Recycling Apparent Waste Into Biologicals: The Case of Umbilical Cord Blood in Italy and Spain

Most public cord blood banking programs are currently facing financial difficulties due to a progressive decline in the number of cord blood transplants performed worldwide and to a high discard rate of the donated units caused by progressively increasing thresholds of the stem cell dose required to perform safe and effective hemopoietic cord blood transplants. Recycling a proportion of unused cord blood units to prepare novel cord blood components obtained with minimal manipulation (platelets, plasma, red blood cells) and to develop more technologically complex products regulated in the US as Cellular and Gene Therapy Products and in Europe as Advanced Therapy Medicinal Products [e.g. virus-specific T cells (VST), natural killer (NK) cells, induced pluripotent stem cells (iPSCs) is a promising strategy to increase the therapeutic value and reduce the financial deficits of public cord blood banking. Based on encouraging preliminary evidences reported in the literature, additional laboratory studies, large multicenter clinical trials and international regulatory harmonization are necessary to achieve these important goals. This article describes organizational, methodological and regulatory advancements developed in Italy and Spain to promote the clinical use of cord blood platelets, plasma and red blood cells.

Fermented milk cream and its properties

Milk fat has an important role in the formation of fla-vour, aroma, texture proper-ties and quality in milk and dairy products, and milk fat is encapsulated in the glob-ule membrane. Milk fat contains triglycerides, phospholipids, free sterols, β-carotene (provitamin A), fat-soluble vitamins (A, D, E, K), sphingomyelins and fatty acids. In addition to its nutritional value, specific short-chain and medium-chain fatty acids found in milk fat are important for health as an energy source for muscle, heart, liver, kid-ney, blood platelets and nervous system. In addition, these fatty acids do not pose a risk of obesity; they also prevent ulcerative colitis, cancer, atherosclerosis and hypertension, and increase natural immunity with their anti-inflammatory and anti-bacterial effects. Fermented cream, which has a high fat content, is becoming more and more popular in many countries with its nutritional value and its use for many purposes in culinary culture. The classification of fer-mented creams, fat content, production method and used starter cultures vary by country. In this study, fer-mented cream produced in Turkey and in the world classification, production stages, the parameters af-fecting the sensory and tex-tural properties was investi-gated.

The Changes of Blood Platelet Reactivity in the Presence of Crude Extracts Isolated from Leaves and Twigs of Elaeagnus Rhamnoides (L.) A. Nelson Measuring in Whole Blood

Uncontrolled blood platelet activation is an important risk factor of cardiovascular disease (CVDs). Various studies on phenolic compounds indicate that they have a protective effect on the cardiovascular system through different mechanisms, including the reduction of blood platelet activation. One of the plants that is particularly rich in phenolic compounds is sea buckthorn (Elaeagnus rhamnoides (L.) A. Nelson). The aim of the present study in vitro was to determine the anti-platelet properties of crude extracts isolated from leaves and twigs of E. rhamnoides (L.) A. Nelson in whole blood using flow cytometric and total thrombus-formation analysis system (T-TAS). The aim of our study was also analyze of blood platelet proteoms in the presence of different sea buckthorn extracts. A significant new finding is a decrease surface expression of P-selectin on blood platelets stimulated by 10 µM ADP and 10 µg/mL collagen, and a decrease surface expression of GPIIb/IIIa active complex on non-activated platelets and platelets stimulated by 10 µM ADP and 10 µg/mL collagen in the presence of sea buckthorn leaf extract (especially at the concentration 50 µg/mL). The twig extract also displayed antiplatelet potential. However, this activity was higher in the leaf extract than in the twig extract in whole blood. In addition, our present findings clearly demonstrate that investigated plant extracts have anticoagulant properties (measured by T-TAS). Therefore, the two tested extracts may be promising candidates for the natural anti-platelet and anticoagulant supplements.

