Introduction to Bioproducts and Bioseparations

2015 ◽  
Author(s):  
Roger G. Harrison ◽  
Paul W. Todd ◽  
Scott R. Rudge ◽  
Demetri P. Petrides
Keyword(s):  
Ex Vivo ◽  
Blood Clot ◽  
Chemical Properties ◽  
Downstream Processing ◽  
Added Value ◽  
Chemical Substances ◽  
Whole Cells ◽  
Living Things ◽  
Insecticide Activity

Bioproducts—chemical substances or combinations of chemical substances that are made by living things—range from methanol to whole cells. They are derived by extraction from whole plants and animals or by synthesis in bioreactors containing cells or enzymes. Bioproducts are sold for their chemical activity: methanol for solvent activity, ethanol for its neurological activity or as a fuel, penicillin for its antibacterial activity, taxol for its anticancer activity, streptokinase (an enzyme) for its blood clot dissolving activity, hexose isomerase for its sugar-converting activity, and whole Bacillus thuringiensis cells for their insecticide activity, to name a few very different examples. The wide variety represented by this tiny list makes it clear that bioseparations must encompass a correspondingly wide variety of methods. The choice of separation method depends on the nature of the product, remembering that purity, yield, and activity are the goals, and the most important of these is activity. This first chapter therefore reviews the chemical properties of bioproducts with themes and examples chosen to heighten awareness of those properties that must be recognized in the selection of downstream processes that result in acceptably high final purity while preserving activity. The final part of this chapter is an introduction to the field of bioseparations, which includes a discussion of the stages of downstream processing, the basic principles of engineering analysis as applied to bioseparations, and the various factors involved in developing a bioproduct for the marketplace. The pharmaceutical, agrichemical, and biotechnology bioproduct industries account for many billion dollars in annual sales—neglecting, of course, commodity foods and beverages. By “bioproduct” we mean chemical substances that are produced in or by a biological process, either in vivo or ex vivo (inside or outside a living organism). Figure 1.1 indicates a clear inverse relationship between bioproduct market size and cost. Owing to intense competition, cost, price, and value are very closely related, except in the case of completely new products that are thoroughly protected by patents, difficult to copy, and of added value to the end user.

scholarly journals Inhibition of Histone Demethylases LSD1 and UTX Regulates ERα Signaling in Breast Cancer

Cancers ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 2027 ◽  
Author(s):  
Rosaria Benedetti ◽  
Carmela Dell’Aversana ◽  
Tommaso De Marchi ◽  
Dante Rotili ◽  
Ning Qing Liu ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Histone Methylation ◽  
Ex Vivo ◽  
Acquired Resistance ◽  
Chemical Properties ◽  
Breast Cancers ◽  
Histone Demethylases ◽  
Lysine Specific Demethylase ◽  
Breast Cancer Model

In breast cancer, Lysine-specific demethylase-1 (LSD1) and other lysine demethylases (KDMs), such as Lysine-specific demethylase 6A also known as Ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX), are co-expressed and co-localize with estrogen receptors (ERs), suggesting the potential use of hybrid (epi)molecules to target histone methylation and therefore regulate/redirect hormone receptor signaling. Here, we report on the biological activity of a dual-KDM inhibitor (MC3324), obtained by coupling the chemical properties of tranylcypromine, a known LSD1 inhibitor, with the 2OG competitive moiety developed for JmjC inhibition. MC3324 displays unique features not exhibited by the single moieties and well-characterized mono-pharmacological inhibitors. Inhibiting LSD1 and UTX, MC3324 induces significant growth arrest and apoptosis in hormone-responsive breast cancer model accompanied by a robust increase in H3K4me2 and H3K27me3. MC3324 down-regulates ERα in breast cancer at both transcriptional and non-transcriptional levels, mimicking the action of a selective endocrine receptor disruptor. MC3324 alters the histone methylation of ERα-regulated promoters, thereby affecting the transcription of genes involved in cell surveillance, hormone response, and death. MC3324 reduces cell proliferation in ex vivo breast cancers, as well as in breast models with acquired resistance to endocrine therapies. Similarly, MC3324 displays tumor-selective potential in vivo, in both xenograft mice and chicken embryo models, with no toxicity and good oral efficacy. This epigenetic multi-target approach is effective and may overcome potential mechanism(s) of resistance in breast cancer.

scholarly journals Reduced Contraction of Blood Clots in Venous Thromboembolism Is a Potential Thrombogenic and Embologenic Mechanism

