72 research outputs found
Outer Limits of Biotechnologies: A Jewish Perspective
A great deal of biomedical research focuses on new biotechnologies such as gene editing, stem cell biology and reproductive medicine, which have created a scientific revolution. While the potential medical benefits of this research may be far-reaching, ethical issues related to non-medical applications of these technologies are demanding. We analyze, from a Jewish legal perspective, some of the ethical conundrums that society faces in pushing the outer limits in researching these new biotechnologies
Creatine kinase expression and creatine phosphate accumulation are developmentally regulated during differentiation of mouse and human monocytes
We have studied the expression of creatine kinase (CK) and the accumulation of creatine phosphate during the differentiation of human and mouse peripheral blood monocytes. Mouse monocytes cultured for 24 h do not contain detectable levels of CK and creatine phosphate. However, resident tissue macrophages and inflammatory elicited macrophages obtained from the peritoneal cavities of mice have 70 and 300 mU per mg protein of CK activity and contain 3 and 6 mol of creatine phosphate per mol of ATP, respectively. The major isozyme of CK in these cells has been identified as the brain form. These findings suggest that the differentiation of monocytes into macrophages is associated with the expression of CK and the accumulation of creatine phosphate. We have found a similar pattern in human monocytes. Human blood monocytes, maintained in culture for 24 or 48 h, do not contain detectable levels of CK or creatine phosphate. Monocyte-derived macrophages (monocytes maintained in tissue cultures for 1 to 2 wk) have up to 100 mU per mg protein of CK activity and contain 0.5 mol of creatine phosphate per mol of ATP. Human macrophages express multiple isozymes of CK including the brain (BB) and possibly the mitochondrial forms of this enzyme. Thus, the expression of CK and the accumulation of creatine phosphate in human monocytes is induced by their in vitro cultivation. The induction of CK during in vitro cultivation occurs independently of the concentration of creatine in the medium. However, the size of the creatine phosphate pool varies with respect to extracellular creatine concentration. Creatine phosphate and CK are not detectable in freshly isolated human lymphocytes, polymorphonuclear leukocytes or erythrocytes, but are found in freshly isolated human platelets
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Determination of the Critical Concentration of Neutrophils Required to Block Bacterial Growth in Tissues
We showed previously that the competition between bacterial killing by neutrophils and bacterial growth in stirred serum-containing suspensions could be modeled as the competition between a first-order reaction (bacterial growth) and a second-order reaction (bacterial killing by neutrophils). The model provided a useful parameter, the critical neutrophil concentration (CNC), below which bacterial concentration increased and above which it decreased, independent of the initial bacterial concentration. We report here that this model applies to neutrophil killing of bacteria in three-dimensional fibrin matrices and in rabbit dermis. We measured killing of 103–108 colony forming units/ml Staphylococcus epidermidis by 105–108 human neutrophils/ml in fibrin gels. The CNC was ∼4 × 106 neutrophils/ml gel in the presence of normal serum and ∼1.6 × 107 neutrophils/ml gel in the presence of C5-deficient serum. Application of our model to published data of others on killing of ∼5 × 107 to 2 × 108 E. coli/ml rabbit dermis yielded CNCs from ∼4 × 106 to ∼8 × 106 neutrophils/ml dermis. Thus, in disparate tissues and tissuelike environments, our model fits the kinetics of bacterial killing and gives similar lower limits (CNCs) to the neutrophil concentration required to control bacterial growth
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Out of Many, One: Collaborating is the Way to Accelerate COVID 19 Vaccine Development
The article titled "Out of Many, One: Collaborating is the Way to Accelerate COVID-19 Vaccine Development," authored by John Loike, Matthew Cobb, Patricia J. Williams, and Robert Pollack, emphasizes the importance of global collaboration in the rapid development of vaccines to combat the COVID-19 pandemic. The authors argue that the traditional competitive approach to drug development, characterized by stringent intellectual property protections, is inadequate in the face of a global health crisis.
They advocate for a temporary suspension of patents and intellectual property rights for COVID-19 vaccines to facilitate open sharing of research data and technologies among pharmaceutical companies and research institutions. This collaboration is deemed essential to expedite the development, testing, and distribution of effective vaccines. The article highlights the need for coordinated efforts to overcome the challenges at each stage of vaccine development, from lab research to clinical trials, and underscores the potential economic benefits of such collaboration, which could ultimately save lives and restore global economic stability.