Ethanolic extract of Ocimum Gratissimum (Scent Leaf) leaves improves haematological parameters in restrain-stressed rats

Stress alters system physiology as well as humoral immunity and hematology, and its problems have become a global issue. The present study was designed to investigate effects of the use of traditional herbs such as Ocimum gratissimum (OG) on hematological parameters in stressed rats. We subjected adult female Wistar rats to prolonged restrained stress for 14 days and administered 100 mg/kg, 200 mg/kg, 400 mg/kg of OG leaf extract and Vitamin C respectively for 14 days. Animals (200-250 g) were sacrificed by cervical dislocation at day 0, day 7 and Day 14 so as to assess the dose and time dependent effect of the extract. Findings revealed that OG leaf extract caused a dose-dependent significant protective effect (P<0.05) on the packed cell volume (PCV), total white blood cell (WBC) count, haemoglobin (Hb), platelet count and total red blood cell (RBC) count compared to Vitamin C treated group. Our study confirms that OG leaf extract is better at improving haematological parameters of restrain stressed rats compared to Vitamin C and it was more effective in animals that were administered 100 mg/kg and 400 mg/kg of the extract. The result indicates that OG leaf extract could alleviate prolonged exposure to stress by lowering PCV, total WBC, total RBC, hemoglobin count and elevating blood platelets.

The Anti-platelet and Anticoagulant Potential of Isorhamnetin and Its New Derivatives Isolated From Sea Buckthorn Fruits in Whole Blood

Blood platelets play a crucial role in hemostasis, the process responsible for keeping blood flowing in the circulatory system. However, unnecessary platelet activation can lead to aggregation at the site of atherosclerotic plaque rapture and the formation of a thrombus, which promotes atherothrombotic diseases. Various dietary components, such as phenolic compounds, are known to demonstrate antiplatelet and anticoagulant properties, and it is possible that these could form an important element in the prophylaxis and therapy of cardiovascular diseases. Our present study examines the biological activity of isorhamnetin (compound 1) and two isorhamnetin derivatives, compound 2 (3-O-beta-glucoside-7-O-alpha-rhamnoside) and compound 3 (3-O-beta-glucoside-7-O-alpha-(3”’-isovaleryl)-rhamnoside), isolated from the phenolic fraction of sea buckthorn fruit, against human washed blood platelets and human whole blood in vitro. The anti-platelet and anticoagulant potential was determined using (A) flow cytometry, (B) the thrombus-formation analysis system (T-TAS) and (C) colorimetry. The tested flavonoids demonstrated anticoagulant and anti-platelet potential, including anti-adhesive activity, with these effects being more intense in compound 2 than isorhamnetin. Compound 2 inhibited GPIIb/IIIa and P-selectin expression on blood platelets from whole blood, and demonstrated anti-adhesion properties in washed blood platelets and anti-coagulant potential in whole blood, measured by T-TAS.

Control of Blood Coagulation by Hemocompatible Material Surfaces—A Review

Hemocompatibility of biomaterials in contact with the blood of patients is a prerequisite for the short- and long-term applications of medical devices such as cardiovascular stents, artificial heart valves, ventricular assist devices, catheters, blood linings and extracorporeal devices such as artificial kidneys (hemodialysis), extracorporeal membrane oxygenation (ECMO) and cardiopulmonary bypass. Although lower blood compatibility of materials and devices can be handled with systemic anticoagulation, its side effects, such as an increased bleeding risk, make materials that have a better hemocompatibility highly desirable, particularly in long-term applications. This review provides a short overview on the basic mechanisms of blood coagulation including plasmatic coagulation and blood platelets, as well as the activation of the complement system. Furthermore, a survey on concepts for tailoring the blood response of biomaterials to improve the hemocompatibility of medical devices is given which covers different approaches that either inhibit interaction of material surfaces with blood components completely or control the response of the coagulation system, blood platelets and leukocytes.