TH Open ◽  
2018 ◽  
Vol 02 (01) ◽  
pp. e104-e115 ◽  
Author(s):  
Alina Peshkova ◽  
Dmitry Malyasyov ◽  
Roman Bredikhin ◽  
Giang Le Minh ◽  
Izabella Andrianova ◽  
...  
Keyword(s):  
Venous Thromboembolism ◽  
Ex Vivo ◽  
Blood Clot ◽  
Chemical Activation ◽  
Healthy Controls ◽  
Activation Markers ◽  
Fold Reduction ◽  
Blood Clots ◽  
Clot Contraction

AbstractContraction (retraction) of the blood clot is a part of the clotting process driven by activated platelets attached to fibrin that can potentially modulate the obstructiveness and integrity of thrombi. The aim of this work was to reveal the pathogenic importance of contraction of clots and thrombi in venous thromboembolism (VTE). We investigated the kinetics of clot contraction in the blood of 55 patients with VTE. In addition, we studied the ultrastructure of ex vivo venous thrombi as well as the morphology and functionality of isolated platelets. Thrombi from VTE patients contained compressed polyhedral erythrocytes, a marker for clot contraction in vivo. The extent and rate of contraction were reduced by twofold in clots from the blood of VTE patients compared with healthy controls. The contraction of clots from the blood of patients with pulmonary embolism was significantly impaired compared with that of those with isolated venous thrombosis, suggesting that less compacted thrombi are prone to embolization. The reduced ability of clots to contract correlated with continuous platelet activation followed by their partial refractoriness. Morphologically, 75% of platelets from VTE patients were spontaneously activated (with filopodia) compared with only 21% from healthy controls. At the same time, platelets from VTE patients showed a 1.4-fold reduction in activation markers expressed in response to chemical activation when compared with healthy individuals. The results obtained suggest that the impaired contraction of thrombi is an underappreciated pathogenic mechanism in VTE that may regulate the obstructiveness and embologenicity of venous thrombi.

A Hybrid Plasminogen Activator Binds to the u-PA Receptor and Has a Reduced Thrombolytic Potency In Vivo

1993 ◽  
Vol 69 (01) ◽  
pp. 050-055 ◽  
Author(s):  
Fred A M Asselbergs ◽  
Rolf Bürgi ◽  
Jessica Hamerman ◽  
Jutta Heim ◽  
Jan van Oostrum ◽  
...  
Keyword(s):  
Growth Factor ◽  
Ex Vivo ◽  
Blood Clot ◽  
Clotting Time ◽  
Jugular Vein Thrombosis ◽  
Thrombolytic Activity ◽  
Vein Thrombosis ◽  
Kringle 2 ◽  
Blood Elements

SummaryFibrinolytic properties of four hybrids of u-PA and t-PA, all containing the u-PA growth factor domain and binding to recombinant human u-PA receptor expressed in CHO cells, were compared. Highest fibrin stimulation was observed with uK2tPA which when compared to t-PA in the rabbit system, had a considerably prolonged circulatory half-life in vivo. Compared to an equimolar dose of t-PA, 0.4 mg/kg uK2tPA caused a similar consumption of α2-antiplasmin and fibrinogen and a considerably greater prolongation of the ex-vivo blood clotting time. Nevertheless, this dose of uK2tPA was inactive in the jugular vein thrombosis assay. This lack of thrombolytic activity is presumably due to the presence of a functional u-PA growth factor domain, which in binding uK2tPA to cellular blood elements possibly retards its penetration into the blood clot and in this manner could neutralize the potential thrombolytic activity of the t-PA kringle 2 and protease domains in uK2tPA.

scholarly journals Mobilisation of Joint-resident Mesenchymal Stromal Cells for Articular Cartilage Regeneration

2017 ◽  
Vol 5 (2_suppl2) ◽  
pp. 2325967117S0007
Author(s):  
Alam Khalil Khan ◽  
Thomas Baboolal ◽  
Owen Wall ◽  
Elena Jone ◽  
Dennis Mcgonagle
Keyword(s):  
Synovial Fluid ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Blood Clot ◽  
Cartilage Regeneration ◽  
Mechanical Agitation ◽  
Cytology Brush