By leveraging the collective knowledge and resources of the global scientific community, the authors believe that the process of developing and deploying a safe and effective COVID-19 vaccine can be significantly accelerated
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Role of facilitative glucose transporters in diffusional water permeability through J774 cells
We have reported previously that in the presence of an osmotic gradient, facilitative glucose transporters (GLUTs) act as a transmembrane pathway for water flow. Here, we find evidence that they also allow water passage in the absence of an osmotic gradient. We applied the linear diffusion technique to measure the diffusional permeability (Pd) of tritiated water (3H-H2O) through plasma membranes of J774 murine macrophage-like cells. Untreated cells had a Pd of 30.9 +/- 1.8 microns/s; the inhibitors of facilitative glucose transport cytochalasin B (10 microM) and phloretin (20 microM) reduced that value to 15.3 +/- 1.8 (50%) and 11.0 +/- 0.7 (62%) microns/s, respectively. In contrast, no significant effect on Pd was observed in cells treated with dihydrocytochalasin B (Pd = 28.4 +/- 1.5 microns/s). PCMBS (3 mM) inhibited glucose uptake by greater than 95%, and 3H-H2O diffusion by approximately 30% (Pd = 22.9 +/- 1.5 microns/s). The combination of cytochalasin B plus pCMBS reduced Pd by about 87% (Pd = 3.9 +/- 0.3 microns/s). Moreover, 1 mM pCMBS did not affect the osmotic water permeability in Xenopus laevis oocytes expressing the brain/erythroid form of facilitative glucose transporters (GLUT1). These results indicate for the first time that about half of the total Pd of J774 cells may be accounted for by water passage across GLUTs. Hence, they highlight the multifunctional properties of these transporters serving as conduits for both water and glucose. Our results also suggest for the first time that pCMBS blocks glucose transport without affecting water permeation through GLUTs. Lastly, because pCMBS decreases the Pd of J774 cells, this suggests the presence in their plasma membranes of another protein(s) exhibiting water channel properties
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Statin Inhibition of Fc Receptor–Mediated Phagocytosis by Macrophages Is Modulated by Cell Activation and Cholesterol
Objectives— An inflammatory response to altered lipoproteins that accumulate in the arterial wall is a major component of the pathogenesis of atherosclerosis. Statins reduce plasma levels of low-density lipoprotein (LDL) and are effective treatments for atherosclerosis. It is hypothesized that they also modulate inflammation. The aim of this study was to examine whether lovastatin inhibits macrophage inflammatory processes and clarify its mechanism of action.
Methods and Results— We examined the effects of statins on phagocytosis of antibody-coated red blood cells by cultured human monocytes and mouse peritoneal macrophages. Lovastatin, simvastatin, and zaragozic acid, a squalene synthase inhibitor, blocked Fc receptor–mediated phagocytosis by cultured human monocytes and mouse peritoneal macrophages. The inhibitory effect of lovastatin on Fc receptor–mediated phagocytosis was prevented completely by addition of mevalonate, farnesyl pyrophosphate, LDL, or cholesterol to the culture medium. The inhibitory effect of zaragozic acid was reversed by addition of LDL, but not by the addition of geranylgeranyl pyrophosphate, to the medium. In addition, the effect of lovastatin on phagocytosis is a function of cell activation because treatment of cells with tumor necrosis factor-α or lipopolysaccharide prevented inhibition of phagocytosis by lovastatin.
Conclusions— The inhibition of Fc receptor–mediated phagocytosis of lovastatin is related to its effect on cholesterol biosynthesis rather than its effect on the formation of isoprenoids
Lipoprotein lipase regulates Fc receptor-mediated phagocytosis by macrophages maintained in glucose-deficient medium
During periods of intense activity such as phagocytosis, macrophages are thought to derive most of their energy from glucose metabolism under both aerobic and anaerobic conditions. To determine whether fatty acids released from lipoproteins by macrophage lipoprotein lipase (LPL) could substitute for glucose as a source of energy for phagocytosis, we cultured peritoneal macrophages from normal and LPL knockout (LPL-KO) mice that had been rescued from neonatal demise by expression of human LPL via the muscle creatine kinase promoter. Normal and LPL-KO macrophages were cultured in medium containing normal (5 mM) or low (1 mM) glucose, and were tested for their capacity to phagocytose IgG-opsonized sheep erythrocytes. LPL-KO macrophages maintained in 1 and 5 mM glucose phagocytosed 67 and 79% fewer IgG-opsonized erythrocytes, respectively, than macrophages from normal mice. Addition of VLDL to LPL-expressing macrophages maintained in 1 mM glucose enhanced the macrophages' phagocytosis of IgG-opsonized erythrocytes, but did not stimulate phagocytosis by LPL-KO macrophages. Inhibition of secreted LPL with a monoclonal anti-LPL antibody or with tetrahydrolipstatin blocked the ability of VLDL to enhance phagocytosis by LPL-expressing macrophages maintained in 1 mM glucose. Addition of oleic acid significantly enhanced phagocytosis by both LPL-expressing and LPL-KO macrophages maintained in 1 mM glucose. Moreover, oleic acid stimulated phagocytosis in cells cultured in non-glucose-containing medium, and increased the intracellular stores of creatine phosphate. Inhibition of oxidative phosphorylation, but not of glycolysis, blocked the capacity of oleic acid to stimulate phagocytosis. Receptor-mediated endocytosis of acetyl LDL by macrophages from LPL-expressing and LPL-KO mice was similar whether the cells were maintained in 5 or 1 mM glucose, and was not augmented by VLDL. We postulate that fatty acids derived from macrophage LPL-catalyzed hydrolysis of triglycerides and phospholipids provide energy for macrophages in areas that have limited amounts of ambient glucose, and during periods of intense metabolic activity