Platelet PD-L1 reflects collective intratumoral PD-L1 expression and predicts immunotherapy response in non-small cell lung cancer

Immune-checkpoint inhibitors (ICI) have transformed oncological therapy. Up to 20% of all non-small cell lung cancers (NSCLCs) show durable responses upon treatment with ICI, however, robust markers to predict therapy response are missing. Here we show that blood platelets interact with lung cancer cells and that PD-L1 protein is transferred from tumor cells to platelets in a fibronectin 1, integrin α5β1 and GPIbα-dependent manner. Platelets from NSCLC patients are found to express PD-L1 and platelet PD-L1 possess the ability to inhibit CD4 and CD8 T-cells. An algorithm is developed to calculate the activation independent adjusted PD-L1 payload of platelets (pPD-L1Adj.), which is found to be superior in predicting the response towards ICI as compared to standard histological PD-L1 quantification on tumor biopsies. Our data suggest that platelet PD-L1 reflects the collective tumor PD-L1 expression, plays important roles in tumor immune evasion and overcomes limitations of histological quantification of often heterogeneous intratumoral PD-L1 expression.

Application of conditioned medium from mesenchymal stromal cells in the protocol for ex vivo production of megakaryocytes and platelets

Introduction. Of interest is the use of a conditioned medium from mesenchymal stromal cells in order to increase the expansion of CD34+ hematopoietic stem cells (HSCs).Aim — to analyze the effi cacy of two methods of ex vivo production of human megakaryocytes and platelets from CD34+ cord blood HSC using conditioned media from mesenchymal stromal cells and IMDM. Methods. Two cultivation methods that differ from each other by medium composition were compared. As a control of antigen expression of the donor, venous blood platelets were used. CD34+ HSCs were isolated from mononuclear fraction of cord blood using the immunomagnetic selection technique. The resulting cells were introduced at a concentration of 1 × 104 cells/mL into 24-well plates and cultured at 39 °C and 10 % CO2 for the first 7 days, after which the conditions were changed to 37 °C and 5 % CO2 and cultured for 14 days. In Group 1, up to day 7, the culture was performed using conditioned medium from mesenchymal stromal cell containing TPO (30 ng/mL), SCF (2 ng/mL), IL-6 (7.5 ng/mL), IL-9 (13.5 ng/mL), and in Group 2 a IMDM medium with the same cytokine cocktail was used. The cells were calculated using haemocytometer. CD34, CD41a, CD42b expression was evaluated using fl ow cytometry. Statistic data was processed with using R-language. The differences were evaluated as statistically signifi cant at signifi cance level p < 0.05.Results. Megakaryocyte production was observed starting from day 7 of culture. The expression level using conditioned medium from mesenchymal stromal cells (Group 1) according to CD41a was 5.84 ± 0.33 % versus 10.43 ± 1.08 % using IMDM medium (Group 2). On day 13 the ratio increased up to 42.05 ± 1.71 % in Group 1 and 61.78 ± 1.71 % in Group 2. CD41a+ megakaryocytes of Group 1 expressed the CD42b marker at the level of 96.85 ± 1.06 % versus 88.7 ± 0.56 % in Group 2. With the application of MSC conditioned medium the average number of nucleated cells was signifi cantly higher on the day 11 and it was equal 326.016 ± 1.86 × 104 cells/mL vs 197.26 ± 10.55 × 104 cells/mL in IMDM medium. Proplatelet formation was observed with microscopy staring from the day 12. The ratio of CD41a+ /CD42b+ platelets was 59.5 ± 3.85 % in conditioned medium, 65.9 ± 8.72 % in IMDM, and 96.11 ± 0.89 % in control platelets derived from venous blood.Conclusion. It was demonstrated that the use of MSC conditioned medium leads to an increase in the expansion of nucleated cells, however it decreases the rate of differentiation in megakaryocytes.