Background and Objectives: Microfracture is a recognized procedure used to treat isolated cartilage injuries or defects, in which bone marrow mesenchymal stromal cells (BM-MSCs) are thought to migrate into the resulting blood clot, leading to subsequent cartilage repair via fibrocartilage formation. The discovery of MSCs in the synovium and synovial fluid (SF) provides a potential mechanism for repairing cartilage from the top down via their migration and homing to the microfracture site, however SF-MSCs low in number and usually lost with joint irrigation. The purpose of this work was threefold; first to test the hypothesis that SF-MSCs can be replaced, and also their numbers further increased by synovial agitation, second that these cells were capable of rapid adhesion to clots and third that the clot composition improve MSC migration. Materials-Methods: Ex-vivo mechanical agitation of human superficial synovium and in vivo intra-operative agitation of synovium in patients undergoing arthroscopy were performed using a Cytology brush and custom designed Synovial brush. Colony-forming unit-fibroblast (CFU-F) assay was performed to quantify released MSCs. Adhesion to clots was studied by comparing Platelet Rich Plasma (PRP), Whole Blood (WB) and Fibrin Glue (FG). Migration studies were performed using passage 2-4 synovial MSCs in trans-well migration assay. MSC migration, over a five hour period, was compared between PRP and pooled human Platelet Lysate (hPL). Results: Ex-vivo mechanical agitating of the synovium with the cytology brush compared to irrigation alone increased MSC number 2.7-fold (n=10, p=0.002). Irrigation during arthroscopy was seen to effectively remove the majority of CFU-Fs for the synovial fluid. Use of a custom designed synovial brush, compared to the cytology brush resulted in a median 65-fold increase in the number of CFU-Fs (n=8, p=0.0148). Trilineage differentiation of released synovial MSCs was at least comparable to donor match synovial fluid MSCs. These MSCs adhered to clots within 30 minutes with no difference seen between clot compositions. Released synovial MSCs demonstrated a trend for a better migration towards hPL compared to PRP. Conclusions: Existing surgical procedures wash away SF-MSCs. Using a novel brushing technique and a custom designed synovial brush, synovial MSCs can be mechanically released in vivo, and these cells were capable of migration and rapid adhesion to clots. Collectively these findings aid in the rapid replenishment of endogenous minimally manipulated MSCs, and provide a one-stage procedure combining synovial brushing with microfracture, as a strategy for cost effective joint repair.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Antithrombotic Effects and Bleeding Time Prolongation with Synthetic Platelet GPIIb/llla Inhibitors in Animal Models of Platelet-Mediated Thrombosis

1994 ◽  
Vol 71 (01) ◽  
pp. 095-102 ◽  
Author(s):  
Désiré Collen ◽  
Hua Rong Lu ◽  
Jean-Marie Stassen ◽  
Ingrid Vreys ◽  
Tsunehiro Yasuda ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Bleeding Time ◽  
Bolus Injection ◽  
Cell Injury ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Femoral Vein ◽  
Thrombotic Occlusion ◽  
Antithrombotic Effects

SummaryCyclic Arg-Gly-Asp (RGD) containing synthetic peptides such as L-cysteine, N-(mercaptoacetyl)-D-tyrosyl-L-arginylglycyl-L-a-aspartyl-cyclic (1→5)-sulfide, 5-oxide (G4120) and acetyl-L-cysteinyl-L-asparaginyl-L-prolyl-L-arginyl-glycyl-L-α-aspartyl-[0-methyltyrosyl]-L-arginyl-L-cysteinamide, cyclic 1→9-sulfide (TP9201) bind with high affinity to the platelet GPIIb/IIIa receptor.The relationship between antithrombotic effect, ex vivo platelet aggregation and bleeding time prolongation with both agents was studied in hamsters with a standardized femoral vein endothelial cell injury predisposing to platelet-rich mural thrombosis, and in dogs with a carotid arterial eversion graft inserted in the femoral artery. Intravenous administration of G4120 in hamsters inhibited in vivo thrombus formation with a 50% inhibitory bolus dose (ID50) of approximately 20 μg/kg, ex vivo ADP-induccd platelet aggregation with ID50 of 10 μg/kg, and bolus injection of 1 mg/kg prolonged the bleeding time from 38 ± 9 to 1,100 ± 330 s. Administration of TP9201 in hamsters inhibited in vivo thrombus formation with ID50 of 30 μg/kg, ex vivo platelet aggregation with an ID50 of 50 μg/kg and bolus injection of 1 mg/kg did not prolong the template bleeding time. In the dog eversion graft model, infusion of 100 μg/kg of G4120 over 60 min did not fully inhibit platelet-mediated thrombotic occlusion but was associated with inhibition of ADP-induccd ex vivo platelet aggregation and with prolongation of the template bleeding time from 1.3 ± 0.4 to 12 ± 2 min. Infusion of 300 μg/kg of TP9201 over 60 min completely prevented thrombotic occlusion, inhibited ex vivo platelet aggregation, but was not associated with prolongation of the template bleeding time.TP9201, unlike G4120, inhibits in vivo platelet-mediated thrombus formation without associated prolongation of the template bleeding time.

