the establishment of a clinical consensus. Here the metabolomes of the two different serum eye drop preparations were compared using proton nuclear magnetic resonance spectroscopy. We found that both PBS and CBS contained several organic compounds, the majority of them already detected in human tears and may be thereby considered lacrimal substitutes. Metabolites having in the multivariate statistical analysis Partial least squares discriminant analysis (PLS-DA) a VIP scores > 1.0 were considered to be significantly different. All the metabolites identified were found to have a p < 0.05 in the univariate analysis. CBS, in particular, showed the highest amount of choline, myo-inositol, glutamine, creatine and β-hydroxybutyrate. These evidences constitute relevant advances towards serum eye drops characterization and confirm that cord blood is a valid alternative source of serum eye drops., (Copyright © 2021. Published by Elsevier Ltd.)

More than 60% of the 1,700 infectious diseases that affect human come from animals and zoonotic pandemics, after starting from sporadic phenomena limited to rural areas, have become a global emergency. The repeated and frequent zoonotic outbreaks such as the most recent COVID-19 pandemic can be attributed also to human activities. In particular, the creation of enormous intensive domestic animal farms, the indiscriminate use of antibiotics, the destruction of forests, the consumption of the meat of wild animals and the illegal animal trade are all factors causing the insurgence and the transmission of zoonotic diseases from animals to humans. The purpose of this study was to explore the knowledge of the One Health concept including the zoonotic risk potentially derived from illegally traded pet animals and wildlife among adolescents in 6 different countries (Italy, Austria, Slovenia, Germany, Mauritius, and Japan). A representative sample of 656 students was recruited and all participants took an anonymous questionnaire. Data were analyzed by ANOVAs to estimate the prevalence of correct health prevention behaviors and to identify the influential factors for these behaviors. After two theoretical-practical lectures, the same anonymous questionnaire was administered for the second time in order to assess the efficacy of the program. The proportion of students who did not know that many diseases affecting humans come from animals is 28.96% while 32.16% of them did not know what a zoonosis is. The circularity of the One Health concept related to the transmission of diseases from animals to humans and vice-versa is not understood from a large prevalence of the adolescents with 31.40 and 59.91% of wrong responses, respectively. Furthermore, rabies is not considered as a dangerous disease by 23.02% of the adolescents. After two theoretical-practical classroom sessions, the correct answers improved to 21.92% according to the different question. More than a third of the student cohort investigated showed a profound ignorance of the zoonotic risks and a poor understanding of the One Health concept. The authors believe that the teaching of health prevention with a One Health approach and a practical training should be included in every school curriculum., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zucca, Rossmann, Dodic, Ramma, Matsushima, Seet, Holtze, Bremini, Fischinger, Morosetti, Sitzia, Furlani, Greco, Meddi, Zambotto, Meo, Pulcini, Palei and Zamaro.)

Objective: The main focus of this in vitro study was to highlight possible differences between outcomes of photobiomodulation performed in the presence or absence of growth factors derived from platelet-rich plasma. Background: Photobiomodulation has garnered increasing attention, thanks to a large number of controlled clinical trials that have proven its efficacy in various oral pathologies. Nevertheless, the mechanism of action is still a matter of debate. Materials and methods: The cell model used was Saos-2ATTC HTB-85, a human osteosarcoma cell line that retains an osteogenic potential matching that of osteoblastic cells. Photobiomodulation was performed with a 645 nm diode laser; we investigated three different fluence values (2, 5, and 10 J/cm 2 ) delivered with 3 different irradiation times (1, 2, and 4 min). The design of the study included a case-control structure. Cell viability was assessed by resazurin reduction assay before laser irradiation. We assessed cell differentiation by Alizarin-red Sigma Aldrich assay 48 h after the last laser irradiation. Results: Results show that the combination of photobiomodulation and platelet-rich plasma can lead to a statistically significant increase in both proliferation and differentiation rates. Conclusions: Only a defined amount of energy, that is, a fluence of 5 J/cm 2 delivered in 2 min and of 10 J/cm 2 in 4 min, was proven to be the most effective in the presence of platelet-rich plasma to induce cell proliferation and calcium deposition.

Mutations in the Plasmodium falciparum chloroquine resistance transporter, PfCRT, are the major determinant of chloroquine resistance in this lethal human malaria parasite. Here, we describe P. falciparum lines subjected to selection by amantadine or blasticidin that carry PfCRT mutations (C101F or L272F), causing the development of enlarged food vacuoles. These parasites also have increased sensitivity to chloroquine and some other quinoline antimalarials, but exhibit no or minimal change in sensitivity to artemisinins, when compared with parental strains. A transgenic parasite line expressing the L272F variant of PfCRT confirmed this increased chloroquine sensitivity and enlarged food vacuole phenotype. Furthermore, the introduction of the C101F or L272F mutation into a chloroquine-resistant variant of PfCRT reduced the ability of this protein to transport chloroquine by approximately 93 and 82%, respectively, when expressed in Xenopus oocytes. These data provide, at least in part, a mechanistic explanation for the increased sensitivity of the mutant parasite lines to chloroquine. Taken together, these findings provide new insights into PfCRT function and PfCRT-mediated drug resistance, as well as the food vacuole, which is an important target of many antimalarial drugs.

Sarco/endoplasmic reticulum Ca(2+)-ATPases (SERCAs) are increasingly being studied for therapeutic interventions in the fields of cancer, heart disease, and infection. Our suggestion a decade ago that artemisinins (the most important antimalarial class) act by inhibiting parasite SERCAs (PfATP6 and orthologues) expressed in Xenopus oocytes stimulated new directions for research away from conventional site-of-action studies of the food vacuole of the parasite. There is, however, still no consensus on how artemisinins act. We have continued to explore the hypothesis that PfATP6 is a key target by confirming that artemisinins inhibit Plasmodium falciparum PfATP6 when it is expressed in yeast and that it is essential for survival of pathogenic asexual-stage parasites. These advances are discussed with their implications for our understanding of how parasites regulate calcium in different stages of asexual development and for the global challenge posed by artemisinin resistance., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Background: The mechanism of action of artemisinins against malaria is unclear, despite their widespread use in combination therapies and the emergence of resistance., Results: Here, we report expression of PfATP6 (a SERCA pump) in yeast and demonstrate its inhibition by artemisinins. Mutations in PfATP6 identified in field isolates (such as S769N) and in laboratory clones (such as L263E) decrease susceptibility to artemisinins, whereas they increase susceptibility to unrelated inhibitors such as cyclopiazonic acid. As predicted from the yeast model, Plasmodium falciparum with the L263E mutation is also more susceptible to cyclopiazonic acid. An inability to knockout parasite SERCA pumps provides genetic evidence that they are essential in asexual stages of development. Thaperoxides are a new class of potent antimalarial designed to act by inhibiting PfATP6. Results in yeast confirm this inhibition., Conclusions: The identification of inhibitors effective against mutated PfATP6 suggests ways in which artemisinin resistance may be overcome.

With the advent of artemisinin resistance, it is timely to revisit the biological basis for the controversial suggestion that this class of antimalarial exerts its activity by inhibiting a calcium ATPase (PfATP6) that is most similar to sarcoplasmic endoplasmic reticulum calcium ATPases (SERCAs). Herein, evidence is discussed that relates to this hypothesis as alternative suggestions for how artemisinins might act have been reviewed elsewhere., (Copyright © 2010 Elsevier Ltd. All rights reserved.)