Preparations from selected cucurbit vegetables as antiplatelet agents

Increased blood platelet activation plays an important role in cardiovascular diseases (CVDs). Recent experiments indicate that certain fruits and vegetables, including onion, garlic, and beetroot, have anti-platelet potential and therefore may reduce the likelihood of CVDs. While vegetables from the Cucuritaceae family are known to exerting beneficial antioxidant and anti-inflammatory effects, their effects on blood platelet activation are poorly understood. Therefore, the aim of the present study was to determine the effect on platelet adhesion of preparations from selected cucurbits: pumpkin (Cucurbita pepo; fruit without seeds), zucchini (Cucurbita pepo convar. giromontina; fruit with seeds), cucumber (Cucumis sativus; fruit with seeds), white pattypan squash (Cucurbita pepo var. patisoniana; fruit without seeds) and yellow pattypan squash (Cucurbita pepo var. patisoniana, fruit without seeds). It also evaluates the activity of these preparations on enzymatic lipid peroxidation in thrombin-activated washed blood platelets by TBARS assay. The study also determines the anti-platelet properties of these five cucurbit preparations in whole blood by flow cytometry and with the total thrombus-formation analysis system (T-TAS) and evaluates the cytotoxicity of the tested preparations against platelets based on LDH activity. The results indicate that the yellow Cucurbita pepo var. patisoniana preparation demonstrated stronger anti-platelet properties than the other tested preparations, reducing the adhesion of thrombin-activated platelets to collagen/fibrinogen, and inhibiting arachidonic acid metabolism and GPIIb/IIIa expression on 10 µM ADP-activated platelets. None of the preparations was found to cause platelet lysis. Our findings provide new information on the anti-platelet activity of the tested cucurbit preparations and their potential for treating CVDs associated with platelet hyperactivity.

Evaluation de la toxicité d’un aphrodisiaque ivoirien d’origine naturelle (aphro) chez le rat

L’usage des plantes à des fins thérapeutiques est très ancien. De nos jours encore, elles sont utilisées par une frange partie de la population mondiale. La présente étude avait pour objectif d’étudier les effets toxiques de l'administration subaiguë d'un phytomédicament ivoirien Aphro vendu sur les marchés chez le rat Wistar albinos. Pour ce faire, le phytomédicament Aphro a été administré quotidiennement par voie orale à trois lots de rats, aux doses de 5,75 ; 11,5 et 23 mg/kg de poids corporel et cela pendant 28 jours. Le taux de croissance des rats, les paramètres hématologiques et biochimiques sanguins ont été étudiés. Les résultats de cette étude ont révélé qu’aux doses de 5,75 ; 11,5 et 23 mg/kg de poids corporel, le phytomédicament ne modifie pas de façon significative le taux d’accroissement de la masse corporelle et les paramètres hématologiques constitués du taux de globules rouges, de globules blancs, de plaquettes sanguines et le taux d’hémoglobine. Par ailleurs, des marqueurs hépatiques et lipidiques sériques des rats traités n'ont pas été perturbés par l’administration du phytomédicament comparativement au lot témoin. En revanche, la créatinine, l’un des marqueurs rénaux étudiés a connu une augmentation significative (p < 0,05) chez les rats traités avec le phytomédicament aux doses de 11,5 et 23 mg/kg de poids corporel par rapport au lot témoin. Cette étude a permis de montrer que l’usage prolongé et à doses élevées de Aphro pourrait entraîner des effets nocifs sur les reins. English title: Assessment of the toxicity of an ivoirian natural aphrodisiac (aphro) on rat The use of plants for therapeutic purposes is very old. Even today, they are used by a part of the world population. The objective of the present study was to evaluate the toxic effects of the subacute administration of an ivorian phytomedicine Aphro sold in the markets in the albino Wistar rat. To do this, the phytomedicine Aphro was administered orally daily to three batches of rats, at doses of 5.75; 11.5 and 23 mg/kg of body weight for 28 days. Rats' growth rate, blood haematological and biochemical parameters were studied. The results of this study showed that at doses of 5.75; 11.5 and 23 mg/kg of body weight, the phytomedicine does not significantly modify the rate of increase in body mass and the hematological parameters consisting of the level of red blood cells, white blood cells, blood platelets and the level hemoglobin. Furthermore, hepatic and serum lipid markers of the treated rats were not disturbed by the administration of the phytomedicine compared to the control group. In contrast, creatinine, one of the renal markers studied, experienced a significant increase (p <0.05) in rats treated with the phytomedicine at doses of 11.5 and 23 mg / kg of body weight compared to the batch witness. This study showed that prolonged use and in high doses of Aphro could cause harmful effects on the kidneys.