Significance of Platelet β-Adrenoceptors for Platelet Responses In Vivo and In Vitro

1992 ◽  
Vol 68 (06) ◽  
pp. 687-693 ◽  
Author(s):  
P T Larsson ◽  
N H Wallén ◽  
A Martinsson ◽  
N Egberg ◽  
P Hjemdahl
Keyword(s):  
Ex Vivo ◽  
High Dose ◽  
Platelet Aggregability ◽  
Adrenoceptor Agonist ◽  
Dose Infusion ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Factor Antigen

SummaryThe significance of platelet β-adrenoceptors for platelet responses to adrenergic stimuli in vivo and in vitro was studied in healthy volunteers. Low dose infusion of the β-adrenoceptor agonist isoprenaline decreased platelet aggregability in vivo as measured by ex vivo filtragometry. Infusion of adrenaline, a mixed α- and β-adrenoceptor agonist, increased platelet aggregability in vivo markedly, as measured by ex vivo filtragometry and plasma β-thromboglobulin levels. Adrenaline levels were 3–4 nM in venous plasma during infusion. Both adrenaline and high dose isoprenaline elevated plasma von Willebrand factor antigen levels β-Blockade by propranolol did not alter our measures of platelet aggregability at rest or during adrenaline infusions, but inhibited adrenaline-induced increases in vWf:ag. In a model using filtragometry to assess platelet aggregability in whole blood in vitro, propranolol enhanced the proaggregatory actions of 5 nM, but not of 10 nM adrenaline. The present data suggest that β-adrenoceptor stimulation can inhibit platelet function in vivo but that effects of adrenaline at high physiological concentrations are dominated by an α-adrenoceptor mediated proaggregatory action.

In Vivo Effect of Suloctidil as an Antiplatelet Agent

1979 ◽  
Vol 41 (03) ◽  
pp. 465-474 ◽  
Author(s):  
Marcia R Stelzer ◽  
Thomas S Burns ◽  
Robert N Saunders
Keyword(s):  
Ex Vivo ◽  
Antiplatelet Agent ◽  
Ratio Method ◽  
Platelet Aggregate ◽  
Permanent Effect ◽  
Platelet Aggregates ◽  
5 Ht Uptake ◽  
High Doses

SummaryThe relationship between the effects of suloctidil in vivo as an antiplatelet agent and in vitro as a modifier of platelet serotonin (5-HT) parameters was investigated. Suloctidil was found to be effective in reducing platelet aggregates formation in the retired breeder rat as determined using the platelet aggregate ratio method (PAR) with an ED50 of 16.1 mg/kg 24 hours post administration. In contrast to the hypothesis that 5-HT depletion is involved in the anti-aggregatory mechanism of suloctidil, no correlation was found between platelet 5- HT content and this antiplatelet activity. Reduction of platelet 5-HT content required multiple injections of high doses (100 mg/kg/day) of suloctidil. Suloctidil administration for 8 days at 100 mg/kg/day, which lowered platelet 5-HT content by 50%, resulted in no permanent effect on ex vivo platelet 5-HT uptake or thrombin-induced release, nor alteration in the plasma 5-HT level. However, these platelets exhibited a short-lived, significant increase in percent leakage of 5-HT after 30 minutes of incubation. Therefore, suloctidil treatment at high doses may with time result in platelet 5-HT depletion, however this effect is probably not related to the primary anti-aggregatory activity of the drug.

Studies on the Haemostatic Defect in a Complicated Syndrome

1995 ◽  
Vol 74 (05) ◽  
pp. 1244-1251 ◽  
Author(s):  
H Stormorken ◽  
H Holmsen ◽  
R Sund ◽  
K S Sakariassen ◽  
T Hovig ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Bleeding Tendency ◽  
Clot Retraction ◽  
Abnormal Finding ◽  
Shear Rates ◽  
Perfusion Chamber ◽  
Coagulant Activity ◽  
5 Ht Uptake ◽  
And Storage

SummaryThe Stormorken syndrome is a multifacetted syndrome including a bleeding tendency. No deviations were found in the coagulation- or fibrinolytic systems. Platelet number was low normal, and size abnormal, whereas EM findings were unremarkable. Survival time was half normal. Clot retraction was initially rapid, but clearly decreased, whereas prothrombin consumption was also initially rapid, but complete. Membrane GP’s were normal, so was AA metabolism, PI-cycle, granule storage and secretion, and c-AMP function, whereas 5-HT uptake and storage was decreased. Optical platelet aggregation was low normal with all physiological agonists. The only clearly abnormal finding was that coagulant activity was present on non stimulated platelets at the same level as kaolin-stimulated normal platelets. This indicated a platelet abnormality which should lead to a thrombogenic, not to a haemorrhagic trait. This paradox may have its origin in rheology, because when challenged with in vivo shear rates in an ex vivo perfusion chamber, platelet cohesion was abnormally low. Further studies to better delineate the membrane abnormality are underway.

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