4. (25 points total) You want to understand how the density of P-selectin on the
ID: 150995 • Letter: 4
Question
4. (25 points total) You want to understand how the density of P-selectin on the endothelial lining of blood vessels affects the rolling interactions of neutrophils (a type of white blood cell) when they are subjected to hydrodynamic drag forces in the blood. You introduce P-selectin into a synthetic lipid bilayer and attach it to a glass slide mounted on the floor of a flow chamber on a microscope stage. This arrangement allows you to measure neutrophil motion at different densities of P-selectin and at different flow rates At high densities of P-selectin, from 40 to 400 molecules per um', the cells stay attached to the membrane floor and roll slowly and irregularly with the flow of the medium. At P-selectin densities from 1 to 15 molecules per m2, the cells either move freely with the medium or become transiently tethered to the membrane floor before beginning moving again. When the system is treated with monoclonal antibodies against P-selectin, the tethering events do not occur, and the cells flow freely with the medium At the lower P-selectin densities, the number of tethering events per unit time is directly proportional to the density of P-selectin. At a low flow rate, roughly equivalent to a cell being subjected to 40 piconewtons of shear, the time cells stay tethered follows a simple exponential decay curve with a cellular off rate, koff, of about 1.0 sec This off rate constant is unaffected by the density of P-selectin a. What is the point of doing the experiment with the monoclonal antibody? (5 points) b. Is a single interaction between one P-selectin molecule and one copy its glycoprotein ligand on the surface of the neutrophil sufficient to tether the cell to the floor of the flow chamber transiently? Explain your reasoning. (10 points) c. If you increase the flow rate by a factor of three, increasing the shear force felt by each cell to about 110 piconewtons, kofr increases to about 3.5 sec1. Explain how this force can affect a dissociation rate constant. (10 points)Explanation / Answer
Answer
a.Monoclonal antibody against P-selectin has been used to block P-selectin. , it will be blocked when P-selectin binds to its mAb. Then, the mobility or the attachment of the cells to membrane surface without P-selectin could be checked.
b.it appears that a high density of P-selectin in required for tethering of the cells.unlikely that a single molecule of P-selectin will be able to tether the cell to the membrane floor, but a single interaction P-selectin molecule and its glycoprotein ligand on neutrophil surface is a interaction to be considered.
c. In certain experiments flowing cells have been found to be unable to attach
if the flow rate is increased by a shear force,
the dissociation will increase and hence the dissociation constant is directly prportional to the dissociation process, the dissociation rate constant should increase.
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