Neuronal MHC-I display in T-cell mediated neurodegeneration
Parkinson’s disease (PD) and other disorders feature the degeneration of ventral midbrain (VM) catecholamine neurons. Recent data suggest that neuroinflammatory mechanisms contribute to a cascade of events leading to chronic neuronal degeneration
Fibrin regulates neutrophil migration in response to interleukin 8, leukotriene B4, tumor necrosis factor, and formyl-methionyl-leucyl-phenylalanine
We have examined the capacity of four different chemoattractants/cytokines to promote directed migration of polymorphonuclear leukocytes (PMN) through three-dimensional gels composed of extracellular matrix proteins. About 20% of PMN migrated through fibrin gels and plasma clots in response to a gradient of interleukin 8 (IL-8) or leukotriene B4 (LTB4). In contrast, < 0.3% of PMN migrated through fibrin gels in response to a gradient of tumor necrosis factor alpha (TNF) or formyl-methionyl-leucyl-phenylalanine (FMLP). All four chemoattractants stimulated PMN to migrate through gels composed of collagen IV or of basement membrane proteins (Matrigel), or through filters to which fibronectin or fibrinogen had been adsorbed. PMN stimulated with TNF or FMLP adhered and formed zones of close apposition to fibrin, as measured by the exclusion of a 10-kD rhodamine-polyethylene glycol probe from the contact zones between PMN and the underlying fibrin gel. By this measure, IL-8- or LTB4-treated PMN adhered loosely to fibrin, since 10 kD rhodamine-polyethylene glycol permeated into the contact zones between these cells and the underlying fibrin gel. PMN stimulated with FMLP and IL-8, or FMLP and LTB4, exhibited very little migration through fibrin gels, and three times as many of these cells excluded 10 kD rhodamine-polyethylene glycol from their zones of contact with fibrin as PMN stimulated with IL-8 or LTB4 alone. These results show that PMN chemotaxis is regulated by both the nature of the chemoattractant and the composition of the extracellular matrix; they suggest that certain combinations of chemoattractants and matrix proteins may limit leukocyte movements and promote their localization in specific tissues in vivo
Activated platelets form protected zones of adhesion on fibrinogen and fibronectin-coated surfaces.
Leukocytes form zones of close apposition when they adhere to ligand-coated surfaces. Because plasma proteins are excluded from these contact zones, we have termed them protected zones of adhesion. To determine whether platelets form similar protected zones of adhesion, gel-filtered platelets stimulated with thrombin or ADP were allowed to adhere to fibrinogen- or fibronectin-coated surfaces. The protein-coated surfaces with platelets attached were stained with either fluorochrome-conjugated goat anti-human fibrinogen or anti-human fibronectin antibodies, or with rhodamine-conjugated polyethylene glycol polymers. Fluorescence microscopy revealed that F(ab')2 anti-fibrinogen (100 kD) did not penetrate into the contact zones between stimulated platelets and the underlying fibrinogen-coated surface, while Fab antifibrinogen (50 kD) and 10 kD polyethylene glycol readily penetrated and stained the substrate beneath the platelets. Thrombin- or ADP-stimulated platelets also formed protected zones of adhesion on fibronectin-coated surfaces. F(ab')2 anti-fibronectin and 10 kD polyethylene glycol were excluded from these adhesion zones, indicating that they are much less permeable than those formed by platelets on fibrinogen-coated surfaces. The permeability properties of protected zones of adhesion formed by stimulated platelets on surfaces coated with both fibrinogen and fibronectin were similar to the zones of adhesion formed on fibronectin alone. mAb 7E3, directed against the alpha IIb beta 3 integrin blocked the formation of protected adhesion zones between thrombin-stimulated platelets and fibrinogen or fibronectin coated surfaces. mAb C13 is directed against the alpha 5 beta 1 integrin on platelets. Stimulated platelets treated with this mAb formed protected zones of adhesion on surfaces coated with fibronectin. These protected zones were impermeable to F(ab')2 antifibronectin but were permeable to 10 kD polyethylene glycol. These results show that activated platelets form protected zones of adhesion and that the size of molecules excluded from these zones depends upon the composition of the matrix proteins to which the platelets adhere. They also show that formation of protected zones of adhesion by platelets requires alpha IIb beta 3 integrins while the permeability properties of these zones of adhesion are regulated by both alpha IIb beta 3 and alpha 5 beta 1 integrins
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